Antiviral antisense oligonucleotide

Abstract

In the present description, provided are an antisense oligonucleotide targeting a specific region of the genomic RNA of SARS-CoV-2, a pharmaceutically acceptable salt thereof or a hydrate of the same, a pharmaceutical composition containing the antisense oligonucleotide, a pharmaceutically acceptable salt thereof or a hydrate of the same, etc.

Description

Antiviral antisense oligonucleotides

[0001] The present invention relates to an antisense oligonucleotide or a pharmaceutically acceptable salt thereof, or a hydrate of these (hereinafter also referred to as "the antisense oligonucleotide, etc.") having an antiviral effect against SARS-CoV-2, SARS-CoV-1, or MERS-CoV; a pharmaceutical composition containing the antisense oligonucleotide, etc.; or a method for treating and / or preventing a viral infection, comprising the step of administering the antisense oligonucleotide, etc. or the pharmaceutical composition to a subject.

[0002] COVID-19 (Coronavirus disease-2019) is a novel infectious disease characterized by pneumonia that was first identified in Wuhan, Hubei Province, China in November 2019 and subsequently declared a pandemic by the WHO in March 2020 (Non-Patent Document 1). The etiology of COVID-19 is a novel virus, the causative virus of which was identified in January 2020 as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). SARS-CoV-2 is evolutionarily closely related to SARS-CoV, the causative virus of the severe acute respiratory syndrome (SARS) epidemic that occurred in 2003, and, like SARS-CoV, belongs to the Betacoronavirus genus of the Coronaviridae family (Non-Patent Document 2).

[0003] World Health Organization (WHO) (Press release). 11 March 2020Nat Microbiol. 2020 Apr;5(4):536-544

[0004] The use of existing antiviral drugs has been proposed for SARS-CoV-2, but their effectiveness has not yet been clinically proven, and no established treatment exists.Furthermore, there is no established treatment for SARS-CoV-1 or MERS-CoV, which, like SARS-CoV-2, belong to the Betacoronavirus genus and cause viral infections.

[0005] In this situation, it is desirable to provide new therapeutic agents that have antiviral effects against SARS-CoV-2, SARS-CoV-1, or MERS-CoV.

[0006] The present invention provides antisense oligonucleotides that target specific regions of the genomic RNA of SARS-CoV-2, or pharmaceutically acceptable salts thereof, or hydrates thereof, as well as pharmaceutical compositions containing the antisense oligonucleotides, or pharmaceutically acceptable salts thereof, or hydrates thereof, as described below.

[0007] (1) An antisense oligonucleotide consisting of 15 to 30 nucleotides, which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of the 5'UTR region, nsp1 region, nsp3 region, nsp4 region, 3C-like proteinase region, nsp6 region, nsp7 region, nsp8 region, nsp9 region, nsp10 region, RNA-dependent RNA polymerase region, helicase region, 3'-to-5' exonuclease region, endoRNase region, 2'-O-ribose methyltransferase region, S region including S1 region and S2 region, ORF3a region, E region, M region, ORF7b region, N region, ORF10 region, and 3'UTR region in the genomic RNA of SARS-CoV-2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof; (1) An antisense oligonucleotide or a pharmaceutically acceptable salt thereof or a hydrate thereof, which has an antiviral effect against a virus selected from the group consisting of SARS-CoV-2, SARS-CoV-1, and MERS-CoV. (2) The antisense oligonucleotide or a pharmaceutically acceptable salt thereof or a hydrate thereof according to (1), wherein the antisense oligonucleotide is a gapmer comprising a central gap region and two wing regions flanking the gap region. (3) The antisense oligonucleotide or a pharmaceutically acceptable salt thereof or a hydrate thereof according to (1) or (2), wherein the virus is SARS-CoV-2 or SARS-CoV-1. (4) At least one target region is selected from the group consisting of the nsp1 region, nsp3 region, 3C-like proteinase region, nsp8 region, RNA-dependent RNA polymerase region, helicase region, 3'-to-5' exonuclease region, endoRNase region, 2'-O-ribose methyltransferase region, S region including the S1 region and the S2 region, M region, ORF7b region, N region, ORF10 region, and 3'UTR region in the genomic RNA of SARS-CoV-2;The antisense oligonucleotide according to any one of (1) to (3), or a pharmaceutically acceptable salt thereof, or a hydrate thereof. (5) The antisense oligonucleotide according to any one of (1) to (4), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein at least one target region is selected from the group consisting of the nsp3 region, the RNA-dependent RNA polymerase region, the helicase region, the 3'-to-5' exonuclease region, the endoRNase region, the 2'-O-ribose methyltransferase region, the S region including the S1 region and the S2 region, and the N region in the genomic RNA of SARS-CoV-2. (6) positions 43 to 89, 242 to 279, 290 to 312, 455 to 477, 704 to 723, 3352 to 3372, 5384 to 5403, 6071 to 6090, 6406 to 6427, 6797 to 6815, 7532 to 7551, 8707 to 8724, 10292 to 10309, 10406 ​​to 10431, 10484 to 10506 of SEQ ID NO: 1 11609-11630, 12023-12045, 12170-12188, 12212-12234, 12314-12339, 12401-12420, 12589- 12608th place, 12839th place - 12867th place, 12898th place - 12922nd place, 12965th place - 12990th place, 13151st - 13175th place, 13271st - 13290th place, 13363rd - 13386th place 13458th to 13502nd, 13642nd to 13661st, 13672nd to 13691st, 13762nd to 13790th, 13894th to 13916th, 14050th to 14069th, 142nd 90th - 14312nd, 14512 - 14531st, 14654th - 14687th, 14698 - 14717th, 14750th - 14777th, 14824th - 14846th, 14854 - 1 4873rd, 14878th to 14909th, 14953rd to 14990th, 14992nd to 15026th, 15037th to 15061st, 15063rd to 15140th, 15172nd to 151st 98th place, 15278th to 15299th, 15454th to 15479th, 15496th to 15518th, 15520th to 15539th, 15622nd to 15644th, 15791st to 15809th,15829th to 15859th, 15886th to 15905th, 15929th to 15950th, 15985th to 16011th, 1605 1st to 16085th, 16189th to 16220th, 16430th to 16451st, 16636th to 16655th, 16822nd to 16845th, 17015th to 17051st, 17080th to 17103rd, 17137th to 17156th, 17215th to 1723th 4th place, 17254th to 17277th, 17564th to 17600th, 17748th to 17765th, 17773th to 17792nd, 17 830-17849th, 17859-17876, 18094-18111, 18196-18218, 18253- 18278th, 18370-18387, 19568-19595, 19622-19639, 19780-19802 , 20107-20130th, 20776-20795th, 20797-20816th, 20888th-20909th, 2145 3rd to 21472nd, 21502nd to 21524th, 22550th to 22569th, 22814th to 22836th, 23093rd to 23rd 113th, 23956th to 23976th, 24302nd to 24324th, 24446th to 24465th, 24467th to 24489th 24620th to 24651st, 24662nd to 24684th, 24962nd to 24982nd, 25104th to 25128th, 2 5364th to 25384th, 25502 to 25520, 26287th to 26325th, 26574th to 26604th, 2678 2-26800th, 27093-27111, 27771-27794, 27806-27823, 28270-2829 complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 4, 28397 to 28418, 28509 to 28538, 28744 to 28784, 28799 to 28820, 28946 to 28972, 28986 to 29005, 29010 to 29031, 29102 to 29130, 29174 to 29196, 29354 to 29373, 29615 to 29634, 29712 to 29731, and 29787 to 29867;The antisense oligonucleotide according to any one of (1) to (5), or a pharmaceutically acceptable salt thereof or a hydrate thereof. (7) positions 47 to 66, 242 to 261, 259 to 278, 292 to 311, 456 to 475, 704 to 723, 3353 to 3372, 5384 to 5403, 6071 to 6090, 6406 to 6425, 6407 to 6426, 6408 to 6427, 6797 to 6815, 7532 to 7551, 8707 to 8724, 10292 to 10309, 10407 to 10426, 10409 to 10428, 10487 to 10506, 11609 to 11628, 12025 to 12044, 12170 to 12188, 12214 to 12233, 12316 to 12335, 12401 to 12420, 12589 to 12608, 12843 to 12862, 12900 to 12919, 12969 to 12988, 1�153 to 13172, 13271 to 13290, 13366 to 13385, 13462 to 13481, 13481 to 13500, 13642 to 13661, 13672 to 13691, 13767 to 13786, 13894 to 13913, 14050 to 14069, 14292 to 14311, 14512 to 14531, 14656 to 14675, 14668 to 14687, 14698 to 14717, 14753 to 14772, 14826 to 14845, 14854 to 14873, 14879 to 14898, 14889 to 14908, 14962 to 14981, 14971 to 14990, 14992 to 15011, 15007 to 15026, 15040 to 15059, 15064 to 15083, 15091 to 15107, 15092 to 15111, 15120 to 15139, 15172 to 15191, 15173 to 15192, 15174 to 15193, 15175 to 15194, 15179 to 15198, 15280 to 15299, 15280 to 1529, position 15457 to 15476, 15496 to 15515, 15497 to 15516, 15498 to 15517 of SEQ ID NO: 115499-15518, 15520-15539, 15623-15642, 15791-15809, 15834-15853, 15886-15905, 15931-15950, 15988-16007, 16052-16071 16066-16085, 16189-16208, 16192-16211, 16193-16212, 16194-16213, 16195-16214, 16196-16215, 16198-16217, 16201-162 20, 16432nd to 16451st, 16636th to 16655th, 16825th to 16844th, 17016th to 17035th, 17032nd to 17051st, 17080th to 17099th, 17083rd to 17102nd, 17084th to 17103rd, 17137th to 1 7156, 17215-17234, 17257-17276, 17564-17583, 17565-17584, 17566-17585, 17567-17586, 17569-17588, 17573-17592, 17576- 17595, 17580-17599, 17748-17765, 17773-17792, 17830-17849, 17859-17876, 18094-18111, 18197-18216, 18257-18276, 18370 18387, 19572-19591, 19622-19639, 19780-19799, 19783-19802, 20107-20126, 20108-20127, 20109-20128, 20110-20129, 201 11th to 20130th, 20776th to 20795th, 20797th to 20816th, 20889th to 20908th, 21453th to 21472nd, 21502th to 21521st, 21503th to 21522nd, 21504th to 21523rd, 22550th to 22569th, 2 2817th to 22836th, 23093rd to 23112th, 23957th to 23976th, 24303rd to 24322nd, 24446th to 24465th, 24469th to 24488th, 24620th to 24639th, 24632nd to 24651st, 24665th to 24684th,24962nd - 24981st, 25107th - 25126th, 25365th - 25384th, 25502nd - 25520th, 26287th - 26306th, 26305th ~26324th place, 26579th place - 26598th place, 26782nd place - 26800th place, 27093rd place - 27111th place, 27774th place - 27793rd place, 27806th place - 27823rd place , 28274th to 28293rd, 28398th to 28417th, 28514th to 28533rd, 28744th to 28763rd, 28745th to 28764th, 28746th ~28765th place, 28747th place - 28766th place, 28748th place - 28767th place, 28752nd place - 28771st place, 28756th place - 28775th place, 28757th place - 28776th place , 28758th to 28777th, 28759th to 28778th, 28762nd to 28781st, 28765th to 28784th, 28799th to 28818th, 28950th ~28969th place, 28986th place - 29005th place, 29010th place - 29029th place, 29106th place - 29125th place, 29176th place - 29195th place, 29354th place - 29373rd place The antisense oligonucleotide according to any one of (1) to (6), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 29615 to 29634, positions 29712 to 29731, and positions 29789 to 29808. (8) positions 704 to 723, 5384 to 5403, 6407 to 6426, 10409 to 10428, 10487 to 10506, 12170 to 12188, 12214 to 12233, 12316 to 12335, 12401 to 12420, 13642 to 13661, 13672 to 13691, 13767 to 13786, 13894 to 13913, 14050 of SEQ ID NO: 1 ~14069th place, 14512th place - 14531st place, 14656th place - 14675th place, 14668th place - 14687th place, 14753rd place - 14772nd place, 14826th place - 14845th place, 14879th place - 14898th place, 148 89th to 14908th, 14962nd to 14981st, 14992nd to 15011th, 15007th to 15026th, 15040th to 15059th, 15064th to 15083rd, 15091st to 15107th,15092-15111, 15120-15139, 15173-15192, 15175-15194, 15179-15198, 15280-15299, 15280-15296, 15457-15476, 15496-155 15, 15498-15517, 15791-15809, 15834-15853, 15886-15905, 15931-15950, 15988-16007, 16052-16071, 16066-16085, 16189- 16208, 16192-16211, 16195-16214, 16201-16220, 16432-16451, 16636-16655, 17016-17035, 17032-17051, 17083-17102, 1708 4th to 17103rd, 17257th to 17276th, 17565th to 17584th, 17576th to 17595th, 17580th to 17599th, 17748th to 17765th, 17773rd to 17792nd, 17830th to 17849th, 18094th to 18111th, 1 8197-18216, 18257-18276, 19622-19639, 19783-19802, 20108-20127, 20111-20130, 20776-20795, 20797-20816, 20889-2090 8, 21502-21521, 21504-21523, 24469-24488, 24620-24639, 24632-24651, 24665-24684, 24962-24981, 25107-25126, 26782-2 6800, 27806-27823, 28274-28293, 28514-28533, 28745-28764, 28746-28765, 28756-28775, 28757-28776, 28758-28777, 28759 Bits ~ 28778, 28765 ~ 28784, 28799 ~ 28818, 28950 ~ 28969, 29010 ~ 290 29 digits, 29106 digits to 29125 digits, 29176 digits to 29195 digits, 29354 digits to 29373 digits, and 29615 digits to 29634 digits,and 29712 to 29731. (9) The antisense oligonucleotide according to any one of (1) to (7), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 5384 to 5403, 6407 to 6426, 13642 to 13661, 13672 to 13691, 15173 to 15192, 15175 to 15194, 15179 to 15198, 15496 to 15515, 15498 to 15517, 161 89th to 16208th, 16192nd to 16211th, 16195th to 16214th, 16201st to 16220th, 17083rd to 17102nd, 17084th to 17103rd, 17565th to 17584th, 17576th to 17595th, 18257th to 18276th, 19783rd to 1980 The antisense oligonucleotide according to any one of (1) to (8), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 2, 20108 to 20127, 20111 to 20130, 21502 to 21521, 21504 to 21523, 24665 to 24684, 28745 to 28764, 28746 to 28765, 28756 to 28775, 28757 to 28776, 28758 to 28777, and 28759 to 28778. (10) The antisense oligonucleotide according to any one of (1) to (9), which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 6407 to 6426, 15173 to 15192, 15496 to 15515, 15498 to 15517, 16195 to 16214, 17084 to 17103, 20111 to 20130, 21502 to 21521, 28757 to 28776, and 28758 to 28777 of SEQ ID NO: 1, or a pharmaceutically acceptable salt thereof, or a hydrate thereof. (11) The antisense oligonucleotide according to any one of (1) to (9), which is complementary to a nucleic acid comprising at least 15 consecutive bases in a target region.The antisense oligonucleotide according to any one of (1) to (10), or a pharmaceutically acceptable salt thereof, or a hydrate thereof. (12) The antisense oligonucleotide according to any one of (1) to (11), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which comprises (a) a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177, (b) a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177 in which one or several bases have been added, deleted, or substituted, or (c) a base sequence having 80% or more sequence identity to a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177, and which inhibits the function of the target region. (13) (a) a base sequence selected from the group consisting of SEQ ID NOs: 7, 9, 12, 19, 20, 23-26, 36-40, 42-44, 46, 47, 49-51, 53-59, 61, 63-68, 70, 74-82, 85, 88-90, 92, 93, 95, 96, 99, 101, 106-110, 112-114, 117, 119, 121, 124, 125-127, 129, 131, 138-143, 149, 152, 153, 155, 157, 158, 162-165, 167-169, and 171-176; (b) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 7, 9, 12, 19, 20, 23-26, 36-40, 42-44, 46, 47, 49-51, 53-59, 61, 63-68, 70, 74-82, 85, 88-90, 92, 93, 95, 96, 99, 101, 106-110, 112-114, 117, 119, 121, 124, 125-127, 129, 131, 138-143, 149, 152, 153, 155, 157, 158, 162-165, 167-169, and 171-176, in which one or several nucleotides have been added, deleted, or substituted; or (c) SEQ ID NOs: 7, 9, 12, 19, 20, 23-26, 36-40, 42-44, 46, 47, 49-51, 53-59, 61, 63-68, 70, 74-82, 85, 88-90, 92, 93, 95, 96, 99, 101, 106-110, 112-114, 117, 119, 121, 124, 125-127, 129, 131, 138-143, 149, 152, 153, 155, 157, 158, 162-165, 167-169,and 171 to 176, and the antisense oligonucleotide or a pharmaceutically acceptable salt thereof, or a hydrate thereof, according to any one of (1) to (12), which comprises a base sequence having 80% or more sequence identity to a base sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165; (b) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165, in which one or several nucleotides have been added, deleted, or substituted; or (c) The antisense oligonucleotide according to any one of (1) to (13), or a pharmaceutically acceptable salt thereof, or a hydrate of the same, which comprises a base sequence having 80% or more sequence identity to a base sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165, and inhibits the function of the target region. (15) The antisense oligonucleotide according to any one of (1) to (14), which comprises: (a) a base sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164; (b) a base sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164 in which one or several bases have been added, deleted, or substituted; or (c) a base sequence having 80% or more sequence identity to a base sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164, and which inhibits the function of the target region, or a pharmaceutically acceptable salt thereof, or a hydrate thereof. (16) The antisense oligonucleotide according to any one of (1) to (14), which comprises the sequence of (a),The antisense oligonucleotide according to any one of (11) to (15), or a pharmaceutically acceptable salt thereof, or a hydrate thereof. (17-1) The antisense oligonucleotide according to any one of (1) to (16), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein the antisense oligonucleotide consists of 20 nucleotides. (17-2) The antisense oligonucleotide according to (17-1), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein the antisense oligonucleotide is a gapmer composed of, from the 5' side to the 3' side, a wing region 5 nucleotides in length, a gap region 10 nucleotides in length, and a wing region 10 nucleotides in length. (18) The antisense oligonucleotide according to any one of (1) to (17-2), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein the wing region contains a 2'-OMe (2'-O-CH3) group and / or a 2'-O-MOE (2'-O-CH2CH2OCH3) group. (19) The antisense oligonucleotide according to (17-1), (17-2), or (18), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein the bond between the second and third nucleosides and the bond between the fourth and fifth nucleosides from the 5' side of the 5' wing region are phosphodiester bonds, and the bond between the first and second nucleosides and the bond between the third and fourth nucleosides from the 5' side of the 3' wing region are phosphodiester bonds, and all other internucleoside bonds are phosphorothioate bonds. (20) (17-1), (17-2), in which the bond between the second and third nucleosides from the 5' side, the bond between the third and fourth nucleosides, and the bond between the fourth and fifth nucleosides from the 5' side in the 5' wing region are phosphodiester bonds, and the bond between the first and second nucleosides from the 5' side in the 3' wing region, the bond between the second and third nucleosides, and the bond between the third and fourth nucleosides from the 5' side are phosphodiester bonds, and all other internucleoside bonds are phosphorothioate bonds.or the antisense oligonucleotide according to (18), or a pharmaceutically acceptable salt thereof, or a hydrate thereof. (21) A pharmaceutical composition comprising the antisense oligonucleotide according to any one of (1) to (20), or a pharmaceutically acceptable salt thereof, or a hydrate thereof. (22) The pharmaceutical composition according to (21), for treating and / or preventing a viral infection selected from the group consisting of SARS-CoV-2, SARS-CoV-1, and MERS-CoV. (23) A method for treating and / or preventing a viral infection selected from the group consisting of SARS-CoV-2, SARS-CoV-1, and MERS-CoV, comprising a step of administering to a subject an effective amount of the nucleic acid according to any one of (1) to (20), or a pharmaceutically acceptable salt thereof, or a hydrate thereof, or the pharmaceutical composition according to any one of (21) to (22).

[0008] The present invention provides an antisense oligonucleotide that targets the genomic RNA of SARS-CoV-2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, as well as a composition comprising the antisense oligonucleotide, or a pharmaceutically acceptable salt thereof, or a hydrate thereof.

[0009] According to a preferred embodiment of the present invention, the antisense oligonucleotides of the present invention can provide a therapeutic and / or prophylactic agent for viruses that has a high viral growth inhibitory effect and / or few side effects.Furthermore, according to a preferred embodiment of the present invention, the antisense oligonucleotides of the present invention can also be effective against known SARS-related coronaviruses (SARSr-CoV) such as SARS-CoV-2 mutants and / or SARS-CoV-1 and / or unknown SARS-related coronaviruses.

[0010] In one embodiment, the present invention relates to an antisense oligonucleotide or a pharmaceutically acceptable salt thereof or a hydrate thereof having an antiviral effect against SARS-CoV-2, SARS-CoV-1, or MERS-CoV (hereinafter, the antisense oligonucleotide of the present invention or a pharmaceutically acceptable salt thereof or a hydrate thereof will also be collectively referred to as the "antisense oligonucleotide of the present invention").

[0011] In one embodiment, the antisense oligonucleotides of the invention consist of 15 to 30 nucleotides that are complementary to a nucleic acid comprising at least 10 consecutive bases in the target region.

[0012] In one embodiment, the antisense oligonucleotides of the invention are complementary to a nucleic acid comprising or consisting of at least 10, e.g., at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, e.g., 20, consecutive bases in the target region.

[0013] As used herein, an antisense oligonucleotide that is "complementary" to a nucleic acid is not limited to an antisense oligonucleotide that forms a Watson-Crick base pair with the target nucleic acid, but also includes an antisense oligonucleotide that forms a wobble base pair. Here, Watson-Crick base pairs refer to base pairs in which hydrogen bonds are formed between adenine-thymine, adenine-uracil, and guanine-cytosine, and wobble base pairs refer to base pairs in which hydrogen bonds are formed between guanine-uracil, inosine-uracil, inosine-adenine, and inosine-cytosine. Furthermore, a "complementary base sequence" does not necessarily have to be 100% complementary to the target base sequence. For example, the "complementary base sequence" may contain one, two, three, four, or five non-complementary bases relative to the target base sequence. Alternatively, the "complementary base sequence" may be one, two, three, four, or five bases shorter than the target base sequence. In one embodiment, an antisense oligonucleotide that is "complementary" to a nucleic acid has at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 100% complementarity to the nucleic acid. Complementarity can be easily determined by one of ordinary skill in the art, for example, by aligning two sequences, counting the number of bases that form Watson-Crick base pairs or wobble base pairs between the sequences, dividing the number of base-paired bases by the total number of bases in the sequences, and multiplying this by 100.

[0014] An example of an antisense oligonucleotide that is "complementary" to a certain nucleic acid is an antisense oligonucleotide that can hybridize to that nucleic acid, for example, under stringent conditions. As used herein, "stringent conditions" may refer to low stringency conditions, moderate stringency conditions, or high stringency conditions. "Low stringency conditions" are, for example, 5x SSC, 5x Denhardt's solution, 0.5% SDS, 50% formamide, and 32°C. "Medium stringency conditions" are, for example, 5x SSC, 5x Denhardt's solution, 0.5% SDS, 50% formamide, and 42°C, or 5x SSC, 1% SDS, 50 mM Tris-HCl (pH 7.5), 50% formamide, and 42°C. "Highly stringent conditions" are, for example, conditions of 5x SSC, 5x Denhardt's solution, 0.5% SDS, 50% formamide, and 50°C, or 0.2x SSC, 0.1% SDS, and 65°C. Under these conditions, it is expected that the higher the temperature, the more efficiently base sequences with higher sequence identity can be obtained. However, several factors are thought to affect the stringency of hybridization, such as temperature, probe concentration, probe length, ionic strength, time, and salt concentration, and those skilled in the art can achieve similar stringency by appropriately selecting these factors.

[0015] When using a commercially available hybridization kit, for example, the AlkPhos Direct Labeling and Detection System (GE Healthcare) can be used. In this case, hybridization can be detected by incubating the membrane with the labeled probe overnight according to the protocol provided with the kit, washing the membrane with a primary wash buffer containing 0.1% (w / v) SDS at 55°C, and then detecting hybridization. Alternatively, when preparing a probe based on a target sequence, if the probe is labeled with digoxigenin (DIG) using a commercially available reagent (e.g., PCR Labeling Mix (Roche Diagnostics)), hybridization can be detected using a DIG Nucleic Acid Detection Kit (Roche Diagnostics), etc.

[0016] The identity of nucleotide sequences can be determined using the BLAST (Basic Local Alignment Search Tool) algorithm by Carlin and Altschul (Proc. Natl. Acad. Sci. USA 872264-2268, 1990; Proc Natl Acad Sci USA 90: 5873, 1993). Programs based on the BLAST algorithm, such as BLASTN and BLASTX, have been developed (Altschul SF, et al: J Mol Biol 215: 403, 1990). When analyzing nucleotide sequences using BLASTN, the parameters are set to, for example, score = 100 and wordlength = 12. When using BLAST and Gapped BLAST programs, the default parameters of each program are used.

[0017] In one embodiment, the antisense oligonucleotides of the invention may be, for example, 15 or more nucleotides in length, 16 or more nucleotides in length, 17 or more nucleotides in length, 18 or more nucleotides in length, 19 or more nucleotides in length, 20 or more nucleotides in length, 21 or more nucleotides in length, 22 or more nucleotides in length, 23 or more nucleotides in length, 24 or more nucleotides in length, 25 or more nucleotides in length, 26 or more nucleotides in length, 27 or more nucleotides in length, 28 or more nucleotides in length, 29 or more nucleotides in length, or 30 nucleotides in length, or 30 or less nucleotides in length, 29 or less nucleotides in length, 28 or less nucleotides in length, 27 or less nucleotides in length, 26 or less nucleotides in length, 25 or less nucleotides in length, 24 or less nucleotides in length, 23 or less nucleotides in length, 22 or less nucleotides in length, 21 or less nucleotides in length, 20 or less nucleotides in length, 19 or less nucleotides in length, 18 or less nucleotides in length, 17 or less nucleotides in length, 16 or less nucleotides in length, or 15 nucleotides in length. Antisense oligonucleotides of the invention may consist of, for example, 15 to 30 nucleotides, 15 to 25 nucleotides, 16 to 24 nucleotides, 17 to 23 nucleotides, 18 to 22 nucleotides, 19 to 21 nucleotides, eg, 20 nucleotides.

[0018] Examples of pharmaceutically acceptable salts of the antisense oligonucleotides of the present invention include alkali metal salts such as sodium salt, potassium salt, and lithium salt, and alkaline earth metal salts such as calcium salt and magnesium salt; metal salts such as aluminum salt, iron salt, zinc salt, copper salt, nickel salt, and cobalt salt; ammonium salt; t-octylamine salt, dibenzylamine salt, morpholine salt, glucosamine salt, phenylglycine alkyl ester salt, ethylenediamine salt, N-methylglucamine salt, guanidine salt, diethylamine salt, triethylamine salt, dicyclohexylamine salt, N,N'-dibenzylethylenediamine salt, chloroprocaine salt, procaine salt, diethanolamine salt, N-benzyl-phenethylamine salt, Examples of suitable salts include organic amine salts such as piperazine salts, tetramethylammonium salts, and tris(hydroxymethyl)aminomethane salts; hydrohalide salts such as hydrofluoride, hydrochloride, hydrobromide, and hydroiodide; inorganic acid salts such as nitrate, perchlorate, sulfate, and phosphate; lower alkane sulfonate salts such as methanesulfonate, trifluoromethanesulfonate, and ethanesulfonate; arylsulfonate salts such as benzenesulfonate and p-toluenesulfonate; organic acid salts such as acetate, malate, fumarate, succinate, citrate, tartrate, oxalate, and maleate; and amino acid salts such as glycine salt, lysine salt, arginine salt, ornithine salt, glutamate, and aspartate. These salts can be prepared by known methods. Alternatively, the antisense oligonucleotides of the present invention may be in the form of their hydrates.

[0019] The antisense oligonucleotides of the present invention are composed of nucleotides as building blocks, and such nucleotides may be ribonucleotides, deoxyribonucleotides, or modified nucleotides.

[0020] A modified nucleotide refers to a ribonucleotide or deoxyribonucleotide in which all or part of the nucleic acid base, sugar moiety, and phosphate linkage moiety that constitute the ribonucleotide or deoxyribonucleotide have been modified.

[0021] Examples of nucleic acid bases include adenine, guanine, hypoxanthine, cytosine, thymine, uracil, and modified bases thereof. Examples of such modified bases include pseudouracil, 3-methyluracil, dihydrouracil, 5-alkylcytosine (e.g., 5-methylcytosine), 5-alkyluracil (e.g., 5-ethyluracil), 5-halouracil (e.g., 5-bromouracil), 6-azapyrimidine, 6-alkylpyrimidine (e.g., 6-methyluracil), 2-thiouracil, 4-thiouracil, 4-acetylcytosine, 5-(carboxyhydroxymethyl)uracil, 5-carboxymethylaminomethyl-2-thiouracil, 5-carboxymethylaminomethyluracil, 1-methyladenine, 1-methylhypoxanthine, 1-methyluracil, 1-methylhydroxy ... Examples of amino acids include xanthine, 2,2-dimethylguanine, 3-methylcytosine, 2-methyladenine, 2-methylguanine, N6-methyladenine, 7-methylguanine, 5-methoxyaminomethyl-2-thiouracil, 5-methylaminomethyluracil, 5-methylcarbonylmethyluracil, 5-methyloxyuracil, 5-methyl-2-thiouracil, 2-methylthio-N6-isopentenyladenine, uracil-5-oxyacetic acid, 2-thiocytosine, purine, 2,6-diaminopurine, 2-aminopurine, isoguanine, indole, imidazole, and xanthine.

[0022] As used herein, thymine "T" and uracil "U" are interchangeable, and since the activity of the antisense oligonucleotide of the present invention is not essentially affected whether "T" or "U" is used, the base sequences shown herein include cases where "T" is replaced with "U," and are represented by the same SEQ ID NO. Also, as used herein, sequences containing modified bases are represented by the same SEQ ID NO as sequences not containing modified bases; for example, "cytosine" and "methylcytosine" are interchangeable, and when "cytosine" is replaced with "methylcytosine," the same SEQ ID NO is used.

[0023] Modifications of the sugar moiety include, for example, modifications of the 2'-position of ribose and modifications of other parts of the sugar. Modifications of the 2'-position of ribose include, for example, replacing the -OH group at the 2'-position of ribose with -OR, -OROR, -R, -R'OR, -SH, -SR, -NH ２ , -NHR, -NR ２ , -N ３ , —CN, —F, —Cl, —Br, —I, for example, —OMe(—O—CH ３ ) or -O-methoxyethyl (-O-MOE: -O-CH ２ CH ２ OCH ３ ) where R represents alkyl, cycloalkyl, acyl, or aryl; and R' represents alkylene.

[0024] Modifications of other sugar moieties include, but are not limited to, substitution of the O at the 4' position of ribose or deoxyribose with S, and cross-linking of the 2' and 4' positions of the sugar, such as LNA (Locked Nucleic Acid) or ENA (2'-O,4'-C-Ethylene-bridged Nucleic Acids).

[0025] Modifications of the phosphate linkage moiety include, for example, substitution of a phosphodiester bond with a phosphorothioate bond, a phosphorodithioate bond, an alkylphosphonate bond, a phosphoramidate bond, or a boranophosphate bond (see, for example, Enya et al.: Bioorganic & Medicinal Chemistry, 2008, 18, 9154-9160) (see, for example, Republished Patent Publication No. 2006 / 129594 and Republished Patent Publication No. 2006 / 038608).

[0026] In this specification, the alkyl is preferably a linear or branched alkyl having 1 to 6 carbon atoms. Specific examples include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, tert-pentyl, n-hexyl, and isohexyl. The alkyl may be substituted, and examples of such substituents include halogen, alkoxy, cyano, and nitro, and the alkyl may be substituted with 1 to 3 of these.

[0027] In this specification, the cycloalkyl is preferably a cycloalkyl having 3 to 12 carbon atoms. Specific examples include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclodecyl, and cyclododecyl.

[0028] In this specification, halogen includes fluorine, chlorine, bromine, and iodine.

[0029] In this specification, examples of alkoxy include straight-chain or branched-chain alkoxy having 1 to 6 carbon atoms, such as methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, tert-butoxy, n-pentyloxy, isopentyloxy, n-hexyloxy, isohexyloxy, etc. In particular, alkoxy having 1 to 3 carbon atoms is preferred.

[0030] In this specification, the aryl is preferably an aryl having 6 to 10 carbon atoms. Specific examples include phenyl, α-naphthyl, and β-naphthyl. Phenyl is particularly preferred. The aryl may be substituted, and examples of such substituents include alkyl, halogen, alkoxy, cyano, and nitro, and the aryl may be substituted with 1 to 3 of these.

[0031] In this specification, the alkylene is preferably a linear or branched alkylene having 1 to 6 carbon atoms, such as methylene, ethylene, trimethylene, tetramethylene, pentamethylene, hexamethylene, 2-(ethyl)trimethylene, and 1-(methyl)tetramethylene.

[0032] As used herein, acyl includes straight-chain or branched-chain alkanoyl or aroyl. Examples of alkanoyl include formyl, acetyl, 2-methylacetyl, 2,2-dimethylacetyl, propionyl, butyryl, isobutyryl, pentanoyl, 2,2-dimethylpropionyl, and hexanoyl. Examples of aroyl include benzoyl, toluoyl, and naphthoyl. Such aroyl may be substituted at any substitutable position, and may be substituted with alkyl.

[0033] In one embodiment, the present invention provides a gapmer antisense oligonucleotide comprising a central gap region and two wing regions (also referred to as the 5' wing region and the 3' wing region, respectively) flanking the 5'- and 3'-ends of the gap region. The gap region is a region recognized by RNase H and is composed of deoxyribonucleotides with unmodified sugar moieties. The wing regions contain at least one modified nucleotide, for example, all of which are modified nucleotides (e.g., ribonucleotides modified at the 2'-position of the ribose). In one embodiment, the nucleosides in the 5' wing region and the 3' wing region each contain at least one, e.g., two or more, three or more, four or more, or five or more sugar moiety modifications, such as 2'-OMe and / or 2'-O-MOE groups. For example, all of the nucleosides in the 5' wing region and the 3' wing region may contain 2'-OMe and / or 2'-O-MOE groups. Additionally, the nucleosides of the 5' and 3' wing regions may contain modifications in the base moiety, such as at least one methylcytosine.

[0034] The length of the gap region is not limited, but may be, for example, 5 to 15, 8 to 12, 9 to 11, or 10 bases long. The lengths of the 5' wing region and the 3' wing region are not limited, but may be, for example, independently 2 to 10, 3 to 8, 4 to 6, or 5 bases long. Herein, the structure of a gapmer may be represented as "abc type." "abc type" means that the gapmer is composed of, from the 5' side to the 3' side, a wing region of nucleotide length a, a gap region of nucleotide length b, and a wing region of nucleotide length c. For example, the gapmer may be a 5-10-5 type, a 4-10-5 type, a 5-10-4 type, a 4-10-4 type, a 5-9-4 type, or a 4-9-4 type gapmer.

[0035] In one embodiment, a gapmer of the invention comprises one or more phosphate linkage modifications, e.g., phosphorothioate linkages, and for example, one or more, two or more, three or more, four or more, five or more, ten or more, fifteen or more, e.g., all, of the internucleotide linkages may be phosphorothioate linkages. In one embodiment, a gapmer of the invention comprises one or more, two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, nine or more, or ten or more (e.g., four to six) phosphodiester linkages, e.g., in the 5' wing region and / or the 3' wing region, and all other internucleoside linkages are phosphorothioate linkages. In one embodiment, the bond between the second and third nucleosides and the bond between the fourth and fifth nucleosides from the 5' side of the 5' wing region are phosphodiester bonds, and the bond between the first and second nucleosides and the third and fourth nucleosides from the 5' side of the 3' wing region are phosphodiester bonds, and all other internucleoside bonds are phosphorothioate bonds. In one embodiment, the bond between the second and third nucleosides, the bond between the third and fourth nucleosides, and the bond between the fourth and fifth nucleosides from the 5' side of the 5' wing region are phosphodiester bonds, and the bond between the first and second nucleosides, the bond between the second and third nucleosides, and the bond between the third and fourth nucleosides from the 5' side of the 3' wing region are phosphodiester bonds, and all other internucleoside bonds are phosphorothioate bonds. In one embodiment, a gapmer of the invention does not include modifications of phosphate linkages and all of the internucleotide linkages may be phosphate linkages.

[0036] In one embodiment, the antisense oligonucleotides of the present invention have an antiviral effect against SARS-CoV-2, SARS-CoV-1, or MERS-CoV, e.g., SARS-CoV-2 or SARS-CoV-1, e.g., SARS-CoV-2. As used herein, SARS-CoV-2 refers to a virus having a genomic RNA sequence consisting of the nucleotide sequence NC_045512.2 (SEQ ID NO: 1) or a mutant thereof. SARS-CoV-1 refers to a virus having a genomic RNA sequence consisting of the nucleotide sequence NC_004718.3 or a mutant thereof. MERS-CoV refers to a virus having a genomic RNA sequence consisting of the nucleotide sequence NC_019843.3 or a mutant thereof.

[0037] Here, a mutant strain refers to an offspring with new properties that arise from a mutation that partially changes the genetic information due to misreading or recombination that occurs during the replication process of viral genes. Mutant strains have some altered properties influenced by the altered genetic information, but the original virus species remains unchanged. In the Coronaviridae family, viruses are considered to belong to the same virus species if they share more than 90% of the amino acid sequence of the conserved replicase region (see the entries for Nidovirales and Coronaviridae in the ICTV 9th report (2011) (https: / / talk.ictvonline.org / ictv-reports / ictv_9th_report / positive-sense-rna-viruses-2011 / w / posrna_viruses / 222 / coronaviridae)).

[0038] In one embodiment, the antisense oligonucleotides of the present invention are effective against not only known SARS-associated coronaviruses (SARSr-CoVs) but also unknown SARS-associated coronaviruses. As used herein, SARS-associated coronaviruses refer to enveloped, single-stranded, positive-stranded RNA viruses belonging to the Betacoronavirus genus (group 2 coronaviruses), which infect mammals including humans and bats. SARS-associated coronaviruses use the angiotensin-converting enzyme 2 (ACE2) receptor to enter cells.

[0039] In one embodiment, the term "antiviral effect" refers to the effect of inhibiting viral proliferation and / or reducing viral infectivity. Whether or not the antisense oligonucleotides of the present invention have an antiviral effect can be tested as described in the Examples section of the present specification. For example, the presence or absence of an antiviral effect can be measured by constructing a plasmid expressing a nucleic acid containing a target sequence, introducing the plasmid and the antisense oligonucleotide of the present invention into cells, and determining whether or not the amount of nucleic acid containing the target sequence expressed from the cells is reduced (e.g., by 5% or more, 10% or more, or 20% or more) compared to when a negative control nucleic acid is introduced. Alternatively, the presence or absence of an antiviral effect can be measured by introducing a virus and the antisense oligonucleotide of the present invention into cells, culturing the cells, and then determining whether or not the amount of virus or nucleic acid derived therefrom in the cells or cell culture supernatant is reduced (e.g., by 5% or more, 10% or more, or 20% or more) compared to when a negative control nucleic acid is introduced.

[0040] In one embodiment, the antisense oligonucleotides of the invention, when tested as described in Example 1 using a SARS-CoV-2 expression plasmid, have a knockdown activity of 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, or 90% or more at either the 5' or 3' side of the Fluc-corrected gapmer, or have the same target sequence as an antisense oligonucleotide with such knockdown activity.

[0041] In one embodiment, the antisense oligonucleotide of the present invention has a knockdown activity of 5% or more, 10% or more, 20% or more, 30% or more, 40% or more, 50% or more, 60% or more, 70% or more, 80% or more, or 90% or more at a concentration of 2, 3, 6, 8, or 10 μM in gapmer knockdown activity assay using SARS-CoV-2 virus when tested as described in Example 2, or has the same target sequence as an antisense oligonucleotide having such knockdown activity.

[0042] The antisense oligonucleotides of the present invention target a sequence in at least one target region selected from the group consisting of the 5'UTR region, nsp1 region, nsp3 region, nsp4 region, 3C-like proteinase region, nsp6 region, nsp7 region, nsp8 region, nsp9 region, nsp10 region, RNA-dependent RNA polymerase region, helicase region, 3'-to-5' exonuclease region, endoRNase region, 2'-O-ribose methyltransferase region, S region including S1 region and S2 region, ORF3a region, E region, M region, ORF7b region, N region, ORF10 region, and 3'UTR region in the genomic RNA of SARS-CoV-2.

[0043] The sequence of the SARS-CoV-2 genomic RNA and the sequence of each region in the genomic RNA can be easily determined by referring to a database (e.g., ncbi.nlm.nih.gov / ). For example, the sequence of the SARS-CoV-2 genomic RNA may be the nucleotide sequence of NC_045512.2 (SEQ ID NO: 1).In addition, in the base sequence of SEQ ID NO: 1, the 5'UTR region is the base sequence of positions 1 to 265 of SEQ ID NO: 1, the nsp1 region is the base sequence of positions 266 to 805 of SEQ ID NO: 1, the nsp3 region is the base sequence of positions 2720 to 8554 of SEQ ID NO: 1, the nsp4 region is the base sequence of positions 8555 to 10054 of SEQ ID NO: 1, the 3C-like The proteinase region is the nucleotide sequence of positions 10055 to 10972 in SEQ ID NO: 1, the nsp6 region is the nucleotide sequence of positions 10973 to 11842 in SEQ ID NO: 1, the nsp7 region is the nucleotide sequence of positions 11843 to 12091 in SEQ ID NO: 1, the nsp8 region is the nucleotide sequence of positions 12092 to 12685 in SEQ ID NO: 1, the nsp9 region is the nucleotide sequence of positions 12686 to 13024 in SEQ ID NO: 1, the nsp10 region is the nucleotide sequence of positions 13025 to 13441 in SEQ ID NO: 1, the RNA-dependent RNA polymerase region is the nucleotide sequence of positions 13442 to 16236 in SEQ ID NO: 1, the helicase region is the nucleotide sequence of positions 16237 to 18039 in SEQ ID NO: 1, and the 3'-to-5' The exonuclease region is the nucleotide sequence of positions 18040 to 19620 in SEQ ID NO: 1, the endoRNase region is the nucleotide sequence of positions 19621 to 20658 in SEQ ID NO: 1, the 2'-O-ribose methyltransferase region is the nucleotide sequence of positions 20659 to 21552 in SEQ ID NO: 1, the S region is the nucleotide sequence of positions 21563 to 25384 in SEQ ID NO: 1 (the S1 region is the nucleotide sequence of positions 21599 to 23617 in SEQ ID NO: 1, the S2 region is the nucleotide sequence of positions 23618 to 25381 in SEQ ID NO: 1), the ORF3a region is the nucleotide sequence of positions 25393 to 26220 in SEQ ID NO: 1, the E region is the nucleotide sequence of positions 262 the M region may be the nucleotide sequence of positions 26523 to 27191 of SEQ ID NO: 1; the ORF7b region may be the nucleotide sequence of positions 27756 to 27887 of SEQ ID NO: 1; the N region may be the nucleotide sequence of positions 28274 to 29533 of SEQ ID NO: 1; the ORF10 region may be the nucleotide sequence of positions 29558 to 29674 of SEQ ID NO: 1; and the 3'UTR region may be the nucleotide sequence of positions 29675 to 29903 of SEQ ID NO: 1.

[0044] In one embodiment, the antisense oligonucleotides of the present invention target regions conserved in the sequences of the SARS-CoV-2 genomic RNA and the SARS-CoV-1 genomic RNA, and therefore may have an antiviral effect against the Betacoronavirus genus, including SARS-CoV-2, SARS-CoV-1, and MERS-CoV. In one embodiment, the antisense oligonucleotides of the present invention target regions conserved in the sequences of the SARS-CoV-2 genomic RNA and the SARS-CoV-1 genomic RNA, among the above target regions, for example, positions 43 to 89, 242 to 279, 290 to 312, 455 to 477, 704 to 723, 3352 to 3372, 5384 to 5403, 6071 to 6090, and 6406 to 6409 of SEQ ID NO: 1. 27th, 6797-6815, 7532-7551, 8707-8724, 10292-10309, 10406-10431, 10484-10506, 11609-11630, 12023-12045, 1 2170-12188th, 12212-12234, 12314-12339, 12401-12420, 12589-12608, 12839-12867, 12898-12922, 12965-12990, 13151-13175, 13271-13290, 13363-13386, 13458-13502, 13642-13661, 13672-13691, 13762-13790, 13894-13916 14050-14069, 14290-14312, 14512-14531, 14654-14687, 14698-14717, 14750-14777, 14824-14846, 14854-148 73rd, 14878th to 14909th, 14953rd to 14990th, 14992nd to 15026th, 15037th to 15061st, 15063rd to 15140th, 15172nd to 15198th, 15278th to 15299th, 154 54th to 15479th, 15496th to 15518th, 15520th to 15539th, 15622nd to 15644th, 15791st to 15809th, 15829th to 15859th, 15886th to 15905th, 15929th to 15950th,15985-16011, 16051-16085, 16189-16220, 16430-16451, 16636-16655, 16822-16845, 17015-17051, 17080-17103, 1 7137-17156, 17215-17234, 17254-17277, 17564-17600, 17748-17765, 17773-17792, 17830-17849, 17859-17876, 18094-1 8111st, 18196th - 18218th, 18253rd - 18278th, 18370th - 18387th, 19568th - 19595th, 19622nd - 19639th, 19780 - 19802nd, 20107th - 20130th, 20776th - 20795th , 20797-20816, 20888-20909, 21453-21472, 21502-21524, 22550-22569, 22814-22836, 23093-23113, 23956-23976, 24 302nd to 24324th, 24446 to 24465, 24467 to 24489, 24620 to 24651, 24662 to 24684, 24962 to 24982, 25104 to 25128, 25364 to 25384, 25 502-25520, 26287-26325, 26574-26604, 26782-26800, 27093-27111, 27771-27794, 27806-27823, 28270-28294, 28397- It is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 28418, 28509 to 28538, 28744 to 28784, 28799 to 28820, 28946 to 28972, 28986 to 29005, 29010 to 29031, 29102 to 29130, 29174 to 29196, 29354 to 29373, 29615 to 29634, 29712 to 29731, and 29787 to 29867.

[0045] In one embodiment, the antisense oligonucleotide of the present invention is located at positions 47 to 66, 242 to 261, 259 to 278, 292 to 311, 456 to 475, 704 to 723, 3353 to 3372, 5384 to 5403, 6071 to 6090, 6406 to 6425, 6407 to 6426, 6408 to 6427, 6797 to 6815, 7532 to 7551, 8707 to 8724, 10292 to 10309, 10407 to 10426, 10409 to 10428, 10487 to 10506, 11609 to 11628, 12025 to 12044, 12170 to 12188, 12214 to 12233, 12316 to 12335, 12401 to 12420, 12589 to 12608, 12843 to 12862, 12900 to 12919, 12969 to 12988, 13153 to 13172, 13271 to 13290, 13366 to 13385, 13462 to 13481, 13481 to 13500, 13642 to 13661, 13672 to 13691, 13767 to 13786, 13894 to 13913, 14050 to 14069, 14292 to 14311, 14512 to 14531, 14656 to 14675, 14668 to 14687, 14698 to 14717, 14753 to 14772, 14826 to 14845, 14854 to 14873, 14879 to 14898, 14889 to 14908, 14962 to 14981, 14971 to 14990, 14992 to 15011, 15007 to 15026, 15040 to 15059, 15064 to 15083, 15091 to 15107, 15092 to 15111, 15120 to 15139, 15172 to 15191, 15173 to 15192, 15174 to 15193, 15175 to 15194, 15179 to 15198, 15280 to 15299, 15280 to 15296, 15457 to 15476, 15496 to 15515, 15497 to 15516, 15498 to 15517, 15499 to 15518, 15520 to 15539 of SEQ ID NO: 1.15623-15642, 15791-15809, 15834-15853, 15886-15905, 15931-15950, 15988-16007, 16052-16071, 16066-16085, 16189-16208 16192-16211, 16193-16212, 16194-16213, 16195-16214, 16196-16215, 16198-16217, 16201-16220, 16432-16451, 16636-166 55, 16825-16844, 17016-17035, 17032-17051, 17080-17099, 17083-17102, 17084-17103, 17137-17156, 17215-17234, 17257-1 7276, 17564-17583, 17565-17584, 17566-17585, 17567-17586, 17569-17588, 17573-17592, 17576-17595, 17580-17599, 17748- 17765, 17773-17792, 17830-17849, 17859-17876, 18094-18111, 18197-18216, 18257-18276, 18370-18387, 19572-19591, 19622 19639, 19780-19799, 19783-19802, 20107-20126, 20108-20127, 20109-20128, 20110-20129, 20111-20130, 20776-20795, 207 97th to 20816th, 20889th to 20908th, 21453rd to 21472nd, 21502nd to 21521st, 21503rd to 21522nd, 21504th to 21523rd, 22550th to 22569th, 22817th to 22836th, 23093rd to 23112th, 2 3957th to 23976th, 24303rd to 24322nd, 24446th to 24465th, 24469th to 24488th, 24620th to 24639th, 24632nd to 24651st, 24665th to 24684th, 24962nd to 24981st, 25107th to 25126th,25365th to 25384th, 25502nd to 25520th, 26287th to 26306th, 26305th to 26324th, 26579th to 265th 98th place, 26782nd to 26800th, 27093rd to 27111th, 27774th to 27793rd, 27806th to 27823rd, 28274th ~28293rd place, 28398th place - 28417th place, 28514th place - 28533rd place, 28744th place - 28763rd place, 28745th place - 28764th place, 28 746th to 28765th, 28747th to 28766th, 28748th to 28767th, 28752nd to 28771st, 28756th to 28775th , 28757 to 28776, 28758 to 28777, 28759 to 28778, 28762 to 28781, 28765 to 28784, 28799 to 28818, 28950 to 28969, 28986 to 29005, 29010 to 29029, 29106 to 29125, 29176 to 29195, 29354 to 29373, 29615 to 29634, 29712 to 29731, and 29789 to 29808.

[0046] In one embodiment, the antisense oligonucleotide of the present invention comprises or consists of any of the following sequences: (a) a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177, (b) a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177 in which one or more bases have been added, deleted, or substituted, or (c) a base sequence having 80%, 85% or more, 86% or more, 87% or more, 88% or more, 89% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more sequence identity to a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177. In one embodiment, the antisense oligonucleotide of the present invention comprises or consists of any of the base sequences of (a) above.

[0047] As used herein, "several" in a base sequence in which one or more bases have been added, deleted, or substituted means two, three, four, five, six, seven, eight, nine, or ten bases.

[0048] In one embodiment, at least one target region of the antisense oligonucleotide of the present invention is selected from the group consisting of the nsp1 region, nsp3 region, 3C-like proteinase region, nsp8 region, RNA-dependent RNA polymerase region, helicase region, 3'-to-5' exonuclease region, endoRNase region, 2'-O-ribose methyltransferase region, S region including S1 region and S2 region, M region, ORF7b region, N region, ORF10 region, and 3'UTR region in the genomic RNA of SARS-CoV-2. For example, the antisense oligonucleotides of the present invention may be used in combination with the oligonucleotides of SEQ ID NO: 1 at positions 704 to 723, 5384 to 5403, 6407 to 6426, 10409 to 10428, 10487 to 10506, 12170 to 12188, 12214 to 12233, 12316 to 12335, 12401 to 12420, 13642 to 13661, 13672 to 13691, 13767th to 13786th, 13894th to 13913th, 14050th to 14069th, 14512th to 14531st, 14656th to 14675th, 14668th to 14687th, 1475 3rd to 14772nd, 14826th to 14845th, 14879th to 14898th, 14889th to 14908th, 14962nd to 14981st, 14992nd to 15011th, 15007th to 15th 026th, 15040th to 15059th, 15064th to 15083rd, 15091st to 15107th, 15092nd to 15111th, 15120th to 15139th, 15173rd to 15192nd , 15175th to 15194th, 15179th to 15198th, 15280th to 15299th, 15280th to 15296th, 15457th to 15476th, 15496th to 15515th, 1549 8th to 15517th, 15791st to 15809th, 15834th to 15853rd, 15886th to 15905th, 15931st to 15950th, 15988th to 16007th, 16052nd to 16th 071st, 16066th to 16085th, 16189th to 16208th, 16192nd to 16211th, 16195th to 16214th, 16201st to 16220th, 16432nd to 16451st,16636th to 16655th, 17016th to 17035th, 17032nd to 17051st, 17083rd to 17102nd, 17084th to 17103rd, 17257th to 17276th, 17565th to 1 7584th, 17576th to 17595th, 17580th to 17599th, 17748th to 17765th, 17773rd to 17792nd, 17830th to 17849th, 18094th to 18111th, 181 97th to 18216th, 18257th to 18276th, 19622nd to 19639th, 19783rd to 19802nd, 20108th to 20127th, 20111th to 20130th, 20776th to 20795th 20797th - 20816th, 20889th - 20908th, 21502nd - 21521st, 21504th - 21523rd, 24469th - 24488th, 24620th - 24639th, 24632nd - 24651st, 24665th to 24684th, 24962nd to 24981st, 25107th to 25126th, 26782nd to 26800th, 27806th to 27823rd, 28274th to 28293rd, 2 8514th to 28533rd, 28745th to 28764th, 28746th to 28765th, 28756th to 28775th, 28757th to 28776th, 28758th to 28777th, 28759th to 287th It is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 78, 28765 to 28784, 28799 to 28818, 28950 to 28969, 29010 to 29029, 29106 to 29125, 29176 to 29195, 29354 to 29373, 29615 to 29634, and 29712 to 29731.

[0049] In one embodiment, the antisense oligonucleotide of the present invention comprises: (a) a base sequence selected from the group consisting of SEQ ID NOs: 7, 9, 12, 19, 20, 23-26, 36-40, 42-44, 46, 47, 49-51, 53-59, 61, 63-68, 70, 74-82, 85, 88-90, 92, 93, 95, 96, 99, 101, 106-110, 112-114, 117, 119, 121, 124, 125-127, 129, 131, 138-143, 149, 152, 153, 155, 157, 158, 162-165, 167-169, and 171-176; (b) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 7, 9, 12, 19, 20, 23-26, 36-40, 42-44, 46, 47, 49-51, 53-59, 61, 63-68, 70, 74-82, 85, 88-90, 92, 93, 95, 96, 99, 101, 106-110, 112-114, 117, 119, 121, 124, 125-127, 129, 131, 138-143, 149, 152, 153, 155, 157, 158, 162-165, 167-169, and 171-176, in which one or several nucleotides have been added, deleted, or substituted; or (c) SEQ ID NOs: 7, 9, 12, 19, 20, 23-26, 36-40, 42-44, 46, 47, 49-51, 53-59, 61, 63-68, 70, 74-82, 85, 88-90, 92, 93, 95, 96, 99, 101, 106-110, 112-114, 117, 119, 121, 124, 125-127, 129, 131, 138-143, 149, 152, 153 or consisting of a nucleotide sequence having 80% or more, 85% or more, 86% or more, 87% or more, 88% or more, 89% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more sequence identity to a nucleotide sequence selected from the group consisting of 155, 157, 158, 162-165, 167-169, and 171-176. In one embodiment, the antisense oligonucleotide of the present invention comprises or consists of any of the nucleotide sequences of (a) above.

[0050] In one embodiment, at least one target region of the antisense oligonucleotide of the present invention is selected from the group consisting of the nsp3 region, the RNA-dependent RNA polymerase region, the helicase region, the 3'-to-5' exonuclease region, the endoRNase region, the 2'-O-ribose methyltransferase region, the S region including the S1 region and the S2 region, and the N region in the genomic RNA of SARS-CoV-2. For example, the antisense oligonucleotides of the present invention may be located at positions 5384 to 5403, 6407 to 6426, 13642 to 13661, 13672 to 13691, 15173 to 15192, 15175 to 15194, 15179 to 15198, 15496 to 15515, 15498 to 15517, 16189 to 16208, 16192 to 16211, 16195 to 16214, 16201 to 16220, 17083 to 17102, 17084 to 17103, 17565 to 17584 in SEQ ID NO: 1. positions, 17576 to 17595, 18257 to 18276, 19783 to 19802, 20108 to 20127, 20111 to 20130, 21502 to 21521, 21504 to 21523, 24665 to 24684, 28745 to 28764, 28746 to 28765, 28756 to 28775, 28757 to 28776, 28758 to 28777, and 28759 to 28778. In one embodiment, the antisense oligonucleotide of the present invention is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 6407 to 6426, 15173 to 15192, 15496 to 15515, 15498 to 15517, 16195 to 16214, 17084 to 17103, 20111 to 20130, 21502 to 21521, 28757 to 28776, and 28758 to 28777 of SEQ ID NO: 1.

[0051] In one embodiment, the antisense oligonucleotide of the present invention comprises: (a) a base sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165; (b) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165, in which one or several nucleotides have been added, deleted, or substituted; or (c) a nucleotide sequence having 80% or more sequence identity to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165, or consisting of any of these sequences. In one embodiment, the antisense oligonucleotide of the present invention comprises or consists of any of the nucleotide sequences in (a) above. In one embodiment, the antisense oligonucleotide of the present invention is: (a) a base sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164; (b) a base sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164 in which one or several bases have been added, deleted, or substituted; or (c) A nucleotide sequence having 80% or more, 85% or more, 86% or more, 87% or more, 88% or more, 89% or more, 90% or more, 91% or more, 92% or more, 93% or more, 94% or more, 95% or more, 96% or more, 97% or more, 98% or more, or 99% or more sequence identity to a nucleotide sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164, or consisting of any of these sequences.In one embodiment, the antisense oligonucleotide of the present invention comprises or consists of any of the base sequences of (a) above.

[0052] In one embodiment, the antisense oligonucleotide of the present invention inhibits the function of a target region. As used herein, "inhibiting the function of a target region" includes one or more of the following: cleavage by RNase H of an RNA containing the target region that has bound to the target region and formed a duplex with the antisense oligonucleotide; inhibition of replication of the RNA containing the target region; inhibition of translation of the target region when it is translated; and inhibition of transcription of the RNA containing the target region. As used herein, the target region may be present not only in the genomic RNA but also in the subgenomic RNA. "Subgenomic RNA" refers to an RNA that is shorter than the genomic RNA and is synthesized by RNA-dependent RNA polymerase using a portion of the (-) strand RNA as a template, which acts as mRNA for viral protein synthesis (translation).

[0053] The antisense oligonucleotides of the present invention can generally be produced according to the methods described in W. Brad Wan et al., Nucleic Acid Research, Vol. 42, No. 22 13456 (2014), etc. In addition, they can be easily synthesized using various automated synthesizers (e.g., AKTA oligopilot plus 10 / 100 (GE Healthcare)), or can be produced by outsourcing to a third party organization (e.g., Promega or Takara).

[0054] In one embodiment, the present invention relates to a pharmaceutical composition comprising one or more of the antisense oligonucleotides of the present invention, or pharmaceutically acceptable salts thereof, or hydrates thereof. When the antisense nucleotides of the present invention are administered to a subject, the pharmaceutical composition of the present invention may contain a carrier that facilitates delivery of the antisense nucleotides. Such carriers are not particularly limited as long as they are pharmaceutically acceptable, and examples include cationic carriers such as cationic liposomes and cationic polymers, or carriers that utilize viral envelopes. Examples of cationic liposomes include liposomes formed from 2-O-(2-diethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol and phospholipids as essential components (hereinafter referred to as "Liposome A"), Oligofectamine (registered trademark) (Invitrogen), Lipofectin (registered trademark) (Invitrogen), Lipofectamine (registered trademark) (Invitrogen), Lipofectamine 2000 (registered trademark) (Invitrogen), DMRIE-C (registered trademark) (Invitrogen), GeneSilencer (registered trademark) (Gene Therapy Systems), TransMessenger (registered trademark) (QIAGEN), TransIT TKO (registered trademark) (Mirus), and Nucleofector II (Lonza). Examples of cationic polymers include JetSI (registered trademark) (manufactured by Qbiogene) and Jet-PEI (registered trademark) (polyethyleneimine, manufactured by Qbiogene). Examples of carriers utilizing viral envelopes include GenomeOne (registered trademark) (HVJ-E liposome, manufactured by Ishihara Sangyo Kaisha). Alternatively, the pharmaceutical device described in Japanese Patent No. 2924179 and the cationic carriers described in Republished Patent Publication Nos. 2006 / 129594 and 2008 / 096690 can also be used.

[0055] In one embodiment, the antisense oligonucleotide of the present invention may be conjugated with a lipid or the like in a pharmaceutical composition to facilitate delivery of the antisense oligonucleotide, for example, with cholesterol as described in Bijsterbosch, MK et al. (2000) Nucleic Acid Res., 28, 2717-2725.

[0056] The pharmaceutical compositions of the present invention may contain pharmaceutically acceptable additives in addition to the antisense oligonucleotide, its pharmaceutically acceptable salt, or hydrate thereof, and optionally the above-mentioned carrier. Examples of such additives include emulsifiers (e.g., C6-C22 fatty acids or pharmaceutically acceptable salts thereof, albumin, and dextran), stabilizers (e.g., cholesterol, phosphatidic acid, mannitol, and sorbitol), isotonicity agents (e.g., sodium chloride, glucose, maltose, lactose, sucrose, and trehalose), and pH adjusters (e.g., hydrochloric acid, sulfuric acid, phosphoric acid, acetic acid, sodium hydroxide, potassium hydroxide, and triethanolamine). These additives may be used alone or in combination. The content of the additive in the composition of the present invention is suitably 90% by weight or less, preferably 70% by weight or less, and more preferably 50% by weight or less.

[0057] The method for preparing the pharmaceutical composition of the present invention is not limited, and can be prepared, for example, by adding the antisense oligonucleotide of the present invention to a dispersion of a carrier and stirring appropriately. In addition, additives can be added at an appropriate step, either before or after the addition of the antisense oligonucleotide of the present invention. The aqueous solvent that can be used when adding the antisense oligonucleotide of the present invention is not particularly limited as long as it is pharmaceutically acceptable, and examples thereof include electrolyte solutions such as water for injection, distilled water for injection, and physiological saline, and sugar solutions such as glucose solution and maltose solution. In addition, conditions such as pH and temperature in such cases can be appropriately selected by those skilled in the art.

[0058] The pharmaceutical composition of the present invention can be, for example, a liquid formulation or a lyophilized formulation thereof. The lyophilized formulation can be prepared by lyophilizing the composition of the present invention in liquid form using standard methods. For example, after appropriate sterilization of the composition of the present invention in liquid form, a predetermined amount can be dispensed into vials, pre-frozen for approximately 2 hours at a temperature ranging from approximately −40°C to −20°C, primary dried under reduced pressure at a temperature ranging from approximately 0°C to 10°C, and then secondary dried under reduced pressure at a temperature ranging from approximately 15°C to 25°C, thereby achieving lyophilization. The interior of the vial can then generally be purged with nitrogen gas, and the vial can then be stoppered to obtain a lyophilized formulation of the composition of the present invention.

[0059] The lyophilized pharmaceutical composition of the present invention can generally be reconstituted and used by adding any appropriate solution (reconstitution liquid). Examples of such reconstitution liquid include water for injection, physiological saline, and other general infusion solutions. The volume of the reconstitution liquid varies depending on the intended use and is not particularly limited, but is suitably 0.5 to 2 times the volume of the liquid before lyophilization or 500 mL or less.

[0060] The dosage of the composition of the present invention can be adjusted taking into consideration the type of antisense oligonucleotide or pharmaceutically acceptable salt thereof or hydrate thereof contained therein, the formulation of the composition, the patient's condition such as age and body weight, the route of administration, and the nature and symptoms of the disease. The amount of the antisense oligonucleotide of the present invention can be, for example, 0.01 mg to 200 mg or 0.1 mg to 20 mg per kg of body weight per administration, preferably 0.2 mg to 10 mg per kg of body weight, more preferably 0.5 mg to 4 mg per kg of body weight, and more preferably 1 mg to 2 mg per kg of body weight per administration. The number and frequency of administration are not limited, but the composition can be administered once, or several times a few days later (e.g., within the next day to one week), for a total of multiple administrations (e.g., two times in total). When administered multiple times, the administration frequency can be once every 1 to 3 days, once every 4 to 6 days, once a week, or once every 2 to 3 weeks. These values ​​may vary depending on the type of disease being targeted, the administration form, and the target molecule. Therefore, in some cases, a lower dose or administration frequency may be sufficient, while in other cases a higher dose or administration frequency may be required.

[0061] The administration form of the composition of the present invention is not particularly limited as long as it is a pharmaceutically acceptable administration form and can be selected depending on the treatment method, and examples thereof include intratracheal administration, pulmonary administration, nasal administration, intravenous administration, intraarterial administration, intramuscular administration, subcutaneous administration, oral administration, intratissue administration, transdermal administration, etc. Furthermore, the dosage form that the composition of the present invention can take is not particularly limited, and examples thereof include inhalants, various injections, oral administration, drip infusions, ointments, lotions, etc.

[0062] The administration form of the pharmaceutical composition of the present invention is preferably intratracheal administration, and the dosage form that the composition of the present invention can take is preferably an inhalant, specifically, for example, an inhalation liquid (e.g., administered with a nebulizer), a powder inhalant (e.g., administered with a DPI (dry powder inhaler)), or an aerosol, preferably an inhalation liquid.

[0063] The subjects to which the antisense oligonucleotides or pharmaceutical compositions of the present invention are administered include, for example, mammals, such as primates such as humans, laboratory animals such as rats, mice, and brown rats, and livestock animals such as pigs, cows, horses, and sheep, and preferably humans.

[0064] In one embodiment, the present invention relates to a method for treating and / or preventing a viral infection selected from the group consisting of SARS-CoV-2, SARS-CoV-1, and MERS-CoV, comprising the step of administering to a subject the antisense oligonucleotide of the present invention, or a pharmaceutically acceptable salt or hydrate thereof, or a pharmaceutical composition thereof. The pharmaceutical composition in this embodiment, as well as the dosage and administration route of the pharmaceutical composition, are as described herein.

[0065] As used herein, treatment of a viral infection includes one or more of alleviating, improving, and remission of a disease caused by the virus or its symptoms (e.g., one or more of shortness of breath, fever, dry cough, headache, chills, and muscle pain). As used herein, prevention of a viral infection includes reducing the risk of developing a disease caused by the virus or its symptoms.

[0066] The present invention will be explained in more detail below with reference to examples and test examples, but the present invention is not limited to the scope shown in the examples.

[0067] Example 1: Measurement of gapmer knockdown activity using SARS-CoV-2 expression plasmid The genomic RNA sequences of SARS-CoV-2 (reference sequence; NC_045512.2) (SEQ ID NO: 1) and SARS-CoV-1 (reference sequence; NC_004718.3) were compared to identify regions conserved between the two species. The regions conserved between the two species are positions 43 to 89, 242 to 279, 290 to 312, 455 to 477, 704 to 723, 3352 to 3372, 5384 to 5403, 6071 to 6090, 6406 to 6427, 6797 to 6815, 7532 to 7551, 8707 to 8724, 10292 to 10309, 10406 ​​to 10431, 10484 to 10506, 11609 to 11630, 12023 to 12024, and 12123 to 12125 of SEQ ID NO: 1. 45th, 12170-12188, 12212-12234, 12314-12339, 12401-12420, 12589-12608, 12839-12867, 12898-12922, 12965-1 2990th, 13151-13175, 13271-13290, 13363-13386, 13458-13502, 13642-13661, 13672-13691, 13762-13790, 13894 ~13916th, 14050-14069, 14290-14312, 14512-14531, 14654-14687, 14698-14717, 14750-14777, 14824-14846, 1 4854 to 14873, 14878 to 14909, 14953 to 14990, 14992 to 15026, 15037 to 15061, 15063 to 15140, 15172 to 15198, 15278 to 15 299th, 15454th to 15479th, 15496th to 15518th, 15520th to 15539th, 15622nd to 15644th, 15791st to 15809th, 15829th to 15859th, 15886th to 15905th, 15929th -15950th, 15985-16011, 16051-16085, 16189-16220, 16430-16451, 16636-16655, 16822-16845, 17015-17051,17080th to 17103rd, 17137th to 17156th, 17215th to 17234th, 17254th to 17277th, 17564th to 17600th, 17748th to 17765th, 17773th to 17792nd, 17830-17849, 17859-17876, 18094-18111, 18196-18218, 18253-18278, 18370-18387, 19568-19595, 1 9622-19639, 19780-19802, 20107-20130, 20776-20795, 20797-20816, 20888-20909, 21453-21472, 215 02nd to 21524th, 22550 to 22569, 22814 to 22836, 23093 to 23113, 23956 to 23976, 24302 to 24324, 24446 to 24465, 24 467th - 24489th, 24620 - 24651, 24662 - 24684, 24962 - 24982, 25104 - 25128, 25364 - 25384, 25502 - 25520 , 26287th to 26325th, 26574th to 26604th, 26782nd to 26800th, 27093th to 27111th, 27771st to 27794th, 27806th to 27823rd, 28270th to 28294th, The nucleotide sequences identified were: positions 28397 to 28418, 28509 to 28538, 28744 to 28784, 28799 to 28820, 28946 to 28972, 28986 to 29005, 29010 to 29031, 29102 to 29130, 29174 to 29196, 29354 to 29373, 29615 to 29634, 29712 to 29731, and 29787 to 29867. Test substance gapmers Nos. G1 to G176 (SEQ ID NOs: 2 to 177) were prepared by targeting sequences contained in these conserved regions. In the test substance gapmer Nos. G1 to G176, all nucleosides in the wing region contain a 2'-OMe (2'-O-CH3) group. In the examples, gapmers with "MOE" at the end of the gamper No. are 2'-O-MOE gapmers (gapmers in which all nucleosides in the wing region contain 2'-O-MOE (2'-O-CH2CH2OCH3)).In Table 1-1 of the Examples, all internucleoside bonds are phosphorothioate bonds. In the Examples, all oligonucleotides are, in principle, 5-10-5 type gapmers, except for the following oligonucleotides: 19mer 4-10-5 type: G110, G114 19mer 5-10-4 type: G11, G19, G61, G115 18mer 4-10-4 type: G13, G14, G81, G84, G85, G88, G90 18mer 5-9-4 type: G117 17mer 4-9-4 type: 133-2'OMe, 133-2'MOE, 134-2'OMe, 134-2'MOE Note that in the above gapmers, "abc type" means that the gapmer is composed of, from the 5' side to the 3' side, a wing region of nucleotide length a, a gap region of nucleotide length b, and a wing region of nucleotide length c. In the 2'-O-MOE gapmer, 2'-O-MOE-5-methyl C was used for the C in the wing region, and 5-methyl dC was used for the C in the gap region. The target sequence and the sequences of test substance gapmers No. G1 to G176 are shown in Table 1-1 below.

[0068] In Table 1-1, two target sequences (5'UTR / nsp1 of G3) indicate that the target sequence of the gapmer contains two regions. The target sequences and sequences of the PS / PS type test substances, gapmer Nos. G58 PO / PS type-1, G58 PO / PS-type-2, G144 PO / PS type-1, and PO / PS type-2, are shown in Table 1-2. In the 5-10-5 type gapmers in Table 1-2, the bonds between the second and third nucleosides and the fourth and fifth nucleosides from the 5' end of the 5' wing region are phosphodiester bonds, and the bonds between the first and second nucleosides and the third and fourth nucleosides from the 5' end of the 3' wing region are phosphodiester bonds, and all other internucleoside bonds are phosphorothioate bonds (PO / PS type-2). The bonds between the second and third nucleosides, the third and fourth nucleosides, and the fourth and fifth nucleosides from the 5' side of the 3' wing region are phosphodiester bonds, and the bonds between the first and second nucleosides, the second and third nucleosides, and the third and fourth nucleosides from the 5' side of the 3' wing region are phosphodiester bonds, and all other internucleoside bonds are phosphorothioate bonds (PO / PS type-1).

[0069] 293T cells (obtained from ATCC) were electroporated with a SARS-CoV-2 expression plasmid (final concentration 400ng / sample), a Firefly luciferase (Fluc) expression plasmid (control plasmid) (final concentration 4ng / sample), and either the test substance gapper or the negative control gapper (final concentration 1.3µM) using the SF Cell Line 4D-Nucleofector X Kit (Lonza) and a 4D-Nucleofector (Lonza) (16.4µL Nucleofector and 3.6µL supplement) with DS-130 selected in the 4D-Nucleofector's built-in pulse program. The SARS-CoV-2 expression plasmid was prepared by synthesizing a DNA artificial gene from a nine-part fragment of SARS-CoV-2 genomic RNA (Table 1-3) and subcloning it into the expression plasmid pcDNA3.1. Synthetic gene synthesis and subcloning into the expression plasmid were outsourced to Thermo Fisher Scientific. However, because plasmid No. 4 was unavailable, SARS-CoV-2 (2019-nCoV) 3C-like proteinase / 3CLpro Gene ORF cDNA clone expression plasmid (Sino Biological) and SARS-CoV-2 (2019-nCoV) nsp8 Gene ORF cDNA clone expression plasmid (Sino Biological) were used to evaluate the activity of some gapmers (referred to as "3CLpro" and "nsp8," respectively, in Tables 1-3). The control plasmid was the pGL4.50 [luc2 / CMV / Hygro] Vector (Promega). The sequence of ASO#129700 (Wheeler et al. 2012. Nature 488: 111-5. doi: 10.1038 / nature11362) was used as a negative control gapmer.The test substance gapmer and negative control gapmer were synthesized by Nippon Bio Services as 2'-OMe gapmers (gapmers in which all nucleosides in the wing moiety contain 2'-OMe (2'-O-CH)) or 2'-O-MOE gapmers (gapmers in which all nucleosides in the wing moiety contain 2'-O-MOE (2'-O-CHCHOCH).

[0070] 293T: 2.0 × 10 per electroporation 5 Cells were used. After electroporation, they were seeded into 24-well plates. Two days after transfection with Gapmer and the corresponding plasmid, RNA was isolated from the cells using NucleoSpin RNA (Macherey-Nagel). Reverse transcription (RT) was performed using the High Capacity cDNA Reverse Transcription Kit with RNase Inhibitor (Thermo Fisher Scientific) and the random primers provided with the kit. qPCR was performed using Fast SYBR Green Master Mix (Thermo Fisher Scientific) according to the manufacturer's protocol, with the RT reaction mixture as the template to measure the SARS-CoV-2 RNA expression level derived from the SARS-CoV-2 expression plasmid. The primers used to detect SARS-CoV-2 RNA derived from the SARS-CoV-2 expression plasmid are listed in Tables 1-3. SARS-CoV-2 RNA expression levels were normalized by the Fluc RNA expression level derived from the control plasmid or the PPIB RNA expression level of the housekeeping gene. The primers used to detect Fluc RNA or PPIB RNA are listed in Tables 1-4. The rate of SARS-CoV-2 RNA expression suppression (knockdown activity of test substance gapmer) was analyzed relative to the level of SARS-CoV-2 RNA expression after introduction of the negative control gapmer. The results are shown in Table 1-5.

[0071]

[0072]

[0073]

[0074] In the above table, "Fluc-corrected 5' side" and "Fluc-corrected 3' side" refer to whether the expression was detected using the 5' side or the 3' side of the two pairs of detection primers (Table 1-3).

[0075] Example 2: Measurement of gapmer knockdown activity using SARS-CoV-2 virus. Human ACE2-stably expressing cells (293T-ACE2) were prepared by infecting 293T cells with a human ACE2-expressing lentiviral vector. Test gapmers or negative control gapmers were applied at final concentrations of 2-10 μM. Using the Amxa Cell Line Nucleofector Kit V (Lonza) (73.6 μL of solution V and 16.4 μL of supplement per sample) and a Nucleofector II (Amaxa), electroporation was performed using the A-023 pulse program on the Nucleofector II. The negative control gapmer was ASO#129700 (Wheeler et al. 2012. Nature 488: 111-5. doi: 10.1038 / nature11362). The test substance gapmers used were some unevaluated sequences among Gapmer Nos. 1 to 135 used in Example 1, and Gapmer Nos. 136 to G176, which were additionally designed based on the activity data of Gapmer Nos. 1 to 135. The test substance gapmers and negative control gapmers were synthesized as 2'-OMe gapmers or 2'-MOE gapmers by outsourcing to Japan Bio Services or Ajinomoto Bio-Pharma Services. 293T-ACE2 was used at a concentration of 6.0 × 10 per electroporation. 5 After electroporation, 5.0 × 10 293T-ACE2 cells were added to a 96-well plate. 4Cells were seeded at 1000 cells / well. The next day, SARS-CoV-2 virus was added at an MOI of 0.01 to infect the cells. SARS-CoV-2 virus was obtained from the National Institute of Infectious Diseases (NIID) and the WK521 strain was used. A recombinant SARS-CoV-2 virus (rSARS-CoV-2-ORF8-Nluc) was used, in which the nano-luciferase gene was engineered into the coding region of the accessory protein ORF8 using a method similar to that described previously (Terada et al. 2019. J Virol 93: e01208-19. doi: 10.1128 / JVI.01208-19). The virus was removed from the medium by medium replacement 1 h after virus addition. 48 h after virus infection, the culture supernatant was collected, and the cells were lysed using Passive Lysis Buffer (Promega). The luciferase activity in the cell lysate was measured using the Luciferase Assay System (Promega) according to the attached protocol, and the amount of SARS-CoV-2 virus in the cells was evaluated (measurement of intracellular virus amount). In addition, the collected culture supernatant was separately plated in a 96-well plate at 3.0 × 10 4 5 μL / well of the culture supernatant was added to 293T-ACE2 cells seeded at 1000 cells / well, and the cells were infected with the virus in the culture supernatant. 24 hours after infection with the culture supernatant, the amount of SARS-CoV-2 virus in the cells was assessed using the same method as for the initial virus infection, and the amount of infectious virus in the culture supernatant was evaluated (measurement of infectious virus amount in the medium). The knockdown activity of the test substance gapmer (inhibition rate of infectious virus amount in the medium: the rate of inhibition of infectious virus amount when the test substance gapmer was introduced relative to the infectious virus amount when the negative control gapmer was introduced) is shown in Table 2.

[0076]

Claims

1. An antisense oligonucleotide consisting of 15 to 30 nucleotides, which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of the 5'UTR region, nsp1 region, nsp3 region, nsp4 region, 3C-like proteinase region, nsp6 region, nsp7 region, nsp8 region, nsp9 region, nsp10 region, RNA-dependent RNA polymerase region, helicase region, 3'-to-5' exonuclease region, endoRNase region, 2'-O-ribose methyltransferase region, S region including S1 region and S2 region, ORF3a region, E region, M region, ORF7b region, N region, ORF10 region, and 3'UTR region in the genomic RNA of SARS-CoV-2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof; An antisense oligonucleotide, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, having an antiviral effect against a virus selected from the group consisting of SARS-CoV-2, SARS-CoV-1, and MERS-CoV.

2. 2. The antisense oligonucleotide according to claim 1, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein the antisense oligonucleotide is a gapmer comprising a central gap region and two wing regions flanking the gap region.

3. The antisense oligonucleotide or a pharmaceutically acceptable salt thereof or a hydrate thereof according to claim 1 or 2, wherein the virus is SARS-CoV-2 or SARS-CoV-1.

4. The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein at least one target region is selected from the group consisting of the nsp1 region, nsp3 region, 3C-like proteinase region, nsp8 region, RNA-dependent RNA polymerase region, helicase region, 3'-to-5' exonuclease region, endoRNase region, 2'-O-ribose methyltransferase region, S region including S1 region and S2 region, M region, ORF7b region, N region, ORF10 region, and 3'UTR region in the genomic RNA of SARS-CoV-2.

5. The antisense oligonucleotide or its pharmaceutically acceptable salt or hydrate thereof according to claim 1 or 2, wherein at least one target region is selected from the group consisting of the nsp3 region, the RNA-dependent RNA polymerase region, the helicase region, the 3'-to-5' exonuclease region, the endoRNase region, the 2'-O-ribose methyltransferase region, the S region including the S1 region and the S2 region, and the N region in the genomic RNA of SARS-CoV-2.

6. The allocation number 1 is 43 to 89, 242 to 279, 290 to 312, 455 to 477, 704 to 723, 3352 to 3372, 5384~5403, 6071~6090, 6406~6427, 6797~6815, 7532~7551, 8707~8 724, 10292-10309, 10406-10431, 10484-10506, 11609-11630, 12023-12045, 12170-12188, 12212-12234, 12314-12339, 12401-12420, 12589-12608, 12839-12867, 12898-12922, 12965-12990, 13151-13175, 13271-13290, 13363-13386, 13458-13502, 13642-13661, 136 72nd to 13691st, 13762nd to 13790th, 13894th to 13916th, 14050th to 14069th, 14290th to 14312th, 14512th to 14531st, 14654th to 14687th, 14698th to 14717th, 14750th to 14777th, 14824th 14846, 14854-14873, 14878-14909, 14953-14990, 14992-15026, 15037-15061, 15063-15140, 15172-15198, 15278-15299, 15454 15479, 15496-15518, 15520-15539, 15622-15644, 15791-15809, 15829-15859, 15886-15905, 15929-15950, 15985-16011, 16051-1 6085, 16189-16220, 16430-16451, 16636-16655, 16822-16845, 17015-17051, 17080-17103, 17137-17156, 17215-17234, 17254-17 277, 17564-17600, 17748-17765, 17773-17792, 17830-17849, 17859-17876, 18094-18111, 18196-18218, 18253-18278, 18370-18387,19568th - 19595th, 19622 - 19639th, 19780 - 19802nd, 20107 - 20130th, 20776 - 20795th, 20797 - 20816th, 20888 -20909th, 21453-21472, 21502-21524, 22550-22569, 22814-22836, 23093-23113, 23956-2 3976th, 24302nd to 24324th, 24446th to 24465th, 24467th to 24489th, 24620th to 24651st, 24662nd to 24684th, 24962nd to 24th 982nd, 25104th to 25128th, 25364th to 25384th, 25502nd to 25520th, 26287th to 26325th, 26574th to 26604th, 26782 to 26800 27093-27111, 27771-27794, 27806-27823, 28270-28294, 28397-28418, 28509-28538, 2 8744th - 28784th, 28799th - 28820th, 28946th - 28972nd, 28986 - 29005th, 29010th - 29031st, 29102nd - 29130th, 291 The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 74 to 29196, positions 29354 to 29373, positions 29615 to 29634, positions 29712 to 29731, and positions 29787 to 29867.

7. The allocation number 1 is 47 to 66, 242 to 261, 259 to 278, 292 to 311, 456 to 475, 704 to 723 bits, 3353-3372 bits, 5384-5403 bits, 6071-6090 bits, 6406-6425 bits, 6407-6426 bits, 6408-6427, 6797-6815, 7532-7551, 8707-8724, 10292-10309, 10407-10426, 10409-10428, 10487-10506, 11609-11628, 12025- 12044, 12170-12188, 12214-12233, 12316-12335, 12401-12420, 12589-12608, 12843-12862, 12900-12919, 12969-12988, 13153 13172, 13271-13290, 13366-13385, 13462-13481, 13481-13500, 13642-13661, 13672-13691, 13767-13786, 13894-13913, 14050 14,069th to 14,292nd to 14,311th, 14,512th to 14,531st, 14,656th to 14,675th, 14,668th to 14,687th, 14,698th to 14,717th, 14,753rd to 14,772nd, 14,826th to 14,845th, 14,854th to 14,873rd, 14,8 79th to 14898th, 14889th to 14908th, 14962th to 14981st, 14971st to 14990th, 14992th to 15011th, 15007th to 15026th, 15040th to 15059th, 15064th to 15083rd, 15091st to 15107th, 15 092 to 15111, 15120 to 15139, 15172 to 15191, 15173 to 15192, 15174 to 15193, 15175 to 15194, 15179 to 15198, 15280 to 15299, 15280 to 15296, 1 5457th to 15476th, 15496th to 15515th, 15497th to 15516th, 15498th to 15517th, 15499th to 15518th, 15520th to 15539th, 15623rd to 15642nd, 15791st to 15809th, 15834th to 15853rd,15886-15905, 15931-15950, 15988-16007, 16052-16071, 16066-16085, 16189-16208, 16192-16211, 16193-16212, 16194-16213 16195-16214, 16196-16215, 16198-16217, 16201-16220, 16432-16451, 16636-16655, 16825-16844, 17016-17035, 17032-170 51, 17080-17099, 17083-17102, 17084-17103, 17137-17156, 17215-17234, 17257-17276, 17564-17583, 17565-17584, 17566-1 7585, 17567-17586, 17569-17588, 17573-17592, 17576-17595, 17580-17599, 17748-17765, 17773-17792, 17830-17849, 17859- 17876, 18094-18111, 18197-18216, 18257-18276, 18370-18387, 19572-19591, 19622-19639, 19780-19799, 19783-19802, 20107 20126, 20108-20127, 20109-20128, 20110-20129, 20111-20130, 20776-20795, 20797-20816, 20889-20908, 21453-21472, 215 02 to 21521, 21503 to 21522, 21504 to 21523, 22550 to 22569, 22817 to 22836, 23093 to 23112, 23957 to 23976, 24303 to 24322, 24446 to 24465, 2 4469th to 24488th, 24620th to 24639th, 24632th to 24651st, 24665th to 24684th, 24962th to 24981st, 25107th to 25126th, 25365th to 25384th, 25502th to 25520th, 26287th to 26306th,26305th to 26324th, 26579th to 26598th, 26782nd to 26800th, 27093rd to 27111th, 27774th to 2779th 3rd place, 27806th to 27823rd, 28274th to 28293rd, 28398th to 28417th, 28514th to 28533rd, 28744th to 28th 763rd, 28745th to 28764th, 28746th to 28765th, 28747th to 28766th, 28748th to 28767th, 28752nd ~28771st, 28756th~28775th, 28757th~28776th, 28758th~28777th, 28759th~28778th, 2876 The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 2 to 28781, 28765 to 28784, 28799 to 28818, 28950 to 28969, 28986 to 29005, 29010 to 29029, 29106 to 29125, 29176 to 29195, 29354 to 29373, 29615 to 29634, 29712 to 29731, and 29789 to 29808.

8. The allocation number 1 is 704-723, 5384-5403, 6407-6426, 10409-10428, 10487-1 0506 bits, 12170 bits to 12188 bits, 12214 bits to 12233 bits, 12316 bits to 12335 bits, 12401 bits to 12420 bits, 13642nd to 13661st, 13672nd to 13691st, 13767th to 13786th, 13894th to 13913th, 14050th to 14069th, 14512th to 14531st, 14656th to 14675th, 14668th to 14687th, 14753th to 14772nd 14826-14845, 14879-14898, 14889-14908, 14962-14981, 14992-15011, 15007-15026, 15040-15059, 15064-15083, 15091-151 07, 15092-15111, 15120-15139, 15173-15192, 15175-15194, 15179-15198, 15280-15299, 15280-15296, 15457-15476, 15496-1 5515, 15498-15517, 15791-15809, 15834-15853, 15886-15905, 15931-15950, 15988-16007, 16052-16071, 16066-16085, 16189 16208, 16192-16211, 16195-16214, 16201-16220, 16432-16451, 16636-16655, 17016-17035, 17032-17051, 17083-17102, 1708 4th to 17103rd, 17257th to 17276th, 17565th to 17584th, 17576th to 17595th, 17580th to 17599th, 17748th to 17765th, 17773rd to 17792nd, 17830th to 17849th, 18094th to 18111th, 18 197th to 18216th, 18257th to 18276th, 19622th to 19639th, 19783rd to 19802th, 20108th to 20127th, 20111th to 20130th, 20776th to 20795th, 20797th to 20816th, 20889th to 20908th,21502nd to 21521st, 21504th to 21523rd, 24469th to 24488th, 24620th to 24639th, 2463 2nd to 24651st, 24665th to 24684th, 24962nd to 24981st, 25107th to 25126th, 26782nd to 26th 800th, 27806th to 27823rd, 28274th to 28293rd, 28514th to 28533rd, 28745th to 28764th, 28746th to 28765th, 28756th to 28775th, 28757th to 28776th, 28758th to 28777th, 28759 The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 28778 to 28778, 28765 to 28784, 28799 to 28818, 28950 to 28969, 29010 to 29029, 29106 to 29125, 29176 to 29195, 29354 to 29373, 29615 to 29634, and 29712 to 29731.

9. positions 5384 to 5403, 6407 to 6426, 13642 to 13661, 13672 to 13691, 15173 to 15192, 15175 to 15194, 15179 to 15198, 15496 to 15515, 15498 to 15517, 1618 9th to 16208th, 16192nd to 16211th, 16195th to 16214th, 16201st to 16220th, 17083rd to 17102nd, 17084th to 17103rd, 17565th to 17584th, 17576th to 17595th, 18257th to 18276th, 19783rd to 198th 3. The antisense oligonucleotide according to claim 1, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 02, 20108 to 20127, 20111 to 20130, 21502 to 21521, 21504 to 21523, 24665 to 24684, 28745 to 28764, 28746 to 28765, 28756 to 28775, 28757 to 28776, 28758 to 28777, and 28759 to 28778.

10. 3. The antisense oligonucleotide according to claim 1, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which is complementary to a nucleic acid comprising at least 10 consecutive bases in at least one target region selected from the group consisting of positions 6407 to 6426, 15173 to 15192, 15496 to 15515, 15498 to 15517, 16195 to 16214, 17084 to 17103, 20111 to 20130, 21502 to 21521, 28757 to 28776, and 28758 to 28777 of SEQ ID NO:

1.

11. 3. The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which is complementary to a nucleic acid comprising at least 15 consecutive bases in a target region.

12. (a) a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177; (b) a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177, in which one or more bases have been added, deleted, or substituted; or (c) The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate of the same, which comprises a base sequence having 80% or more sequence identity to a base sequence selected from the group consisting of SEQ ID NOs: 2 to 177 and inhibits the function of the target region.

13. (a) a base sequence selected from the group consisting of SEQ ID NOs: 7, 9, 12, 19, 20, 23 to 26, 36 to 40, 42 to 44, 46, 47, 49 to 51, 53 to 59, 61, 63 to 68, 70, 74 to 82, 85, 88 to 90, 92, 93, 95, 96, 99, 101, 106 to 110, 112 to 114, 117, 119, 121, 124, 125 to 127, 129, 131, 138 to 143, 149, 152, 153, 155, 157, 158, 162 to 165, 167 to 169, and 171 to 176; (b) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 7, 9, 12, 19, 20, 23-26, 36-40, 42-44, 46, 47, 49-51, 53-59, 61, 63-68, 70, 74-82, 85, 88-90, 92, 93, 95, 96, 99, 101, 106-110, 112-114, 117, 119, 121, 124, 125-127, 129, 131, 138-143, 149, 152, 153, 155, 157, 158, 162-165, 167-169, and 171-176, in which one or several nucleotides have been added, deleted, or substituted; or (c) SEQ ID NOs: 7, 9, 12, 19, 20, 23-26, 36-40, 42-44, 46, 47, 49-51, 53-59, 61, 63-68, 70, 74-82, 85, 88-90, 92, 93, 95, 96, 99, 101, 106-110, 112-114, 117, 119, 121, 124, 125-127, 129, 131, 138-143, 1 3. The antisense oligonucleotide according to claim 1, or a pharmaceutically acceptable salt thereof, or a hydrate of the same, which comprises a base sequence having 80% or more sequence identity to a base sequence selected from the group consisting of 49, 152, 153, 155, 157, 158, 162 to 165, 167 to 169, and 171 to 176, and which inhibits the function of the target region.

14. (a) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165; (b) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165, in which one or several nucleotides have been added, deleted, or substituted; or (c) The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which comprises a base sequence having 80% or more sequence identity to a base sequence selected from the group consisting of SEQ ID NOs: 9, 12, 36, 37, 61, 63, 64, 68, 70, 81, 82, 85, 88, 95, 96, 101, 106, 114, 119, 121, 124, 129, 131, 141, 157, 158, 162, 163, 164, and 165, and inhibits the function of the target region.

15. (a) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164; (b) a nucleotide sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164, in which one or several nucleotides have been added, deleted, or substituted; or (c) The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, which comprises a base sequence having 80% or more sequence identity to a base sequence selected from the group consisting of SEQ ID NOs: 12, 61, 68, 70, 85, 96, 124, 129, 163, and 164, and inhibits the function of the target region.

16. The antisense oligonucleotide according to claim 12, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, comprising the sequence (a).

17. 3. The antisense oligonucleotide according to claim 1, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein the antisense oligonucleotide consists of 20 nucleotides.

18. The wing region is 2'-OMe (2'-O-CH 3 ) group and / or 2'-O-MOE(2'-O-CH 2 CH 2 OCH 3 3. The antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, comprising a .

19. The antisense oligonucleotide according to claim 17, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein the bond between the second and third nucleosides from the 5' side and the bond between the fourth and fifth nucleosides from the 5' side of the 5' wing region are phosphodiester bonds, and the bond between the first and second nucleosides from the 5' side and the bond between the third and fourth nucleosides from the 3' wing region are phosphodiester bonds, and all other internucleoside bonds are phosphorothioate bonds.

20. An antisense oligonucleotide or a pharmaceutically acceptable salt thereof according to claim 18, wherein the bond between the second and third nucleosides and the bond between the fourth and fifth nucleosides from the 5' side of the 5' wing region are phosphodiester bonds, and the bond between the first and second nucleosides and the bond between the third and fourth nucleosides from the 5' side of the 3' wing region are phosphodiester bonds, and all other bonds between nucleosides are phosphorothioate bonds.

21. The antisense oligonucleotide according to claim 17, or a pharmaceutically acceptable salt thereof, or a hydrate thereof, wherein the bonds between the second and third nucleosides from the 5' side, the bond between the third and fourth nucleosides, and the bond between the fourth and fifth nucleosides from the 5' side of the 5' wing region are phosphodiester bonds, and the bonds between the first and second nucleosides, the bond between the second and third nucleosides, and the bond between the third and fourth nucleosides from the 5' side of the 3' wing region are phosphodiester bonds, and all other internucleoside bonds are phosphorothioate bonds.

22. The antisense oligonucleotide or a pharmaceutically acceptable salt thereof described in claim 18, or a hydrate thereof, wherein the bonds between the second and third nucleosides from the 5' side of the 5' wing region, the bond between the third and fourth nucleosides, and the bond between the fourth and fifth nucleosides are phosphodiester bonds, and the bonds between the first and second nucleosides from the 5' side of the 3' wing region, the bond between the second and third nucleosides, and the bond between the third and fourth nucleosides are phosphodiester bonds, and all other bonds between nucleosides are phosphorothioate bonds.

23. A pharmaceutical composition comprising the antisense oligonucleotide according to claim 1 or 2, or a pharmaceutically acceptable salt thereof, or a hydrate thereof.

24. 24. The pharmaceutical composition of claim 23 for treating and / or preventing a viral infection selected from the group consisting of SARS-CoV-2, SARS-CoV-1, and MERS-CoV.