Composition for treating HIV infection

Abstract

The present disclosure provides a composition for treating viral infections. In one mode, the present disclosure provides a composition that is for treating viral infections in subjects, and that includes a nucleic acid construct having, in an activatable manner, a nucleotide sequence encoding Ag85B protein and a nucleotide sequence encoding an antigen protein included in a virus belonging to the genus Lintivirus or a portion of the protein. In one embodiment, the composition according to the present disclosure is characterized by being administered after a viral infection.

Description

Compositions for treating HIV infection

[0001] The present disclosure provides compositions and related technologies for treating viral infections, more particularly, technologies relating to nucleic acid constructs operably comprising a nucleic acid sequence encoding an antigenic protein or a portion thereof contained in a virus belonging to the genus Lentivirus and a nucleic acid sequence encoding the Ag85B protein.

[0002] Despite the significant resources invested in the development of an effective human immunodeficiency virus (HIV) vaccine over the past 30 years, this task remains elusive. Antiretroviral therapy has dramatically reduced HIV-related morbidity and mortality, but it cannot cure HIV (Non-Patent Document 1). There have been two cases in which HIV remission was achieved through cell transplantation. The development of an effective vaccine could prevent the spread of HIV infection, but vaccine development has met with little success over the past 30 years. The only successful HIV vaccine to date was evaluated in the RV144 clinical trial, which demonstrated an overall efficacy of only 31% (Non-Patent Document 2).

[0003] Cohen, M. S. et al. Prevention of HIV-1 infection with early antiretroviral therapy. N. Engl. J. Med. 365, 493-505 (2011). Rerks-Ngarm, S. et al. Vaccination with ALVAC and AIDSVAX to prevent HIV-1 infection in Thailand. N. Engl. J. Med. 361, 2209-2220 (2009).

[0004] The present inventors discovered that a nucleic acid construct operably comprising a nucleic acid sequence encoding an antigen protein or a part thereof contained in a virus belonging to the genus Lentivirus and a nucleic acid sequence encoding the Ag85B protein can treat viral infection in a subject, and thus completed the present invention.

[0005] Thus, the present invention provides, for example, the following items: (Item 1) A composition for treating a viral infection in a subject, comprising a nucleic acid construct operably comprising a nucleic acid sequence encoding an antigenic protein or a portion thereof contained in a virus belonging to the Lentivirus genus and a nucleic acid sequence encoding an Ag85B protein. (Item 2) The composition according to the above items, characterized in that the composition is administered after viral infection. (Item 3) The composition according to any one of the above items, wherein the treatment for viral infection is to substantially eliminate the virus from the subject after viral infection. (Item 4) The composition according to any one of the above items, wherein the treatment for viral infection is to substantially eliminate the virus from the subject after viral infection. (Item 5) The composition according to any one of the above items, wherein the nucleic acid sequence encoding the antigenic protein or a portion thereof is a nucleic acid sequence encoding an attenuated virus. (Item 6) The composition according to any one of the above items, wherein the attenuated virus is a nef-deficient attenuated virus. (Item 7) The composition of any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is integrated into the nucleic acid sequence encoding the attenuated virus. (Item 8) The composition of any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is integrated in place of the deleted nef gene in the nucleic acid sequence encoding the nef-deficient attenuated virus. (Item 9) The composition of any one of the above items, wherein the virus is an AIDS virus. (Item 10) The composition of any one of the above items, wherein the virus is an attenuated virus that infects humans. (Item 11) The composition of any one of the above items, wherein the virus is an attenuated AIDS virus selected from the group consisting of HIV, SIV, SHIV, and FIV. (Item 12) The composition of any one of the above items, wherein the virus is an attenuated HIV. (Item 13) The composition of any one of the above items, wherein the virus is a nef-deficient attenuated HIV.(Item 14) The composition of any one of the above items, wherein the nucleic acid sequence encoding the antigen protein or a portion thereof comprises a nucleic acid sequence at least 90% identical to a nucleic acid sequence comprising nucleotides at positions 1 to 9633 and 10612 to 11022 of SEQ ID NO: 3, or a nucleic acid sequence comprising nucleotides at positions 1 to 8786 and 9765 to 15182 of SEQ ID NO: 4. (Item 15) The composition of any one of the above items, wherein the nucleic acid construct comprises a nucleic acid sequence at least 90% identical to the nucleic acid sequence set forth in SEQ ID NO: 3 or 4. (Item 16) The composition of any one of the above items, wherein the nucleic acid construct comprises the nucleic acid sequence set forth in SEQ ID NO: 3 or 4. (Item 17) The composition of any one of the above items, wherein the nucleic acid construct is administered once. (Item 18) The composition of any one of the above items, wherein the nucleic acid construct is administered two or more times. (Item 18-A) The composition of any one of the above items, wherein the nucleic acid construct is administered after administration of an anti-HIV drug. (Item 18-B) The composition of any one of the preceding items, wherein the anti-HIV drug is selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors, and is preferably tenofovir, emtricidavine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. (Item 18-C) The composition of any one of the preceding items, wherein the nucleic acid construct is administered after administration of an anti-HIV drug and after discontinuing administration of the HIV drug. (Item 18-D) The composition according to any one of the preceding items, wherein the nucleic acid construct is administered two or more times after administration of an anti-HIV drug. (Item 18-E) The composition according to any one of the preceding items, wherein the nucleic acid construct is administered three, four, five or more times after administration of an anti-HIV drug.(Item 18-F) The composition according to any one of the preceding items, wherein the two or more administrations of the nucleic acid construct are administered at intervals of at least one week or more. (Item 18-G) The composition according to any one of the preceding items, wherein the two or more administrations of the nucleic acid construct are administered at intervals of one to four weeks. (Item 18-H) The composition according to any one of the preceding items, wherein the two or more administrations of the nucleic acid construct are administered at intervals of one to three weeks. (Item 18-I) The composition according to any one of the preceding items, wherein the viral load in the patient's plasma is monitored during the two or more administrations of the nucleic acid construct, and the nucleic acid construct is administered again when it reaches a predetermined value. (Item 18-J) The nucleic acid construct is administered at a dose of 5.0 x 10 per administration. ５ T.C.I.D. ５０ or more or 1.0 x 10 ６ T.C.I.D. ５０ The composition according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 5.0 x 10 or more per administration. ５ T.C.I.D. ５０ or more or 1.0 x 10 ６ T.C.I.D. ５０ The composition according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, or once every two months. (Item 19) The composition according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of once to four weeks. (Item 19A) The composition according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of once or more. (Item 20) The nucleic acid construct is administered at a dose of 1.0 x 10 ３ T.C.I.D. ５０ (Item 21) The composition according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 1.0 x 10 or more. ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID５０ 22. The composition according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of less than 1.0 x 10 ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ 23. The composition according to any one of the preceding items, wherein the nucleic acid construct is administered once a week at a dose of less than 1.0 x 10 ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ 24. The composition according to any one of the preceding items, wherein the nucleic acid construct is administered once at a dose of less than 5.0 x 10, and if a virus is detected in the body, the nucleic acid construct is further administered. ４ T.C.I.D. ５０ Above 5.0 x 10 ６ T.C.I.D. ５０ The composition according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 5.0 x 10 ４ T.C.I.D. ５０20. The composition according to any one of the preceding items, wherein the subject is administered a single dose of at least 100 mg / kg / day. (Item 26) The composition according to any one of the preceding items, wherein the subject has already been administered an anti-HIV drug. (Item 27) The composition according to any one of the preceding items, wherein the subject has not yet been administered an anti-HIV drug, or the anti-HIV drug will be discontinued at the time of initiation of administration of the composition. (Item 28) The composition according to any one of the preceding items, wherein the subject has been administered an anti-HIV drug selected from the group consisting of a reverse transcriptase inhibitor, a protease inhibitor, an integrase inhibitor, and a CCR5 inhibitor. (Item 29) The composition of any one of the above items, wherein the anti-HIV drug is an anti-HIV drug selected from the group consisting of tenofovir, emtricidavine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. (Item 30) The composition of any one of the above items, wherein the composition completely eliminates the AIDS virus in the subject. (Item 31) The composition of any one of the above items, wherein the composition has not been administered prior to viral infection. (Item 32) The composition of any one of the above items, wherein the subject has previously been vaccinated with a BCG vaccine. (Item 1A) A method for treating a viral infection in a subject, the method comprising administering an effective amount of a nucleic acid construct operably comprising a nucleic acid sequence encoding an antigenic protein or a portion thereof contained in a virus belonging to the genus Lentivirus and a nucleic acid sequence encoding an Ag85B protein. (Item 2A) The method according to any one of the above items, characterized in that the nucleic acid construct is administered after viral infection. (Item 3A) The method according to any one of the above items, wherein the treatment of the viral infection substantially eliminates the virus from the subject after viral infection.(Item 4A) The method of any one of the above items, wherein the treatment for viral infection is administered for the purpose of substantially eliminating the virus from the subject after viral infection. (Item 5A) The method of any one of the above items, wherein the nucleic acid sequence encoding the antigen protein or a portion thereof is a nucleic acid sequence encoding an attenuated virus. (Item 6A) The method of any one of the above items, wherein the attenuated virus is a nef-deficient attenuated virus. (Item 7A) The method of any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is integrated into the nucleic acid sequence encoding the attenuated virus. (Item 8A) The method of any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is integrated in the position of the deleted nef gene in the nucleic acid sequence encoding the nef-deficient attenuated virus. (Item 9A) The method of any one of the above items, wherein the virus is an AIDS virus. (Item 10A) The method of any one of the above items, wherein the virus is an attenuated version of a virus that infects humans. (Item 11A) The method of any one of the above items, wherein the virus is an attenuated AIDS virus selected from the group consisting of HIV, SIV, SHIV, and FIV. (Item 12A) The method of any one of the above items, wherein the virus is an attenuated HIV. (Item 13A) The method of any one of the above items, wherein the virus is a nef-deficient attenuated HIV. (Item 14A) The method of any one of the above items, wherein the nucleic acid sequence encoding the antigenic protein or a portion thereof comprises a nucleic acid sequence at least 90% identical to a nucleic acid sequence comprising nucleotides 1 to 9633 and 10612 to 11022 of SEQ ID NO:3, or a nucleic acid sequence comprising nucleotides 1 to 8786 and 9765 to 15182 of SEQ ID NO:4. (Item 15A) The method of any one of the above items, wherein the nucleic acid construct comprises a nucleic acid sequence at least 90% identical to the nucleic acid sequence set forth in SEQ ID NO:3 or 4. (Item 16A) The method according to any one of the preceding items, wherein the nucleic acid construct comprises the nucleic acid sequence set forth in SEQ ID NO: 3 or 4.(Item 17A) The method according to any one of the preceding items, wherein the nucleic acid construct is administered once. (Item 18A) The method according to any one of the preceding items, wherein the nucleic acid construct is administered two or more times. (Item 18A-A) The method according to any one of the preceding items, wherein the nucleic acid construct is administered after administration of an anti-HIV drug. (Item 18A-B) The method of any one of the above items, wherein the anti-HIV drug is selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors, and is preferably selected from tenofovir, emtricidadine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. (Item 18A-C) The method of any one of the above items, wherein the nucleic acid construct is administered after administration of the anti-HIV drug and after discontinuing administration of the HIV drug. (Items 18A-D) The method according to any one of the above items, wherein the nucleic acid construct is administered two or more times after administration of an anti-HIV drug. (Items 18A-E) The method according to any one of the above items, wherein the nucleic acid construct is administered three, four, five or more times, or more, after administration of an anti-HIV drug. (Items 18A-F) The method according to any one of the above items, wherein the two or more administrations of the nucleic acid construct are administered at least one week apart. (Items 18A-G) The method according to any one of the above items, wherein the two or more administrations of the nucleic acid construct are administered one to four weeks apart. (Items 18A-H) The method according to any one of the above items, wherein the two or more administrations of the nucleic acid construct are administered one to three weeks apart.(Item 18A-I) The method according to any one of the preceding items, wherein the viral load in the plasma of the patient is monitored during two or more administrations of the nucleic acid construct, and when the viral load reaches a predetermined value, the nucleic acid construct is administered again. (Item 18A-J) The nucleic acid construct is administered in an amount of 5.0 x 10 per administration. ５ T.C.I.D. ５０ or more or 1.0 x 10 ６ T.C.I.D. ５０ The method according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 5.0 x 10 or more per administration. ５ T.C.I.D. ５０ or more or 1.0 x 10 ６ T.C.I.D. ５０ The method according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, or once every two months, or at intervals of one to four weeks. (Item 19A) The method according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of more than one week. (Item 20A) The method according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of more than one week. ３ T.C.I.D. ５０ The method according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 1.0 x 10 or more. ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ The method according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of less than 1.0 x 10 ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ The method according to any one of the preceding items, wherein the nucleic acid construct is administered once a week at a dose of less than 1.0 x 10 ３ T.C.I.D.５０ Above 5.0 x 10 ４ TCID ５０ The method according to any one of the preceding items, wherein the nucleic acid construct is administered once at a dose of less than 5.0 x 10, and if a virus is detected in the body, the nucleic acid construct is further administered. ４ T.C.I.D. ５０ Above 5.0 x 10 ６ T.C.I.D. ５０ The method according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 5.0 x 10 ４ T.C.I.D. ５０The method of any one of the preceding items, wherein the composition is administered once at a dose of at least 100 mg / kg / day. (Item 26A) The method of any one of the preceding items, wherein the subject has already been administered an anti-HIV drug. (Item 27A) The method of any one of the preceding items, wherein the subject has not yet been administered an anti-HIV drug, or the anti-HIV drug will be discontinued at the time of initiation of administration of the composition. (Item 28A) The method of any one of the preceding items, wherein the subject has been administered an anti-HIV drug selected from the group consisting of a reverse transcriptase inhibitor, a protease inhibitor, an integrase inhibitor, and a CCR5 inhibitor. (Item 29A) The method of any one of the above items, wherein the anti-HIV drug is selected from the group consisting of tenofovir, emtricidavine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. (Item 30A) The method of any one of the above items, wherein the composition completely eliminates the AIDS virus in the subject. (Item 31A) The method of any one of the above items, wherein the composition is not administered prior to viral infection. (Item 32A) The method of any one of the above items, wherein the subject has previously been vaccinated with a BCG vaccine. (Item 1B) Use of a nucleic acid construct operably comprising a nucleic acid sequence encoding an antigenic protein or a portion thereof contained in a virus belonging to the genus Lentivirus and a nucleic acid sequence encoding an Ag85B protein in the manufacture of a medicament for treating a viral infection in a subject. (Item 2B) The use according to the above items, characterized in that the nucleic acid construct is administered after viral infection. (Item 3B) The use according to any one of the above items, wherein the treatment of viral infection substantially eliminates the virus from the subject after viral infection.(Item 4B) The use according to any one of the above items, wherein the treatment for viral infection is administered for the purpose of substantially eliminating the virus from the subject after viral infection. (Item 5B) The use according to any one of the above items, wherein the nucleic acid sequence encoding the antigen protein or a portion thereof is a nucleic acid sequence encoding an attenuated virus. (Item 6B) The use according to any one of the above items, wherein the attenuated virus is a nef-deficient attenuated virus. (Item 7B) The use according to any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is integrated into the nucleic acid sequence encoding the attenuated virus. (Item 8B) The use according to any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is integrated in the position of the deleted nef gene in the nucleic acid sequence encoding the nef-deficient attenuated virus. (Item 9B) The use according to any one of the above items, wherein the virus is an AIDS virus. (Item 10B) The use according to any one of the above items, wherein the virus is an attenuated virus of a virus that infects humans. (Item 11B) The use according to any one of the above items, wherein the virus is an attenuated AIDS virus selected from the group consisting of HIV, SIV, SHIV, and FIV. (Item 12B) The use according to any one of the above items, wherein the virus is an attenuated HIV. (Item 13B) The use according to any one of the above items, wherein the virus is a nef-deficient attenuated HIV. (Item 14B) The use according to any one of the above items, wherein the nucleic acid sequence encoding the antigenic protein or a portion thereof comprises a nucleic acid sequence at least 90% identical to a nucleic acid sequence comprising nucleotides 1 to 9633 and 10612 to 11022 of SEQ ID NO:3, or a nucleic acid sequence comprising nucleotides 1 to 8786 and 9765 to 15182 of SEQ ID NO:4. (Item 15B) The use according to any one of the above items, wherein the nucleic acid construct comprises a nucleic acid sequence at least 90% identical to the nucleic acid sequence set forth in SEQ ID NO:3 or 4. (Item 16B) The use according to any one of the preceding items, wherein the nucleic acid construct comprises the nucleic acid sequence set forth in SEQ ID NO: 3 or 4.(Item 17B) The use according to any one of the preceding items, wherein the nucleic acid construct is administered once. (Item 18B) The use according to any one of the preceding items, wherein the nucleic acid construct is administered two or more times. (Item 18B-A) The use according to any one of the preceding items, wherein the nucleic acid construct is administered after administration of an anti-HIV drug. (Item 18B-B) The use according to any one of the preceding items, wherein the anti-HIV drug is selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors, and is preferably selected from tenofovir, emtricidadine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. (Item 18B-C) The use according to any one of the preceding items, wherein the nucleic acid construct is administered after administration of the anti-HIV drug and after discontinuing administration of the HIV drug. (Item 18B-D) The use according to any one of the preceding items, wherein the nucleic acid construct is administered two or more times after administration of an anti-HIV drug. (Item 18B-E) The use according to any one of the preceding items, wherein the nucleic acid construct is administered three, four, five or more times, or more, after administration of an anti-HIV drug. (Item 18B-F) The use according to any one of the preceding items, wherein the two or more administrations of the nucleic acid construct are administered at least one week apart. (Item 18B-G) The use according to any one of the preceding items, wherein the two or more administrations of the nucleic acid construct are administered one to four weeks apart. (Item 18B-H) The use according to any one of the preceding items, wherein the two or more administrations of the nucleic acid construct are administered one to three weeks apart.(Item 18B-I) The use according to any one of the preceding items, wherein the nucleic acid construct is administered two or more times while monitoring the viral load in the patient's plasma, and is administered again when the viral load reaches a predetermined value. (Item 18B-J) The nucleic acid construct is administered in an amount of 5.0 x 10 per administration. ５ T.C.I.D. ５０ or more or 1.0 x 10 ６ T.C.I.D. ５０ The use according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 5.0 x 10 or more per administration. ５ T.C.I.D. ５０ or more or 1.0 x 10 ６ T.C.I.D. ５０ The use according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, or once every two months. (Item 19B) The use according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of once to four weeks. (Item 19B-A) The use according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of once or more. (Item 20B) The nucleic acid construct is administered at a dose of 1.0 x 10 ３ T.C.I.D. ５０ The use according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 1.0 x 10 or more. ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ The use according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of less than 1.0 x 10 ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ The use according to any one of the preceding items, wherein the nucleic acid construct is administered once a week at a dose of less than 1.0 x 10 ３ T.C.I.D.５０ Above 5.0 x 10 ４ TCID ５０ The use according to any one of the preceding items, wherein the nucleic acid construct is administered once at a dose of less than 5.0 x 10, and if a virus is detected in the body, the nucleic acid construct is further administered. ４ T.C.I.D. ５０ Above 5.0 x 10 ６ T.C.I.D. ５０ The use according to any one of the preceding items, characterized in that the nucleic acid construct is administered at a dose of 5.0 x 10 ４ T.C.I.D. ５０The use according to any one of the preceding items, wherein the composition is administered once at a dose of at least 100 mg / kg / day. (Item 26B) The use according to any one of the preceding items, wherein the subject is already receiving an anti-HIV drug. (Item 27B) The use according to any one of the preceding items, wherein the subject has not yet received an anti-HIV drug, or the anti-HIV drug will be discontinued when administration of the composition is initiated. (Item 28B) The use according to any one of the preceding items, wherein the subject is receiving an anti-HIV drug selected from the group consisting of a reverse transcriptase inhibitor, a protease inhibitor, an integrase inhibitor, and a CCR5 inhibitor. (Item 29B) The use of any one of the above items, wherein the anti-HIV drug is an anti-HIV drug selected from the group consisting of tenofovir, emtricidavine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. (Item 30B) The use of any one of the above items, wherein the composition completely eliminates the AIDS virus in the subject. (Item 31B) The use of any one of the above items, wherein the composition has not been administered prior to viral infection. (Item 32B) The use according to any one of the preceding items, wherein the subject has previously been vaccinated with a BCG vaccine. (Item 1C) A nucleic acid construct for treating a viral infection in a subject, operably comprising a nucleic acid sequence encoding an antigenic protein or a part thereof contained in a virus belonging to the genus Lentivirus and a nucleic acid sequence encoding an Ag85B protein. (Item 2C) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered after viral infection. (Item 3C) The nucleic acid construct according to any one of the preceding items, wherein the treatment of viral infection substantially eliminates the virus from the subject after viral infection.(Item 4C) The nucleic acid construct of any one of the above items, wherein the treatment for viral infection is administered for the purpose of substantially eliminating the virus from the subject after viral infection. (Item 5C) The nucleic acid construct of any one of the above items, wherein the nucleic acid sequence encoding the antigen protein or a portion thereof is a nucleic acid sequence encoding an attenuated virus. (Item 6C) The nucleic acid construct of any one of the above items, wherein the attenuated virus is a nef-deficient attenuated virus. (Item 7C) The nucleic acid construct of any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is integrated into the nucleic acid sequence encoding the attenuated virus. (Item 8C) The nucleic acid construct of any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is integrated in the position of the deleted nef gene in the nucleic acid sequence encoding the nef-deficient attenuated virus. (Item 9C) The nucleic acid construct of any one of the above items, wherein the virus is an AIDS virus. (Item 10C) The nucleic acid construct of any one of the above items, wherein the virus is an attenuated virus of a virus that infects humans. (Item 11C) The nucleic acid construct of any one of the above items, wherein the virus is an attenuated AIDS virus selected from the group consisting of HIV, SIV, SHIV, and FIV. (Item 12C) The nucleic acid construct of any one of the above items, wherein the virus is an attenuated HIV. (Item 13C) The nucleic acid construct of any one of the above items, wherein the virus is a nef-deficient attenuated HIV. (Item 14C) The nucleic acid construct of any one of the above items, wherein the nucleic acid sequence encoding the antigenic protein or a portion thereof comprises a nucleic acid sequence at least 90% identical to a nucleic acid sequence comprising nucleotides at positions 1 to 9633 and 10612 to 11022 of SEQ ID NO:3, or a nucleic acid sequence comprising nucleotides at positions 1 to 8786 and 9765 to 15182 of SEQ ID NO:4. (Item 15C) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct comprises a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence set forth in SEQ ID NO: 3 or 4.(Item 16C) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct comprises the nucleic acid sequence set forth in SEQ ID NO: 3 or 4. (Item 17C) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered once. (Item 18C) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered two or more times. (Item 18C-A) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered after administration of an anti-HIV drug. (Item 18C-B) The nucleic acid construct according to any one of the preceding items, wherein the anti-HIV drug is selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors, and is preferably tenofovir, emtricidadine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. (Item 18C-C) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered after administration of an anti-HIV drug and then discontinuing administration of the HIV drug. (Items 18C-D) The nucleic acid construct of any one of the above items, wherein the nucleic acid construct is administered two or more times after administration of an anti-HIV drug. (Items 18C-E) The nucleic acid construct of any one of the above items, wherein the nucleic acid construct is administered three, four, five or more times, or more, after administration of an anti-HIV drug. (Items 18C-F) The nucleic acid construct of any one of the above items, wherein the two or more administrations of the nucleic acid construct are administered at intervals of at least one week or more. (Items 18C-G) The nucleic acid construct of any one of the above items, wherein the two or more administrations of the nucleic acid construct are administered at intervals of one to four weeks.(Item 18C-H) The nucleic acid construct according to any one of the preceding items, wherein the two or more administrations of the nucleic acid construct are administered at intervals of 1 to 3 weeks. (Item 18C-I) The nucleic acid construct according to any one of the preceding items, wherein the viral load in the patient's plasma is monitored during the two or more administrations of the nucleic acid construct, and the nucleic acid construct is administered again when it reaches a predetermined value. (Item 18C-J) The nucleic acid construct is administered at a dose of 5.0 x 10 per administration. ５ T.C.I.D. ５０ or more or 1.0 x 10 ６ T.C.I.D. ５０ The nucleic acid construct according to any one of the preceding items is administered at a dose of 5.0 x 10 or more. ５ T.C.I.D. ５０ or more or 1.0 x 10 ６ T.C.I.D. ５０ The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, or once every two months. (Item 19C) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of once to four weeks. (Item 19C-A) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, once every two months, or at intervals of once or more. (Item 20C) The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 1.0 x 10 ３ T.C.I.D. ５０ The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 1.0 x 10 or more. ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of less than 1.0 x 10 ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID５０ The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered once a week at a dose of less than 1.0 x 10 ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered once at a dose of less than 5.0 x 10, and if a virus is detected in the body, the nucleic acid construct is further administered. ４ T.C.I.D. ５０ Above 5.0 x 10 ６ T.C.I.D. ５０ The nucleic acid construct according to any one of the preceding items, wherein the nucleic acid construct is administered at a dose of 5.0 x 10 ４ T.C.I.D. ５０The nucleic acid construct of any one of the preceding items, wherein the subject is administered a single time at a dose of at least 100 mg / kg / day. (Item 26C) The nucleic acid construct of any one of the preceding items, wherein the subject has already been administered an anti-HIV drug. (Item 27C) The nucleic acid construct of any one of the preceding items, wherein the subject has not yet been administered an anti-HIV drug, or the anti-HIV drug will be discontinued at the time of initiation of administration of the composition. (Item 28C) The nucleic acid construct of any one of the preceding items, wherein the subject has been administered an anti-HIV drug selected from the group consisting of a reverse transcriptase inhibitor, a protease inhibitor, an integrase inhibitor, and a CCR5 inhibitor. (Item 29C) The nucleic acid construct of any one of the preceding items, wherein the anti-HIV drug is an anti-HIV drug selected from the group consisting of tenofovir, emtricidavine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. (Item 30C) The nucleic acid construct of any one of the preceding items, wherein the composition completely eliminates the AIDS virus in the subject. (Item 31C) The nucleic acid construct of any one of the preceding items, wherein the composition has not been administered prior to viral infection. (Item 32C) The nucleic acid construct of any one of the preceding items, wherein the subject has previously been vaccinated with a BCG vaccine. (Item 1D) The composition, nucleic acid molecule, method, use or nucleic acid construct of any one of the preceding items, wherein the composition, nucleic acid molecule, method, use or nucleic acid construct is for treating infection with a virus belonging to the lentivirus genus, preferably an AIDS virus, or a viral infection caused by such a virus.

[0006] It is contemplated that the present disclosure may provide one or more of the above-described features in combinations other than those explicitly stated. Still further embodiments and advantages of the present disclosure will be recognized by those skilled in the art upon reading and understanding the following detailed description, if necessary.

[0007] Figure 1 shows an outline of the experiment in Example 3. Figure 2 shows the plasma viral load and CD4 count in monkeys (#137) administered only an anti-HIV drug. ＋ Figure 3 shows the dynamics of the plasma viral load and CD4 T cell count in monkeys (#139) administered only anti-HIV drugs. ＋ Figure 4 shows the dynamics of T cell counts. Plasma viral load and CD4 counts in a monkey (#142) administered two doses of low-dose SHIV-Ag85B. ＋ Figure 5 shows the dynamics of T cell counts. Plasma viral load and CD4 counts in a monkey (#141) administered a single high-dose SHIV-Ag85B. ＋ Figure 6 shows the dynamics of T cell counts. Plasma viral load and CD4 counts in a monkey (#140) administered multiple doses of high-dose SHIV-Ag85B. ＋ Figure 7 shows the dynamics of T cell counts. Plasma viral load and CD4 counts in monkeys (#138) administered a single dose of low-dose and high-dose SHIV-Ag85B, respectively. ＋The dynamics of T cell counts are shown. Figure 8 shows the administration schedule for each individual in Example 2. Figure 9A shows the time course of plasma viral load in a monkey (#138) administered two doses of low-dose SHIV-Ag85B in Example 3. Figure 9B shows the time course of plasma viral load in a monkey (#142) administered a single dose of high-dose SHIV-Ag85B in Example 3. Figure 10 shows that in one monkey (#141) administered a high dose of SHIV-Ag85B in Example 3, only a short-term effect was observed initially, but when the drug was administered whenever virus appeared in the plasma, a long-term suppressive effect was observed from the fifth inoculation onwards. Figure 11A shows the results for an untreated monkey (#137) administered only an anti-HIV drug in Example 4. Figure 11B shows the results for an untreated monkey (#139) administered only an anti-HIV drug in Example 4. Fig. 12 shows an experimental protocol for verifying the change in plasma viral load when the composition according to the present disclosure is administered after inoculation with HIV virus in Example 4. Fig. 13 shows the results in Example 4 showing that the anti-HIV drug is effective regardless of individual differences. Fig. 14 shows the results of administering a high dose (5.0 x 10 ４ T.C.I.D. ５０ 15 shows the plasma viral load after four administrations of high-dose SHIV-Ag85B at weekly intervals in monkeys (#164, #165, and #167) administered with SHIV-Ag85B at a high dose (5.0 × 10 ４ T.C.I.D. ５０ 16 shows the results of plasma viral loads in monkeys (#162, #166, and #168) administered with SHIV-NI (100 mg / kg / day). Figure 16 shows the results of plasma viral loads in the control group not containing Ag85B in Example 5, two weeks after the drug withdrawal.

[0008] The present disclosure will be described below. Throughout this specification, singular expressions should be understood to include the plural concept unless otherwise specified. Therefore, singular articles (e.g., "a," "an," "the," etc. in English) should be understood to include the plural concept unless otherwise specified. Furthermore, it should be understood that terms used in this specification are used in the sense commonly used in the relevant field unless otherwise specified. Therefore, unless otherwise defined, all technical and scientific terms used in this specification have the same meaning as commonly understood by those skilled in the art to which this disclosure belongs. In the event of conflict, the present specification (including definitions) will prevail.

[0009] (Definition of Terms) The definitions of terms used particularly in this specification are listed below.

[0010] As used herein, "about" means ±10% of the indicated value.

[0011] As used herein, "Ag85B" refers to an immunogenic protein secreted by acid-fast bacteria. Acid-fast bacteria are bacteria that are resistant to acid when stained with a dye, meaning that the dye is not bleached by acid. Acid-fast bacteria (mycobacteria) are broadly classified into Mycobacterium tuberculosis, Mycobacterium leprae, and non-tuberculous acid-fast bacteria. A typical source of Ag85B is Mycobacterium tuberculosis. Ag85B includes antigen 85-B, 85B, extracellular alpha-antigen, antigen 85 complex B, Ag85B, mycolyl transferase 85B, EC 2.3.1. It is also known as Fibronectin-binding protein B, 30 kDa extracellular protein, fbpB, A85B, Major Secretary Protein Antigen 85B, etc. A representative accession number is Q847N4. See https: / / www.uniprot.org / uniprotkb / Q847N4 / entry. A representative nucleic acid sequence ID is AY207396g, and a representative protein sequence ID is AAO62005.1. Although shown herein as SEQ ID NO: 1 (nucleic acid sequence) and SEQ ID NO: 2 (amino acid sequence), it is understood that any immunogenic protein secreted by acid-fast bacteria is within the scope of the present disclosure, without being limited thereto.

[0012] As used herein, "operably" refers to a state in which the transcription or translation of a nucleic acid sequence is under the control of an expression control element, the transcription or translation of the nucleic acid sequence is appropriately controlled, and functional expression is achieved.

[0013] As used herein, the term "Th1-type immune response" refers to a cellular immune response mediated by T lymphocytes, and a response resulting from cytokines and / or chemokines produced by activated T cells.

[0014] As used herein, "Th2-type immune response" refers to a humoral immune response mediated by secretory antibodies produced by B cells.

[0015] As used herein, the term "attenuated virus" refers to a virus that has reduced pathogenicity compared to the parent virus but retains the ability to induce an immune response. Whether or not a virus is attenuated can be confirmed by its toxicity to cells and / or pathogenicity to animals. It is preferably determined through animal experiments, etc. In general, the term "attenuation" refers to artificially reducing the toxicity of a pathogen by mutating a gene so that the pathogen loses pathogenicity but retains immunogenicity. Attenuation is generally achieved by UV irradiation, chemical treatment, or in vitro serial high-passage culture, which reduces pathogen toxicity. Toxicity is reduced by artificially modifying a gene, for example, by deleting specific nucleotides in a known sequence.

[0016] As used herein, the term "AIDS virus" refers to a virus that causes acquired immunodeficiency syndrome (AIDS), and includes human immunodeficiency virus (HIV), simian immunodeficiency virus (SIV), simian / human immunodeficiency (chimeric) virus (SHIV), feline immunodeficiency virus (FIV), etc.

[0017] As used herein, "simian / human immunodeficiency (chimeric) virus (SHIV)" refers to a simian / human immunodeficiency chimeric virus in which at least some of the SIV genes have been replaced with those of HIV-1. In addition to the env gene, the vpr, rev, tat, and vpu genes of SHIV may also be replaced with those of HIV-1.

[0018] Nef (negative factor) is a protein of approximately 27 kDa that is encoded on the 3' side of the HIV-1 gene. Originally reported as a factor that suppresses viral proliferation, it has since been shown to enhance viral proliferation in primary cultured cells and in vivo, and is believed to be related to HIV-1 pathogenicity. HIV-1 Nef protein, anchor domain superfamily (IPR027480), HIV-1 Nef protein, core domain superfamily (IPR027481), etc. are known. Although various vaccine viruses attenuated by genetic disruption of key regulatory genes such as nef, vpx, vpr, and vif have been used, most studies have used the moderately attenuated prototype vaccine strain SIVmac239Δnef. Nef-deficient attenuated simian immunodeficiency virus (SIV) and simian-human immunodeficiency virus (SHIV) have proven highly effective as vaccines in nonhuman primate models, they are not sufficiently safe to be used as templates for HIV vaccines in humans. While several studies have demonstrated that inserting cytokine or chemokine genes into attenuated, genetically deleted SIV or SHIV improves immunogenicity and enhances viral defense compared to safer, less toxic strains, the present disclosure has unexpectedly found that the nef-deficient attenuated HIV-Ag85B can, upon administration after infection, substantially eliminate the virus in patients already infected with the virus present in their bodies, thereby curing the disease to a state in which the virus is completely eliminated from the body (typically, to a state in which the virus is below the limit of detection by a highly sensitive detection method such as PCR and no clinical symptoms are observed). Furthermore, the present disclosure has been found to provide a therapeutic composition for HIV that is practically safe and less toxic to infected subjects, and can effectively induce a Th1-type immune response.

[0019] The SIV infection model in non-human primates used herein is important for analyzing the mechanism of acquired immunodeficiency syndrome (AIDS) and determining the effectiveness of HIV vaccines or HIV therapeutic interventions. As reported by the present inventors, SIVmac and SHIV89.6P are pathogenic in cynomolgus macaques from Indonesia, Malaysia, and the Philippines. Plasma viral load, peripheral CD4 ＋ Comparison of parameters such as T cell counts, patterns of viral antigen-specific immune responses, and disease outcome has shown that SIV and SHIV can cause disease in cynomolgus monkeys of different origins. Compared with Indian rhesus macaques, cynomolgus macaques of Asian origin are similarly tolerant to various strains of SIVmac and SHIV and have longer survival times after SIV or SHIV infection, indicating that cynomolgus macaques are a model that closely resembles the human course of HIV-1 disease.

[0020] As used herein, "complete elimination of the virus" refers to a state in which the virus has been completely removed from the living body. A state in which the virus has been completely removed from the living body means that the virus is below the detection limit of a highly sensitive detection method such as PCR, and no clinical symptoms are observed.

[0021] As used herein, the term "vaccine" refers to a substance containing or encoding an antigen that provides active immunity against a substance containing the antigen but does not cause disease. As used herein, the term "vaccine" refers to a substance used prophylactically. As used herein, the term "DNA vaccine" refers to a nucleic acid encoding a vaccine antigen, and is generally used in the form of DNA (particularly, plasmid DNA), hence the general term. In some embodiments, nucleic acids are incorporated into viral vectors or the like for delivery into the body, and in such cases, it is understood that the vaccine may be provided in a nucleic acid form other than DNA. In such cases, the vaccine is also referred to as a "nucleic acid vaccine." DNA vaccines typically take the form of plasmid DNA, and when administered subcutaneously in the form of plasmid DNA, the plasmid DNA is taken up by subcutaneous cells and produces the target antigen protein within those cells.

[0022] As used herein, the term "antigen" (also referred to as Ag) refers to any substance that can be specifically bound by an antibody molecule. As used herein, the term "immunogen" refers to an antigen that can initiate lymphocyte activation resulting in an antigen-specific immune response.

[0023] As used herein, "remission" refers to a state in which the virus has been completely eliminated and the patient is healthy without the need for administration of therapeutic drugs.

[0024] As used herein, "treating" a "viral infection" refers to reducing at least one symptom of the viral infection, slowing the progression of at least one symptom, or ameliorating at least one symptom.

[0025] As used herein, "substantially eliminating the virus" means that the virus is clinically undetectable and it is determined that the disease has not developed.

[0026] As used herein, the term "subject" refers to a mammal, including a human patient, who is a target of the treatments of the present disclosure.

[0027] As used herein, the term "nucleic acid construct" refers to a DNA or RNA molecule that comprises a nucleotide sequence that encodes a protein.

[0028] As used herein, the term "deficiency" refers to the loss of a gene. Deficiency also includes the loss of a part of a gene, which is therefore non-functional.

[0029] As used herein, "X number of doses" refers to X number of doses administered during the observation period.

[0030] As used herein, "TCID" ５０ " refers to 50% tissue culture infectious dose, which means the amount of virus that infects 50% of cells.

[0031] As used herein, the phrase "already administered with an anti-HIV drug" refers to administration of an anti-HIV drug before administration of the nucleic acid construct of the present disclosure.

[0032] As used herein, the phrase "has not yet been administered an anti-HIV drug" refers to the fact that an anti-HIV drug has not been administered at the time the nucleic acid construct of the present disclosure is administered.

[0033] As used herein, the term "reverse transcriptase inhibitor" refers to a drug that blocks the enzymatic function of reverse transcriptase, prevents the synthesis of double-stranded viral DNA, and prevents the proliferation of HIV.

[0034] As used herein, the term "protease inhibitor" refers to a drug that binds to the enzymatic active site of a protease and eliminates its activity. HIV functional proteins are first produced as complex proteins, and only function after being cleaved at a specific site by HIV's own protease. Protease inhibitors eliminate the activity of the protease, preventing the virus from reaching its mature form and making it infective.

[0035] As used herein, the term "integrase inhibitor" refers to a drug that inhibits integrase, thereby inhibiting the reaction in which HIV integrates its own genes into the host chromosome, thereby preventing infection. This prevents the production of new infectious virus particles and therefore prevents the spread of infection.

[0036] As used herein, the term "CCR5 inhibitor" refers to a drug that inhibits CC chemokine receptor 5 (CCR5), a coreceptor used by HIV when it enters cells.

[0037] As used herein, "complete elimination of the AIDS virus" refers to the determination that viruses such as the AIDS virus cannot be detected clinically or by any other detection method.

[0038] As used herein, "nucleic acid construct," "construct," "construct" or "gene construct" are used interchangeably and refer to a nucleic acid molecule that comprises a collection of nucleic acids that are isolated from naturally occurring genes or that are combined and juxtaposed in a manner that does not occur in nature.

[0039] Amino acids may be referred to herein by either their commonly known three-letter symbols or the one-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Nucleotides may also be referred to by their commonly accepted one-letter codes. Herein, comparisons of similarity, identity, and homology between amino acid sequences and base sequences are calculated using the sequence analysis tool BLAST with default parameters. Identity searches can be performed, for example, using NCBI's BLAST 2.2.28 (published April 2, 2013) (Proc. Natl. Acad. Sci. USA 90:5873-5877, 1993). Identity values ​​herein generally refer to values ​​obtained when aligned using the above-mentioned BLAST under default conditions. However, if a higher value is obtained by changing the parameters, the highest value is used as the identity value. When identity is evaluated in multiple regions, the highest value among them is used as the identity value. Similarity is a numerical value that takes into account not only identity but also similar amino acids. When comparing amino acid sequences with BLAST, the algorithm Blastp can be used with default settings. Measurement results are quantified as Positives or Identities. The homology of amino acid sequences or nucleotide sequences can be determined using the BLAST algorithm by Karlin and Altschul. Based on this algorithm, programs called BLASTN and BLASTX have been developed (Altschul et al. J. Mol. Biol. 215:403-410, 1990). When analyzing nucleotide sequences using BLASTN based on BLAST, parameters are set, for example, as score = 100 and wordlength = 12. When analyzing amino acid sequences using BLASTX based on BLAST, parameters are set, for example, as score = 50 and wordlength = 3. When using BLAST and Gapped BLAST programs, the default parameters of each program are used. Specific techniques for these analysis methods are known (http: / / www.ncbi.nlm.nih.gov.).

[0040] The nucleic acids or proteins used in the present disclosure may include sequences in which one or more amino acids or nucleotides have been substituted, deleted, and / or added to the target amino acid or base sequence. Here, "one or more" in the full-length amino acid sequence of the chimeric protein typically refers to 50 amino acids or less, preferably 30 amino acids or less, and more preferably 10 amino acids or less (e.g., 5 amino acids or less, 3 amino acids or less, or 1 amino acid or less). Furthermore, in the amino acid sequence of a domain, "one or more" typically refers to 6 amino acids or less, preferably 5 amino acids or less, and more preferably 4 amino acids or less (e.g., 3 amino acids or less, 2 amino acids or less, or 1 amino acid or less). To maintain the biological activity of the chimeric protein of the present disclosure, it is desirable that the mutated amino acid residue be mutated to another amino acid whose amino acid side chain properties are conserved. For example, the properties of the amino acid side chains include hydrophobic amino acids (A, I, L, M, F, P, W, Y, V), hydrophilic amino acids (R, D, N, C, E, Q, G, H, K, S, T), amino acids with aliphatic side chains (G, A, V, L, I, P), amino acids with hydroxyl-containing side chains (S, T, Y), amino acids with sulfur-containing side chains (C, M), amino acids with carboxylic acid- and amide-containing side chains (D, N, E, Q), amino acids with base-containing side chains (R, K, H), and amino acids with aromatic-containing side chains (H, F, Y, W) (the characters in parentheses represent the single-letter symbols of the amino acids). These are also referred to herein as "conservative substitutions." It is known that proteins having an amino acid sequence modified by deletion, addition, and / or substitution of one or more amino acid residues with other amino acids maintain their biological activity (Mark, D.F. et al., Proc. Natl. Acad. Sci. USA (1984) 81, 5662-5666; Zoller, M.J. & Smith, M. Nucleic Acids Research (1982) 10, 6487-6500; Wang, A. et al., Science 224, 1431-1433; Dalbadie-McFarland, G. et al., Proc. Natl. Acad. Sci. USA (1982) 79, 6409-6413).Therefore, in one embodiment of the present disclosure, "several" may be, for example, 10, 8, 6, 5, 4, 3, or 2, or any of these values ​​or less. Chimeric proteins with deletions or other modifications can be prepared, for example, by site-directed mutagenesis, random mutagenesis, or biopanning using an antibody phage library. In the present disclosure, "70% or more" may be, for example, 70, 75, 80, 85, 90, 95, 96, 97, 98, or 99% or more, "80% or more" may be, for example, 80, 85, 90, 95, 96, 97, 98, or 99% or more, and "90% or more" may be, for example, 90, 95, 96, 97, 98, or 99% or more, or may be within a range of any two of these values. "Homology" may be calculated by methods known in the art as the percentage of homologous amino acids in two or more amino acid sequences. Before calculating the percentage, the amino acid sequences of the amino acid sequences to be compared are aligned, and gaps are introduced into some of the amino acid sequences if necessary to maximize the percentage of identical amino acids. Methods for alignment, percentage calculation, comparison, and related computer programs are well known in the art (e.g., BLAST, GENETYX, etc.). In the case of "identity," the percentage of identical amino acids is calculated, and in the case of "similarity," the percentage of similar amino acids is calculated. Similar amino acids include, but are not limited to, amino acids that can be conservatively substituted.

[0041] Portions of the constructs specifically disclosed herein are also encompassed within the scope of the present disclosure. As used herein, the terms "portion," "fragment," or "fragment" refer to a polypeptide or polynucleotide having a sequence length of 1 to n-1 relative to the full-length polypeptide or polynucleotide (length n). The length of the fragment can be varied appropriately depending on the purpose. For example, the lower limit of the length for a polypeptide can be 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 40, 50, or more amino acids, and lengths represented by integers not specifically recited herein (e.g., 11) may also be suitable as lower limits. Furthermore, for polynucleotides, the lower limit can be 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 40, 50, 75, 100, or more nucleotides, and lengths represented by integers not specifically recited herein (e.g., 11) may also be suitable as lower limits. As used herein, such fragments are understood to fall within the scope of the present disclosure, for example, if the full-length fragment functions as a vaccine, as long as the fragment itself also functions as a vaccine. This disclosure also encompasses "functional equivalents" of those specifically disclosed herein (constructs, etc.). As used herein, the term "functional equivalent" refers to any entity that has the same intended function but a different structure compared to the original entity of interest.

[0042] In accordance with the present disclosure, the term "activity" as used herein refers to the function of a molecule in the broadest sense. Activity generally includes, but is not limited to, the biological, biochemical, physical, or chemical function of a molecule. Activity includes, for example, enzymatic activity, the ability to interact with other molecules, and the ability to activate, promote, stabilize, inhibit, suppress, or destabilize the function of other molecules, stability, and the ability to localize to a specific subcellular location. Where applicable, the term also relates to the function of a protein complex in the broadest sense. As used herein, "biological activity" includes, for example, activation of a light reaction.

[0043] As used herein, the term "functional equivalent" refers to any entity that has the same intended function as the original entity but a different structure. Therefore, functional equivalents of "HIV-encoded proteins" or chimeras thereof are not HIV-encoded proteins or chimeras themselves, but include mutants or variants (e.g., amino acid sequence variants, etc.) of HIV-encoded proteins or chimeras that have the biological activity of HIV-encoded proteins or chimeras, as well as those that can be transformed into HIV-encoded proteins or antibodies thereof or mutants or variants of HIV-encoded proteins or chimeras at the time of action (e.g., nucleic acids encoding HIV-encoded proteins or chimeras, or HIV-encoded proteins or chimeric mutants or variants, and vectors, cells, etc. containing such nucleic acids). Functional equivalents of the present disclosure can include insertions, substitutions, and / or deletions of one or more amino acids in the amino acid sequence, or additions to one or both termini. As used herein, the phrase "insertion, substitution, and / or deletion of one or more amino acids in an amino acid sequence, or addition to one or both termini thereof" means that the amino acid sequence has been modified by a well-known technical method such as site-directed mutagenesis, or by natural mutation, resulting in the substitution, etc., of a number of amino acids to the extent that would occur naturally. The modified amino acid sequence may be one in which, for example, 1 to 30, preferably 1 to 20, more preferably 1 to 9, even more preferably 1 to 5, and particularly preferably 1 to 2 amino acids have been inserted, substituted, or deleted, or added to one or both termini thereof. The modified amino acid sequence may preferably be an amino acid sequence having one or more (preferably one or several, or 1, 2, 3, or 4) conservative substitutions in the amino acid sequence of a protein encoded by the HIV virus.

[0044] As used herein, "treatment" refers to preventing, preferably maintaining, more preferably alleviating, and even more preferably eliminating a certain disease or disorder (e.g., caused by a viral infection) from worsening when that condition occurs, and includes the possibility of exerting an effect of improving symptoms of the patient's disease or one or more symptoms associated with the disease. Preliminary diagnosis followed by appropriate treatment is called "companion treatment," and diagnostic agents used for this purpose are sometimes called "companion diagnostic agents."

[0045] As used herein, the term "therapeutic agent" broadly refers to any drug capable of treating a target condition (e.g., retinal degenerative disease, etc.). In one embodiment of the present disclosure, the "therapeutic agent" may be a pharmaceutical composition containing an active ingredient and one or more pharmacologically acceptable carriers. The pharmaceutical composition can be prepared, for example, by mixing the active ingredient with the carrier using any method known in the technical field of pharmaceuticals. Furthermore, the therapeutic agent may be in any form used for treatment, and may be the active ingredient alone or a mixture of the active ingredient with any other ingredient. Furthermore, the shape of the carrier is not particularly limited, and may be, for example, a solid or liquid (e.g., a buffer solution).

[0046] The present disclosure may be provided as a kit. As used herein, the term "kit" refers to a unit in which the components to be provided (e.g., multiple nucleic acid constructs, lyophilized drugs and administration buffers, instructions, etc.) are provided, usually in two or more compartments. Such kits preferably advantageously include instructions or manuals describing how to use the components to be provided (e.g., nucleic acid constructs) or how to handle reagents. The present specification also includes instructions describing how to use the kit.

[0047] As used herein, the term "active ingredient" refers to an ingredient contained in a composition, etc. of the present disclosure in an amount necessary to achieve the intended effect of treatment or progression inhibition, and other ingredients may also be contained as long as the effect is not impaired to below the desired level. Furthermore, the medicament, composition, etc. of the present disclosure may be formulated. Furthermore, the route of administration of the medicament, composition, etc. of the present disclosure may be either oral or parenteral, and can be appropriately determined depending on the form of the formulation, etc. The construct of the present disclosure may be used as an active ingredient.

[0048] As used herein, "instructions" (including package inserts and labels used by the U.S. FDA) are written instructions to a physician or other user on how to use the present disclosure. The instructions include instructions for administering the medicament or the like of the present disclosure. The instructions may also include instructions for intravenous administration (e.g., by injection) as the administration site. The instructions are prepared in accordance with a format specified by the regulatory agency of the country in which the present disclosure is implemented (e.g., the Ministry of Health, Labor and Welfare in Japan, the Food and Drug Administration (FDA) in the United States, etc.), and clearly state that they have been approved by the regulatory agency. The instructions are so-called package inserts or labels, and are typically provided in paper form, but are not limited thereto, and may also be provided in the form of electronic media (e.g., a website provided on the Internet, a PDF, or email).

[0049] (Preferred Embodiments) A description of preferred embodiments will be given below, but it should be understood that these embodiments are merely examples of the present disclosure and that the scope of the present disclosure is not limited to such preferred embodiments. It should also be understood that those skilled in the art can easily make modifications, changes, etc. within the scope of the present disclosure by referring to the following preferred examples. Those skilled in the art can combine any of these embodiments as appropriate.

[0050] In one aspect, the present disclosure provides a composition for treating a viral infection in a subject, the composition comprising a nucleic acid construct operably comprising a nucleic acid sequence encoding an antigenic protein or a portion thereof contained in a virus belonging to the Lentivirus genus, and a nucleic acid sequence encoding an Ag85B protein, as well as methods, uses, and the like related thereto.

[0051] In one preferred embodiment, the subject of the present disclosure is a subject following a viral infection, and the composition of the present disclosure is administered following the infection.

[0052] In preferred embodiments, the treatment of a viral infection of the present disclosure results in the substantial elimination, preferably complete elimination, of the virus from said subject following viral infection.

[0053] Here, "substantially eliminating" the virus means that the AIDS virus or other virus cannot be detected clinically and it is determined that the disease has not developed.

[0054] Here, "complete disappearance" of the virus means that the AIDS virus or other viruses cannot be detected by any detection method (for example, even by highly sensitive PCR tests), and no clinical symptoms are observed.

[0055] Thus, in a preferred embodiment, the present disclosure provides a composition that is administered to a virally infected subject for the purpose of substantially eliminating the virus.

[0056] In one embodiment, the nucleic acid sequence encoding the antigen protein or a portion thereof used is a nucleic acid sequence encoding an attenuated virus, and preferably the attenuated virus is a nef-deficient attenuated virus. Since the nef gene is a gene that controls the immune evasion of the AIDS virus, deleting this gene attenuates the virus through the immune system of the host. Conventionally, attenuated viruses have not been considered for use with such adjuvant antigens.

[0057] In one embodiment, the nucleic acid sequence encoding the Ag85B protein used in the present disclosure is incorporated into a nucleic acid sequence encoding the attenuated virus of the present disclosure. Because the nef gene is a gene that controls immune evasion by the AIDS virus, deleting this gene attenuates the virus through the body's immune system. Addition of Ag85B induces strong cellular immunity, inducing a strong immune response and further attenuating the virus.

[0058] In one embodiment, the nucleic acid sequence encoding the Ag85B protein used in the present disclosure is integrated into the nucleic acid sequence encoding the nef-deficient attenuated virus at the location of the deleted nef gene. Because the nef gene is a gene that controls the immune evasion of the AIDS virus, deleting this portion attenuates the virus through the body's immune function. Addition of Ag85B induces strong cellular immunity, inducing a strong immune response and further attenuating the virus.

[0059] In one embodiment, the virus used is the AIDS virus.

[0060] In one embodiment, the virus used is an attenuated version of a virus that infects humans.

[0061] In one embodiment, the virus used is an attenuated form of the AIDS virus selected from the group consisting of human immunodeficiency virus (HIV), simian immunodeficiency virus (SIV), simian / human immunodeficiency chimeric virus (SHIV), and feline immunodeficiency virus (FIV).

[0062] In a preferred embodiment, the virus is an HIV virus.

[0063] Human immunodeficiency virus (HIV) is genetically classified into two main types: type I HIV (HIV-1) and type II HIV (HIV-2). HIV-1 is the most prevalent type in the world, including Japan, and is further classified into three types: group M (Main), which accounts for the majority of cases, and groups O (Outlier) and N (New), which are found in limited areas. Group M is further classified into nine subtypes (A-D, F-H, J, and K). On the other hand, HIV-2 is classified into subtypes A-G, and has been found mainly in West Africa, but cases of infection have also been reported in France, the United States, the West Indies, and South Korea.

[0064] In certain embodiments, the virus is a strain of HIV-1, including, but not limited to, strains BaL, IIIB, RF, GB8, U455, ROD, and variants thereof.

[0065] In a preferred embodiment, the virus may be an attenuated HIV. In a more preferred embodiment, the virus may be a nef-deficient attenuated HIV.

[0066] In one embodiment, the virus used is a nef-deficient attenuated SHIV. Because the nef gene is responsible for the immune evasion of the AIDS virus, deleting this gene attenuates the virus through the body's immune system. Addition of Ag85B induces strong cellular immunity, leading to a strong immune response and further attenuating the virus.

[0067] The nucleic acid sequence encoding the antigen protein or a portion thereof used is at least 90% identical, at least 95% identical, at least 96% identical, at least 97% identical, at least 98% identical, or at least 99% identical to the nucleic acid sequence comprising nucleotides 1 to 9633 and 10612 to 11022 of SEQ ID NO:3, or the nucleic acid sequence comprising nucleotides 1 to 8786 and 9765 to 15182 of SEQ ID NO:4.

[0068] In a preferred embodiment, the nucleic acid construct used is a nucleic acid sequence that is at least 90% identical, at least 95% identical, at least 96% identical, at least 97% identical, at least 98% identical, or at least 99% identical to the nucleic acid sequence set forth in SEQ ID NO: 3 or 4, and preferably the nucleic acid construct used comprises the nucleic acid sequence set forth in SEQ ID NO: 3 or 4. More advantageously, the nucleic acid sequence set forth in SEQ ID NO: 3 or 4 has one or several modifications (particularly, the modifications may be conservative substitutions, and the modifications may be located at positions that do not affect activity), and particularly advantageously, the nucleic acid sequence set forth in SEQ ID NO: 3 or 4. In particular, the construct having the sequence of SEQ ID NO: 3 or 4 of the present disclosure has been demonstrated to be safer and less toxic, and to be the first to provide a therapeutic composition that is practical for use against HIV, and is of great clinical significance.

[0069] In the amino acid sequence encoded by the construct of the present disclosure, "one or several" typically refers to six or fewer amino acids, preferably five or fewer amino acids, and more preferably four or fewer amino acids (e.g., three or fewer, two or fewer, or one amino acid). To maintain the biological activity of the chimeric protein of the present disclosure, it is desirable for the mutated amino acid residue to be mutated to another amino acid that preserves the properties of the amino acid side chain. When modified, the properties of the amino acid side chain are preferably conservative substitutions. In one embodiment of the present disclosure, "several" may be, for example, 10, 8, 6, 5, 4, 3, or 2, or any of these values ​​or less. Chimeric proteins with deletions or other modifications can be prepared, for example, by site-directed mutagenesis, random mutagenesis, or biopanning using an antibody phage library. Site-directed mutagenesis can be performed using, for example, the KOD-Plus-Mutagenesis Kit (TOYOBO CO., LTD.).

[0070] Dosage and Administration In various embodiments, the compositions of the present disclosure may be used in a variety of dosage and administration regimes.

[0071] In one embodiment, for compositions of the present disclosure, the nucleic acid construct is administered once, or alternatively, two or more times, for example, three, four, five or more times.

[0072] Although a single administration is preferred for convenience, repeated administration, high-dose administration, or a combination thereof may be used to achieve a reliable therapeutic effect. Those skilled in the art can appropriately determine the number, frequency, and amount of administration.

[0073] For example, in the present disclosure, the nucleic acid construct or composition is administered once a week, once every two weeks, once every three weeks, once a month, or once every two months, once every three months, once every four months, once every six months, or once a year.

[0074] In one embodiment, the composition or nucleic acid construct of the present disclosure is present in a concentration of 1.0 x 10 ３ T.C.I.D. ５０ Alternatively, in another embodiment, the composition or nucleic acid construct of the present disclosure is administered at a dose of 1.0 x 10 １ T.C.I.D. ５０ That's it, 2.0 x 10 １ T.C.I.D. ５０ That's it, 3.0 x 10 １ T.C.I.D. ５０ That's it, 4.0 x 10 １ T.C.I.D. ５０ That's it, 5.0 x 10 １ T.C.I.D. ５０ That's it, 6.0 x 10 １ T.C.I.D. ５０ That's it, 7.0 x 10 １ T.C.I.D. ５０ That's it, 8.0 x 10 １ T.C.I.D. ５０ That's it, 9.0 x 10 １ T.C.I.D. ５０ That's it, 1.0 x 10 ２ T.C.I.D. ５０ That's it, 2.0 x 10 ２ T.C.I.D. ５０ That's it, 3.0 x 10 ２ T.C.I.D. ５０ That's it, 4.0 x 10 ２ T.C.I.D. ５０ That's it, 5.0 x 10 ２ T.C.I.D. ５０That's it, 6.0 x 10 ２ T.C.I.D. ５０ That's it, 7.0 x 10 ２ T.C.I.D. ５０ That's it, 8.0 x 10 ２ T.C.I.D. ５０ That's it, 9.0 x 10 ２ T.C.I.D. ５０ That's it, 1.0 x 10 ３ T.C.I.D. ５０ That's it, 2.0 x 10 ３ T.C.I.D. ５０ That's it, 3.0 x 10 ３ T.C.I.D. ５０ That's it, 4.0 x 10 ３ T.C.I.D. ５０ That's it, 5.0 x 10 ３ T.C.I.D. ５０ That's it, 6.0 x 10 ３ T.C.I.D. ５０ That's it, 7.0 x 10 ３ T.C.I.D. ５０ That's it, 8.0 x 10 ３ T.C.I.D. ５０ That's it, 9.0 x 10 ３ T.C.I.D. ５０ That's it, 1.0 x 10 ４ T.C.I.D. ５０ That's it, 2.0 x 10 ４ T.C.I.D. ５０ That's it, 3.0 x 10 4 T.C.I.D. ５０ That's it, 4.0 x 10 4 T.C.I.D. ５０ That's it, 5.0 x 10 4 T.C.I.D. ５０ That's it, 6.0 x 10 4 T.C.I.D. ５０ That's it, 7.0 x 10 4 T.C.I.D. ５０ That's it, 8.0 x 10 4 T.C.I.D. ５０ That's it, 9.0 x 10 4 T.C.I.D. ５０ or more, or 1.0 x 10 ５ T.C.I.D. ５０ That's all.

[0075] In one embodiment, the dose of the nucleic acid construct or composition is 1.0 x 10 ３ T.C.I.D. ５０Above 5.0 x 10 ４ Less than TCID ５０ It is administered in doses less than 100 mg / kg.

[0076] In some embodiments, the low dose, the high dose, or both may be administered two or more times weekly, e.g., two, three, four, five, or more times. In certain embodiments, a low dose may be administered followed by a high dose. In certain embodiments, the high dose may be administered multiple times, e.g., four times.

[0077] In one embodiment, the nucleic acid construct or composition is 1.0 x 10 ３ T.C.I.D. ５０ Above 5.0 x 10 ４ TCID ５０ The nucleic acid construct is administered once in a dose of less than 100 mg / kg, and if a virus is detected in the body, a further administration of the nucleic acid construct is performed. The virus detection can be carried out by an antigen test, a PCR test, or the like.

[0078] In one embodiment, the nucleic acid construct or composition is administered at a low dose of 5.0 x 10 ４ T.C.I.D. ５０ Above 5.0 x 10 ６ T.C.I.D. ５０ It is administered in the following doses:

[0079] In one embodiment, the nucleic acid construct or composition is administered at a high dose of 5.0 x 10 ４ T.C.I.D. ５０ The above doses are administered once.

[0080] In one embodiment, the nucleic acid construct or composition can be administered to a subject who has already received an anti-HIV drug, such as a reverse transcriptase inhibitor, a protease inhibitor, an integrase inhibitor, or a CCR5 inhibitor, such as tenofovir, emtricidadine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof.

[0081] In one embodiment, the nucleic acid construct or composition may be administered to a subject who has not yet received an anti-HIV drug, or to a subject whose anti-HIV drug has been discontinued when administration of the composition is initiated.

[0082] In one embodiment, the other anti-HIV drug used is selected from the group consisting of a reverse transcriptase inhibitor, a protease inhibitor, an integrase inhibitor, and a CCR5 inhibitor.

[0083] In one embodiment, the anti-HIV drug that can be used in conjunction with the nucleic acid construct or composition of the present disclosure can be an anti-HIV drug selected from the group consisting of tenofovir, emtricidavine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof.

[0084] In one embodiment, the nucleic acid construct or composition completely eliminates the AIDS virus in a subject, something that has not been previously available. Even in individuals who are unable to completely eliminate the virus, continued administration of the nucleic acid construct or composition of the present disclosure can continuously suppress onset and maintain health.

[0085] In one embodiment, the subject has not been administered a nucleic acid construct or composition prior to viral infection, and it has not previously been known to completely eliminate the target infectious virus, such as the AIDS virus, in such a subject.

[0086] (Method of Producing Constructs) The constructs of the present disclosure can be produced as follows, representative examples of which are described in the Examples.

[0087] Virus is isolated from a patient and constructed from molecules other than nef (e.g., gag, pol, vif, vpx, vpr, vpu, tat, env, rev, or a combination thereof). The Ag85B gene is then inserted into the nef region. Ag85B can be isolated from the BCG vaccine and inserted into the viral nucleic acid sequence using an integrating plasmid.

[0088] The construct may be prepared by incorporating Ag85B into a known virus strain. For example, in the case of type I HIV (HIV-1) virus, the HIV-1 NL432 strain is typically used, but other examples include, but are not limited to, the BaL strain, IIIB strain, RF strain, GB8 strain, U455 strain, ROD strain, and mutant strains thereof.

[0089] (Use of Construct) The construct of the present disclosure can be used to treat HIV. In one aspect, the present disclosure provides a composition for treating a viral infection in a subject, comprising a nucleic acid construct operably comprising a nucleic acid sequence encoding an antigenic protein or a portion thereof contained in a virus belonging to the lentivirus genus and a nucleic acid sequence encoding an Ag85B protein. The nucleic acid construct of the present disclosure is capable of treating a viral infection without relying on a Th2-type immune response.

[0090] In some embodiments, the nucleic acid sequence encoding the antigenic protein or portion thereof may comprise a nucleic acid sequence that is at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to the nucleic acid sequence comprising nucleotides 1 to 9633 and 10612 to 11022 of SEQ ID NO:3, or the nucleic acid sequence comprising nucleotides 1 to 8786 and 9765 to 15182 of SEQ ID NO:4.

[0091] (Medical Use) In a further aspect, the present disclosure provides a pharmaceutical composition for treating a viral infection in a subject, the pharmaceutical composition comprising a nucleic acid construct operably comprising a nucleic acid sequence encoding an antigenic protein or a portion thereof contained in a virus belonging to the lentivirus genus and a nucleic acid sequence encoding an Ag85B protein. Treatment of viral infection can be achieved by inducing a Th1-type immune response without relying on a Th2-type immune response. Treatment after HIV infection has been achieved for the first time by the present invention.

[0092] The compositions of the present disclosure can completely eliminate viruses in subjects. Particularly in HIV infection, complete elimination of viruses from subjects has not been achieved until now. Complete elimination of viruses is achieved by a Th1-type immune response.

[0093] In a preferred embodiment, the subject has previously been vaccinated with the BCG vaccine. Since BCG vaccination is expected to enhance the adjuvant effect of Ag85B, it is expected that the therapeutic effect of HIV-Ag85B against HIV will be high in the BCG-vaccinated group. The BCG vaccine is not administered in some countries (such as the United States) due to contraindications, and is not necessarily required. A sufficient effect is also expected in the BCG-unvaccinated group.

[0094] (Medicine, Therapy) In one aspect, the present disclosure provides a therapy by administering a composition or medicament of the present disclosure. In one embodiment, the composition or nucleic acid construct of the present disclosure is administered by injection. In another embodiment, the composition or nucleic acid construct of the present disclosure is administered intracorporeally. In certain embodiments, the composition or nucleic acid construct of the present disclosure may be provided together with a preservation solution. In some embodiments, the preservation solution may be a buffer solution. In other embodiments, the composition or nucleic acid construct of the present disclosure may be provided in a container. In certain embodiments, the container that stores the composition or nucleic acid construct of the present disclosure may be a syringe.

[0095] Administration of the agents associated with the attenuated viruses of the present disclosure can be by any suitable means that results in a concentration of the therapeutic agent, in combination with other ingredients, effective to ameliorate or mitigate HIV. The agent may be contained in any suitable amount in any suitable carrier substance, and will generally be present in an amount of 1-95% by weight of the total weight of the composition. The composition may be provided in a dosage form suitable for parenteral (e.g., subcutaneous, intravenous, intramuscular, or intraperitoneal) administration routes. Pharmaceutical compositions can be formulated in accordance with conventional pharmaceutical practice (see, e.g., Remington: The Science and Practice of Pharmacy (20th ed.), ed. A. R. Gennaro, Lippincott Williams & Wilkins, 2000 and Encyclopedia of Pharmaceutical Technology, eds. J. Swarbrick and J. C. Boylan, 1988-1999, Marcel Dekker, New York). The pharmaceutical compositions of the present disclosure can be formulated to release the active compound approximately immediately after administration, or at a predetermined time or period after administration.

[0096] When used for parenteral administration, the present disclosure may be formulated in a unit-dose ampule, multi-dose container, or tube, and may contain additives such as stabilizers, buffers, preservatives, and isotonicity agents. For parenteral administration, the formulation may be formulated as a powder that can be reconstituted with an appropriate carrier (e.g., sterile water) at the time of use. Parenteral administration includes intravenous administration, intramuscular administration, subcutaneous administration, etc., with intravenous administration being preferred. The constructs and active ingredients described herein may be administered with a carrier. A carrier includes a diluent, adjuvant, excipient, or vehicle. The composition may also contain minor amounts of wetting or emulsifying agents, or pH buffering agents, as needed. These compositions may take the form of solutions, suspensions, emulsions, tablets, pills, capsules, powders, sustained-release formulations, combinations thereof, and the like.

[0097] Pharmaceutically acceptable carriers that can be used in the present disclosure refer to vehicles containing the constructs or agents described herein that can be injected into a subject without side effects. Pharmaceutically acceptable carriers include sterile liquids, such as water and oils, including oils of petroleum, animal, plant, or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil, and combinations thereof. Suitable pharmaceutically acceptable carriers include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol, and combinations thereof. Other examples of suitable pharmaceutical carriers are described in "Remington's Pharmaceutical Sciences" by E. W. Martin. Administration of the compositions of the present disclosure may be by any route typically used for inoculation of a composition of the present disclosure, including topical, subcutaneous, intravenous, intramuscular, intradermal, intraperitoneal, oral, inhalation, or a combination thereof.

[0098] The compositions described herein can be formulated into nucleic acid medicines.The nucleic acid medicines described herein include the vectors described herein, which contain DNA encoding the full-length genomic RNA molecule of an infectious, non-pathogenic and / or attenuated virus, operably linked to a promoter suitable for expression in eukaryotic cells, and are appropriately formulated.

[0099] In one specific embodiment, the construct, medicament, etc. of the present disclosure are administered once or twice or more times during the treatment period. As described in the Examples, it has been confirmed that the medicament, etc. of the present disclosure exerts its effects by administering it at least once, and patient compliance is also considered to be good.

[0100] (General Techniques) The molecular biological techniques, biochemical techniques, and microbiological techniques used herein are well known and commonly used in the art, and are described, for example, in Current Protocols in Molecular Biology (http: / / onlinelibrary.wiley.com / book / 10.1002 / 0471142727) and Molecular Cloning: A Laboratory Manual (Fourth Edition) (http: / / www.molecularcloning.com), the relevant portions of which (possibly in their entirety) are incorporated herein by reference.

[0101] The present disclosure has been described above by showing preferred embodiments for ease of understanding. Hereinafter, the present disclosure will be described based on examples. However, the above description and the following examples are provided for illustrative purposes only and are not intended to limit the present disclosure. Therefore, the scope of the present disclosure is not limited to the embodiments or examples specifically described herein, but is limited only by the scope of the claims.

[0102] Example 1: Construction of the Construct SHIV-NM3rN, which contains the HIV-1 NL432 gene in the SIVmac239 background, was used as the starting material. Recombinant SHIV was constructed according to previously reported methods (15, 16). The SHIV-nef vector (SHIV-NI) was constructed from an infectious molecular clone of SHIV-NM3rN (48). The source of the env gene for SHIV-NI was the X4-tropic virus HIV-1 NL432. In SHIV-NI, the nef gene was replaced with unique restriction enzyme sites, such as ClaI and ApaI. The Ag85B gene was amplified by PCR using Mycobacterium kansasii as a template and the primers 5'-ATATCGATACCATGTTCTCCCGTCCCGGGCT-3' (ClaI) (SEQ ID NO: 5) and 5'-AGGGCCCCCTAGCGGGCGCCCAGGCTGG-3' (ApaI) (SEQ ID NO: 6). The PCR product was then digested with restriction enzymes at the ClaI and ApaI sites. This plasmid was designated pSHIV-Ag85B. SHIV-Ag85B was prepared by transfecting 293T cells with pSHIV-Ag85B using FuGENE 6 Transfection Reagent (Roche Diagnostics, Indianapolis, IN), and the culture supernatant 48 hours after transfection was stored in liquid nitrogen until use.

[0103] Example 2: Construct Performance Verification (Detection of Ag85B Protein) M8166 cells were infected with SHIV-Ag85B at an MOI of 0.1 and cultured for 1 hour. The cells were washed three times with phosphate-buffered saline (PBS) and then cultured for an additional 48 hours in culture medium. After three additional washes with PBS, the cells were lysed in PBS containing 1.5 M urea, 2% NP-40, and 5% 2-mercaptoethanol. Cells were then separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, electroblotted onto a nitrocellulose membrane, and blocked with 5% nonfat dry milk in PBS containing 0.01% Tween 20 (PBST). After three washes with PBST, the membrane was incubated with rabbit anti-Ag85B polyclonal antibody for 2 hours. The membrane was washed three times with NBT / BCIP (Roche Diagnostics, Mannheim, Germany) and then incubated with alkaline phosphatase-conjugated anti-rabbit IgG (New England Biolabs, Beverly, MA).

[0104] In vivo stability of the inserted Ag85B gene. ６ Proviral DNA was extracted from PBMCs. When the virus was reisolated, it was isolated from CD8 cells co-cultured with M8166 cells. ＋ Depleted PBMCs were also monitored. Cellular DNA was extracted using a DNeasy tissue kit (QIAGEN). To confirm the stability of the inserted Ag85B gene in SHIV-Ag85B, a proviral DNA fragment encompassing the inserted Ag85B gene in SHIV-Ag85B was PCR amplified using primers whose sequences are as described above.

[0105] (Example 3) Evaluation of nucleic acid medicine for treating SHIV-Ag85B The outline of the experiment in this example is shown in Figure 1. In this example, the amount of virus in plasma and CD4 ＋ The transition of T cell counts was examined.

[0106] (Materials and Methods) (Virus Strains) In this study, SHIV-Ag85B, SHIV-NI, and SHIV89.6P were used. These virus strains were propagated in PBMCs from cynomolgus monkeys. PBMCs isolated by standard Ficoll density gradient separation were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum, 2 mM L-glutamine, and 100 units / ml of IL-2 (Shionogi Pharmaceuticals). They were stimulated with phytohemagglutinin for 72 hours and then infected with SHIV-Ag85B, SHIV-NI, or SHIV89.6P at a multiplicity of infection (MOI) of 0.1. Half of the culture medium was replaced with fresh medium every three days, and cell-free supernatants were collected 6 to 9 days after infection. The TCID of each SHIV was measured using M8166 cells. ５０ The TCID of the virus stock was measured. ５０ The value was 5 × 10 for SHIV-Ag85B. ４ , 4.7 × 10 in SHIV-NI ４ , 3 × 10 in SHIV89.6P ５ It was.

[0107] (Animals) Adult monkeys (from Indonesia, the Philippines, and Malaysia) that were negative for simian SIV, type D retrovirus, T-cell lymphoma virus, simian foamy virus, Epstein-Barr virus, cytomegalovirus, and B virus were used. Seven monkeys were infected with SHIV89.6P50 at 10 ４ T.C.I.D. ５０ Then, one or two weeks after SHIV89.6P infection (5 weeks after #137), anti-HIV drugs (tenofovir 20 mg / kg, emtricitabine 40 mg / kg, or dolutegravir 2.5 mg / kg) were administered subcutaneously once daily. After discontinuation of anti-HIV drug administration, five monkeys were inoculated intravenously with SHIV-Ag85B (high-dose SHIV-Ag85B inoculation; 8 × 10 ４ T.C.I.D. ５０ , low dose SHIV-Ag85B inoculation; 10 ４ T.C.I.D. ５０ Blood was collected periodically using sodium citrate as an anticoagulant and CD4 ＋Lymphoid tissue samples were obtained by biopsy and used for measurement of proviral DNA load and pathological examination.

[0108] Preparation of DNA samples and amplification of the SHIV gag gene by nested PCR. To measure proviral DNA in monkeys inoculated with SHIV-Ag85B, a fragment of the gag gene segment was amplified using nested PCR. Proviral DNA was extracted from PBMCs of inoculated monkeys. Cellular DNA was extracted using DNeasy tissue kits (QIAGEN). Nested PCR was performed using TaKaRa Ex Taq (Takara Bio Inc., Shiga, Japan). The initial and nested PCR protocols are described in references 49 and 50. The primers used in this study were Outer SIVgag-F (5'-CCATTAGTGCCAACAGGCTCAG-3' (SEQ ID NO: 7)) and Outer SIVgag-R (5'-CCCCAGTTGGATCCATCTCCTG-3' (SEQ ID NO: 8)) in the first round of PCR, and Nested SIVgag-F (5'-ACTGTCTGCGTCATCTGGTG-3' (SEQ ID NO: 9)) and Nested SIVgag-R (5'-GTCCCAATCTGCAGCCTCCTC-3' (SEQ ID NO: 10)) in the second round of PCR. After the second round of amplification, 10 μl of the nested PCR product was loaded onto a 1.0% agarose gel and stained with ethidium bromide to visualize the DNA bands. Following initial amplification with the outer gag primer pair, the minimum level of plasmid SIV DNA detected by this PCR method was 100 copies. Further amplification with nested / internal gag primers routinely detects a single copy of plasmid DNA (49, 50).

[0109] Plasma Viral RNA Levels. SHIV infection levels were monitored by measuring viral RNA levels in plasma using highly sensitive quantitative real-time RT-PCR, as previously described (23, 51, 52). Viral RNA was isolated from plasma using the MagNA PureCompact Nucleic Acid Isolation Kit (Roche Diagnostics). Real-time RT-PCR was performed using the QuantiTec Probe RT-PCR Kit (Qiagen) and a LightCycler 480 thermocycler (Roche Diagnostics, Rotkreuz, Switzerland). The gag gene of SIVmac239 was amplified with probe 5'-FAM-TGTCCACCTGCCATTAAGTCCCGA-TAMRA-3' (where FAM is 6-carboxyfluorescein and TAMRA is 6-carboxytetramethylrhodamine) (SEQ ID NO:11) and primers 5'-TGGAAGAAAGACCTCCAGAAAATG-3' (SEQ ID NO:12) and 5'-CAAGTGCAGTTAGCAAGCGAGGAT-3' (SEQ ID NO:13). The detection limit was calculated to be 1000 viral RNA copies / ml.

[0110] (Proviral DNA content) DNA samples were extracted from PBMCs and lymphoid tissues using a DNeasy tissue kit (QIAGEN) according to the manufacturer's protocol. Ultrasensitive digital PCR was performed using a QX200 Droplet Digital PCR system (Bio-Rad). A 20-μl reaction mixture was prepared containing 2 μl of DNA sample, ddPCR supermix for probes (dUTP-free) (Bio-Rad), 900 nM of each primer, 200 nM of probe, and demineralized water. This mixture was placed in a DG8 cartridge with 70 μl of droplet-generating oil (Bio-Rad), and droplets were formed using a droplet generator (Bio-Rad). The droplets were then transferred to a 96-well microplate. PCR amplification was performed with the following program: initial denaturation and stabilization at 95°C for 10 min, 40 cycles of denaturation at 94°C for 30 s, and annealing / extension at 57°C for 60 s, followed by 10 min at 98°C. Droplets were then sorted and analyzed on a QX200 droplet reader (Bio-Rad) using QuantaSoft v1.6 (Bio-Rad) software. Samples were considered only if ≥20,000 droplets were read. Cell numbers were monitored by high-sensitivity quantitative real-time PCR as previously described (53). DNA samples were extracted from PBMCs and lymphoid tissues using the DNADNeasy tissue kit (QIAGEN) according to the manufacturer's protocol. Cell numbers were confirmed by detecting cellular IL-4 sequences using rhesus IL-4-specific primers 5′-TGTGCTCCGGCAGTTCTACA-3′ (SEQ ID NO: 14) and 5′-CCGTTTCAGGAATCGGATCA-3′ (SEQ ID NO: 15) and probe 5′-FAM-TGCACAGCAGTTCCACAGGCAACAAG-TAMRA-3′ (SEQ ID NO: 16).

[0111] (CD4 ＋T cell counts) 100 μl of whole blood from each cynomolgus monkey was stained with fluorescently labeled monoclonal antibodies: anti-CD3 (clone SP34-2, Alexa700; BD) and anti-CD4 (clone L200, PerCP-Cy5.5; BD). Flow cytometry was performed using a FACSCanto II flow cytometer (BD). Data were analyzed using FACSDiVa software.

[0112] (Results) (Administration of anti-HIV drugs only) In monkeys (#137 and #139) administered only anti-HIV drugs, the plasma viral load remained at a low level, but when the administration of anti-HIV drugs was discontinued, the plasma viral load increased (Figures 2 and 3). In addition, in monkeys (#137 and #139) inoculated with SHIV89.6P and administered anti-HIV drugs, the CD4 ＋ T cell counts were very low (Figures 2 and 3).

[0113] (Two doses of low-dose SHIV-Ag85B) In monkey #138 that received two doses of low-dose SHIV-Ag85B, almost no virus was detected in the plasma even after discontinuation of anti-HIV drug administration, and the plasma viral load reached below the detection limit 14 weeks after the first SHIV-Ag85B administration (Fig. 4). In monkey #138 that received two doses of low-dose SHIV-Ag85B, the CD4 ＋ T cell counts remained at normal levels (Figure 4).

[0114] (Single administration of high-dose SHIV-Ag85B) In monkey #142 that received a single dose of high-dose SHIV-Ag85B, the plasma viral load remained below the detection limit throughout the observation period after discontinuation of anti-HIV drug administration (Fig. 5). In monkey #142 that received a single dose of high-dose SHIV-Ag85B, the CD4 ＋ T cell counts remained at normal levels (Figure 5).

[0115] (Five administrations of high-dose SHIV-Ag85B) In a monkey (#141) administered with a high dose of SHIV-Ag85B, the plasma viral load decreased to a low level after two administrations of high-dose SHIV-Ag85B, but after a while the plasma viral load increased (Fig. 6). When a low-dose SHIV-Ag85B was administered again (on days 91, 147, and 154 after infection), the plasma viral load decreased. Even in such cases, it is thought that multiple administrations of SHIV-Ag85B can maintain the plasma viral load at a low level. CD4 ＋ T cells remained at normal levels (Fig. 6).

[0116] (One dose of low-dose and one dose of high-dose SHIV-Ag85B were administered.) In monkey #140, which received low-dose SHIV-Ag85B on day 66 after infection, the plasma viral load increased from day 77 onward. High-dose SHIV-Ag85B was administered on day 98 after infection, but the plasma viral load remained elevated (Figure 7). CD4 ＋ T cell counts remained at low levels throughout the observation period (Figure 7).

[0117] The experimental method of this example was carried out in accordance with FIG.

[0118] Specifically, the administration of anti-HIV drugs was started: after infection with SHIV89.6P and after the peak of the virus in plasma (excluding #137). Administration method: Once daily; subcutaneous administration Anti-HIV drugs: Tenofovir 20 mg / kg Emtricitabine 40 mg / kg Dolutegravir 2.5 mg / kg Reference (Goswami R, et al. Analytical Treatment Interruption after Short-Term Antiretroviral Therapy in a Postnatally Simian-Human Immunodeficiency Virus-Infected Infant Rhesus Macaque Model. mBio. 2019 Sep 5; 10(5): e01971-19. doi: 10.1128 / mBio. 01971-19. , Nishimura Y, et al. , Prevention and treatment of SHIVAD8 infection by a potent d-peptide HIV entry inhibitor. Proc Natl Acad Sci USA. 2020 Sep 8;117(36):22436-22442. doi: 10.1073 / pnas. 2009700117)

[0119] Specifically, the procedure was essentially the same as that for SHIV described above. The administration schedule for each individual is shown in Figure 8.

[0120] In monkey #138 that received two doses of low-dose SHIV-Ag85B and monkey #142 that received a single dose of high-dose SHIV-Ag85B, plasma viral loads remained below the limit of detection for a long period after inoculation (Fig. 9A and 9B). In one monkey (#141) that received high-dose SHIV-Ag85B, only a short-term effect was observed initially, but administration whenever virus appeared in plasma demonstrated a long-term suppressive effect from the fifth inoculation onward (Fig. 10).

[0121] The explanation for the above results is as follows.

[0122] The monkey (#138) that received two doses of low-dose SHIV-Ag85B showed a low dose (1x10 ４ When SHIV-Ag85B (TCID50) was administered on the 4th and 11th days after discontinuation of treatment, the plasma virus remained below the detection limit, and CD4 ＋ The cells also returned to normal levels and were maintained.

[0123] The explanation for the above results is as follows.

[0124] The monkey (#142) that received a single high-dose SHIV-Ag85B injection had a high-dose SHIV-Ag85B (5x10 ４ TCID50) was administered 7 days after discontinuing the drug, and plasma virus levels remained below the detection limit. ＋ The cells also returned to normal levels and were maintained.

[0125] The monkey (#141) given high-dose SHIV-Ag85B had a high-dose SHIV-Ag85B (5x10 ４ When 100 mg of 1000 TCID50 was administered 7 days after discontinuing treatment, the virus in the plasma remained below the detection limit for only a short period of time. Therefore, when the virus appeared in the plasma, it was administered every time it appeared. A long-term suppressive effect was observed from the fifth vaccination onwards. ＋ The cells also returned to normal levels and were maintained.

[0126] (Example 4) Further clinical trials with SHIV-Ag85B Cynomolgus monkeys are administered high doses of SHIV-Ag85B four times weekly. ＋ The change in the number of T cells was measured. (1) Effect of drug withdrawal To examine the effect of drug withdrawal, the number of inoculations and the inoculation dose of SHIV-Ag85B were examined, and an experiment similar to that in Example 3 was performed. Drug withdrawal was performed by discontinuing the administration for 3 days after 8 weeks of administration.

[0127] The results are shown in FIG. 11 (#137 is FIG. 11A, #139 is FIG. 11B).

[0128] The explanation for the above results is as follows.

[0129] Figure 11A shows the results of an untreated monkey (#137) that received only anti-HIV drugs. Virus levels in plasma appeared 7 days after the drug withdrawal and then increased. CD4 counts were inversely proportional to the levels. ＋ The cells decreased and died 170 days after infection.

[0130] Figure 11B shows the results of an untreated monkey (#139) that received only anti-HIV drugs. Plasma virus appeared 110 days after the drug withdrawal and then increased. CD4 ＋ The cells decreased and died 500 days after infection.

[0131] (2) Optimization of Treatment Protocol An outline of the experiment in this example is shown in Figure 12. In this example, the change in plasma viral load when the composition according to the present disclosure was administered after inoculation with the HIV virus was examined.

[0132] (Protocol) Medication will begin one week after SHIV infection. SHIV-Ag85B (5x10 ４ TCID50) was administered.

[0133] Validation Methods Plasma Viral RNA Levels SHIV infection levels were monitored by measuring viral RNA levels in plasma using highly sensitive quantitative real-time RT-PCR as previously described (23, 51, 52). Viral RNA was isolated from plasma using the MagNA PureCompact Nucleic Acid Isolation Kit (Roche Diagnostics). Real-time RT-PCR was performed using the QuantiTec Probe RT-PCR Kit (Qiagen) and a LightCycler 480 thermocycler (Roche Diagnostics, Rotkreuz, Switzerland). The gag gene of SIVmac239 was amplified with probe 5'-FAM-TGTCCACCTGCCATTAAGTCCCGA-TAMRA-3' (where FAM is 6-carboxyfluorescein and TAMRA is 6-carboxytetramethylrhodamine) (SEQ ID NO:11) and primers 5'-TGGAAGAAAGACCTCCAGAAAATG-3' (SEQ ID NO:12) and 5'-CAAGTGCAGTTAGCAAGCGAGGAT-3' (SEQ ID NO:13). The detection limit was calculated to be 1000 viral RNA copies / ml.

[0134] (CD4 ＋ T cell counts) 100 μl of whole blood from each cynomolgus monkey was stained with fluorescently labeled monoclonal antibodies: anti-CD3 (clone SP34-2, Alexa700; BD) and anti-CD4 (clone L200, PerCP-Cy5.5; BD). Flow cytometry was performed using a FACSCanto II flow cytometer (BD). Data were analyzed using FACSDiVa software.

[0135] (Results) The results are shown in Figure 13. It was shown that the anti-HIV drug used was effective regardless of individual differences.

[0136] Example 5: Treatment optimization In this example, a four-dose SHIV-Ag85B study was conducted, essentially following the protocol described above.

[0137] (SHIV-Ag85B 4 doses) High dose (5.0 x 10 ４ T.C.I.D. ５０ In monkeys (#164, #165, and #167) administered SHIV-Ag85B (Figure 14), plasma viral loads remained below the limit of detection when high-dose SHIV-Ag85B was administered four times at weekly intervals. In the control group (no Ag85B), plasma virus levels appeared two weeks after the drug withdrawal, but in the SHIV-Ag85B-treated group, plasma virus levels remained below the limit of detection.

[0138] (SHIV-NI 4 doses) High dose (5.0 x 10 ４ T.C.I.D. ５０ In monkeys (#162, #166, and #168) administered SHIV-NI (Figure 15), plasma viral load remained below the limit of detection after four high-dose SHIV-NI administrations in #168, but increased in #162 and #166 (Figure 15). Plasma virus appeared in the control group without Ag85B two weeks after the drug withdrawal, but plasma virus remained below the limit of detection in the SHIV-Ag85B-treated group (Figure 15 for the control group without Ag85B, Figure 14 for the SHIV-Ag85B-vaccinated group, and Figure 16 for the untreated group).

[0139] (Example 6) Clinical trial using HIV-Ag85B In this example, HIV-Ag85B (SEQ ID NO: 4) is administered subcutaneously or intravenously 4 to 8 times at 1 to 2 week intervals to HIV-infected humans (BCG vaccinated group and BCG unvaccinated group) undergoing treatment with an anti-HIV therapeutic drug, and the therapeutic effect of HIV-Ag85B is confirmed.

[0140] Based on Examples 1 to 5, plasma viral load and CD4 ＋ Since BCG vaccination is expected to enhance the adjuvant effect of Ag85B, it is expected that the therapeutic effect of HIV-Ag85B against HIV will be high in the BCG-vaccinated group.

[0141] (Example 7) Formulation example When a formulation is made, it can be produced by the following method.

[0142] (Example 1) An appropriate amount of freeze-dried nucleic acid construct can be directly added with an appropriate volume of physiological saline to prepare a solution preparation for injection.

[0143] (Example 2) A solution for injection can be prepared by adding an appropriate volume of an isotonic 5% glucose solution to an appropriate amount of the freeze-dried nucleic acid construct.

[0144] (Example 3) A suitable amount of nucleic acid construct dissolved in water can be added with a suitable volume of electrolyte correction solution such as Otsuka Saline Injection 10% to adjust the NaCl concentration to 0.9% to prepare a solution preparation for injection.

[0145] (Example 4) A suitable amount of a nucleic acid construct dissolved in water can be freeze-dried to give a freeze-dried preparation of the sodium salt of the nucleic acid construct.

[0146] References 1. Cohen, M. S. et al. Prevention of HIV-1 infection with early antiretroviral therapy. N. Engl. J. Med. 365, 493-505 (2011). 2. Gupta, R. K. et al. HIV-1 remission following CCR5Delta32 / Delta32 haematopoietic stem-cell transplantation. Nature 568, 244-248 (2019). 3. Hutter, G. et al. Long-term control of HIV by CCR5 Delta32 / Delta32 stem-cell transplantation. N. Engl. J. Med. 360, 692-698 (2009). 4. Burton, D. R. et al. A blueprint for HIV vaccine discovery. Cell Host Microbe 12, 396-407 (2012). 5. Haynes, B. F. et al. HIV-host interactions: implications for vaccine design. Cell Host Microbe 19, 292-303 (2016). 6. Rerks-Ngarm, S. et al. Vaccination with ALVAC and AIDSVAX to prevent HIV-1 infection in Thailand. N. Engl. J. Med. 361, 2209-2220 (2009). 7. Kestler, H. W. 3rd et al. Importance of the nef gene for maintenance of high virus loads and for development of AIDS. Cell 65, 651-662 (1991). 8. Daniel, M. D., Kirchhoff, F., Czajak, S. C., Sehgal, P. K. & Desrosiers, R. C.Protective effects of a live attenuated SIV vaccine with a deletion in the nef gene. Science 258, 1938-1941 (1992). 9. Johnson, R. P. & Desrosiers, R. C. Protective immunity induced by live attenuated simian immunodeficiency virus. Curr. Opin. Immunol. 10, 436-443 (1998). 10. Koff, W. C. et al. HIV vaccine design: insights from live attenuated SIV vaccines. Nat. Immunol. 7, 19-23 (2006). 11. Picker, L. J., Hansen, S. G. & Lifson, J. D. New paradigms for HIV / AIDS vaccine development. Annu. Rev. Med. 63, 95-111 (2012). 12. Baba, T. W. et al. Pathogenicity of live, attenuated SIV after mucosal infection of neonatal macaques. Science 267, 1820-1825 (1995). 13. Baba, T. W. et al. Live attenuated, multiply deleted simian immunodeficiency virus causes AIDS in infant and adult macaques. Nat. Med. 5, 194-203 (1999). 14. Wyand, M. S., Manson, K. H., Lackner, A. A. & Desrosiers, R. C. Resistance of neonatal monkeys to live attenuated vaccine strains of simian immunodeficiency virus. Nat. Med.3, 32-36 (1997). 15. Shimizu, Y. et al. A genetically engineered live-attenuated simian-human immunodeficiency virus that co-expresses the RANTES gene improves the magnitude of cellular immunity in rhesus macaques. Virology 361, 68-79 (2007). 16. Shimizu, Y. et al. Induction of immune response in macaque monkeys infected with simian-human immunodeficiency virus having the TNF-alpha gene at an early stage of infection. Virology 343, 151-161 (2005). 17. Stahl-Hennig, C. et al. Replication, immunogenicity, and protective properties of live-attenuated simian immunodeficiency viruses expressing interleukin-4 or interferon-gamma. Virology 305, 473-485 (2003). 18. Takamura, S., Matsuo, K., Takebe, Y. & Yasutomi, Y. Ag85B of mycobacteria elicits effective CTL responses through activation of robust Th1 immunity as a novel adjuvant in DNA vaccine. J. Immunol. 175, 2541-2547 (2005). 19. Mori, H. et al.Administration of Ag85B showed therapeutic effects to Th2-type cytokine-mediated acute phase atopic dermatitis by inducing regulatory T cells. Arch. Dermatol. Res. 301, 151-157 (2009). 20. Tsujimura, Y. et al. Effects of mycobacteria major secretion protein, Ag85B, on allergic inflammation in the lung. PLoS ONE 9, e106807 (2014). 21. Tsujimura, Y. & Yasutomi, Y. Allergy vaccines using a mycobacterium-secreted antigen, Ag85B, and an IL-4 antagonist. Methods Mol. Biol. 1403, 723-738 (2016). 22. Watanabe, K. et al. Recombinant Ag85B vaccine by taking advantage of characteristics of human parainfluenza type 2 virus vector showed Mycobacteriaspecific immune responses by intranasal immunization. Vaccine 32, 1727-1735 (2014). 23. Okamura, T. et al. Simian immunodeficiency virus SIVmac239 infection and simian human immunodeficiency virus SHIV89.6P infection result in progression to AIDS in cynomolgus macaques of Asian origin. J. Gen. Virol. 97, 3413-3426 (2016). 24. Alexopoulou, L., Holt, A.C., Medzhitov, R. & Flavell, R. A. Recognition of doublestranded RNA and activation of NF-kappaB by Toll-like receptor 3. Nature 413, 732-738 (2001). 25. Kato, H. et al. Differential roles of MDA5 and RIG-I helicases in the recognition of RNA viruses. Nature 441, 101-105 (2006). 26. Fukazawa, Y. et al. Lymph node T cell responses predict the efficacy of live attenuated SIV vaccines. Nat. Med. 18, 1673-1681 (2012). 27. Reynolds, M. R. et al. Macaques vaccinated with live-attenuated SIV control replication of heterologous virus. J. Exp. Med. 205, 2537-2550 (2008). 28. Villinger, F. et al. Induction of long-term protective effects against heterologous challenge in SIVhu-infected macaques. Virology 278, 194-206 (2000). 29. Giavedoni, L. D., Velasquillo, M. C., Parodi, L. M., Hubbard, G. B. & Hodara, V. L. Expression of IL-18 by SIV does not modify the outcome of the antiviral immune response. Virology 303, 327-337 (2002). 30. Giavedoni, L. D. & Yilma, T.Construction and characterization of replicationcompetent simian immunodeficiency virus vectors that express gamma interferon. J. Virol. 70, 2247-2251 (1996). 31. Berg, R. K. et al. Genomic HIV RNA induces innate immune responses through RIG-I-dependent sensing of secondary-structured RNA. PLoS ONE 7, e29291 (2012). 32. Bosinger, S. E. & Utay, N. S. Type I interferon: understanding its role in HIV pathogenesis and therapy. Curr. HIV / AIDS Rep. 12, 41-53 (2015). 33. Co, J. G., Witwer, K. W., Gama, L., Zink, M. C. & Clements, J. E. Induction of innate immune responses by SIV in vivo and in vitro: differential expression and function of RIG-I and MDA5. J. Infect. Dis. 204, 1104-1114 (2011). 34. Mogensen, T. H., Melchjorsen, J., Larsen, C. S. & Paludan, S. R. Innate immune recognition and activation during HIV infection. Retrovirology 7, 54 (2010). 35. Borducchi, E. N. et al. Ad26 / MVA therapeutic vaccination with TLR7 stimulation in SIV-infected rhesus monkeys. Nature 540, 284-287 (2016).36. Vaccari, M. et al. Adjuvant-dependent innate and adaptive immune signatures of risk of SIVmac251 acquisition. Nat. Med. 22, 762-770 (2016). 37. Goulder, P. J. & Watkins, D. I. Impact of MHC class I diversity on immune control of immunodeficiency virus replication. Nat. Rev. Immunol. 8, 619-630 (2008). 38. Loffredo, J. T. et al. Mamu-B*08-positive macaques control simian immunodeficiency virus replication. J. Virol. 81, 8827-8832 (2007). 39. Muhl, T., Krawczak, M., Ten Haaft, P., Hunsmann, G. & Sauermann, U. MHC class I alleles influence set-point viral load and survival time in simian immunodeficiency virus-infected rhesus monkeys. J. Immunol. 169, 3438-3446 (2002). 40. Yant, L. J. et al. The high-frequency major histocompatibility complex class I allele Mamu-B*17 is associated with control of simian immunodeficiency virus SIVmac239 replication. J. Virol. 80, 5074-5077 (2006). 41. Saito, Y., Naruse, T. K., Akari, H., Matano, T. & Kimura, A.Diversity of MHC class I haplotypes in cynomolgus macaques. Immunogenetics 64, 131-141 (2012). 42. Hansen, S. G. et al. Profound early control of highly pathogenic SIV by an effector memory T-cell vaccine. Nature 473, 523-527 (2011). 43. Hansen, S. G. et al. Effector memory T cell responses are associated with protection of rhesus monkeys from mucosal simian immunodeficiency virus challenge. Nat. Med. 15, 293-299 (2009). 44. Hansen, S. G. et al. Immune clearance of highly pathogenic SIV infection. Nature 502, 100-104 (2013). 45. Martinez-Navio, J. M. et al. Adeno-associated virus delivery of anti-HIV monoclonal antibodies can drive long-term virologic suppression. Immunity 50, 567-575.e565 (2019). 46. Borducchi, E. N. et al. Antibody and TLR7 agonist delay viral rebound in SHIVinfected monkeys. Nature 563, 360-364 (2018). 47. Lim, S. Y. et al. TLR7 agonists induce transient viremia and reduce the viral reservoir in SIV-infected rhesus macaques on antiretroviral therapy. Sci. Transl.Med. 10, eaao4521 (2018). 48. Igarashi, T. et al. Infectivity and immunogenicity of SIVmac / HIV-1 chimeric viruses (SHIVs) with deletions in two or three genes (vpr, nef and vpx). Microbiol. Immunol. 42, 71-74 (1998). 49. Unger, R. E. et al. Detection of simian immunodeficiency virus DNA in macrophages from infected rhesus macaques. J. Med. Primatol. 21, 74-81 (1992). 50. Yoshino, N. et al. Intradermal delivery of recombinant vaccinia virus vector DIs induces gut-mucosal immunity. Scand. J. Immunol. 72, 98-105 (2010). 51. Mori, K. et al. Quintuple deglycosylation mutant of simian immunodeficiency virus SIVmac239 in rhesus macaques: robust primary replication, tightly contained chronic infection, and elicitation of potent immunity against the parental wild-type strain. J. Virol. 75, 4023-4028 (2001). 52. Enose, Y. et al. Protection by intranasal immunization of a nef-deleted, nonpathogenic SHIV against intravaginal challenge with a heterologous pathogenic SHIV.Virology 298, 306-316 (2002). 53. Sugimoto, C. et al. Glycosylation of simian immunodeficiency virus influences immune-tissue targeting during primary infection, leading to immunodeficiency or viral control. J. Virol. 86, 9323-9336 (2012). 54. Amara, R. R. et al. Different patterns of immune responses but similar control of a simian-human immunodeficiency virus 89.6P mucosal challenge by modified vaccinia virus Ankara (MVA) and DNA / MVA vaccines. J. Virol. 76, 7625-7631 (2002). 55. Montefiori, D. C. Measuring HIV neutralization in a luciferase reporter gene assay. Methods Mol. Biol. 485, 395-405 (2009). 56. Pollara, J. et al. Bridging Vaccine-Induced HIV-1 Neutralizing and Effector Antibody Responses in Rabbit and Rhesus Macaque Animal Models. J. Virol. 93, e02119-18 (2019). 57. Yamamoto, T. et al. Virus inhibition activity of effector memory CD8(+) T cells determines simian immunodeficiency virus load in vaccinated monkeys after vaccine breakthrough infection. J. Virol.86, 5877-5884 (2012). 58. Yamamoto, T. et al. STING agonists activate latent infected cells and enhance SIV-specific responses ex vivo in naturally SIV-controlled cynomolgus macaques. Sci. Rep. 9, 5917 (2019). 59. Kuromatsu, I., Matsuo, K., Takamura, S., Kim, G., Takebe, Y., Kawamura, J. and Yasutomi, Y. Induction of effective antitumor immune responses by using DNA of an αAg from mycobacteria. Cancer Gene Ther. 2001;8:483-490. (Note) As described above, the present disclosure has been illustrated using preferred embodiments of the present disclosure, but the present disclosure should not be construed as limited to these embodiments. It is understood that the scope of the present disclosure should be interpreted solely by the scope of the claims. It is understood that those skilled in the art will be able to implement equivalent embodiments based on the description of the present disclosure and common general technical knowledge from the description of specific preferred embodiments of the present disclosure. It is understood that the contents of the patents, patent applications, and literature cited in this specification are to be incorporated by reference into this specification as if the contents themselves were specifically set forth in this specification. This application claims priority to Japanese Patent Application No. 2022-140218, filed with the Japan Patent Office on September 2, 2022, the entire contents of which are incorporated herein by reference as necessary.

[0147] The present disclosure is useful in the pharmaceutical industry.

[0148] SEQ ID NO: 1: Nucleic acid sequence of Ag85B SEQ ID NO: 2: Amino acid sequence of Ag85B SEQ ID NO: 3: Nucleic acid sequence of SHIV-Ag85B SEQ ID NO: 4: Nucleic acid sequence of HIV-Ag85B SEQ ID NO: 5: ClaI primer SEQ ID NO: 6: ApaI primer SEQ ID NO: 7: Outer SIVgag-F primer SEQ ID NO: 8: Outer SIVgag-R primer SEQ ID NO: 9: Nested SIVgag-F primer SEQ ID NO: 10: Nested SIVgag-R primer SEQ ID NO: 11: Probe for gag of SIVmac239 SEQ ID NO: 12: Primer 1 for gag of SIVmac239 SEQ ID NO: 13: Primer 2 for gag of SIVmac239 SEQ ID NO: 14: Primer 1 for IL-4 SEQ ID NO: 15: Primer 2 for IL-4 SEQ ID NO: 16: Probe for IL-4

Claims

1. A composition for treating viral infection in a subject, comprising a nucleic acid construct activatably containing a nucleic acid sequence encoding an antigen protein or a portion thereof contained in a virus belonging to the genus Lentivirus, and a nucleic acid sequence encoding the Ag85B protein.

2. The composition according to claim 1, characterized in that the composition is administered after viral infection.

3. The composition according to claim 2, wherein the treatment for the viral infection substantially eliminates the virus from the subject after the viral infection.

4. The composition according to claim 2, wherein the treatment for the viral infection is administered to the subject after the viral infection for the purpose of substantially eliminating the virus.

5. The composition according to any one of claims 1 to 4, wherein the nucleic acid sequence encoding the antigen protein or a part thereof is a nucleic acid sequence encoding an attenuated virus.

6. The composition according to claim 5, wherein the attenuated virus is a NEF-deficient attenuated virus.

7. The composition according to claim 5, wherein the nucleic acid sequence encoding the Ag85B protein is incorporated into the nucleic acid sequence encoding the attenuated virus.

8. The composition according to claim 6, wherein the nucleic acid sequence encoding the Ag85B protein is incorporated into the position of the deleted nef gene in the nucleic acid sequence encoding the nef-deficient attenuated virus.

9. The composition according to any one of claims 1 to 4, wherein the virus is the AIDS virus.

10. The composition according to any one of claims 1 to 4, wherein the virus is a weakened virus of a virus that infects humans.

11. The composition according to any one of claims 1 to 4, wherein the virus is an attenuated AIDS virus selected from the group consisting of HIV, SIV, SHIV, and FIV.

12. The composition according to any one of claims 1 to 4, wherein the virus is a weakened HIV.

13. The composition according to any one of claims 1 to 4, wherein the virus is a nef-deficient attenuated HIV.

14. The composition according to any one of claims 1 to 4, wherein the nucleic acid sequence encoding the antigen protein or a portion thereof includes a nucleic acid sequence that is at least 90% identical to a nucleic acid sequence including nucleotides at positions 1 to 9633 and 10612 to 11022 of SEQ ID NO: 3, or a nucleic acid sequence including nucleotides at positions 1 to 8786 and 9765 to 15182 of SEQ ID NO:

4.

15. The composition according to any one of claims 1 to 4, wherein the nucleic acid construct comprises a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence described in SEQ ID NO: 3 or 4.

16. The composition according to any one of claims 1 to 4, wherein the nucleic acid construct comprises the nucleic acid sequence described in SEQ ID NO: 3 or 4.

17. The composition according to any one of claims 1 to 4, characterized in that the nucleic acid construct is administered once.

18. The composition according to any one of claims 1 to 4, characterized in that the nucleic acid construct is administered two or more times.

19. The composition according to claim 18, characterized in that the nucleic acid construct is administered once a week, once every two weeks, once every three weeks, once a month, or once every two months.

20. The nucleic acid construct is 1.0 × 10 ３ TCID ５０ The composition according to any one of claims 1 to 4, characterized in that it is administered in the above dose.

21. The nucleic acid construct is 1.0 × 10 ３ TCID ５０ The above 5.0 x 10 ４ TCID ５０ The composition according to any one of claims 1 to 4, characterized in that it is administered in a dose less than a certain amount.

22. The nucleic acid construct is 1.0 × 10 ３ TCID ５０ to 5.0 × 10 ４ of TCID ５０ The composition according to claim 21, which is administered at a dose less than [the above value] once a week.

23. The nucleic acid construct is 1.0 × 10 ３ TCID ５０ The above 5.0 x 10 ４ TCID ５０ The composition according to claim 21, characterized in that it is administered once in a dose of less than a certain amount, and if a virus is detected in the body, the nucleic acid construct is further administered.

24. The nucleic acid construct is 5.0 × 10 ４ TCID ５０ The above 5.0 x 10 ６ TCID ５０ The composition according to any one of claims 1 to 4, characterized in that it is administered in the following doses.

25. The nucleic acid construct is 5.0 × 10 ４ TCID ５０ The composition according to claim 24, characterized in that it is administered once in the above dose.

26. The composition according to any one of claims 1 to 4, characterized in that the subject has already been administered an anti-HIV drug.

27. The composition according to any one of claims 1 to 4, wherein the subject has not yet been administered an anti-HIV drug, or the anti-HIV drug is discontinued at the start of administration of the composition.

28. The anti-HIV drug is characterized by being administered as an anti-HIV drug selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors. The composition according to any one of claims 1 to 4, characterized in that the subject has already been administered an anti-HIV drug, or The composition according to any one of claims 1 to 4, wherein the subject has not yet been administered an anti-HIV drug, or the anti-HIV drug is discontinued at the start of administration of the composition.

29. The anti-HIV drug administered is characterized by being selected from the group consisting of tenofovir, emtricidabine, dolutegravir, zidovudine, lamivudine, sanilvudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination thereof. The composition according to any one of claims 1 to 4, characterized in that the subject has already been administered an anti-HIV drug, or The composition according to any one of claims 1 to 4, wherein the subject has not yet been administered an anti-HIV drug, or the anti-HIV drug is discontinued at the start of administration of the composition.

30. The composition according to any one of claims 1 to 4, wherein the composition completely eliminates the AIDS virus in the subject.

31. The composition according to any one of claims 1 to 4, wherein the composition is not administered before viral infection.

32. The composition according to any one of claims 1 to 4, wherein the subject has previously received BCG vaccination.