Compositions for treating HIV infections

Abstract

The present application provides a composition for the treatment of viral infections. In one embodiment, the present application provides a composition for treating a viral infection in a subject, the composition operably containing a nucleic acid construct encoding a nucleic acid sequence encoding an antigen protein contained in a virus belonging to the genus lentiviral, or a portion thereof, and a nucleic acid sequence encoding an Ag85B protein. In one embodiment, the composition of the present application is characterized by being administered after infection with a virus.

Description

Technical Field

[0001] The present application provides compositions and related techniques for treating viral infections. More specifically, a technique regarding a nucleic acid construct is provided, which operably contains 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. Background Art

[0002] In the past 30 years, despite significant resources being devoted to the development of an effective human immunodeficiency virus (HIV) vaccine, these efforts have not been successful. Although antiretroviral therapy has led to a dramatic reduction in HIV-related morbidity and mortality, it cannot cure HIV (Non-Patent Document 1). There have been two cases of remission of HIV achieved through cell transplantation. If an effective vaccine could be developed, it would prevent the spread of HIV infection, but in the past 30 years, almost all vaccine development efforts have failed. The only successful HIV vaccine to date was evaluated using the RV144 clinical trial, with an overall efficacy of only 31% (Non-Patent Document 2). Prior Art Documents Non-Patent Documents

[0003] Non-Patent Document 1: Cohen, M.S. et al. Prevention of HIV-1 infection with early antiretroviral therapy. N. Engl. J. Med. 365, 493 - 505 (2011). Non-Patent Document 2: 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). Summary of the Invention Means for Solving the Problem

[0004] The present inventors have found that a nucleic acid construct operably containing 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 can treat viral infections in a subject, thereby completing the present invention.

[0005] Accordingly, the present invention provides, for example, the following items. (Item 1) A composition for treating a viral infection in a subject, operably comprising a nucleic acid construct encoding a nucleic acid sequence of 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 2) The composition according to the above item, 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 of the 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 of the viral infection is administered for the purpose of substantially eliminating 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 part 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-deleted attenuated virus. (Item 7) The composition according to any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is assembled into the nucleic acid sequence encoding the attenuated virus. (Item 8) The composition according to any one of the above items, wherein the nucleic acid sequence encoding the Ag85B protein is assembled at the position of the deleted nef gene in the nucleic acid sequence encoding the nef-deleted attenuated virus. (Item 9) The composition according to any one of the above items, wherein the virus is a human immunodeficiency virus. (Item 10) The composition according to any one of the above items, wherein the virus is an attenuated virus of a virus infecting humans. (Item 11) The composition according to any one of the above items, wherein the virus is an attenuated virus of a human immunodeficiency virus selected from the group consisting of HIV, SIV, SHIV, and FIV. (Item 12) The composition according to any one of the above items, wherein the virus is an attenuated HIV. (Item 13) The composition according to any one of the above items, wherein the virus is a nef-deleted attenuated HIV. (Item 14) For the composition according to any one of the above items, the nucleic acid sequence encoding the antigenic protein or a part thereof contains a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence comprising nucleotides 1 to 9633 and nucleotides 10612 to 11022 of SEQ ID NO: 3 or the nucleic acid sequence comprising nucleotides 1 to 8786 and nucleotides 9765 to 15182 of SEQ ID NO: 4. (Item 15) For the composition according to any one of the above items, the nucleic acid construct contains a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence described in SEQ ID NO: 3 or 4. (Item 16) For the composition according to any one of the above items, the nucleic acid construct contains the nucleic acid sequence described in SEQ ID NO: 3 or 4. (Item 17) For the composition according to any one of the above items, it is characterized in that the nucleic acid construct is administered once. (Item 18) For the composition according to any one of the above items, it is characterized in that the nucleic acid construct is administered two or more times. (Item 18-A) For the composition according to any one of the above items, it is characterized in that the nucleic acid construct is administered after administering an anti-HIV drug. (Item 18-B) The composition according to 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, preferably from tenofovir, emtricitabine, 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 of these substances. (Item 18-C) The composition according to any one of the above items, characterized in that after administering the anti-HIV drug, the nucleic acid construct is administered after interrupting the administration of the HIV drug. (Item 18-D) The composition according to any one of the above items, characterized in that after administering the anti-HIV drug, the nucleic acid construct is administered more than 2 times. (Item 18-E) The composition according to any one of the above items, characterized in that after administering the anti-HIV drug, the nucleic acid construct is administered 3 times, 4 times, 5 times or more, or more than these times. (Item 18-F) The composition according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is carried out at least 1 week apart. (Item 18-G) The composition according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is carried out at intervals of 1 to 4 weeks. (Item 18-H) The composition according to any one of the above items, characterized in that the administrations of the nucleic acid construct are carried out at intervals of 1 to 3 weeks for more than 2 times. (Item 18-I) The composition according to any one of the above items, characterized in that during the administrations of the nucleic acid construct for more than 2 times, the plasma viral load of the patient is observed, and if it reaches a specified value, the administration is carried out again. (Item 18-J) The composition according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 5.0×10 5 TCID 50 or more or 1.0×10 6 TCID 50 or more. (Item 18-K) The composition according to any one of the above items, characterized in that the administrations of the nucleic acid construct for more than 2 times are carried out at intervals of 1 week to 4 weeks at a dose of 5.0×10 5 TCID 50 or more or 1.0×10 6 TCID 50 or more. (Item 19) The composition according to any one of the above items, characterized in that the nucleic acid construct is administered once a week, once every 2 weeks, once every 3 weeks, once a month or once every 2 months. (Item 19A) The composition according to any one of the above items, characterized in that the nucleic acid construct is administered at intervals of once a week, once every 2 weeks, once every 3 weeks, once a month or once every 2 months or at intervals longer than these. (Item 20) The composition according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID 50 or more. (Item 21) The composition according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 or less. (Item 22) The composition according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID50 above and less than 5.0×10 4 TCID 50 of the dosage, the nucleic acid construct is administered once a week. (Item 23) The composition according to any one of the above items, characterized in that, at 1.0×10 3 TCID 50 above and less than 5.0×10 4 TCID 50 of the dosage, the nucleic acid construct is administered once, and when the virus is detected in vivo, the nucleic acid construct is further administered. (Item 24) The composition according to any one of the above items, characterized in that, at 5.0×10 4 TCID 50 above, 5.0×10 6 TCID 50 or less of the dosage, the nucleic acid construct is administered. (Item 25) The composition according to any one of the above items, characterized in that, at 5.0×10 4 TCID 50 above of the dosage, the nucleic acid construct is administered once. (Item 26) The composition according to any one of the above items, characterized in that the subject has been administered an anti-HIV drug. (Item 27) The composition according to any one of the above items, characterized in that the subject has not been administered an anti-HIV drug or the anti-HIV drug has been discontinued at the start of administration of the composition. (Item 28) The composition according to any one of the above items, characterized in that, regarding the anti-HIV drug, an anti-HIV drug selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors is administered. (Item 29) The composition according to any one of the above items, characterized in that, regarding the anti-HIV drug, an anti-HIV drug selected from the group consisting of tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances is administered. (Item 30) The composition according to any one of the above items, characterized in that the composition completely clears the HIV in the subject. (Item 31) The composition according to any one of the above items, the composition is not administered before viral infection. (Item 32) The composition according to any one of the above items, the subject has been previously vaccinated with the BCG vaccine. (Item 1A) A method, which is a method for treating a viral infection of a subject, the method comprising the step of administering an effective amount of a nucleic acid construct, the nucleic acid construct operably containing 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. (Item 2A) The method according to the above item, characterized in that the nucleic acid construct is administered after viral infection. (Item 3A) The method according to any one of the above items, characterized in that the treatment of the viral infection is to substantially eliminate the virus from the subject after viral infection. (Item 4A) The method according to any one of the above items, the treatment of the viral infection is administered for the purpose of substantially eliminating the virus from the subject after viral infection. (Item 5A) The method according to any one of the above items, the nucleic acid sequence encoding the antigen protein or a part thereof is a nucleic acid sequence encoding an attenuated virus. (Item 6A) The method according to any one of the above items, the attenuated virus is a nef-deleted attenuated virus. (Item 7A) The method according to any one of the above items, the nucleic acid sequence encoding the Ag85B protein is assembled into the nucleic acid sequence encoding the attenuated virus. (Item 8A) The method according to any one of the above items, the nucleic acid sequence encoding the Ag85B protein is assembled at the position of the deleted nef gene in the nucleic acid sequence encoding the nef-deleted attenuated virus. (Item 9A) The method according to any one of the above items, the virus is the AIDS virus. (Item 10A) The method according to any one of the above items, wherein the virus is an attenuated virus of a virus that infects humans. (Item 11A) The method according to any one of the above items, wherein the virus is an attenuated virus of an AIDS virus selected from the group consisting of HIV, SIV, SHIV, and FIV. (Item 12A) The method according to any one of the above items, wherein the virus is attenuated HIV. (Item 13A) The method according to any one of the above items, wherein the virus is nef-deleted attenuated HIV. (Item 14A) The method according to any one of the above items, wherein the nucleic acid sequence encoding the antigen protein or a part thereof contains a nucleic acid sequence that is at least 90% 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. (Item 15A) The method according to any one of the above items, wherein the nucleic acid construct contains a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence described in SEQ ID NO: 3 or 4. (Item 16A) The method according to any one of the above items, wherein the nucleic acid construct contains the nucleic acid sequence described in SEQ ID NO: 3 or 4. (Item 17A) The method according to any one of the above items, characterized in that the nucleic acid construct is administered once. (Item 18A) The method according to any one of the above items, characterized in that the nucleic acid construct is administered two or more times. (Item 18A-A) The method according to any one of the above items, characterized in that the nucleic acid construct is administered after administering an anti-HIV drug. (Item 18A-B) The method according to 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, preferably selected from tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir mixture, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances. (Item 18A-C) The method according to any one of the above items, wherein after administering the anti-HIV drug, the nucleic acid construct is administered after interrupting the administration of the HIV drug. (Item 18A-D) The method according to any one of the above items, wherein after administering the anti-HIV drug, the nucleic acid construct is administered more than 2 times. (Item 18A-E) The method according to any one of the above items, wherein after administering the anti-HIV drug, the nucleic acid construct is administered 3 times, 4 times, 5 times or more, or more than these times. (Item 18A-F) The method according to any one of the above items, wherein the administration of the nucleic acid construct more than 2 times is carried out at least 1 week apart. (Item 18A-G) The method according to any one of the above items, wherein the administration of the nucleic acid construct more than 2 times is carried out at intervals of 1 to 4 weeks. (Item 18A-H) The method according to any one of the above items, wherein the administration of the nucleic acid construct more than 2 times is carried out at intervals of 1 to 3 weeks. (Item 18A-I) The method according to any one of the above items, wherein during the administration of the nucleic acid construct more than 2 times, the plasma viral load of the patient is observed, and if a specified value is reached, administration is carried out again. (Item 18A-J) The method according to any one of the above items, wherein at each administration, 5.0×10 5 TCID 50 or more, or 1.0×10 6 TCID 50 or more of the nucleic acid construct is administered. (Item 18A-K) According to the method described in any one of the above items, it is characterized in that the nucleic acid construct is administered more than twice at a dose of 5.0×10 5 TCID 50 or more or 1.0×10 6 TCID 50 or more, and the administrations are carried out at intervals of 1 to 4 weeks. (Item 19A) According to the method described in any one of the above items, it is 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. (Item 19A) According to the method described in any one of the above items, it is characterized in that the nucleic acid construct is administered at intervals of once a week, once every two weeks, once every three weeks, once a month or once every two months or at intervals longer than these. (Item 20A) According to the method described in any one of the above items, it is characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID 50 or more. (Item 21A) According to the method described in any one of the above items, it is characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 . (Item 22A) According to the method described in any one of the above items, it is characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 , and the nucleic acid construct is administered once a week. (Item 23A) According to the method described in any one of the above items, it is characterized in that the nucleic acid construct is administered once at a dose of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 , and when the virus is detected in the body, the nucleic acid construct is further administered. (Item 24A) According to the method described in any one of the above items, it is characterized in that at a dose of 5.0×104 TCID 50 The above, 5.0×10 6 TCID 50 The nucleic acid construct is administered at a dosage of 5.0×10 or less. (Item 25A) The method according to any one of the above items, characterized in that the nucleic acid construct is administered once at a dosage of 5.0×10 4 TCID 50 or more. (Item 26A) The method according to any one of the above items, characterized in that the subject has been administered an anti-HIV drug. (Item 27A) The method according to any one of the above items, wherein the subject has not been administered an anti-HIV drug or the anti-HIV drug has been discontinued at the start of administration of the composition. (Item 28A) The method according to any one of the above items, characterized in that, with respect to the anti-HIV drug, an anti-HIV drug selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors is administered. (Item 29A) The method according to any one of the above items, characterized in that, with respect to the anti-HIV drug, an anti-HIV drug selected from the group consisting of tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances is administered. (Item 30A) The method according to any one of the above items, wherein the composition completely clears the AIDS virus in the subject. (Item 31A) The method according to any one of the above items, wherein the composition is not administered before viral infection. (Item 32A) The method according to any one of the above items, wherein the subject has been previously vaccinated with the BCG vaccine. (Item 1B) Use of a nucleic acid construct in the manufacture of a drug for treating viral infections in a subject, the nucleic acid construct operably containing 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. (Item 2B) For use as described in the above item, it is characterized in that the nucleic acid construct is administered after viral infection. (Item 3B) For use as described in any one of the above items, the treatment of viral infection is to substantially eliminate the virus from the subject after viral infection. (Item 4B) For use as described in any one of the above items, the treatment of viral infection is to administer a drug with the aim of substantially eliminating the virus from the subject after viral infection. (Item 5B) For use as described in any one of the above items, the nucleic acid sequence encoding the antigen protein or a part thereof is a nucleic acid sequence encoding an attenuated virus. (Item 6B) For use as described in any one of the above items, the attenuated virus is a nef - deleted attenuated virus. (Item 7B) For use as described in any one of the above items, the nucleic acid sequence encoding the Ag85B protein is assembled into the nucleic acid sequence encoding the attenuated virus. (Item 8B) For use as described in any one of the above items, the nucleic acid sequence encoding the Ag85B protein is assembled at the position of the deleted nef gene in the nucleic acid sequence encoding the nef - deleted attenuated virus. (Item 9B) For use as described in any one of the above items, the virus is a human immunodeficiency virus. (Item 10B) For use as described in any one of the above items, the virus is an attenuated virus of a virus that infects humans. (Item 11B) For use as described in any one of the above items, the virus is an attenuated virus of a human immunodeficiency virus selected from the group consisting of HIV, SIV, SHIV, and FIV. (Item 12B) For use as described in any one of the above items, the virus is an attenuated HIV. (Item 13B) For use as described in any one of the above items, the virus is a nef - deleted attenuated HIV. (Item 14B) For use according to any one of the above items, the nucleic acid sequence encoding the antigenic protein or a part thereof contains a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence comprising nucleotides 1 to 9633 and nucleotides 10612 to 11022 of SEQ ID NO: 3 or the nucleic acid sequence comprising nucleotides 1 to 8786 and nucleotides 9765 to 15182 of SEQ ID NO: 4. (Item 15B) For use according to any one of the above items, the nucleic acid construct contains a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence described in SEQ ID NO: 3 or 4. (Item 16B) For use according to any one of the above items, the nucleic acid construct contains the nucleic acid sequence described in SEQ ID NO: 3 or 4. (Item 17B) For use according to any one of the above items, it is characterized in that the nucleic acid construct is administered once. (Item 18B) For use according to any one of the above items, it is characterized in that the nucleic acid construct is administered two or more times. (Item 18B-A) For use according to any one of the above items, it is characterized in that the nucleic acid construct is administered after administering an anti-HIV drug. (Item 18B-B) For use according to any one of the above items, the anti-HIV drug is selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors, preferably tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances. (Item 18B-C) For use according to any one of the above items, it is characterized in that the nucleic acid construct is administered after administering an anti-HIV drug and after discontinuing the administration of the HIV drug. (Item 18B-D) For use according to any one of the above items, it is characterized in that the nucleic acid construct is administered two or more times after administering an anti-HIV drug. (Item 18B-E) Use according to any one of the above items, characterized in that after administering an anti-HIV drug, the nucleic acid construct is administered 3 times, 4 times, 5 times or more, or more than these times. (Item 18B-F) Use according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is carried out at intervals of at least 1 week or more. (Item 18B-G) Use according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is carried out at intervals of 1 week to 4 weeks. (Item 18B-H) Use according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is carried out at intervals of 1 to 3 weeks. (Item 18B-I) Use according to any one of the above items, characterized in that during the administration of the nucleic acid construct more than 2 times, the plasma viral load of the patient is observed, and if it reaches a specified value, administration is carried out again. (Item 18B-J) Use according to any one of the above items, characterized in that at each time 5.0×10 5 TCID 50 or more or 1.0×10 6 TCID 50 or more of the dose, the nucleic acid construct is administered. (Item 18B-K) Use according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is at each time 5.0×10 5 TCID 50 or more or 1.0×10 6 TCID 50 or more, and is administered at intervals of 1 week to 4 weeks. (Item 19B) Use according to any one of the above items, 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. (Item 19B-A) Use according to any one of the above items, characterized in that the nucleic acid construct is administered at intervals of once a week, once every two weeks, once every three weeks, once a month or once every two months or more than these intervals. (Item 20B) The use according to any one of the above items, characterized in that the nucleic acid construct is administered at a dosage of 1.0×10 3 TCID 50 or more. (Item 21B) The use according to any one of the above items, characterized in that the nucleic acid construct is administered at a dosage of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 . (Item 22B) The use according to any one of the above items, characterized in that the nucleic acid construct is administered once a week at a dosage of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 . (Item 23B) The use according to any one of the above items, characterized in that the nucleic acid construct is administered once at a dosage of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 , and when the virus is detected in vivo, the nucleic acid construct is further administered. (Item 24B) The use according to any one of the above items, characterized in that the nucleic acid construct is administered at a dosage of 5.0×10 4 TCID 50 or more and 5.0×10 6 TCID 50 or less. (Item 25B) The use according to any one of the above items, characterized in that the nucleic acid construct is administered once at a dosage of 5.0×10 4 TCID 50 or more. (Item 26B) The use according to any one of the above items, characterized in that the subject has been administered an anti-HIV drug. (Item 27B) The use according to any one of the above items, wherein the subject has not been administered an anti-HIV drug or the anti-HIV drug has been discontinued at the start of administration of the composition. (Item 28B) The use according to any one of the above items, characterized in that, with respect to the anti-HIV drug, an anti-HIV drug selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors is administered. (Item 29B) The use according to any one of the above items, characterized in that, with respect to the anti-HIV drug, an anti-HIV drug selected from the group consisting of tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir mixture, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances is administered. (Item 30B) The use according to any one of the above items, wherein the composition completely clears the AIDS virus in the subject. (Item 31B) The use according to any one of the above items, wherein the composition is not administered before viral infection. (Item 32B) The use according to any one of the above items, wherein the subject has been previously vaccinated with the BCG vaccine. (Item 1C) A nucleic acid construct for treating viral infection in a subject, operably containing 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. (Item 2C) The nucleic acid construct according to the above item, characterized in that the nucleic acid construct is administered after viral infection. (Item 3C) The nucleic acid construct according to any one of the above items, wherein the treatment of the viral infection is to substantially eliminate the virus from the subject after viral infection. (Item 4C) The nucleic acid construct according to any one of the above items, wherein the treatment of the viral infection is administered for the purpose of substantially eliminating the virus from the subject after viral infection. (Item 5C) The nucleic acid construct according to any one of the above items, wherein the nucleic acid sequence encoding the antigen protein or a part thereof is a nucleic acid sequence encoding an attenuated virus. (Item 6C) The nucleic acid construct according to any one of the above items, wherein the attenuated virus is a nef-deleted attenuated virus. (Item 7C) For the nucleic acid construct according to any one of the above items, the nucleic acid sequence encoding the Ag85B protein is assembled into the nucleic acid sequence encoding the attenuated virus. (Item 8C) For the nucleic acid construct according to any one of the above items, the nucleic acid sequence encoding the Ag85B protein is assembled at the position of the deleted nef gene in the nucleic acid sequence encoding the nef-deleted attenuated virus. (Item 9C) For the nucleic acid construct according to any one of the above items, the virus is a human immunodeficiency virus. (Item 10C) For the nucleic acid construct according to any one of the above items, the virus is an attenuated virus of a virus that infects humans. (Item 11C) For the nucleic acid construct according to any one of the above items, the virus is an attenuated virus of a human immunodeficiency virus selected from the group consisting of HIV, SIV, SHIV, and FIV. (Item 12C) For the nucleic acid construct according to any one of the above items, the virus is attenuated HIV. (Item 13C) For the nucleic acid construct according to any one of the above items, the virus is nef-deleted attenuated HIV. (Item 14C) For the nucleic acid construct according to any one of the above items, the nucleic acid sequence encoding the antigen protein or a part thereof contains a nucleic acid sequence that is at least 90% 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. (Item 15C) For the nucleic acid construct according to any one of the above items, the nucleic acid construct contains a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence described in SEQ ID NO: 3 or 4. (Item 16C) For the nucleic acid construct according to any one of the above items, the nucleic acid construct contains the nucleic acid sequence described in SEQ ID NO: 3 or 4. (Item 17C) For the nucleic acid construct according to any one of the above items, it is characterized in that the nucleic acid construct is administered once. (Item 18C) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered more than 2 times. (Item 18C-A) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered after administering an anti-HIV drug. (Item 18C-B) The anti-HIV drug for the nucleic acid construct according to any one of the above items is selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors, preferably tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances. (Item 18C-C) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered after discontinuing the administration of the anti-HIV drug after administering the anti-HIV drug. (Item 18C-D) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered more than 2 times after administering an anti-HIV drug. (Item 18C-E) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered 3 times, 4 times, 5 times or more, or more than these times after administering an anti-HIV drug. (Item 18C-F) The nucleic acid construct according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is carried out at least 1 week apart. (Item 18C-G) The nucleic acid construct according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is carried out at intervals of 1 to 4 weeks. (Item 18C-H) The nucleic acid construct according to any one of the above items, characterized in that the administration of the nucleic acid construct more than 2 times is carried out at intervals of 1 to 3 weeks. (Item 18C-I) The nucleic acid construct according to any one of the above items, characterized in that, during more than two administrations of the nucleic acid construct, the plasma viral load of the patient is observed, and if it reaches a specified value, administration is carried out again. (Item 18C-J) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 5.0×10 5 TCID 50 or more or 1.0×10 6 TCID 50 or more. (Item 18C-K) The nucleic acid construct according to any one of the above items, characterized in that more than two administrations of the nucleic acid construct are carried out at intervals of 1 to 4 weeks at a dose of 5.0×10 5 TCID 50 or more or 1.0×10 6 TCID 50 or more. (Item 19C) The nucleic acid construct according to any one of the above items, 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. (Item 19C-A) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered at intervals of once a week, once every two weeks, once every three weeks, once a month or once every two months or longer intervals. (Item 20C) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID 50 or more. (Item 21C) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 or more. (Item 22C) The nucleic acid construct according to any one of the above items, characterized in that the nucleic acid construct is administered at a dose of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50The dosage of the nucleic acid construct is administered once a week. (Item 23C) The nucleic acid construct according to any one of the above items, characterized in that it is 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 The nucleic acid construct is administered once, and when the virus is detected in vivo, the nucleic acid construct is further administered. (Item 24C) The nucleic acid construct according to any one of the above items, characterized in that it is 5.0×10 4 TCID 50 or more and 5.0×10 6 TCID 50 or less. The nucleic acid construct is administered. (Item 25C) The nucleic acid construct according to any one of the above items, characterized in that it is 5.0×10 4 TCID 50 or more. The nucleic acid construct is administered once. (Item 26C) The nucleic acid construct according to any one of the above items, characterized in that the subject has been administered an anti-HIV drug. (Item 27C) The nucleic acid construct according to any one of the above items, the subject has not been administered an anti-HIV drug or the anti-HIV drug has been discontinued when starting to administer the composition. (Item 28C) The nucleic acid construct according to any one of the above items, characterized in that for the anti-HIV drug, an anti-HIV drug selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors is administered. (Item 29C) The nucleic acid construct according to any one of the above items, characterized in that for the anti-HIV drug, an anti-HIV drug selected from the group consisting of tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir mixture, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances is administered. (Item 30C) The nucleic acid construct according to any one of the above items, wherein the composition completely clears the human immunodeficiency virus in the subject. (Item 31C) The nucleic acid construct according to any one of the above items, wherein the composition is not administered before viral infection. (Item 32C) The nucleic acid construct according to any one of the above items, wherein the subject has been previously vaccinated with the BCG vaccine. (Item 1D) The composition, nucleic acid molecule, method, use or nucleic acid construct according to any one of the above items, wherein the composition, nucleic acid molecule, method, use or nucleic acid construct treats an infection with a virus belonging to the genus Lentivirus, preferably human immunodeficiency virus, or a viral infection caused by these.

[0006] In the present application, it is intended to provide, in addition to the explicitly described combinations, further combinations of one or more of the above features. More embodiments and advantages of the present application can be recognized by those skilled in the art if the following detailed description is read as needed. Description of the Drawings

[0007] Figure 1 Figure 1 Shows an overview of the experiment in Example 3. Figure 2 Figure 2 Shows the dynamics of plasma viral load and CD4 + T cell numbers in the monkey (#137) administered only the anti-HIV drug. Figure 3 Figure 3 Shows the dynamics of plasma viral load and CD4 + T cell numbers in the monkey (#139) administered only the anti-HIV drug. Figure 4 Figure 4 Shows the dynamics of plasma viral load and CD4 + T cell numbers in the monkey (#142) administered low doses of SHIV-Ag85B twice. Figure 5 Figure 5 Shows the dynamics of plasma viral load and CD4 + T cell numbers in the monkey (#141) administered a high dose of SHIV-Ag85B once. Figure 6 Figure 6 Shows the dynamics of plasma viral load and CD4 + T cell numbers in the monkey (#140) administered high doses of SHIV-Ag85B multiple times.​​​​​​​​​​​​ Figure 7 Figure 7 Shows the plasma viral load and CD4 + T cell numbers dynamics in monkeys (#138) administered low and high doses of SHIV-Ag85B once respectively. Figure 8 Figure 8 Shows the dosing schedule for each individual in Example 2. Figure 9A Figure 9A Shows the trend of change in plasma viral load in monkey (#138) administered low dose of SHIV-Ag85B twice in Example 3. Figure 9B Figure 9B Shows the trend of change in plasma viral load in monkey (#142) administered high dose of SHIV-Ag85B once in Example 3. Figure 10 Figure 10 In, for one monkey (#141) administered SHIV-Ag85B of a certain dose in Example 3, only short-term effects were seen initially. Each time the virus appeared in the plasma, dosing was carried out, and a long-term inhibitory effect was shown after the 5th inoculation. Figure 11A Figure 11A Is the result of untreated monkeys (#137) administered only anti-HIV drugs in Example 4. Figure 11B Figure 11B Is the result of untreated monkeys (#139) administered only anti-HIV drugs in Example 4. Figure 12 Figure 12 Is the experimental protocol for verifying the trend of change in plasma viral load when administering the composition of the present application after inoculating with HIV virus in Example 4. Figure 13 Figure 13 Is the result showing that anti-HIV drugs function regardless of individual differences in Example 4. Figure 14 Figure 14 Shows the graph of plasma viral load after administering high dose (5.0×10 4 TCID 50 ) of SHIV-Ag85B to monkeys (#164, #165 and #167) at 1-week intervals for 4 times. Figure 15 Figure 15 Is for the monkeys in Example 5 administered high dose (5.0×10​​​​​​​​​​​​​​​​​​​​​​4 TCID 50 Results of plasma viral load when administering a high dose of SHIV-NI four times to monkeys (#162, #166, and #168) with SHIV-NI Figure 16 ) Figure 16 Results of plasma viral load in the control group without Ag85B in Example 5 two weeks after drug withdrawal. Detailed implementation mode

[0008] The present application will be described below. Throughout this specification, unless otherwise specifically mentioned, an expression in the singular form should be understood to also include the concept of its plural form. Therefore, for articles in the singular form (such as "a", "an", "the", etc. in English), unless otherwise specifically mentioned, they should be understood to also include the concept of their plural forms. In addition, for the terms used in this specification, unless otherwise specifically mentioned, they should be understood to be used in the meaning commonly used in the field. Therefore, unless otherwise defined, all professional terms and scientific and technical terms used in this specification have the same meaning as that commonly understood by those skilled in the art to which this application belongs. In case of contradiction, this specification (including definitions) shall prevail.

[0009] (Definition of terms) The following gives examples of the definitions of terms specifically used in this specification.

[0010] In this specification, "about" means ±10% of the indicated value.

[0011] ​In this specification, "Ag85B" refers to an immunogenic protein secreted by acid-fast bacilli. Acid-fast bacilli refer to the property that when staining bacteria with pigments, the pigments are not decolorized by acids, that is, they show resistance to acids. Acid-fast bacilli (Mycobacteria) are roughly classified into Mycobacterium tuberculosis, Mycobacterium leprae, and non-tuberculous mycobacteria. A representative source of Ag85B is Mycobacterium tuberculosis. Ag85B is also known as antigen 85-B, 85B, extracellular alpha-antigen, Antigen 85 complex B, Ag85B, Mycolyltransferase 85B, EC 2.3.1.-, Fibronectin-binding protein B, 30kDa extracellular protein, fbpB, A85B, Major Secretory Protein Antigen 85B, etc. The representative Accession Number is Q847N4. Refer to https: / / www.uniprot.org / uniprotkb / Q847N4 / entry. For the ID of the nucleic acid sequence, AY207396g is representative, and for the ID of the protein sequence, AAO62005.1 is representative. In this specification, it is shown as Sequence No. 1 (nucleic acid sequence) and Sequence No. 2 (amino acid sequence), but it is not limited thereto. It can be understood that as long as it is an immunogenic protein secreted by acid-fast bacilli, it is within the scope of this application.

[0012] In this specification, "operable" means that the transcription or translation of a nucleic acid sequence is under the regulation of an expression regulatory element, and the transcription or translation of the nucleic acid sequence is appropriately regulated and functionally expressed.

[0013] In this specification, "Th1-type immune response" is a cell-mediated immune response mediated by T lymphocytes, and refers to a response caused by cytokines and / or chemokines produced by activated T cells.

[0014] In this specification, "Th2-type immune response" refers to a humoral immune response mediated by secreted antibodies produced by B cells.

[0015] In the present specification, an "attenuated virus" refers to a virus that, although having reduced pathogenicity compared to the parental virus, retains the ability to induce an immune response. Whether a virus has been attenuated can be confirmed by determining its toxicity to cells and / or its pathogenicity to animals. Preferably, it is determined through animal experiments or the like. Generally, a term such as "attenuation" refers to artificially reducing the toxicity of a pathogen by mutating the genes in such a way that the pathogen loses its pathogenicity but retains its immunogenicity. Usually, the toxicity of a pathogen is attenuated by UV irradiation, chemical treatment, or continuous high-order passage culture in vitro to achieve attenuation. The genes are artificially altered, for example, by deleting specific nucleotides in a known sequence to weaken the toxicity.

[0016] In the present specification, "AIDS virus" refers to a virus that causes acquired immunodeficiency syndrome (AIDS), and examples thereof include: human immunodeficiency virus (HIV), simian immunodeficiency virus (SIV), simian / human immunodeficiency (chimeric) virus (SHIV), feline immunodeficiency virus (FIV), and the like.

[0017] In the present specification, "simian / human immunodeficiency (chimeric) virus (SHIV)" refers to a simian / human immunodeficiency chimeric virus in which at least a part of the SIV gene is replaced with an HIV-1 gene. For SHIV, in addition to the env gene, the vpr, rev, tat, and vpu genes can be further replaced with HIV-1.

[0018] Nef (negative factor) is a protein of approximately 27 kDa and is encoded on the 3'-end side of the HIV-1 gene. It was originally reported as a factor that inhibits virus proliferation, but it was later found to promote virus proliferation in primary cultured cells and in vivo, and is related to the pathogenicity of HIV-1. There are known HIV-1 Nef proteins, the anchor domain superfamily (IPR027480), HIV-1 Nef proteins, the core domain superfamily (IPR027481), etc. Various vaccine viruses attenuated by disrupting major regulatory genes such as nef, vpx, vpr, and vif are used. In almost all studies, the moderately attenuated prototype vaccine strain SIVmac239Δnef is used. Nef-deleted attenuated simian immunodeficiency virus (SIV) and simian-human immunodeficiency virus (SHIV) have been proven to be highly effective as vaccines in non-human primate models, but as templates for human HIV vaccines, they cannot be said to have sufficient safety. According to multiple studies, it has been found that although it has been proven that when cytokine or chemokine genes are inserted into attenuated and genetically defective SIV or SHIV, compared with safe and low-toxic virus strains, their immunogenicity is improved and the virus defense ability is enhanced, the nef-deleted attenuated HIV-Ag85B of the present application unexpectedly substantially disappears in patients already infected and with the virus present in the body through post-infection administration, that is, it is cured until the virus is completely cleared from the organism (representatively, a state where it is below the detection limit even by highly sensitive detection methods such as PCR and no clinical symptoms are shown). In addition, it is determined that the nef-deleted attenuated HIV-Ag85B of the present application or its functional equivalent is not only highly safe and low-toxic for post-infection subjects, but can also effectively induce a Th1-type immune response, and can provide a therapeutic composition that can practically tolerate HIV.

[0019] The SIV infection model in non-human primates used in this specification is crucial for the analysis of the pathogenesis of acquired immunodeficiency syndrome (AIDS) and the determination of the effectiveness of HIV vaccines or HIV treatment interventions. As also reported by the present inventors, in cynomolgus monkeys in Indonesia, Malaysia, and the Philippines, they have the pathogenicity of SIVmac and SHIV89.6P. Comparing plasma viral load, peripheral blood CD4 +Results of parameters such as the number of T cells, the pattern of virus antigen-specific immune responses, and the outcome of the disease showed that SIV and SHIV caused disease in cynomolgus monkeys even with different origins. Compared with Indian rhesus monkeys, cynomolgus monkeys from Asia were equally tolerant to various virus strains of SIVmac and SHIV, and had a longer survival period after infection with SIV or SHIV, indicating that cynomolgus monkeys are a model close to the course of HIV-1 disease development in humans.

[0020] In this specification, "complete virus clearance" refers to the state where the virus has been completely cleared from the organism. The state where the virus has been completely cleared from the organism means that it is below the detection limit even according to highly sensitive detection methods such as PCR, and no clinical symptoms are shown.

[0021] In this specification, a "vaccine" refers to a substance containing an antigen or a substance encoding an antigen that provides active immunity against the substance containing the antigen without causing disease. In this specification, the vaccine is a substance for prophylactic use. In this specification, a "DNA vaccine" refers to a nucleic acid encoding a vaccine antigen. Since DNA (especially plasmid DNA) is mainly used, such a general name is used. In addition, as an implementation method using nucleic acids, there is also a method of assembling into a virus vector and then delivering it to the organism. In this case, it can be understood that it can also be provided in the form of nucleic acids other than DNA. Such a situation is also referred to as a "nucleic acid vaccine". Usually, a DNA vaccine takes the form of plasmid DNA. When the DNA vaccine is administered subcutaneously in the form of plasmid DNA, the plasmid DNA is taken up by subcutaneous cells, and the target antigen protein is produced in the cells.

[0022] In this specification, an "antigen" (antigen; also referred to as Ag) refers to any substrate that can be specifically bound by an antibody molecule. In this specification, an "immunogen" refers to an antigen that can cause the activation of lymphocytes that produce an antigen-specific immune response.

[0023] In this specification, "remission" refers to a healthy state where the virus has been completely cleared and no therapeutic drugs need to be administered.

[0024] In this specification, the "treatment" of "viral infection" refers to alleviating at least one symptom caused by viral infection, delaying the development of at least one symptom, or relieving at least one symptom.

[0025] In this specification, "substantially disappearing the virus" refers to the situation where the virus cannot be detected clinically and is determined not to have developed the disease.

[0026] In this specification, an "object" refers to a mammal including a human patient that is the treatment object of this application.

[0027] In this specification, "nucleic acid construct" refers to a DNA or RNA molecule containing a nucleotide sequence encoding a protein.

[0028] In this specification, "deletion" means the disappearance of a gene. Deletion also includes the disappearance of a part of a gene so that it cannot function.

[0029] In this specification, "administration X times" means administration X times during the observation period.

[0030] In this specification, "TCID 50 " refers to the 50% tissue culture infective dose, which means the amount of virus that infects 50% of the cells.

[0031] In this specification, "administered anti-HIV drug" means an anti-HIV drug that has been administered before the nucleic acid construct of the present application is administered.

[0032] In this specification, "not yet administered anti-HIV drug" means that no anti-HIV drug has been administered when the nucleic acid construct of the present application is administered.

[0033] In this specification, "reverse transcriptase inhibitor" refers to an agent that inhibits the enzymatic function of reverse transcriptase, prevents the synthesis of double-stranded viral DNA, and prevents the proliferation of HIV.

[0034] In this specification, "protease inhibitor" refers to an agent that binds to the enzymatic active site of protease to abolish its activity. The functional proteins of HIV are first produced as polyproteins and only function after being cleaved at specific sites by the protease of HIV itself. The protease inhibitor abolishes the activity of protease, and as a result, the virus does not become a complete form and loses its infectivity.

[0035] In this specification, "integrase inhibitor" refers to an agent that inhibits the reaction of HIV integrating its own gene into the host chromosome by inhibiting integrase, and has the effect of preventing the establishment of infection. Thus, since no new infectious virus particles are produced, the spread of infection is prevented.

[0036] In this specification, "CCR5 inhibitor" refers to an agent that inhibits the C-C chemokine receptor 5 (CCR5), an auxiliary receptor used when HIV invades cells.

[0037] In this specification, "complete clearance of HIV" means that it is determined that HIV and other viruses cannot be detected not only clinically but also by any detection method.

[0038] In this specification, "nucleic acid construct", "construct", "construct", or "gene construct" can be used interchangeably, and it is a nucleic acid molecule containing a collection of nucleic acids that are separated from naturally occurring genes or combined and juxtaposed in a non-naturally occurring form.

[0039] For amino acids, in this specification, they can be referred to by either the commonly known three-letter symbols or the single-letter symbols recommended by the IUPAC-IUB Biochemical Nomenclature Commission. Nucleotides can also be referred to by the commonly recognized single-letter codes. In this specification, for the comparison of similarity, identity, and homology of amino acid sequences and base sequences, BLAST, a sequence analysis tool, is used to perform the calculation with default parameters. The search for identity can be performed, for example, using BLAST 2.2.28 of NCBI (released in April 2013) (Proc. Natl. Acad. Sci. USA 90: 5873-5877, 1993). The identity value in this specification generally refers to the value obtained when performing sequence alignment using the above BLAST under default conditions. However, when a higher value appears by changing the parameters, the highest value is taken as the identity value. When evaluating identity in multiple regions, the highest value among them is taken as the identity value. Similarity is a value that includes similar amino acids in addition to identity in the calculation. For the algorithm when comparing amino acid sequences using BLAST, Blastp can be used with the default settings. The measurement results are quantified as similarity (Positives) or identity (Identities). The homology of amino acid sequences and base sequences can be determined by the BLAST algorithm from 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 base sequences by BLASTN based on BLAST, the parameters are set, for example, as score = 100 and wordlength = 12. In addition, when analyzing amino acid sequences by BLASTX based on BLAST, the 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. The specific techniques of these analysis methods are well known (http: / / www.ncbi.nlm.nih.gov.).

[0040] For the nucleic acids or proteins used in the present application, the amino acid or nucleotide sequence of the subject may include a sequence in which one or more amino acids or nucleotides are substituted, deleted, and / or added. Among them, in the amino acid sequence of the total length of the chimeric protein, "one or more" is usually within 50 amino acids, preferably within 30 amino acids, and more preferably within 10 amino acids (for example, within 5 amino acids, within 3 amino acids, 1 amino acid). In addition, in the amino acid sequence of the domain, "one or more" is usually within 6 amino acids, preferably within 5 amino acids, and more preferably within 4 amino acids (for example, within 3 amino acids, within 2 amino acids, 1 amino acid). When maintaining the biological activity of the chimeric protein of the present application, among the mutated amino acid residues, it is desirable to mutate into other amino acids in which the properties of the amino acid side chain are retained. Examples of the properties of the amino acid side chain 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 atom-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 side chains (H, F, Y, W) (the parentheses all represent the single-letter symbols of amino acids). These are also referred to as "conservative substitutions" in this specification. In addition, it is well known that a protein having an amino acid sequence modified by deletion, addition, and / or substitution of one or more amino acid residues for a certain amino acid sequence maintains its 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 application, "several" may be, for example, 10, 8, 6, 5, 4, 3, or 2, or may be any value or less. The chimeric protein to which deletion or the like is applied can be produced, for example, by site-directed mutagenesis, random mutagenesis, or biopanning using an antibody phage library.In the present application, "more than 70%" can be, for example, 70, 75, 80, 85, 90, 95, 96, 97, 98, 99% or more, "more than 80%" can be, for example, 80, 85, 90, 95, 96, 97, 98, 99% or more, "more than 90%" can be, for example, 90, 95, 96, 97, 98, 99% or more, and can also be within the range of any two of these values. For "homology", the proportion of the number of identical amino acids in the amino acid sequences between two or more sequences can be calculated according to methods well-known in the art. Before calculating the proportion, the amino acid sequences of the amino acid sequence groups to be compared are aligned, and in order to maximize the proportion of identical amino acids, gaps are introduced into some amino acid sequences if necessary. The methods for alignment, the methods for calculating the proportion, the comparison methods, and the computer programs related to these have been well-known in the art (for example, BLAST, GENETYX, etc.). For "identity", the proportion of identical amino acids is calculated, and for "similarity", the proportion of similar amino acids is calculated. Examples of similar amino acids include amino acids that can be conservatively substituted, but are not limited to these.

[0041] In the present application, a part of the constructs specifically shown in the present application is also included in the scope of the present application. In this specification, "a part", "fragment" means a polypeptide or polynucleotide having a sequence length from 1 to n - 1 with respect to the full-length polypeptide or polynucleotide (length n). The length of the fragment can be appropriately changed according to its purpose. For example, as the lower limit of its length, in the case of a polypeptide, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 40, 50 or more amino acids can be cited, and lengths represented by integers not specifically exemplified here (such as 11, etc.) can also be suitably used as the lower limit. In addition, in the case of a polynucleotide, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 40, 50, 75, 100 or more nucleotides can be cited, and lengths represented by integers not specifically exemplified here (such as 11, etc.) can also be suitably used as the lower limit. In this specification, such a fragment can be understood to be included in the scope of the present application as long as the fragment itself has the function of a vaccine when the full-length functions as a vaccine. In the present application, "functional equivalents" of the content (constructs, etc.) specifically shown in the present application are also included in the present application. In this specification, "functional equivalents" means any substance that has the same purpose function but different structures with respect to the original entity as the object.

[0042] According to the present application, the term "activity" in this specification refers to the function of a molecule in the broadest sense. The activity is not intentionally limited, but generally includes the biological function, biochemical function, physical function or chemical function of the molecule. Activity includes, for example, enzyme activity, the ability to interact with other molecules, and the ability to activate, promote, stabilize, hinder, inhibit or destabilize the functions of other molecules, stability, and the ability to localize to a specific intracellular location. When applicable, the term is additionally related to the function of a protein complex in the broadest sense. In this specification, "biological activity" includes the activation of photoreactions, etc.

[0043] In this specification, a "functional equivalent" refers to any substance that has the same function for the purpose but a different structure relative to the original entity that is the object. Therefore, the functional equivalent of a "protein encoded by the HIV virus" or its chimera is not the protein encoded by the HIV virus or its chimera itself, but a mutant or modified form (such as an amino acid sequence modifier, etc.) of the protein encoded by the HIV virus or its chimera, and can be understood to include substances having the biological effects of the protein encoded by the HIV virus or its chimera, and substances that can be changed into the protein encoded by the HIV virus or its antibody itself or a mutant or modified form of the protein encoded by the HIV virus or its chimera at the time of action (such as including nucleic acids encoding the protein encoded by the HIV virus or its chimera or a mutant or modified form of the protein encoded by the HIV virus or its chimera, and vectors, cells, etc. containing such nucleic acids). As a functional equivalent of the present application, a sequence in which one or more amino acids are inserted, substituted, and / or deleted or added to one or both ends thereof can be used. In this specification, "inserting, substituting, and / or deleting one or more amino acids in the amino acid sequence or adding one or more amino acids to one or both ends thereof" means modification by known technical methods such as site-directed mutagenesis or by natural mutations, by substitution of a plurality of amino acids to the extent that can occur naturally. The modified amino acid sequence can be a sequence in which, for example, 1 to 30, preferably 1 to 20, more preferably 1 to 9, further preferably 1 to 5, and particularly preferably 1 to 2 amino acids are inserted, substituted, or deleted or added to one or both ends thereof. The modified amino acid sequence preferably can be an amino acid sequence having one or more (preferably one or more or 1, 2, 3, or 4) conservative substitutions in the amino acid sequence of the protein encoded by the HIV virus.

[0044] In this specification, "treatment" refers to, with respect to a certain disease or damage (such as damage caused by viral infection), when it becomes such a state, preventing the deterioration of such a disease or damage, preferably maintaining the status quo, more preferably alleviating it, and further preferably causing it to subside, including being able to exert an effect of improving the symptoms of the patient's disease or one or more symptoms accompanying the disease. Sometimes, performing a diagnosis in advance and carrying out appropriate treatment is called "companion treatment", and the diagnostic drug used for this is called "companion diagnostic drug".

[0045] In this specification, "therapeutic agent" generally refers to all agents capable of treating a target state (such as retinal degenerative diseases, etc.). In one embodiment of the present application, the "therapeutic agent" can be a pharmaceutical composition containing an active ingredient and one or more pharmacologically acceptable carriers. The pharmaceutical composition can be manufactured by, for example, mixing the active ingredient with the above-mentioned carrier by any method known in the technical field of pharmaceutics. In addition, as long as the therapeutic agent is a drug for treatment, its usage form is not limited, and it can be a single active ingredient or a mixture of the active ingredient and any component. In addition, the shape of the above-mentioned carrier is not particularly limited, and it can be, for example, solid or liquid (such as buffer solution).

[0046] The composition of the present application can be provided as a kit. In this specification, a "kit" refers to a unit that provides parts that should generally be provided in two or more divided areas (such as multiple nucleic acid constructs, components obtained by lyophilizing a drug, a buffer solution for administration, instructions, etc.). It is advantageous that such a kit preferably has instructions or a manual describing how to use the provided parts (such as nucleic acid constructs) or how to handle the reagents. This specification further includes instructions such as those describing the usage method.

[0047] The "active ingredient" in this specification refers to the ingredient contained in the composition of the present application, etc. in a necessary amount to obtain the desired therapeutic or inhibitory development effect, etc., and other ingredients can also be contained as long as the effect is not impaired to a level lower than the expected level. In addition, the drugs, compositions, etc. of the present application can also be formulated drugs, compositions. In addition, the administration route of the drugs, compositions, etc. of the present application can be either oral or parenteral, and can be appropriately set according to the form of the preparation, etc. The construct of the present application can be used as an active ingredient.

[0048] In this specification, the "Instructions" (including package inserts for drugs, labels adopted by the US FDA, etc.) describe the method of using the present application for doctors or other users. The Instructions contain words indicating the administration of the drugs of the present application, etc. In addition, the Instructions also contain words indicating intravenous administration, etc. (e.g., by injection, etc.) as the administration site. The Instructions are prepared in accordance with the format specified by the regulatory authorities of the country where the present application is implemented (e.g., the Ministry of Health, Labour and Welfare in Japan, the Food and Drug Administration (FDA) in the US, etc.), and clearly state the gist of being recognized by the regulatory authorities. The Instructions are so-called package inserts, labels, and are usually provided in the form of paper, but are not limited thereto. For example, they can also be provided in the form of electronic media (e.g., providing a homepage, PDF, email via the Internet).

[0049] (Preferred Embodiment) The following describes the description of the preferred embodiment, but this embodiment is an example of the present application, and it should be understood that the scope of the present application is not limited to such a preferred embodiment. In addition, those skilled in the art should understand that, with reference to the following preferred examples, modifications, changes, etc. within the scope of the present application can be easily made. Regarding these embodiments, those skilled in the art can appropriately combine any embodiments.

[0050] In one aspect, the present application provides a composition for treating viral infections in a subject, and related methods and uses, wherein the composition, etc. operably contains a nucleic acid construct 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.

[0051] In a preferred embodiment, the subject of the present application is a subject after being infected with a virus, and the composition of the present application is administered after the infection.

[0052] In a preferred embodiment, the treatment of the viral infection in the present application is to substantially eliminate the virus from the subject after being infected with the virus, preferably to completely eliminate it.

[0053] Among them, the virus "substantially" "disappears" means that viruses such as the AIDS virus cannot be detected clinically and is determined not to have developed.

[0054] Among them, the virus "completely" "disappears" means that it is determined that the virus such as the AIDS virus cannot be detected by any detection method (e.g., even by highly sensitive PCR examination) and no clinical symptoms are shown.

[0055] Therefore, in a preferred embodiment, the present application provides a composition for administration for the purpose of substantially eliminating a virus from a virus-infected subject.

[0056] In one embodiment, the nucleic acid sequence encoding the antigenic protein or a part thereof to be used is a nucleic acid sequence encoding an attenuated virus. Preferably, the attenuated virus is an Nef-deleted attenuated virus. Since the Nef gene is a gene responsible for immune escape of the AIDS virus, when this part is deleted, the virus is attenuated through the immune function of the organism. It has not been envisioned to use an attenuated virus together with such an adjuvant antigen.

[0057] In one embodiment, the nucleic acid sequence encoding the Ag85B protein used in the present application is assembled into the nucleic acid sequence encoding the attenuated virus of the present application. Since the Nef gene is a gene responsible for immune escape of the AIDS virus, when this part is deleted, the virus is attenuated through the immune function of the organism. If Ag85B is added, strong cellular immunity is induced, a strong immune response is induced, and the virus is further attenuated.

[0058] In one embodiment, the above-mentioned nucleic acid sequence encoding the Ag85B protein used in the present application is assembled at the position of the deleted Nef gene in the nucleic acid sequence encoding the Nef-deleted attenuated virus. Since the Nef gene is a gene responsible for immune escape of the AIDS virus, when this part is deleted, the virus is attenuated through the immune function of the organism. If Ag85B is added, strong cellular immunity is induced, a strong immune response is induced, and the virus is further attenuated.

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

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

[0061] In one embodiment, the virus used is an attenuated virus 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 the 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 viral type in the world, including Japan, and is classified into three groups: group M (Main), which accounts for the majority, and groups O (Outlier) and N (New), which have been identified in limited regions. Group M is further classified into nine subtypes (A - D, F - H, J, K). On the other hand, HIV-2 is classified into subtypes A - G and is mainly confined to West Africa, although infection cases 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, such as, but not limited to, BaL strain, IIIB strain, RF strain, GB8 strain, U455 strain, ROD strain, and mutants thereof.

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

[0066] In one embodiment, the virus used is a nef-deleted attenuated SHIV. Since the nef gene is responsible for the immune escape of the AIDS virus, when this part is deleted, the virus is attenuated by the immune function of the organism. If Ag85B is added, it induces a strong cellular immune response, inducing a strong immune reaction, and the virus is further attenuated.

[0067] The nucleic acid sequence encoding the antigenic protein or a part thereof used is at least 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to the nucleic acid sequence comprising nucleotides 1 - 9633 and 10612 - 11022 of SEQ ID NO: 3 or the nucleic acid sequence comprising nucleotides 1 - 8786 and 9765 - 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, at least 99% identical to the nucleic acid sequence set forth in SEQ ID NO: 3 or 4. Preferably, the nucleic acid construct used includes the nucleic acid sequence set forth in SEQ ID NO: 3 or 4. More preferably, it is a nucleic acid sequence having one or several modifications in the nucleic acid sequence set forth in SEQ ID NO: 3 or 4 (in particular, the modification can be a conservative substitution, and the modification position can be a position that has no effect on the activity), and particularly preferably, it is the nucleic acid sequence set forth in SEQ ID NO: 3 or 4. In particular, constructs having the sequence of SEQ ID NO: 3 or 4 of the present application have been proven to not only have high safety and low toxicity, but also be able to provide, for the first time, a therapeutically useful composition that is tolerable to HIV, which is of great clinical significance.

[0069] In the amino acid sequence encoded by the construct of the present application, "one or several" is usually within 6 amino acids, preferably within 5 amino acids, and more preferably within 4 amino acids (for example, within 3 amino acids, within 2 amino acids, 1 amino acid). When the chimeric protein maintains the biological activity of the present application, it is desirable that the mutated amino acid residue is mutated to another amino acid in which the nature of the amino acid side chain is retained. When being modified, as the nature of the amino acid side chain, a conservative substitution is preferred. In one embodiment of the present application, "several" can be, for example, 10, 8, 6, 5, 4, 3, or 2, or can be any value below these. The chimeric protein to which deletions, etc. are applied can be produced, for example, by site-directed mutagenesis, random mutagenesis, or biopanning using an antibody phage library. As a method for site-directed mutagenesis, for example, the KOD-Plus-Mutagenesis kit (TOYOBO CO., LTD.) can be used.

[0070] (Dosage and Administration) In various embodiments, the compositions of the present application can be used in various dosages and administrations.

[0071] In one embodiment, with respect to the composition of the present application, the nucleic acid construct is administered once or more than twice, for example, 3 times, 4 times, 5 times, or more than these times.

[0072] From the perspective of simplicity, administration once is preferred, but in order to achieve an exact therapeutic effect, repeated administration, high-dose administration, or a combination of these can also be used. Those skilled in the art can appropriately determine the number of administrations, frequency, and dose.

[0073] For example, in the present application, the nucleic acid construct or composition is administered once a week, once every two weeks, once every three weeks, once a month, 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 application is administered in an amount of 1.0×10 3 TCID 50 or more. Alternatively, in other embodiments, the composition or nucleic acid construct of the present application is 1.0×10 1 TCID 50 or more, 2.0×10 1 TCID 50 or more, 3.0×10 1 TCID 50 or more, 4.0×10 1 TCID 50 or more, 5.0×10 1 TCID 50 or more, 6.0×10 1 TCID 50 or more, 7.0×10 1 TCID 50 or more, 8.0×10 1 TCID 50 or more, 9.0×10 1 TCID 50 or more, 1.0×10 2 TCID 50 or more, 2.0×10 2 TCID 50 or more, 3.0×10 2 TCID 50 or more, 4.0×10 2 TCID 50 or more, 5.0×10 2 TCID 50 or more, 6.0×10 2 TCID 50 or more, 7.0×10 2 TCID 50 or more, 8.0×10 2 TCID 50 or more, 9.0×10 2 TCID 50 or more, 1.0×10 3 TCID 50 or more, 2.0×10 3 TCID 50 or more, 3.0×10 3 TCID 50 or more, 4.0×10 3 TCID 50 or more, 5.0×10 3 TCID50 Above, 6.0×10 3 TCID 50 Above, 7.0×10 3 TCID 50 Above, 8.0×10 3 TCID 50 Above, 9.0×10 3 TCID 50 Above, 1.0×10 4 TCID 50 Above, 2.0×10 4 TCID 50 Above, 3.0×10 4 TCID 50 Above, 4.0×10 4 TCID 50 Above, 5.0×10 4 TCID 50 Above, 6.0×10 4 TCID 50 Above, 7.0×10 4 TCID 50 Above, 8.0×10 4 TCID 50 Above, 9.0×10 4 TCID 50 Above, or 1.0×10 5 TCID 50 Above.

[0075] In one embodiment, the dosage of the nucleic acid construct or composition is administered at a dosage of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 of the dosage.

[0076] In some embodiments, it can also be administered more than twice a week at a low dosage, a high dosage, or both dosages, such as administered 2, 3, 4, 5 times or more than these times. In certain embodiments, after administering the low dosage, the high dosage can also be administered. In certain embodiments, the high dosage can also be administered multiple times, such as administered 4 times.

[0077] In one embodiment, the nucleic acid construct or composition is at 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50When the nucleic acid construct is administered once at a certain dosage and the virus is detected in the body, the above nucleic acid construct is further administered. The detection of the virus can be carried out by antigen examination, PCR examination, etc.

[0078] In one embodiment, the nucleic acid construct or composition is administered at a low dosage of 5.0×10 4 TCID 50 or more and 5.0×10 6 TCID 50 or less.

[0079] In one embodiment, the nucleic acid construct or composition is administered once at a high dosage of 5.0×10 4 TCID 50 or more.

[0080] In one embodiment, for the subject of the nucleic acid construct or composition, it can be administered to a subject who has been administered an anti-HIV drug. Examples of the anti-HIV drug that has been administered include reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors. For example, it can be mentioned: tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir mixture, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances, etc.

[0081] In one embodiment, for the subject of the nucleic acid construct or composition, it can be administered to a subject who has not been administered an anti-HIV drug or a subject who has discontinued the anti-HIV drug when starting to administer the above composition.

[0082] In one embodiment, the other anti-HIV drugs used can be selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors.

[0083] In one embodiment, the anti-HIV drugs that can be used together with the nucleic acid construct or composition of the present application can be anti-HIV drugs selected from the group consisting of tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir mixture, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances.

[0084] In one embodiment, the nucleic acid construct or composition completely clears the human immunodeficiency virus (HIV) in a subject, and such a substance has never been provided. In addition, in individuals in whom complete clearance cannot be achieved, the onset of the disease can also be continuously suppressed and health can be maintained by continuously administering the nucleic acid construct or composition of the present application.

[0085] In one embodiment, the nucleic acid construct or composition is not administered to the subject before viral infection. It has never been known to completely clear the infecting virus of the subject such as the human immunodeficiency virus in such a subject.

[0086] (Method for manufacturing the construct) The construct of the present application can be manufactured as follows. Representative examples are described in the Examples.

[0087] The virus is isolated from the patient's body, and the virus is produced from molecules other than nef (for example, gag, pol, vif, vpx, vpr, vpu, tat, env, rev, or a combination thereof). At this time, the Ag85B gene is inserted into the nef region. Ag85B can be genetically isolated by the BCG vaccine. By assembling the plasmid, it is inserted into the nucleic acid sequence of the virus.

[0088] The construct can be produced by assembling Ag85B into a known virus strain. For example, if it is a type I HIV (HIV-1) virus, typically the HIV-1 NL432 strain can be used. In addition, examples include: BaL strain, IIIB strain, RF strain, GB8 strain, U455 strain, ROD strain, and mutant strains thereof, etc., but are not limited to these.

[0089] (Use of the construct) The construct of the present application can be used for the treatment of HIV. In one aspect, the present application provides a composition for treating a viral infection in a subject, the composition containing a nucleic acid construct, the nucleic acid construct operably containing 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. The nucleic acid construct of the present application can treat viral infections independently of the Th2-type immune response.

[0090] In some embodiments, the nucleic acid sequence encoding the antigen protein or a part thereof may contain 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 further aspects, the present application provides a pharmaceutical composition for treating viral infections in a subject, the pharmaceutical composition operably comprising a nucleic acid construct encoding an antigen protein or a portion thereof contained in a virus belonging to the genus Lentivirus and a nucleic acid sequence encoding an Ag85B protein. The treatment of viral infections does not rely on a Th2-type immune response and can be achieved by inducing a Th1-type immune response. Treatment after HIV infection has been achieved for the first time by the present invention.

[0092] The composition of the present application is capable of completely clearing the virus in a subject. Particularly in the case of HIV infection, complete clearance of the virus from the subject has not been achieved to date. Complete clearance of the virus can be achieved by a Th1-type immune response.

[0093] In a preferred embodiment, the subject has been previously vaccinated with the BCG vaccine. Since it is expected that vaccination with the BCG vaccine enhances the adjuvant effect of Ag85B, a high therapeutic effect against HIV obtained from HIV-Ag85B is anticipated in the BCG vaccination group. For the BCG vaccine, there are also countries (such as the United States) where vaccination has not been carried out due to contraindications, etc., and it is not necessarily required to vaccinate with the BCG vaccine. Sufficient effects are also expected in the group not vaccinated with BCG.

[0094] (Drug, treatment method) In one aspect, the present application provides a treatment method using the composition or drug of the present application. In one embodiment, the composition or nucleic acid construct of the present application is administered by injection. In other embodiments, the composition or nucleic acid construct of the present application is administered in vivo. In a specific embodiment, the composition or nucleic acid construct of the present application can be provided together with a preservation solution. In some embodiments, the preservation solution can be a buffer solution. In other embodiments, the composition or nucleic acid construct of the present application can be provided in a state contained in a container. In a specific embodiment, the container containing the composition or nucleic acid construct of the present application can be a syringe.

[0095] Administration of an agent related to the attenuated virus of the present application can be carried out by any suitable means of achieving a concentration of a therapeutic agent effective for improving or alleviating HIV, in combination with other components. The agent can be contained in any suitable amount in any suitable carrier substance and is usually present in an amount of 1 to 95% by weight of the total weight of the composition. The composition can be provided in a dosage form suitable for parenteral (e.g., subcutaneous, intravenous, intramuscular or intraperitoneal) administration routes. The pharmaceutical composition can be formulated according to conventional pharmaceutical practice (e.g., see Remington: The Science and Practice of Pharmacy (20th edition), A.R. Gennaro, Lippincott Williams and Wilkins, 2000 and Encyclopedia of Pharmaceutical Technology, J. Swarbrick and J.C. Boylan, 1988 - 1999, Marcel Dekker, New York). The pharmaceutical composition of the present application can be formulated to release the active compound almost immediately after administration or at a predetermined time point or after a predetermined time from the start of administration.

[0096] When the present application is used for parenteral administration, it can be formulated in a state contained in a unit dose ampoule or a multi-dose container or vial. In addition, additives such as stabilizers, buffers, preservatives, and permeants can also be contained. In addition, when the formulation for parenteral administration is used, it can also be formulated as a powder that can be redissolved with a suitable carrier (such as sterilized water). Examples of parenteral administration include intravenous administration, intramuscular administration, subcutaneous administration, etc., and intravenous administration is preferred. The active ingredient of the construct described in this specification can be administered together with a carrier. The so-called carrier includes diluents, adjuvants, excipients or vehicles. In addition, if necessary, the composition can contain trace amounts of wetting agents or emulsifying agents or pH buffers. These compositions can be in the form of solutions, suspensions, emulsions, tablets, pills, capsules, powders, sustained release formulations, combinations thereof, etc.

[0097] A pharmaceutically acceptable carrier that can be used in the present application refers to a medium containing the construct or agent described in this specification that can be injected into the body of an object without side effects. Pharmaceutically acceptable carriers include sterile liquids such as water and oils. Oils include oils of petroleum origin, animal origin, plant origin, 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, wheat 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 E.W. Martin's Remington's Pharmaceutical Sciences. Administration of the compositions of the present application can be carried out by any route typically used for inoculation of the compositions of the present application, including local routes, subcutaneous routes, intravenous routes, intramuscular routes, intradermal routes, intraperitoneal routes, oral routes, inhalation routes, or combinations thereof.

[0098] The compositions described in this specification can be formulated into nucleic acid drugs. The nucleic acid drugs described in this specification contain the vectors described in this specification containing the following DNA, which encodes a full-length gene RNA molecule of an infectious non-pathogenic and / or attenuated virus operably linked to a promoter suitable for expression in eukaryotic cells.

[0099] In a specific embodiment, the constructs, drugs, etc. of the present application are administered one or more times during treatment. As described in the examples, it is considered that the drugs, etc. of the present application confirm their effects by being administered at least once, and the compliance of patients is also relatively good.

[0100] (General technology) The molecular biology techniques, biochemistry techniques, and microbiology techniques used in this specification are well-known and commonly used in the art. For example, they are described 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), etc. These are incorporated by reference in relevant parts (which may be all) of this specification.

[0101] The preferred embodiments are shown above for easy understanding to illustrate the present application. The present application is described below based on examples, but the above description and the following examples are provided for illustrative purposes only and not for the purpose of limiting the present application. Therefore, the scope of the present application is not limited to the embodiments and examples specifically described in this specification, but is only limited by the claims. Example

[0102] (Example 1) Method for preparing a construct SHIV-NM3rN having the HIV-1 NL4-3-2 gene in the SIVmac239 background was used as the starting material. The recombinant SHIV was constructed according to the method reported previously (15, 16). The SHIV-nef vector (SHIV-NI) was a vector constructed from the infectious molecular clone of SHIV-NM3rN (48). The source of the env gene of SHIV-NI was HIV-1 NL4-3-2 as an X4-tropic virus. In SHIV-NI, the nef gene was replaced at unique restriction enzyme sites such as ClaI and ApaI. For the Ag85B gene, Mycobacterium kansasii was used as a template, and primers 5'-ATATCGATACCATGTTCTCCCGTCCCGGGCT-3′ (ClaI) (SEQ ID NO: 5) and 5'-AGGGCCCCTAGCGGGCGCCCAGGCTGG-3' (ApaI) (SEQ ID NO: 6) were used for amplification by PCR. Subsequently, the PCR product was digested with restriction enzymes at the ClaI and ApaI sites. This plasmid was designated pSHIV-Ag85B. pSHIV-Ag85B was transfected into 293T cells using the FuGENE 6 transfection reagent (Roche Diagnostics, Indianapolis, IN) to prepare SHIV-Ag85B, and the culture supernatant 48 hours after transfection was stored in liquid nitrogen until use.

[0103] (Example 2) Confirmation of the performance of the construct (Detection of Ag85B protein) Infect M8166 cells with SHIV-Ag85B at an MOI of 0.1 and culture for 1 hour. After washing the cells 3 times with phosphate-buffered saline (PBS), further culture in the culture medium for 48 hours. After further washing 3 times with PBS, lyse the cells with PBS containing 1.5 M urea, 2% NP-40, and 5% 2-mercaptoethanol. Then, separate by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transfer to a nitrocellulose membrane by electroblotting. Block with 5% non-fat dry milk in PBS containing 0.01% Tween 20 (PBST). After washing 3 times with PBST, incubate the membrane with rabbit anti-Ag85B polyclonal antibody for 2 hours. After washing the membrane 3 times with NBT / BCIP (Roche Diagnostics, Mannheim, Germany), incubate with alkaline phosphatase-labeled anti-rabbit IgG (New England Biolabs, Beverly, MA).

[0104] (In vivo stability of the inserted Ag85B gene) Extract proviral DNA from 1×10 6 PBMCs of the inoculated macaque monkeys. When separating the virus, also monitor the CD8 + depleted PBMCs co-cultured with M8166 cells. Extract cellular DNA using the DNeasy Tissue Kit (QIAGEN). To confirm the stability of the inserted Ag85B gene in SHIV-Ag85B, amplify the proviral DNA fragment containing the inserted Ag85B gene in SHIV-Ag85B using primers. The sequences of the primers are as described above.

[0105] (Example 3) Evaluation of SHIV-Ag85B therapeutic nucleic acid drug The outline of the experiment in this example is as Figure 1 shown. In this example, the changing trends of plasma viral load and CD4 + T cell count were verified when administering the composition of the present invention after inoculating with the HIV virus.

[0106] (Materials and methods) (Virus strain) In this study, SHIV-Ag85B, SHIV-NI, and SHIV89.6P were used. These viral strains were propagated using PBMC from cynomolgus monkeys. PBMC isolated by standard Ficoll density gradient separation were cultured in RPMI1640 supplemented with 10% fetal bovine serum, 2 mM L-glutamine, and 100 units / ml of IL-2 (Shionogi & Co., Ltd.), and stimulated with phytohemagglutinin for 72 hours. Cells were 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 3 days, and cell-free supernatants were harvested on days 6 - 9 post-infection. Using M8166 cells, the TCID 50 of each SHIV was measured in the virus stock 50 . The TCID 4 value of the virus stock was 5×10 4 for SHIV-Ag85B, 4.7×10 5 for SHIV-NI, and 3×10

[0107] (Animals) Adult monkeys (from Indonesia, the Philippines, and Malaysia) negative for simian SIV, D-type retrovirus, T-cell lymphoma virus, simian foamy virus, Epstein-Barr virus, cytomegalovirus, and B virus were used. 7 monkeys were intravenously inoculated with 50 TCID 4 of SHIV89.6P. One or two weeks (5 weeks only for #137) after infection with SHIV89.6P, anti-HIV drugs (tenofovir 20 mg / kg, emtricitabine 40 mg / kg, or dolutegravir 2.5 mg / kg) were administered subcutaneously once a day. After discontinuation of anti-HIV drug administration, 5 monkeys were intravenously inoculated with SHIV-Ag85B (high-dose SHIV-Ag85B inoculation; 8×10 50 TCID 4 , low-dose SHIV-Ag85B inoculation; 10 50 TCID 4 , and 10 50 TCID + ). Blood was collected regularly using sodium citrate as an anticoagulant for determination of CD4

[0108] (Preparation of DNA samples and amplification of the SHIV gag gene by nested PCR) To determine proviral DNA in monkeys inoculated with SHIV-Ag85B, a fragment of the gag gene was amplified using nested PCR. Proviral DNA was extracted from the PBMC 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 specifications for the initial and nested PCRs are described in References 49 and 50 (49 and 50). For the primers used in this study, Outer SIVgag-F (5'-CCATTAGTGCCAACAGGCTCAG-3' (SEQ ID NO: 7)) and Outer SIVgag-R (5'-CCCCAGTTGGATCCATCTCCTG-3' (SEQ ID NO: 8)) were used 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)) were used in the second round of PCR. After the second amplification, 10 μl of the product amplified by nested PCR was added to a 1.0% agarose gel and stained with ethidium bromide to visualize the DNA bands. By the first amplification using the outer gag primer pair, the lowest concentration of plasmid SIV DNA detected by this PCR method was 100 copies. Further, when amplified with nested / internal gag primers, single-copy plasmid DNA could be routinely detected (49, 50).

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

[0110] (amount of proviral DNA) DNA specimens were extracted from PBMC and lymphoid tissues using the DNeasy Tissue Kit (QIAGEN) according to the manufacturer's specifications. Ultra-high sensitivity digital PCR was performed using the QX200 Droplet Digital PCR system (Bio-Rad). A 20-μl reaction mixture containing 2 μl of the DNA specimen, ddPCR supermix for probes (without dUTP) (Bio-Rad), 900 nM of each primer, 200 nM of the probe, and deionized water was prepared. This mixture was placed in a DG8 cartridge together with 70 μl of droplet generation oil (Bio-Rad), and droplets were formed using a droplet generator (Bio-Rad). Subsequently, the droplets were transferred to a 96-well microplate. PCR amplification was performed according to the following program: initial denaturation and stabilization at 95°C for 10 minutes, denaturation at 94°C for 30 seconds, annealing / extension at 57°C for 60 seconds for 40 cycles, and then at 98°C for 10 minutes. Subsequently, the droplets were sorted and analyzed using QuantaSoft v1.6 (Bio-Rad) software with a QX200 droplet reader (Bio-Rad). For the specimens, only the cases where more than 20,000 droplets were read were considered. As described above, cell numbers were monitored by high-sensitivity real-time quantitative PCR (53). DNA specimens were extracted from PBMC and lymphoid tissues using the DNA DNeasy Tissue Kit (QIAGEN) according to the manufacturer's specifications. The cellular IL-4 sequence was detected and cell numbers were confirmed using cynomolgus monkey IL-4-specific primers 5'-TGTGCTCCGGCAGTTCTACA-3' (SEQ ID NO: 14) and 5'-CCGTTTCAGGAATCGGATCA-3' (SEQ ID NO: 15) and probe 5'-FAM-TGCACAGCAGTTCCACAGGCACAAG-TAMRA-3' (SEQ ID NO: 16).

[0111] (CD4 + T cell count) One hundred microliters of whole blood from each cynomolgus monkey was stained using fluorescently labeled monoclonal antibodies: anti-CD3 (clone SP34-2, Alexa700, BD), 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) (Only administered anti-HIV drugs) In monkeys (#137 and #139) administered only anti-HIV drugs, the plasma viral load was maintained at a low level, but when the administration of anti-HIV drugs was discontinued, the plasma viral load increased ( Figure 2 and Figure 3 ). In addition, in monkeys (#137 and #139) inoculated with SHIV89.6P and administered anti-HIV drugs, the number of CD4 + T cells was very small during the observation period ( Figure 2 and Figure 3 ).

[0113] (Two administrations of low-dose SHIV-Ag85B) In the monkey (#138) administered two low doses of SHIV-Ag85B, almost no virus could be detected in the plasma after the administration of anti-HIV drugs was discontinued, and the plasma viral load reached below the detection limit 14 weeks after the first administration of SHIV-Ag85B ( Figure 4 ). In addition, in the monkey (#138) administered two low doses of SHIV-Ag85B, the number of CD4 + T cells was maintained at a normal level ( Figure 4 ).

[0114] (One administration of high-dose SHIV-Ag85B) In the monkey (#142) administered one high dose of SHIV-Ag85B, the plasma viral load was maintained below the detection limit throughout the observation period after the administration of anti-HIV drugs was discontinued ( Figure 5 ). In addition, in the monkey (#142) administered one high dose of SHIV-Ag85B, the number of CD4 + T cells was maintained at a normal level ( Figure 5 ).

[0115] (Five administrations of high-dose SHIV-Ag85B) In the monkey (#141) administered high-dose SHIV-Ag85B, if two high doses of SHIV-Ag85B were administered, the plasma viral load decreased to a low level, but later the plasma viral load increased again ( Figure 6 ). When low-dose SHIV-Ag85B was administered again (on days 91, 147, and 154 after infection), the plasma viral load decreased. In such an example, it was also considered that the plasma viral load could be maintained at a low level by multiple administrations of SHIV-Ag85B. CD4 + T cells were maintained at a normal level ( Figure 6 ).

[0116] (Administering low and high doses of SHIV-Ag85B once respectively) In the monkey (#140) administered with a low dose of SHIV-Ag85B on the 66th day after infection, the plasma viral load increased after the 77th day after infection. On the 98th day after infection, a high dose of SHIV-Ag85B was administered, but the plasma viral load was still on the rise ( Figure 7 ). CD4 + T cell counts were at low levels throughout the observation period ( Figure 7 ).

[0117] The experimental method of this example is based on Figure 1 .

[0118] Specifically as follows. Start administering anti-HIV drugs: After the peak of the virus appears in the plasma after infection with SHIV89.6P (except for #137). Administration method: Once a day, subcutaneous administration Anti-HIV drugs: Tenofovir 20 mg / kg Emtricitabine 40 mg / kg Dolutegravir 2.5 mg / kg References (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 in rhesus macaques by a potent d-peptide HIV entry inhibitor. Proc Natl Acad Sci U S A. 2020 Sep 8;117(36):22436-22442. Doi:10.1073 / pnas.2009700117)

[0119] Specifically, it is implemented in substantially the same steps as the above-mentioned SHIV. The dosing schedule in each individual is as described in Figure 8 It is described.

[0120] In the monkeys (#138) administered a low dose of SHIV-Ag85B twice and the monkeys (#142) administered a high dose of SHIV-Ag85B once, the plasma viral load was maintained below the detection limit for a long time after inoculation with SHIV-Ag85B ( Figure 9A and Figure 9B ). In addition, regarding one monkey (#141) administered a high dose of SHIV-Ag85B, only a short-term effect was seen initially, and a long-term inhibitory effect was seen after the 5th inoculation when administered each time the virus appeared in the plasma ( Figure 10 ).

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

[0122] For the monkeys (#138) administered a low dose of SHIV-Ag85B twice, when administered a low dose (1x10 4 TCID 50 ) of SHIV-Ag85B on the 4th and 11th days after discontinuation of the therapeutic drug, the virus in the plasma was maintained below the detection limit, and the CD4 + cells also recovered and were maintained at normal values.

[0123] The explanation of the above results is as described below.

[0124] For the monkeys (#142) administered a high dose of SHIV-Ag85B once, when administered a high dose of SHIV-Ag85B (5x10 4 TCID 50 ) on the 7th day after discontinuation of the therapeutic drug, the virus in the plasma was maintained below the detection limit, and the CD4 + cells also recovered and were maintained at normal values.

[0125] For the monkeys (#141) administered a high dose of SHIV-Ag85B, when administered a high dose of SHIV-Ag85B (5x10 4 TCID 50 ) on the 7th day after discontinuation of the therapeutic drug, the virus in the plasma was maintained below the detection limit only in the short term. Therefore, when administered each time the virus appeared in the plasma, a long-term inhibitory effect was seen after the 5th inoculation. The CD4 + cells also recovered and were maintained at normal values.

[0126] (Example 4) Further clinical trial of SHIV-Ag85B Cynomolgus monkeys were administered a high dose of SHIV-Ag85B 4 times per week. Thereafter, the plasma viral load and CD4 in the cynomolgus monkeys were measured+ The changing trend of the number of T cells. (1) Influence of drug withdrawal Regarding the influence of drug withdrawal, the number of inoculations and the dosage of SHIV-Ag85B were studied, and the same experiment as in Example 3 was conducted. Drug withdrawal was carried out by suspending drug administration for 3 days after 8 weeks of drug administration.

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

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

[0129] Figure 11A As a control for untreated monkeys (#137) given only anti-HIV drugs. On the 7th day after drug withdrawal, virus appeared in the plasma, and its content increased thereafter. CD4 + cells decreased in an inverse proportion to it, and the monkey died 170 days after infection.

[0130] Figure 11B As a control for untreated monkeys (#139) given only anti-HIV drugs. On the 110th day after drug withdrawal, virus appeared in the plasma, and its content increased thereafter. CD4 + cells decreased in an inverse proportion to it, and the monkey died 500 days after infection.

[0131] (2) Optimization of the treatment regimen The experimental outline of this example is as Figure 12 shown. In this example, the changing trend of the viral load in the plasma when the composition involved in this application was administered after inoculating with the HIV virus was verified.

[0132] (Protocol) Start drug administration from the 1st week after infection with SHIV. Starting from the 7th day after discontinuing the therapeutic drug, administer SHIV-Ag85B (5x10 4 TCID 50 ) 4 times every 1 week.

[0133] (Verification technique) (Amount of viral RNA in plasma) As described above, the amount of viral RNA in plasma was measured using highly sensitive real-time quantitative RT-PCR to monitor the level of SHIV infection (23, 51, 52). Viral RNA was isolated from plasma using the MagNA PureCompact nucleic acid isolation kit (Roche Diagnostics). For real-time RT-PCR, the QuantiTec probe RT-PCR kit (Qiagen) and the LightCycler 480 thermal cycler (Roche Diagnostics, Rotkreuz, Switzerland) were used. The gag gene of SIVmac239 was amplified using the probe 5'-FAM-TGTCCACCTGCCATTAAGTCCCGA-TAMRA-3' (where FAM is 6-carboxyfluorescein and TAMRA is 6-carboxytetramethylrhodamine) (SEQ ID NO: 11), the primers 5'-TGGAAGAAAGACCTCCAGAAAATG-3' (SEQ ID NO: 12) and 5'-CAAGTGCAGTTAGCAAGCGAGGAT-3' (SEQ ID NO: 13). The limit of detection was counted as 1000 viral RNA copies / ml.

[0134] (CD4 + T cell count) One hundred microliters of whole blood from each cynomolgus monkey was stained using fluorescently labeled monoclonal antibodies anti-CD3 (clone SP34-2, Alexa700, BD), 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 were as Figure 13 shown. It was shown that the anti-HIV drugs used functioned regardless of individual differences.

[0136] (Example 5: Treatment optimization) In this example, a trial of administering SHIV-Ag85B four times was conducted. It was substantially the same as the above protocol.

[0137] (Four administrations of SHIV-Ag85B) In monkeys (#164, #165, and #167) administered a high dose (5.0 × 10 4 TCID 50 ) of SHIV-Ag85B, when a high dose of SHIV-Ag85B was administered four times at 1-week intervals, the plasma viral load was maintained below the limit of detection throughout the observation period ( Figure 14). In the control group without Ag85B, virus appeared in the plasma 2 weeks after drug withdrawal, while the virus in the plasma of the SHIV-Ag85B administration group remained below the detection limit.

[0138] (4 doses of SHIV-NI) At high doses (5.0×10 4 TCID 50 In monkeys (#162, #166, and #168) that were given SHIV-NI four times, the plasma viral load remained below the detection limit in #168, but increased in #162 and #166 ( Figure 15 In the control group without Ag85B, the virus appeared in the plasma 2 weeks after drug withdrawal, while the virus in the plasma of the SHIV-Ag85B-administered group was maintained below the detection limit (the control group without Ag85B was Figure 15 The SHIV-Ag85B vaccination group was Figure 14 The untreated group Figure 16 ).

[0139] (Example 6) Clinical Trial of HIV-Ag85B In this example, HIV-Ag85B (SEQ ID NO. 4) was subcutaneously or intravenously administered 4 to 8 times at intervals of 1 to 2 weeks to HIV-infected humans (BCG vaccination group and non-BCG vaccination group) who were receiving anti-HIV treatment drugs after stopping the anti-HIV treatment drugs. The therapeutic effect caused by HIV-Ag85B was confirmed.

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

[0141] (Example 7) Formulation Example When the pharmaceutical preparation is prepared, it can be produced according to the following production method.

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

[0143] (Example 2) An appropriate volume of an isotonic solution of 5% glucose solution can be directly added to an appropriate amount of freeze-dried nucleic acid construct to prepare a solution preparation for injection.

[0144] (Example 3) An appropriate volume of an electrolyte modifier solution such as Otsuka Salt Injection 10% etc. can be added to an appropriate amount of a nucleic acid construct dissolved in water to prepare an injection solution preparation with a 0.9% NaCl concentration.

[0145] (Example 4) An appropriate amount of a nucleic acid construct dissolved in water can be lyophilized to obtain a lyophilized 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 Mycobacteria - specific 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 double-stranded 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 replication - competent 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 SHIV-infected 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 latently 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. (Annotation) As described above, the preferred embodiments of the present application are used to illustrate the present application, but the present application should not be construed as being limited to such embodiments. It is understood that the scope of the present application should be construed only in accordance with the claims. It is understood that those skilled in the art can implement equivalent scopes based on the description of the specific preferred embodiments of the present application and the knowledge of the art. For the patents, patent applications, and other documents cited in this specification, it is understood that their contents should be cited as references relative to the present specification in the same manner as the contents specifically described in this specification. This application claims priority from Japanese Patent Application No. 2022-140218 filed with the Japan Patent Office on September 2, 2022, and all of its contents are incorporated herein by reference as needed. Industrial Applicability

[0147] The present application is useful in the field of the pharmaceutical industry. Free Text of Sequence Listing

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

Claims

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

2. The composition according to claim 1, wherein The composition is administered after the subject is infected with the virus.

3. The composition according to claim 2, wherein the treatment of the viral infection is to substantially eliminate the virus from the subject after the subject is infected with the virus.

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

7. The composition according to claim 5 or 6, wherein the nucleic acid sequence encoding the Ag85B protein is assembled 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 assembled at the position of the deleted nef gene in the nucleic acid sequence encoding the nef - deleted attenuated virus.

9. The composition according to any one of claims 1 to 8, wherein the virus is a human immunodeficiency virus.

10. The composition according to any one of claims 1 to 9, wherein the virus is an attenuated virus of a virus that infects humans.

11. The composition according to any one of claims 1 to 10, wherein the virus is an attenuated virus of a human immunodeficiency virus selected from the group consisting of HIV, SIV, SHIV, and FIV.

12. The composition according to any one of claims 1 to 11, wherein the virus is an attenuated HIV.

13. The composition according to any one of claims 1 to 12, wherein the virus is a nef - deleted attenuated HIV.

14. The composition according to any one of claims 1 to 13, wherein the nucleic acid sequence encoding the antigen protein or a part thereof contains a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence comprising nucleotides 1 - 9633 and 10612 - 11022 of SEQ ID NO: 3 or the nucleic acid sequence comprising nucleotides 1 - 8786 and 9765 - 15182 of SEQ ID NO:

4.

15. The composition according to any one of claims 1 to 14, wherein the nucleic acid construct contains a nucleic acid sequence that is at least 90% identical to the nucleic acid sequence recited in SEQ ID NO: 3 or 4.

16. The composition according to any one of claims 1 to 15, wherein the nucleic acid construct contains the nucleic acid sequence recited in SEQ ID NO: 3 or 4.

17. The composition according to any one of claims 1 to 16, characterized in that, The nucleic acid construct is administered once.

18. The composition according to any one of claims 1 to 16, characterized in that, The nucleic acid construct is administered more than twice.

19. The composition according to claim 18, 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.

20. The composition according to any one of claims 1 to 19, characterized in that, Administer the nucleic acid construct at a dosage of 1.0×10 3 TCID 50 or greater.

21. The composition according to any one of claims 1 to 20, characterized in that, at a dosage of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 administer the nucleic acid construct.

22. The composition according to claim 21, wherein at a dose of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 administer the nucleic acid construct once a week.

23. The composition according to claim 21, wherein Administer the nucleic acid construct once at a dosage of 1.0×10 3 TCID 50 or more and less than 5.0×10 4 TCID 50 When the virus is detected in vivo, further administer the nucleic acid construct.

24. The composition according to any one of claims 1 to 20, characterized in that, Administer the nucleic acid construct at a dosage of 5.0×10 4 TCID 50 or more and 5.0×10 6 TCID 50 or less.

25. The composition according to claim 24, characterized in that, Administer the nucleic acid construct once at a dosage of 5.0×10 4 TCID 50 or greater.

26. The composition according to any one of claims 1 to 25, characterized in that, The subject has been administered an anti - HIV drug.

27. The composition according to any one of claims 1 to 25, wherein the subject has not been administered an anti-HIV drug or the anti-HIV drug has been discontinued at the start of administration of the composition.

28. The composition according to claim 26 or 27, characterized in that, Regarding the anti-HIV drug, an anti-HIV drug selected from the group consisting of reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and CCR5 inhibitors is administered.

29. The composition according to claim 26 or 27, characterized in that, Regarding the anti-HIV drug, an anti-HIV drug selected from the group consisting of tenofovir, emtricitabine, dolutegravir, zidovudine, lamivudine, stavudine, didanosine, abacavir, nevirapine, efavirenz, etravirine, rilpivirine, saquinavir, indinavir, nelfinavir, lopinavir / ritonavir combination, atazanavir, fosamprenavir, darunavir, ritonavir, raltegravir, elvitegravir, maraviroc, or any combination of these substances is administered.

30. The composition according to any one of claims 1 to 29, wherein the composition completely clears the AIDS virus in the subject.

31. The composition according to any one of claims 1 to 30, wherein the composition is not administered before viral infection.

32. The composition according to any one of claims 1 to 31, wherein the subject has previously been vaccinated with the BCG vaccine.

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