Influenza Virus NS Vectors Expressing theMycobacterium tuberculosisESAT-6 Protein Induce CD4⁺Th1 Immune Response and Protect Animals against Tuberculosis Challenge

Abstract

Infection with Mycobacterium tuberculosis remains a major cause of morbidity and mortality all over the world. Since the effectiveness of the only available tuberculosis vaccine, Mycobacterium bovis bacillus CalmetteGuérin (BCG), is suboptimal, there is a strong demand to develop new tuberculosis vaccines. As tuberculosis is an airborne disease, the intranasal route of vaccination might be preferable. Live influenza virus vaccines might be considered as potential vectors for mucosal immunization against various viral or bacterial pathogens, including M. tuberculosis. We generated several subtypes of attenuated recombinant influenza A viruses expressing the 6-kDa early secretory antigenic target protein (ESAT-6) of M. tuberculosis from the NS1 reading frame. We were able to demonstrate the potency of influenza virus NS vectors to induce an M. tuberculosisspecific Th1 immune response in mice. Moreover, intranasal immunization of mice and guinea pigs with such vectors induced protection against mycobacterial challenge, similar to that induced by BCG vaccination.Since reverse genetics methods have been developed for negative-strand RNA viruses (30, 33), influenza viruses can be considered as viral vectors for immunization against different pathogens. The influenza A virus contains a segmented genome consisting of eight negative-strand RNA fragments. Among them, the smallest fragment (NS), encoding two proteins (NS1 and Nep), is a suitable target for genetic manipulation. NS1 is the only nonstructural protein of the influenza virus and has been shown to tolerate relatively long insertions of more than 250 amino acids (23). Moreover, as the NS1 protein is produced in large quantities in infected cells, expression of foreign sequences from the NS1 reading frame should result in a strong immune response against the inserted antigen. It was demonstrated that immunization of mice with NS influenza virus vectors could trigger a CD8 ϩ T-cell response, especially when two vectors belonging to different influenza virus subtypes were used for prime-boost immunizations (12, 39). However, influenza virus NS vectors might be less efficient for the induction of insert-specific T-helper and antibody responses due to the intracellular localization of the NS1 protein leading to a less efficient presentation of linked antigen through the major histocompatibility complex class II pathway.In this work, we tested the potential of recombinant influenza virus NS vectors expressing a Mycobacterium tuberculosis antigen to induce T-helper response after intranasal immunization of mice. It is accepted that control over tuberculosis infection depends on the recruitment of antigen-specific T cells, mainly CD4 ϩ cells, to the lungs and the release of cytokines, particularly gamma interferon (IFN-␥), to trigger the bacterium-killing mechanisms in activated macrophages (14). We constructed a recombinant influenza virus NS gene for the expression of the 6-kDa early secretory antigenic target protein (ESAT-6) derived from M. tuberculosis and introduc...

Evaluation of Recombinant Influenza Virus-Simian Immunodeficiency Virus Vaccines in Macaques

“…Although we show proof of principle that at least three SIV-specific CD8 T-cell responses (one Gag response and two Tat responses, all presented by Mane-A*10) can be boosted simultaneously by this recombinant influenza virus strategy, it could be cumbersome to construct sufficient numbers of influenza virus vectors to broadly cover relevant HIV/SIV T-cell epitopes in outbred populations. Expression of whole heterologous proteins from influenza virus proteins is technically possible and has been achieved with expression of the enhanced green fluorescence protein, interleukin-2, and tuberculosis proteins (21,37,56); however, the stability of such vectors will require careful study. We are currently constructing influenza virus vectors expressing whole SIV proteins.…”

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“…These vaccine candidates are capable of inducing the enhanced Th1 systemic and the local mucosal immune response at the infection entry. The current research data prove that the vector technology based on the modification of the NS segment of influenza virus enables the generation of highly immunogenic recombinant virus strains that express the foreign sequences from the NS1 reading frame [90][91][92][93][94][95][96]. Clinical trials showed that viruses with the modified NS gene are safe and highly immunogenic [97].…”

Проблема Создания Универсальной Противогриппозной Вакцины

“…Такие вакцины способны вызывать усиленный Тh1 системный и локальный мукозальный иммунный ответ во входных воротах инфекции. Полученные к настоящему времени ре-зультаты свидетельствуют, что векторная технология на основе модификаций сегмента NS вируса гриппа позволяет получать высокоиммуногенные рекомби-нантные штаммы, экспрессирующие посторонние антигенные последовательности с рамки считывания белка NS1 [90][91][92][93][94][95][96]. Вирусы с модифицированным ге-ном NS прошли клинические испытания, в которых показана их безопасность и высокая иммуногенность [97].…”