A novel hyper-immunogenic low virulent BCG vaccine against tuberculosis

摘要 由细菌病原体分枝杆菌引起的结核病(TB)的人类发病率和死亡率 结核病(Mtb)仍然是全世界的重大健康问题。卡米特盖林杆菌 卡介苗(BCG)仍然是唯一被批准的结核病疫苗。尽管卡介苗被认为是安全的和部分 对肺外儿童结核病有效，其预防儿童和成人肺结核的能力 仍然值得怀疑。此外，还有一种担忧是，卡介苗不会在 接种疫苗的个体。然而，我们对结核分枝杆菌和卡介苗衍生的重组疫苗的研究结果强烈 提示卡介苗可以改进，以更有效地对抗结核病。 这项建议的目标是通过合理删除基因来改善卡介苗。我们计划按顺序删除 卡介苗的三个基因，即sapm，zmp1和nuoG，通过同源重组产生三个 卡介苗(BCG-TKO)基因敲除株。这些基因编码的蛋白质使卡介苗能够逃避宿主免疫。 防止吞噬小体-溶酶体融合、自噬、细胞凋亡和其他相关过程的反应 抗原提呈细胞(APC)，如树突状细胞和巨噬细胞。我们假设，删除 这些基因将允许BCG-TKO菌株被APC有效地处理，这将导致 将抗原递呈给免疫细胞，增强体内的免疫原性和疗效。此外，我们 预计这些基因的缺失将降低卡介苗的毒力，从而使卡介苗-TKO成为 艾滋病病毒是安全的。我们计划实现三个目标：1)通过同源基因构建BCG-TKO菌株 重组，2)在SCID小鼠模型中分析BCG-TKO株的免疫原性和安全性，以及 3)观察卡介苗TKO在感染和人源化感染HIV小鼠模型中的疗效。 感染。总体而言，我们预计这项提议将产生一种高效的第三代卡介苗疫苗。 即使是在感染艾滋病毒的婴儿身上，也适用于治疗结核病。

SUMMARY Human morbidity and mortality due to tuberculosis (TB), caused by the bacterial pathogen Mycobacterium tuberculosis (Mtb), continue to be of significant health concern throughout the world. Bacille Calmette-Guerin (BCG) still remains the only approved vaccine against TB. Although BCG is considered safe and partially effective against extra-pulmonary childhood TB, its ability to protect against childhood and adult pulmonary TB is still questionable. In addition, there is a concern that BCG does not induce long lasting immune responses in the immunized individuals. However, our findings with Mtb- and BCG- derived recombinant vaccines strongly suggest that BCG can be improved to be more efficacious against TB. The goal of this proposal is to improve BCG by rationally deleting genes. We plan to sequentially delete three genes of BCG namely sapM, zmp1 and nuoG, through homologous recombination, to result in a triple knockout BCG (BCG-TKO) strain. These genes encode proteins that enable BCG to evade host immune response by preventing phagosome-lysosomal fusion, autophagy, apoptosis and other related processes in the antigen presenting cells (APCs) such as dendritic cells and macrophages. We hypothesize that deletion of these genes will allow the BCG-TKO strain to be efficiently processed by APCs, which will lead to increased antigen presentation to the immune cells and enhanced in vivo immunogenicity and efficacy. Additionally, we anticipate that deletion of these genes will reduce the virulence of the BCG, thus making the BCG-TKO to be HIV safe. We plan to accomplish our goal with three aims: 1) construct a BCG-TKO strain through homologous recombination, 2) analyze the immunogenicity and safety of the BCG-TKO strain in a SCID mouse model, and 3) investigate the efficacy of the BCG-TKO in the regular and humanized mouse models with and without HIV infection. Overall, we expect that this proposal will produce a highly efficacious, third generation BCG vaccine against TB that will be suitable for administration even in HIV infected infants.