Bahrami Hamid, Mosavat Arman, Soleimanpour Saman, Farsiani Hadi, Valizadeh Narges, Rezaee Seyed Abdolrahim, Amini Abbas Ali
Cancer and Immunology Research Center, Research Institute for Health Development, Kurdistan University of Medical Sciences, Sanandaj, Iran.
Blood Borne Infections Research Center, Academic Center for Education, Culture, and Research (ACECR), Razavi Khorasan, Mashhad, Iran.
Microb Pathog. 2025 Oct;207:107894. doi: 10.1016/j.micpath.2025.107894. Epub 2025 Jul 10.
Tuberculosis (TB) is still a life-threatening infection. Mycobacterium tuberculosis (Mtb) Ag85 is the most potent immunogenic factor for introducing protective vaccines against TB. To enhance its immunogenicity, Mtb Ag85B was fused to the Fc fragment of human IgG1 to produce Ag85B:hFcγ1 and its immunogenicity was assessed in a mouse model. The Ag85B:hFcγ1 was designed and made in the Pichia pastoris expression system. Then, the production and purity of Ag85B:hFcγ1 were confirmed using ELISA and Western blotting. Co-localisation assay showed that Ag85B:hFcγ1 can be localised with hFcγRI (CD64), facilitating a proper Th1 response. Immunisation assays in a mouse model showed a high IFN-γ production as the hallmark of cell-mediated immunity (CMI) in the sera of the treated animals compared with the control ones (p = 0.02). However, the concentration of IL-17 did not reach the sensitivity of the assays. Functional co-localisation revealed that the fused hFcγ1 with Ag85 can bind to CD64 and induce cross-presentation toward Th1 responses by producing higher IFN-γ. In conclusion, Ag85B:hFcγ1 with high glycosylation seems more immunogenic than our previous studies Mtb multistage and Fc fusion proteins-multi molecules. APC targeting was used to favour cross-presentation using the Fc tag.
结核病（TB）仍然是一种危及生命的感染。结核分枝杆菌（Mtb）Ag85是用于研发抗结核保护性疫苗的最有效的免疫原性因子。为增强其免疫原性，将Mtb Ag85B与人IgG1的Fc片段融合以产生Ag85B:hFcγ1，并在小鼠模型中评估其免疫原性。Ag85B:hFcγ1在毕赤酵母表达系统中设计并制备。然后，使用酶联免疫吸附测定（ELISA）和蛋白质免疫印迹法确认Ag85B:hFcγ1的产量和纯度。共定位分析表明，Ag85B:hFcγ1可与hFcγRI（CD64）共定位，促进适当的Th1反应。小鼠模型中的免疫测定显示，与对照动物相比，处理动物血清中作为细胞介导免疫（CMI）标志的干扰素-γ（IFN-γ）产生量很高（p = 0.02）。然而，白细胞介素-17（IL-17）的浓度未达到测定的灵敏度。功能共定位表明，与Ag85融合的hFcγ1可与CD64结合，并通过产生更高水平的IFN-γ诱导向Th1反应的交叉呈递。总之，具有高糖基化的Ag85B:hFcγ1似乎比我们之前研究的Mtb多阶段和Fc融合蛋白 - 多分子更具免疫原性。使用Fc标签通过抗原呈递细胞（APC）靶向来促进交叉呈递。
