Immunization And Safety Of Recombinant Respiratory Syncytial Virus Fusion Protein TB10.4-F1

Respiratory syncytial virus (RSV) is the most major pathogens of lower respiratory tract infection in infants worldwide, which are likely to cause bronchiolitis, pneumonia, or death. There is no useful vaccine to prevent RSV infection. In this study, the F1 protein of RSV, fused with the TB10.4 proteins of Mycobacterium tuberculosis was expressed in E. coli, and immunised BALB/c mice combined with mucosal adjuvants non-toxic E. coli heat-labile enterotoxin (LT). Finally, we evaluated the immunogenicity and safty of the fusion protein.We obtained the recombinant fusion protein TB10.4-F1 and protein TB10.4, which contain His-tag in E. coli, by constructed the recombinant vector of TB10.4-F1/pET30a and TB10.4/pET28a, and proved by western-blot with anti-His tag antibody. The recombinant inclusion bodies protein was purified with nickel affinity chromatography under denaturing conditions. The purity of TB10.4-F1 and TB10.4 was 80% and 95%, respectively.The purified TB10.4-F1 and TB10.4 were immunised BALB/c mice combined with LT. Twenty five mice were divided into 5 groups, each group were immunized at week 0,2,4 as the following:PBS group; LT group; TB10.4 group; TB10.4-F1 group; TB10.4-F1+LT group. The results showed that:①the specific IgG of TB10.4-F1+LT group and TB10.4-F1 group were higher than other groups obviously, which suggested that fusion protein could induce a high level of serum antibody;②compared with the TB10.4 group, the IgG1/IgG2a ratio of TB10.4-F1+LT group and TB10.4-F1 group was closer to 1, indicating that T cell immune response was more balanced;③TB10.4-F1 fusion protein had immune toxicity in lung biopsy, which can cause lung telangiectasia, eosinophilia;④the IgG1/IgG2a ratio of TB10.4 group was greater than 1, which represented that the Th2-type response is dominant.Mice were challenged with 3×105 pfu of RSV by intranasally after immunization.5 days later, mice were killed for detection. The results suggeted that:①mice weight of TB10.4-F1+LT group and TB10.4-F1 group decreased quickly, three mice in TB10.4-F1+LT group were dead due to respiratory failure;②the douche of the respiratory tract of TB10.4-F1+LT group and TB10.4-F1 group had a good IgA response, and TB10.4-F1+LT group was higher than TB10.4-F1 group (P<0.05), indicating that LT can promote production of secretory IgA on the respiratory mucosa;③lung biopsy showed that TB10.4-F1+LT group and TB10.4-F1 group were induced serious Immune toxicological reaction after challenge, indicating that the recombinant protein TB10.4-F1 not only failed in protection, but increased the disease in mice.This study investigated the immunogenicity and safety of fusion protein TB10.4-F1,which will provide a theoretical basis for developing new RSV vaccines.