Hohmann E L, Oletta C A, Loomis W P, Miller S I
Infectious Disease Unit, Massachusetts General Hospital, Boston 02114, USA.
Proc Natl Acad Sci U S A. 1995 Mar 28;92(7):2904-8. doi: 10.1073/pnas.92.7.2904.
Attenuated Salmonella are useful oral vaccine vectors capable of carrying multiple heterologous antigen genes, but optimal expression of foreign antigens has not yet been achieved. We hypothesized that Salmonella phoP-activated genes, which are transcriptionally activated within antigen-processing macrophages, could prove useful for delivery of heterologous antigens to the immune system. We have created a suicide vector that allows the stable chromosomal insertion of heterologous antigen genes within the phoP-activated gene C (pagC) of Salmonella and permits the expression of heterologous antigens as fusion proteins between the first 84 amino acids of PagC and the chosen antigen. The Escherichia coli phoA gene encoding alkaline phosphatase was cloned into this vector; the resultant plasmid was used to construct Salmonella typhimurium strains that express PagC-alkaline phosphatase fusion proteins from a single chromosomal gene copy. Such strains were administered orally and i.p. as vaccines to BALB/c mice and compared with control strains expressing alkaline phosphatase constitutively. After 3 weeks, mouse sera were analyzed for IgG responses to S. typhimurium lipopolysaccharide and alkaline phosphatase. Remarkably, though all mice had comparable antibody responses to lipopolysaccharide, only mice immunized with strains bearing phoP-activated fusion genes had antibody responses to the heterologous antigen. We conclude that expression of a heterologous antigen from an S. typhimurium in vivo-induced promoter that is activated within macrophages markedly enhances the immunogenicity of a model antigen expressed from a single chromosomal gene copy.
减毒沙门氏菌是一种有用的口服疫苗载体，能够携带多个异源抗原基因，但尚未实现外源抗原的最佳表达。我们推测，沙门氏菌phoP激活基因在抗原加工巨噬细胞内被转录激活，可能有助于将异源抗原递送至免疫系统。我们构建了一种自杀载体，可使异源抗原基因稳定地插入沙门氏菌的phoP激活基因C（pagC）内，并允许异源抗原作为PagC前84个氨基酸与所选抗原之间的融合蛋白表达。将编码碱性磷酸酶的大肠杆菌phoA基因克隆到该载体中；所得质粒用于构建鼠伤寒沙门氏菌菌株，这些菌株从单个染色体基因拷贝表达PagC-碱性磷酸酶融合蛋白。将这些菌株作为疫苗经口和腹腔注射给BALB/c小鼠，并与组成型表达碱性磷酸酶的对照菌株进行比较。3周后，分析小鼠血清中针对鼠伤寒沙门氏菌脂多糖和碱性磷酸酶的IgG反应。值得注意的是，尽管所有小鼠对脂多糖的抗体反应相当，但只有用携带phoP激活融合基因的菌株免疫的小鼠对异源抗原有抗体反应。我们得出结论，在巨噬细胞内被激活的鼠伤寒沙门氏菌体内诱导型启动子表达异源抗原，可显著增强从单个染色体基因拷贝表达的模型抗原的免疫原性。
