Kim Seung Jo, Suh Dongchul, Park Sang Eun, Park Jeong-Sook, Byun Hyang-Min, Lee Chan, Lee Sun Young, Kim Inho, Oh Yu-Kyoung
Pundang CHA General Hospital, Sungnam, Kyonggi-do, South Korea.
Virology. 2003 Sep 15;314(1):84-91. doi: 10.1016/s0042-6822(03)00417-3.
To increase the potency of DNA vaccines, we constructed genetic fusion vaccines encoding antigen, secretion signal, and/or chemokine RANTES. The DNA vaccines encoding secreted hepatitis B surface antigen (HBsAg) were constructed by inserting HBsAg gene into an expression vector with an endoplasmic reticulum (ER)-targeting secretory signal sequence. The plasmid encoding secretory HBsAg (pER/HBs) was fused to cDNA of RANTES, generating pER/HBs/R. For comparison, HBsAg genes were cloned into pVAX1 vector with no signal sequence (pHBs), and further linked to the N-terminus of RANTES (pHBs/R). Immunofluorescence study showed the cytoplasmic localization of HBsAg protein expressed from pHBs and pHBs/R, but not from pER/HBs and pER/HBs/R at 48 h after transfection. In mice, RANTES-fused DNA vaccines more effectively elicited the levels of HBsAg-specific IgG antibodies than pHBs. All the DNA vaccines induced higher levels of IgG(2a) rather than IgG(1) antibodies. Of RANTES-fused vaccines, pER/HBs/R encoding the secreted fusion protein revealed much higher humoral and CD8(+) T cell-stimulating responses compared to pHBs/R. These results suggest that the immunogenicity of DNA vaccines could be enhanced by genetic fusion to a secretory signal peptide sequence and RANTES.
为提高DNA疫苗的效力，我们构建了编码抗原、分泌信号和/或趋化因子RANTES的基因融合疫苗。通过将乙型肝炎表面抗原（HBsAg）基因插入带有内质网（ER）靶向分泌信号序列的表达载体中，构建了编码分泌型HBsAg的DNA疫苗。将编码分泌型HBsAg的质粒（pER/HBs）与RANTES的cDNA融合，产生pER/HBs/R。作为对照，将HBsAg基因克隆到无信号序列的pVAX1载体中（pHBs），并进一步连接到RANTES的N端（pHBs/R）。免疫荧光研究显示，转染后48小时，pHBs和pHBs/R表达的HBsAg蛋白定位于细胞质中，而pER/HBs和pER/HBs/R表达的HBsAg蛋白则不是。在小鼠中，与pHBs相比，融合RANTES的DNA疫苗能更有效地诱导HBsAg特异性IgG抗体水平。所有DNA疫苗诱导的IgG(2a)水平均高于IgG(1)抗体。在融合RANTES的疫苗中，编码分泌型融合蛋白的pER/HBs/R与pHBs/R相比，显示出更高的体液免疫和CD8(+) T细胞刺激反应。这些结果表明，通过与分泌信号肽序列和RANTES进行基因融合，可以增强DNA疫苗的免疫原性。
