Kim S J, Lee C, Lee S Y, Kim I, Park J S, Sasagawa T, Ko J J, Park S E, Oh Y-K
Comprehensive Gynecologic Cancer Center, Pundang CHA General Hospital, Sungnam, Kyonggi-do, South Korea.
Gene Ther. 2003 Aug;10(15):1268-73. doi: 10.1038/sj.gt.3301997.
To increase the potency of human papillomavirus (HPV) DNA vaccines, we constructed a series of HPV16 L1 vaccines genetically fused with a secretion signal and/or immune cell-recruiting RANTES. The DNA vaccines encoding secretory HPV L1 were constructed by inserting HPV L1 gene into a vector with an ER-targeting secretory signal sequence. The expression plasmid encoding secretory HPV L1 (pER/L1) was fused with cDNA of RANTES, generating pER/L1/R. For comparison, HPV L1 genes were cloned into pVAX1 vector with no signal sequence (pL1), and further linked to the N-terminus (pL1/R) or C-terminus of RANTES (pR/L1). The secretion of L1 proteins was observed in the pER/L1, pER/L1/R, and pR/L1-transfected cells, except the pL1/R-transfected group. Cytoplasmic localization of L1 protein was observed in the cells transfected with pL1/R, but not with pER/L1/R at 48 h after transfection. In mice, RANTES-fused vaccines more effectively elicited the levels of HPV16 L1-specific IgG and IgG2a antibodies than pL1. Of RANTES-fused vaccines, pER/L1/R encoding the secreted fusion protein induced the highest humoral and CD8(+) T-cell-stimulating responses. These results suggest that the immunogenicity of HPV L1 DNA vaccines could be enhanced by genetic fusion to a chemokine and secretory signal peptide sequences.
为提高人乳头瘤病毒（HPV）DNA疫苗的效力，我们构建了一系列与人乳头瘤病毒16型L1疫苗基因融合的分泌信号和/或免疫细胞募集RANTES的疫苗。通过将HPV L1基因插入带有内质网靶向分泌信号序列的载体中构建编码分泌型HPV L1的DNA疫苗。将编码分泌型HPV L1的表达质粒（pER/L1）与RANTES的cDNA融合，生成pER/L1/R。为作比较，将HPV L1基因克隆到无信号序列的pVAX1载体（pL1）中，并进一步连接到RANTES的N端（pL1/R）或C端（pR/L1）。除pL1/R转染组外，在pER/L1、pER/L1/R和pR/L1转染的细胞中均观察到L1蛋白的分泌。转染48小时后，在pL1/R转染的细胞中观察到L1蛋白的细胞质定位，但在pER/L1/R转染的细胞中未观察到。在小鼠中，与pL1相比，融合RANTES的疫苗更有效地引发了HPV16 L1特异性IgG和IgG2a抗体水平。在融合RANTES的疫苗中，编码分泌型融合蛋白的pER/L1/R诱导了最高的体液和CD8(+) T细胞刺激反应。这些结果表明，通过与趋化因子和分泌信号肽序列进行基因融合，可以增强HPV L1 DNA疫苗的免疫原性。
