Suter M, Lew A M, Grob P, Adema G J, Ackermann M, Shortman K, Fraefel C
Institute of Virology, University of Zurich, Winterthurerstrasse 266a, CH-8057 Zurich, Switzerland.
Proc Natl Acad Sci U S A. 1999 Oct 26;96(22):12697-702. doi: 10.1073/pnas.96.22.12697.
This study aimed to exploit bacterial artificial chromosomes (BAC) as large antigen-capacity DNA vaccines (BAC-VAC) against complex pathogens, such as herpes simplex virus 1 (HSV-1). The 152-kbp HSV-1 genome recently has been cloned as an F-plasmid-based BAC in Escherichia coli (fHSV), which can efficiently produce infectious virus progeny upon transfection into mammalian cells. A safe modification of fHSV, fHSVDeltapac, does not give rise to progeny virus because the signals necessary to package DNA into virions have been excluded. However, in mammalian cells fHSVDeltapac DNA can still replicate, express the HSV-1 genes, cause cytotoxic effects, and produce virus-like particles. Because these functions mimic the lytic cycle of the HSV-1 infection, fHSVDeltapac was expected to stimulate the immune system as efficiently as a modified live virus vaccine. To test this hypothesis, mice were immunized with fHSVDeltapac DNA applied intradermally by gold-particle bombardment, and the immune responses were compared with those induced by infection with disabled infectious single cycle HSV-1. Immunization with either fHSVDeltapac or disabled infectious single cycle HSV-1 induced the priming of HSV-1-specific cytotoxic T cells and the production of virus-specific antibodies and conferred protection against intracerebral injection of wild-type HSV-1 at a dose of 200 LD(50). Protection probably was cell-mediated, as transfer of serum from immunized mice did not protect naive animals. We conclude that BAC-VACs per se, or in combination with genetic elements that support replicative amplification of the DNA in the cell nucleus, represent a useful new generation of DNA-based vaccination strategies for many viral and nonviral antigens.
本研究旨在利用细菌人工染色体（BAC）作为大容量抗原的DNA疫苗（BAC-VAC）来对抗复杂病原体，如单纯疱疹病毒1型（HSV-1）。最近，152千碱基对的HSV-1基因组已被克隆为基于F质粒的大肠杆菌BAC（fHSV），将其转染到哺乳动物细胞后可高效产生有感染性的病毒后代。fHSV的一种安全修饰形式fHSVDeltapac不会产生子代病毒，因为将DNA包装到病毒粒子中的必要信号已被去除。然而，在哺乳动物细胞中，fHSVDeltapac DNA仍可复制、表达HSV-1基因、引起细胞毒性作用并产生病毒样颗粒。由于这些功能模拟了HSV-1感染的裂解周期，因此预计fHSVDeltapac能像改良活病毒疫苗一样有效地刺激免疫系统。为了验证这一假设，通过金颗粒轰击将fHSVDeltapac DNA皮内注射免疫小鼠，并将免疫反应与感染失活的单周期感染性HSV-1诱导的免疫反应进行比较。用fHSVDeltapac或失活的单周期感染性HSV-1免疫均可引发HSV-1特异性细胞毒性T细胞的启动以及病毒特异性抗体的产生，并能保护小鼠免受200个半数致死量（LD50）的野生型HSV-1脑内注射。这种保护可能是细胞介导的，因为来自免疫小鼠的血清转移并不能保护未免疫的动物。我们得出结论，BAC-VAC本身或与支持细胞核中DNA复制扩增的遗传元件结合，代表了一种针对许多病毒和非病毒抗原的有用的新一代基于DNA的疫苗接种策略。
