Use of helper-dependent adenoviral vectors of alternative serotypes permits repeat vector administration
Use of helper-dependent adenoviral vectors of alternative serotypes permits repeat vector administration
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DOI：
10.1038/sj.gt.3300995
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发表时间：
1999-09-01
期刊：
影响因子：
5.1
通讯作者：
Graham, FL
中科院分区：
文献类型：
--
作者：
Parks, RJ;Evelegh, CM;Graham, FL
我们已经开发了一种基于血清型2的新的辅助腺病毒（Ad），Ad 2LCBcCARP，用于Cre/loxP系统（Parks等，Proc Natl Acad Sci USA，1996; 93：13565-13570），以产生缺失所有蛋白质编码序列的Ad载体（辅助依赖性Ad载体（hdAd））。比较Ad 2LC 8 cCARP和我们的原始辅助病毒（基于血清5型，Ad 5LC 8 cluc），这两种辅助病毒以相似的效率扩增hdAd，并且在大规模制备载体后导致相似的产量和纯化。辅助病毒依赖性系统的一个重要特征是，除病毒体DNA外，所有病毒体组分均来自辅助病毒。因此，在Ad 2LC 8 cCARP的帮助下产生的载体不被针对Ad 5的抗体中和，并且用Ad 5辅助物产生的载体对针对Ad 2的中和抗体具有抗性。静脉内注射基于Ad 2的hdAd后小鼠肝脏中的转基因表达的分析表明，该载体可以有效地使肝脏增殖，并产生高水平的外源转基因，类似于用Ad 5辅助病毒产生的hdAd所表达的那些。用hdAd 2免疫小鼠产生Ad 2中和抗体，其不与hdAdS交叉反应。为了确定是否可以通过顺序使用替代的Ad血清型来实现成功的重复Ad载体施用，我们用hdAd 2（hSEAP）注射小鼠，3个月后用相同或不同血清型的表达lacZ的hdAd注射。重复施用hdAd 2导致转基因表达与幼稚动物相比减少30至100倍。相反，当第二载体是不同的血清型时，没有观察到转基因表达的降低。这些结果表明，有效的载体再施用可以通过基于替代血清型顺序使用hdAd来实现。
We have developed a new helper adenovirus (Ad) based on serotype 2, Ad2LCBcCARP, for use in the Cre/loxP system (Parks et al. Proc Natl Acad Sci USA, 1996; 93: 13565-13570) to generate Ad vectors deleted of all protein coding sequences (helper-dependent Ad vectors (hdAd)). A comparison of Ad2LC8cCARP and our original helper Virus (based on serotype 5, Ad5LC8cluc) showed that the two helper viruses amplified hdAd with a similar efficiency, and resulted in a similar yield and purify after large-scale preparation of vector In vitro, the resulting hdAd2 had a similar transduction efficiency and expression kinetics of transgene (beta-gal) as the hdAdS. An important feature of the helper-dependent system is that all virion components, except the virion DNA, derive from the helper virus. Consequently, vectors produced with help from Ad2LC8cCARP were not neutralized by antibodies against Ad5, and vectors produced with Ad5 helper were resistant to neutralizing antibodies against Ad2. Analysis of transgene expression in mouse liver after intravenous injection of the Ad2-based hdAd showed that the vector could efficiently transduce the liver, and produce high levels of a foreign transgene, similar to those expressed by the hdAd generated with the Ad5 helper virus. Mice immunized with hdAd2 produced Ad2-neutralizing antibodies, which did not crossreact with hdAdS. To determine if successful repeat Ad vector administration could be achieved by sequential use of alternative Ad serotypes, we injected mice with hdAd2 (hSEAP) followed 3 months later by a lacZ-expressing hdAd of either the same or different serotype. Repeated administration of hdAd2 resulted in a 30- to 100-fold reduction in transgene expression compared with naive animals. In contrast no decrease in transgene expression was observed when the second vector was of a different serotype. These results demonstrate that effective vector readministration can be achieved by the sequential use of hdAds based on alternative serotypes.