In vivo model of adeno-associated virus vector persistence and rescue
In vivo model of adeno-associated virus vector persistence and rescue
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DOI：
10.1128/jvi.70.5.3235-3241.1996
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发表时间：
1996-05-01
影响因子：
5.4
通讯作者：
Flotte, TR
中科院分区：
文献类型：
--
作者：
Afione, SA;Conrad, CK;Flotte, TR
基于人类DNA病毒的基因治疗载体可以从随后感染相应野生型（wt）辅助病毒的个体中动员或抢救出来。这种现象已经在体外用腺病毒（Ad）和腺相关病毒（AAV）载体进行了有效的模拟，但此前尚未在体内进行过研究。在目前的研究中，我们已经建立了一个体内模型来研究重组AAV载体（AAV- cftr）与wt AAV2型（AAV2）和宿主范围突变Ad （Ad2HR405）的相互作用（D. E. Brough， S. a . Rice， S. Sell, and D. F. Klessig， J. Virol. 55:206-212, 1985）， AAV- cftr被给予恒河猴鼻子或肺的呼吸上皮细胞。在给药后3个月的时间点从输注部位收集原代细胞，以确认载体DNA的持久性。载体DNA以外体形式存在，仅通过添加wt AAV2和Ad才能在体外获救。在体内救援研究中，在感染wt-AAV2和Ad2HR405之前或之后给予载体，检测AAV-CFTR的脱落情况。同时给药后，在鼻部发现Ad2HR405和wt-AAV2感染。当将高滴度的病毒直接注射到下呼吸道时，wt-AAV2在肺部发生复制。后一种情况下也观察到AAV-CFTR载体的挽救。尽管这些研究是在每组中使用少量动物进行的，但似乎AAV- cftr DNA在灵长类动物呼吸道中持续存在，并且该模型可能对重组AAV载体救援的研究有用。
Gene therapy vectors based on human DNA viruses could be mobilized or rescued from individuals who are subsequently infected with the corresponding wild-type (wt) helper viruses. This phenomenon has been effectively modeled in vitro with both adenovirus (Ad) and adeno-associated virus (AAV) vectors but has not previously been studied in vivo. In the current study, we have developed an in vivo model to study the interactions of a recombinant AAV vector (AAV-CFTR) with wt AAV type 2 (AAV2) and a host range mutant Ad (Ad2HR405) for which monkey cells are permissive (D. E. Brough, S. A. Rice, S. Sell, and D. F. Klessig, J. Virol. 55:206-212, 1985), AAV-CFTR was administered to the respiratory epithelium of the nose or lung of rhesus macaques. Primary cells were harvested from the infusion site at time points up to 3 months after vector administration to confirm vector DNA persistence. Vector DNA was present in episomal form and could be rescued in vitro only by addition of wt AAV2 and Ad. In in vivo rescue studies, vector was administered before or after wt-AAV2 and Ad2HR405 infection, and the shedding of AAV-CFTR was examined. Ad2HR405 and wt-AAV2 infections were established in the nose with concomitant administration. wt-AAV2 replication occurred in the lung when virus was administered directly at a high titer to the lower respiratory tract. AAV-CFTR vector rescue was also observed in the latter setting. Although these studies were performed with small numbers of animals within each group, it appears that AAV-CFTR DNA persists in the primate respiratory tract and that this model may be useful for studies of recombinant AAV vector rescue.