Department of Microbiology and Immunology, University of Rochester School of Medicine and Dentistry, 601 Elmwood Avenue, Box 672, Rochester, NY, 14642, USA.
Cytotechnology. 2006 Jul;51(3):133-40. doi: 10.1007/s10616-006-9023-6. Epub 2006 Nov 18.
Recombinant adenoviruses (Ad) are being explored as promising delivery systems for gene therapy and vaccination. However, there is a concern about the possibility of generating replication-competent adenoviruses (RCA) using the human embryonic kidney 293 cell line. We have constructed a new cell line named the UR cell line which can be used to produce Ad vectors free of RCA. This cell line is based on the human embryonic lung HEL 299 cell. We first constructed a shuttle plasmid which encodes the E1A/E1B sequence that is necessary for adenovirus replication. The shuttle plasmid was then transfected into HEL 299 cells. The presence of the E1A/E1B sequence and protein expression in the stably transformed UR cells was confirmed. Viruses produced in UR cells were still RCA-free after ten test passages, while adenovirus produced in 293 cells had generated RCA during the fourth passage. We conclude that the UR cell line is sufficiently stable, can effectively produce a virus yield comparable with 293 cells, and does not generate RCA formation during Ad propagation.
重组腺病毒（Ad）被探索作为基因治疗和疫苗接种的有前途的传递系统。然而，人们担心使用人胚肾 293 细胞系可能会产生复制能力的腺病毒（RCA）。我们构建了一种新的细胞系，命名为 UR 细胞系，可用于生产无 RCA 的 Ad 载体。该细胞系基于人胚肺 HEL 299 细胞。我们首先构建了一个穿梭质粒，其编码腺病毒复制所必需的 E1A/E1B 序列。然后将穿梭质粒转染到 HEL 299 细胞中。在稳定转化的 UR 细胞中证实了 E1A/E1B 序列的存在和蛋白表达。在 UR 细胞中产生的病毒在经过十次测试传代后仍然没有 RCA，而在 293 细胞中产生的腺病毒在第四代时已经产生了 RCA。我们得出结论，UR 细胞系足够稳定，能够有效地产生与 293 细胞相当的病毒产量，并且在 Ad 繁殖过程中不会产生 RCA 形成。
