Miura Y, Yoshida K, Nishimoto T, Hatanaka K, Ohnami S, Asaka M, Douglas J T, Curiel D T, Yoshida T, Aoki K
Genetics Division, National Cancer Center Research Institute, Tokyo, Japan.
Gene Ther. 2007 Oct;14(20):1448-60. doi: 10.1038/sj.gt.3303007. Epub 2007 Aug 16.
Targeting of gene transfer at the level of cell entry is one of the most attractive challenges in vector development. However, attempts to redirect adenovirus vectors to alternative receptors by engineering the capsid-coding region have shown limited success because proper targeting ligand-receptor systems on the cells of interest are generally unknown. Systematic approaches to generate adenovirus vectors targeting any given cell type need to be developed to achieve this goal. Here, we constructed an adenovirus library that was generated by a Cre-lox-mediated in vitro recombination between an adenoviral fiber-modified plasmid library and genomic DNA to display random peptides on a fiber knob. As proof of concept, we screened the adenovirus display library on a glioma cell line and observed selection of several particular peptide sequences. The targeted vector carrying the most frequently isolated peptide significantly enhanced gene transduction in the glioma cell line but not in many other cell lines. Because the insertion of a pre-selected peptide into a fiber knob often fails to generate an adenovirus vector, the selection of targeting peptides is highly useful in the context of the adenoviral capsid. This vector-screening system can facilitate the development of a targeted adenovirus vector for a variety of applications in medicine.
在载体开发中，在细胞进入水平上进行基因转移靶向是最具吸引力的挑战之一。然而，通过改造衣壳编码区将腺病毒载体重定向至其他受体的尝试取得的成功有限，因为目标细胞上合适的靶向配体 - 受体系统通常并不清楚。需要开发系统性方法来生成靶向任何给定细胞类型的腺病毒载体以实现这一目标。在此，我们构建了一个腺病毒文库，该文库通过腺病毒纤维修饰质粒文库与基因组DNA之间的Cre-lox介导的体外重组产生，以在纤维钮上展示随机肽段。作为概念验证，我们在胶质瘤细胞系上筛选了腺病毒展示文库，并观察到几种特定肽序列的选择。携带最常分离肽段的靶向载体显著增强了在胶质瘤细胞系中的基因转导，但在许多其他细胞系中则不然。由于将预先选择的肽段插入纤维钮中常常无法产生腺病毒载体，因此在腺病毒衣壳的背景下选择靶向肽非常有用。这种载体筛选系统可促进用于医学中各种应用的靶向腺病毒载体的开发。
