Cancer Cell-specific Transfection of hCas9 Gene Using Ad5F35 Vector
Cancer Cell-specific Transfection of hCas9 Gene Using Ad5F35 Vector
复制标题
DOI：
10.21873/anticanres.15164
复制
发表时间：
2021-08-01
影响因子：
2
通讯作者：
Gotoh, Akinobu
中科院分区：
文献类型：
--
作者：
Matsunaga, Wataru;Hamada, Katsuyuki;Gotoh, Akinobu
背景资料：成簇的规则间隔的短回文重复相关蛋白9（CRISPR-Cas9）被认为具有广阔的临床潜力。然而，Cas9的脱靶效应是其应用的主要问题。因此，我们假设，如果人类密码子优化的化脓性链球菌Cas9（hCas 9）可以在癌细胞中特异性表达，则脱靶基因编辑的不良影响可能会最小化。材料和方法：构建了嵌合腺病毒载体Ad 5 F35-MKp-hCas 9，并将其感染人膀胱癌细胞系。在感染后3-4天内进行hCas 9基因表达的确认。结果：在Ad 5 F35-MKphCas 9感染的膀胱癌细胞中观察到hCas 9基因表达，但在非恶性细胞中未观察到。结论：Ad 5 F35-MKphCas 9载体能够在膀胱癌细胞中特异性表达hCas 9基因。这些发现可能有助于最大限度地降低基因编辑脱靶效应的风险。
Background: The clustered regularly interspaced short palindromic repeats-associated protein 9 (CRISPR-Cas9) is thought to have promising clinical potential. However, the off-target effects of Cas9 are a major concern for its application. Therefore, we hypothesized that the adverse effects of off-target gene editing might be minimized if the human codon-optimized Streptococcus pyogenes Cas9 (hCas9) could be specifically expressed in cancer cells. Materials and Methods: We constructed a chimeric adenoviral vector, Ad5F35-MKp-hCas9, and infected human bladder cancer cell lines with this vector. The confirmation of hCas9 gene expression was performed in 3-4 days after from infection. Results: hCas9 gene expression was observed in Ad5F35-MKphCas9 infected bladder cancer cells but not in non-malignant cells. Conclusion: Our study showed that the Ad5F35-MKphCas9 vector is capable of expressing the hCas9 gene with high specificity in bladder cancer cells. These findings may help in minimizing the risk of off-target effects of gene editing.