Ottawa Hospital Research Institute, Ottawa, ON, K1H 8L6, Canada.
Faculty of Medicine and Health Sciences, Department of Microbiology and Infectious Diseases, Université de Sherbrooke, Sherbrooke, QC, J1E 4K8, Canada.
Nat Commun. 2023 May 26;14(1):3035. doi: 10.1038/s41467-023-38651-x.
The large coding potential of vaccinia virus (VV) vectors is a defining feature. However, limited regulatory switches are available to control viral replication as well as timing and dosing of transgene expression in order to facilitate safe and efficacious payload delivery. Herein, we adapt drug-controlled gene switches to enable control of virally encoded transgene expression, including systems controlled by the FDA-approved rapamycin and doxycycline. Using ribosome profiling to characterize viral promoter strength, we rationally design fusions of the operator element of different drug-inducible systems with VV promoters to produce synthetic promoters yielding robust inducible expression with undetectable baseline levels. We also generate chimeric synthetic promoters facilitating additional regulatory layers for VV-encoded synthetic transgene networks. The switches are applied to enable inducible expression of fusogenic proteins, dose-controlled delivery of toxic cytokines, and chemical regulation of VV replication. This toolbox enables the precise modulation of transgene circuitry in VV-vectored oncolytic virus design.
痘苗病毒（VV）载体的大编码潜力是其定义特征。然而，可用的调控开关有限，无法控制病毒复制以及转基因表达的时间和剂量，以促进安全有效的有效载荷传递。在此，我们采用药物控制的基因开关来控制病毒编码的转基因表达，包括受 FDA 批准的雷帕霉素和强力霉素控制的系统。我们使用核糖体图谱分析来描述病毒启动子的强度，然后合理设计不同药物诱导系统的操纵元件与 VV 启动子的融合，以产生合成启动子，从而产生强大的诱导表达，同时具有不可检测的基线水平。我们还生成嵌合合成启动子，为 VV 编码的合成转基因网络提供额外的调控层。这些开关可用于实现融合蛋白的诱导表达、毒性细胞因子的剂量控制传递以及 VV 复制的化学调控。该工具包可实现 VV 载体溶瘤病毒设计中转基因电路的精确调制。
