van der Meulen Karen, Smets Greet, Rüdelsheim Patrick
PERSEUS bv, Sint-Martens-Latem, Belgium.
Appl Biosaf. 2023 Jun 1;28(2):102-122. doi: 10.1089/apb.2022.0037. Epub 2023 Jun 5.
Viral RNA replicons are self-amplifying RNA molecules generated by deleting genetic information of one or multiple structural proteins of wild-type viruses. Remaining viral RNA is used as such (naked replicon) or packaged into a viral replicon particle (VRP), whereby missing genes or proteins are supplied via production cells. Since replicons mostly originate from pathogenic wild-type viruses, careful risk consideration is crucial.
A literature review was performed compiling information on potential biosafety risks of replicons originating from positive- and negative-sense single-stranded RNA viruses (except retroviruses).
For naked replicons, risk considerations included genome integration, persistence in host cells, generation of virus-like vesicles, and off-target effects. For VRP, the main risk consideration was formation of primary replication competent virus (RCV) as a result of recombination or complementation. To limit the risks, mostly measures aiming at reducing the likelihood of RCV formation have been described. Also, modifying viral proteins in such a way that they do not exhibit hazardous characteristics in the unlikely event of RCV formation has been reported.
Despite multiple approaches developed to reduce the likelihood of RCV formation, scientific uncertainty remains on the actual contribution of the measures and on limitations to test their effectiveness. In contrast, even though effectiveness of each individual measure is unclear, using multiple measures on different aspects of the system may create a solid barrier. Risk considerations identified in the current study can also be used to support risk group assignment of replicon constructs based on a purely synthetic design.
病毒RNA复制子是通过删除野生型病毒一种或多种结构蛋白的遗传信息而产生的自我扩增RNA分子。剩余的病毒RNA可直接使用(裸复制子)或包装成病毒复制子颗粒(VRP),缺失的基因或蛋白则通过生产细胞提供。由于复制子大多源自致病性野生型病毒,因此仔细考虑风险至关重要。
进行了一项文献综述,汇编了有关源自正链和负链单链RNA病毒(逆转录病毒除外)的复制子潜在生物安全风险的信息。
对于裸复制子,风险考虑因素包括基因组整合、在宿主细胞中的持久性、病毒样囊泡的产生以及脱靶效应。对于VRP,主要的风险考虑因素是由于重组或互补而形成具有初级复制能力的病毒(RCV)。为了降低风险,大多描述了旨在降低RCV形成可能性的措施。此外,还报道了对病毒蛋白进行修饰,使其在不太可能形成RCV的情况下不表现出有害特征。
尽管已开发出多种方法来降低RCV形成的可能性,但对于这些措施的实际作用以及测试其有效性的局限性仍存在科学不确定性。相比之下,即使每项措施的有效性尚不清楚,但在系统的不同方面使用多种措施可能会形成一个坚实的屏障。本研究中确定的风险考虑因素也可用于支持基于纯合成设计的复制子构建体的风险组分配。