NSP16-deficient SARS-CoV- 2

Live attenuated vaccines might elicit mucosal and sterilizing immunity against SARS-CoV-2 that the existing mRNA, adenoviral vector and inactivated vaccines fail to induce. Here, we describe a candidate live attenuated vaccine strain of SARS-CoV-2 in which the NSP16 gene, which encodes 2′-O-methyltransferase, is catalytically disrupted by a point mutation. This virus, designated d16, was severely attenuated in hamsters and transgenic mice, causing only asymptomatic and nonpathogenic infection. A single dose of d16 administered intranasally resulted in sterilizing immunity in both the upper and lower respiratory tracts of hamsters, thus preventing viral spread in a contact-based transmission model. It also robustly stimulated humoral and cell-mediated immune responses, thus conferring full protection against lethal challenge with SARS-CoV-2 in a transgenic mouse model. The neutralizing antibodies elicited by d16 effectively cross-reacted with several SARS-CoV-2 variants. Secretory immunoglobulin A was detected in the blood and nasal wash of vaccinated mice. Our work provides proof-of-principle evidence for harnessing NSP16-deficient SARS- CoV-2 for the development of live attenuated vaccines and paves the way for further preclinical studies of d16 as a prototypic vaccine strain, to which new features might be introduced to improve safety, transmissibility, immunogenicity and efficacy.减毒活疫苗可诱导现有mRNA疫苗、腺病毒载体疫苗及灭活疫苗无法诱发的针对严重急性呼吸综合征冠状病毒2（SARS-CoV-2）的黏膜免疫与清除性免疫（sterilizing immunity）。本研究报道一株候选SARS-CoV-2减毒活疫苗株，其编码2'-O-甲基转移酶（2′-O-methyltransferase）的非结构蛋白16（NSP16）基因通过点突变（point mutation）实现了催化功能失活。该病毒被命名为d16，在仓鼠与转基因小鼠中呈现显著减毒特性，仅能引发无症状且无致病性的感染。经鼻内接种单剂d16即可在仓鼠的上下呼吸道中诱导清除性免疫，从而在接触传播模型中阻断病毒扩散。该疫苗株还可强效刺激体液免疫与细胞免疫应答，在转基因小鼠模型中可完全抵御SARS-CoV-2的致死性攻毒。d16诱导的中和抗体可与多种SARS-CoV-2变异株实现有效交叉反应，免疫小鼠的血液与鼻腔灌洗液中可检测到分泌型免疫球蛋白A（Secretory immunoglobulin A）。本研究为利用NSP16缺陷型SARS-CoV-2开发减毒活疫苗提供了原理验证证据，并为将d16作为原型疫苗株开展进一步临床前研究铺平了道路，后续可通过引入新的改造优化其安全性、传播性、免疫原性与保护效力。