Liu Ruikang, Americo Jeffrey L, Cotter Catherine A, Earl Patricia L, Erez Noam, Peng Chen, Moss Bernard
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892.
Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, MD 20892
Proc Natl Acad Sci U S A. 2021 Mar 23;118(12). doi: 10.1073/pnas.2026785118.
Modified vaccinia virus Ankara (MVA) is a replication-restricted smallpox vaccine, and numerous clinical studies of recombinant MVAs (rMVAs) as vectors for prevention of other infectious diseases, including COVID-19, are in progress. Here, we characterize rMVAs expressing the S protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Modifications of full-length S individually or in combination included two proline substitutions, mutations of the furin recognition site, and deletion of the endoplasmic retrieval signal. Another rMVA in which the receptor binding domain (RBD) is flanked by the signal peptide and transmembrane domains of S was also constructed. Each modified S protein was displayed on the surface of rMVA-infected cells and was recognized by anti-RBD antibody and soluble hACE2 receptor. Intramuscular injection of mice with the rMVAs induced antibodies, which neutralized a pseudovirus in vitro and, upon passive transfer, protected hACE2 transgenic mice from lethal infection with SARS-CoV-2, as well as S-specific CD3+CD8+IFNγ+ T cells. Antibody boosting occurred following a second rMVA or adjuvanted purified RBD protein. Immunity conferred by a single vaccination of hACE2 mice prevented morbidity and weight loss upon intranasal infection with SARS-CoV-2 3 wk or 7 wk later. One or two rMVA vaccinations also prevented detection of infectious SARS-CoV-2 and subgenomic viral mRNAs in the lungs and greatly reduced induction of cytokine and chemokine mRNAs. A low amount of virus was found in the nasal turbinates of only one of eight rMVA-vaccinated mice on day 2 and none later. Detection of low levels of subgenomic mRNAs in turbinates indicated that replication was aborted in immunized animals.
安卡拉痘苗病毒（MVA）是一种复制受限的天花疫苗，目前有多项关于重组MVA（rMVA）作为预防包括COVID-19在内的其他传染病载体的临床研究正在进行。在此，我们对表达严重急性呼吸综合征冠状病毒2（SARS-CoV-2）S蛋白的rMVA进行了表征。全长S的单独或组合修饰包括两个脯氨酸取代、弗林蛋白酶识别位点的突变以及内质网回收信号的缺失。还构建了另一种rMVA，其中受体结合域（RBD）两侧为S的信号肽和跨膜域。每种修饰的S蛋白都展示在rMVA感染细胞的表面，并被抗RBD抗体和可溶性人血管紧张素转换酶2（hACE2）受体识别。给小鼠肌肉注射rMVA可诱导产生抗体，这些抗体在体外可中和假病毒，经被动转移后可保护hACE2转基因小鼠免受SARS-CoV-2致死性感染，同时还可诱导产生S特异性CD3⁺CD8⁺IFNγ⁺ T细胞。第二次接种rMVA或佐剂化纯化RBD蛋白后出现抗体增强。对hACE2小鼠进行单次疫苗接种所赋予的免疫力可预防3周或7周后经鼻感染SARS-CoV-2时的发病和体重减轻。接种一次或两次rMVA还可防止在肺部检测到传染性SARS-CoV-2和亚基因组病毒mRNA，并大大减少细胞因子和趋化因子mRNA的诱导。在第2天，仅在8只接种rMVA的小鼠中的1只的鼻甲中发现少量病毒，之后未再检测到。鼻甲中低水平亚基因组mRNA的检测表明免疫动物中的复制被中止。
