One or two injections of MVA-vectored vaccine shields hACE2 transgenic mice from SARS-CoV-2 upper and lower respiratory tract infection

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.

Recombinant DNA methods have revolutionized the engineering of vaccines against microbial pathogens, thereby creating opportunities to control the current COVID-19 pandemic (1). The main categories of recombinant vaccines are protein, nucleic acid (DNA and RNA), virus vectors (replicating and nonreplicating), and genetically modified live viruses. Each approach has advantages and drawbacks with regard to manufacture, stability, cold-chain requirements, mode of inoculation, and immune stimulation. Recombinant proteins have been successfully deployed as vaccines against a variety of diseases (2–5). DNA vaccines have been licensed for veterinary purposes (6, 7), although none are in regular human use. Recently developed messenger RNA (mRNA) vaccines are in use for COVID-19 and are in preclinical development for other infectious diseases (8). At least 12 virus vector vaccines based on adenovirus, fowlpox virus, vaccinia virus (VACV), and yellow fever virus have veterinary applications, but, so far, only two have been marketed for humans (9), although numerous clinical trials, particularly with attenuated adenovirus and VACV, are listed online in ClinicalTrials.gov.A variety of recombinant approaches utilizing the spike (S) protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2; abbreviated CoV-2) as immunogen are being explored to quell the COVID-19 pandemic (10). Vaccines based on mRNA and adenovirus vectors have demonstrated promising results in clinical trials and have received emergency regulatory approval (11–14). Other candidate CoV-2 vaccines, including ones based on vesicular stomatitis virus (15), an alphavirus-derived replicon RNA (16), an inactivated recombinant Newcastle Disease virus (17), and modified VACV Ankara (MVA) (18, 19) are at early stages of evaluation.Experiments with virus vectors for vaccination were carried out initially with VACV (20, 21), providing a precedent for a multitude of other virus vectors (9). The majority of current VACV vaccine studies employ the MVA strain, which was attenuated by more than 500 passages in chicken embryo fibroblasts during which numerous genes were deleted or mutated, resulting in an inability to replicate in human and most other mammalian cells (22). Despite the inability to complete a productive infection, MVA is capable of highly expressing recombinant genes and inducing immune responses (23, 24). MVA is a licensed smallpox vaccine, and numerous clinical studies of recombinant MVA (rMVA) vectors are in progress or have been completed. Protection has been obtained with MVA-based SARS-CoV-1 and Middle East respiratory syndrome CoV (MERS-CoV) in animals (25–28), and an MVA-based MERS-CoV vaccine was shown to be safe and immunogenic in a phase 1 clinical trial (29). Currently, two clinical trials for MVA-based CoV-2 vaccines are in the recruiting phase (ClinicalTrials.gov). Here, we show that one or two immunizations with rMVAs expressing the CoV-2 S proteins elicit strong neutralizing antibody responses, induce CD8+ T cells, and protect susceptible transgenic mice against a lethal intranasal challenge with CoV-2 virus, supporting clinical testing of related rMVA vaccines.