Rotavirus as an Expression Platform of the SARS-CoV-2 Spike Protein

Philip, Asha A.; Patton, John T.

doi:10.1101/2021.02.18.431835

Cited by 8 publications

(3 citation statements)

References 46 publications

(64 reference statements)

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“…With the exception of T2A_RBM, NSP3 mutants expressing SARS-CoV-2 spike peptides cross-reacted with the SARS-CoV-2 RBD antibody (Fig. 6B), supporting the previously proposed viability of using NSP3 as a tagging system [52] and demonstrating its application to other RV strains. Follow up studies using sera from COVID-19 patients are needed to confirm antibody responses to the antigens produced by the NSP3 mutants.…”

Section: Discussionsupporting

confidence: 76%

“…Previous studies have shown that SA11 NSP3 is able to tolerate insertions of foreign sequences at its C-terminus [45, 51, 52]; we have demonstrated that this is also possible using the bovine RF strain NSP3. This is advantageous as the current live attenuated pentavalent RV vaccine, RotaTeq, utilises a bovine RV backbone reassorted with different human strains and the use of a non-primate origin RV vector helps to circumvent pre-existing immunity [10, 69, 95].…”

Section: Discussionmentioning

confidence: 58%

“…The C-terminus of the SA11 NSP3 open reading frame (ORF) was fused to a porcine teschovirus translational 2A element followed by various reporters including UnaG, mKate, mRuby or TagBFP to successfully yield two uncoupled proteins without compromising virus replication [51]. A more recent study showed that the C-terminus of SA11 NSP3 can express different peptides of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein with minimal impact on endpoint titres [52]. This has highlighted the potential to use RVs as expression vectors for development of polyvalent vaccines for enteric viruses.…”

Section: Introductionmentioning

confidence: 99%

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Engineering a Vaccine Platform using Rotavirus A to Express SARS-CoV-2 Spike Epitopes

Diebold

Gonzalez

Venditti

et al. 2022

Preprint

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Human rotavirus (RV) vaccines used worldwide have been developed using live attenuated platforms. The recent development of a reverse genetics system for RVs has delivered the possibility of engineering chimeric viruses expressing heterologous peptides from other virus species to generate polyvalent vaccines. We tested the feasibility of this using two approaches. Firstly, we inserted short SARS-CoV-2 spike peptides into the hypervariable region of the simian SA11 RV strain viral protein (VP) 4. Secondly, we fused the receptor binding domain (RBD) of the SARS-CoV-2 spike protein, or the shorter receptor binding motif (RBM) nested within the RBD, to the C-terminus of non-structural protein (NSP) 3 of the bovine RF strain RV, with or without an intervening T2A peptide. Mutating the hypervariable region of SA11 VP4 impeded viral replication, and for these mutants no cross-reactivity with spike antibodies was detected. To rescue NSP3 mutants, we established a plasmid-based reverse genetics system for the bovine RF strain. Except for the RBD mutant, all NSP3 mutants delivered endpoint titres and replication kinetics comparable to that of the WT virus. In ELISAs, cell lysates of an NSP3 mutant expressing the RBD peptide showed cross reactivity with a SARS-CoV-2 RBD antibody. 3D bovine gut enteroids were susceptible to infection by all NSP3 mutants but only RBM mutant showed cross reactivity with SARS-CoV-2 RBD antibody. The tolerability of large peptide insertions in the NSP3 segment highlights the potential for this approach in the development of vaccine vectors targeting multiple enteric pathogens simultaneously.

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Section: Discussionsupporting

confidence: 76%

Section: Discussionmentioning

confidence: 58%

Section: Introductionmentioning

confidence: 99%

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Engineering a Vaccine Platform using Rotavirus A to Express SARS-CoV-2 Spike Epitopes

Diebold

Gonzalez

Venditti

et al. 2022

Preprint

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Mucosal and systemic neutralizing antibodies to norovirus and rotavirus by oral immunization with recombinant rotavirus in infant mice

Sánchez-Tacuba

Feng

Costantini

et al. 2022

Preprint

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Rotaviruses (RVs) preferentially replicate in the small intestine, frequently cause severe diarrheal disease, and following enteric infection generally induce variable levels of protective systemic and mucosal immune responses in humans and other animals. Rhesus rotavirus (RRV) is a simian RV that was previously used as a human RV vaccine and has been extensively studied in mice. Although RRV replicates poorly in the suckling mouse intestine, infection induces a robust and protective antibody response. The recent availability of plasmid-based RV reverse genetics systems has enabled the generation of recombinant RVs expressing foreign proteins. However, recombinant RVs have not yet been experimentally tested as potential vaccine vectors to immunize against other gastrointestinal pathogens in vivo. This is a missed opportunity because several live-attenuated RV vaccines are already widely administered to infants and young children worldwide. To explore the feasibility of using RV as a dual vaccine vector, we rescued a replication-competent recombinant RRV harboring bicistronic gene segment 7 that encodes both the native RV NSP3 protein and a human norovirus (HuNoV) VP1 protein from the predominant genotype GII.4 (rRRV-HuNoV-VP1). The rRRV-HuNoV-VP1 expressed HuNoV VP1 in infected cells in vitro and importantly, elicited both systemic and local antibody responses to HuNoV following oral infection of suckling mice. Serum IgG and fecal IgA from infected suckling mice bound to and neutralized both RV and HuNoV. These findings have encouraging practical implications for the design of RV-based next-generation multivalent enteric vaccines to target HuNoV and other human enteric pathogens while providing immunity to RV.

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Human Rotavirus Reverse Genetics Systems to Study Viral Replication and Pathogenesis

Komoto¹,

Fukuda²,

Murata³

et al. 2021

Viruses

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Human rotaviruses (HuRVAs) are highly important causes of acute gastroenteritis in infants and young children worldwide. A lack of reliable and reproducible reverse genetics systems for HuRVAs has limited a proper understanding of HuRVA biology and also the rational design of live-attenuated vaccines. Since the development of the first reverse genetics system for RVAs (partially plasmid-based reverse genetics system) in 2006, there have been many efforts with the goal of generating infectious recombinant HuRVAs entirely from cloned cDNAs. However, the establishment of a HuRVA reverse genetics system was very challenging until 2019. This review article provides an overview of the historical background of the recent development of long-awaited HuRVA reverse genetics systems, beginning with the generation of recombinant human-simian reassortant RVAs with the aid of a helper virus in 2006 and the generation of recombinant animal (simian) RVAs in a helper virus-free manner in 2017, and culminating in the generation of recombinant HuRVAs entirely from plasmid cDNAs in 2019. Notably, the original HuRVA reverse genetics system has already been optimized to increase the efficiency of virus generation. Although the application of HuRVA reverse genetics systems has only just been initiated, these technologies will help to answer HuRVA research questions regarding viral replication and pathogenicity that could not be addressed before, and to develop next-generation vaccines and intestine-specific rotaviral vectors.

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Rotavirus as an Expression Platform of the SARS-CoV-2 Spike Protein

Cited by 8 publications

References 46 publications

Engineering a Vaccine Platform using Rotavirus A to Express SARS-CoV-2 Spike Epitopes

Engineering a Vaccine Platform using Rotavirus A to Express SARS-CoV-2 Spike Epitopes

Mucosal and systemic neutralizing antibodies to norovirus and rotavirus by oral immunization with recombinant rotavirus in infant mice

Human Rotavirus Reverse Genetics Systems to Study Viral Replication and Pathogenesis

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