COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has significantly impacted public health and the economy worldwide. Most of the currently licensed COVID-19 vaccines act by inhibiting the receptor-binding function of the SARS-CoV-2 spike protein. The constant emergence of SARS-CoV-2 variants resulting from mutations in the receptor-binding domain (RBD) leads to vaccine immune evasion and underscores the importance of broadly acting COVID-19 vaccines. Inactivated whole virus vaccines can elicit broader immune responses to multiple epitopes of several antigens and help overcome such immune evasions. We prepared a psoralen-inactivated SARS-CoV-2 vaccine (SARS-CoV-2 PsIV) and evaluated its immunogenicity and efficacy in nonhuman primates (NHPs) when administered with the Advax-CpG adjuvant. We also evaluated the SARS-CoV-2 PsIV as a booster shot in animals vaccinated with a DNA vaccine that can express the full-length spike protein. The Advax-CpG-adjuvanted SARS-CoV-2 PsIV elicited a dose-dependent neutralizing antibody response in the NHPs, as measured using a serum microneutralization assay against the SARS-CoV-2 Washington strain and the Delta variant. The animals vaccinated with the DNA vaccine followed by a boosting dose of the SARS-CoV-2 PsIV exhibited the highest neutralizing antibody responses and were able to quickly clear infection after an intranasal challenge with the SARS-CoV-2 Delta variant. Overall, the data show that the Advax-CpG-adjuvanted SARS-CoV-2 PsIV, either by itself or as a booster shot following nucleic acid (NA) vaccines, has the potential to protect against emerging variants.

Viral proteins are known to be methylated by host protein arginine methyltransferases (PRMTs) necessary for the viral life cycle, but it remains unknown whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) proteins are methylated. Herein, we show that PRMT1 methylates SARS-CoV-2 nucleocapsid (N) protein at residues R95 and R177 within RGG/RG motifs, preferred PRMT target sequences. We confirmed arginine methylation of N protein by immunoblotting viral proteins extracted from SARS-CoV-2 virions isolated from cell culture. Type I PRMT inhibitor (MS023) or substitution of R95 or R177 with lysine inhibited interaction of N protein with the 5'-UTR of SARS-CoV-2 genomic RNA, a property required for viral packaging. We also defined the N protein interactome in HEK293 cells, which identified PRMT1 and many of its RGG/RG substrates, including the known interacting protein G3BP1 as well as other components of stress granules (SGs), which are part of the host antiviral response. Methylation of R95 regulated the ability of N protein to suppress the formation of SGs, as R95K substitution or MS023 treatment blocked N-mediated suppression of SGs. Also, the coexpression of methylarginine reader Tudor domain-containing protein 3 quenched N protein-mediated suppression of SGs in a dose-dependent manner. Finally, pretreatment of VeroE6 cells with MS023 significantly reduced SARS-CoV-2 replication. Because type I PRMT inhibitors are already undergoing clinical trials for cancer treatment, inhibiting arginine methylation to target the later stages of the viral life cycle such as viral genome packaging and assembly of virions may represent an additional therapeutic application of these drugs.
Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.