Inhibition of Phosphodiesterase 12 results in Antiviral Activity against several RNA Viruses including SARS-CoV-2.

Phosphodiesterase 12 (PDE12) is a negative regulator of the type 1 interferon (IFN) response and here we show that PDE12 inhibitors (lead compounds 63 and 17) are associated with increased RNAseL activity, are well tolerated at the therapeutic range and inhibit, both in vitro and in vivo, the replication of several RNA viruses including hepatitis C virus (HCV), dengue virus (DENV), West Nile Virus (WNV) and SARS-CoV-2.

Rapid selection of P323L in the SARS-CoV-2 polymerase (NSP12) in humans and non-human primate models and confers a large plaque phenotype

The mutational landscape of SARS-CoV-2 varies at both the dominant viral genome sequence and minor genomic variant population. An early change associated with transmissibility was the D614G substitution in the spike protein. This appeared to be accompanied by a P323L substitution in the viral polymerase (NSP12), but this latter change was not under strong selective pressure. Investigation of P323L/D614G changes in the human population showed rapid emergence during the containment phase and early surge phase of wave 1 in the UK. This rapid substitution was from minor genomic variants to become part of the dominant viral genome sequence. A rapid emergence of 323L but not 614G was observed in a non-human primate model of COVID-19 using a starting virus with P323 and D614 in the dominant genome sequence and 323L and 614G in the minor variant population. In cell culture, a recombinant virus with 323L in NSP12 had a larger plaque size than the same recombinant virus with P323. These data suggest that it may be possible to predict the emergence of a new variant based on tracking the distribution and frequency of minor variant genomes at a population level, rather than just focusing on providing information on the dominant viral genome sequence e.g., consensus level reporting. The ability to predict an emerging variant of SARS-CoV-2 in the global landscape may aid in the evaluation of medical countermeasures and non-pharmaceutical interventions.

Identification and quantification of SARS-CoV-2 leader subgenomic mRNA gene junctions in nasopharyngeal samples shows phasic transcription in animal models of COVID-19 and aberrant pattens in humans

IntroductionSARS-CoV-2 has a complex strategy for the transcription of viral subgenomic mRNAs (sgmRNAs), which are targets for nucleic acid diagnostics. Each of these sgRNAs has a unique 5 sequence, the leader-transcriptional regulatory sequence gene junction (leader-TRS-junction), that can be identified using sequencing. ResultsHigh resolution sequencing has been used to investigate the biology of SARS-CoV-2 and the host response in cell culture models and from clinical samples. LeTRS, a bioinformatics tool, was developed to identify leader-TRS-junctions and be used as a proxy to quantify sgmRNAs for understanding virus biology. This was tested on published datasets and clinical samples from patients and longitudinal samples from animal models with COVID-19. DiscussionLeTRS identified known leader-TRS-junctions and identified novel species that were common across different species. The data indicated multi-phasic abundance of sgmRNAs in two different animal models, with spikes in sgmRNA abundance reflected in human samples, and therefore has implications for transmission models and nucleic acid-based diagnostics.

A COVID-19 vaccine candidate using SpyCatcher multimerization of the SARS-CoV-2 spike protein receptor-binding domain induces potent neutralising antibody responses

There is dire need for an effective and affordable vaccine against SARS-CoV-2 to tackle the ongoing pandemic. In this study, we describe a modular virus-like particle vaccine candidate displaying the SARS-CoV-2 spike glycoprotein receptor-binding domain (RBD) using SpyTag/SpyCatcher technology (RBD-SpyVLP). Low doses of RBD-SpyVLP in a prime-boost regimen induced a strong neutralising antibody response in mice and pigs that was superior to convalescent human sera. We evaluated antibody quality using ACE2 blocking and neutralisation of cell infection by pseudovirus or wild-type SARS-CoV-2. Using competition assays with a monoclonal antibody panel, we showed that RBD-SpyVLP induced a polyclonal antibody response that recognised all key epitopes on the RBD, reducing the likelihood of selecting neutralisation-escape mutants. The induction of potent and polyclonal antibody responses by RBD-SpyVLP provides strong potential to address clinical and logistic challenges of the COVID-19 pandemic. Moreover, RBD-SpyVLP is highly resilient, thermostable and can be lyophilised without losing immunogenicity, to facilitate global distribution and reduce cold-chain dependence.

Structural basis for the neutralization of SARS-CoV-2 by an antibody from a convalescent patient

The COVID-19 pandemic has had unprecedented health and economic impact, but currently there are no approved therapies. We have isolated an antibody, EY6A, from a late-stage COVID-19 patient and show it neutralises SARS-CoV-2 and cross-reacts with SARS-CoV-1. EY6A Fab binds tightly (KD of 2 nM) the receptor binding domain (RBD) of the viral Spike glycoprotein and a 2.6[A] crystal structure of an RBD/EY6A Fab complex identifies the highly conserved epitope, away from the ACE2 receptor binding site. Residues of this epitope are key to stabilising the pre-fusion Spike. Cryo-EM analyses of the pre-fusion Spike incubated with EY6A Fab reveal a complex of the intact trimer with three Fabs bound and two further multimeric forms comprising destabilized Spike attached to Fab. EY6A binds what is probably a major neutralising epitope, making it a candidate therapeutic for COVID-19.