Accelerated waning of the humoral response to COVID-19 vaccines in obesity.

Obesity is associated with an increased risk of severe Coronavirus Disease 2019 (COVID-19) infection and mortality. COVID-19 vaccines reduce the risk of serious COVID-19 outcomes; however, their effectiveness in people with obesity is incompletely understood. We studied the relationship among body mass index (BMI), hospitalization and mortality due to COVID-19 among 3.6 million people in Scotland using the Early Pandemic Evaluation and Enhanced Surveillance of COVID-19 (EAVE II) surveillance platform. We found that vaccinated individuals with severe obesity (BMI > 40 kg/m2) were 76% more likely to experience hospitalization or death from COVID-19 (adjusted rate ratio of 1.76 (95% confidence interval (CI), 1.60-1.94). We also conducted a prospective longitudinal study of a cohort of 28 individuals with severe obesity compared to 41 control individuals with normal BMI (BMI 18.5-24.9 kg/m2). We found that 55% of individuals with severe obesity had unquantifiable titers of neutralizing antibody against authentic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus compared to 12% of individuals with normal BMI (P = 0.0003) 6 months after their second vaccine dose. Furthermore, we observed that, for individuals with severe obesity, at any given anti-spike and anti-receptor-binding domain (RBD) antibody level, neutralizing capacity was lower than that of individuals with a normal BMI. Neutralizing capacity was restored by a third dose of vaccine but again declined more rapidly in people with severe obesity. We demonstrate that waning of COVID-19 vaccine-induced humoral immunity is accelerated in individuals with severe obesity. As obesity is associated with increased hospitalization and mortality from breakthrough infections, our findings have implications for vaccine prioritization policies.

Translation of in vitro-transcribed RNA therapeutics.

In vitro transcribed, modified messenger RNAs (IVTmRNAs) have been used to vaccinate billions of individuals against the SARS-CoV-2 virus, and are currently being developed for many additional therapeutic applications. IVTmRNAs must be translated into proteins with therapeutic activity by the same cellular machinery that also translates native endogenous transcripts. However, different genesis pathways and routes of entry into target cells as well as the presence of modified nucleotides mean that the way in which IVTmRNAs engage with the translational machinery, and the efficiency with which they are being translated, differs from native mRNAs. This review summarises our current knowledge of commonalities and differences in translation between IVTmRNAs and cellular mRNAs, which is key for the development of future design strategies that can generate IVTmRNAs with improved activity in therapeutic applications.

Development of a colorimetric assay for the detection of SARS-CoV-2 3CLpro activity.

Diagnostic testing continues to be an integral component of the strategy to contain the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) global pandemic, the causative agent of Coronavirus Disease 2019 (COVID-19). The SARS-CoV-2 genome encodes the 3C-like protease (3CLpro) which is essential for coronavirus replication. This study adapts an in vitro colorimetric gold nanoparticle (AuNP) based protease assay to specifically detect the activity of SARS-CoV-2 3CLpro as a purified recombinant protein and as a cellular protein exogenously expressed in HEK293T human cells. We also demonstrate that the specific sensitivity of the assay for SARS-CoV-2 3CLpro can be improved by use of an optimised peptide substrate and through hybrid dimerisation with inactive 3CLpro mutant monomers. These findings highlight the potential for further development of the AuNP protease assay to detect SARS-CoV-2 3CLpro activity as a novel, accessible and cost-effective diagnostic test for SARS-CoV-2 infection at the point-of-care. Importantly, this versatile assay could also be easily adapted to detect specific protease activity associated with other viruses or diseases conditions.

A protease-activatable luminescent biosensor and reporter cell line for authentic SARS-CoV-2 infection.

Efforts to define serological correlates of protection against COVID-19 have been hampered by the lack of a simple, scalable, standardised assay for SARS-CoV-2 infection and antibody neutralisation. Plaque assays remain the gold standard, but are impractical for high-throughput screening. In this study, we show that expression of viral proteases may be used to quantitate infected cells. Our assays exploit the cleavage of specific oligopeptide linkers, leading to the activation of cell-based optical biosensors. First, we characterise these biosensors using recombinant SARS-CoV-2 proteases. Next, we confirm their ability to detect viral protease expression during replication of authentic virus. Finally, we generate reporter cells stably expressing an optimised luciferase-based biosensor, enabling viral infection to be measured within 24 h in a 96- or 384-well plate format, including variants of concern. We have therefore developed a luminescent SARS-CoV-2 reporter cell line, and demonstrated its utility for the relative quantitation of infectious virus and titration of neutralising antibodies.