Continued Virus-Specific Antibody-Secreting Cell Production, Avidity Maturation and B Cell Evolution in Patients Hospitalized with COVID-19.

Understanding the development and sustainability of the virus-specific protective immune response to infection with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) remains incomplete with respect to the appearance and disappearance of virus-specific antibody-secreting cells (ASCs) in circulation. Therefore, we performed cross-sectional and longitudinal analyses of peripheral blood mononuclear cells and plasma collected from 55 hospitalized patients up to 4 months after onset of COVID-19 symptoms. Spike (S)- and nucleocapsid (N)-specific IgM and IgG ASCs appeared within 2 weeks accompanied by flow cytometry increases in double negative plasmablasts consistent with a rapid extrafollicular B cell response. Total and virus-specific IgM and IgG ASCs peaked at 3-4 weeks and were still being produced at 3-4 months accompanied by increasing antibody avidity consistent with a slower germinal center B cell response. N-specific ASCs were produced for longer than S-specific ASCs and avidity maturation was greater for antibody to N than S. Patients with more severe disease produced more S-specific IgM and IgG ASCs than those with mild disease and had higher levels of N- and S-specific antibody. Women had more B cells in circulation than men and produced more S-specific IgA and IgG and N-specific IgG ASCs. Flow cytometry analysis of B cell phenotypes showed an increase in circulating B cells at 4-6 weeks with decreased percentages of switched and unswitched memory B cells. These data indicate ongoing antigen-specific stimulation, maturation, and production of ASCs for several months after onset of symptoms in patients hospitalized with COVID-19.

Overcoming anti-PEG antibody mediated accelerated blood clearance of PEGylated liposomes by pre-infusion with high molecular weight free PEG.

Antibodies that specifically bind polyethylene glycol (PEG), i.e. anti-PEG antibodies (APA), are associated with reduced efficacy and increased risk of serious adverse events for several PEGylated therapeutics. Here, we explored the concept of using free PEG molecules to saturate circulating APA. Surprisingly, we found that 40 kDa free PEG effectively restored the prolonged circulation of PEGylated liposomes in the presence of high titers of pre-existing APA for at least 48 h in mice. In contrast, lower molecular weight free PEG (≤10 kDa) failed to restore circulation beyond a few hours. These in vivo results were consistent with estimates from a minimal physiologically based pharmacokinetic model. Importantly, the infusion of free PEG appeared to be safe in mice previously sensitized by injection of PEGylated liposomes, and free PEG did not elicit excess APA production even in mice with pre-existing adaptive immunity against PEG. Our results support further investigation of high molecular weight free PEG as a potential method to control and overcome high titers of APA, restoring the prolonged circulation of PEGylated liposomes and possibly other PEGylated therapeutics.