Non-classical CD45RBlo memory B-cells are the majority of circulating antigen-specific B-cells following mRNA vaccination and COVID-19 infection. (preprint)

Resting memory B-cells can be divided into classical and non-classical groups based on differential expression of markers such as CD27 and CD11c, while activated memory B-cells express a combination of markers, making their ontogeny hard to determine. Here by longitudinal analysis of COVID-19, bacterial sepsis, and BNT162b2 mRNA vaccine recipients by mass cytometry and CITE-seq we describe a three-branch structure of resting B-cell memory consisting of “classical” CD45RB+ memory and two branches of CD45RBlo memory further defined by expression of CD23 and CD11c respectively. Stable differences in CD45RB upon activation allowed tracking of activated B-cells and plasmablasts derived from CD45RB+ classical and CD45RBlo non-classical memory B-cells. In both COVID-19 patients and mRNA vaccination, CD45RBlo B-cells formed the majority of SARS-CoV2 specific memory B-cells and correlated with serum antibodies while CD45RB+ memory was most strongly activated by bacterial Sepsis. These results suggest that diverse non-classical CD45RBlo memory B-cells consisting of branches of CD11c+Tbet+ and CD23+ fractions form a critical part of responses to viral infection and vaccination.

Classification of patients with COVID-19 by blood RNA endotype: A prospective cohort study (preprint)

Background: Although the development of vaccines has considerably reduced the severity of COVID-19, its incidence is still high. Hence, a targeted approach based on RNA endotypes of a population should be developed to help design biomarker-based therapies for COVID-19. Objectives: We evaluated the major RNAs transcribed in blood cells during COVID-19 using PCR to further elucidate its pathogenesis and determine predictive phenotypes in COVID-19 patients. Study design: In a discovery cohort of 40 patients with COVID-19, 26,354 RNAs were measured on day 1 and day 7. Five RNAs associated with disease severity and prognosis were derived. In a validation cohort of 153 patients with COVID-19 treated in the intensive care unit, we focused on prolactin (PRL), and toll-like receptor 3 (TLR3) among RNAs, which have a strong association with prognosis, and evaluated the accuracy for predicting survival of PRL-to-TL3 ratios (PRL/TLR3) with the areas under the ROC curves (AUC). The validation cohort was divided into two groups based on the cut-off value in the ROC curve with the maximum AUC. The two groups were defined by high PRL/TLR3 (n=47) and low PRL/TLR3 groups (n=106) and the clinical outcomes were compared. Results: In the validation cohort, the AUC for PRL/TLR3 was 0.79, showing superior prognostic ability compared to severity scores such as APACHE II and SOFA. The high PRL/TLR3 group had a significantly higher 28-day mortality than the low PRL/TLR3 group (17.0% vs 0.9%, P<0.01). Conclusions: A new RNA endotype classified using high PRL/TLR3 was associated with mortality in COVID-19 patients.

An engineered ACE2 decoy broadly neutralizes Omicron subvariants and shows therapeutic effect in SARS-CoV-2-infected cynomolgus macaques (preprint)

The Omicron variant continuously evolves under the humoral immune pressure obtained by vaccination and SARS-CoV-2 infection and the resultant Omicron subvariants exhibit further immune evasion and antibody escape. Engineered ACE2 decoy composed of high-affinity ACE2 and IgG1 Fc domain is an alternative modality to neutralize SARS-CoV-2 and we previously reported its broad spectrum and therapeutic potential in rodent models. Here, we show that engineered ACE2 decoy retains the neutralization activity against Omicron subvariants including the currently emerging XBB and BQ.1 which completely evade antibodies in clinical use. The culture of SARS-CoV-2 under suboptimal concentration of neutralizing drugs generated SARS-CoV-2 mutants escaping wild-type ACE2 decoy and monoclonal antibodies, whereas no escape mutant emerged against engineered ACE2 decoy. As the efficient drug delivery to respiratory tract infection of SARS-CoV-2, inhalation of aerosolized decoy treated mice infected with SARS-CoV-2 at a 20-fold lower dose than the intravenous administration. Finally, engineered ACE2 decoy exhibited the therapeutic efficacy for COVID-19 in cynomolgus macaques. Collectively, these results indicate that engineered ACE2 decoy is the promising therapeutic strategy to overcome immune-evading SARS-CoV-2 variants and that liquid aerosol inhalation can be considered as a non-invasive approach to enhance efficacy in the treatment of COVID-19.

Resistin Forms A Network With Inflammatory Cytokines And Endothelial Damage Markers And Provides Prognostic And Therapeutic Information For COVID-19 (preprint)

Background: Resistin increases in septic subjects and is associated with severity and prognosis. Its role in Coronavirus disease 2019 (COVID-19) is unknown. We investigated relationships between resistin and the severity, prognosis and time to wean off mechanical ventilation (MV) in two cohorts. Methods: : Plasma resistin was available for 306 mild-to-critical COVID-19 patients on days 1, 4 and 8 from the Massachusetts General Hospital Emergency Department COVID-19 (MGH) cohort public proteomics data. The relationship between resistin and severity (World Health Organization COVID-19 outcomes) and the prognosis were evaluated. A cohort of 62 critical COVID-19 patients (Osaka cohort) was used to evaluate the relationship between resistin on days 1 (day of ICU admission), 2–3, 6–8 and 11–15 and the prognosis and time to wean off MV. Correlations among resistin, inflammatory cytokines and endothelial damage markers were evaluated. Results: : In the MGH cohort, day 1 resistin was associated with severity and predicted the prognosis in an ROC analysis (AUC, 0.739; 95% CI, 0.659–0.819). Twenty-eight-day non-survivors showed significantly greater resistin levels than 28-day survivors on days 1, 4 and 8. In the Osaka cohort, a Cox proportional hazards model (time dependent) showed a significant relationship between resistin and time to wean off MV (crude hazard ratio, 0.702 [95% CI, 0.508–0.969]). Resistin formed a network with inflammatory cytokines and endothelial damage markers. Conclusions: : Resistin was associated with severity, prognosis and time to wean off MV in COVID-19 patients. Resistin formed a network with inflammatory cytokines and endothelial damage markers, suggesting its contribution to the pathogenesis of COVID-19.