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BACKGROUND. SARS-CoV-2-specific antibodies may protect from reinfection and disease, providing rationale for administration of plasma containing SARS-CoV-2-neutralizing antibodies (nAbs) as a treatment for COVID-19. Clinical factors and laboratory assays to streamline plasma donor selection, and the durability of nAb responses, are incompletely understood. METHODS. Potential convalescent plasma donors with virologically documented SARS-CoV-2 infection were tested for serum IgG against SARS-CoV-2 spike protein S1 domain and against nucleoprotein (NP), and for nAb. RESULTS. Among 250 consecutive persons, including 27 (11%) requiring hospitalization, who were studied a median of 67 days since symptom onset, 97% were seropositive on 1 or more assays. Sixty percent of donors had nAb titers >1:80. Correlates of higher nAb titers included older age (adjusted OR [AOR] 1.03 per year of age, 95% CI 1.00-1.06), male sex (AOR 2.08, 95% CI 1.13-3.82), fever during illness (AOR 2.73, 95% CI 1.25-5.97), and disease severity represented by hospitalization (AOR 6.59, 95% CI 1.32-32.96). Receiver operating characteristic analyses of anti-S1 and anti-NP antibody results yielded cutoffs that corresponded well with nAb titers, with the anti-S1 assay being slightly more predictive. nAb titers declined in 37 of 41 paired specimens collected a median of 98 days (range 77-120) apart (P< 0.001). Seven individuals (2.8%) were persistently seronegative and lacked T cell responses. CONCLUSION. nAb titers correlated with COVID-19 severity, age, and sex. SARS-CoV-2 IgG results can serve as useful surrogates for nAb testing. Functional nAb levels declined, and a small proportion of convalescent individuals lacked adaptive immune responses. FUNDING. The project was supported by the Frederick National Laboratory for Cancer Research with support from the NIAID under contract number 75N91019D00024, and was supported by the Fred Hutchinson Joel Meyers Endowment, FastGrants, a New Investigator award from the American Society for Transplantation and Cellular Therapy, and NIH contracts 75N93019C0063, 75N91019D00024, and HHSN272201800013C, and NIH grants T32-AI118690, T32-AI007044, K08-AI119142, and K23-AI140918., Introduction Since its emergence in late 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has spread globally, with devastating effects in many communities. At this time, remdesivir remains the [...]

Abstract Background The mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) gene results in CF. The most common mutation, ΔF508-CFTR, is a temperature-sensitive, trafficking mutant with reduced chloride transport and exaggerated immune response. The ΔF508-CFTR is misfolded, ubiquitinated, and prematurely degraded by proteasome mediated- degradation. We recently demonstrated that selective inhibition of proteasomal pathway by the FDA approved drug PS-341 (pyrazylcarbonyl-Phe-Leuboronate, a.k.a. Velcade or bortezomib) ameliorates the inflammatory pathophysiology of CF cells. This proteasomal drug is an extremely potent, stable, reversible and selective inhibitor of chymotryptic threonine protease-activity. The apprehension in considering the proteasome as a therapeutic target is that proteasome inhibitors may affect proteostasis and consecutive processes. The affect on multiple processes can be mitigated by nanoparticle mediated PS-341 lung-delivery resulting in favorable outcome observed in this study. Results To overcome this challenge, we developed a nano-based approach that uses drug loaded biodegradable nanoparticle (PLGA-PEGPS-341) to provide controlled and sustained drug delivery. The in vitro release kinetics of drug from nanoparticle was quantified by proteasomal activity assay from days 1-7 that showed slow drug release from day 2-7 with maximum inhibition at day 7. For in vivo release kinetics and biodistribution, these drug-loaded nanoparticles were fluorescently labeled, and administered to C57BL6 mice by intranasal route. Whole-body optical imaging of the treated live animals demonstrates efficient delivery of particles to murine lungs, 24 hrs post treatment, followed by biodegradation and release over time, day 1-11. The efficacy of drug release in CF mice (Cftr-/-) lungs was determined by quantifying the changes in proteasomal activity (~2 fold decrease) and ability to rescue the Pseudomonas aeruginosa LPS (Pa-LPS) induced inflammation, which demonstrates the rescue of CF lung disease in murine model. Conclusion We have developed a novel drug delivery system to provide sustained delivery of CF "correctors" and "anti-inflammatories" to the lungs. Moreover, we demonstrate here the therapeutic efficacy of nano-based proteostasis-modulator to rescue Pa-LPS induced CF lung disease.

The environmental, genetic, and/or age-related changes in proteostasis induce inflammation, oxidative stress, and apoptosis. We quantified the correlation of protein expression of critical proteostasis mediators to severity of chronic lung disease using lung tissue samples from control and chronic obstructive pulmonary disease (COPD) subjects (GOLD stage 0-IV) and cigarette smoke (CS)-induced murine model. The human bronchial epithelial cells, HEK-293, and Beas2B cells were used for in vitro experiments to verify the mechanisms. Our data verifies the correlation of higher expression of valosin-containing protein (VCP) retrograde translocation complex (VCP-Rma1-gp78) with severity of emphysema in COPD lung tissues and over-expression of inflammatory, ER stress and apoptotic mediators like NFκB, GADD-153/CHOP, and p-eIF2α. Moreover, subjects with severe emphysema had a higher accumulation of ubiquitinated proteins and deubiquitinating enzyme, UCHL-1, indicating towards the aggregation of misfolded or damaged proteins. The modulation of both protein degradation and synthesis rates by CS-extract substantiates the pathogenetic role of proteostasis-imbalance in emphysema and COPD. We identified that VCP also mediates proteasomal degradation of HDAC2 and Nrf2, as a potential mechanism for increased oxidative stress and corticosteroid resistance in COPD subjects with emphysema. Next, we confirmed that higher VCP expression associates with increased inflammation and apoptosis using in vitro and murine models. Our data clearly shows aberrant proteostasis in COPD subjects with severe emphysema. In addition, we evaluate therapeutic efficacy of salubrinal (ER stress inhibitor) to correct the proteostasis-imbalance based on its ability to control VCP expression and ubiquitin accumulation. Overall, our data demonstrate for the first time the critical role of proteostasis-imbalance in pathogenesis of severe emphysema. [ABSTRACT FROM AUTHOR]

Ocozocoautla de Espinosa virus (OCEV) is a novel, uncultured arenavirus. We found that the OCEV glycoprotein mediates entry into grivet and bat cells through transferrin receptor 1 (TfR1) binding but that OCEV glycoprotein precursor (GPC)-pseudotyped retroviruses poorly entered 53 human cancer cell lines. Interestingly, OCEV and Tacaribe virus could use bat, but not human, TfR1. Replacing three human TfR1 amino acids with their bat ortholog counterparts transformed human TfR1 into an efficient OCEV and Tacaribe virus receptor. [ABSTRACT FROM AUTHOR]

Simian hemorrhagic fever virus (SHFV) causes a severe and almost uniformly fatal viral hemorrhagic fever in Asian macaques but is thought to be nonpathogenic for humans. To date, the SHFV life cycle is almost completely uncharacterized on the molecular level. Here, we describe the first steps of the SHFV life cycle. Our experiments indicate that SHFV enters target cells by lowpH- dependent endocytosis. Dynamin inhibitors, chlorpromazine, methyl-β-cyclodextrin, chloroquine, and concanamycin A dramatically reduced SHFV entry efficiency, whereas the macropinocytosis inhibitors EIPA, blebbistatin, and wortmannin and the caveolin-mediated endocytosis inhibitors nystatin and filipin III had no effect. Furthermore, overexpression and knockout study and electron microscopy results indicate that SHFV entry occurs by a dynamin-dependent clathrin-mediated endocytosislike pathway. Experiments utilizing latrunculin B, cytochalasin B, and cytochalasin D indicate that SHFV does not hijack the actin polymerization pathway. Treatment of target cells with proteases (proteinase K, papain, α-chymotrypsin, and trypsin) abrogated entry, indicating that the SHFV cell surface receptor is a protein. Phospholipases A2 and D had no effect on SHFV entry. Finally, treatment of cells with antibodies targeting CD163, a cell surface molecule identified as an entry factor for the SHFVrelated porcine reproductive and respiratory syndrome virus, diminished SHFV replication, identifying CD163 as an important SHFV entry component. [ABSTRACT FROM AUTHOR]