Signal Transduction and Targeted Therapy

Neutralizing monoclonal antibodies (NAbs) with prophylactic and therapeutic efficacy have demonstrated fundamental importance in the control of SARS-CoV-2 transmission. However, their wide application has been largely limited by high cost and inconvenience in administration. Here, we developed an intranasal spray containing two synergetic human NAbs that could broadly neutralize the emerging Omicron variants in vitro. A unique synergetic neutralizing mechanism was identified that the two NAbs bound to exclusive epitopes on the RBD and structurally compensate each other in blocking the Spike-ACE2 interaction. Importantly, when given at low dosages for three consecutive days through the intranasal mucosal route, this cocktail showed significant improvement in the emergency preventive and therapeutic effects in hamsters challenged with authentic Omicron BA.1. Further, we performed an investigator-initiated trail in healthy volunteers (ChiCTR2200066525) to study the safety and pharmacokinetics of the antibody cocktail administrated as nasal spray. The nasal spray is generally safe and well tolerated without treatment related severe abnormal effects. The antibody cocktail nasal spray demonstrated nasal concentrations higher than the IC90 of neutralization activity against Omicron BA.4/5 even at 24 hours post dosing. Furthermore, nasal samples from the study subjects demonstrated potent neutralization activity against Omicron BA.4/5 in an ex vivo pseudovirus neutralization assay. Together, we provide a novel approach for NAb regimens, a potentially highly effective product with broad applicable perspective in depressing the infection risk of new epidemic variant and ameliorating the heavy medical burden of hospital. One Sentence SummaryAn intranasal spray of two synergetic antibodies cocktail neutralizing Omicron BA.4/5 and an initial clinical evaluation in healthy volunteers.

The adaptive immunity that protects patients from coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is not well characterized. In particular, the asymptomatic patients have been found to induce weak and transient SARS-CoV-2 antibody responses, but the underlying mechanisms remain unknown; meanwhile, the protective immunity that guide the recovery of these asymptomatic patients is also not well studied. Here, we characterized SARS-CoV-2-specific B-cell and T-cell responses in 10 asymptomatic patients and 49 patients with other disease severity (mild, n = 10, moderate, n = 32, severe, n = 7) and found that asymptomatic or mild symptomatic patients failed to mount virus-specific germinal center (GC) B cell responses that result in robust and long-term humoral immunity, assessed by GC response indicators including follicular helper T (TFH) cell and memory B cell responses as well as serum CXCL13 levels. Alternatively, these patients mounted potent virus-specific TH1 and CD8+ T cell responses. In sharp contrast, patients of moderate or severe disease induced vigorous virus-specific GC B cell responses and associated TFH responses; however, the virus-specific TH1 and CD8+ T cells were minimally induced in these patients. These results therefore uncovered the protective immunity in COVID-19 patients and revealed the strikingly dichotomous and incomplete adaptive immunity in COVID-19 patients with different disease severity, providing important insights into rational design of COVID-19 vaccines.

There are currently no specific vaccine or drugs proven to be effective against COVID-19. Traditional Chinese herbal medicine has been integrated into the official therapeutic protocol against COVID-19 in China. Qing Fei Pai Du Tang (QFPDT) is a Chinese multi-herbal formula newly developed and specifically optimized for the treatment of COVID-19. Therapeutic administration of QFPDT resulted in an improved cure rate in mild to critically-ill patients. However, the immunological mechanism for the efficacy of QFPDT has been poorly understood. Furthermore, the feasibility of prophylactic use in uninfected individuals remain unconfirmed. We thus examined whether the administration of QFPDT at a dose lower than recommended for therapeutic use alters hematological and/or immunological measures in healthy individuals. We found that QFPDT elevates the plasma levels of IL-1{beta}, IL-18, TNF-, and IL-8, which are key mediators of acute inflammatory responses to ssRNA viruses. No apparent adverse effects were observed during the trial. Our finding suggests that the pharmacological action of QFPDT is associated with the upregulation of a subset of proinflammatory cytokines despite its clinical benefits for COVID-19 patients. We should therefore be careful in its prophylactic use in uninfected individuals until we have a better understanding of the immunopharmacological action of QFPDT through further clinical studies with larger cohorts.

The comparison between traditional Chinese medicine Jinzhen Oral Liquid (JZOL) and western medicine in treating children with acute bronchitis (AB) showed encouraging outcomes. This trial evaluated the efficacy and safety of the JZOL for improving cough and expectoration in children with AB. 480 children were randomly assigned to take JZOL or Ambroxol Hydrochloride and Clenbuterol Hydrochloride Oral Solution for 7 days. The primary outcome was time-to-cough resolution. The median time-to-cough resolution in both groups was 5.0 days and the antitussive onset median time was only 1 day. This head to head randomized controlled trial showed that JZOL was not inferior to cough suppressant and phlegm resolving western medicine in treating cough and sputum and could comprehensively treat respiratory and systemic discomfort symptoms. Combined with clinical trials, the mechanism of JZOL against AB was uncovered by network target analysis, it was found that the pathways in TRP channels like IL-1{beta}/IL1R/TRPV1/TRPA1, NGF/TrkA/TRPV1/TRPA1 and PGE2/EP/PKA/TRPV1/TRPA1 might play important roles. Animal experiments further confirmed that inflammation and immune regulatory effect of JZOL in the treatment of AB were of vital importance and TRP channels was the key mechanism of action.

IntroductionCOVID-19 has affected many people worldwide. Effective treatments have been reported using medicinal plants which their multi functionality may beneficial. Considering the importance of innate immune response and pathologic effect of cytokine storm in COVID-19 patients, this study examined the effect of Redcurrant (Ribes rubrum) fruit powder capsules on inflammatory cytokines of IL-6 and TNF as primary objectives. Moreover, its effect on the COVID-19 patients recovery symptoms was evaluated. MethodsWe analyzed the clinical effect on recovery symptoms, hematological parameters, and the serum levels of interlucin-6 and TNF in the placebo-controlled and treated patients with Redcurrant fruit powder. ResultsRedcurrant fruit powder capsules significantly decreased pro-inflammatory cytokines of IL-6 and TNF-, and reduced the clinical recovery time of COVID-19 and fatigue (from 7.3 {+/-} 2.09 to 2.9 {+/-} 1.28 days) as well as cough (from 5.67 {+/-} 2.4 to 2.4 {+/-} 1.7) days compared to the control group. ConclusionRedcurrant fruit powder might be a promising herbal treatment of COVID-19, and since no considerable side effect is observed in short-term usage, it might be beneficial to start treatment as early as possible in suspected cases of COVID-19.

BackgroundPatients with sepsis-induced acute lung injury (ALI)/acute respiratory distress syndrome (ARDS) commonly suffer from severe pulmonary thrombosis, but clinical trials of anti-coagulant therapies in sepsis and ARDS patients have failed. ARDS patients with thrombocytopenia also exhibit increased mortality, and widespread pulmonary thrombosis is often seen in coronavirus disease 2019 (COVID-19) ARDS patients. MethodsEmploying different amounts of microbeads to induce various levels of pulmonary thrombosis. Acute lung injury was induced by either lipopolysaccharide i.p. or cecal ligation and puncture. Endothelial cell (EC)-targeted nanoparticle coupled with CDH5 promoter was employed to delivery plasmid DNA expressing the CRISPR/Cas9 system for EC-specific gene knockout or expressing Alox15 for EC-specific overexpression. Additionally, thrombocytopenia was induced by genetic depletion of platelets using DTRPf4Cre mice by breeding Pf4Cre mice into the genetic background of DTR mice. ResultsWe show that while severe pulmonary thrombosis or thrombocytopenia augments sepsis-induced ALI, the induction of mild pulmonary thrombosis conversely reduces endothelial cell (EC) apoptosis, ALI, and mortality via sustained expression of endothelial arachidonate 15-lipoxygenase (Alox15). Endothelial Alox15 knockout via EC-targeted nanoparticle delivery of CRISPR/Cas9 plasmid DNA in adult mice abolished the protective impact of mild lung thrombosis. Conversely, overexpression of endothelial Alox15 inhibited the increases in ALI caused by severe pulmonary thrombosis. The clinical relevance of the findings was validated by the observation of reduced ALOX15-expressing ECs in lung autopsy samples of ARDS patients. Additionally, restoration of pulmonary thrombosis in thrombocytopenic mice also normalized endotoxemia-induced ALI. ConclusionWe have demonstrated that moderate levels of thrombosis protect against sepsis-induced inflammatory lung injury via endothelial Alox15. Overexpression of Alox5 inhibits severe pulmonary thrombosis-induced increase of ALI. Thus, activation of ALOX15 signaling represents a promising therapeutic strategy for treatment of ARDS, especially in sub-populations of patients with thrombocytopenia and/or severe pulmonary thrombosis.

An excessive immune response contributes to SARS-CoV, MERS-CoV and SARS-CoV-2 pathogenesis and lethality, but the mechanism remains unclear. In this study, the N proteins of SARS-CoV, MERS-CoV and SARS-CoV-2 were found to bind to MASP-2, the key serine protease in the lectin pathway of complement activation, resulting in aberrant complement activation and aggravated inflammatory lung injury. Either blocking the N protein:MASP-2 interaction or suppressing complement activation can significantly alleviate N protein-induced complement hyper-activation and lung injury in vitro and in vivo. Complement hyper-activation was also observed in COVID-19 patients, and a promising suppressive effect was observed when the deteriorating patients were treated with anti-C5a monoclonal antibody. Complement suppression may represent a common therapeutic approach for pneumonia induced by these highly pathogenic coronaviruses. One Sentence SummaryThe lectin pathway of complement activation is a promising target for the treatment of highly pathogenic coronavirus induced pneumonia.

LEAF-4L6715 is a liposomal formulation encapsulating transcrocetin (TC) developed to enhance the diffusion of oxygen in the body. Here, we report the final results of the phase I/II clinical trial (NCT04378920; EUDRACT2020-001393-30) initiated to identify an optimal regimen and to assess the activity of TC in the context of acute respiratory distress syndrome (ARDS). More specifically, LEAF-4L6715 was developed to treat patients with ARDS due to severe SARS-CoV-2 infection who have a ratio of partial arterial pressure to inspired fraction of oxygen (PaO2/FiO2 ratio) <200 treated with artificial ventilation support in an intensive care unit. A total of 37 patients were treated (across 6 dosing cohorts) with LEAF-4L6715 given as an intravenous infusion for over 90 minutes. The dose of LEAF-4L6715 was increased until the transaminase levels were elevated and 4 grade 3 events occurred among 8 patients. The recommended dosage was determined to be a fixed concentration of 300 mg administered every 12 hours. An improvement in the PaO2/FiO2 ratio and SOFA score was observed. The overall 28-day survival rate of 81%. This study identified the recommended dose for LEAF 4L6715 and the dose-limiting toxicity and showed an overall favorable risk/benefit profile. These preliminary findings are promising for the activity of LEAF-4L6715 but will require confirmation in a randomized phase III trial.

BackgroundCytokine release syndrome (CRS) is a potentially life-threatening complication of chimeric antigen receptor T (CAR-T) cell therapy. Recent studies indicated critical roles of macrophages and monocytes in CAR-T induced CRS. Here, we report rapid dissipation of CAR-T induced CRS in two patients after receiving Tripterygium glycosides (TG). Effects of triptolide, the major active component of TG, on macrophages and monocytes were examined in animal models. MethodsTwo patients with CRS after CAR-T cell therapy (for hematological malignancy) received TG (50 mg, p.o.). Flow cytometry analysis and single cell RNA sequencing (scRNAseq) were conducted to examine the effects of TG on immune cells. Potential effects of triptolide were also examined ex vivo using patient-derived monocytes, as well as in mice. FindingsRapid alleviation of fever and cytokine storm was observed within 72 hours after TG treatment. Blood concentration of triptolide ranged from 21 to 154 ng/mL during treatment. Flow cytometry and scRNAseq showed selective depletion of monocytes with minimal impact on CAR-T cells in both patients. In ex vivo experiments with patient-derived monocytes, triptolide dramatically inhibited the synthesis of pro-inflammatory cytokines (e.g., IL-6, IL-10, and IP-10). Triptolide also rapidly and selectively depleted peritoneal concanavalin A activated macrophages and monocytes in mice. InterpretationTG could be a promising treatment for CAR-T induced CRS, as well as other diseases with similar mechanisms, e.g., hemophagocytic lymphohistiocytosis and COVID-19. Our preliminary findings require further verification with properly designed clinical trials.

BackgroundNovaferon, a novel protein drug approved for the treatment of chronic hepatitis B in China, exhibits potent antiviral activities. We aimed to determine the anti-SARS-CoV-2 effects of Novaferon in vitro, and conducted a randomized, open-label, parallel group study to explore the antiviral effects of Novaferon for COVID-19. MethodsIn laboratory, the inhibition of Novaferon on viral replication in cells infected with SARS-CoV-2, and on SARS-CoV-2 entry into healthy cells was determined. Antiviral effects of Novaferon were evaluated in COVID-19 patients with treatment of Novaferon, Novaferon plus Lopinavir/Ritonavir, or Lopinavir/Ritonavir. The primary endpoint was the SARS-CoV-2 clearance rates on day 6 of treatment, and the secondary endpoint was the time to the SARS-CoV-2 clearance in COVID-19 patients ResultsNovaferon inhibited the viral replication in infected cells (EC50=1.02 ng/ml), and protected healthy cells from SARS-CoV-2 infection (EC50=0.1 ng/ml). Results from the 89 enrolled COVID-19 patients showed that both Novaferon and Novaferon plus Lopinavir/Ritonavir groups had significantly higher SARS-CoV-2 clearance rates on day 6 than the Lopinavir/Ritonavir group (50.0% vs.24.1%, p = 0.0400, and 60.0% vs.24.1%, p = 0.0053). Median time to SARS-CoV-2 clearance were 6 days, 6 days, and 9 days for three groups respectively, suggesting a 3-dayreduction of time to SARS-CoV-2 clearance in both Novaferon and Novaferon plus Lopinavir/Ritonavir groups compared with Lopinavir/Ritonavir group. ConclusionsNovaferon exhibited anti-SARS-CoV-2 effects in vitro and in COVID-19 patients. These data justified the further evaluation of Novaferon.

The human coronavirus HCoV-19 infection can cause acute respiratory distress syndrome (ARDS), hypercoagulability, hypertension, extrapulmonary multiorgan dysfunction. Effective antiviral and anti-coagulation agents with safe clinical profiles are urgently needed to improve the overall prognosis. We screened an FDA approved drug library and found that an anticoagulant agent dipyridamole (DIP) suppressed HCoV-19 replication at an EC50 of 100 nM in vitro. It also elicited potent type I interferon responses and ameliorated lung pathology in a viral pneumonia model. In analysis of twelve HCoV-19 infected patients with prophylactic anti-coagulation therapy, we found that DIP supplementation was associated with significantly increased platelet and lymphocyte counts and decreased D-dimer levels in comparison to control patients. Two weeks after initiation of DIP treatment, 3 of the 6 severe cases (60%) and all 4 of the mild cases (100%) were discharged from the hospital. One critically ill patient with extremely high levels of D-dimer and lymphopenia at the time of receiving DIP passed away. All other patients were in clinical remission. In summary, HCoV-19 infected patients could potentially benefit from DIP adjunctive therapy by reducing viral replication, suppressing hypercoagulability and enhancing immune recovery. Larger scale clinical trials of DIP are needed to validate these therapeutic effects.

IntroductionThe Covid-19 pandemic, caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), has triggered a serious global health crisis, resulting in millions of reported deaths since its initial identification in China in November 2019. The global disparities in immunization access emphasize the urgent need for ongoing research into therapeutic interventions. This study focuses on the potential use of molecular dihydrogen (H2) inhalation as an adjunctive treatment for Covid-19. H2 therapy shows promise in inhibiting intracellular signaling pathways associated with inflammation, particularly when administered early in conjunction with nasal oxygen therapy. MethodsThis Phase I study, characterized by an open-label, prospective, monocentric, and single ascending dose design, seeks to assess the safety and tolerability of the procedure in individuals with confirmed SARS-CoV-2 infection. Employing a 3+3 design, the study includes three exposure durations (target durations): 1 day (D1), 3 days (D2), and 6 days (D3). ResultsWe concluded that the Maximum Tolerated Duration is at least three days. Every patient showed clinical improvement and excellent tolerance to H2 therapy. Discussion/conclusionTo the best of our knowledge, this phase 1 clinical trial is the first to establish the safety of inhaling a mixture of H2 (3.6%) and N2 (96.4%) in hospitalized Covid-19 patients. The original device and method employed ensure the absence of explosion risk. The encouraging outcomes observed in the 12 patients included in the study justify further exploration through larger, controlled clinical trials. QuestionWhat is the Maximum Tolerated Duration of inhalation for a gaseous mixture including 3.6% of molecular dihydrogen in moderate COVID-19 patients? FindingsThe Maximum Tolerated Duration is at least three days. All patients showed clinical improvement and excellent tolerance to H2 therapy. To the best of our knowledge, this phase 1 clinical trial is the first to establish the safety of inhaling a mixture of H2 (3.6%) and N2 (96.4%) in hospitalized Covid-19 patients. MeaningA gaseous mixture including 3.6% H2, considered in the literature to have promising anti-inflammatory potential, and presenting no risk of explosion, can be used in patients with moderate COVID 19 for at least three days.

BackgroundAs increasing cases of COVID-19 around world, urgent need for effective COVID-19-specific therapeutic drugs is necessary; therefore, we conducted a pilot randomized-controlled study to evaluate the efficacy of 99mTc-MDP for COVID-19 therapeutic treatment. MethodsA total of 21 mild patients with COVID-19 were enrolled in this pilot RCT from February 2020 through March 2020, and then were assigned, in a 1:1 ratio, into control (11 patients) and 99mTc-MDP group (10 patients). Patients in the control group received routine treatment and patients assigned to the 99mTc-MDP group received a combination of routine treatment and an administration of 99mTc-MDP injection of 5ml/day. Both of the patients in the control and 99mTc-MDP groups were treated for 7 days with the primary end point of CT-based radiological pulmonary changes during 7-day follow-up. FindingsFrom baseline to the day 7, 8 (80%) of 10 mild patients in the 99mTc-MDP group had a significant radiological improvement in lung and a decline in inflammatory infiltration, whereas only 1 (9.1%) of 11 patients in the control group had a radiological improvement in lung. None of the patients in the 99mTc-MDP group had disease progression from mild to severe, as well as an inflammatory cytokine storm, and 2 mild patients (18.2%) in the control group developed severe. During days 7 through 14, the number of patients with radiological improvement in the 99mTc-MDP group remained consistent, and only 1 additional case (22%) in the control group were reported. ConclusionIn this randomized pilot study, 99mTc-MDP had an effective inhibitory effect on the inflammatory disease progression for the therapy of COVID-19, and it can accelerate the absorption of pulmonary inflammation in a short period of time during the process of treatment.

This study aims to develop new molecular structures as potential therapeutic agents against COVID-19, utilizing both in silico and in vitro studies. Potential targets of cepharanthine (CEP) against COVID-19 to reveal its underlying mechanism of action were evaluated using in silico screening experiments. A library of new molecules was docked into the receptor binding domain of the SARS-CoV-2 spike glycoprotein complex with its receptor, human ACE2, to identify promising compounds. Receptor-oriented docking was performed using the most likely macromolecular targets, aimed at inhibiting key viral replication pathways and reducing inflammatory processes in damaged tissues. The hit molecules showed potential inhibition of Mpro and PLpro proteases of SARS-CoV-2, which are involved in viral replication. They also showed a potential inhibitory effect on Janus kinase (Jak3), which mediates intracellular signaling responsible for inflammatory processes. The in vitro study examined the effects of the selected hit molecules on the generation of superoxide anions and the release of elastase in activated neutrophils, which are factors that exacerbate tissue inflammation and worsen the clinical manifestations of COVID-19. It was demonstrated that 2-((5-((4-isopropylphenyl)sulfonyl)-6-oxo-1,6-dihydropyrimidin-2-yl)thio)-N-(3-methoxyphenyl)acetamide (Hit15) inhibited virus infection by 43.0% at 10 M using pseudovirus assay and suppressed fMLF/CB-induced superoxide anion generation and elastase release in human neutrophils with IC50 values 1.43 and 1.28 M, respectively. Hit15 showed promising activity against coronavirus that can be further developed into a therapeutic agent. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=183 SRC="FIGDIR/small/24316825v1_ufig1.gif" ALT="Figure 1"> View larger version (51K): org.highwire.dtl.DTLVardef@13607f2org.highwire.dtl.DTLVardef@43a597org.highwire.dtl.DTLVardef@12c7a63org.highwire.dtl.DTLVardef@11711c7_HPS_FORMAT_FIGEXP M_FIG C_FIG

ObjectiveS-Adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH) are indicators of global transmethylation and may play an important role as markers of severity of COVID-19. MethodsThe levels of plasma SAM and SAH were determined in patients admitted with COVID-19 (n = 56, mean age = 61). Lung injury was identified by computed tomography (CT) in accordance with the CT0-4 classification. ResultsSAM was found to be a potential marker of lung damage risk in COVID-19 patients (SAM > 80 nM; CT3,4 vs. CT 0-2: relative ratio (RR) was 3.0; p = 0.0029). SAM/SAH > 6.0 was also found to be a marker of lung injury (CT2-4 vs. CT0,1: RR = 3.47, p = 0.0004). Interleukin-6 (IL-6) levels were associated with SAM ({rho} = 0.44, p = 0.01) and SAH ({rho} = 0.534, p = 0.001) levels. ConclusionsHigh SAM levels and high methylation index are associated with the risk of lung injury in COVID-19 patients. The association of SAM and SAH with IL-6 indicates an important role of transmethylation in the development of cytokine imbalance in COVID-19 cases.

We used a new strategy to screen cytokines associated with SARS-CoV-2 infection. Cytokines that can classify populations in different states of SARS-CoV-2 infection were first screened in cross-sectional serum samples from 184 subjects by 2 statistical analyses. The resultant cytokines were then analyzed for their interrelationships and fluctuating features in sequential samples from 38 COVID-19 patients. Three cytokines, M-CSF, IL-8 and SCF, which were clustered into 3 different correlation groups and had relatively small fluctuations during SARS-CoV-2 infection, were selected for the construction of a multiclass classification model. This model discriminated healthy individuals and asymptomatic and nonsevere patients with accuracy of 77.4% but was not successful in classifying severe patients. Further searching led to a single cytokine, hepatocyte growth factor (HGF), which classified severe from nonsevere COVID-19 patients with a sensitivity of 84.6% and a specificity of 97.9% under a cutoff value of 1128 pg/ml. The level of this cytokine did not increase in nonsevere patients but was significantly elevated in severe patients. Considering its potent antiinflammatory function, we suggest that HGF might be a new candidate therapy for critical COVID-19. In addition, our new strategy provides not only a rational and effective way to focus on certain cytokine biomarkers for infectious diseases but also a new opportunity to probe the modulation of cytokines in the immune response.

We detected active alveolar regrowth in the lung of a 58-year-old COVID-19 patient who underwent lung transplantation due to severe lung hemorrhage. Specifically, immunohistological and scanning electronic microscopy analyses revealed that alveolar type II epithelial cells (AT2 cells) accumulate in response to viral pneumonia and that these AT2 cells actively proliferate and differentiate into squamous AT1-like alveolar epithelial cells. Thus, our work establishes that alveolar regrowth does occur in post-COVID-19 injury adult human lungs.

BackgroundCurrently, effective vaccines or specific therapeutic agents against COVID-19 are not available. However, in China, traditional Chinese herbal medicines have provided therapeutic benefit to patients with COVID-19. Jinhua Qinggan granule (JHQGG) is a Chinese multi-herbal formula previously developed for the treatment of H1N1 influenza and has been encouraged for patients clinically suspected of COVID-19 during medical observation. However, the immunological mechanism for the efficacy of JHQGG has not been confirmed. ObjectivesWe thus examined whether the administration of JHQGG affects hematological and immunological measures in healthy individuals. MethodWe enrolled 18 healthy volunteers, all of whom tested negative for antibodies to SARS-CoV-2. Peripheral blood samples were collected 1 h after oral administration of JHQGG and subjected to hematological, biochemical, and cytokine tests. ResultsJHQGG rapidly induced a significant decrease in the plasma level of IL-6 and an increase in the plasma level of IFN-{gamma}. ConclusionsOur finding suggests that the therapeutic efficacy of JHQGG against COVID-19 is, in part, associated with its rapid immunomodulatory activity.

Neutralizing antibodies could be antivirals against COVID-19 pandemics. Here, we report the isolation of four human-origin monoclonal antibodies from a convalescent patient in China. All of these isolated antibodies display neutralization abilities in vitro. Two of them (B38 and H4) block the binding between RBD and vial cellular receptor ACE2. Further competition assay indicates that B38 and H4 recognize different epitopes on the RBD, which is ideal for a virus-targeting mAb-pair to avoid immune escape in the future clinical applications. Moreover, therapeutic study on the mouse model validated that these two antibodies can reduce virus titers in the infected mouse lungs. Structure of RBD-B38 complex revealed that most residues on the epitope are overlapped with the RBD-ACE2 binding interface, which explained the blocking efficacy and neutralizing capacity. Our results highlight the promise of antibody-based therapeutics and provide the structural basis of rational vaccine design. One Sentence SummaryA pair of human neutralizing monoclonal antibodies against COVID-19 compete cellular receptor binding but with different epitopes, and with post-exposure viral load reduction activity.

Hermansky-Pudlak syndrome (HPS) is a group of rare genetic disorders, with several subtypes leading to fatal adult-onset pulmonary fibrosis (PF) and no effective treatment. Circulating biomarkers detecting early PF have not been identified. We investigated whether endocannabinoids could serve as blood biomarkers of PF in HPS. We measured endocannabinoids in the serum of HPS, IPF, and healthy human subjects and in a mouse model of HPSPF. Pulmonary function tests (PFT) were correlated with endocannabinoid measurements. In a pale ear mouse model of bleomycin-induced HPSPF, serum endocannabinoid levels were measured with and without treatment with zevaquenabant (MRI-1867), a peripheral CB1R and iNOS antagonist. In three separate cohorts, circulating anandamide levels were increased in HPS-1 patients with or without PF, compared to healthy volunteers. This increase was not observed in IPF patients or in HPS-3 patients, who do not have PF. Circulating anandamide (AEA) levels were negatively correlated with PFT. Furthermore, a longitudinal study over the course of 5-14 years with HPS-1 patients indicated that circulating AEA levels begin to increase with the fibrotic lung process even at the subclinical stages of HPSPF. In pale ear mice with bleomycin-induced HpsPF, serum AEA levels were significantly increased in the earliest stages of PF and remained elevated at a later fibrotic stage. Zevaquenabant treatment reduced the increased AEA levels and attenuated progression in bleomycin-induced HpsPF. Circulating AEA may be a prognostic blood biomarker for PF in HPS-1 patients. Further studies are indicated to evaluate endocannabinoids as potential surrogate biomarkers in progressive fibrotic lung diseases.