ABSTRACT
Acute pancreatitis (AP) is a devastating disease characterized by an inflammatory disorder of the pancreas. P-selectin glycoprotein ligand-1 (PSGL-1) plays a crucial role in the initial steps of the adhesive at process to inflammatory sites, blockade of PSGL-1 might confer potent anti-inflammatory effects. In this study, we generated two non-human primate derived monoclonal antibodies capable of efficiently targeting human PSGL-1, RH001-6 and RH001-22, which were screened from immunized rhesus macaques. We found that RH001-6, can effectively block the binding of P-selectin to PSGL-1, and abolish the adhesion of leukocytes to endothelial cells in vitro. In vivo, we verified that RH001-6 relieved inflammatory responses and pancreatic injury in both caerulein and l-arginine induced AP models. We also evaluated the safety profile after RH001-6 treatment in mice, and verified that RH001-6 did not cause any significant pathological damages in vivo. Taken together, we developed a novel non-human primate derived PSGL-1 blocking antibody with high-specificity, named RH001-6, which can interrupt the binding of PSGL-1 and P-selectin and attenuate inflammatory responses during AP. Therefore, RH001-6 is highly potential to be further developed into therapeutics against acute inflammatory diseases, such as AP.
ABSTRACT
Objective:To investigate the efficacy of a SARS-CoV-2 recombinant protein vaccine as a booster dose.Methods:A new immunogen, namely RBD-sc-trimer, was designed by tandem repeating of single receptor binding domain (RBD) of SARS-CoV-2 spike (S) protein to mimic the trimeric form of RBD presented by the virus. The RBD-sc-trimer protein was expressed as a His-tagged fusion protein using a baculovirus expression system and purified by nickel affinity column. The purified protein was identified by Western blot. Its in vitro binding activity to human angiotensin converting enzyme 2 (hACE2) was analyzed by ELISA. The immunogenicity of RBD-sc-trimer as well as RBD proteins of other forms including RBD dimer (RBD-Fc), RBD monomer (RBD) and S protein trimer (S trimer) as a booster dose was evaluated in BALB/c mice. Results:In terms of both binding and neutralizing antibodies against SARS-CoV-2, RBD-sc-trimer showed an immunogenicity that was superior to that of RBD-Fc and RBD and close to the level of S trimer. The antibody response induced by RBD-sc-trimer was characterized as Th1-biased. Moreover, it displayed a stronger cross-neutralization activity against SARS-CoV-2 Beta, Delta and Omicron variants. The titer of neutralizing antibody against Omicron induced by RBD-sc-trimer only decreased by 9.1 folds relative to the prototype strain, while the antibody response induced by RBD-Fc and S trimer decreased by 68.4 and 70.8 folds, respectively.Conclusions:The recombinant protein, RBD-sc-trimer, which was capable of eliciting stronger humoral response in mice as a booster dose and showed the superiority in raising cross-reactive antibodies against SARS-CoV-2 variants over non-trimeric RBD forms, should be considered as an optimal immunogen for the development of more effective SARS-CoV-2 vaccines.
ABSTRACT
The spread of SARS-CoV-2 virus in the ongoing global pandemics has led to infections of millions of people and losses of many lives. The rapid, accurate and convenient SARS-CoV-2 virus detection is crucial for controlling and stopping the pandemics. Diagnosis of patients in the early stage infection are so far limited to viral nucleic acid or antigen detection in human nasopharyngeal swab or saliva samples. Here we developed a method for rapid and direct optical measurement of SARS-CoV-2 virus particles in one step nearly without any sample preparation using a spike protein specific nanoplasmonic resonance sensor. We demonstrate that we can detect as few as 30 virus particles in one step within 15 minutes and can quantify the virus concentration linearly in the range of 103 vp/ml to 106 vp/ml. Measurements shown on both generic microplate reader and a handheld smartphone connected device suggest that our low-cost and rapid detection method may be adopted quickly under both regular clinical environment and resource-limited settings.
ABSTRACT
The coronavirus induced disease 19 (COVID-19) caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a worldwide threat to human lives, and neutralizing antibodies present a great therapeutic potential in curing affected patients. We purified more than one thousand memory B cells specific to SARS-CoV-2 S1 or RBD (receptor binding domain) antigens from 11 convalescent COVID-19 patients, and a total of 729 naturally paired heavy and light chain fragments were obtained by single B cell cloning technology. Among these, 178 recombinant monoclonal antibodies were tested positive for antigen binding, and the top 13 binders with Kd below 0.5 nM are all RBD binders. Importantly, all these 13 antibodies could block pseudoviral entry into HEK293T cells overexpressing ACE2, with the best ones showing IC50s around 2-3 nM. We further identified 8 neutralizing antibodies against authentic virus with IC50s within 10 nM. Among these, 414-1 blocked authentic viral entry at IC50 of 1.75 nM and in combination with 105-38 could achieve IC50 as low as 0.45 nM. Meanwhile, we also found that 3 antibodies could cross-react with the SARS-CoV spike protein. Altogether, our study provided a panel of potent human neutralizing antibodies for COVID19 as therapeutics candidates for further development.
