Biology subjects

Anand, S. P.

Publications and source records attributed to Anand, S. P..

9 recordsLinked to original sources

bioRxiv2021

Structural Basis and Mode of Action for Two Broadly Neutralizing Antibodies Against SARS-CoV-2 Emerging Variants of Concern

Emerging variants of concern for the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can transmit more efficiently and partially evade protective immune responses, thus necessitating continued refinement of antibody therapies and immunogen design. Here we elucidate the structural basis and mode of action for two potent SARS-CoV-2 Spike (S) neutralizing monoclonal antibodies CV3-1 and CV3-25 that remained effective against emerging variants of concern in vitro and in vivo. CV3-1 bound to the (485-GFN-487) loop within the receptor-binding domain (RBD) in the "RBD-up" position and triggered potent shedding of the S1 subunit. In contrast, CV3-25 inhibited membrane fusion by binding to an epitope in the stem helix region of the S2 subunit that is highly conserved among {beta}-coronaviruses. Thus, vaccine immunogen designs that incorporate the conserved regions in RBD and stem helix region are candidates to elicit pan-coronavirus protective immune responses.

microbiology↗

bioRxiv2021

Impact of temperature on the affinity of SARS-CoV-2 Spike for ACE2

The seasonal nature in the outbreaks of respiratory viral infections with increased transmission during low temperatures has been well established. The current COVID-19 pandemic makes no exception, and temperature has been suggested to play a role on the viability and transmissibility of SARS-CoV-2. The receptor binding domain (RBD) of the Spike glycoprotein binds to the angiotensin-converting enzyme 2 (ACE2) to initiate viral fusion. Studying the effect of temperature on the receptor-Spike interaction, we observed a significant and stepwise increase in RBD-ACE2 affinity at low temperatures, resulting in slower dissociation kinetics. This translated into enhanced interaction of the full Spike to ACE2 receptor and higher viral attachment at low temperatures. Interestingly, the RBD N501Y mutation, present in emerging variants of concern (VOCs) that are fueling the pandemic worldwide, bypassed this requirement. This data suggests that the acquisition of N501Y reflects an adaptation to warmer climates, a hypothesis that remains to be tested.

microbiology↗

bioRxiv2021

Live imaging of SARS-CoV-2 infection in mice reveals neutralizing antibodies require Fc function for optimal efficacy

Neutralizing antibodies (NAbs) are effective in treating COVID-19 but the mechanism of immune protection is not fully understood. Here, we applied live bioluminescence imaging (BLI) to monitor the real-time effects of NAb treatment in prophylaxis and therapy of K18-hACE2 mice intranasally infected with SARS-CoV-2-nanoluciferase. We could visualize virus spread sequentially from the nasal cavity to the lungs and thereafter systemically to various organs including the brain, which culminated in death. Highly potent NAbs from a COVID-19 convalescent subject prevented, and also effectively resolved, established infection when administered within three days. In addition to direct Fab-mediated neutralization, Fc effector interactions of NAbs with monocytes, neutrophils and natural killer cells were required to effectively dampen inflammatory responses and limit immunopathology. Our study highlights that both Fab and Fc effector functions of NAbs are essential for optimal in vivo efficacy against SARS-CoV-2.

immunology↗

bioRxiv2021

A single BNT162b2 mRNA dose elicits antibodies with Fc-mediated effector functions and boost pre-existing humoral and T cell responses

The standard dosing of the Pfizer/BioNTech BNT162b2 mRNA vaccine validated in clinical trials includes two doses administered three weeks apart. While the decision by some public health authorities to space the doses because of limiting supply has raised concerns about vaccine efficacy, data indicate that a single dose is up to 90% effective starting 14 days after its administration. We analyzed humoral and T cells responses three weeks after a single dose of this mRNA vaccine. Despite the proven efficacy of the vaccine at this time point, no neutralizing activity were elicited in SARS-CoV-2 naive individuals. However, we detected strong anti-receptor binding domain (RBD) and Spike antibodies with Fc-mediated effector functions and cellular responses dominated by the CD4+ T cell component. A single dose of this mRNA vaccine to individuals previously infected by SARS-CoV-2 boosted all humoral and T cell responses measured, with strong correlations between T helper and antibody immunity. Neutralizing responses were increased in both potency and breadth, with distinctive capacity to neutralize emerging variant strains. Our results highlight the importance of vaccinating uninfected and previously-infected individuals and shed new light into the potential role of Fc-mediated effector functions and T cell responses in vaccine efficacy. They also provide support to spacing the doses of two-vaccine regimens to vaccinate a larger pool of the population in the context of vaccine scarcity against SARS-CoV-2.

immunology↗

bioRxiv2021

Longitudinal analysis of humoral immunity against SARS-CoV-2 Spike in convalescent individuals up to 8 months post-symptom onset

Functional and lasting immune responses to the novel coronavirus (SARS-CoV-2) are currently under intense investigation as antibody titers in plasma have been shown to decline during convalescence. Since the absence of antibodies does not equate to absence of immune memory, we sought to determine the presence of SARS-CoV-2-specific memory B cells in COVID-19 convalescent patients. In this study, we report on the evolution of the overall humoral immune responses on 101 blood samples obtained from 32 COVID-19 convalescent patients between 16 and 233 days post-symptom onset. Our observations indicate that anti-Spike and anti-RBD IgM in plasma decay rapidly, whereas the reduction of IgG is less prominent. Neutralizing activity in convalescent plasma declines rapidly compared to Fc-effector functions. Concomitantly, the frequencies of RBD-specific IgM+ B cells wane significantly when compared to RBD-specific IgG+ B cells, which increase over time, and the number of IgG+ memory B cells which remain stable thereafter for up to 8 months after symptoms onset. With the recent approval of highly effective vaccines for COVID-19, data on the persistence of immune responses are of central importance. Even though overall circulating SARS-CoV-2 Spike-specific antibodies contract over time during convalescence, we demonstrate that RBD-specific B cells increase and persist up to 8 months post symptom onset. We also observe modest increases in RBD-specific IgG+ memory B cells and importantly, detectable IgG and sustained Fc-effector activity in plasma over the 8-month period. Our results add to the current understanding of immune memory following SARS-CoV-2 infection, which is critical for the prevention of secondary infections, vaccine efficacy and herd immunity against COVID-19.

microbiology↗

bioRxiv2020

High-throughput detection of antibodies targeting the SARS-CoV-2 Spike in longitudinal convalescent plasma samples

BackgroundThe SARS-CoV-2 virus is the cause of the ongoing coronavirus disease 2019 (COVID-19) pandemic, infecting millions of people and causing more than a million deaths. The SARS-CoV-2 Spike glycoproteins mediate viral entry and represent the main target for antibody responses. Humoral responses were shown to be important for preventing and controlling infection by coronaviruses. A promising approach to reduce the severity of COVID-19 is the transfusion of convalescent plasma. However, longitudinal studies revealed that the level of antibodies targeting the receptor-binding domain (RBD) of the SARS-CoV-2 Spike declines rapidly after the resolution of the infection. Study Design and MethodsTo extend this observation beyond the RBD domain, we performed a longitudinal analysis of the persistence of antibodies targeting the full-length SARS-CoV-2 Spike in the plasma from 15 convalescent donors. We generated a 293T cell line constitutively expressing the SARS-CoV-2 Spike and used it to develop a high-throughput flow cytometry-based assay to detect SARS-CoV-2 Spike specific antibodies in the plasma of convalescent donors. Results and ConclusionWe found that the level of antibodies targeting the full-length SARS-CoV-2 Spike declines gradually after the resolution of the infection. This decline was not related to the number of donations, but strongly correlated with the decline of RBD-specific antibodies and the number of days post-symptom onset. These findings help to better understand the decline of humoral responses against the SARS-CoV-2 Spike and provide important information on when to collect plasma after recovery from active infection for convalescent plasma transfusion.

microbiology↗

bioRxiv2020

Real-time conformational dynamics of SARS-CoV-2 spikes on virus particles

SARS-CoV-2 spike (S) mediates entry into cells and is critical for vaccine development against COVID-19. Structural studies have revealed distinct conformations of S, but real-time information that connects these structures, is lacking. Here we apply single-molecule Forster Resonance Energy Transfer (smFRET) imaging to observe conformational dynamics of S on virus particles. Virus-associated S dynamically samples at least four distinct conformational states. In response to hACE2, S opens sequentially into the hACE2-bound S conformation through at least one on-path intermediate. Conformational preferences of convalescent plasma and antibodies suggest mechanisms of neutralization involving either competition with hACE2 for binding to RBD or allosteric interference with conformational changes required for entry. Our findings inform on mechanisms of S recognition and conformations for immunogen design.

microbiology↗

bioRxiv2020

Interaction of human ACE2 to membrane-bound SARS-CoV-1 and SARS-CoV-2 S glycoproteins

A novel severe acute respiratory (SARS)-like coronavirus (SARS-CoV-2) is responsible for the current global coronavirus disease 2019 (COVID-19) pandemic, infecting millions of people and causing hundreds of thousands of deaths. The viral entry of SARS-CoV-2 depends on an interaction between the receptor binding domain of its trimeric Spike glycoprotein and the human angiotensin converting enzyme 2 (ACE2) receptor. A better understanding of the Spike/ACE2 interaction is still required to design anti-SARS-CoV-2 therapeutics. Here, we investigated the degree of cooperativity of ACE2 within both the SARS-CoV-2 and the closely related SARS-CoV-1 membrane-bound S glycoproteins. We show that there exist differential inter-protomer conformational transitions between both Spike trimers. Interestingly, the SARS-CoV-2 spike exhibits a positive cooperativity for monomeric soluble ACE2 binding when compared to the SARS-CoV-1 spike, which might have more structural restrains. Our findings can be of importance in the development of therapeutics that block the Spike/ACE2 interaction.

microbiology↗

bioRxiv2020

Cross-sectional evaluation of humoral responses against SARS-CoV-2 Spike

The SARS-CoV-2 virus is responsible for the current worldwide coronavirus disease 2019 (COVID-19) pandemic, infecting millions of people and causing hundreds of thousands of deaths. The Spike glycoprotein of SARS-CoV-2 mediates viral entry and is the main target for neutralizing antibodies. Understanding the antibody response directed against SARS-CoV-2 is crucial for the development of vaccine, therapeutic and public health interventions. Here we performed a cross-sectional study on 106 SARS-CoV-2-infected individuals to evaluate humoral responses against the SARS-CoV-2 Spike. The vast majority of infected individuals elicited anti-Spike antibodies within 2 weeks after the onset of symptoms. The levels of receptor-binding domain (RBD)-specific IgG persisted overtime, while the levels of anti-RBD IgM decreased after symptoms resolution. Some of the elicited antibodies cross-reacted with other human coronaviruses in a genus-restrictive manner. While most of individuals developed neutralizing antibodies within the first two weeks of infection, the level of neutralizing activity was significantly decreased over time. Our results highlight the importance of studying the persistence of neutralizing activity upon natural SARS-CoV-2 infection.

microbiology↗