Biology subjects

Rosen, L. E.

Publications and source records attributed to Rosen, L. E..

9 recordsLinked to original sources

bioRxiv2023

Identifying and overcoming the sampling challenges in relative binding free energy calculations of a model protein:protein complex

Relative alchemical binding free energy calculations are routinely used in drug discovery projects to optimize the affinity of small molecules for their drug targets. Alchemical methods can also be used to estimate the impact of amino acid mutations on protein:protein binding affinities, but these calculations can involve sampling challenges due to the complex networks of protein and water interactions frequently present in protein:protein interfaces. We investigate these challenges by extending a GPU-accelerated open-source relative free energy calculation package (Perses) to predict the impact of amino acid mutations on protein:protein binding. Using the well-characterized model system barnase:barstar, we describe analyses for identifying and characterizing sampling problems in protein:protein relative free energy calculations. We find that mutations with sampling problems often involve charge-changes, and inadequate sampling can be attributed to slow degrees of freedom that are mutation-specific. We also explore the accuracy and efficiency of current state-of-the-art approaches--alchemical replica exchange and alchemical replica exchange with solute tempering--for overcoming relevant sampling problems. By employing sufficiently long simulations, we achieve accurate predictions (RMSE 1.61, 95% CI: [1.12, 2.11] kcal/mol), with 86% of estimates within 1 kcal/mol of the experimentally-determined relative binding free energies and 100% of predictions correctly classifying the sign of the changes in binding free energies. Ultimately, we provide a model workflow for applying protein mutation free energy calculations to protein:protein complexes, and importantly, catalog the sampling challenges associated with these types of alchemical transformations. Our free open-source package (Perses) is based on OpenMM and available at https://github.com/choderalab/perses.

biophysics↗

bioRxiv2022

The potent broadly neutralizing antibody VIR-3434 controls Hepatitis B and D Virus infection and reduces HBsAg in humanized mice

Background & AimsChronic hepatitis B is a major global public health problem, and coinfection with hepatitis delta virus (HDV) worsens disease outcome. Here, we describe a hepatitis B virus (HBV) surface antigen (HBsAg)-targeting monoclonal antibody (mAb) with the potential to promote functional cure of chronic hepatitis B and D to address this unmet medical need. MethodsHBsAg-specific mAbs were isolated from memory B cells of HBV vaccinated individuals. In vitro neutralization was determined against HBV and HDV enveloped with HBsAg representing eight HBV genotypes. Human liver-chimeric mice were treated twice weekly with a candidate mAb starting three weeks post HBV inoculation (spreading phase) or during stable HBV or HBV/HDV coinfection (chronic phase). ResultsFrom a panel of human anti-HBs mAbs, VIR-3434 was selected and engineered for pre-clinical development. VIR-3434 targets a putative conserved, conformational epitope within the antigenic loop of HBsAg and neutralized HBV and HDV infection with >12,000-fold higher potency than Hepatitis B Immunoglobulins in vitro. Neutralization was pan-genotypic against strains representative of HBV genotypes A-H. In the spreading phase of HBV infection in human liver-chimeric mice, a parental mAb of VIR-3434 (HBC34) prevented HBV dissemination and intrahepatic HBV RNA and cccDNA increase. In the chronic phase of HBV infection or co-infection with HDV, HBC34 treatment decreased circulating HBsAg by >1 log and HDV RNA by >2 logs. ConclusionsThis in vitro and in vivo characterization identified the potent anti-HBs mAb VIR-3434, which reduces circulating HBsAg and HBV/HDV viremia in human liver-chimeric mice. VIR-3434 is currently in clinical development for treatment of patients with chronic hepatitis B or D. Lay summaryChronic infection with hepatitis B virus places approximately 290 million individuals worldwide at risk for severe liver disease and cancer. Currently available treatments result in low rates of functional cure or require lifelong therapy that does not eliminate the risk of liver disease. We isolated and characterized a potent, human antibody that neutralizes hepatitis B and D viruses and reduces infection in a mouse model. This antibody could provide a new treatment for patients with chronic hepatitis B and D. HighlightsO_LIIdentification of a human mAb VIR-3434 that potently neutralizes HBV and HDV C_LIO_LIVIR-3434 targets a conserved, conformational epitope of the HBsAg antigenic loop C_LIO_LIVIR-3434 treatment blocks intrahepatic HBV spread in human liver-chimeric mice C_LIO_LIVIR-3434 treatment reduces circulating HBsAg and HDV RNA in co-infected mice C_LIO_LIData have enabled clinical development of VIR-3434 against chronic hepatitis B/D C_LI

microbiology↗

bioRxiv2021

Structural basis of SARS-CoV-2 Omicron immune evasion and receptor engagement

The SARS-CoV-2 Omicron variant of concern evades antibody mediated immunity with an unprecedented magnitude due to accumulation of numerous spike mutations. To understand the Omicron antigenic shift, we determined cryo-electron microscopy and X-ray crystal structures of the spike and RBD bound to the broadly neutralizing sarbecovirus monoclonal antibody (mAb) S309 (the parent mAb of sotrovimab) and to the human ACE2 receptor. We provide a structural framework for understanding the marked reduction of binding of all other therapeutic mAbs leading to dampened neutralizing activity. We reveal electrostatic remodeling of the interactions within the spike and those formed between the Omicron RBD and human ACE2, likely explaining enhanced affinity for the host receptor relative to the prototypic virus.

biochemistry↗

bioRxiv2021

Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift

The recently emerged SARS-CoV-2 Omicron variant harbors 37 amino acid substitutions in the spike (S) protein, 15 of which are in the receptor-binding domain (RBD), thereby raising concerns about the effectiveness of available vaccines and antibody therapeutics. Here, we show that the Omicron RBD binds to human ACE2 with enhanced affinity relative to the Wuhan-Hu-1 RBD and acquires binding to mouse ACE2. Severe reductions of plasma neutralizing activity were observed against Omicron compared to the ancestral pseudovirus for vaccinated and convalescent individuals. Most (26 out of 29) receptor-binding motif (RBM)-directed monoclonal antibodies (mAbs) lost in vitro neutralizing activity against Omicron, with only three mAbs, including the ACE2-mimicking S2K146 mAb1, retaining unaltered potency. Furthermore, a fraction of broadly neutralizing sarbecovirus mAbs recognizing antigenic sites outside the RBM, including sotrovimab2, S2X2593 and S2H974, neutralized Omicron. The magnitude of Omicron-mediated immune evasion and the acquisition of binding to mouse ACE2 mark a major SARS-CoV-2 mutational shift. Broadly neutralizing sarbecovirus mAbs recognizing epitopes conserved among SARS-CoV-2 variants and other sarbecoviruses may prove key to controlling the ongoing pandemic and future zoonotic spillovers.

immunology↗

bioRxiv2021

A human antibody that broadly neutralizes betacoronaviruses protects against SARS-CoV-2 by blocking the fusion machinery

The repeated spillovers of {beta}-coronaviruses in humans along with the rapid emergence of SARS-CoV-2 escape variants highlight the need to develop broad coronavirus therapeutics and vaccines. Five monoclonal antibodies (mAbs) were isolated from COVID-19 convalescent individuals and found to cross-react with multiple {beta}-coronavirus spike (S) glycoproteins by targeting the stem helix. One of these mAbs, S2P6, cross-reacts with more than twenty human and animal {beta}-coronavirus S glycoproteins and broadly neutralizes SARS-CoV-2 and pseudotyped viruses from the sarbecovirus, merbecovirus and embecovirus subgenera. Structural and functional studies delineate the molecular basis of S2P6 cross-reactivity and broad neutralization and indicate that this mAb blocks viral entry by inhibiting membrane fusion. S2P6 protects hamsters challenged with SARS-CoV-2 (including the B.1.351 variant of concern) through direct viral neutralization and Fc-mediated effector functions. Serological and B cell repertoire analyses indicate that antibodies targeting the stem helix are found in some convalescent donors and vaccinees but are predominantly of narrow specificity. Germline reversion of the identified cross-reactive mAbs revealed that their unmutated ancestors are specific for the endemic OC43 or HKU1 viruses and acquired enhanced affinity and breadth through somatic mutations. These data demonstrate that conserved epitopes in the coronavirus fusion machinery can be targeted by protective antibodies and provide a framework for structure-guided design of pan-{beta}-coronavirus vaccines eliciting broad protection.

immunology↗

bioRxiv2021

Antibodies to the SARS-CoV-2 receptor-binding domain that maximize breadth and resistance to viral escape

An ideal anti-SARS-CoV-2 antibody would resist viral escape1-3, have activity against diverse SARS-related coronaviruses4-7, and be highly protective through viral neutralization8-11 and effector functions12,13. Understanding how these properties relate to each other and vary across epitopes would aid development of antibody therapeutics and guide vaccine design. Here, we comprehensively characterize escape, breadth, and potency across a panel of SARS-CoV-2 antibodies targeting the receptor-binding domain (RBD), including S3094, the parental antibody of the late-stage clinical antibody VIR-7831. We observe a tradeoff between SARS-CoV-2 in vitro neutralization potency and breadth of binding across SARS-related coronaviruses. Nevertheless, we identify several neutralizing antibodies with exceptional breadth and resistance to escape, including a new antibody (S2H97) that binds with high affinity to all SARS-related coronavirus clades via a unique RBD epitope centered on residue E516. S2H97 and other escape-resistant antibodies have high binding affinity and target functionally constrained RBD residues. We find that antibodies targeting the ACE2 receptor binding motif (RBM) typically have poor breadth and are readily escaped by mutations despite high neutralization potency, but we identify one potent RBM antibody (S2E12) with breadth across sarbecoviruses closely related to SARS-CoV-2 and with a high barrier to viral escape. These data highlight functional diversity among antibodies targeting the RBD and identify epitopes and features to prioritize for antibody and vaccine development against the current and potential future pandemics.

microbiology↗

bioRxiv2021

Structural basis for broad sarbecovirus neutralization by a human monoclonal antibody

The recent emergence of SARS-CoV-2 variants of concern (VOC) and the recurrent spillovers of coronaviruses in the human population highlight the need for broadly neutralizing antibodies that are not affected by the ongoing antigenic drift and that can prevent or treat future zoonotic infections. Here, we describe a human monoclonal antibody (mAb), designated S2x259, recognizing a highly conserved cryptic receptor-binding domain (RBD) epitope and cross-reacting with spikes from all sarbecovirus clades. S2x259 broadly neutralizes spike-mediated entry of SARS-CoV-2 including the B.1.1.7, B.1.351, P.1 and B.1.427/B.1.429 VOC, as well as a wide spectrum of human and zoonotic sarbecoviruses through inhibition of ACE2 binding to the RBD. Furthermore, deep-mutational scanning and in vitro escape selection experiments demonstrate that S2x259 possesses a remarkably high barrier to the emergence of resistance mutants. We show that prophylactic administration of S2x259 protects Syrian hamsters against challenges with the prototypic SARS-CoV-2 and the B.1.351 variant, suggesting this mAb is a promising candidate for the prevention and treatment of emergent VOC and zoonotic infections. Our data unveil a key antigenic site targeted by broadly-neutralizing antibodies and will guide the design of pan-sarbecovirus vaccines.

immunology↗

bioRxiv2021

SARS-CoV-2 immune evasion by variant B.1.427/B.1.429

SARS-CoV-2 entry is mediated by the spike (S) glycoprotein which contains the receptor-binding domain (RBD) and the N-terminal domain (NTD) as the two main targets of neutralizing antibodies (Abs). A novel variant of concern (VOC) named CAL.20C (B.1.427/B.1.429) was originally detected in California and is currently spreading throughout the US and 29 additional countries. It is unclear whether antibody responses to SARS-CoV-2 infection or to the prototypic Wuhan-1 isolate-based vaccines will be impacted by the three B.1.427/B.1.429 S mutations: S13I, W152C and L452R. Here, we assessed neutralizing Ab responses following natural infection or mRNA vaccination using pseudoviruses expressing the wildtype or the B.1.427/B.1.429 S protein. Plasma from vaccinated or convalescent individuals exhibited neutralizing titers, which were reduced 3-6 fold against the B.1.427/B.1.429 variant relative to wildtype pseudoviruses. The RBD L452R mutation reduced or abolished neutralizing activity of 14 out of 35 RBD-specific monoclonal antibodies (mAbs), including three clinical-stage mAbs. Furthermore, we observed a complete loss of B.1.427/B.1.429 neutralization for a panel of mAbs targeting the N-terminal domain due to a large structural rearrangement of the NTD antigenic supersite involving an S13I-mediated shift of the signal peptide cleavage site. These data warrant closer monitoring of signal peptide variants and their involvement in immune evasion and show that Abs directed to the NTD impose a selection pressure driving SARS-CoV-2 viral evolution through conventional and unconventional escape mechanisms.

immunology↗

bioRxiv2020

The circulating SARS-CoV-2 spike variant N439K maintains fitness while evading antibody-mediated immunity

SARS-CoV-2 can mutate to evade immunity, with consequences for the efficacy of emerging vaccines and antibody therapeutics. Herein we demonstrate that the immunodominant SARS-CoV-2 spike (S) receptor binding motif (RBM) is the most divergent region of S, and provide epidemiological, clinical, and molecular characterization of a prevalent RBM variant, N439K. We demonstrate that N439K S protein has enhanced binding affinity to the hACE2 receptor, and that N439K virus has similar clinical outcomes and in vitro replication fitness as compared to wild- type. We observed that the N439K mutation resulted in immune escape from a panel of neutralizing monoclonal antibodies, including one in clinical trials, as well as from polyclonal sera from a sizeable fraction of persons recovered from infection. Immune evasion mutations that maintain virulence and fitness such as N439K can emerge within SARS-CoV-2 S, highlighting the need for ongoing molecular surveillance to guide development and usage of vaccines and therapeutics.

microbiology↗