Biology subjects

Barman, S.

Publications and source records attributed to Barman, S..

At least 19 recordsLinked to original sources

bioRxiv2026

Deep phenomics of human PBMCs using a 31-marker spectral flow cytometry panel

Functional cytokines orchestrate leukocyte-mediated innate and adaptive immune responses within the host following infection or immunization. In contrast, lymphocyte-mediated activation-induced markers (AIMs) are predominantly associated with adaptive immune responses. Regulatory T cells (Tregs) primarily function to balance the immune response by preventing excessive inflammation and maintaining self-tolerance. Currently, conventional intracellular cytokine tracking or cytokine-independent AIMs methodology is available for studying antigen-specific lymphocyte or T cell responses in humans. However, this approach faces challenges due to the difficulty in simultaneously deep immunophenotyping the key functional cytokine signatures, AIMs, and nuclear signature of FoxP3+ Tregs. Using spectral flow cytometry, we developed a 31-parameter panel to profile simultaneously major immune cell subsets (B cells, NK cells, DCs, monocytes) along with T cells, memory phenotypes, functional cytokines by tracking established Th1 markers IFN{gamma}, TNF and IL-2; Th2 markers IL-4/5/13 and the Th17 marker, IL-17, AIMs and FoxP3+ Tregs in human PBMCs in steady state as well as in mitogen stimulated conditions. For AIMs, CD69, CD134 (OX40) and CD137 (4-1BB) are included and validated. We also re-validated our panel to profile antigen specific Th1/2/17 cells or AIMs distribution after peptide stimulation. This optimized multicolor spectral flow panel allows a detailed immune profiling of functional cytokines, AIMs and FoxP3+ Tregs in the T cell compartment from human and might be useful for exploratory analysis of how these functional cytokines or AIMs or Tregs shape host immunity after infection or vaccination. Our panel is highly customizable, allowing researchers to tailor it to their specific requirements.

immunology↗

bioRxiv2026

An immunophenotype-coupled transcriptomic atlas of mouse hematopoietic progenitors

Decoding the bone marrow (BM) hematopoietic progenitor compartment is central to understanding the development of mature immune cells in health and disease. Single-cell multimodal approaches are transforming the analysis of this heterogeneous compartment by simultaneously capturing transcriptomic and proteomic information. Still, integrated studies covering the full spectrum of murine hematopoietic stem and progenitor cells (HSPCs) remain scarce. Here, we report a Cellular Indexing of Transcriptomes and Epitopes by sequencing (CITE-seq) based atlas of 9,281 low-frequency mouse BM HSPCs, profiled using a 123 antibody-derived tag (ADT) panel coupled with genome-wide single-cell transcriptomes. Multimodal analysis resolved 28 progenitor stages across eight blood lineages, each defined by distinctive surface marker and transcription factor (TF) profiles, and provided novel marker combinations to dissect transcriptionally closely related progenitor stages. Classical mouse progenitor surface markers, such as CD117, CD34, and CD115, were detected in accordance with well-established lineage relationships, whereas CD43 and CD48 showed, unexpectedly, broader expression patterns. Neglected surface molecules such as CD27, CD36, CD106, and PIR-A/B are nominated as novel stage-specific markers. ADT versus transcriptome comparison revealed transcription-translation inconsistencies for markers, including CD54 and CD68. A CITE-seq-guided gating strategy translates these population definitions into FACS-compatible panels for prospective isolation and functional interrogation of previously unresolved progenitor stages. Comprehensive TF network analysis delineated lineage- and stage-specific transcriptional programs. This contributes to clarifying unresolved ontogenies and functions, including the lympho-myeloid dual origin of pDCs or rapid neutrophil turnover. This new and comprehensive resource enables exploration of mouse hematopoietic progenitor states driving discoveries in hematology and immunology.

immunology↗

bioRxiv2025

CSF single-cell RNA sequencing reveals clonally expanded CD4+ stem cell-like memory T cells in GAD65-antibody associated neurological syndromes

BackgroundGlutamic acid decarboxylase (GAD) antibody-associated autoimmune neurological syndromes (AINS) are a spectrum of autoimmune-mediated CNS disorders. While antibodies targeting the 65 kDa isoform of GAD are of high diagnostic value, T cell mediated cytotoxicity has been identified as a key component of disease pathogenesis. The precise pathophysiological mechanisms by which the disease is triggered and maintained, however, remain incompletely understood. MethodsWe performed single-cell transcriptome and immune repertoire sequencing (sc-seq) in CSF and blood of 8 anti-GAD65 AINS patients compared to 8 non-inflammatory controls. Monoclonal antibodies (mAbs) were synthesized from B cell receptor (BCR) data to evaluate the B cellular immune response. FindingsWe identified an increase and expansion of activated CD4+ stem cell-like memory T cells (TSCM) in the CSF of anti-GAD65 AINS patients. Expanded T cells showed increased expression of proinflammatory genes. The mAb analysis revealed a high frequency of GAD65-reactive BCRs in the CSF of anti-GAD65 AINS patients with increased somatic hypermutations compared to non-GAD-reactive BCRs and BCRs from controls. ConclusionsSc-seq identified clonally expanded CD4+ TSCM in the CSF of anti-GAD65 AINS patients harboring cytotoxic properties likely contributing to disease pathogenesis. GAD-reactive B cells circulate in the CSF of anti-GAD65 AINS patients further supporting the concept of an antigen-specific intrathecal immune response. Future studies need to clarify the actual pathogenicity of these immune cells and the link between T and B cellular immune mechanisms in the pathogenesis of anti-GAD65 AINS. FundingGerman Research Foundation (ERARE18-202 UltraAIE), German Federal Ministry of Education and Research (CONNECT GENERATE (2.0); 01GM1908A and 01GM2208A).

immunology↗

bioRxiv2025

Deep Phenomics of Pattern Recognition Receptor Agonist Specific Activation of Human Blood

Human whole blood (WB) immunophenotyping may represent the in vivo immunological state with better fidelity than artificially isolated peripheral blood mononuclear cells (PBMCs). We used a deep phenomics modeling approach to elucidate the quantitative differences in major immune cell lineages in WB and PBMC compartments in a steady-state in vitro setting. We studied functional innate immune responses induced by pattern recognition receptor agonist adjuvants (PRRa). Using an optimized 49-parameter CyTOF panel and implementing machine learning (ML) algorithms, we mapped PRRa-mediated CD69, CD40, CD80, CD86 and CCR7 activation at the nodal innate immune subsets level. We also portrayed cellular origin of innate functional chemokine CCL4 and intracellular cytokine IFN{gamma} production. We mapped neutrophils as the primary source of TLR7/8 agonist (TLR7/8a) and STING agonist mediated CCL4 responses in WB. Notably, in the PBMC fraction, where neutrophils are limited, natural killer (NK) cells became the major source of innate CCL4 production. TLR7/8a-mediated IFN{gamma} induction by early NK cells was mapped in PBMCs, which was limited in WB. Considering such distinctions, we hypothesized that deep phenomics employing a clinical sample that has not been manipulated, i.e., WB, may be additive in translating in vitro innate fingerprinting into in vivo biology.

immunology↗

bioRxiv2024

The role of secondary structures of peptide polymers on antimicrobial efficacy and antibiotic potentiation

The rise of antibiotic resistance, biofilm formation, and dormant bacterial populations poses serious global health threats. Synthetic antimicrobial peptide (AMP) mimics offer promising alternatives, though the impact of secondary structures in polymeric AMP mimics on antimicrobial efficacy is underexplored. This study investigates chirality-controlled -peptide polymers (D-PP and DL-PP), synthesized via ring-opening polymerization of allylglycine N-carboxy anhydrides and post-polymerization modification through thiol-ene click chemistry. D-PP adopts a stable helical structure under biomimetic conditions, whereas DL-PP remains random. This helical structure enhanced D-PPs antibacterial and antibiotic potentiation activities, amplifying antibiotic efficacy by 2- to 256-fold across various classes--including tetracyclines, ansamycins, fusidanes, macrolides, cephalosporins, and monobactams--against multidrug-resistant Gram-negative pathogens, while maintaining low hemolytic activity and high protease stability. Mechanistic investigations revealed that D-PP exhibited greater membrane interaction. D-PP and antibiotic combinations eradicated dormant bacterial populations and disrupted biofilms with minimal antimicrobial resistance development. This study paves the way for the rational design of polypeptide-based antimicrobial agents, harnessing chirality and secondary structural features to enhance the efficacy of synthetic antimicrobial peptide mimics.

microbiology↗

bioRxiv2024

A climate and population dependent diffusion model forecasts the spread of Aedes Albopictus mosquitoes in Europe

Vectors of Dengue, Chikungunya, Zika, and Yellow Fever are emerging in new areas, posing increasing public health risks. Aedes albopictus, a key vector for these diseases, is expanding its range beyond its tropical and subtropical origins, driven by suitable climate, population mobility, trade, and urbanization. Since its introduction to Europe, Ae. albopictus has rapidly spread and triggered recurrent outbreaks. Past model attempts have handled vector suitability and vector introduction as independent drivers. Here we develop a novel, highly predictive spatio-temporal vector diffusion model based on minimum temperature, median temperature, relative humidity and human population as predictors. The model predicts areas of presence or absence with an accuracy of 99% and 79% for new established vector populations. The model explains how short- and long-range spread of Ae. albopictus interacts with vector suitability. These results show that the expansion of Ae. albopictus in Europe is predictable and closely linked to climate suitability, and human population. The new model integrates in one simultaneous model framework the climate and mobility drivers, providing a better basis for anticipating future outbreaks in situations of dependent interacting co-drivers. Significance statementThe Tiger mosquito, Aedes albopictus, which spreads diseases like Dengue, Chikungunya, Zika and Yellow fever, has accelerated its presence in Europe over the past decades. This poses a public health risk as Europe face an upsurge of autochthonous arbovirus transmission events. The change is partly due to conducive environments, climate change and human mobility and trade. We develop a novel model explaining the spread of the mosquito over the years 2010-2023, observing a change in recorded mosquito presence areas from 138 regions in 2010 to 537 regions in 2023. The model incorporates patterns in both space and time, capturing with high accuracy how the mosquito spreads from region to region based on climate suitability, geographical diffusion and human population density.

molecular biology↗

bioRxiv2024

Olive phenolic compounds, potent tankyrase 1 inhibitor exhibit anti colon cancer effects by blocking Wnt/β-catenin pathway

Colon cancer (CC) needs special attention to develop novel targets and therapeutic approaches, as the total number of morbidity and mortality is growing rapidly. Although Olive has been long reported for its anti-CC activities but the mode is not completely understood yet. Here, we targeted tankysare 1 (TNKS), an important mediator of the Wnt/{beta}-catenin pathway, crucial in colon carcinogenesis. Bioinformatics analysis revealed that the mutation and copy number alterations of TNKS is found to be 12% whereas the proteins expression of TNKS is moderate to high in tumor groups vs normal in CC datasets. On the more, TNKS and its positively associated genes are found to be concomitant with several carcinogenesis related biological processes and signaling pathways. Further, three olive phytochemicals (apigenin, luteolin, and quercetin), which were shortlisted by employing filters like drug likeness and toxicity screening, molecular docking and finally molecular dynamics simulations, can target tankyrase 1. These ployphenols demonstrated cytotoxic effect in CC (HT-29) cells either alone or in combination. Further, these compounds successfully reduced {beta}-catenin level and its target genes (CCND1, CDK4, and MYC). Overall, inhibition of the Wnt/{beta}-catenin pathway by targeting tankyrase 1 was revealed as one of the anti-CC mechanisms of olive polyphenols.

cancer biology↗

bioRxiv2024

Immune-profiling of T helper 1 (Th1), Th2 and Th17 signatures in murine splenocytes by targeting intracellular cytokines

Functional cytokines shape both innate and adaptive immune responses in the host after infection or immunization. Deep immunophenotyping of the key functional cytokine signatures associated with T cells in murine lymphoid tissue, especially in the spleen, is challenging. Using spectral flow cytometry, we developed a 17-parameter panel to profile major immune cell subsets along with T cells, memory phenotypes and functional cytokines in murine splenocytes in steady state as well as in stimulated conditions. This panel dissects the memory T cell compartment via CD62L and CD44 expression after mitogen stimulation. To profile T helper (Th) cells distribution after mitogen stimulation, established Th1 markers IFN{gamma}, TNF and IL-2; Th2 markers IL-4/5 and the Th17 marker, IL-17, are included. This optimized multicolor spectral flow panel allows a detailed immune profiling of functional cytokines in the murine T cell compartment and might be useful for exploratory analysis of how these functional cytokines shape host immunity after infection or vaccination. Our panel could be easily modified, if researchers wish to tailor the panel to their specific needs.

immunology↗

bioRxiv2023

ATOM-1: A Foundation Model for RNA Structure and Function Built on Chemical Mapping Data

RNA-based medicines and RNA-targeting drugs are emerging as promising new approaches for treating disease. Optimizing these therapeutics by naive experimental screening is a time-consuming and expensive process, while rational design requires an accurate understanding of the structure and function of RNA. To address this design challenge, we present ATOM-1, the first RNA foundation model trained on chemical mapping data, enabled by data collection strategies purposely developed for machine learning training. Using small probe neural networks on top of ATOM-1 embeddings, we demonstrate that this model has developed rich internal representations of RNA. Trained on limited amounts of additional data, these small networks achieve state-of-the-art accuracy on key RNA prediction tasks, suggesting that this approach can enable the design of therapies across the RNA landscape.

bioinformatics↗

bioRxiv2022

Reduced SARS-CoV-2 mRNA vaccine immunogenicity and protection in mice with diet-induced obesity and insulin resistance.

BackgroundObesity and Type 2 Diabetes Mellitus (T2DM) are associated with an increased risk of severe outcomes from infectious diseases, including COVID-19. These conditions are also associated with distinct responses to immunization, including an impaired response to widely used SARS-CoV-2 mRNA vaccines. ObjectiveTo establish a connection between reduced immunization efficacy via modeling the effects of metabolic diseases on vaccine immunogenicity that is essential for the development of more effective vaccines for this distinct vulnerable population. MethodsWe utilized a murine model of diet-induced obesity and insulin resistance to model the effects of comorbid T2DM and obesity on vaccine immunogenicity and protection. ResultsMice fed a high-fat diet (HFD) developed obesity, hyperinsulinemia, and glucose intolerance. Relative to mice fed a normal diet (ND), HFD mice vaccinated with a SARS-CoV-2 mRNA vaccine exhibited significantly lower anti-spike IgG titers, predominantly in the IgG2c subclass, associated with a lower type 1 response, along with a 3.83-fold decrease in neutralizing titers. Furthermore, enhanced vaccine-induced spike-specific CD8+ T cell activation and protection from lung infection against SARS-CoV-2 challenge were seen only in ND mice but not in HFD mice. ConclusionWe demonstrate impaired immunity following SARS-CoV-2 mRNA immunization in a murine model of comorbid T2DM and obesity, supporting the need for further research into the basis for impaired anti-SARS-CoV-2 immunity in T2DM and investigation of novel approaches to enhance vaccine immunogenicity among those with metabolic diseases. Capsule summaryObesity and type 2 diabetes impair SARS-CoV-2 mRNA vaccine efficacy in a murine model.

immunology↗

bioRxiv2022

Identification of novel natural product inhibitors of BRD4 using high throughput virtual screening and MD simulation

Bromodomains are evolutionarily conserved structural motifs that recognize acetylated lysine residues on histone tails. They play a crucial role in shaping chromatin architecture and regulating gene expression in various biological processes. Mutations in bromodomains containing proteins leads to multiple human diseases, which makes them attractive target for therapeutic intervention. Extensive studies have been done on BRD4 as a target for several cancers, such as Acute Myeloid Leukemia (AML) and Burkitt Lymphoma. Several potential inhibitors have been identified against the BRD4 bromodomain. However, most of these inhibitors have drawbacks such as nonspecificity and toxicity, decreasing their appeal and necessitating the search for novel non-toxic inhibitors. This study aims to address this need by virtually screening natural compounds from the NPASS database against the Kac binding site of BRD4-BD1 using high throughput molecular docking followed by similarity clustering, pharmacokinetic screening, MD simulation, and MM-PBSA binding free energy calculations. Using this approach, we identified five natural product inhibitors having a similar or better binding affinity to the BRD4 bromodomain compared to JQ1 (previously reported inhibitor of BRD4). Further systematic analysis of these inhibitors resulted in the top three hits: NPC268484 (Palodesangren-B), NPC295021 (Candidine), and NPC313112 (Buxifoliadine-D). Collectively, our in silico results identified some promising natural products that have the potential to act as potent BRD4-BD1 inhibitors and can be considered for further validation through future in vitro and in vivo studies.

bioinformatics↗

bioRxiv2022

Shaping neonatal immunization by tuning delivery of synergistic adjuvants via nanocarriers

Adjuvanted nanocarrier-based vaccines hold substantial potential for applications in novel early-life immunization strategies. Here, via mouse and human age-specific in vitro modelling, we identified the combination of a small molecule STING agonist (2'3'-cyclic GMP-AMP, cGAMP) and a TLR7/8 agonist (CL075) to drive synergistic activation of neonatal dendritic cells and precision CD4 Th cell expansion via the IL-12/IFN{gamma} axis. We further demonstrate that vaccination of neonatal mice with quadrivalent influenza recombinant hemagglutinin (rHA) and an admixture of two polymersome (PS) nanocarriers separately encapsulating-cGAMP (cGAMP-PS) and CL075 (CL075-PS) drove robust T helper (Th)1 bias, high frequency of T follicular helper (TFH) cells and germinal center B (GC B) cells along with IgG2c-skewed humoral response in vivo. Dual loaded cGAMP/CL075-PSs did not outperform admixed cGAMP-PSs and CL075-PSs in vivo. These data validate an optimally designed adjuvantation system, via age-selected small molecule synergy and a multi-component nanocarrier formulation, as an effective approach to induce type 1 immune responses in early life.

immunology↗

bioRxiv2022

Molecular insights into the recognition of acetylated histone modifications by the BRPF2 bromodomain

HBO1 (HAT bound to ORC), a member of the MYST family of histone acetyltransferases (HATs), was initially identified as a binding partner of the origin recognition complex (ORC) that acetylates free histone H3, H4, and nucleosomal H3. It functions as a quaternary complex with the BRPF (BRPF1/2/3) scaffolding protein and two accessory proteins, ING4/5 and Eaf6. BRPF2 interaction with HBO1 has been shown to be important for regulating H3K14 acetylation during embryonic development. However, how the BRPF2 directs the HBO1 HAT complex to chromatin to regulate its HAT activity towards nucleosomal substrates remains unclear. Our findings reveal novel interacting partners of the BRPF2 bromodomain that recognizes different acetyllysine residues on the N-terminus of histone H4, H3, and H2A and preferentially binds to H4K5ac, H4K8ac, and H4K5acK12ac modifications. Further, mutational analysis of BRPF2 bromodomain coupled with ITC binding and pull-down assays on the histone substrates identified critical residues responsible for acetyllysine binding. Moreover, the BRPF2 bromodomain could enrich H4K5ac mark-bearing mononucleosomes compared to other acetylated H4 marks. Consistent with this, ChIP-seq analysis revealed that BRPF2 strongly co-localizes with HBO1 at histone H4K5ac and H4K8ac marks near the TSS in the genome. Together, our study provides novel insights into how the histone binding function of the BRPF2 bromodomain directs the recruitment of the HBO1 HAT complex to chromatin to regulate gene expression.

biochemistry↗

bioRxiv2022

Engineering SARS-CoV-2 neutralizing antibodies for increased potency and reduced viral escape

The rapid spread of SARS-CoV-2 variants poses a constant threat of escape from monoclonal antibody and vaccine countermeasures. Mutations in the ACE2 receptor binding site on the surface S protein have been shown to disrupt antibody binding and prevent viral neutralization. Here, we use a directed evolution-based approach to engineer three neutralizing antibodies for enhanced binding to S protein. The engineered antibodies showed increased in vitro functional activity in terms of neutralization potency and/or breadth of neutralization against viral variants. Deep mutational scanning revealed that higher binding affinity reduced the total number of viral escape mutations. Studies in the Syrian hamster model showed two examples where the affinity matured antibody provided superior protection compared to the parental antibody. These data suggest that monoclonal antibodies for anti-viral indications could benefit from in vitro affinity maturation to reduce viral escape pathways and appropriate affinity maturation in vaccine immunization could help resist viral variation.

bioengineering↗

bioRxiv2021

A combination of potently neutralizing monoclonal antibodies isolated from an Indian convalescent donor protects against the SARS-CoV-2 delta variant

Although efficacious vaccines have significantly reduced the morbidity and mortality due to COVID-19, there remains an unmet medical need for treatment options, which monoclonal antibodies (mAbs) can potentially fill. This unmet need is exacerbated by the emergence and spread of SARS-CoV-2 variants of concern (VOCs) that have shown some resistance to vaccine responses. Here we report the isolation of two highly potently neutralizing mAbs (THSC20.HVTR04 and THSC20.HVTR26) from an Indian convalescent donor, that neutralize SARS-CoV-2 VOCs at picomolar concentrations including the delta variant (B.1.617.2). These two mAbs target non-overlapping epitopes on the receptor-binding domain (RBD) of the spike protein thereby preventing the virus attachment to its host receptor, human angiotensin converting enzyme-2 (hACE2). Furthermore, the mAb cocktail demonstrated protection against the Delta variant at low antibody doses when passively administered in the K18 hACE2 transgenic mice model, highlighting their potential as cocktail for prophylactic and therapeutic applications. Developing the capacity to rapidly discover and develop mAbs effective against highly transmissible pathogens like coronaviruses at a local level, especially in a low- and middle-income country (LMIC) such as India, will enable prompt responses to future pandemics as an important component of global pandemic preparedness. HighlightsO_LIIdentification of an Indian convalescent donor prior to emergence of SARS-CoV-2 Delta variant whose plasma demonstrated neutralization breadth across SARS-CoV-2 variants of concern (VOCs). C_LIO_LITwo (THSC20.HVTR04 and THSC20.HVTR26) monoclonal antibodies isolated from peripheral memory B cells potently neutralize SARS-CoV-2 VOCs: Alpha, Beta, Gamma, Delta and VOIs: Kappa and Delta Plus. C_LIO_LITHSC20.HVTR04 and THSC20.HVTR26 target non-competing epitopes on the receptor binding domain (RBD) and represent distinct germline lineages. C_LIO_LIPassive transfer of THSC20.HVTR04 and THSC20.HVTR26 mAbs demonstrated protection against Delta virus challenge in K18-hACE2 mice at low antibody doses. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/474152v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1f1b55corg.highwire.dtl.DTLVardef@1b9b438org.highwire.dtl.DTLVardef@e6d2a6org.highwire.dtl.DTLVardef@f92cd_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2021

Molecular insights into antibody-mediated protection against the prototypic simian immunodeficiency virus

SIVmac239 infection of macaques is a favored model of human HIV infection. However, the SIVmac239 envelope (Env) trimer structure, glycan occupancy, and the targets and ability of neutralizing antibodies (nAbs) to protect against SIVmac239 remain unknown. Here, we report the isolation of SIVmac239 nAbs that recognize a glycan hole and the V1/V4 loop. A high-resolution structure of a SIVmac239 Env trimer-nAb complex shows many similarities to HIV and SIVcpz Envs, but with distinct V4 features and an extended V1 loop. Moreover, SIVmac239 Env has a higher glycan shield density than HIV Env that may contribute to poor or delayed nAb responses in SIVmac239-infected macaques. Passive transfer of a nAb protects macaques from repeated low dose intraveneous SIVmac239 challenge at serum titers comparable to those described for protection of humans against HIV infection. Our results provide structural insights for vaccine design and shed light on antibody-mediated protection in the SIV model.

immunology↗

bioRxiv2021

Broadening a SARS-CoV-1 neutralizing antibody for potent SARS-CoV-2 neutralization through directed evolution

The emergence of SARS-CoV-2 underscores the need for strategies to rapidly develop neutralizing monoclonal antibodies that can function as prophylactic and therapeutic agents and to help guide vaccine design. Here, we demonstrate that engineering approaches can be used to refocus an existing neutralizing antibody to a related but resistant virus. Using a rapid affinity maturation strategy, we engineered CR3022, a SARS-CoV-1 neutralizing antibody, to bind SARS-CoV-2 receptor binding domain with >1000-fold improved affinity. The engineered CR3022 neutralized SARS-CoV-2 and provided prophylactic protection from viral challenge in a small animal model of SARS-CoV-2 infection. Deep sequencing throughout the engineering process paired with crystallographic analysis of an enhanced antibody elucidated the molecular mechanisms by which engineered CR3022 can accommodate sequence differences in the epitope between SARS-CoV-1 and SARS-CoV-2. The workflow described provides a blueprint for rapid broadening of neutralization of an antibody from one virus to closely related but resistant viruses.

bioengineering↗

bioRxiv2021

One-shot identification of SARS-CoV-2 S RBD escape mutants using yeast screening

The potential emergence of SARS-CoV-2 Spike (S) escape mutants is a threat to reduce the efficacy of existing vaccines and neutralizing antibody (nAb) therapies. An understanding of the antibody/S escape mutations landscape is urgently needed to preemptively address this threat. Here we describe a rapid method to identify escape mutants for nAbs targeting the S receptor binding site. We identified escape mutants for five nAbs, including three from the public germline class VH3-53 elicited by natural COVID-19 infection. Escape mutations predominantly mapped to the periphery of the ACE2 recognition site on the RBD with K417, D420, Y421, F486, and Q493 as notable hotspots. We provide libraries, methods, and software as an openly available community resource to accelerate new therapeutic strategies against SARS-CoV-2. One Sentence SummaryWe present a facile method to identify antibody escape mutants on SARS-CoV-2 S RBD.

biochemistry↗