B cell receptor (BCR) sequencing is a powerful tool for interrogating immune responses to infection and vaccination, but it provides limited information about the antigen specificity of the sequenced ...BCRs. Here, we present LIBRA-seq (linking B cell receptor to antigen specificity through sequencing), a technology for high-throughput mapping of paired heavy- and light-chain BCR sequences to their cognate antigen specificities. B cells are mixed with a panel of DNA-barcoded antigens so that both the antigen barcode(s) and BCR sequence are recovered via single-cell next-generation sequencing. Using LIBRA-seq, we mapped the antigen specificity of thousands of B cells from two HIV-infected subjects. The predicted specificities were confirmed for a number of HIV- and influenza-specific antibodies, including known and novel broadly neutralizing antibodies. LIBRA-seq will be an integral tool for antibody discovery and vaccine development efforts against a wide range of antigen targets.
Display omitted
•LIBRA-seq: high-throughput mapping of BCR sequence to antigen specificity•Identified HIV- and influenza-specific B cells in two HIV-infected subjects•Predicted antigen reactivity for thousands of single B cells•Identified a previously unknown broadly neutralizing HIV antibody
LIBRA-seq enables high-throughput mapping of B cell receptor sequence to antigen specificity at the single-cell level.
Characterization of single antibody lineages within infected individuals has provided insights into the development of Env-specific antibodies. However, a systems-level understanding of the humoral ...response against HIV-1 is limited. Here, we interrogated the antibody repertoires of multiple HIV-infected donors from an infection-naive state through acute and chronic infection using next-generation sequencing. This analysis revealed the existence of “public” antibody clonotypes that were shared among multiple HIV-infected individuals. The HIV-1 reactivity for representative antibodies from an identified public clonotype shared by three donors was confirmed. Furthermore, a meta-analysis of publicly available antibody repertoire sequencing datasets revealed antibodies with high sequence identity to known HIV-reactive antibodies, even in repertoires that were reported to be HIV naive. The discovery of public antibody clonotypes in HIV-infected individuals represents an avenue of significant potential for better understanding antibody responses to HIV-1 infection, as well as for clonotype-specific vaccine development.
Display omitted
•Within-donor longitudinal antibody repertoire to HIV-1 infection was analyzed by NGS•Public antibody clonotypes shared among multiple HIV-infected individuals were uncovered•A public antibody clonotype shared by three donors was confirmed to be HIV reactive•Antibody sequences from HIV-naive repertoires are similar to known HIV antibodies
The overall antibody repertoires of HIV-infected subjects are considered to be unique. Setliff et al. analyze the longitudinal antibody repertoire of HIV-1-infected individuals to uncover the existence of “public” HIV-reactive antibodies in multiple subjects. Antibody sequences with high identity to known HIV-reactive antibodies were identified even in HIV-naive repertoires.
A leading strategy for developing a prophylactic HIV-1 vaccine is the elicitation of antibodies that can neutralize a large fraction of circulating HIV-1 variants. However, a major challenge that has ...limited the effectiveness of current vaccine candidates is the extensive global diversity of the HIV-1 envelope protein (Env), the sole target for HIV-neutralizing antibodies. To address this challenge, various strategies incorporating Env diversity into the vaccine formulation have been proposed. Here, we assessed the potential of two such strategies that utilize a nanoparticle-based vaccine platform to elicit broadly neutralizing antibody responses. The nanoparticle immunogens developed here consisted of different formulations of Envs from strains BG505 (clade A) and CZA97 (clade C), attached to the N-termini of bacterial ferritin. Single-antigen nanoparticle cocktails, as well as mosaic nanoparticles bearing both Env trimers, elicited high antibody titers in mice and guinea pigs. Furthermore, serum from guinea pigs immunized with nanoparticle immunogens achieved autologous, and in some cases heterologous, tier 2 neutralization, although significant differences between mosaic and single-antigen nanoparticles were not observed. These results provide insights into the ability of different vaccine strategies for incorporating Env sequence diversity to elicit neutralizing antibodies, with implications for the development of broadly protective HIV-1 vaccines.
Development of novel technologies for the discovery of human monoclonal antibodies has proven invaluable in the fight against infectious diseases. Among the diverse antibody repertoires elicited by ...infection or vaccination, often only rare antibodies targeting specific epitopes of interest are of potential therapeutic value. Current antibody discovery efforts are capable of identifying B cells specific for a given antigen; however, epitope specificity information is usually only obtained after subsequent monoclonal antibody production and characterization. Here we describe LIBRA-seq with epitope mapping, a next-generation sequencing technology that enables residue-level epitope determination for thousands of single B cells simultaneously. By utilizing an antigen panel of point mutants within the HIV-1 Env glycoprotein, we identified and confirmed antibodies targeting multiple sites of vulnerability on Env, including the CD4-binding site and the V3-glycan site. LIBRA-seq with epitope mapping is an efficient tool for high-throughput identification of antibodies against epitopes of interest on a given antigen target.
Despite prolific efforts to characterize the antibody response to human immunodeficiency virus type 1 (HIV-1) and hepatitis C virus (HCV) mono-infections, the response to chronic co-infection with ...these two ever-evolving viruses is poorly understood. Here, we investigate the antibody repertoire of a chronically HIV-1/HCV co-infected individual using linking B cell receptor to antigen specificity through sequencing (LIBRA-seq). We identify five HIV-1/HCV cross-reactive antibodies demonstrating binding and functional cross-reactivity between HIV-1 and HCV envelope glycoproteins. All five antibodies show exceptional HCV neutralization breadth and effector functions against both HIV-1 and HCV. One antibody, mAb688, also cross-reacts with influenza and coronaviruses, including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We examine the development of these antibodies using next-generation sequencing analysis and lineage tracing and find that somatic hypermutation established and enhanced this reactivity. These antibodies provide a potential future direction for therapeutic and vaccine development against current and emerging infectious diseases. More broadly, chronic co-infection represents a complex immunological challenge that can provide insights into the fundamental rules that underly antibody-antigen specificity.
Display omitted
•LIBRA-seq is applied to characterize antibody responses to HIV-1/HCV co-infection•A diverse set of functional HIV-1/HCV cross-reactive antibodies are identified•A glycan-directed antibody exhibits exceptionally broad anti-viral recognition•Antibody somatic hypermutation is a major determinant for HIV-1/HCV cross-reactivity
Pilewski et al. uses LIBRA-seq to identify multiple, functional HIV-1/HCV cross-reactive antibodies with diverse epitope specificities. This study identifies a glycan-reactive, broadly anti-viral antibody recognizing the NTD of SARS-CoV-2, in addition to antigens from multiple other viruses. The authors further show that somatic hypermutation both establishes and enhances HIV-1/HCV cross-reactivity.
Public antibody clonotypes shared among multiple individuals have been identified for several pathogens. However, little is known about the determinants of antibody “publicness”. Here, we ...characterize the sequence and functional properties of antibodies from a public clonotype targeting the CD4 binding site on HIV-1 Env. Our results showed that HIV-1 specificity for the public antibodies studied here, comprising sequences from three individuals, was modulated by the VH, but not VL, germline gene. Non-native pairing of public heavy and light chains from different individuals suggested functional complementation of sequences within this public antibody clonotype. The strength of antigen recognition appeared to be dependent on the specific antibody light chain used, but not on other sequence features such as native-antibody or germline sequence identity. Understanding the determinants of antibody clonotype “publicness” can provide insights into the fundamental rules of host-pathogen interactions at the population level, with implications for clonotype-specific vaccine development.
Display omitted
•Defined antibody features that affect the antigen specificity of a public clonotype•Public antibodies exhibited high sequence similarity both within and among donors•Revealed functional complementation of heavy and light chains from different donors
Immunology; Virology
PDF
