Discordant Antigenic Properties of Soluble and Virion SARS-CoV-2 Spike Proteins.

Efforts to develop vaccine and immunotherapeutic countermeasures against the COVID-19 pandemic focus on targeting the trimeric spike (S) proteins of SARS-CoV-2. Vaccines and therapeutic design strategies must impart the characteristics of virion S from historical and emerging variants onto practical constructs such as soluble, stabilized trimers. The virus spike is a heterotrimer of two subunits: S1, which includes the receptor binding domain (RBD) that binds the cell surface receptor ACE2, and S2, which mediates membrane fusion.

Evidence of variable human Fcγ receptor-Fc affinities across differentially-complexed IgG.

Antibody-mediated effector functions are widely considered to unfold according to an associative model of IgG-Fcγ receptor (FcγR) interactions. The associative model presupposes that Fc receptors cannot discriminate antigen-bound IgG from free IgG in solution and have equivalent affinities for each. Therefore, the clustering of Fcγ receptors (FcγR) in the cell membrane, cross-activation of intracellular signaling domains, and the formation of the immune synapse are all the result of avid interactions between the Fc region of IgG and FcγRs that collectively overcome the individually weak, transient interactions between binding partners.

Replacement of Antiretroviral Therapy with HIV Broadly Neutralizing Antibodies to Maximize the Effectiveness of Chemotherapy in HIV Patients with Lung Cancer.

Non-small cell lung cancer (NSCLC) is the most fatal non-AIDS defining cancer in people living with HIV (PWH) on antiretroviral therapy (ART). Treatment of malignancies in PWH requires concomitant cancer therapy and ART, which can lead to potential drug-drug interactions (DDIs) and overlapping toxicities. In this study, we hypothesize that replacement of ART with HIV broadly neutralizing antibodies (bNAbs) during cancer chemotherapy (chemo) may maintain HIV suppression and tumor inhibition while minimizing DDIs and overlapping toxicities.

Identifying CCR5 coreceptor populations permissive for HIV-1 entry and productive infection: implications for in vivo studies.

Background: The chemokine receptor CCR5 is the major coreceptor for HIV-1 cell entry. We previously observed that not all CCR5 mAbs reduce HIV-1 infection, suggesting that only some CCR5 populations are permissive for HIV-1 entry. This study aims to better understand the relevant conformational states of the cellular coreceptor, CCR5, involved in HIV entry.

Structure and Fc-Effector Function of Rhesusized Variants of Human Anti-HIV-1 IgG1s.

Passive transfer of monoclonal antibodies (mAbs) of human origin into Non-Human Primates (NHPs), especially those which function predominantly by a Fc-effector mechanism, requires an preparation step, in which the human mAb is reengineered to an equivalent NHP IgG subclass. This can be achieved by changing both the Fc and Fab sequence while simultaneously maintaining the epitope specificity of the parent antibody. This Ab reengineering process, referred to as rhesusization, can be challenging because the simple grafting of the complementarity determining regions (CDRs) into an NHP IgG subclass may impact the functionality of the mAb.

Safety and immunogenicity of an HIV-1 gp120-CD4 chimeric subunit vaccine in a phase 1a randomized controlled trial.

A major challenge for HIV vaccine development is to raise anti-envelope antibodies capable of recognizing and neutralizing diverse strains of HIV-1. Accordingly, a full length single chain (FLSC) of gp120-CD4 chimeric vaccine construct was designed to present a highly conserved CD4-induced (CD4i) HIV-1 envelope structure that elicits cross-reactive anti-envelope humoral responses and protective immunity in animal models of HIV infection. IHV01 is the FLSC formulated in aluminum phosphate adjuvant.

Rational Engraftment of Quaternary-Interactive Acidic Loops for Anti-HIV-1 Antibody Improvement.

Broadly neutralizing antibodies (bNAbs) are the focus of increasing interest for human immunodeficiency virus type 1 (HIV-1) prevention and treatment. Although several bNAbs are already under clinical evaluation, the development of antibodies with even greater potency and breadth remains a priority. Recently, we reported a novel strategy for improving bNAbs against the CD4-binding site (CD4bs) of gp120 by engraftment of the elongated framework region 3 (FR3) from VRC03, which confers the ability to establish quaternary interactions with a second gp120 protomer.

Performance of nucleocapsid and spike-based SARS-CoV-2 serologic assays.

There is an urgent need for an accurate antibody test for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We have developed 3 ELISA methods, trimer spike IgA, trimer spike IgG, and nucleocapsid IgG, for detecting anti-SARS-CoV-2 antibodies. We evaluated their performance along with four commercial ELISAs, EDI™ Novel Coronavirus COVID-19 ELISA IgG and IgM, Euroimmun Anti-SARS-CoV-2 ELISA IgG and IgA, and one lateral flow assay, DPP® COVID-19 IgM/IgG System (Chembio).

Specificity and Performance of Nucleocapsid and Spike-based SARS-CoV-2 Serologic Assays.

There is an urgent need for an accurate antibody test for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In this paper, we have developed 3 ELISA methods, trimer spike IgA, trimer spike IgG, and nucleocapsid IgG, for detecting anti-SARS-CoV-2 antibodies. We evaluated their performance in comparison with four commercial ELISAs, EDI™ Novel Coronavirus COVID-19 ELISA IgG and IgM, Euroimmun Anti-SARS-CoV-2 ELISA IgG and IgA, and one lateral flow assay, DPP® COVID-19 IgM/IgG System (Chembio).

Antigen-Induced Allosteric Changes in a Human IgG1 Fc Increase Low-Affinity Fcγ Receptor Binding.

Antibody structure couples adaptive and innate immunity via Fab (antigen binding) and Fc (effector) domains that are connected by unique hinge regions. Because antibodies harbor two or more Fab domains, they are capable of crosslinking multi-determinant antigens, which is required for Fc-dependent functions through associative interactions with effector ligands, including C1q and cell surface Fc receptors. The modular nature of antibodies, with distal ligand binding sites for antigen and Fc-ligands, is reminiscent of allosteric proteins, suggesting that allosteric interactions might contribute to Fc-mediated effector functions.

Stoichiometric Analyses of Soluble CD4 to Native-like HIV-1 Envelope by Single-Molecule Fluorescence Spectroscopy.

Analyses of HIV-1 envelope (Env) binding to CD4, and the conformational changes the interactions induce, inform the molecular mechanisms and factors governing HIV-1 infection. To address these questions, we used a single-molecule detection (SMD) approach to study the nature of reactions between soluble CD4 (sCD4) and soluble HIV-1 trimers. SMD of these reactions distinguished a mixture of one, two, or three CD4-bound trimer species.

Concurrent Exposure of Neutralizing and Non-neutralizing Epitopes on a Single HIV-1 Envelope Structure.

The trimeric envelope spikes on the HIV-1 virus surface initiate infection and comprise key targets for antiviral humoral responses. Circulating virions variably present intact envelope spikes, which react with neutralizing antibodies; and altered envelope structures, which bind non-neutralizing antibodies. Once bound, either type of antibody can enable humoral effector mechanisms with the potential to control HIV-1 infection .

Broadly Neutralizing Antibodies against HIV: Back to Blood.

After years of continuous exposure to HIV envelope antigens, a minority of HIV-infected individuals develop a cognate polyclonal humoral response comprising very potent and extremely cross-reactive neutralizing antibodies [broadly neutralizing antibodies (bNAbs)]. Isolated bNAbs derived from memory B cell pools have been the focus of intense studies over the past decade. However, it is not yet known how to translate the features of bNAbs into practical HIV prevention methods.

Identification of Near-Pan-neutralizing Antibodies against HIV-1 by Deconvolution of Plasma Humoral Responses.

Anti-HIV-1 envelope broadly neutralizing monoclonal antibodies (bNAbs) isolated from memory B cells may not fully represent HIV-1-neutralizing profiles measured in plasma. Accordingly, we characterized near-pan-neutralizing antibodies extracted directly from the plasma of two "elite neutralizers." Circulating anti-gp120 polyclonal antibodies were deconvoluted using proteomics to guide lineage analysis of bone marrow plasma cells.

Patterns of conserved gp120 epitope presentation on attached HIV-1 virions.

A complete picture of HIV antigenicity during early replication is needed to elucidate the full range of options for controlling infection. Such information is frequently gained through analyses of isolated viral envelope antigens, host CD4 receptors, and cognate antibodies. However, direct examination of viral particles and virus-cell interactions is now possible via advanced microscopy techniques and reagents.

Conserved signatures indicate HIV-1 transmission is under strong selection and thus is not a "stochastic" process.

Recently, Oberle et al. published a paper in Retrovirology evaluating the question of whether selection plays a role in HIV transmission. The Oberle study found no obvious genotypic or phenotypic differences between donors and recipients of epidemiologically linked pairs from the Swiss cohort.

Survivors Remorse: antibody-mediated protection against HIV-1.

It is clear that antibodies can play a pivotal role in preventing the transmission of HIV-1 and large efforts to identify an effective antibody-based vaccine to quell the epidemic. Shortly after HIV-1 was discovered as the cause of AIDS, the search for epitopes recognized by neutralizing antibodies became the driving strategy for an antibody-based vaccine. Neutralization escape variants were discovered shortly thereafter, and, after almost three decades of investigation, it is now known that autologous neutralizing antibody responses and their selection of neutralization resistant HIV-1 variants can lead to broadly neutralizing antibodies in some infected individuals.

An HIV gp120-CD4 Immunogen Does Not Elicit Autoimmune Antibody Responses in Cynomolgus Macaques.

A promising concept for human immunodeficiency virus (HIV) vaccines focuses immunity on the highly conserved transition state structures and epitopes that appear when the HIV glycoprotein gp120 binds to its receptor, CD4. We are developing chimeric antigens (full-length single chain, or FLSC) in which gp120 and CD4 sequences are flexibly linked to allow stable intrachain complex formation between the two moieties (A. DeVico et al.

Paring Down HIV Env: Design and Crystal Structure of a Stabilized Inner Domain of HIV-1 gp120 Displaying a Major ADCC Target of the A32 Region.

Evidence supports a role of antibody-dependent cellular cytotoxicity (ADCC) toward transitional epitopes in the first and second constant (C1-C2) regions of gp120 (A32-like epitopes) in preventing HIV-1 infection and in vaccine-induced protection. Here, we describe the first successful attempt at isolating the inner domain (ID) of gp120 as an independent molecule that encapsulates the A32-like region within a minimal structural unit of the HIV-1 Env. Through structure-based design, we developed ID2, which consists of the ID expressed independently of the outer domain and stabilized in the CD4-bound conformation by an inter-layer disulfide bond.

Enhanced Immune Responses to HIV-1 Envelope Elicited by a Vaccine Regimen Consisting of Priming with Newcastle Disease Virus Expressing HIV gp160 and Boosting with gp120 and SOSIP gp140 Proteins.

Newcastle disease virus (NDV) expressing HIV-1 BaL gp160 was evaluated either alone or with monomeric BaL gp120 and BaL SOSIP gp140 protein in a prime-boost combination in guinea pigs to enhance envelope (Env)-specific humoral and mucosal immune responses. We showed that a regimen consisting of an NDV prime followed by a protein boost elicited stronger serum and mucosal Th-1-biased IgG responses and neutralizing antibody responses than NDV-only immunizations. Additionally, these responses were higher after the gp120 than after the SOSIP gp140 protein boost.

Conformational Masking and Receptor-Dependent Unmasking of Highly Conserved Env Epitopes Recognized by Non-Neutralizing Antibodies That Mediate Potent ADCC against HIV-1.

The mechanism of antibody-mediated protection is a major focus of HIV-1 vaccine development and a significant issue in the control of viremia. Virus neutralization, Fc-mediated effector function, or both, are major mechanisms of antibody-mediated protection against HIV-1, although other mechanisms, such as virus aggregation, are known. The interplay between virus neutralization and Fc-mediated effector function in protection against HIV-1 is complex and only partially understood.

Synergistic Inhibition of R5 HIV-1 by the Fusion Protein (FLSC) IgG1 Fc and Maraviroc in Primary Cells: Implications for Prevention and Treatment.

Background: Antiretroviral (ARV) drugs targeting retroviral enzymes have been extensively employed to treat HIV-1 infection. Drawbacks of this approach include cost, toxicity, and the eventual emergence of resistant strains that threaten prophylactic and/or therapeutic efficacy. Accordingly, efforts to develop next-generation ARV approaches are warranted, particularly if they can offer a higher threshold of resistance.

λ Light Chain Bias Associated With Enhanced Binding and Function of Anti-HIV Env Glycoprotein Antibodies.

The humoral response to human immunodeficiency virus (HIV) remains incompletely understood. In this report, we describe biased λ light chain use during the HIV Env glycoprotein (Env) response in HIV infection and vaccination. We examined HIV Env binding (and neutralization) in the context of light chain use in subjects with acute HIV infection, chronic HIV infection, and among HIV vaccinees.