Vaccine induction of heterologous HIV-1-neutralizing antibody B cell lineages in humans.

A critical roadblock to HIV vaccine development is the inability to induce B cell lineages of broadly neutralizing antibodies (bnAbs) in humans. In people living with HIV-1, bnAbs take years to develop. The HVTN 133 clinical trial studied a peptide/liposome immunogen targeting B cell lineages of HIV-1 envelope (Env) membrane-proximal external region (MPER) bnAbs (NCT03934541). Here, we report MPER peptide-liposome induction of polyclonal HIV-1 B cell lineages of mature bnAbs and their precursors, the most potent of which neutralized 15% of global tier 2 HIV-1 strains and 35% of clade B strains with lineage initiation after the second immunization. Neutralization was enhanced by vaccine selection of improbable mutations that increased antibody binding to gp41 and lipids. This study demonstrates proof of concept for rapid vaccine induction of human B cell lineages with heterologous neutralizing activity and selection of antibody improbable mutations and outlines a path for successful HIV-1 vaccine development.

Mutation-guided vaccine design: A process for developing boosting immunogens for HIV broadly neutralizing antibody induction.

A major goal of HIV-1 vaccine development is the induction of broadly neutralizing antibodies (bnAbs). Although success has been achieved in initiating bnAb B cell lineages, design of boosting immunogens that select for bnAb B cell receptors with improbable mutations required for bnAb affinity maturation remains difficult. Here, we demonstrate a process for designing boosting immunogens for a V3-glycan bnAb B cell lineage. The immunogens induced affinity-matured antibodies by selecting for functional improbable mutations in bnAb precursor knockin mice. Moreover, we show similar success in prime and boosting with nucleoside-modified mRNA-encoded HIV-1 envelope trimer immunogens, with improved selection by mRNA immunogens of improbable mutations required for bnAb binding to key envelope glycans. These results demonstrate the ability of both protein and mRNA prime-boost immunogens for selection of rare B cell lineage intermediates with neutralizing breadth after bnAb precursor expansion, a key proof of concept and milestone toward development of an HIV-1 vaccine.

Host immunity associated with spontaneous suppression of viremia in therapy-naïve young rhesus macaques following neonatal SHIV infection.

IMPORTANCE: Despite the advent of highly active anti-retroviral therapy, people are still dying from HIV-related causes, many of whom are children, and a protective vaccine or cure is needed to end the HIV pandemic. Understanding the nature and activation states of immune cell subsets during infection will provide insights into the immunologic milieu associated with viremia suppression that can be harnessed via therapeutic strategies to achieve a functional cure, but these are understudied in pediatric subjects. We evaluated humoral and adaptive host immunity associated with suppression of viremia in rhesus macaques infected soon after birth with a pathogenic SHIV. The results from our study provide insights into the immune cell subsets and functions associated with viremia control in young macaques that may translate to pediatric subjects for the design of future anti-viral strategies in HIV-1-infected infants and children and contribute to an understudied area of HIV-1 pathogenesis in pediatric subjects.

Neonatal SHIV infection in rhesus macaques elicited heterologous HIV-1-neutralizing antibodies.

Infants and children infected with human immunodeficiency virus (HIV)-1 have been shown to develop neutralizing antibodies (nAbs) against heterologous HIV-1 strains, characteristic of broadly nAbs (bnAbs). Thus, having a neonatal model for the induction of heterologous HIV-1 nAbs may provide insights into the mechanisms of neonatal bnAb development. Here, we describe a neonatal model for heterologous HIV-1 nAb induction in pathogenic simian-HIV (SHIV)-infected rhesus macaques (RMs). Viral envelope (env) evolution showed mutations at multiple sites, including nAb epitopes. All 13 RMs generated plasma autologous HIV-1 nAbs. However, 8/13 (62%) RMs generated heterologous HIV-1 nAbs with increasing potency over time, albeit with limited breadth, and mapped to multiple nAb epitopes, suggestive of a polyclonal response. Moreover, plasma heterologous HIV-1 nAb development was associated with antigen-specific, lymph-node-derived germinal center activity. We define a neonatal model for heterologous HIV-1 nAb induction that may inform future pediatric HIV-1 vaccines for bnAb induction in infants and children.

mRNA-encoded HIV-1 Env trimer ferritin nanoparticles induce monoclonal antibodies that neutralize heterologous HIV-1 isolates in mice.

The success of nucleoside-modified mRNAs in lipid nanoparticles (mRNA-LNP) as COVID-19 vaccines heralded a new era of vaccine development. For HIV-1, multivalent envelope (Env) trimer protein nanoparticles are superior immunogens compared with trimers alone for priming of broadly neutralizing antibody (bnAb) B cell lineages. The successful expression of complex multivalent nanoparticle immunogens with mRNAs has not been demonstrated. Here, we show that mRNAs can encode antigenic Env trimers on ferritin nanoparticles that initiate bnAb precursor B cell expansion and induce serum autologous tier 2 neutralizing activity in bnAb precursor VH + VL knock-in mice. Next-generation sequencing demonstrates acquisition of critical mutations, and monoclonal antibodies that neutralize heterologous HIV-1 isolates are isolated. Thus, mRNA-LNP can encode complex immunogens and may be of use in design of germline-targeting and sequential boosting immunogens for HIV-1 vaccine development.

Long-Term Recovery of the Adaptive Immune System in Rhesus Macaques After Total Body Irradiation.

PURPOSE: Ionizing radiation causes acute damage to hematopoietic and immune cells, but the long-term immunologic consequences of irradiation are poorly understood. We therefore performed a prospective study of the delayed immune effects of radiation using a rhesus macaque model. METHODS AND MATERIALS: Ten macaques received 4 Gy high-energy x-ray total body irradiation (TBI) and 6 control animals received sham irradiation. TBI caused transient lymphopenia that resolved over several weeks. Once white blood cell counts recovered, flow cytometry was used to immunophenotype the circulating adaptive immune cell populations 4, 9, and 21 months after TBI. Data were fit using a mixed-effects model to determine age-dependent, radiation-dependent, and interacting effects. T cell receptor (TCR) sequencing and quantification of TCR Excision Circles were used to determine relative contributions of thymopoiesis and peripheral expansion to T cell repopulation. Two years after TBI, the cohort was vaccinated with a 23-valent pneumococcal polysaccharide vaccine and a tetravalent influenza hemagglutinin vaccine. RESULTS: Aging, but not TBI, led to significant changes in the frequencies of dendritic cells, CD4 and CD8 T cells, and B cells. However, irradiated animals exhibited increased frequencies of central memory T cells and decreased frequencies of naïve T cells. These consequences of irradiation were time-dependent and more prolonged in the CD8 T cell population. Irradiation led to transient increases in CD8+ T cell TCR Excision Circles and had no significant effect on TCR sequence entropy, indicating T cell recovery was partially mediated by thymopoiesis. Animals that were irradiated and then vaccinated showed normal immunoglobulin G binding and influenza neutralization titers in response to the 4 protein antigens but weaker immunoglobulin G binding titers to 10 of the 23 polysaccharide antigens. CONCLUSIONS: These findings indicate that TBI causes subtle but long-lasting immune defects that are evident years after recovery from lymphopenia.

Ability of nucleoside-modified mRNA to encode HIV-1 envelope trimer nanoparticles.

The success of nucleoside-modified mRNAs in lipid nanoparticles (mRNA-LNP) as COVID-19 vaccines heralded a new era of vaccine development. For HIV-1, multivalent envelope (Env) trimer protein nanoparticles are superior immunogens compared to trimers alone for priming of broadly neutralizing antibody (bnAb) B cell lineages. The successful expression of complex multivalent nanoparticle immunogens with mRNAs has not been demonstrated. Here we show that mRNAs can encode antigenic Env trimers on ferritin nanoparticles that initiate bnAb precursor B cell expansion and induce serum autologous tier 2 neutralizing activity in bnAb precursor VH + VL knock-in mice. Next generation sequencing demonstrated acquisition of critical mutations, and monoclonal antibodies that neutralized heterologous HIV-1 isolates were isolated. Thus, mRNA-LNP can encode complex immunogens and are of use in design of germline-targeting and sequential boosting immunogens for HIV-1 vaccine development.

Fab-dimerized glycan-reactive antibodies are a structural category of natural antibodies.

Natural antibodies (Abs) can target host glycans on the surface of pathogens. We studied the evolution of glycan-reactive B cells of rhesus macaques and humans using glycosylated HIV-1 envelope (Env) as a model antigen. 2G12 is a broadly neutralizing Ab (bnAb) that targets a conserved glycan patch on Env of geographically diverse HIV-1 strains using a unique heavy-chain (VH) domain-swapped architecture that results in fragment antigen-binding (Fab) dimerization. Here, we describe HIV-1 Env Fab-dimerized glycan (FDG)-reactive bnAbs without VH-swapped domains from simian-human immunodeficiency virus (SHIV)-infected macaques. FDG Abs also recognized cell-surface glycans on diverse pathogens, including yeast and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike. FDG precursors were expanded by glycan-bearing immunogens in macaques and were abundant in HIV-1-naive humans. Moreover, FDG precursors were predominately mutated IgM+IgD+CD27+, thus suggesting that they originated from a pool of antigen-experienced IgM+ or marginal zone B cells.

Metastability Gap in the Phase Diagram of Monoclonal IgG Antibody.

Crystallization of IgG antibodies has important applications in the fields of structural biology, biotechnology, and biopharmaceutics. However, a rational approach to crystallize antibodies is still lacking. In this work, we report a method to estimate the solubility of antibodies at various temperatures. We experimentally determined the full phase diagram of an IgG antibody. Using the full diagram, we examined the metastability gaps, i.e., the distance between the crystal solubility line and the liquid-liquid coexistence curve, of IgG antibodies. By comparing our results to the partial phase diagrams of other IgGs reported in literature, we found that IgG antibodies have similar metastability gaps. Thereby, we present an equation with two phenomenological parameters to predict the approximate location of the solubility line of IgG antibodies with respect to their liquid-liquid coexistence curves. We have previously shown that the coexistence curve of an antibody solution can be readily determined by the polyethylene glycol-induced liquid-liquid phase separation method. Combining the polyethylene glycol-induced liquid-liquid phase separation measurements and the phenomenological equation in this article, we provide a general and practical means to predict the thermodynamic conditions for crystallizing IgG antibodies in the solution environments of interest.