Scientific Publications

Clinical trial participants may differ from the source population due to the demands of trial participation and self-selection, inadvertent selection of a lower-risk group, or both. We investigated the HIV risk status of volunteers in a Simulated Vaccine Efficacy Trial (SiVET) nested within a prospective observational cohort of fisher folks in South Western Uganda.

Induction of broadly neutralizing antibodies (bnAbs) is a major HIV vaccine goal. Germline-targeting immunogens aim to initiate bnAb induction by activating bnAb germline precursor B cells. Critical unmet challenges are to determine whether bnAb precursor naïve B cells bind germline-targeting immunogens and occur at sufficient frequency in humans for reliable vaccine responses. Using deep mutational scanning and multitarget optimization, we developed a germline-targeting immunogen (eOD-GT8) for diverse VRC01-class bnAbs. We then used the immunogen to isolate VRC01-class precursor naïve B cells from HIV-uninfected donors. Frequencies of true VRC01-class precursors, their structures, and their eOD-GT8 affinities support this immunogen as a candidate human vaccine prime. These methods could be applied to germline targeting for other classes of HIV bnAbs and for Abs to other pathogens.

The HIV-1 envelope glycoprotein trimer is covered by an array of N-linked glycans that shield it from immune surveillance. The high density of glycans on the trimer surface imposes steric constraints limiting the actions of glycan-processing enzymes, so that multiple under-processed structures remain on specific areas. These oligomannose glycans are recognized by broadly neutralizing antibodies (bNAbs) that are not thwarted by the glycan shield but, paradoxically, target it. Our site-specific glycosylation analysis of a soluble, recombinant trimer (BG505 SOSIP.664) maps the extremes of simplicity and diversity of glycan processing at individual sites and reveals a mosaic of dense clusters of oligomannose glycans on the outer domain. Although individual sites usually minimally affect the global integrity of the glycan shield, we identify examples of how deleting some glycans can subtly influence neutralization by bNAbs that bind at distant sites. The network of bNAb-targeted glycans should be preserved on vaccine antigens.

Antibody effector functions, such as antibody-dependent cellular cytotoxicity, complement deposition, and antibody-dependent phagocytosis, play a critical role in immunity against multiple pathogens, particularly in the absence of neutralizing activity. Two modifications to the IgG constant domain (Fc domain) regulate antibody functionality: changes in antibody subclass and changes in a single N-linked glycan located in the CH2 domain of the IgG Fc. Together, these modifications provide a specific set of instructions to the innate immune system to direct the elimination of antibody-bound antigens. While it is clear that subclass selection is actively regulated during the course of natural infection, it is unclear whether antibody glycosylation can be tuned, in a signal-specific or pathogen-specific manner. Here, we show that antibody glycosylation is determined in an antigen- and pathogen-specific manner during HIV infection. Moreover, while dramatic differences exist in bulk IgG glycosylation among individuals in distinct geographical locations, immunization is able to overcome these differences and elicit antigen-specific antibodies with similar antibody glycosylation patterns. Additionally, distinct vaccine regimens induced different antigen-specific IgG glycosylation profiles, suggesting that antibody glycosylation is not only programmable but can be manipulated via the delivery of distinct inflammatory signals during B cell priming. These data strongly suggest that the immune system naturally drives antibody glycosylation in an antigen-specific manner and highlights a promising means by which next-generation therapeutics and vaccines can harness the antiviral activity of the innate immune system via directed alterations in antibody glycosylation in vivo.  .

Live attenuated vaccines such as SIV with a deleted nef gene have provided the most robust protection against subsequent vaginal challenge with wild-type (WT) SIV in the SIV-rhesus macaque model of HIV-1 transmission to women. Hence, identifying correlates of this protection could enable design of an effective HIV-1 vaccine. One such prechallenge correlate of protection from vaginal challenge has recently been identified as a system with three components: 1) IgG Abs reacting with the viral envelope glycoprotein trimeric gp41; 2) produced by plasma cells in the submucosa and ectopic tertiary lymphoid follicles in the ectocervix and vagina; and 3) concentrated on the path of virus entry by the neonatal FcR in the overlying epithelium. We now examine the mucosal production of the Ab component of this system after vaginal challenge. We show that vaginal challenge immediately elicits striking increases in plasma cells not only in the female reproductive tract but also at other mucosal sites, and that these increases correlate with low but persistent replication at mucosal sites. We describe vaginal ectopic follicles that are structurally and functionally organized similar to follicles in secondary lymphoid organs, and we provide inferential evidence for a key role of the female reproductive tract epithelium in facilitating Ab production, affinity maturation, and class switch recombination. Vaccination thus accesses an epithelial-immune system axis in the female reproductive tract to respond to exposure to mucosal pathogens. Designing strategies to mimic this system could advance development of an effective HIV-1 vaccine.

Significantly higher levels of plasma CXCL13 [chemokine (C-X-C motif) ligand 13] were associated with the generation of broadly neutralizing antibodies (bnAbs) against HIV in a large longitudinal cohort of HIV-infected individuals. Germinal centers (GCs) perform the remarkable task of optimizing B-cell Ab responses. GCs are required for almost all B-cell receptor affinity maturation and will be a critical parameter to monitor if HIV bnAbs are to be induced by vaccination. However, lymphoid tissue is rarely available from immunized humans, making the monitoring of GC activity by direct assessment of GC B cells and germinal center CD4(+) T follicular helper (GC Tfh) cells problematic. The CXCL13-CXCR5 [chemokine (C-X-C motif) receptor 5] chemokine axis plays a central role in organizing both B-cell follicles and GCs. Because GC Tfh cells can produce CXCL13, we explored the potential use of CXCL13 as a blood biomarker to indicate GC activity. In a series of studies, we found that plasma CXCL13 levels correlated with GC activity in draining lymph nodes of immunized mice, immunized macaques, and HIV-infected humans. Furthermore, plasma CXCL13 levels in immunized humans correlated with the magnitude of Ab responses and the frequency of ICOS(+) (inducible T-cell costimulator) Tfh-like cells in blood. Together, these findings support the potential use of CXCL13 as a plasma biomarker of GC activity in human vaccine trials and other clinical settings.

The envelope glycoprotein trimer (Env) on the surface of HIV-1 recognizes CD4(+) T cells and mediates viral entry. During this process, Env undergoes substantial conformational rearrangements, making it difficult to study in its native state. Soluble stabilized trimers have provided valuable insights into the Env structure, but they lack the hydrophobic membrane proximal external region (MPER, an important target of broadly neutralizing antibodies), the transmembrane domain, and the cytoplasmic tail. Here we present (i) a cryogenic electron microscopy (cryo-EM) structure of a clade B virus Env, which lacks only the cytoplasmic tail and is stabilized by the broadly neutralizing antibody PGT151, at a resolution of 4.2 angstroms and (ii) a reconstruction of this form of Env in complex with PGT151 and MPER-targeting antibody 10E8 at a resolution of 8.8 angstroms. These structures provide new insights into the wild-type Env structure.

Some studies suggest that hormonal contraception, pregnancy, and/or breastfeeding may influence rates of HIV disease progression.

A prophylactic HIV-1 vaccine is a global health priority.

The specificity of CD8(+) T cells is critical for early control of founder/transmitted and reactivated HIV-1. To tackle HIV-1 variability and escape, we designed vaccine immunogen HIVconsv assembled from 14 highly conserved regions of mainly Gag and Pol proteins. When administered to HIV-1-negative human volunteers in trial HIV-CORE 002, HIVconsv vaccines elicited CD8(+) effector T cells which inhibited replication of up to 8 HIV-1 isolates in autologous CD4(+) cells. This inhibition correlated with interferon-γ production in response to Gag and Pol peptide pools, but direct evidence of the inhibitory specificity was missing. Here, we aimed to define through recognition of which epitopes these effectors inhibit HIV-1 replication.

Innate lymphoid cells (ILCs) play a central role in the response to infection by secreting cytokines crucial for immune regulation, tissue homeostasis, and repair. Although dysregulation of these systems is central to pathology, the impact of HIV-1 on ILCs remains unknown. We found that human blood ILCs were severely depleted during acute viremic HIV-1 infection and that ILC numbers did not recover after resolution of peak viremia. ILC numbers were preserved by antiretroviral therapy (ART), but only if initiated during acute infection. Transcriptional profiling during the acute phase revealed upregulation of genes associated with cell death, temporally linked with a strong IFN acute-phase response and evidence of gut barrier breakdown. We found no evidence of tissue redistribution in chronic disease and remaining circulating ILCs were activated but not apoptotic. These data provide a potential mechanistic link between acute HIV-1 infection, lymphoid tissue breakdown, and persistent immune dysfunction.

The elicitation of HIV-1 broadly neutralizing antibodies following envelope glycoprotein (Env) vaccination is exceedingly difficult. Suboptimal engagement of naïve B cells is suggested to limit these low frequency events, especially at the conserved CD4bs. Here, we analyzed CD4bs-directed monoclonal antibodies (mAbs) elicited by YU2 gp140-foldon trimers in a non-human primate by selective sorting using CD4bs 'knock out' trimers. Following two inoculations, the CD4bs-directed mAbs efficiently recognized the eliciting immunogen in their affinity-maturing state but did not recognize CD4bs-defective probes. We reverted these mAbs to their most likely inferred germline (igL) state, leaving the HCDR3 unaltered, to establish correlates of in vitro affinity to in vivo activation. Most igL-reverted mAbs bound the eliciting gp140 immunogen, indicating that CD4bs-directed B cells possessing reasonable affinity existed in the naïve repertoire. We detected relatively high affinities for the majority of the igL mAbs to gp120 and of Fabs to gp140, which, as expected, increased when the antibodies 'matured' following vaccination. Affinity increases were associated with slower off-rates as well as with acquisition of neutralizing capacity. These data reveal in vitro binding properties associated with in vivo activation that result in functional archiving of antigen-specific B cells elicited by a complex glycoprotein antigen following immunization.

Broadly neutralizing antibodies (bnAbs) directed against the mannose-patch on the HIV envelope glycoprotein gp120 have several features that make them desirable targets for vaccine design. The PGT125-131 bnAb family is of particular interest due to its superior breadth and potency. The overlapping epitopes recognized by this family are intricate and neutralization requires interaction with at least two N-linked glycans (N332/N334, N295 or N301) in addition to backbone-mediated contact with the (323)IGDIR(327) motif of the V3 loop. We have recently shown that this bnAb family consists of two distinct antibody classes that can bind alternate arrangements of glycans in the mannose-patch in the absence of N332 thereby limiting viral escape. This led us to further investigate viral resistance and escape mechanisms to the PGT125-131 bnAb family.