Faculty Bibliography

We studied the contribution of the light chain to functions of human monoclonal antibodies (mAbs) by measuring the relationships between the rate of mutations and cross-reactivity, binding affinity and neutralization activity. We analyzed 12 mAbs of two clonal families specific to the V2 region of HIV-1 derived from two chronically HIV-1 infected individuals. The clonal mAbs exhibited a range of reactivities, and the clones with superior properties were associated with the rate of mutations and the presence of particular mutated residues in the light chains, but not in the heavy chains. Our observations suggest that for some antibodies, the light chains play a vital role in antibody evolution toward more efficient ones and also suggest the importance of optimal residues rather than the rate of mutations in the variable fragment of the antibody.

The repertoire of antibodies (Abs) produced upon vaccination against a particular antigenic site is rarely studied due to the complexity of the immunogens. We received such an opportunity when one rhesus macaque was immunized six times at 0, 4, 10, 16, 32, and 143 weeks with C4-447 peptide containing the 8-mer epitope for human monoclonal Ab (mAb) 447-52D specific to the V3 region of gp120 HIV-1. Strong anti-V3 antibody responses reached 50% binding titer in serum of 10^-5 at week 10 that declined to 10^-3 by week 70. After an additional boost of C4-447 peptide at week 143, titers rebounded to 10^-5 at week 146, or 2.7 years after the first immunization. Using the blood sample at week 146, we produced 41 V3-specific recombinant mAbs by single B cell isolation and cloning. Sequence analysis revealed 21B cell lineages, single and clonally related, based on immunoglobulin gene usage and CDR3s. The broad repertoire of Abs directed to a small antigenic site shows the targeting potency of a vaccine-elicited immune response in rhesus macaques.

The role of vaccine-induced anti-V2 Abs was tested in three protection experiments in rhesus macaques. In an experiment using immunogens similar to those in the RV144 vaccine trial (Anti-envelope [Env]), nine rhesus macaques were coimmunized with gp160_92TH023 DNA and SIV gag and gp120_A244 and gp120_MN proteins. In two V2-focused experiments (Anti-V2 and Anti-V2 Mucosal), nine macaques in each group were immunized with V1V2_92TH023 DNA, V1V2_A244 and V1V2_CasaeA2 proteins, and cyclic V2_CaseA2 peptide. DNA and protein immunogens, formulated in Adjuplex, were given at 0, 4, 12, and 20 weeks, followed by intrarectal SHIV_BaL.P4 challenges. Peak plasma viral loads (PVL) of 10⁶-10⁷ copies/ml developed in all nine sham controls. Overall, PVL was undetectable in one third of immunized macaques, and two animals tightly controlled the virus with the Anti-V2 Mucosal vaccine strategy. In the Anti-Env study, Abs that captured or neutralized SHIV_BaL.P4 inversely correlated with PVL. Conversely, no correlation with PVL was found in the Anti-V2 experiments with nonneutralizing plasma Abs that only captured virus weakly. Titers of Abs against eight V1V2 scaffolds and cyclic V2 peptides were comparable between controllers and noncontrollers as were Ab-dependent cellular cytotoxicity and Ab-dependent cell-mediated virus inhibition activities against SHIV-infected target cells and phagocytosis of gp120-coated beads. The Anti-Env experiment supports the role of vaccine-elicited neutralizing and nonneutralizing Abs in control of PVL. However, the two V2-focused experiments did not support a role for nonneutralizing V2 Abs alone in controlling PVL, as neither Ab-dependent cellular cytotoxicity, Ab-dependent cell-mediated virus inhibition, nor phagocytosis correlated inversely with heterologous SHIV_BaL.P4 infection.

In the only successful RV144 vaccine trial to date, high levels of antibodies (Abs) against the V2 region of the virus envelope protein gp120 correlated with reduced HIV-1 infection. The protective role of V2 Abs has not yet been determined, and the antiviral function of V2 Abs that mediate protection against HIV-1 in humans or SHIV infection in rhesus macaques remains unclear. V2 Abs do not neutralize resistant tier 2 viruses; their Fc-mediated activities are modest and similar to those of another anti-envelope Abs, and inhibition of the gp120-α4β7 integrin interaction is ineffective in both animals and clinical trials. Moreover, in protection experiments in monkeys, levels of V1V2 vaccine-induced V2 Abs do not correlate with plasma viral load. Together, these observations suggest that V2 Abs may not control SHIV infection in rhesus macaques and that V2 Abs may instead be a surrogate marker of other protective immune responses.

Monoclonal antibody 1C10 targets the V3 loop of HIV-1 and neutralizes a broad range of clade B viruses. However, the mode of interaction between 1C10 and the V3 loop remains unclear, because the crystallization of 1C10 and the V3 peptide was unsuccessful because of the flexible nature of 1C10 and the V3 peptide. In this study, we predicted which amino acid residues of 1C10 contact the V3 loop using a deep learning (DL)-based method. Inputs from ROSIE for docking simulation and FastContact, Naccess, and PDBtools, to approximate interactions were processed by Chainer for DL, and outputs were obtained as probabilities of contact residues. Using this DL algorithm, D95, D97, P100a and D100b of CDRH3, D53 and D56 of CDRH2, and D61 of FR3 were highly ranked as contact residues of 1C10. Substitution of these residues to alanine significantly decreased the affinity of 1C10 to the V3 peptide. Moreover, the higher the rank of residue, the more the binding activity was diminished. This study demonstrates that the prediction of contact residues using a DL-based approach is precise and useful for the analysis of antibody-antigen interactions.

BACKGROUND: = 6.7 d, beta-emitter) are two long-lived clinically proven radioisotopes for cancer treatment which might be more effective in killing infected cells systemically and in CNS. METHODS:Lu (4-50 μCi)-2556 mAb. RESULTS:Ac-2556 showing the least non-specific killing. CONCLUSION/CONCLUSIONS:These results indicate that RIT holds promise as a novel treatment option for the eradication of HIV-infected cells that merits further study in combination with cART and reactivation drugs.

Most human immunodeficiency virus (HIV) vaccine trials have lacked efficacy and empirical vaccine lead targets are scarce. Thus far, the only independent correlate of reduced risk of HIV-1 acquisition in humans is elevated levels of V2-specific antibodies identified in the modestly protective RV144 vaccine trial. Ten years after RV144, human and non-human primate vaccine studies have reassessed the potential contribution of V2-specific antibodies to vaccine efficacy. In addition, studies of natural HIV-1 infection in humans have provided insight into the development of V1V2-directed antibody responses and their impact on clinical parameters and disease progression. Functionally diverse anti-V2 monoclonal antibodies were isolated and their structurally distinct V2 epitope regions characterized. After RV144, a plethora of research studies were performed using different model systems, immunogens, protocols, and challenge viruses. These diverse studies failed to provide a clear picture regarding the contribution of V2 antibodies to vaccine efficacy. Here, we summarize the biological functions and clinical findings associated with V2-specific antibodies and discuss their impact on HIV vaccine research.

Most human immunodeficiency virus (HIV) vaccine trials have lacked efficacy and empirical vaccine lead targets are scarce. Thus far, the only independent correlate of reduced risk of HIV-1 acquisition in humans is elevated levels of V2-specific antibodies identified in the modestly protective RV144 vaccine trial. Ten years after RV144, human and non-human primate vaccine studies have reassessed the potential contribution of V2-specific antibodies to vaccine efficacy. In addition, studies of natural HIV-1 infection in humans have provided insight into the development of V1V2-directed antibody responses and their impact on clinical parameters and disease progression. Functionally diverse anti-V2 monoclonal antibodies were isolated and their structurally distinct V2 epitope regions characterized. After RV144, a plethora of research studies were performed using different model systems, immunogens, protocols, and challenge viruses. These diverse studies failed to provide a clear picture regarding the contribution of V2 antibodies to vaccine efficacy. Here, we summarize the biological functions and clinical findings associated with V2-specific antibodies and discuss their impact on HIV vaccine research.

INTRODUCTION/BACKGROUND:In Cameroon, a manifold diversity of HIV strains exists with CRF02_AG and unique recombinant forms (URFs) being the predominant strains. In recent years, a steady increase in URFs and clade F2 viruses has been monitored through partial genome sequencing. There is an information gap in the characterization of emerging URFs along the full genome, which is needed to address the challenges URFs pose towards diagnosis, treatment and HIV-1 vaccine design. METHOD/METHODS:Eighteen Cameroonian URFs from samples collected between the years 2000 and 2015 were studied using a newly developed near full genome sequencing (NFGS) protocol based on variable nested RT-PCRs with a versatile primer set. Near full genomes were characterized for recombination patterns and sequence signatures with possible impact on antiretroviral treatment or Env-directed immune responses. Third-generation sequencing (3GS) of near full or half genomes (HGs) gave insight into intra-patient URF diversity. RESULTS:The characterized URFs were composed of a broad variety of subtypes and recombinants including A, F, G, CRF01_AE, CRF02_AG and CRF22_01A1. Phylogenetic analysis unveiled dominant CRF02_AG and F2 recombination patterns. 3GS indicated a high intra-patient URF diversity with up to four distinct viral sub-populations present in plasma at the same time. URF pol genomic analysis revealed a number of accessory drug resistance mutations (DRMs) in the ART-naïve participants. Genotypic env analysis suggests CCR5 usage in 14/18 samples and identified deviations at residues, critical for gp120/gp41 interphase and CD4 binding site broadly neutralizing antibodies in more than half of the studied URFs. V1V2 sites of immune pressure in the human RV144 vaccine study varied in more than a third of URFs. CONCLUSIONS:This study identified novel mosaic patterns in URFs in Cameroon. In line with the regional predominance of CRF_02AG and the increased prevalence of clade F2, prominent CRF_02AG and F2 background patterns were observed underlying the URFs. In the context of the novel mosaic genomes, the impact of the identified accessory DRMs and Env epitope variations on treatment and immune control remains elusive. The evolving diversity of HIV-1 URFs in Cameroon requires continuous monitoring to respond to the increasing challenges for diagnosis, antiretroviral treatment and prevention.

The results of the RV144 vaccine clinical trial showed a correlation between high level of anti-V1V2 antibodies (Abs) and a decreased risk of acquiring HIV-1 infection. This turned the focus of HIV vaccine design to the induction of elevated levels of anti-V2 Abs to increase vaccine efficacy. In plasma samples from HIV-1 infected Cameroonian individuals, we observed broad variations in levels of anti-V2 Abs, and 6 of the 79 plasma samples tested longitudinally displayed substantial deficiency of V2 Abs. Sequence analysis of the V2 region from plasma viruses and multivariate analyses of V2 characteristics showed a significant difference in several features between V2-deficient and V2-reactive plasma Abs. These results suggest that HIV vaccine immunogens containing a shorter V2 region with fewer glycosylation sites and higher electrostatic charges can be beneficial for induction of a higher level of anti-V2 Abs and thus contribute to HIV vaccine efficacy.