Antibody-Mediated Internalization of Infectious HIV-1 Virions Differs among Antibody Isotypes and Subclasses.
ABSTRACT: Emerging data support a role for antibody Fc-mediated antiviral activity in vaccine efficacy and in the control of HIV-1 replication by broadly neutralizing antibodies. Antibody-mediated virus internalization is an Fc-mediated function that may act at the portal of entry whereby effector cells may be triggered by pre-existing antibodies to prevent HIV-1 acquisition. Understanding the capacity of HIV-1 antibodies in mediating internalization of HIV-1 virions by primary monocytes is critical to understanding their full antiviral potency. Antibody isotypes/subclasses differ in functional profile, with consequences for their antiviral activity. For instance, in the RV144 vaccine trial that achieved partial efficacy, Env IgA correlated with increased risk of HIV-1 infection (i.e. decreased vaccine efficacy), whereas V1-V2 IgG3 correlated with decreased risk of HIV-1 infection (i.e. increased vaccine efficacy). Thus, understanding the different functional attributes of HIV-1 specific IgG1, IgG3 and IgA antibodies will help define the mechanisms of immune protection. Here, we utilized an in vitro flow cytometric method utilizing primary monocytes as phagocytes and infectious HIV-1 virions as targets to determine the capacity of Env IgA (IgA1, IgA2), IgG1 and IgG3 antibodies to mediate HIV-1 infectious virion internalization. Importantly, both broadly neutralizing antibodies (i.e. PG9, 2G12, CH31, VRC01 IgG) and non-broadly neutralizing antibodies (i.e. 7B2 mAb, mucosal HIV-1+ IgG) mediated internalization of HIV-1 virions. Furthermore, we found that Env IgG3 of multiple specificities (i.e. CD4bs, V1-V2 and gp41) mediated increased infectious virion internalization over Env IgG1 of the same specificity, while Env IgA mediated decreased infectious virion internalization compared to IgG1. These data demonstrate that antibody-mediated internalization of HIV-1 virions depends on antibody specificity and isotype. Evaluation of the phagocytic potency of vaccine-induced antibodies and therapeutic antibodies will enable a better understanding of their capacity to prevent and/or control HIV-1 infection in vivo.
Project description:The RV144 prime-boost regimen demonstrated efficacy against HIV acquisition while VAX003 and VAX004 did not. Although these trials differed by risk groups, immunization regimens, and immunogens, antibody responses may have contributed to the differences observed in vaccine efficacy. We assessed HIV-specific IgG, both total and subclass, and IgA binding to HIV envelope (Env): gp120 proteins and Cyclic V2 (CycV2) and CycV3 peptides and gp70 V1 V2 scaffolds in these 3 HIV vaccine trials. After two protein immunizations, IgG responses to 92TH023 gp120 (contained in ALVAC-HIV vaccine) were significantly higher in RV144 but responses to other Env were higher in the VAX trials lacking ALVAC-HIV. IgG responses declined significantly between vaccinations. All trials induced antibodies to gp70 V1 V2 but VAX004 responses to 92TH023 gp70 V1 V2 were weak. All CycV2 responses were undetectable in VAX004 while 92TH023 gp70 V1 V2 was detected in both RV144 and VAX003 but MN CycV2 was detected only in VAX003. Multiple protein vaccinations in VAX trials did not improve magnitude or durability of V1 V2 and CycV2 antibodies. Herpes simplex virus glycoprotein D (gD) peptide at the N terminus of AIDSVAX® B/E and B/B gp120 proteins induced antibodies in all trials, although significantly higher in VAX trials. gD peptide induced IgA, IgG1, IgG2, and IgG3 but not IgG4. Multiple protein vaccinations decreased IgG3 and increased IgG4 changing subclass contribution to total IgG. Although confounded by different modes of HIV transmission, higher Env-specific IgA and IgG4 binding antibodies induced in the VAX trials compared to RV144 raises the hypothesis that these differences may have contributed to different vaccine efficacy results.
Project description:The RV144 vaccine efficacy clinical trial showed a reduction in HIV-1 infections by 31%. Vaccine efficacy was associated with stronger binding antibody responses to the HIV Envelope (Env) V1V2 region, with decreased efficacy as responses wane. High levels of Ab-dependent cellular cytotoxicity (ADCC) together with low plasma levels of Env-specific IgA also correlated with decreased infection risk. We investigated whether B cell priming from RV144 vaccination impacted functional antibody responses to HIV-1 following infection. Antibody responses were assessed in 37 vaccine and 63 placebo recipients at 6, 12, and 36 months following HIV diagnosis. The magnitude, specificity, dynamics, subclass recognition and distribution of the binding antibody response following infection were different in RV144 vaccine recipients compared to placebo recipients. Vaccine recipients demonstrated increased IgG1 binding specifically to V1V2, as well as increased IgG2 and IgG4 but decreased IgG3 to HIV-1 Env. No difference in IgA binding to HIV-1 Env was detected between the vaccine and placebo recipients following infection. RV144 vaccination limited the development of broadly neutralizing antibodies post-infection, but enhanced Fc-mediated effector functions indicating B cell priming by RV144 vaccination impacted downstream antibody function. However, these functional responses were not associated with clinical markers of disease progression. These data reveal that RV144 vaccination primed B cells towards specific binding and functional antibody responses following HIV-1 infection.
Project description:Different HIV-1 antigen specificities appear in sequence after HIV-1 transmission and the immunoglobulin G (IgG) subclass responses to HIV antigens are distinct from each other. The initial predominant IgG subclass response to HIV-1 infection consists of IgG1 and IgG3 antibodies with a noted decline in some IgG3 antibodies during acute HIV-1 infection. Thus, we postulate that multiple antigen-specific IgG3 responses may serve as surrogates for the relative time since HIV-1 acquisition.We determined the magnitude, peak, and half-life of HIV-1 antigen-specific IgG1 and IgG3 antibodies in 41 HIV-1-infected individuals followed longitudinally from acute infection during the first appearance of HIV-1-specific antibodies through approximately 6 months after infection.We used quantitative HIV-1-binding antibody multiplex assays and exponential decay models to estimate concentrations of IgG1 and IgG3 antibodies to eight different HIV-1 proteins including gp140 Env, gp120 Env, gp41 Env, p66 reverse transcriptase, p31 Integrase, Tat, Nef, and p55 Gag proteins during acute/recent HIV-1 infection.Among HIV-1-specific IgG3 responses, anti-gp41 IgG3 antibodies were the first to appear. We found that anti-gp41 Env IgG3 and anti-p66 reverse transcriptase IgG3 antibodies, in addition to anti-Gag IgG3 antibodies, each consistently and measurably declined after acute infection, in contrast to the persistent antigen-specific IgG1 responses.The detailed measurements of the decline in multiple HIV-specific IgG3 responses simultaneous with persistent IgG1 responses during acute and recent HIV-1 infection could serve as markers for detection of incident HIV infection.
Project description:Although passive immunization with anti-HIV-1 Env IgG1 neutralizing monoclonal antibodies (nmAbs) prevented simian-human immunodeficiency virus (SHIV) infection in rhesus monkeys, IgA nmAbs have not been tested. Here, we sought to determine whether human anti-HIV-1 dimeric (d)IgA1, dIgA2, and IgG1 differ in their ability to prevent mucosal R5 SHIV acquisition in rhesus monkeys.DIgA1, dIgA2, and IgG1 versions of nmAb HGN194 were applied intrarectally in three rhesus monkey groups 30 min before intrarectal SHIV challenge.After a control pharmacokinetic study confirmed that nmAb concentrations in rectal fluids over time were similar for all HGN194 isotypes, control and nmAb-treated animals were challenged intrarectally with an R5 SHIV, and viral loads were monitored.Unexpectedly, dIgA1 provided the best protection in vivo--although all nmAbs showed similar neutralizing activity in vitro. Five out of the six dIgA1-treated rhesus monkeys remained virus-free compared to only one out of six animals given dIgA2 (P=0.045 by log-rank test) and two out of six rhesus monkeys treated with IgG1 forms of the nmAb (P=0.12). Protection correlated significantly with virion capture activity by a given nmAb form, as well as inhibition of transcytosis of cell-free virus across an epithelial cell layer in vitro.Our data imply that dIgA1-mediated capturing of virions in mucosal secretions and inhibition of transcytosis can provide significant prevention of lentiviral acquisition--over and above direct virus neutralization. Vaccine strategies that induce mucosal IgA, especially IgA1, should be developed as a first line of defense against HIV-1, a virus predominantly transmitted mucosally.
Project description:HVTN 505 is a preventative vaccine efficacy trial testing DNA followed by recombinant adenovirus serotype 5 (rAd5) in circumcised, Ad5-seronegative men and transgendered persons who have sex with men in the United States. Identified immune correlates of lower HIV-1 risk and a virus sieve analysis revealed that, despite lacking overall efficacy, vaccine-elicited responses exerted pressure on infecting HIV-1 viruses. To interrogate the mechanism of the antibody correlate of HIV-1 risk, we examined antigen-specific antibody recruitment of Fc? receptors (Fc?Rs), antibody-dependent cellular phagocytosis (ADCP), and the role of anti-envelope (anti-Env) IgG3. In a prespecified immune correlates analysis, antibody-dependent monocyte phagocytosis and antibody binding to Fc?RIIa correlated with decreased HIV-1 risk. Follow-up analyses revealed that anti-Env IgG3 breadth correlated with reduced HIV-1 risk, anti-Env IgA negatively modified infection risk by Fc effector functions, and that vaccine recipients with a specific Fc?RIIa single-nucleotide polymorphism locus had a stronger correlation with decreased HIV-1 risk when ADCP, Env-Fc?RIIa, and IgG3 binding were high. Additionally, Fc?RIIa engagement correlated with decreased viral load setpoint in vaccine recipients who acquired HIV-1. These data support a role for vaccine-elicited anti-HIV-1 Env IgG3, antibody engagement of FcRs, and phagocytosis as potential mechanisms for HIV-1 prevention.
Project description:Generating durable humoral immunity through vaccination depends upon effective interaction of follicular helper T cells (Tfh) with germinal center (GC) B cells. Th1 polarization of Tfh cells is an important process shaping the success of Tfh-GC B cell interactions by influencing co-stimulatory and cytokine-dependent Tfh help to B cells. However, the question remains whether adjuvant-dependent modulation of Tfh cells enhances HIV-1 vaccine-induced anti-Envelope (Env) antibody responses. We investigated whether an HIV-1 vaccine platform designed to increase the number of Th1-polarized Tfh cells enhances the magnitude and quality of anti-Env antibodies. Utilizing a novel interferon-induced protein (IP)-10-adjuvanted HIV-1 DNA prime, followed by an MPLA+QS-21-adjuvanted Env protein boost in macaques (DIP-10 PALFQ), we observed higher anti-Env serum IgG titers with greater cross-clade reactivity, specificity to V1V2, and effector functions when compared to macaques primed with DNA lacking IP-10 and boosted with MPLA+alum-adjuvanted Env protein (DPALFA) The DIP-10 PALFQ vaccine regimen elicited higher anti-Env IgG1 and lower IgG4 antibodies in serum, showing for the first time that adjuvants can dramatically impact the IgG subclass profile in macaques. The DIP-10 PALFQ regimen also increased vaginal and rectal IgA antibodies to a greater extent. Within lymph nodes, we observed augmented GC B cell responses and promotion of Th1 gene expression profiles in GC Tfh cells. The frequency of GC Tfh cells correlated with both the magnitude and avidity of anti-Env serum IgG. Together, these data suggest that adjuvant-induced stimulation of Th1-Tfh cells is an effective strategy for enhancing the magnitude and quality of anti-Env antibody response. Overall design: We report enrichment of Th1 transcriptional profiles in TFH cells following vaccination with DIP-10PALFQ Comparision of transciptome profiles across CD4 subsets
Project description:The phase III RV144 HIV-1 vaccine trial estimated vaccine efficacy (VE) to be 31.2%. This trial demonstrated that the presence of HIV-1-specific IgG-binding Abs to envelope (Env) V1V2 inversely correlated with infection risk, while the presence of Env-specific plasma IgA Abs directly correlated with risk of HIV-1 infection. Moreover, Ab-dependent cellular cytotoxicity responses inversely correlated with risk of infection in vaccine recipients with low IgA; therefore, we hypothesized that vaccine-induced Fc receptor-mediated (FcR-mediated) Ab function is indicative of vaccine protection. We sequenced exons and surrounding areas of FcR-encoding genes and found one FCGR2C tag SNP (rs114945036) that associated with VE against HIV-1 subtype CRF01_AE, with lysine at position 169 (169K) in the V2 loop (CRF01_AE 169K). Individuals carrying CC in this SNP had an estimated VE of 15%, while individuals carrying CT or TT exhibited a VE of 91%. Furthermore, the rs114945036 SNP was highly associated with 3 other FCGR2C SNPs (rs138747765, rs78603008, and rs373013207). Env-specific IgG and IgG3 Abs, IgG avidity, and neutralizing Abs inversely correlated with CRF01_AE 169K HIV-1 infection risk in the CT- or TT-carrying vaccine recipients only. These data suggest a potent role of Fc-? receptors and Fc-mediated Ab function in conferring protection from transmission risk in the RV144 VE trial.
Project description:An established sex bias in HIV pathogenesis is linked to immune responses. Recently we reported a vaccine-induced sex bias: vaccinated female but not male rhesus macaques exhibited delayed SIV acquisition. This outcome was correlated with SIV Env-specific rectal IgA, rectal memory B cells, and total rectal plasma cells. To uncover additional contributing factors, using samples from the same study, we investigated memory B cell population dynamics in blood, bone marrow, and rectal tissue during immunization and postchallenge; IgG subtypes and Ab avidity; and regulatory B (Breg) cell frequency and function. Few sex differences were seen in Env-specific memory B cell, plasmablast, or plasma cell frequencies in the three compartments. Males had higher IgG Ab titers and avidity indices than females. However, females had elevated levels of Env-specific IgG1, IgG2, and IgG3 Abs compared with males. gp140-specific IgG3 Abs of females but not males were correlated with Ab-dependent cell-mediated cytotoxicity activity against gp120 targets (p = 0.026) and with Ab-dependent phagocytic activity (p = 0.010). IgG3 Ab of females but not males also correlated with decreased peak viremia (p = 0.028). Peripheral blood CD19(+)CD25(+) Breg cells suppressed T cell proliferation compared with CD19(+)CD25(-) cells (p = 0.031) and exhibited increased IL-10 mRNA expression (p = 0.031). Male macaques postvaccination (p = 0.018) and postinfection (p = 0.0048) exhibited higher Breg frequencies than females. Moreover, male Breg frequencies correlated with peak viremia (p = 0.0071). Our data suggest that vaccinated females developed better Ab quality, contributing to better functionality. The elevated Breg frequencies in males may have facilitated SIV acquisition.
Project description:Analysis of correlates of risk of infection in the RV144 HIV-1 vaccine efficacy trial demonstrated that plasma IgG against the HIV-1 envelope (Env) variable region 1 and 2 inversely correlated with risk, whereas HIV-1 Env-specific plasma IgA responses directly correlated with risk. In the secondary analysis, antibody-dependent cellular cytotoxicity (ADCC) was another inverse correlate of risk, but only in the presence of low plasma IgA Env-specific antibodies. Thus, we investigated the hypothesis that IgA could attenuate the protective effect of IgG responses through competition for the same Env binding sites. We report that Env-specific plasma IgA/IgG ratios are higher in infected than in uninfected vaccine recipients in RV144. Moreover, Env-specific IgA antibodies from RV144 vaccinees blocked the binding of ADCC-mediating mAb to HIV-1 Env glycoprotein 120 (gp120). An Env-specific monomeric IgA mAb isolated from an RV144 vaccinee also inhibited the ability of natural killer cells to kill HIV-1-infected CD4(+) T cells coated with RV144-induced IgG antibodies. We show that monomeric Env-specific IgA, as part of postvaccination polyclonal antibody response, may modulate vaccine-induced immunity by diminishing ADCC effector function.
Project description:<label>BACKGROUND</label>A phase 1 trial of a clade B HIV vaccine in HIV-uninfected adults evaluated the safety and immunogenicity of a DNA prime co-expressing GM-CSF (Dg) followed by different numbers and intervals of modified vaccinia Ankara Boosts (M). Both vaccines produce virus-like particles presenting membrane-bound Env.<label>METHODS</label>Four US sites randomized 48 participants to receiving 1/10th the DNA dose as DgDgMMM given at 0, 2, 4, 6 and 8 months, or full dose DgDgM_M or DgDgMM_M regimens, given at 0, 2, 4, and 8 months, and 0, 2, 4, 6, and 10 months, respectively. Peak immunogenicity was measured 2 weeks post-last vaccination.<label>RESULTS</label>All regimens were well tolerated and safe. Full dose DgDgM_M and DgDgMM_M regimens generated Env-specific IgG to HIV-1 Env in >90%, IgG3 in >80%, and IgA in <20% of participants. Responses to gp140 and gp41 targets were more common and of higher magnitude than to gp120 and V1V2. The gp41 antibody included reactivity to the conserved immunodominant region with specificities known to mediate virus capture and phagocytosis and did not cross-react with a panel of intestinal flora antigens. The 3rd dose of MVA increased the avidity of elicited antibody (7.5% to 39%), the ADCC response to Bal gp120 (14% to 64%), and the one-year durability of the IgG3 responses to gp41 by 4-fold (13% vs. 3.5% retention of peak response). The co-expressed GM-CSF did not enhance responses over those in trials testing this vaccine without GM-CSF.<label>CONCLUSION</label>This DNA/MVA prime-boost regimen induced durable, functional humoral responses that included ADCC, high antibody avidity, and Env IgG1 and IgG3 binding responses to the immunodominant region of gp41. The third, spaced MVA boost improved the overall quality of the antibody response. These products without co-expressed GM-CSF but combined with protein boosts will be considered for efficacy evaluation.<label>TRIAL REGISTRATION</label>ClinicalTrials.gov NCT01571960.