Skip to main content
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems
  • Log in
  • My alerts
  • My Cart

Main menu

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Minireviews
    • JVI Classic Spotlights
    • Archive
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About JVI
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
  • ASM
    • Antimicrobial Agents and Chemotherapy
    • Applied and Environmental Microbiology
    • Clinical Microbiology Reviews
    • Clinical and Vaccine Immunology
    • EcoSal Plus
    • Infection and Immunity
    • Journal of Bacteriology
    • Journal of Clinical Microbiology
    • Journal of Microbiology & Biology Education
    • Journal of Virology
    • mBio
    • Microbiology and Molecular Biology Reviews
    • Microbiology Resource Announcements
    • Microbiology Spectrum
    • Molecular and Cellular Biology
    • mSphere
    • mSystems

User menu

  • Log in
  • My alerts
  • My Cart

Search

  • Advanced search
Journal of Virology
publisher-logosite-logo

Advanced Search

  • Home
  • Articles
    • Current Issue
    • Accepted Manuscripts
    • COVID-19 Special Collection
    • Minireviews
    • JVI Classic Spotlights
    • Archive
  • For Authors
    • Submit a Manuscript
    • Scope
    • Editorial Policy
    • Submission, Review, & Publication Processes
    • Organization and Format
    • Errata, Author Corrections, Retractions
    • Illustrations and Tables
    • Nomenclature
    • Abbreviations and Conventions
    • Publication Fees
    • Ethics Resources and Policies
  • About the Journal
    • About JVI
    • Editor in Chief
    • Editorial Board
    • For Reviewers
    • For the Media
    • For Librarians
    • For Advertisers
    • Alerts
    • RSS
    • FAQ
  • Subscribe
    • Members
    • Institutions
Pathogenesis and Immunity

Two Families of Env Antibodies Efficiently Engage Fc-Gamma Receptors and Eliminate HIV-1-Infected Cells

Sai Priya Anand, Jérémie Prévost, Sophie Baril, Jonathan Richard, Halima Medjahed, Jean-Philippe Chapleau, William D. Tolbert, Sharon Kirk, Amos B. Smith III, Bruce D. Wines, Stephen J. Kent, P. Mark Hogarth, Matthew S. Parsons, Marzena Pazgier, Andrés Finzi
Viviana Simon, Editor
Sai Priya Anand
aCentre de Recherche du CHUM, Montreal, QC, Canada
bDepartment of Microbiology and Immunology, McGill University, Montreal, QC, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jérémie Prévost
aCentre de Recherche du CHUM, Montreal, QC, Canada
cDépartement de Microbiologie, Infectiologie et Immunologie, Université de Montréal, Montreal, QC, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Sophie Baril
aCentre de Recherche du CHUM, Montreal, QC, Canada
cDépartement de Microbiologie, Infectiologie et Immunologie, Université de Montréal, Montreal, QC, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jonathan Richard
aCentre de Recherche du CHUM, Montreal, QC, Canada
cDépartement de Microbiologie, Infectiologie et Immunologie, Université de Montréal, Montreal, QC, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Halima Medjahed
aCentre de Recherche du CHUM, Montreal, QC, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Jean-Philippe Chapleau
aCentre de Recherche du CHUM, Montreal, QC, Canada
cDépartement de Microbiologie, Infectiologie et Immunologie, Université de Montréal, Montreal, QC, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
William D. Tolbert
dInfectious Diseases Division, Uniformed Services University of the Health Sciences, Bethesda, Maryland, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Sharon Kirk
eDepartment of Chemistry, School of Arts and Sciences, University of Pennsylvania, Philadelphia, Pennsylvania, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Amos B. Smith III
eDepartment of Chemistry, School of Arts and Sciences, University of Pennsylvania, Philadelphia, Pennsylvania, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Bruce D. Wines
fImmune Therapies Group Burnet Institute, Melbourne, Victoria, Australia
gDepartment of Clinical Pathology, University of Melbourne, Melbourne, Victoria, Australia
hDepartment of Immunology and Pathology, Monash University, Melbourne, Victoria, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Stephen J. Kent
iDepartment of Microbiology and Immunology, Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Victoria, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
  • ORCID record for Stephen J. Kent
P. Mark Hogarth
fImmune Therapies Group Burnet Institute, Melbourne, Victoria, Australia
gDepartment of Clinical Pathology, University of Melbourne, Melbourne, Victoria, Australia
hDepartment of Immunology and Pathology, Monash University, Melbourne, Victoria, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Matthew S. Parsons
iDepartment of Microbiology and Immunology, Peter Doherty Institute for Infection and Immunity, University of Melbourne, Melbourne, Victoria, Australia
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Marzena Pazgier
dInfectious Diseases Division, Uniformed Services University of the Health Sciences, Bethesda, Maryland, USA
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Andrés Finzi
aCentre de Recherche du CHUM, Montreal, QC, Canada
bDepartment of Microbiology and Immunology, McGill University, Montreal, QC, Canada
cDépartement de Microbiologie, Infectiologie et Immunologie, Université de Montréal, Montreal, QC, Canada
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
Viviana Simon
Icahn School of Medicine at Mount Sinai
Roles: Editor
  • Find this author on Google Scholar
  • Find this author on PubMed
  • Search for this author on this site
DOI: 10.1128/JVI.01823-18
  • Article
  • Figures & Data
  • Info & Metrics
  • PDF
Loading

ABSTRACT

HIV-1 conceals epitopes of its envelope glycoproteins (Env) recognized by antibody (Ab)-dependent cellular cytotoxicity (ADCC)-mediating antibodies. These Abs, including anti-coreceptor binding site (CoRBS) and anti-cluster A antibodies, preferentially recognize Env in its “open” conformation. The binding of anti-CoRBS Abs has been shown to induce conformational changes that further open Env, allowing interaction of anti-cluster A antibodies. We explored the possibility that CoRBS Abs synergize with anti-cluster A Abs to engage Fc-gamma receptors to mediate ADCC. We found that binding of anti-CoRBS and anti-cluster A Abs to the same gp120 is required for interaction with soluble dimeric FcγRIIIa in enzyme-linked immunosorbent assays (ELISAs). We also found that Fc regions of both Abs are required to optimally engage FcγRIIIa and mediate robust ADCC. Taken together, our results indicate that these two families of Abs act together in a sequential and synergistic fashion to promote FcγRIIIa engagement and ADCC.

IMPORTANCE The “open” CD4-bound conformation of HIV-1 envelope glycoproteins is the primary target of antibody-dependent cellular cytotoxicity (ADCC)-mediating antibodies present in HIV-positive (HIV+) sera, such as anti-coreceptor binding site and anti-cluster A antibodies. Here we report that the binding of these two families of antibodies is required to engage FcγRIIIa and mediate ADCC.

INTRODUCTION

Increasing evidence suggests that antibody (Ab)-dependent cellular cytotoxicity (ADCC) is an important host response that decreases human immunodeficiency virus type 1 (HIV-1) infection and replication (1–7). Analysis of the RV144 vaccine trial data suggested that increased ADCC activity was associated with decreased HIV-1 acquisition (8), and antibodies (Abs) with potent ADCC activity were isolated from RV144 vaccinees (9). The Fc portions of IgGs on opsonized targets trigger effector responses, including ADCC, by the ligation and cross-linking of specific Fc-gamma receptors (FcγR) expressed on effector cells (10). FcγRIIIa (CD16a) is the principal activating Fc receptor on natural killer (NK) cells, which are major mediators of ADCC.

There are many factors that can contribute to the ability of Abs to engage effector cells to clear infected cells, such as (i) the strength of IgG interactions with FcγRs; (ii) the density of IgG opsonization; and (iii) the orientation of the Fc portion (11, 12). Additionally, the interaction of FcγRs with Abs bound to distinct epitopes may have additive or synergistic effects on host immune responses. Previous reports have shown improved neutralization efficacy with two or more monoclonal Abs (MAbs) targeting the Ebola virus (13), Hepatitis C virus (14) and HIV-1 (15). Using influenza A virus as a model, it has been shown that the combination of MAbs targeting different epitopes enhanced the induction of ADCC in an additive manner (16). These findings exemplify the complex interactions present in a polyclonal Ab response in vivo.

In the context of HIV-1, it has been shown that antibodies targeting the CD4-bound conformation of viral envelope glycoproteins (Env) mediate strong ADCC activity (17–20). It should be noted that these Abs require infected cells to present Env in an “open” conformation for ADCC to be achievable (18, 19, 21, 22). Several families of Abs can bind epitopes exposed in this conformation of Env, including anti-cluster A, anti-coreceptor binding site (anti-CoRBS), anti-V3, and anti-gp41 Abs (17). To avoid exposure of these CD4-induced epitopes, however, HIV-1 efficiently internalizes Env (76) and prevents the accumulation of the CD4 receptor on the surface of infected cells through Nef and Vpu accessory proteins (20). Moreover, Vpu decreases Env levels at the cell surface by antagonizing the BST-2 host restriction factor which otherwise tethers budding viral particles (23, 24) (the impact of these accessory proteins on ADCC responses was recently reviewed in references 25–27).

In agreement with the susceptibility of the CD4-bound conformation of Env to ADCC, it has been shown that small-molecule CD4-mimetics (CD4mc) “force” Env to adopt a similar conformation that enhances susceptibility of infected cells to ADCC triggered by antibodies within sera, breast milk, and cervicovaginal fluids from HIV-1-infected subjects (21, 28–30). The mechanism via which CD4mc facilitate anti-HIV-1 ADCC includes a synergistic interaction of Env with both CD4mc and anti-CoRBS Abs present in HIV-1-positive (HIV-1+) sera. The sequential binding of CD4mc and anti-CoRBS Abs opens the Env trimer and facilitates recognition by the anti-cluster A family of Abs. The members of the latter family of Abs recognize epitopes located in the gp120 inner domain which are occluded in the “closed” native trimer (12, 20, 31–34) and are known to be responsible for the majority of the ADCC activity present in HIV-1+ sera (17, 19, 20, 31, 35). Accordingly, the addition of anti-cluster A Fab fragments has been shown to block most ADCC activity present in HIV-1+ sera (35). Interestingly, addition of Fab fragments from the 17b antibody, which belongs to the anti-CoRBS family of anti-Env antibodies (36), also significantly blocked ADCC activity of HIV-1+ sera and in some cases to the same extent as anti-cluster A Fab fragments (35). Taken together, these studies suggested that there is a high degree of cooperation between anti-cluster A and anti-CoRBS Abs in the induction of effector functions. To mediate ADCC, Abs must bridge their Fab-bound cognate antigen with effector cells capable of recognizing Ab Fc via FcγR. While it was demonstrated that anti-CoRBS Abs facilitate the binding of anti-cluster A Abs to Env (29), it is still not known whether these two families of Abs also coordinate for FcγR engagement. Here we took advantage of the availability of a recently characterized dimeric recombinant soluble FcγRIIIa protein (11, 37–40) to evaluate whether these two families of anti-Env Abs coordinate for FcγR engagement and ADCC responses.

RESULTS

Anti-CoRBS and anti-cluster A Abs synergize to engage a soluble recombinant dimeric form of FcγRIIIa.We recently developed an enzyme-linked immunosorbent assay (ELISA) to model the cross-linking of FcγRs by Abs, a process essential to initiate ADCC responses. Using this assay, we showed that recombinant FcγR dimer engagement of Abs induced by the RV144 vaccine regimen correlated with a functional measure of ADCC (40). As such, we implemented this ELISA to assess anti-cluster A and anti-CoRBS Ab engagement with recombinant soluble dimeric FcγRIIIa (rsFcγRIIIa). Initially, ELISA plates were coated with recombinant gp120 and the ability of gp120-bound anti-cluster A (A32) and/or anti-CoRBS (17b) Abs to engage soluble rsFcγRIIIa was determined. Despite detectable binding of both A32 and 17b to the gp120-coated plates (Fig. 1C), we noted poor binding of rsFcγRIIIa to either Ab alone or to the two Abs in combination (Fig. 1A). Since we recently reported that anti-CoRBS Abs facilitate anti-cluster A Ab binding upon addition of CD4mc BNM-III-170 (29), we evaluated if this small-molecule CD4mc could facilitate rsFcγRIIIa engagement. Plates were coated with recombinant gp120, and then antibody and rsFcγRIIIa binding levels were measured by ELISA in the presence of BNM-III-170. Consistent with the CD4-induced (CD4i) nature of anti-cluster A and anti-CoRBS Abs and the capacity of BNM-III-170 to stabilize gp120 in a CD4-bound conformation (41), both Abs bound gp120 slightly better in the presence of BNM-III-170 (Fig. 1C). Interestingly, only the combination of A32 and 17b resulted in efficient engagement between the Abs and rsFcγRIIIa (Fig. 1A). To confirm that the CD4-bound conformation of gp120 facilitates rsFcγRIIIa engagement by anti-cluster A and anti-CoRBS Abs, this Env conformation was stabilized by deleting the V1, V2, V3, and V5 variable regions (ΔV1V2V3V5) (42, 43). Using this stabilized recombinant gp120, we observed that neither anti-cluster A (A32; N5-i5) nor anti-CoRBS (17b; N12-i2) Abs could independently engage the rsFcγRIIIa. We noted, however, that utilizing any combination of Abs exhibiting these two specificities resulted in a dramatic increase in rsFcγRIIIa engagement (Fig. 1B)—despite similar overall levels of binding of the Abs to the CD4-bound-stabilized gp120 (Fig. 1D). Cumulatively, these data indicate that anti-cluster A and anti-CoRBS Abs synergize to engage with FcγRIIIa.

FIG 1
  • Open in new tab
  • Download powerpoint
FIG 1

Anti-CoRBS and anti-cluster A Abs synergize to engage dimeric FcγRIIIa. Indirect ELISA was performed using recombinant full-length YU2 gp120 protein (0.4 µg/ml) (A and C) or using recombinant YU2 ΔV1V2V3V5 gp120 protein (0.25 µg/ml) (B and D). gp120-coated wells were incubated with a total concentration of 1 µg/ml of primary antibodies (A and C) with or without BNM-III-170 (25 µM). Antibody binding to gp120 was detected using biotin-tagged dimeric rsFcγRIIIa (0.1 µg/ml) followed by the addition of HRP-conjugated streptavidin (A and B) or using HRP-conjugated anti-human secondary antibody (C and D). Data in graphs represent relative light units (R.L.U.) determined from at least 5 independent experiments done in quadruplicate, with the error bars indicating means ± standard errors of the means (SEM). Statistical significance was tested using an unpaired t test or a Mann-Whitney U test based on statistical normality (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, nonsignificant).

Anti-CoRBS and anti-cluster A Abs must recognize the same gp120 monomer in order to engage with dimeric FcγRIIIa.In theory, due to their nonoverlapping epitopes, anti-cluster A and anti-CoRBS Abs could bind the same gp120-coated monomer or two different adjacent gp120-coated monomers to facilitate rsFcγRIIIa engagement. To distinguish between these two possibilities, we coated equivalent amounts of CD4-bound stabilized (ΔV1V2V3V5) gp120 mutants that were competent to engage with anti-cluster A but had decreased binding to anti-CoRBS (R419D) Abs or that were competent to bind anti-CoRBS but had decreased binding to anti-cluster A (W69A) Abs (Fig. 2B). Strikingly, A32 and 17b were able to engage rsFcγRIIIa only when bound to the CD4-bound stabilized gp120 wild type (WT) (having intact epitopes for anti-cluster and CoRBS Abs) (Fig. 2A). Indeed, despite similar overall levels of binding of A32 and 17b to wells coated with an equimolar ratio of R419D and W69A CD4-bound stabilized recombinant gp120s, poor rsFcγRIIIa engagement was observed. These data suggest that A32 and 17b must bind the same gp120 to efficiently engage with FcγRIIIa by ELISA.

FIG 2
  • Open in new tab
  • Download powerpoint
FIG 2

Anti-CoRBS and anti-cluster A Abs must engage with the same gp120 subunit in order to interact with dimeric FcγRIIIa. Indirect ELISA was performed using recombinant YU2 ΔV1V2V3V5 gp120 protein (WT), its mutated counterpart (W69A or R419D), or an equimolar mixture of both mutated gp120 proteins (W69A and R419D). gp120-coated wells were incubated with a total concentration of 1 µg/ml of primary antibodies. Antibody binding to gp120 was detected using (A) biotin-tagged dimeric rsFcγRIIIa (0.1 µg/ml) followed by the addition of HRP-conjugated streptavidin or using (B) HRP-conjugated anti-human secondary antibody. Graphs represent relative light units (R.L.U.) determined from 5 independent experiments done in quadruplicate, with the error bars indicating means ± SEM. Statistical significance was tested using an unpaired t test or a Mann-Whitney U test based on statistical normality (*, P < 0.05; ****, P < 0.0001; ns, nonsignificant).

Anti-CoRBS and anti-cluster A Abs present in HIV-1+ sera are required for efficient engagement of FcγRIIIa.Since the two families of Abs tested in this study are easily elicited following HIV-1 infection (19, 29, 31, 44), we evaluated their contribution to the capacity of sera from 15 chronically HIV-1-infected individuals to bind to the rsFcγRIIIa by ELISA using the gp120 stabilized in the CD4-bound conformation (ΔV1V2V3V5). Compared to sera from 5 healthy HIV-1-uninfected donors, rsFcγRIIIa was efficiently engaged by Abs present in most HIV-1+ sera tested (Fig. 3A). Interestingly, the addition of A32 or 17b Fab fragments to the HIV-1+ sera significantly blocked the binding of rsFcγRIIIa (Fig. 3A). Addition of A32 and 17b fragments in combination significantly diminished rsFcγRIIIa engagement to the level seen with sera from healthy uninfected (HIV−) individuals (Fig. 3A). Similarly, gp120 mutants with an impaired ability to engage anti-cluster A (W69A) and/or anti-CoRBS (R419D) Abs failed to facilitate rsFcγRIIIa engagement with bound Abs (Fig. 3B). As such, the data suggest that anti-cluster A and anti-CoRBS Abs represent the majority of Abs able to recruit FcγRIIIa in HIV-1+ sera toward the gp120 stabilized in the CD4-bound conformation.

FIG 3
  • Open in new tab
  • Download powerpoint
FIG 3

Dimeric FcγRIIIa engagement by HIV+ sera is driven by anti-CoRBS and anti-cluster A Abs. (A) Indirect ELISA using recombinant YU2 ΔV1V2V3V5 gp120 protein (0.25 µg/ml). gp120-coated wells were incubated with sera from 15 chronically HIV-1-infected individuals with or without A32 Fab and/or 17b Fab fragments (1 µg/ml). (B) Indirect ELISA using recombinant YU2 ΔV1V2V3V5 gp120 protein (WT) or its mutated counterparts (W69A or R419D or the double mutant W69A/R419D). Serum binding to gp120 was detected using biotin-tagged dimeric rsFcγRIIIa (0.1 µg/ml) followed by the addition of HRP-conjugated streptavidin. Sera from 5 healthy individuals (HIV−) were used as a negative control. Graphs represent relative light units (R.L.U.) determined from 3 independent experiments done in quadruplicate, with the error bars indicating means ± SEM. Statistical significance was tested using a paired t test or a Wilcoxon matched-pair signed-rank test (based on statistical normality) to compare HIV+ serum data sets and a Mann-Whitney U test to compare HIV+ sera with HIV− sera (****, P < 0.0001; ns, nonsignificant).

Anti-CoRBS and anti-cluster A Abs are required for efficient binding of FcγRIIIa to infected cells.The work described above and prior studies used plate-bound Env proteins to measure engagement of FcγRIIIa. We therefore developed a more physiologically appropriate system to measure this interaction, using HIV-1-infected cells. Primary CD4+ T cells were infected with primary transmitted/founder (T/F) virus CH58 (CH58 T/F) that was either wild type (WT) or defective for Nef and Vpu expression (N−U−). The N−U− virus was used as a means to present Env in the CD4-bound conformation at the surface of infected cells (19, 20). Infected cells were identified by intracellular p24 staining. As previously shown (29), cells infected with the wild-type virus and therefore exposing Env in its untriggered closed conformation were poorly recognized by A32 or 17b in the absence of the CD4mc BNM-III-170. The presence of BNM-III-170 enhanced recognition of the infected cells by 17b but not A32 (Fig. 4C). In agreement with the ability of 17b to facilitate A32 binding to Env in the presence of CD4mc (29) (Fig. 5), recognition of infected cells in the presence of BNM-III-170 was significantly higher for the combination of 17b and A32 than for 17b alone. Cobinding of 17b and A32 to CD4mc-bound Env translated to efficient and significant engagement of rsFcγRIIIa (Fig. 4A and B). As expected, cells infected with the N−U− virus presented Env in its CD4-bound conformation. As such, N−U− virus-infected cells were efficiently recognized by 17b and by A32 and by the combination of the two Abs (Fig. 4G and H). Interestingly, however, only the combination of A32 and 17b was able to engage dimeric rsFcγRIIIa (Fig. 4E and F). Thus, anti-cluster A and anti-CoRBS Abs are required to efficiently bind rsFcγRIIIa to infected cells.

FIG 4
  • Open in new tab
  • Download powerpoint
FIG 4

Anti-CoRBS and anti-cluster A Abs are required for efficient engagement of FcγRIIIa to infected cells. Cell surface staining of primary CD4+ T cells infected with CH58 T/F virus strains that were either WT (A to D) or defective (E to H) for Nef and Vpu expression (N-U-) was performed with various concentrations of A32 and 17b alone or in combination (1:1 ratio). (A to D) Staining with CH58 T/F WT virus was done with or without BNM-III-170 (50 µM). Antibody binding was detected using a biotin-tagged dimeric rsFcγRIIIa (0.2 µg/ml) followed by the addition of Alexa Fluor 647-conjugated streptavidin (A, B, E, and F) or using Alexa Fluor 647-conjugated anti-human secondary antibody (C, D, G, and H). (A, C, E, and G) Graphs represent mean fluorescence intensities (MFI) in the infected (p24+) population determined from 5 independent experiments, with the error bars indicating means ± SEM. (B, D, F, and H) Areas under the curve (AUC) were calculated based on MFI data sets using GraphPad Prism software. Statistical significance was tested using an unpaired t test or a Mann-Whitney U test based on statistical normality (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, nonsignificant).

FIG 5
  • Open in new tab
  • Download powerpoint
FIG 5

Presence of CD4mc in combination with Fab′2 but not Fab′1 fragments of anti-CoRBS Abs is sufficient to expose cluster A epitopes. Cell surface staining of primary CD4+ T cells infected with CH58 T/F WT virus was performed with (A) Alexa Fluor 647-conjugated N5i5 or (B) Alexa Fluor 647-conjugated A32 with 1.25 µg/ml of CoRBS antibodies with or without BNM-III-170 (50 µM). Graphs represent the mean fluorescence intensities (MFI) in the infected (p24+) population determined from 5 independent experiments, with the error bars indicating means ± SEM. Statistical significance was tested using an unpaired t test (*, P < 0.05; ****, P < 0.0001; ns, nonsignificant).

Anti-CoRBS and anti-cluster A Abs are required for efficient ADCC-mediated killing of HIV-1-infected cells.To evaluate whether engagement of rsFcγRIIIa translates to ADCC, primary CD4+ T cells from at least five healthy uninfected (HIV−) donors were infected with CH58 T/F WT and N−U− viruses and evaluated for their susceptibility to ADCC mediated by effector cells present in autologous peripheral blood mononuclear cells (PBMCs). ADCC was assessed using an assay measuring the elimination of infected (p24+) cells (19–21, 30, 45). In agreement with the lack of rsFcγRIIIa binding to wild-type-infected cells treated with A32, 17b, or A32/17b in the absence of BNM-III-170 CD4mc (Fig. 4A), only low ADCC activity was observed for A32, 17b, or A32/17b in the absence of BNM-III-170 (Fig. 6A and B). Significant ADCC activity was observed only upon addition of the small-molecule CD4mc BNM-III-170 with the combination of A32 and 17b Abs as predicted by rsFcγRIIIa binding. As previously reported, cells exposing Env in its CD4-bound conformation due to Nef and Vpu deletions were susceptible to ADCC mediated by A32 (19, 20) but not 17b (17). However, the combination of A32 and 17b further enhanced ADCC (Fig. 6C and D). Taken together, these data suggest that anti-cluster A and anti-CoRBS Abs cooperate for ADCC activity against infected cells by better engaging FcγRIIIa.

FIG 6
  • Open in new tab
  • Download powerpoint
FIG 6

ADCC against HIV-1-infected cells expressing Env in its CD4-bound conformation is enhanced by the combination of anti-CoRBS and anti-cluster A Abs. Primary CD4+ T cells from at least 5 different healthy donors were infected with CH58 T/F virus strains that were either WT (A and B) or defective (C and D) for Nef and Vpu expression (N-U-) and used as target cells and autologous PBMC as effector cells in a FACS-based ADCC assay (29). (A and B) Analysis of ADCC with CH58 T/F WT was done with or without BNM-III-170 (50 µM). Shown in panels A and C are the percentages of ADCC-mediated killing obtained with various concentrations of A32 and 17b alone or in combination (1:1 ratio). (B and D) Areas under the curve (AUC) were calculated based on ADCC data sets using GraphPad Prism software. Data are representative of results from 5 independent experiments, with the error bars indicating means ± SEM. Statistical significance was tested using an unpaired t test or a Mann-Whitney U test based on statistical normality (*, P < 0.05; ***, P < 0.001; ****, P < 0.0001; ns, nonsignificant).

Fc regions of anti-cluster A and anti-CoRBS Abs are required to engage FcγRIIIa and mediate ADCC.To evaluate how the combination of anti-cluster A and anti-CoRBS Abs facilitates engagement of rsFcγRIIIa, we introduced double leucine-to-alanine substitutions at positions 234 and 235 (LALA) of the Fc region of anti-CoRBS (N12i2) and anti-cluster A (N5i5) Abs. The LALA mutation significantly decreases Ab engagement with FcγRIIIa (46–48). First, we evaluated the ability of the unmutated Abs and their LALA counterparts to engage rsFcγRIIIa by ELISA. As shown in Fig. 7A, only the combination of Fc-competent anti-cluster A and anti-CoRBS Abs engaged rsFcγRIIIa, despite similar levels of Ab binding to coated gp120 under conditions employing single Abs or combinations of LALA mutant Abs (Fig. 7B). These results were corroborated by experiments assessing rsFcγRIIIa engagement with Abs bound to cells infected with wild-type and N−U− viruses. In cells infected with the wild-type virus, anti-cluster A and anti-CoRBS Abs were able to efficiently engage the rsFcγRIIIa upon small-molecule CD4mc BNM-III-170 addition, but only if Fc-competent Abs were used (Fig. 7C). Similarly, only combinations of Fc-competent anti-cluster A and anti-CoRBS Abs were able to engage rsFcγRIIIa to Nef−Vpu− virus-infected cells (Fig. 7D). Importantly, rsFcγRIIIa engagement translated into ADCC activity (Fig. 7E and F). Likewise, even though Fab′2 fragments of anti-CoRBS Abs sufficiently expose anti-cluster A epitopes in the presence of CD4mc (29) (Fig. 5), only the combination of anti-cluster A Abs with full IgG anti-CoRBS Abs, and not Fab'2 fragments, optimally recruited rsFcγRIIIa and mediated potent ADCC in the presence of CD4mc (Fig. 8 and 9). Thus, the data suggest that a threshold of FcγRIIIa engagement is required to mediate ADCC. Moreover, our results indicate that both Fc regions of these two families of Abs are required to engage FcγRIIIa to mediate ADCC.

FIG 7
  • Open in new tab
  • Download powerpoint
FIG 7

Introduction of LALA mutations in the Fc portion of both CoRBS and anti-cluster A antibodies decreases engagement of dimeric FcγRIIIa and ADCC against HIV-1-infected cells. (A and B) Indirect ELISA using recombinant YU2 ΔV1V2V3V5 gp120 protein (0.25 µg/ml) with a total concentration of 1 µg/ml of primary antibodies. Antibody binding to gp120 was detected using (A) a biotin-tagged dimeric rsFcγRIIIa (0.1 µg/ml) followed by the addition of HRP-conjugated streptavidin or using (B) HRP-conjugated anti-human secondary antibody. (A and B) Graphs represent relative light units (R.L.U.) determined from at least 5 independent experiments done in quadruplicate, with the error bars indicating means ± SEM. (C and D) Cell surface staining of primary CD4+ T cells infected with CH58 T/F virus strains that were either (C) WT or (D) defective for Nef and Vpu expression (N-U-) with a total concentration of 2.5 µg/ml of primary antibodies. (C) Staining with CH58 T/F WT virus was done in the presence of BNM-III-170 (50 µM). (C and D) Antibody binding was detected using a biotin-tagged dimeric rsFcγRIIIa (0.2 µg/ml) followed by the addition of Alexa Fluor 647-conjugated streptavidin. Graphs represent mean fluorescence intensities (MFI) in the infected (p24+) population determined from 5 independent experiments, with the error bars indicating means ± SEM. (E and F) Primary CD4+ T cells from at least 5 different healthy donors infected with CH58 T/F virus strains that were either (E) WT or (F) defective for Nef and Vpu expression (N-U-) were used as target cells and autologous PBMC as effector cells in a FACS-based ADCC assay (29). (E) Analysis of ADCC with CH58 T/F WT was done in the presence of BNM-III-170 (50 µM). Shown in panels E and F are the percentages of ADCC-mediated killing obtained with a total concentration of 2.5 µg/ml of antibodies. Data are representative of results from 5 independent experiments, with the error bars indicating means ± SEM. Statistical significance was tested using an unpaired t test or a Mann-Whitney U test based on statistical normality (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, nonsignificant).

FIG 8
  • Open in new tab
  • Download powerpoint
FIG 8

The Fc portions from both anti-CoRBS and anti-cluster A Abs are required to engage dimeric FcγRIIIa. Cell surface staining of primary CD4+ T cells infected with CH58 T/F WT virus was performed with a total concentration of 2.5 µg/ml of primary antibodies with or without BNM-III-170 (50 µM). Antibody binding was detected either by (A and B) using biotin-tagged dimeric rsFcγRIIIa (0.2 µg/ml) followed by the addition of Alexa Fluor 647-conjugated streptavidin or by (C and D) using Alexa Fluor 647-conjugated anti-human secondary antibody. Graphs represent the mean fluorescence intensities (MFI) in the infected (p24+) population determined from 5 independent experiments, with the error bars indicating means ± SEM. Statistical significance was tested using an unpaired t test or a Mann-Whitney U test based on statistical normality (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, nonsignificant).

FIG 9
  • Open in new tab
  • Download powerpoint
FIG 9

The Fc portions from both anti-CoRBS and anti-cluster A Abs are required to mediate potent ADCC against HIV-1-infected cells. Primary CD4+ T cells from at least five different healthy donors were infected with CH58 T/F WT virus and used as target cells. Autologous PBMCs were used as effector cells in a FACS-based ADCC assay. Shown in panels A to D are the percentages of ADCC-mediated killing obtained with a total concentration of 2.5 µg/ml of antibodies in the absence (black bars) or presence (gray bars) of BNM-III-170 (50 µM). Data are representative of results from 5 independent experiments, with the error bars indicating means ± SEM. Statistical significance was tested using an unpaired t test (***, P < 0.001; ****, P < 0.0001).

DISCUSSION

HIV-1 has evolved several mechanisms to evade the humoral response elicited against its envelope glycoproteins, including conformational masking of vulnerable epitopes, high sequence variability in the variable loops, and extensive glycosylation (49–51). The conformational dynamics of Env also constitute a major barrier for Env inactivation by Abs. The functional Env trimer mainly exists in an untriggered closed conformation (state 1) that cannot be seen by nonneutralizing Abs (nnAbs). However, Env has intrinsic access to the open CD4-bound conformation (state 3) through one necessary conformational intermediate (state 2) in which only one protomer is competent for CD4 binding (52, 53). Env from primary tier 2 viruses are more closed, with elevated state 1 occupancy, explaining why they are resistant to binding and neutralization by nnAbs (53). CD4 and CD4mc interaction modifies the conformational landscape toward state 3 by lowering the energy barrier to open states 2 and 3, rendering Env vulnerable to nnAb attack (18, 52–54). The majority of the antibodies elicited during natural infection are strain-specific neutralizing or nnAbs (55) and are thought to play a minimal role in controlling viral replication. Recent studies, however, showed that these antibodies can exert constant selection pressure and alter the course of HIV-1-infection in vivo (56, 57). Nonneutralizing Abs with Fc-effector function are present in HIV-1+ sera and have been shown to mediate potent ADCC activity against cells presenting Env in its open CD4-bound conformation (17–21, 26, 30, 58–60). Among the different nnAbs tested for their ADCC activity, the members of the anti-cluster A family of Abs were identified as being the most potent against target cells expressing Env in this open conformation (17, 31, 33, 35). Of note, these Abs can be readily elicited in nonhuman primates with monomeric gp120 immunogens (22). Additional nnAbs such as anti-CoRBS Abs, which are unable to mediate potent ADCC activity on their own (17), were shown to facilitate engagement of anti-cluster A Abs and potentiate their ADCC activity (29). Here we provide evidence that anti-cluster A and anti-CoRBS Abs collaborate in engaging the dimeric form of the FcγRIIIa receptor when bound to the cognate epitopes within the same monomeric gp120. We utilized a novel probe, a covalent FcγRIIIa dimer, as a surrogate of FcγRIIIa present on effector cells (11, 37, 39, 40, 61) and showed that sequential binding of antibodies specific for these two CD4i epitope targets is required for effective receptor engagement and potent ADCC. Recent structural studies defined the Env epitopes recognized by anti-cluster A and anti-CoRBS Abs at the atomic level and mapped the cluster A epitope region to the C1-C2 regions of the gp120 inner domain in its CD4-bound conformation (12, 32, 33, 62) and CoRBS to the bridging sheet and the base of the V3 loop of the CD4-triggered gp120 (63–65). These two epitopes are not overlapping. In order to validate our findings of collaboration in effective engagement of the dimeric Fc-gamma receptor by antibodies specific for these two epitope targets, we prepared a model of the putative immune complex that needs to be formed on the gp120 antigen in order to engage a covalent FcγRIIIa dimer (Fig. 10). We used the crystallographic dimer of FcγRIIa-HR (PDB code 3RY5), a high-responder polymorphism FcγRIIa (66), as a template to generate the putative FcγRIIIa dimer. As predicted, the anti-cluster A N5-i5 and anti-CoRBS 17b antibodies bound to nonoverlapping sites on the same gp120 protomer such that their Fc domains were brought in close proximity (Fig. 10). Simple adjustment of Fc positions, which could be mediated by the IgG1 hinge region, oriented the Fcs to permit interactions with the FcγRIIIa dimer (Fig. 10). Taken together, the in silico modeling data clearly support the possibility of the formation of complexes in which the anti-cluster A and anti-CoRBS Abs are bound to the same gp120 antigen and their Fc domains engaged in an interaction with the dimeric Fc gamma receptor. However, whether the same configuration is maintained on trimeric Env remains to be shown.

FIG 10
  • Open in new tab
  • Download powerpoint
FIG 10

Model of the dimeric rsFcγRIIIa engagement shown by anti-CoRBS and anti-cluster A Abs bound to their cognate epitopes within the same gp120 subunit. The model was built using representative anti-CoRBS and anti-cluster A Abs for which crystal structures of Fab-antigen complexes are available and the dimeric rsFcγRIIIa receptor as described in Materials and Methods. Monomeric gp120 (shown in gray and black for the inner and outer domains, respectively) immobilized on a plate surface is triggered by the small-molecule CD4 mimetic compound BNM-III-170 to expose the coreceptor binding site (top panel). gp120 in the BNM-III-170-bound conformation is recognized first by anti-CoRBS Ab (17b or N12i2) followed by anti-cluster A Ab (A32 or N5i5) binding (middle panel). The proximity of the CoRBS and cluster A regions within the gp120 antigen permits easy engagement of Ab Fc domains in the complex with the dimeric rsFcγRIIIa receptor (bottom panel). Only minimal adjustment of positions (possibly mediated by the IgG hinge region) of anti-CoRBS and anti-cluster A bound to the same gp120 molecule is required for engagement of the dimeric rsFcγRIIIa receptor. In vivo, an array of identical complexes that involve two Abs bound to the same gp120 antigen and the dimeric rsFcγRIIIa receptor is presumably required to effectively stimulate an effector cell.

Although our results revealed that anti-cluster A and anti-CoRBS Abs collaborate to trigger anti-HIV-1 ADCC, note that a collaboration between these Abs is required for ADCC only in the context of CD4mc. Indeed, target cells infected with N−U− viruses bind A32 and are susceptible to ADCC through A32 alone. Similarly, gp120-coated CD4+ T cells are susceptible to ADCC triggered by A32 alone (45, 67). The exact mechanisms behind the differential requirements for anti-CoRBS Abs for ADCC against target cells infected with wild-type virus in the presence of CD4mc and target cells coated with gp120 or infected with N−U− viruses remain undetermined. One possible contributor to this phenomenon is the minimal density and/or proximity of Env on cells infected with wild-type virus, increasing the need for multiple Abs per Env spike (68). Another possible contributor is that the conformational changes in Env induced by cell surface CD4 (present on target cells coated with gp120 or infected with N−U− viruses) are sufficient to allow binding to cell surface coreceptors and negate the need for anti-CoRBS Abs to reveal the A32 epitope (12, 20). The conformational changes in Env induced by CD4mc, however, are not sufficient to allow coreceptor binding to Env, thus necessitating anti-CoRBS Ab binding to reveal the A32 epitope (29).

Taken together, our data identified a complex interplay between two families of easy-to-elicit antibodies, present in most HIV-1-infected individuals, with good potential to eliminate HIV-1-infected cells, provided that the Env in the target cell is in a more open conformation. Interestingly, small-molecule CD4-mimetics have been shown to “open up” the Env trimer (21, 22, 30); therefore, these data suggest that strategies aimed at stabilizing open Env conformations with CD4mc might expedite the design of new strategies aimed at fighting HIV-1 infection.

MATERIALS AND METHODS

Ethics statement.Written informed consent was obtained from all study participants (the Montreal Primary HIV Infection Cohort [69, 70] and the Canadian Cohort of HIV Infected Slow Progressors [71, 72]), and research adhered to the ethical guidelines of Centre de Recherche du CHUM (CRCHUM) and was reviewed and approved by the CRCHUM institutional review board (Ethics Committee approval number CE16.164-CA). Research adhered to the standards indicated by the Declaration of Helsinki. All participants were adults and provided informed written consent prior to enrollment in accordance with Institutional Review Board approval. All sera were heat inactivated for 30 min at 56°C and stored at 4°C until use in subsequent experiments.

Isolation of primary cells.CD4 T lymphocytes were purified from resting PBMCs by negative selection and activated as previously described (20, 21). Briefly, PBMCs were obtained by leukapheresis. CD4+ T lymphocytes were purified using immunomagnetic beads per the instructions of the manufacturer (StemCell Technologies). CD4+ T lymphocytes were activated with phytohemagglutinin-L (10 µg/ml) for 48 h and then maintained in RPMI 1640 complete medium supplemented with recombinant interleukin-2 (rIL-2) (100 U/ml).

Viral production and infections.Vesicular stomatitis virus G (VSVG)-pseudotyped viruses were produced and titrated as described previously (19). Viruses were used to infect activated primary CD4 T cells from healthy HIV-1-negative donors by spin infection at 800 × g for 1 h in 96-well plates at 25°C.

CD4-mimetics.The small-molecule CD4-mimetic compound BNM-III-170 was synthesized as described previously (41). The compounds were dissolved in dimethyl sulfoxide (DMSO) at a stock concentration of 10 mM and diluted to 25 μM in blocking buffer for the indirect ELISAs and to 50 μM in phosphate-buffered saline (PBS) for cell surface staining or in RPMI 1640 complete medium for ADCC assays.

Antibodies and sera.Anti-cluster A MAbs A32 and N5i5 were conjugated with Alexa-Fluor 647 probe (Thermo Fisher Scientific) per the manufacturer’s protocol and used for cell surface staining of HIV-1-infected primary CD4+ T cells. Additionally, the following anti-Env MAbs were also used for cell surface staining: N12i2 LALA, N5i5 LALA, the Fab fragments, Fab′2 fragments, or full CoRBS MAbs 17b and N12-i2. Goat anti-human Alexa Fluor-647 secondary Ab (Thermo Fisher Scientific) was used to determine overall antibody and serum binding and AquaVivid (Thermo Fisher Scientific) as a viability dye. Alexa Fluor 647-conjugated streptavidin was used to determine levels of biotin-tagged dimeric recombinant soluble FcγRIIIa (V158) binding. Sera from HIV-infected and healthy donors were collected, heat inactivated, and conserved as previously described (19).

Protein production of recombinant proteins.FreeStyle 293F cells (Invitrogen) were grown in FreeStyle 293F medium (Invitrogen) to a density of 1 × 106 cells/ml at 37°C with 8% CO2 with regular agitation (150 rpm). Cells were transfected with a codon-optimized plasmid expressing His6-tagged wild-type or mutant HIV-1 YU2 gp120 using ExpiFectamine 293 transfection reagent, as directed by the manufacturer (Invitrogen). At 1 week later, the cells were pelleted and discarded. The supernatants were filtered (Thermo Fisher Scientific) (0.22-μm-pore-size filter), and the gp120 glycoproteins were purified by the use of nickel affinity columns, as directed by the manufacturer (Invitrogen). Monomeric gp120 was subsequently purified by fast protein liquid chromatography (FPLC), as reported previously (43). The gp120 preparations were dialyzed against phosphate-buffered saline (PBS) and stored in aliquots at −80°C. To assess purity, recombinant proteins were loaded on SDS-PAGE polyacrylamide gels in the absence of beta-mercaptoethanol and stained with Coomassie blue.

The biotin-tagged dimeric recombinant soluble FcγRIIIa (V158) protein was produced and characterized as described previously (37) with additional purification performed using a Superdex 200 10/300 GL column (GE Life Sciences).

ELISA-based rsFcγRIIIa dimer-binding assay.Nonsaturating concentrations of recombinant gp120 glycoproteins in their full-length form (0.4 μg/ml) or lacking the V1, V2, V3, and V5 variable regions (0.25 μg/ml) were prepared in PBS as well as in bovine serum albumin (BSA) (2 μg/ml) as a negative control and were adsorbed to plates (MaxiSorp; Nunc) overnight at 4°C. Coated wells were subsequently blocked with blocking buffer (Tris-buffered saline [TBS] containing 0.1% Tween 20 and 1% [wt/vol] BSA) for 1 h at room temperature. Anti-HIV-1 Env monoclonal antibodies (1 μg/ml) or HIV+ sera (1:1,000 dilution) were diluted in blocking buffer and incubated with the gp120-coated wells for 1 h at room temperature. When indicated, 25 μM BNM-III-170 (or equivalent amounts of DMSO) was diluted with the antibodies. Plates were washed four times with washing buffer (TBS containing 0.1% Tween 20). This was followed by incubation with 0.1 μg/ml purified dimeric rsFcγRIIIa-biotin in blocking buffer for 1 h at room temperature. Horseradish peroxidase (HRP)-conjugated antibody specific for the Fc region of human IgG (Pierce) or high-sensitivity streptavidin-HRP (Thermo Fisher) (1/1,000 in blocking buffer) was added for 1 h at room temperature, followed by four washes. HRP enzyme activity was determined after the addition of a 1:1 mix of Western Lightning oxidizing and luminol reagents (Perkin Elmer Life Sciences). Light emission was measured with an LB 941 TriStar luminometer (Berthold Technologies).

Flow cytometry analysis.Cell surface staining was performed as previously described (19, 21, 45). Cells infected with HIV-1 primary isolates were identified by intracellular staining of HIV-1 p24 using a Cytofix/Cytoperm fixation/permeabilization kit (BD Biosciences) and the phycoerythrin (PE)-conjugated anti-p24 MAb, clone KC57 (Beckman Coulter). The percentage of infected cells (p24+ cells) was determined by gating the living cell population based on viability dye staining with AquaVivid (Thermo Fisher Scientific). Samples were analyzed on an LSRII cytometer (BD Biosciences), and data analysis was performed using FlowJo vX.0.7 (Tree Star).

FACS-based ADCC assay.Measurement of ADCC using the fluorescence-activated cell sorter (FACS)-based assay was performed at 48 h postinfection as previously described (20, 45, 67, 73). Briefly, infected primary CD4+ T cells were stained with AquaVivid viability dye and cell proliferation dye (eFluor670; eBioscience) and used as target cells. Autologous PBMC effector cells, stained with another cellular marker (cell proliferation dye eFluor450; eBioscience), were added at an effector/target ratio of 10:1 in 96-well V-bottom plates (Corning). The indicated concentrations of ADCC-mediating MAbs (0.3125, 0.625, 1.25, 2.5, or 5 μg/ml) with 50 μM BNM-III-170, or equivalent amounts of DMSO, when specified, were added to appropriate wells, and cells were incubated for 15 min at room temperature. The plates were subsequently centrifuged for 1 min at 300 × g and were incubated at 37°C and 5% CO2 for 5 h before being fixed in a 2% PBS–formaldehyde solution. Samples were acquired on an LSRII cytometer (BD Biosciences), and data analysis was performed using FlowJo vX.0.7 (Tree Star). The percentage of ADCC was calculated by gating on infected live target cells with the following formula: (percent p24+ cells in targets plus effectors) − (percent p24+ cells in targets plus effectors plus Abs)/(percent p24+ cells in targets).

In silico modeling.Models of the engagement of dimeric rsFcγRIIIa receptor by anti-CoRBS and anti-cluster A Abs IgGs bound to the target monomeric gp120 immobilized on a plate surface were built based on available crystal structures as follows. The crystal structures of complexes of N5-i5 Fab-gp12093TH057 coree-d1d2CD4 (PDB code 4H8W) (12) and 17b Fab-gp120HXBC2 core-d1d2CD4 (PDB code 1RZJ) (63) were superimposed based on gp120 with the IgG for each antibody built by overlaying murine antibody subclass IgG1 (PDB code 1IGY) (74) onto Fabs from the two complexes. The model was assembled assuming monovalent binding of IgG1, and the dimeric rsFcγRIIIa receptor was built using the crystallographic dimer of FcγRIIa-HR (PDB code 3RY5; high-responder polymorphism FcγRIIa) (66) as a template. The interaction between N5-i5 and 17b Fcs and dimeric rsFcγRIIIa was modeled based on the complex of FcγRIIIa bound to the Fc of IgG1 (PDB code 1E4K) (75).

Statistical analyses.Statistics were analyzed using GraphPad Prism version 6.01 (GraphPad, San Diego, CA, USA). Every data set was tested for statistical normality, and this information was used to apply the appropriate (parametric or nonparametric) statistical test. P values of <0.05 were considered significant (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001).

ACKNOWLEDGMENTS

We thank Dominique Gauchat of the CRCHUM Flow Cytometry Platform for technical assistance and Mario Legault for cohort coordination and clinical samples.

This work was supported by CIHR foundation grant 352417 to A.F. Support for this work was also provided by NIH via NIH grants NIAID R01 to A.F. and M.P. (AI129769) and NIAID R01 to M.P. (AI116274). A.F. is the recipient of a Canada Research Chair on Retroviral Entry (no. RCHS0235). J.R. is the recipient of Mathilde Krim Fellowships in Basic Biomedical Research from amfAR. J.P. is the recipient of a CIHR PhD fellowship award. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We declare that we have no competing interests.

The views expressed in this presentation are those of the authors and do not reflect the official policy or position of the Uniformed Services University, U.S. Army, the Department of Defense, or the U.S. Government.

FOOTNOTES

    • Received 11 October 2018.
    • Accepted 7 November 2018.
    • Accepted manuscript posted online 14 November 2018.
  • Copyright © 2019 American Society for Microbiology.

All Rights Reserved.

REFERENCES

  1. 1.↵
    1. Alpert MD,
    2. Harvey JD,
    3. Lauer WA,
    4. Reeves RK,
    5. Piatak M, Jr,
    6. Carville A,
    7. Mansfield KG,
    8. Lifson JD,
    9. Li W,
    10. Desrosiers RC,
    11. Johnson RP,
    12. Evans DT
    . 2012. ADCC develops over time during persistent infection with live-attenuated SIV and is associated with complete protection against SIV(mac)251 challenge. PLoS Pathog 8:e1002890. doi:10.1371/journal.ppat.1002890.
    OpenUrlCrossRefPubMed
  2. 2.↵
    1. Banks ND,
    2. Kinsey N,
    3. Clements J,
    4. Hildreth JE
    . 2002. Sustained antibody-dependent cell-mediated cytotoxicity (ADCC) in SIV-infected macaques correlates with delayed progression to AIDS. AIDS Res Hum Retroviruses 18:1197–1205. doi:10.1089/08892220260387940.
    OpenUrlCrossRefPubMedWeb of Science
  3. 3.↵
    1. Baum LL,
    2. Cassutt KJ,
    3. Knigge K,
    4. Khattri R,
    5. Margolick J,
    6. Rinaldo C,
    7. Kleeberger CA,
    8. Nishanian P,
    9. Henrard DR,
    10. Phair J
    . 1996. HIV-1 gp120-specific antibody-dependent cell-mediated cytotoxicity correlates with rate of disease progression. J Immunol 157:2168–2173.
    OpenUrlAbstract
  4. 4.↵
    1. Chung AW,
    2. Isitman G,
    3. Navis M,
    4. Kramski M,
    5. Center RJ,
    6. Kent SJ,
    7. Stratov I
    . 2011. Immune escape from HIV-specific antibody-dependent cellular cytotoxicity (ADCC) pressure. Proc Natl Acad Sci U S A 108:7505–7510. doi:10.1073/pnas.1016048108.
    OpenUrlAbstract/FREE Full Text
  5. 5.↵
    1. Forthal DN,
    2. Landucci G,
    3. Haubrich R,
    4. Keenan B,
    5. Kuppermann BD,
    6. Tilles JG,
    7. Kaplan J
    . 1999. Antibody-dependent cellular cytotoxicity independently predicts survival in severely immunocompromised human immunodeficiency virus-infected patients. J Infect Dis 180:1338–1341. doi:10.1086/314988.
    OpenUrlCrossRefPubMedWeb of Science
  6. 6.↵
    1. Mabuka J,
    2. Nduati R,
    3. Odem-Davis K,
    4. Peterson D,
    5. Overbaugh J
    . 2012. HIV-specific antibodies capable of ADCC are common in breastmilk and are associated with reduced risk of transmission in women with high viral loads. PLoS Pathog 8:e1002739. doi:10.1371/journal.ppat.1002739.
    OpenUrlCrossRefPubMed
  7. 7.↵
    1. Sun Y,
    2. Asmal M,
    3. Lane S,
    4. Permar SR,
    5. Schmidt SD,
    6. Mascola JR,
    7. Letvin NL
    . 2011. Antibody-dependent cell-mediated cytotoxicity in simian immunodeficiency virus-infected rhesus monkeys. J Virol 85:6906–6912. doi:10.1128/JVI.00326-11.
    OpenUrlAbstract/FREE Full Text
  8. 8.↵
    1. Haynes BF,
    2. Gilbert PB,
    3. McElrath MJ,
    4. Zolla-Pazner S,
    5. Tomaras GD,
    6. Alam SM,
    7. Evans DT,
    8. Montefiori DC,
    9. Karnasuta C,
    10. Sutthent R,
    11. Liao HX,
    12. DeVico AL,
    13. Lewis GK,
    14. Williams C,
    15. Pinter A,
    16. Fong Y,
    17. Janes H,
    18. DeCamp A,
    19. Huang Y,
    20. Rao M,
    21. Billings E,
    22. Karasavvas N,
    23. Robb ML,
    24. Ngauy V,
    25. de Souza MS,
    26. Paris R,
    27. Ferrari G,
    28. Bailer RT,
    29. Soderberg KA,
    30. Andrews C,
    31. Berman PW,
    32. Frahm N,
    33. De Rosa SC,
    34. Alpert MD,
    35. Yates NL,
    36. Shen X,
    37. Koup RA,
    38. Pitisuttithum P,
    39. Kaewkungwal J,
    40. Nitayaphan S,
    41. Rerks-Ngarm S,
    42. Michael NL,
    43. Kim JH
    . 2012. Immune-correlates analysis of an HIV-1 vaccine efficacy trial. N Engl J Med 366:1275–1286. doi:10.1056/NEJMoa1113425.
    OpenUrlCrossRefPubMedWeb of Science
  9. 9.↵
    1. Bonsignori M,
    2. Pollara J,
    3. Moody MA,
    4. Alpert MD,
    5. Chen X,
    6. Hwang KK,
    7. Gilbert PB,
    8. Huang Y,
    9. Gurley TC,
    10. Kozink DM,
    11. Marshall DJ,
    12. Whitesides JF,
    13. Tsao CY,
    14. Kaewkungwal J,
    15. Nitayaphan S,
    16. Pitisuttithum P,
    17. Rerks-Ngarm S,
    18. Kim JH,
    19. Michael NL,
    20. Tomaras GD,
    21. Montefiori DC,
    22. Lewis GK,
    23. Devico A,
    24. Evans DT,
    25. Ferrari G,
    26. Liao HX,
    27. Haynes BF
    . 2012. Antibody-dependent cellular cytotoxicity-mediating antibodies from an HIV-1 vaccine efficacy trial target multiple epitopes and preferentially use the VH1 gene family. J Virol 86:11521–11532. doi:10.1128/JVI.01023-12.
    OpenUrlAbstract/FREE Full Text
  10. 10.↵
    1. Hogarth PM,
    2. Pietersz GA
    . 2012. Fc receptor-targeted therapies for the treatment of inflammation, cancer and beyond. Nat Rev Drug Discov 11:311–331. doi:10.1038/nrd2909.
    OpenUrlCrossRefPubMed
  11. 11.↵
    1. Wines BD,
    2. Billings H,
    3. McLean MR,
    4. Kent SJ,
    5. Hogarth PM
    . 2017. Antibody functional assays as measures of Fc receptor-mediated immunity to HIV—new technologies and their impact on the HIV vaccine field. Curr HIV Res 15:202–215. doi:10.2174/1570162X15666170320112247.
    OpenUrlCrossRef
  12. 12.↵
    1. Acharya P,
    2. Tolbert WD,
    3. Gohain N,
    4. Wu X,
    5. Yu L,
    6. Liu T,
    7. Huang W,
    8. Huang CC,
    9. Kwon YD,
    10. Louder RK,
    11. Luongo TS,
    12. McLellan JS,
    13. Pancera M,
    14. Yang Y,
    15. Zhang B,
    16. Flinko R,
    17. Foulke JS, Jr,
    18. Sajadi MM,
    19. Kamin-Lewis R,
    20. Robinson JE,
    21. Martin L,
    22. Kwong PD,
    23. Guan Y,
    24. DeVico AL,
    25. Lewis GK,
    26. Pazgier M
    . 2014. Structural definition of an antibody-dependent cellular cytotoxicity response implicated in reduced risk for HIV-1 infection. J Virol 88:12895–12906. doi:10.1128/JVI.02194-14.
    OpenUrlAbstract/FREE Full Text
  13. 13.↵
    1. Howell KA,
    2. Brannan JM,
    3. Bryan C,
    4. McNeal A,
    5. Davidson E,
    6. Turner HL,
    7. Vu H,
    8. Shulenin S,
    9. He S,
    10. Kuehne A,
    11. Herbert AS,
    12. Qiu X,
    13. Doranz BJ,
    14. Holtsberg FW,
    15. Ward AB,
    16. Dye JM,
    17. Aman MJ
    . 2017. Cooperativity enables non-neutralizing antibodies to neutralize Ebolavirus. Cell Rep 19:413–424. doi:10.1016/j.celrep.2017.03.049.
    OpenUrlCrossRef
  14. 14.↵
    1. Carlsen TH,
    2. Pedersen J,
    3. Prentoe JC,
    4. Giang E,
    5. Keck ZY,
    6. Mikkelsen LS,
    7. Law M,
    8. Foung SK,
    9. Bukh J
    . 2014. Breadth of neutralization and synergy of clinically relevant human monoclonal antibodies against HCV genotypes 1a, 1b, 2a, 2b, 2c, and 3a. Hepatology 60:1551–1562. doi:10.1002/hep.27298.
    OpenUrlCrossRefPubMed
  15. 15.↵
    1. Walker LM,
    2. Huber M,
    3. Doores KJ,
    4. Falkowska E,
    5. Pejchal R,
    6. Julien JP,
    7. Wang SK,
    8. Ramos A,
    9. Chan-Hui PY,
    10. Moyle M,
    11. Mitcham JL,
    12. Hammond PW,
    13. Olsen OA,
    14. Phung P,
    15. Fling S,
    16. Wong CH,
    17. Phogat S,
    18. Wrin T,
    19. Simek MD,
    20. Koff WC,
    21. Wilson IA,
    22. Burton DR,
    23. Poignard P
    . 2011. Broad neutralization coverage of HIV by multiple highly potent antibodies. Nature 477:466–470. doi:10.1038/nature10373.
    OpenUrlCrossRefPubMedWeb of Science
  16. 16.↵
    1. He W,
    2. Tan GS,
    3. Mullarkey CE,
    4. Lee AJ,
    5. Lam MM,
    6. Krammer F,
    7. Henry C,
    8. Wilson PC,
    9. Ashkar AA,
    10. Palese P,
    11. Miller MS
    . 2016. Epitope specificity plays a critical role in regulating antibody-dependent cell-mediated cytotoxicity against influenza A virus. Proc Natl Acad Sci U S A 113:11931–11936. doi:10.1073/pnas.1609316113.
    OpenUrlAbstract/FREE Full Text
  17. 17.↵
    1. Ding S,
    2. Veillette M,
    3. Coutu M,
    4. Prevost J,
    5. Scharf L,
    6. Bjorkman PJ,
    7. Ferrari G,
    8. Robinson JE,
    9. Sturzel C,
    10. Hahn BH,
    11. Sauter D,
    12. Kirchhoff F,
    13. Lewis GK,
    14. Pazgier M,
    15. Finzi A
    . 2016. A highly conserved residue of the HIV-1 gp120 inner domain is important for antibody-dependent cellular cytotoxicity responses mediated by anti-cluster A antibodies. J Virol 90:2127–2134. doi:10.1128/JVI.02779-15.
    OpenUrlAbstract/FREE Full Text
  18. 18.↵
    1. Prevost J,
    2. Richard J,
    3. Ding S,
    4. Pacheco B,
    5. Charlebois R,
    6. Hahn BH,
    7. Kaufmann DE,
    8. Finzi A
    . 2018. Envelope glycoproteins sampling states 2/3 are susceptible to ADCC by sera from HIV-1-infected individuals. Virology 515:38–45. doi:10.1016/j.virol.2017.12.002.
    OpenUrlCrossRef
  19. 19.↵
    1. Veillette M,
    2. Coutu M,
    3. Richard J,
    4. Batraville LA,
    5. Dagher O,
    6. Bernard N,
    7. Tremblay C,
    8. Kaufmann DE,
    9. Roger M,
    10. Finzi A
    . 2015. The HIV-1 gp120 CD4-bound conformation is preferentially targeted by antibody-dependent cellular cytotoxicity-mediating antibodies in sera from HIV-1-infected individuals. J Virol 89:545–551. doi:10.1128/JVI.02868-14.
    OpenUrlAbstract/FREE Full Text
  20. 20.↵
    1. Veillette M,
    2. Desormeaux A,
    3. Medjahed H,
    4. Gharsallah NE,
    5. Coutu M,
    6. Baalwa J,
    7. Guan Y,
    8. Lewis G,
    9. Ferrari G,
    10. Hahn BH,
    11. Haynes BF,
    12. Robinson JE,
    13. Kaufmann DE,
    14. Bonsignori M,
    15. Sodroski J,
    16. Finzi A
    . 2014. Interaction with cellular CD4 exposes HIV-1 envelope epitopes targeted by antibody-dependent cell-mediated cytotoxicity. J Virol 88:2633–2644. doi:10.1128/JVI.03230-13.
    OpenUrlAbstract/FREE Full Text
  21. 21.↵
    1. Richard J,
    2. Veillette M,
    3. Brassard N,
    4. Iyer SS,
    5. Roger M,
    6. Martin L,
    7. Pazgier M,
    8. Schön A,
    9. Freire E,
    10. Routy J-P,
    11. Smith AB, III,
    12. Park J,
    13. Jones DM,
    14. Courter JR,
    15. Melillo BN,
    16. Kaufmann DE,
    17. Hahn BH,
    18. Permar SR,
    19. Haynes BF,
    20. Madani N,
    21. Sodroski JG,
    22. Finzi A
    . 2015. CD4 mimetics sensitize HIV-1-infected cells to ADCC. Proc Natl Acad Sci U S A 112:E2687–E2694. doi:10.1073/pnas.1506755112.
    OpenUrlAbstract/FREE Full Text
  22. 22.↵
    1. Madani N,
    2. Princiotto AM,
    3. Mach L,
    4. Ding S,
    5. Prevost J,
    6. Richard J,
    7. Hora B,
    8. Sutherland L,
    9. Zhao CA,
    10. Conn BP,
    11. Bradley T,
    12. Moody MA,
    13. Melillo B,
    14. Finzi A,
    15. Haynes BF,
    16. Smith AB, III,
    17. Santra S,
    18. Sodroski J
    . 2018. A CD4-mimetic compound enhances vaccine efficacy against stringent immunodeficiency virus challenge. Nat Commun 9:2363. doi:10.1038/s41467-018-04758-9.
    OpenUrlCrossRef
  23. 23.↵
    1. Neil SJ,
    2. Zang T,
    3. Bieniasz PD
    . 2008. Tetherin inhibits retrovirus release and is antagonized by HIV-1 Vpu. Nature 451:425–430. doi:10.1038/nature06553.
    OpenUrlCrossRefPubMedWeb of Science
  24. 24.↵
    1. Van Damme N,
    2. Goff D,
    3. Katsura C,
    4. Jorgenson RL,
    5. Mitchell R,
    6. Johnson MC,
    7. Stephens EB,
    8. Guatelli J
    . 2008. The interferon-induced protein BST-2 restricts HIV-1 release and is downregulated from the cell surface by the viral Vpu protein. Cell Host Microbe 3:245–252. doi:10.1016/j.chom.2008.03.001.
    OpenUrlCrossRefPubMedWeb of Science
  25. 25.↵
    1. Veillette M,
    2. Richard J,
    3. Pazgier M,
    4. Lewis GK,
    5. Parsons MS,
    6. Finzi A
    . 2016. Role of HIV-1 envelope glycoproteins conformation and accessory proteins on ADCC responses. Curr HIV Res 14:9–23.
    OpenUrlCrossRefPubMed
  26. 26.↵
    1. Richard J,
    2. Prevost J,
    3. Alsahafi N,
    4. Ding S,
    5. Finzi A
    . 2018. Impact of HIV-1 envelope conformation on ADCC responses. Trends Microbiol 26:253–265. doi:10.1016/j.tim.2017.10.007.
    OpenUrlCrossRef
  27. 27.↵
    1. Forthal DN,
    2. Finzi A
    . 13 November 2018. Antibody-dependent cellular cytotoxicity (ADCC) in HIV infection. AIDS doi:10.1097/QAD.0000000000002011.
    OpenUrlCrossRef
  28. 28.↵
    1. Lee WS,
    2. Richard J,
    3. Lichtfuss M,
    4. Smith AB, III,
    5. Park J,
    6. Courter JR,
    7. Melillo BN,
    8. Sodroski JG,
    9. Kaufmann DE,
    10. Finzi A,
    11. Parsons MS,
    12. Kent SJ
    . 2016. Antibody-dependent cellular cytotoxicity against reactivated HIV-1-infected cells. J Virol 90:2021–2030. doi:10.1128/JVI.02717-15.
    OpenUrlAbstract/FREE Full Text
  29. 29.↵
    1. Richard J,
    2. Pacheco B,
    3. Gohain N,
    4. Veillette M,
    5. Ding S,
    6. Alsahafi N,
    7. Tolbert WD,
    8. Prevost J,
    9. Chapleau JP,
    10. Coutu M,
    11. Jia M,
    12. Brassard N,
    13. Park J,
    14. Courter JR,
    15. Melillo B,
    16. Martin L,
    17. Tremblay C,
    18. Hahn BH,
    19. Kaufmann DE,
    20. Wu X,
    21. Smith AB, III,
    22. Sodroski J,
    23. Pazgier M,
    24. Finzi A
    . 9 September 2016. Co-receptor binding site antibodies enable CD4-mimetics to expose conserved anti-cluster A ADCC epitopes on HIV-1 envelope glycoproteins. EBioMedicine doi:10.1016/j.ebiom.2016.09.004.
    OpenUrlCrossRef
  30. 30.↵
    1. Richard J,
    2. Prevost J,
    3. von Bredow B,
    4. Ding S,
    5. Brassard N,
    6. Medjahed H,
    7. Coutu M,
    8. Melillo B,
    9. Bibollet-Ruche F,
    10. Hahn BH,
    11. Kaufmann DE,
    12. Smith AB, III,
    13. Sodroski J,
    14. Sauter D,
    15. Kirchhoff F,
    16. Gee K,
    17. Neil SJ,
    18. Evans DT,
    19. Finzi A
    . 2017. BST-2 expression modulates small CD4-mimetic sensitization of HIV-1-infected cells to antibody-dependent cellular cytotoxicity. J Virol doi:10.1128/JVI.00219-17.
    OpenUrlAbstract/FREE Full Text
  31. 31.↵
    1. Guan Y,
    2. Pazgier M,
    3. Sajadi MM,
    4. Kamin-Lewis R,
    5. Al-Darmarki S,
    6. Flinko R,
    7. Lovo E,
    8. Wu X,
    9. Robinson JE,
    10. Seaman MS,
    11. Fouts TR,
    12. Gallo RC,
    13. DeVico AL,
    14. Lewis GK
    . 2013. Diverse specificity and effector function among human antibodies to HIV-1 envelope glycoprotein epitopes exposed by CD4 binding. Proc Natl Acad Sci U S A 110:E69–E78. doi:10.1073/pnas.1217609110.
    OpenUrlAbstract/FREE Full Text
  32. 32.↵
    1. Tolbert WD,
    2. Gohain N,
    3. Veillette M,
    4. Chapleau JP,
    5. Orlandi C,
    6. Visciano ML,
    7. Ebadi M,
    8. DeVico AL,
    9. Fouts TR,
    10. Finzi A,
    11. Lewis GK,
    12. Pazgier M
    . 2016. 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. Structure 24:697–709. doi:10.1016/j.str.2016.03.005.
    OpenUrlCrossRef
  33. 33.↵
    1. Tolbert WD,
    2. Gohain N,
    3. Alsahafi N,
    4. Van V,
    5. Orlandi C,
    6. Ding S,
    7. Martin L,
    8. Finzi A,
    9. Lewis GK,
    10. Ray K,
    11. Pazgier M
    . 2017. Targeting the late stage of HIV-1 entry for antibody-dependent cellular cytotoxicity: structural basis for Env epitopes in the C11 region. Structure 25:1719–1731.e4. doi:10.1016/j.str.2017.09.009.
    OpenUrlCrossRef
  34. 34.↵
    1. Finzi A,
    2. Xiang SH,
    3. Pacheco B,
    4. Wang L,
    5. Haight J,
    6. Kassa A,
    7. Danek B,
    8. Pancera M,
    9. Kwong PD,
    10. Sodroski J
    . 2010. Topological layers in the HIV-1 gp120 inner domain regulate gp41 interaction and CD4-triggered conformational transitions. Mol Cell 37:656–667. doi:10.1016/j.molcel.2010.02.012.
    OpenUrlCrossRefPubMedWeb of Science
  35. 35.↵
    1. Ferrari G,
    2. Pollara J,
    3. Kozink D,
    4. Harms T,
    5. Drinker M,
    6. Freel S,
    7. Moody MA,
    8. Alam SM,
    9. Tomaras GD,
    10. Ochsenbauer C,
    11. Kappes JC,
    12. Shaw GM,
    13. Hoxie JA,
    14. Robinson JE,
    15. Haynes BF
    . 2011. An HIV-1 gp120 envelope human monoclonal antibody that recognizes a C1 conformational epitope mediates potent antibody-dependent cellular cytotoxicity (ADCC) activity and defines a common ADCC epitope in human HIV-1 serum. J Virol 85:7029–7036. doi:10.1128/JVI.00171-11.
    OpenUrlAbstract/FREE Full Text
  36. 36.↵
    1. Wyatt R,
    2. Moore J,
    3. Accola M,
    4. Desjardin E,
    5. Robinson J,
    6. Sodroski J
    . 1995. Involvement of the V1/V2 variable loop structure in the exposure of human immunodeficiency virus type 1 gp120 epitopes induced by receptor binding. J Virol 69:5723–5733.
    OpenUrlAbstract/FREE Full Text
  37. 37.↵
    1. Wines BD,
    2. Vanderven HA,
    3. Esparon SE,
    4. Kristensen AB,
    5. Kent SJ,
    6. Hogarth PM
    . 2016. Dimeric FcgammaR ectodomains as probes of the Fc receptor function of anti-influenza virus IgG. J Immunol 197:1507–1516. doi:10.4049/jimmunol.1502551.
    OpenUrlAbstract/FREE Full Text
  38. 38.↵
    1. Lee WS,
    2. Kristensen AB,
    3. Rasmussen TA,
    4. Tolstrup M,
    5. Ostergaard L,
    6. Sogaard OS,
    7. Wines BD,
    8. Hogarth PM,
    9. Reynaldi A,
    10. Davenport MP,
    11. Emery S,
    12. Amin J,
    13. Cooper DA,
    14. Kan VL,
    15. Fox J,
    16. Gruell H,
    17. Parsons MS,
    18. Kent SJ
    . 1 August 2017. Anti-HIV-1 ADCC antibodies following latency reversal and treatment interruption. J Virol doi:10.1128/JVI.00603-17.
    OpenUrlCrossRef
  39. 39.↵
    1. Vanderven HA,
    2. Wragg K,
    3. Ana-Sosa-Batiz F,
    4. Kristensen AB,
    5. Jegaskanda S,
    6. Wheatley AK,
    7. Wentworth D,
    8. Wines BD,
    9. Hogarth PM,
    10. Rockman S, INSIGHT FLU005 Pilot Study Writing Group,
    11. Kent SJ
    . 31 May 2018. Anti-influenza hyperimmune immunoglobulin enhances Fc-functional antibody immunity during human influenza infection. J Infect Dis doi:10.1093/infdis/jiy328.
    OpenUrlCrossRef
  40. 40.↵
    1. McLean MR,
    2. Madhavi V,
    3. Wines BD,
    4. Hogarth PM,
    5. Chung AW,
    6. Kent SJ
    . 2017. Dimeric Fcgamma receptor enzyme-linked immunosorbent assay to study HIV-specific antibodies: a new look into breadth of Fcgamma receptor antibodies induced by the RV144 Vaccine Trial. J Immunol 199:816–826. doi:10.4049/jimmunol.1602161.
    OpenUrlAbstract/FREE Full Text
  41. 41.↵
    1. Melillo B,
    2. Liang S,
    3. Park J,
    4. Schon A,
    5. Courter JR,
    6. LaLonde JM,
    7. Wendler DJ,
    8. Princiotto AM,
    9. Seaman MS,
    10. Freire E,
    11. Sodroski J,
    12. Madani N,
    13. Hendrickson WA,
    14. Smith AB, III.
    2016. Small-molecule CD4-mimics: structure-based optimization of HIV-1 entry inhibition. ACS Med Chem Lett 7:330–334. doi:10.1021/acsmedchemlett.5b00471.
    OpenUrlCrossRefPubMed
  42. 42.↵
    1. Kwon YD,
    2. Finzi A,
    3. Wu X,
    4. Dogo-Isonagie C,
    5. Lee LK,
    6. Moore LR,
    7. Schmidt SD,
    8. Stuckey J,
    9. Yang Y,
    10. Zhou T,
    11. Zhu J,
    12. Vicic DA,
    13. Debnath AK,
    14. Shapiro L,
    15. Bewley CA,
    16. Mascola JR,
    17. Sodroski JG,
    18. Kwong PD
    . 2012. Unliganded HIV-1 gp120 core structures assume the CD4-bound conformation with regulation by quaternary interactions and variable loops. Proc Natl Acad Sci U S A 109:5663–5668. doi:10.1073/pnas.1112391109.
    OpenUrlAbstract/FREE Full Text
  43. 43.↵
    1. Coutu M,
    2. Finzi A
    . 2015. HIV-1 gp120 dimers decrease the overall affinity of gp120 preparations for CD4-induced ligands. J Virol Methods 215–216:37–44. doi:10.1016/j.jviromet.2015.02.017.
    OpenUrlCrossRef
  44. 44.↵
    1. Decker JM,
    2. Bibollet-Ruche F,
    3. Wei X,
    4. Wang S,
    5. Levy DN,
    6. Wang W,
    7. Delaporte E,
    8. Peeters M,
    9. Derdeyn CA,
    10. Allen S,
    11. Hunter E,
    12. Saag MS,
    13. Hoxie JA,
    14. Hahn BH,
    15. Kwong PD,
    16. Robinson JE,
    17. Shaw GM
    . 2005. Antigenic conservation and immunogenicity of the HIV coreceptor binding site. J Exp Med 201:1407–1419. doi:10.1084/jem.20042510.
    OpenUrlAbstract/FREE Full Text
  45. 45.↵
    1. Richard J,
    2. Prevost J,
    3. Baxter AE,
    4. von Bredow B,
    5. Ding S,
    6. Medjahed H,
    7. Delgado GG,
    8. Brassard N,
    9. Sturzel CM,
    10. Kirchhoff F,
    11. Hahn BH,
    12. Parsons MS,
    13. Kaufmann DE,
    14. Evans DT,
    15. Finzi A
    . 2018. Uninfected bystander cells impact the measurement of HIV-specific antibody-dependent cellular cytotoxicity responses. mBio 9:e00358-18. doi:10.1128/mBio.00358-18.
    OpenUrlAbstract/FREE Full Text
  46. 46.↵
    1. Hezareh M,
    2. Hessell AJ,
    3. Jensen RC,
    4. van de Winkel JG,
    5. Parren PW
    . 2001. Effector function activities of a panel of mutants of a broadly neutralizing antibody against human immunodeficiency virus type 1. J Virol 75:12161–12168. doi:10.1128/JVI.75.24.12161-12168.2001.
    OpenUrlAbstract/FREE Full Text
  47. 47.↵
    1. Arduin E,
    2. Arora S,
    3. Bamert PR,
    4. Kuiper T,
    5. Popp S,
    6. Geisse S,
    7. Grau R,
    8. Calzascia T,
    9. Zenke G,
    10. Kovarik J
    . 2015. Highly reduced binding to high and low affinity mouse Fc gamma receptors by L234A/L235A and N297A Fc mutations engineered into mouse IgG2a. Mol Immunol 63:456–463. doi:10.1016/j.molimm.2014.09.017.
    OpenUrlCrossRefPubMed
  48. 48.↵
    1. Hessell AJ,
    2. Hangartner L,
    3. Hunter M,
    4. Havenith CE,
    5. Beurskens FJ,
    6. Bakker JM,
    7. Lanigan CM,
    8. Landucci G,
    9. Forthal DN,
    10. Parren PW,
    11. Marx PA,
    12. Burton DR
    . 2007. Fc receptor but not complement binding is important in antibody protection against HIV. Nature 449:101–104. doi:10.1038/nature06106.
    OpenUrlCrossRefPubMedWeb of Science
  49. 49.↵
    1. Kwong PD,
    2. Doyle ML,
    3. Casper DJ,
    4. Cicala C,
    5. Leavitt SA,
    6. Majeed S,
    7. Steenbeke TD,
    8. Venturi M,
    9. Chaiken I,
    10. Fung M,
    11. Katinger H,
    12. Parren PW,
    13. Robinson J,
    14. Van Ryk D,
    15. Wang L,
    16. Burton DR,
    17. Freire E,
    18. Wyatt R,
    19. Sodroski J,
    20. Hendrickson WA,
    21. Arthos J
    . 2002. HIV-1 evades antibody-mediated neutralization through conformational masking of receptor-binding sites. Nature 420:678–682. doi:10.1038/nature01188.
    OpenUrlCrossRefPubMedWeb of Science
  50. 50.↵
    1. Wyatt R,
    2. Kwong PD,
    3. Desjardins E,
    4. Sweet RW,
    5. Robinson J,
    6. Hendrickson WA,
    7. Sodroski JG
    . 1998. The antigenic structure of the HIV gp120 envelope glycoprotein. Nature 393:705–711. doi:10.1038/31514.
    OpenUrlCrossRefPubMedWeb of Science
  51. 51.↵
    1. Wyatt R,
    2. Sodroski J
    . 1998. The HIV-1 envelope glycoproteins: fusogens, antigens, and immunogens. Science 280:1884–1888. doi:10.1126/science.280.5371.1884.
    OpenUrlAbstract/FREE Full Text
  52. 52.↵
    1. Ma X,
    2. Lu M,
    3. Gorman J,
    4. Terry DS,
    5. Hong X,
    6. Zhou Z,
    7. Zhao H,
    8. Altman RB,
    9. Arthos J,
    10. Blanchard SC,
    11. Kwong PD,
    12. Munro JB,
    13. Mothes W
    . 2018. HIV-1 Env trimer opens through an asymmetric intermediate in which individual protomers adopt distinct conformations. Elife doi:10.7554/eLife.34271.
    OpenUrlCrossRef
  53. 53.↵
    1. Munro JB,
    2. Gorman J,
    3. Ma X,
    4. Zhou Z,
    5. Arthos J,
    6. Burton DR,
    7. Koff WC,
    8. Courter JR,
    9. Smith AB, III,
    10. Kwong PD,
    11. Blanchard SC,
    12. Mothes W
    . 2014. Conformational dynamics of single HIV-1 envelope trimers on the surface of native virions. Science 346:759–763. doi:10.1126/science.1254426.
    OpenUrlAbstract/FREE Full Text
  54. 54.↵
    1. Herschhorn A,
    2. Ma X,
    3. Gu C,
    4. Ventura JD,
    5. Castillo-Menendez L,
    6. Melillo B,
    7. Terry DS,
    8. Smith AB, III,
    9. Blanchard SC,
    10. Munro JB,
    11. Mothes W,
    12. Finzi A,
    13. Sodroski J
    . 2016. Release of gp120 restraints leads to an entry-competent intermediate state of the HIV-1 envelope glycoproteins. mBio 7:e01598-16. doi:10.1128/mBio.01598-16.
    OpenUrlAbstract/FREE Full Text
  55. 55.↵
    1. Kwong PD,
    2. Mascola JR
    . 2012. Human antibodies that neutralize HIV-1: identification, structures, and B cell ontogenies. Immunity 37:412–425. doi:10.1016/j.immuni.2012.08.012.
    OpenUrlCrossRefPubMedWeb of Science
  56. 56.↵
    1. Moody MA,
    2. Gao F,
    3. Gurley TC,
    4. Amos JD,
    5. Kumar A,
    6. Hora B,
    7. Marshall DJ,
    8. Whitesides JF,
    9. Xia SM,
    10. Parks R,
    11. Lloyd KE,
    12. Hwang KK,
    13. Lu X,
    14. Bonsignori M,
    15. Finzi A,
    16. Vandergrift NA,
    17. Alam SM,
    18. Ferrari G,
    19. Shen X,
    20. Tomaras GD,
    21. Kamanga G,
    22. Cohen MS,
    23. Sam NE,
    24. Kapiga S,
    25. Gray ES,
    26. Tumba NL,
    27. Morris L,
    28. Zolla-Pazner S,
    29. Gorny MK,
    30. Mascola JR,
    31. Hahn BH,
    32. Shaw GM,
    33. Sodroski JG,
    34. Liao HX,
    35. Montefiori DC,
    36. Hraber PT,
    37. Korber BT,
    38. Haynes BF
    . 2015. Strain-specific V3 and CD4 binding site autologous HIV-1 neutralizing antibodies select neutralization-resistant viruses. Cell Host Microbe 18:354–362. doi:10.1016/j.chom.2015.08.006.
    OpenUrlCrossRefPubMed
  57. 57.↵
    1. Horwitz JA,
    2. Bar-On Y,
    3. Lu CL,
    4. Fera D,
    5. Lockhart AAK,
    6. Lorenzi JCC,
    7. Nogueira L,
    8. Golijanin J,
    9. Scheid JF,
    10. Seaman MS,
    11. Gazumyan A,
    12. Zolla-Pazner S,
    13. Nussenzweig MC
    . 2017. Non-neutralizing antibodies alter the course of HIV-1 infection in vivo. Cell 170:637–648. e610. doi:10.1016/j.cell.2017.06.048.
    OpenUrlCrossRef
  58. 58.↵
    1. Alsahafi N,
    2. Ding S,
    3. Richard J,
    4. Markle T,
    5. Brassard N,
    6. Walker B,
    7. Lewis GK,
    8. Kaufmann DE,
    9. Brockman MA,
    10. Finzi A
    . 2016. Nef proteins from HIV-1 elite controllers are inefficient at preventing antibody-dependent cellular cytotoxicity. J Virol 90:2993–3002. doi:10.1128/JVI.02973-15.
    OpenUrlAbstract/FREE Full Text
  59. 59.↵
    1. Prevost J,
    2. Zoubchenok D,
    3. Richard J,
    4. Veillette M,
    5. Pacheco B,
    6. Coutu M,
    7. Brassard N,
    8. Parsons MS,
    9. Ruxrungtham K,
    10. Bunupuradah T,
    11. Tovanabutra S,
    12. Hwang KK,
    13. Moody MA,
    14. Haynes BF,
    15. Bonsignori M,
    16. Sodroski J,
    17. Kaufmann DE,
    18. Shaw GM,
    19. Chenine AL,
    20. Finzi A
    . 2017. Influence of the envelope gp120 Phe 43 cavity on HIV-1 sensitivity to antibody-dependent cell-mediated cytotoxicity responses. J Virol doi:10.1128/JVI.02452-16.
    OpenUrlAbstract/FREE Full Text
  60. 60.↵
    1. Prevost J,
    2. Richard J,
    3. Medjahed H,
    4. Alexander A,
    5. Jones J,
    6. Kappes JC,
    7. Ochsenbauer C,
    8. Finzi A
    . 13 June 2018. Incomplete downregulation of CD4 expression affects HIV-1 Env conformation and antibody-dependent cellular cytotoxicity responses. J Virol doi:10.1128/JVI.00484-18.
    OpenUrlAbstract/FREE Full Text
  61. 61.↵
    1. Madhavi V,
    2. Kulkarni A,
    3. Shete A,
    4. Lee WS,
    5. McLean MR,
    6. Kristensen AB,
    7. Ghate M,
    8. Wines BD,
    9. Hogarth PM,
    10. Parsons MS,
    11. Kelleher A,
    12. Cooper DA,
    13. Amin J,
    14. Emery S,
    15. Thakar M,
    16. Kent SJ
    , ENCORE1 Study Group. 2017. Effect of combination antiretroviral therapy on HIV-1-specific antibody-dependent cellular cytotoxicity responses in subtype B- and subtype C-infected cohorts. J Acquir Immune Defic Syndr 75:345–353. doi:10.1097/QAI.0000000000001380.
    OpenUrlCrossRef
  62. 62.↵
    1. Gohain N,
    2. Tolbert WD,
    3. Acharya P,
    4. Yu L,
    5. Liu T,
    6. Zhao P,
    7. Orlandi C,
    8. Visciano ML,
    9. Kamin-Lewis R,
    10. Sajadi MM,
    11. Martin L,
    12. Robinson JE,
    13. Kwong PD,
    14. DeVico AL,
    15. Ray K,
    16. Lewis GK,
    17. Pazgier M
    . 2015. Cocrystal structures of antibody N60-i3 and antibody JR4 in complex with gp120 define more cluster A epitopes involved in effective antibody-dependent effector function against HIV-1. J Virol 89:8840–8854. doi:10.1128/JVI.01232-15.
    OpenUrlAbstract/FREE Full Text
  63. 63.↵
    1. Huang CC,
    2. Venturi M,
    3. Majeed S,
    4. Moore MJ,
    5. Phogat S,
    6. Zhang MY,
    7. Dimitrov DS,
    8. Hendrickson WA,
    9. Robinson J,
    10. Sodroski J,
    11. Wyatt R,
    12. Choe H,
    13. Farzan M,
    14. Kwong PD
    . 2004. Structural basis of tyrosine sulfation and VH-gene usage in antibodies that recognize the HIV type 1 coreceptor-binding site on gp120. Proc Natl Acad Sci U S A 101:2706–2711. doi:10.1073/pnas.0308527100.
    OpenUrlAbstract/FREE Full Text
  64. 64.↵
    1. Zhang Y,
    2. Guo J,
    3. Huang L,
    4. Tian J,
    5. Yao X,
    6. Liu H
    . 2018. The molecular mechanism of two coreceptor binding site antibodies X5 and 17b neutralizing HIV-1: insights from molecular dynamics simulation. Chem Biol Drug Des 92:1357–1365. doi:10.1111/cbdd.13201.
    OpenUrlCrossRef
  65. 65.↵
    1. Huang CC,
    2. Tang M,
    3. Zhang MY,
    4. Majeed S,
    5. Montabana E,
    6. Stanfield RL,
    7. Dimitrov DS,
    8. Korber B,
    9. Sodroski J,
    10. Wilson IA,
    11. Wyatt R,
    12. Kwong PD
    . 2005. Structure of a V3-containing HIV-1 gp120 core. Science 310:1025–1028. doi:10.1126/science.1118398.
    OpenUrlAbstract/FREE Full Text
  66. 66.↵
    1. Ramsland PA,
    2. Farrugia W,
    3. Bradford TM,
    4. Sardjono CT,
    5. Esparon S,
    6. Trist HM,
    7. Powell MS,
    8. Tan PS,
    9. Cendron AC,
    10. Wines BD,
    11. Scott AM,
    12. Hogarth PM
    . 2011. Structural basis for Fc gammaRIIa recognition of human IgG and formation of inflammatory signaling complexes. J Immunol 187:3208–3217. doi:10.4049/jimmunol.1101467.
    OpenUrlAbstract/FREE Full Text
  67. 67.↵
    1. Richard J,
    2. Veillette M,
    3. Batraville LA,
    4. Coutu M,
    5. Chapleau JP,
    6. Bonsignori M,
    7. Bernard N,
    8. Tremblay C,
    9. Roger M,
    10. Kaufmann DE,
    11. Finzi A
    . 2014. Flow cytometry-based assay to study HIV-1 gp120 specific antibody-dependent cellular cytotoxicity responses. J Virol Methods 208:107–114. doi:10.1016/j.jviromet.2014.08.003.
    OpenUrlCrossRefPubMed
  68. 68.↵
    1. Klein JS,
    2. Bjorkman PJ
    . 2010. Few and far between: how HIV may be evading antibody avidity. PLoS Pathog 6:e1000908. doi:10.1371/journal.ppat.1000908.
    OpenUrlCrossRefPubMed
  69. 69.↵
    1. Fontaine J,
    2. Chagnon-Choquet J,
    3. Valcke HS,
    4. Poudrier J,
    5. Roger M
    . 2011. High expression levels of B lymphocyte stimulator (BLyS) by dendritic cells correlate with HIV-related B-cell disease progression in humans. Blood 117:145–155. doi:10.1182/blood-2010-08-301887.
    OpenUrlAbstract/FREE Full Text
  70. 70.↵
    1. Fontaine J,
    2. Coutlee F,
    3. Tremblay C,
    4. Routy JP,
    5. Poudrier J,
    6. Roger M
    . 2009. HIV infection affects blood myeloid dendritic cells after successful therapy and despite nonprogressing clinical disease. J Infect Dis 199:1007–1018. doi:10.1086/597278.
    OpenUrlCrossRefPubMed
  71. 71.↵
    1. Peretz Y,
    2. Ndongala ML,
    3. Boulet S,
    4. Boulassel MR,
    5. Rouleau D,
    6. Cote P,
    7. Longpre D,
    8. Routy JP,
    9. Falutz J,
    10. Tremblay C,
    11. Tsoukas CM,
    12. Sekaly RP,
    13. Bernard NF
    . 2007. Functional T cell subsets contribute differentially to HIV peptide-specific responses within infected individuals: correlation of these functional T cell subsets with markers of disease progression. Clin Immunol 124:57–68. doi:10.1016/j.clim.2007.04.004.
    OpenUrlCrossRefPubMed
  72. 72.↵
    1. Kamya P,
    2. Boulet S,
    3. Tsoukas CM,
    4. Routy JP,
    5. Thomas R,
    6. Cote P,
    7. Boulassel MR,
    8. Baril JG,
    9. Kovacs C,
    10. Migueles SA,
    11. Connors M,
    12. Suscovich TJ,
    13. Brander C,
    14. Tremblay CL,
    15. Bernard N
    . 2011. Receptor-ligand requirements for increased NK cell polyfunctional potential in slow progressors infected with HIV-1 coexpressing KIR3DL1*h/*y and HLA-B*57. J Virol 85:5949–5960. doi:10.1128/JVI.02652-10.
    OpenUrlAbstract/FREE Full Text
  73. 73.↵
    1. Richard J,
    2. Veillette M,
    3. Ding S,
    4. Zoubchenok D,
    5. Alsahafi N,
    6. Coutu M,
    7. Brassard N,
    8. Park J,
    9. Courter JR,
    10. Melillo B,
    11. Smith AB, III,
    12. Shaw GM,
    13. Hahn BH,
    14. Sodroski J,
    15. Kaufmann DE,
    16. Finzi A
    . 2016. Small CD4 mimetics prevent HIV-1 uninfected bystander CD4 + T cell killing mediated by antibody-dependent cell-mediated cytotoxicity. EBioMedicine 3:122–134. doi:10.1016/j.ebiom.2015.12.004.
    OpenUrlCrossRefPubMed
  74. 74.↵
    1. Harris LJ,
    2. Skaletsky E,
    3. McPherson A
    . 1998. Crystallographic structure of an intact IgG1 monoclonal antibody. J Mol Biol 275:861–872. doi:10.1006/jmbi.1997.1508.
    OpenUrlCrossRefPubMedWeb of Science
  75. 75.↵
    1. Sondermann P,
    2. Huber R,
    3. Oosthuizen V,
    4. Jacob U
    . 2000. The 3.2-A crystal structure of the human IgG1 Fc fragment-Fc gammaRIII complex. Nature 406:267–273. doi:10.1038/35018508.
    OpenUrlCrossRefPubMedWeb of Science
  76. 76.↵
    1. von Bredow B,
    2. Arias JF,
    3. Heyer LN,
    4. Gardner MR,
    5. Farzan M,
    6. Rakasz EG,
    7. Evans DT
    . 2015. Envelope glycoprotein internalization protects human and simian immunodeficiency virus-infected cells from antibody-dependent cell-mediated cytotoxicity. J Virol 89:10648–10655. doi:10.1128/JVI.01911-15.
    OpenUrlAbstract/FREE Full Text
PreviousNext
Back to top
Download PDF
Citation Tools
Two Families of Env Antibodies Efficiently Engage Fc-Gamma Receptors and Eliminate HIV-1-Infected Cells
Sai Priya Anand, Jérémie Prévost, Sophie Baril, Jonathan Richard, Halima Medjahed, Jean-Philippe Chapleau, William D. Tolbert, Sharon Kirk, Amos B. Smith III, Bruce D. Wines, Stephen J. Kent, P. Mark Hogarth, Matthew S. Parsons, Marzena Pazgier, Andrés Finzi
Journal of Virology Jan 2019, 93 (3) e01823-18; DOI: 10.1128/JVI.01823-18

Citation Manager Formats

  • BibTeX
  • Bookends
  • EasyBib
  • EndNote (tagged)
  • EndNote 8 (xml)
  • Medlars
  • Mendeley
  • Papers
  • RefWorks Tagged
  • Ref Manager
  • RIS
  • Zotero
Print

Alerts
Sign In to Email Alerts with your Email Address
Email

Thank you for sharing this Journal of Virology article.

NOTE: We request your email address only to inform the recipient that it was you who recommended this article, and that it is not junk mail. We do not retain these email addresses.

Enter multiple addresses on separate lines or separate them with commas.
Two Families of Env Antibodies Efficiently Engage Fc-Gamma Receptors and Eliminate HIV-1-Infected Cells
(Your Name) has forwarded a page to you from Journal of Virology
(Your Name) thought you would be interested in this article in Journal of Virology.
CAPTCHA
This question is for testing whether or not you are a human visitor and to prevent automated spam submissions.
Share
Two Families of Env Antibodies Efficiently Engage Fc-Gamma Receptors and Eliminate HIV-1-Infected Cells
Sai Priya Anand, Jérémie Prévost, Sophie Baril, Jonathan Richard, Halima Medjahed, Jean-Philippe Chapleau, William D. Tolbert, Sharon Kirk, Amos B. Smith III, Bruce D. Wines, Stephen J. Kent, P. Mark Hogarth, Matthew S. Parsons, Marzena Pazgier, Andrés Finzi
Journal of Virology Jan 2019, 93 (3) e01823-18; DOI: 10.1128/JVI.01823-18
del.icio.us logo Digg logo Reddit logo Twitter logo CiteULike logo Facebook logo Google logo Mendeley logo
  • Top
  • Article
    • ABSTRACT
    • INTRODUCTION
    • RESULTS
    • DISCUSSION
    • MATERIALS AND METHODS
    • ACKNOWLEDGMENTS
    • FOOTNOTES
    • REFERENCES
  • Figures & Data
  • Info & Metrics
  • PDF

KEYWORDS

ADCC
CD4-mimetics
Env
FcγRIIIa
HIV-1
anti-cluster A antibodies
anti-coreceptor binding site antibodies
nonneutralizing antibodies

Related Articles

Cited By...

About

  • About JVI
  • Editor in Chief
  • Editorial Board
  • Policies
  • For Reviewers
  • For the Media
  • For Librarians
  • For Advertisers
  • Alerts
  • RSS
  • FAQ
  • Permissions
  • Journal Announcements

Authors

  • ASM Author Center
  • Submit a Manuscript
  • Article Types
  • Ethics
  • Contact Us

Follow #Jvirology

@ASMicrobiology

       

 

JVI in collaboration with

American Society for Virology

ASM Journals

ASM journals are the most prominent publications in the field, delivering up-to-date and authoritative coverage of both basic and clinical microbiology.

About ASM | Contact Us | Press Room

 

ASM is a member of

Scientific Society Publisher Alliance

 

American Society for Microbiology
1752 N St. NW
Washington, DC 20036
Phone: (202) 737-3600

Copyright © 2021 American Society for Microbiology | Privacy Policy | Website feedback

Print ISSN: 0022-538X; Online ISSN: 1098-5514