Enhanced phagocytic activity of HIV-specific antibodies correlates with natural production of immunoglobulins with skewed affinity for FcγR2a and FcγR2b - PubMed (original) (raw)

Enhanced phagocytic activity of HIV-specific antibodies correlates with natural production of immunoglobulins with skewed affinity for FcγR2a and FcγR2b

Margaret E Ackerman et al. J Virol. 2013 May.

Abstract

While development of an HIV vaccine that can induce neutralizing antibodies remains a priority, decades of research have proven that this is a daunting task. However, accumulating evidence suggests that antibodies with the capacity to harness innate immunity may provide some protection. While significant research has focused on the cytolytic properties of antibodies in acquisition and control, less is known about the role of additional effector functions. In this study, we investigated antibody-dependent phagocytosis of HIV immune complexes, and we observed significant differences in the ability of antibodies from infected subjects to mediate this critical effector function. We observed both quantitative differences in the capacity of antibodies to drive phagocytosis and qualitative differences in their FcγR usage profile. We demonstrate that antibodies from controllers and untreated progressors exhibit increased phagocytic activity, altered Fc domain glycosylation, and skewed interactions with FcγR2a and FcγR2b in both bulk plasma and HIV-specific IgG. While increased phagocytic activity may directly influence immune activation via clearance of inflammatory immune complexes, it is also plausible that Fc receptor usage patterns may regulate the immune response by modulating downstream signals following phagocytosis--driving passive degradation of internalized virus, release of immune modulating cytokines and chemokines, or priming of a more effective adaptive immune response.

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Figures

Fig 1

Fig 1

Differential effector function of antibodies from HIV-positive subjects. (A and B) ADCVI activities of antibodies from different subject groups in assays utilizing primary monocytes (A) or NK cells (B) as effectors. (C) Binding titers (EC50, in μg/ml) of anti-envelope antibodies. (D) Antibody neutralization as determined by viral inhibition in the absence of effector cells. **, P < 0.005; ***, P < 0.0005.

Fig 2

Fig 2

Differential phagocytic function measured at high throughput. (A and B) Representative histograms of bead uptake by THP-1 cells for antibody samples with low (A) and high (B) phagocytosis activity. (C) Representative phagocytosis dose-response curves for HIV-negative subjects (neg), controllers (ctr), untreated subjects (un), and treated subjects (tx). Phagocytic activity is presented as iMFI. (D) Phagocytic potency of antibodies from HIV-infected subjects. Reciprocal log PC50s (1/concentration at which half-maximal phagocytosis was observed) for gp120-coated bead uptake by a monocytic cell line are shown. (E and F) Correlation of viral inhibition in primary monocytes (E) or NK cells (F) with phagocytic potential (1/PC50) determined by the THP-1 phagocytosis assay. **, P < 0.005; ***, P < 0.0005.

Fig 3

Fig 3

Phagocytic dependence on specific FcγR. (A) Phagocytosis by cells pretreated with FcγR2a-, FcγR2b-, and FcγR3a-blocking antibodies or left untreated was determined. The ratio of bead uptake under blocked and unblocked conditions is presented. The effect of blocking FcγR3a (B), FcγR2a (C), and FcγR2b (D) was determined within each subject class, exposing differences between patient groups in reliance on these receptors. *, P < 0.05; ***, P < 0.0005.

Fig 4

Fig 4

Plasma IgG exhibits differential phagocytosis activity among subject groups. (A to C) Representative flow cytometric dot plot (A) and histograms (B and C) of competitive uptake of green fluorescent beads (B) coated with bulk antibody from a low-activity subject relative to uptake of red fluorescent beads (C) opsonized with bulk antibody from a high-activity subject. (D and E) Beads opsonized with bulk antibody from negative subjects were competed against other HIV-negative subjects (Neg), chronically infected subjects (Chr), or controllers (Ctr) (D) or all other subjects (E). Data presented are ratios of phagocytosis of red to green beads. *, P < 0.05; **, P < 0.005; ***, P < 0.0005.

Fig 5

Fig 5

Plasma antibody subclass and glycosylation. (A to D) Fraction of plasma IgG of the IgG1 (A), IgG2 (B), IgG3 (C), and IgG4 (D) subclasses. (E and F) Plasma IgG glycosylation as determined by ELISA for fucose (E) or terminal mannose (F). OD, optical density. *, P < 0.05; **, P < 0.005; ***, P < 0.0005.

Fig 6

Fig 6

Differential phagocytosis is associated with binding to FcγR2. (A) Ratio of binding to the activating FcγR2a relative to the inhibitory FcγR2b as determined by ELISA. (B) Correlation of log phagocytic potency (1/PC50) with binding to FcγR2a/FcγR2b (Biacore analysis). *, P < 0.05; **, P < 0.005.

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