Monoclonal Antibodies Specific for the V2, V3, CD4-Binding Site, and g

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INTRODUCTION

The RV144 vaccine trial was the first clinical study to achieve any degree of protection, reaching a modest efficacy level of 31.2% in HIV vaccine recipients (1, 2). The study revealed that a high titer of plasma anti-V2 antibodies (Abs), particularly immunoglobulin G3 (IgG3) Abs, correlated with a reduced risk of HIV-1 infection (2, 3). The protective V1-V2 IgG3 response was shown to wane quickly, corresponding to the waning vaccine efficacy (4). The concept of a possible role of anti-V2 Abs in protection from HIV-1 infection was further supported by sieve analysis of virus sequences derived from recipients of the RV144 vaccine or a placebo, which suggested that anti-V2 Abs exerted immune pressure on viruses with particular sequence signatures in V2 (5). These results stimulated numerous nonhuman primate studies exploring the potential role of V2 Abs in vaccine-elicited protection. Results revealing serum binding of Abs to V2 were correlated with delayed simian immunodeficiency virus (SIV) acquisition following vaccination with a DNA/Ad5-recombinant regimen and intrarectal challenge (6). High-avidity Abs with respect to the V1-V2 region elicited by an SIV human papillomavirus pseudovirion-based vaccine approach correlated with delayed SIV acquisition following repeated low-dose vaginal challenges (7). Protection against intrarectal SIV acquisition following sequential administration of adenovirus- or adenovirus/poxvirus-based vaccines was correlated with serum V2 binding Abs (8). Recently, an SIV rhesus macaque study mimicking the RV144 vaccine regimen demonstrated a correlation of protective efficacy with mucosal IgG Abs against the V2 region (9). Together with the results of the RV144 trial, these repeated findings associating V2 Abs with protection suggest that this immune response has a significant impact on HIV/SIV acquisition.

However, the mechanism by which anti-V2 Abs might exert a protective effect is unknown. In the RV144 trial, neutralizing Abs (nAbs) were not correlated with protection (2, 10). Further, although monoclonal Abs (MAbs) specific for the V2 region are extensively cross-reactive with gp120s derived from subtypes A, B, and C (11), they exhibit weak neutralizing activities and do so only against tier 1 viruses (11,–15). Thus, investigations have focused on nonneutralizing Ab activities. One possible mechanism is the inhibition of gp120 interaction with the α4β7 integrin expressed on Th17 cells by anti-V2 Abs. This interaction is believed to facilitate infection of activated mucosal CD4⁺ T cells. V2 MAbs target an epitope that overlaps a conserved three-amino-acid motif in the V2 region that is the binding site for α4β7 integrin and may therefore block gp120 interaction with this integrin (12, 16). In support of this hypothesis, mouse anti-V2 MAbs have been shown to inhibit binding of biotinylated gp120_MN to the α4β7⁺ β1⁻ RPMI-8866 B lymphoma cell line (14). Two human anti-V2 MAbs from our laboratory, 697 and 2158, also inhibited binding of gp120_MN to CD4⁺ cells and activated CD8⁺ cells (17). The extent to which such blocking by V2-specific Abs occurs in vivo is not known; however, it has been reported that the envelopes of most HIV strains do not bind to α4β7 (18), a result that may be dependent upon the nature of the carbohydrate residues on the Env protein. The absence of complex carbohydrates on the viral envelope together with enriched oligomannose-type glycans results in greater binding to α4β7 (19). Thus, more experiments are required to determine if blocking of Env-α4β7 binding contributes to the protective function of anti-V2 MAbs.

A spectrum of Fc-mediated nonneutralizing Ab activities have been associated with HIV and SIV vaccine protective efficacy. These include Ab-dependent cell-mediated viral inhibition (ADCVI) (20,–24), Ab-dependent cellular cytotoxicity (ADCC) (22,–28), Ab-dependent cellular phagocytosis (ADCP) (29), and Ab-dependent complement deposition (ADCD) (29). ADCP and ADCD have undergone extensive investigation only recently. In fact, as state-of-the-art high-throughput technologies are now in use, nonneutralizing Ab activities can be grouped, and protective correlations with polyfunctional Ab activities have been demonstrated (30). With regard to anti-V2 MAbs, a few have been shown to mediate low-level ADCC activity. CH58 and CH59, derived from recipients of the RV144 vaccine, and 697 and 2158, derived from chronically infected individuals, displayed specific killing of virus-infected cells and of target cells pulsed with gp140_SF162, respectively (15, 31). In general, however, using sera of HIV-infected people, the variable loops of the viral envelope have been reported not to represent a major ADCC determinant (32). MAbs 697 and 2158 have also been shown to mediate ADCP (31). ADCD by V2 MAbs has not, to our knowledge, been assessed.

In light of the weak or absent neutralizing and ADCC activity mediated by anti-V2 MAbs, we investigated a panel of V2 MAbs for their ability to mediate ADCP, a mechanism potentially associated with protective efficacy. ADCP may contribute more to vaccine-elicited protection than ADCC, particularly in mucosal tissues (33). ADCP was enhanced in recipients of the RV144 vaccine with a high level of IgG3/IgG1 Abs to V1-V2 (34) and was associated with protection in nonhuman primates (29). As ADCP was shown to be a prominent activity of the MAb V2 panel, we further compared it with that of other MAbs specific for V3, the CD4bs, and gp41. All four categories of MAbs mediated ADCP activity. The levels of activity differed between the panels of MAbs and also depended on the target antigens displayed on the beads, with dominant activity mediated by anti-gp41 and some anti-V2 MAbs.

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RESULTS

Titration of biotinylated gp120 for coating the beads.

The phagocytic assay used here is based on the ability of an Ab to recognize antigen-coated beads and mediate phagocytosis by THP-1 cells. Given that the amount of gp120 bound to the surface of the beads is an important factor for assay sensitivity, we titrated the biotinylated recombinant gp120_BaL used for coating from 30 to 0.0015 μg/ml using 3-fold dilutions (Fig. 1). The phagocytic activity mediated by human anti-V2 MAb 2158, but not that by the irrelevant human MAb 1418 against parvovirus B19 as a negative control, was dose dependent. There was some plateauing of phagocytic activity, corresponding to maximal activity at 3.3 to 30.0 μg/ml. Thereafter, we used 1.7 μg/ml for coating the beads with gp120 whereas gp41 was used at 0.6 μg/ml to match the molecular weight of gp120.

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FIG 1

Titration of gp120_BaL coated onto beads. Ten serial 3-fold dilutions of recombinant gp120_BaL were used for coating the beads to test phagocytosis mediated by MAb 2158 (anti-V2) and negative-control MAb 1418 (anti-parvovirus B19). A suboptimal dose of 1.7 μg/ml of gp120 was used for coating the beads in the study.

Binding of MAbs to gp120 and gp41.

All 27 MAbs (Table 1) were screened by enzyme-linked immunosorbent assay (ELISA) for their reactivity to gp120 (20 MAbs), SOSIP (27 MAbs), or gp41 (7 MAbs) to confirm their binding specificity. The majority of MAbs displayed strong binding to the respective proteins, with a few exceptions: 697 (anti-V2), 447-52D (anti-V3, GPGR specific), and 654 (anti-CD4bs) bound relatively weakly to gp120_ZM109, while some anti-gp41 MAbs bound weakly to SOSIP (Table 2).

TABLE 1

List of human anti-HIV-1 gp120 and gp41 MAbs used in the study

MAb no. or description	MAb designation	Specificity	Isotype	Reference
1	697	V2	IgG1(λ)	12
2	830A	V2	IgG3(κ)	35
3	1357	V2	IgG1(κ)	36
4	1361	V2	IgG1(κ)	36
5	1393A	V2	IgG1(κ)	35
6	2158	V2	IgG1(κ)	37
7	447-52D	V3	IgG3(λ)	38
8	2191	V3	IgG1(λ)	39
9	2219	V3	IgG1(λ)	39
10	2442	V3	IgG1(λ)	39
11	2558	V3	IgG1(λ)	40
12	3074	V3	IgG1(λ)	41
13	3869	V3	IgG1(λ)	42
14	4682	V3	IgG1(λ)	43
15	559/64-D	CD4bs	IgG1(κ)	44
16	654-D	CD4bs	IgG1(λ)	45
17	729-D	CD4bs	IgG1(κ)	46
18	1202	CD4bs	IgG1(κ)	45
19	1331E	CD4bs	IgG1(κ)	36
20	1570	CD4bs	IgG1(λ)	47
21	126-7	gp41	IgG1(λ)	48
22	167	gp41	IgG1(λ)	48
23	240	gp41	IgG1(κ)	48
24	246	gp41	IgG1(κ)	48
25	1281	gp41	IgG1(λ)	36
26	1342	gp41	IgG1(λ)	36
27	1367	gp41	IgG1(λ)	36
Ctr.^a	1418	Parvovirus B19	IgG1(κ)	49

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^aCtr., control.

TABLE 2

ADCP and binding activities of MAbs tested in the study^a

Site and MAb no.	MAb designation	ADCP score					ELISA OD
Site and MAb no.	MAb designation	gp120_BaL	gp120_ZM109	SOSIP	DS-SOSIP	BSA	gp120_BaL	gp120_ZM109	SOSIP	BSA
V2
1	697	1.13	1.20	1.06	1.14	0.90	3.5	0.7	3.9	0.1
2	830A	1.34	1.36	1.11	1.23	0.99	3.4	3.5	3.8	0.1
3	1357	1.29	1.25	1.06	1.02	1.02	3.4	3.1	3.5	0.1
4	1361	1.30	1.30	1.10	1.17	0.98	3.4	3.4	3.5	0.1
5	1393A	1.24	1.27	1.11	1.20	0.97	3.5	3.6	2.9	0.1
6	2158	1.24	1.36	1.12	1.28	0.96	3.4	3.5	3.6	0.1
V3
7	447-52D	1.39	1.28	1.06	1.56	0.98	3.5	1.6	3.8	0.1
8	2191	1.19	1.21	1.07	1.20	0.97	3.4	3.5	4.1	0.1
9	2219	1.18	1.14	1.10	1.20	1.03	3.6	3.8	3.5	0.1
10	2442	1.11	1.18	1.09	1.23	1.08	3.5	3.0	3.7	0.1
11	2558	1.15	1.11	1.07	1.24	0.92	3.6	3.7	3.8	0.1
12	3074	1.23	1.05	1.13	1.36	1.00	3.6	3.6	3.7	0.1
13	3869	1.09	1.10	1.01	1.19	0.89	3.5	3.5	4	0.1
14	4682	1.12	1.12	1.10	1.25	1.01	3.5	3.6	4.2	0.1
CD4bs
15	559/64	1.04	1.09	1.01	1.08	0.94	3.3	2.6	3.5	0.1
16	654	1.22	1.13	0.98	1.10	0.93	3.6	0.4	2.7	0.1
17	729	1.12	1.14	1.00	1.12	0.99	3.6	3.6	3.2	0.1
18	1202	1.12	1.14	0.98	1.04	1.04	3.5	3.5	2.4	0.1
19	1331E	1.33	1.19	1.04	1.14	1.02	3.7	3.6	3.8	0.1
20	1570	1.21	1.08	1.02	1.26	0.95	3.5	3.5	4.1	0.1
gp41		gp41_ZA.1197	gp41_HXB2	SOSIP	DS-SOSIP	BSA	gp41_ZA.1197	gp41_HXB2	SOSIP	BSA
21	126-7	1.31	1.80	0.94	0.83	1.09	3.6	3.7	1.2	0.1
22	167	1.34	1.45	1.06	1.24	0.96	3.3	3.5	1.7	0.1
23	240	1.35	1.78	1.02	1.20	1.01	3.6	3.6	3.7	0.1
24	246	1.38	1.52	1.06	1.07	0.88	3.6	3.5	3.7	0.1
25	1281	1.23	1.74	1.08	1.21	1.03	3.8	3.9	1.4	0.1
26	1342	1.36	1.40	1.09	1.17	0.99	3.8	3.8	1.3	0.1
27	1367	1.32	1.53	1.00	1.20	0.92	3.6	3.5	0.9	0.1

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^aCells with boldface highlighting indicate the ADCP scores and ELISA optical density (OD) values above the cutoff, which was determined by averaging the BSA values plus 2× the standard deviation (SD). ADCP scores are shown at the peak of activity at 0.2 μg/ml.

Phagocytic activity of MAbs against gp120-, gp41-, and SOSIP-coated beads.

The main goal of this study was to determine whether the phagocytic activity mediated by human anti-V2 MAbs has any special characteristics compared to that mediated by MAbs against V3, CD4bs, and gp41 which could explain the possible protective function of these V2 Abs observed in the RV144 vaccine clinical trial (2). We tested 25 IgG1 and two IgG3 MAbs (Table 1) produced by hybridoma cell lines which preserved the Fc fragments of the original donors. The V2, V3, and CD4bs MAbs were first tested against two recombinant gp120 proteins (BaL [clade B] and ZM109F.PB4 [clade C]) for phagocytic activity, while the gp41 MAbs were tested against two gp41 proteins (HXB2 [clade B] and ZA.1197 [clade C]) (Fig. 2). The recombinant proteins were produced in 293T cells which preserved typical human glycosylation. All MAbs were titrated at between 1 and 0.0016 μg/ml, and the majority mediated phagocytosis with a peak of activity at 0.2 μg/ml (Fig. 2; Table 2). All MAbs were reactive with the antigens and exhibited strong binding to gp120, gp41, and SOSIP (Table 2).

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FIG 2

Phagocytosis activity of human anti-HIV-1 envelope MAbs. Four panels of MAbs against V2, V3, CD4bs, and gp41 were tested in five dilutions starting at 1 μg/ml. The beads were coated with gp120_BaL (A to C) or gp41_HXB2 (D) (both clade B) or with gp120_ZM109F.PB4 (E to G) or gp41_ZA.1197MB (H) (both clade C). All MAbs were IgG1, with the exception of two IgG3 MAbs, 830A and 447 (labeled γ3). Phagocytosis data indicate fold increases of scores over those determined for beads coated with BSA.

To further evaluate the ADCP activity of MAbs against beads coated with proteins which mimic the trimers on the virus surface, we utilized two recombinant proteins: BG505 SOSIP.664 and its version with a stabilized conformation (201C 433C variant), BG505 DS-SOSIP.664 gp140 (coated on the beads at 1.7 μg/ml). V2 and V3 MAbs exhibited higher ADCP activity against the SOSIP trimer than the anti-CD4bs MAbs (Fig. 3A to toC).C). Interestingly, V2 and V3 MAbs mediated better ADCP against DS-SOSIP (Fig. 3E and andF),F), the version of the trimer that does not change conformation upon interaction with CD4 (50). CD4bs MAbs again mediated lower ADCP than the V2 and V3 MAbs, although some exhibited moderate activity against DS-SOSIP (Fig. 3G). Only some of the gp41 MAbs mediated ADCP activity against SOSIP-coated beads (Fig. 3D), but most showed higher levels against DS-SOSIP (Fig. 3H). Overall, the ADCP activity levels of the gp41 MAbs were much lower against these trimer mimics than against gp41 proteins (Fig. 2D and andHH).

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FIG 3

Phagocytosis activity of MAbs against BG505 SOSIP.664 and BG505 DS-SOSIP.664 gp140 trimers. Four panels of MAbs against V2, V3, CD4bs, and gp41 (from left to right) were tested using beads coated with BG505 SOSIP.664 (A to D) and with BG505 DS-SOSIP.664 (E to H). All MAbs were IgG1, except two IgG3(γ3) MAbs (labeled in black in panels A, B, E, and F).

In order to evaluate the differences in ADCP activity mediated by the various MAbs, analyses of the area under the curve (AUC) were conducted (Fig. 4). The ADCP activity of anti-V2 MAbs was comparable to the activity of anti-V3 and anti-CD4bs in the assay using beads coated with gp120_BaL (Fig. 4A) and was significantly higher (P < 0.009) using gp120_ZM109F.PB4 (Fig. 4B). The anti-gp41 MAbs were tested against two gp41 proteins representing strains that were different from the gp120s used to coat the beads but also belonging to either clade B (HXB2) (Fig. 4A) or clade C (ZA.1197) (Fig. 4B). The ADCP activities of anti-gp41 MAbs were significantly higher than those of the V2, V3, and CD4bs MAbs tested against gp120 from clade B and clade C (P < 0.006) (Fig. 4A and andBB).

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FIG 4

Area under the curve analysis of ADCP activity of MAbs. Panels of IgG1 MAbs were tested against beads coated with gp120_BaL and gp41_HXB2, clade B (A); gp120_ZM109F.PB4 and gp41_ZA.1197MB, clade C (B); SOSIP, clade A (C); and DS-SOSIP, clade A (D). The two IgG3 MAbs shown in Fig. 2 were excluded from this analysis. The statistical significance of the differences in the results was determined using the two-way nonparametric Mann-Whitney test.

AUC analysis of ADCP against the SOSIP trimer showed comparable ADCP activities for all 4 classes of MAb (Fig. 4C). ADCP activity levels of the MAbs against DS-SOSIP were somewhat higher for the V2 and V3 MAbs (Fig. 4B). The V3 MAbs exhibited significantly elevated ADCP activity compared to the CD4bs Abs (P = 0.0012) (Fig. 4D) which was also comparable to that of the V2 and gp41 MAbs.

ADCP activity of IgG3 and IgG1 MAbs.

The Fc components of IgG subclasses use different Fcγ receptors and may mediate different activities. Indeed, two IgG3 MAbs tested here, the anti-V2 MAb (830A) and the anti-V3 MAb (447-52D), exhibited dominant ADCP activity in the panels of anti-V2 and anti-V3 MAbs shown (Fig. 2A, ,B,B, ,E,E, and andFF and and3A,3A, ,E,E, and andF).F). The reason for the poor ADCP activity of the 447-52D antibody against SOSIP-coated beads (Fig. 3B) is not known but presumably reflects poor binding to the clade A antigen.

In order to assess the ability of IgG3 MAbs to potently engage the THP-1 effector cells in the ADCP assay due to strong binding to Fcγ receptors and not to other factors, three anti-V2 MAbs were produced as recombinant Abs in the form of IgG1 and IgG3 variants and were compared in the ADCP assay. The 830A MAb was originally IgG3 and was switched to IgG1, while MAbs 1361 and 2158 were originally IgG1 and were switched to IgG3.

The three pairs of IgG1 and IgG3 recombinant variants were titrated and tested using beads coated with HIV gp120 proteins of BaL (clade B), ZM109F.PB4 (clade C), and CM244 (clade AE) (Fig. 5A to toC),C), as well as beads coated with clade A SOSIP and DS-SOSIP (Fig. 5D and andE).E). While limited ADCP activity was seen with the SOSIP-coated beads, in all other assays the V2 MAbs that were originally IgG1 but had been switched to IgG3 exhibited elevated ADCP activity in the IgG3 form (Fig. 5A to toCC and andE).E). In contrast, the 830A antibody, originally IgG3 but switched to IgG1, retained the same high ADCP activity level in its IgG1 form. AUC analysis revealed that the ADCP activities for the IgG3 variants tended to be higher than those for the corresponding IgG1 variants for all three gp120 molecules tested as well as for DS-SOSIP (Fig. 5F). The high activity of the 830A IgG1 MAb presumably prevented attainment of statistical significance. A larger panel of antibodies will be necessary to confirm the superior functioning of IgG3 over IgG1 recombinant Abs in mediating phagocytic activity in vitro.

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FIG 5

ADCP activity of three pairs of recombinant anti-V2 MAbs with Fc regions switched between IgG1 and IgG3. MAbs 2158 and 1361 were switched from IgG1 to IgG3, while MAb 830A was switched from IgG3 to IgG1; IgG1 is indicated as an open symbol (blue) and as IgG3 is a solid symbol (red). The MAbs were tested against beads coated with gp120_BaL (A), gp120_ZM109F.PB4 (B), gp120_CM244 (C), SOSIP (D), and DS-SOSIP (E). (F) The data were analyzed using area under the curve data.

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DISCUSSION

This study addressed the issue of whether anti-V2 MAbs can mediate ADCP activity, since V2 Abs had been previously shown to contribute to vaccine-induced protection in nonhuman primate experiments (29) and human vaccine trials (2, 3). If so, it would suggest a possible mechanism for the correlation of anti-V2 Abs with the reduced risk of HIV-1 infection observed in nonhuman primate vaccinees and in recipients of the RV144 vaccine. The results of the studies described here showed that anti-V2 MAbs mediate ADCP activity in a dose-dependent fashion. The anti-V2 MAbs exhibited ADCP activity similar to that of anti-V3 and CD4bs MAbs against clade B gp120 but displayed elevated activity against clade C gp120 compared to the other two MAb groups, suggesting broader recognition of exposed epitopes. Anti-gp41 MAbs were more effective overall in mediating ADCP against their appropriate gp41-coated beads than the other MAbs against their gp120 targets, but the results were comparable using gp140 trimers. The gp41 epitopes recognized are likely less exposed on SOSIP and DS-SOSIP trimers due to the presence of gp120, which can shield most of them. Anti-V2 MAbs also exhibited significant ADCP activity against the DS-SOSIP trimer (but not the SOSIP trimer), as did anti-V3 and anti-gp41 MAbs but not anti-CD4bs MAbs. Our results support the notion that the ADCP activity of anti-V2 Abs may have contributed to protection from HIV-1 infection in the RV144 trial.

The Ab isotype/subclass can modulate both epitope specificity and antiviral activity (51). Moreover, IgG3 has been shown to mediate better Fc effector function than other IgG subclasses (52). Therefore, in further evaluating the anti-V2 MAbs, we examined subclass differences. Results of experiments using three pairs of anti-V2 MAbs, with each pair including IgG1 and IgG3 variants (Fig. 5), were consistent with the possible contribution of ADCP to the protective efficacy against HIV-1 acquisition observed in the RV144 trial. Analysis of the RV144 data revealed that the presence of vaccine-elicited Abs of the IgG3 subclass specific for V1-V2 fusion proteins was correlated with reduced risk of HIV-1 infection (4). Given that the IgG3 anti-V2 MAb 830A does not display higher neutralizing activity than IgG1 anti-V2 MAbs (11), the ADCP activity of the IgG3 anti-V2 MAbs shown here, which tended to be higher than that of the corresponding IgG1 MAbs, suggests that this Fc-mediated activity may contribute more to protective efficacy than the weak neutralizing activity of anti-V2 MAbs against tier 1 viruses (11). Indeed, the protective role of anti-V2 Abs against HIV-1 infection may eventually prove to be polyfunctional. It may include other functions not explored here, such as additional Fc receptor-related activities and/or the inhibition of the gp120/α4β7 interaction which has been shown previously despite some remaining controversy (14, 17, 18).

Given that one of the main goals of the vaccine field is to elicit broadly neutralizing Abs (bnAbs), we sought to determine if an antigen designed to elicit such bnAbs would also be a target for a nonneutralizing activity, such as ADCP. As shown in Fig. 3 and and4,4, ADCP activity was poor against SOSIP targets whereas the conformationally constrained DS-SOSIP was a better target for this activity. It has previously been shown that nonneutralizing CD4bs MAbs poorly bound to SOSIP trimers (53), so it was not surprising that the CD4bs group of MAbs poorly mediated ADCP against these antigens. Interestingly, ADCP was better mediated by anti-V2, anti-V3, and anti-gp41 MAbs against DS-SOSIP targets, suggesting that the lack of conformational changes to the trimer resulted in better accessibility of the MAbs to the targeted epitopes. This may indicate that nonneutralizing Abs can act better against a consistent, relatively immobile antigen.

In order to control for day-to-day variability in ADCP assays, we normalized the phagocytic scores to background values (obtained with antigen-coated beads and THP-1 cells without serum) and then reported all results as the fold increase in the normalized value in comparison to that of ADCP against bovine serum albumin (BSA)-coated beads. Thus, the results show relatively modest values unlike the higher phagocytic scores reported without normalization (54). The modest values may also result from the small amount of gp120 and gp41 coated onto the beads. The assay was validated by the dependence of phagocytic activity on the dose of antigen coated on the beads as shown in Fig. 1. We chose a suboptimal dose of gp120 and a similar molar concentration of gp41 for coating the beads; this may mimic the low density of trimeric spikes on the virus envelope. Thus, despite the relatively low density of Env proteins on the beads, the observed ADCP activity mediated by the MAbs supports the concept that this function may also take place in vivo against HIV-1 infection.

ADCP can play an important role in antiviral immunity because it can engage several effector cells expressing three isoforms of FcγRs, including FcγIIa with medium affinity and also FcγIa and FcγIIIa (55). ADCP has been shown to have a protective function, clearing influenza and West Nile viruses (56, 57). In HIV infection, nonneutralizing Abs develop in less time and with less somatic hypermutation than neutralizing Abs (58, 59). Thus, they can respond significantly more quickly than bnAbs, making them very important in the initial immunological response to HIV infection. However, while ADCP develops relatively early in HIV infection (60), due to decreased Fc receptor expression on phagocytic cells with disease progression, its clearing efficiency can be affected (61). Therefore, ADCP mediated by vaccine-induced Abs against HIV-1 may be a particularly important protective correlate, as has been shown in vaccinated rhesus macaques (29).

In conclusion, we have demonstrated that human MAbs specific to several regions of the HIV-1 envelope, in spite of undetectable or weakly neutralizing activity, mediated ADCP activity in an in vitro assay. Depending on the HIV clade tested, the ADCP activity of anti-V2 MAbs displays some dominance over that of anti-V3 and anti-CD4bs MAbs. The activity of anti-gp41 MAbs is the most dominant using monomers but is comparable with that of other MAbs using gp140 trimers. The IgG3 MAbs are particularly effective in mediating ADCP compared to the corresponding IgG1 variants. Moving forward, elicitation of nonneutralizing Ab activities such as ADCP should be sought in conjunction with bnAbs in HIV vaccination strategies, as they appear to contribute to protection and are potentially more easily elicited than bnAbs.

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MATERIALS AND METHODS

Monoclonal antibodies.

Twenty-seven human MAbs specific for V2 (n = 6), V3 (n = 8), CD4bs (n = 6), and gp41 (n = 7) were tested in this study; the panel contained 25 IgG1 MAbs and 2 IgG3 MAbs (Table 1). All of these MAbs were derived from chronically HIV-1-infected individuals and were generated using the hybridoma technology based on fusion of Epstein-Barr virus (EBV)-transformed lymphocytes with the heteromyeloma cell line SHM-D33 (62). In light of the significant changes that differential glycosylation can impart to Fc effector function, especially ADCP (31), it was important that all Abs were produced in the same manner, with the exception of MAb 1361, which was used in recombinant form (r1361). Twenty-three MAbs were derived from individuals living primarily in the New York City area who were presumably infected with subtype B viruses, while four MAbs (2558, 3074, 3869, and 4682) originated from individuals infected with non-clade B viruses and living in Cameroon.

Recombinant proteins.

Three recombinant gp120s, BaL (clade B), ZM109F.PB4 (clade C), and CM244 (CRF01-AE), and two gp41 proteins, HXB2 (clade B) and ZA.1197MB (clade C), were purchased from Immune Technology Corp. Bovine serum albumin (BSA) was purchased from Sigma. gp140 trimers of two recombinant proteins, BG505 SOSIP.664 (clade A) and conformationally fixed BG505 DS-SOSIP.664 (201C 433C variant), were produced by transiently transfected 293F cells as previously described (63). Briefly, 293F cells were transfected using polyethylenimine HCl MAX (PEI-MAX) (1 mg/ml) in water mixed with expression plasmids for SOSIP.664 or DS-SOSIP.664, Furin (kindly provided by Peter Kwong and M. Gordon Joyce), and Opti-MEM. For one flask, 600 μg of Env plasmid, 150 μg of Furin plasmid, and 3 mg of PEI-MAX were added to 550 ml of growth media. Culture supernatants were harvested 72 h after transfection.

Binding assay (ELISA).

The MAbs were screened by standard ELISA against the recombinant gp120 and gp41 antigens used in the ADCP assay to confirm their binding activity. In short, proteins were coated directly onto Immulon 2HB plates at a concentration of 1 μg/ml. Following overnight incubation, the plates were washed with phosphate-buffered saline (PBS) containing 0.05% Tween 20 and blocked with assay diluent (PBS containing 2.5% bovine serum albumin and 7.5% fetal bovine serum). Monoclonal Abs (1 μg/ml) were added and incubated for 1.5 h at 37°C. Plates were washed again before bound Abs were detected by incubation with alkaline phosphatase-conjugated goat anti-human IgG (γ specific) (Southern Biotech), followed by washing and addition of substrate to develop color. The plates were read at 405 nm.

Ab-dependent cellular phagocytosis (ADCP) assay.

ADCP activity was measured as previously described (64) with minor modifications. In short, gp120 (BaL [clade B], ZM109 [clade C], and CM244 [clade AE]), gp41 (ZA.1197MB, clade C), gp41 (HXB2, clade B), BG505 SOSIP.664, BG505 DS-SOSIP.664, and BSA were biotinylated with a Biotin-XX microscale protein labeling kit (Life Technologies, Grand Island, NY) and incubated with a 100-fold dilution of 1 μg yellow-green streptavidin-fluorescent beads (Life Technologies) for 25 min at room temperature in the dark. The diameter of the beads is 1 μm, which is approximately 5 times the size of a virus. Dilutions of each MAb were added to 400,000 THP-1 cells (monocytic cell line) in a 96-well U-bottom plate. The bead-gp120 mixture was further diluted 5-fold in R10 media, and 50 μl was added to the cells and incubated for 3 h at 37°C. At the end of incubation, 70 μl 2% paraformaldehyde was added to fix the samples. Cells were then assayed for fluorescent bead uptake by flow cytometry using a BD Biosciences LSRII flow cytometer. The phagocytic score of each sample was calculated by multiplying the percentage of bead-positive cells (frequency) by the degree of phagocytosis measured as mean fluorescence intensity (MFI) and dividing by 10⁶. Values were normalized to background values (cells and beads without serum) by dividing the phagocytic score of the test sample by the phagocytic score of the background sample to ensure consistency in the values obtained on different assay days. As indicated in the figure legends, we report either this normalized phagocytosis score or, alternatively, the fold increase in phagocytosis against beads coated with test antigen compared to phagocytosis against BSA-coated beads.

Production of recombinant IgG1 and IgG3 anti-V2 MAbs.

Two anti-V2 IgG1 MAbs, 1361 and 2158, were switched to IgG3, and one anti-V2 IgG3 MAb, 830A, was changed to IgG1. The recombinant Abs were produced according to methods used in our laboratory (65, 66). In brief, total RNAs were isolated from hybridoma cell lines producing three MAbs, and cDNAs were synthesized by reverse transcription of mRNAs. The variable genes of heavy chains (VH) and light chains (VL) were amplified by PCR using specific primers for different immunoglobulin (Ig) genes of Abs and cDNAs synthesized from different Abs as the templates. The PCR products were then purified, sequenced, and digested with restriction enzymes which were introduced by the primers. VH genes were digested with restriction enzymes AgeI and SalI, while VL genes were digested with AgeI and BsiWI for kappa chain and XhoI for lambda chain. Finally, the digested VH and VL genes were ligated into IgG-AbVec expression vectors (kindly provided by Michel C. Nussenzweig, Rockefeller University, New York, NY). The VH genes were separately cloned into Igγ1 and Igγ3 expression vectors containing IgG1 or IgG3 Ig constant regions. The IgG3 expression vector was converted from an Igγ1 vector by replacing Cγ1 with a Cγ3 gene sequence. Both the VH and VL plasmids were used for cotransfection of 293T cells to produce recombinant IgG1 and IgG3 MAbs after 4 days of culture. The culture supernatants were harvested, and IgG Abs were purified by the use of protein G chromatography columns (GE Healthcare). The IgG concentrations were determined by quantitative ELISA (67).

Statistical analysis.

Area under the curve (AUC) analysis was used to determine the ADCP activity of MAbs, and the total areas were compared by the two-way nonparametric Mann Whitney test to determine statistical significance (GraphPad Prism).

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ACKNOWLEDGMENTS

We thank Peter Kwong and M. Gordon Joyce for the BG505 SOSIP.664 and BG505 DS-SOSIP.664 plasmids and for the VRC 3965 pCDNA3.1(-) Furin (human) plasmid for coexpression with SOSIP.664 and for helpful advice on the expression and purification of the trimers. We thank Yan Zheng and Xiaomei Liu for technical assistance and Irene Kalisz, Advanced BioScience Laboratories, Inc., for preparation of the SOSIP and DS-SOSIP trimers.

This study was supported by NIH grants AI112546 (M.K.G.) and P01 AI100151 (S.Z.-P.); by the Intramural Research Program of the National Institutes of Health, National Cancer Institute; and by the Department of Medicine at the Icahn School of Medicine at Mount Sinai.

We have no financial conflicts of interest to declare.

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REFERENCES

Rerks-Ngarm S, Pitisuttithum P, Nitayaphan S, Kaewkungwal J, Chiu J, Paris R, Premsri N, Namwat C, de Souza M, Adams E, Benenson M, Gurunathan S, Tartaglia J, McNeil JG, Francis DP, Stablein D, Birx DL, Chunsuttiwat S, Khamboonruang C, Thongcharoen P, Robb ML, Michael NL, Kunasol P, Kim JH. 2009. Vaccination with ALVAC and AIDSVAX to prevent HIV-1 infection in Thailand. N Engl J Med361:2209–2220. doi:10.1056/NEJMoa0908492. [PubMed] [CrossRef] [Google Scholar]
Haynes BF, Gilbert PB, McElrath MJ, Zolla-Pazner S, Tomaras GD, Alam SM, Evans DT, Montefiori DC, Karnasuta C, Sutthent R, Liao HX, DeVico AL, Lewis GK, Williams C, Pinter A, Fong Y, Janes H, DeCamp A, Huang Y, Rao M, Billings E, Karasavvas N, Robb ML, Ngauy V, de Souza MS, Paris R, Ferrari G, Bailer RT, Soderberg KA, Andrews C, Berman PW, Frahm N, De Rosa SC, Alpert MD, Yates NL, Shen X, Koup RA, Pitisuttithum P, Kaewkungwal J, Nitayaphan S, Rerks-Ngarm S, Michael NL, Kim JH. 2012. Immune-correlates analysis of an HIV-1 vaccine efficacy trial. N Engl J Med366:1275–1286. doi:10.1056/NEJMoa1113425. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Zolla-Pazner S, deCamp AC, Cardozo T, Karasavvas N, Gottardo R, Williams C, Morris DE, Tomaras G, Rao M, Billings E, Berman P, Shen X, Andrews C, O'Connell RJ, Ngauy V, Nitayaphan S, de Souza M, Korber B, Koup R, Bailer RT, Mascola JR, Pinter A, Montefiori D, Haynes BF, Robb ML, Rerks-Ngarm S, Michael NL, Gilbert PB, Kim JH. 2013. Analysis of V2 antibody responses induced in vaccinees in the ALVAC/AIDSVAX HIV-1 vaccine efficacy trial. PLoS One8:e53629. doi:10.1371/journal.pone.0053629. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Yates NL, Liao HX, Fong Y, Decamp A, Vandergrift NA, Williams WT, Alam SM, Ferrari G, Yang ZY, Seaton KE, Berman PW, Alpert MD, Evans DT, O'Connell RJ, Francis D, Sinangil F, Lee C, Nitayaphan S, Rerks-Ngarm S, Kaewkungwal J, Pitisuttithum P, Tartaglia J, Pinter A, Zolla-Pazner S, Gilbert PB, Nabel GJ, Michael NL, Kim JH, Montefiori DC, Haynes BF, Tomaras GD. 2014. Vaccine-induced Env V1-V2 IgG3 correlates with lower HIV-1 infection risk and declines soon after vaccination. Sci Transl Med6:228ra39. doi:10.1126/scitranslmed.3007730. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Rolland M, Edlefsen PT, Larsen BB, Tovanabutra S, Sanders-Buell E, Hertz T, deCamp AC, Carrico C, Menis S, Magaret CA, Ahmed H, Juraska M, Chen L, Konopa P, Nariya S, Stoddard JN, Wong K, Zhao H, Deng W, Maust BS, Bose M, Howell S, Bates A, Lazzaro M, O'Sullivan A, Lei E, Bradfield A, Ibitamuno G, Assawadarachai V, O'Connell RJ, deSouza MS, Nitayaphan S, Rerks-Ngarm S, Robb ML, McLellan JS, Georgiev I, Kwong PD, Carlson JM, Michael NL, Schief WR, Gilbert PB, Mullins JI, Kim JH. 2012. Increased HIV-1 vaccine efficacy against viruses with genetic signatures in Env V2. Nature490:417–420. doi:10.1038/nature11519. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Roederer M, Keele BF, Schmidt SD, Mason RD, Welles HC, Fischer W, Labranche C, Foulds KE, Louder MK, Yang ZY, Todd JP, Buzby AP, Mach LV, Shen L, Seaton KE, Ward BM, Bailer RT, Gottardo R, Gu W, Ferrari G, Alam SM, Denny TN, Montefiori DC, Tomaras GD, Korber BT, Nason MC, Seder RA, Koup RA, Letvin NL, Rao SS, Nabel GJ, Mascola JR. 2014. Immunological and virological mechanisms of vaccine-mediated protection against SIV and HIV. Nature505:502–508. [PMC free article] [PubMed] [Google Scholar]
Gordon SN, Doster MN, Kines RC, Keele BF, Brocca-Cofano E, Guan Y, Pegu P, Liyanage NP, Vaccari M, Cuburu N, Buck CB, Ferrari G, Montefiori D, Piatak M Jr, Lifson JD, Xenophontos AM, Venzon D, Robert-Guroff M, Graham BS, Lowy DR, Schiller JT, Franchini G. 2014. Antibody to the gp120 V1/V2 loops and CD4+ and CD8+ T cell responses in protection from SIVmac251 vaginal acquisition and persistent viremia. J Immunol193:6172–6183. doi:10.4049/jimmunol.1401504. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Barouch DH, Liu J, Li H, Maxfield LF, Abbink P, Lynch DM, Iampietro MJ, SanMiguel A, Seaman MS, Ferrari G, Forthal DN, Ourmanov I, Hirsch VM, Carville A, Mansfield KG, Stablein D, Pau MG, Schuitemaker H, Sadoff JC, Billings EA, Rao M, Robb ML, Kim JH, Marovich MA, Goudsmit J, Michael NL. 2012. Vaccine protection against acquisition of neutralization-resistant SIV challenges in rhesus monkeys. Nature482:89–93. doi:10.1038/nature10766. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Vaccari M, Gordon SN, Fourati S, Schifanella L, Liyanage NP, Cameron M, Keele BF, Shen X, Tomaras GD, Billings E, Rao M, Chung AW, Dowell KG, Bailey-Kellogg C, Brown EP, Ackerman ME, Vargas-Inchaustegui DA, Whitney S, Doster MN, Binello N, Pegu P, Montefiori DC, Foulds K, Quinn DS, Donaldson M, Liang F, Lore K, Roederer M, Koup RA, McDermott A, Ma ZM, Miller CJ, Phan TB, Forthal DN, Blackburn M, Caccuri F, Bissa M, Ferrari G, Kalyanaraman V, Ferrari MG, Thompson D, Robert-Guroff M, Ratto-Kim S, Kim JH, Michael NL, Phogat S, Barnett SW, Tartaglia J, Venzon D, Stablein DM, et al. 2016. Adjuvant-dependent innate and adaptive immune signatures of risk of SIVmac251 acquisition. Nature Med22:762–770. doi:10.1038/nm.4105. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Montefiori DC, Karnasuta C, Huang Y, Ahmed H, Gilbert P, de Souza MS, McLinden R, Tovanabutra S, Laurence-Chenine A, Sanders-Buell E, Moody MA, Bonsignori M, Ochsenbauer C, Kappes J, Tang H, Greene K, Gao H, LaBranche CC, Andrews C, Polonis VR, Rerks-Ngarm S, Pitisuttithum P, Nitayaphan S, Kaewkungwal J, Self SG, Berman PW, Francis D, Sinangil F, Lee C, Tartaglia J, Robb ML, Haynes BF, Michael NL, Kim JH. 2012. Magnitude and breadth of the neutralizing antibody response in the RV144 and Vax003 HIV-1 vaccine efficacy trials. J Infect Dis206:431–441. doi:10.1093/infdis/jis367. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, Pan R, Williams C, Wang XH, Volsky B, O'Neal T, Spurrier B, Sampson JM, Li L, Seaman MS, Kong XP, Zolla-Pazner S. 2012. Functional and immunochemical cross-reactivity of V2-specific monoclonal antibodies from human immunodeficiency virus type 1-infected individuals. Virology427:198–207. doi:10.1016/j.virol.2012.02.003. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, Moore JP, Conley AJ, Karwowska S, Sodroski J, Williams C, Burda S, Boots LJ, Zolla-Pazner S. 1994. Human anti-V2 monoclonal antibody that neutralizes primary but not laboratory isolates of HIV-1. J Virol68:8312–8320. [PMC free article] [PubMed] [Google Scholar]
Corti D, Langedijk JP, Hinz A, Seaman MS, Vanzetta F, Fernandez-Rodriguez BM, Silacci C, Pinna D, Jarrossay D, Balla-Jhagjhoorsingh S, Willems B, Zekveld MJ, Dreja H, O'Sullivan E, Pade C, Orkin C, Jeffs SA, Montefiori DC, Davis D, Weissenhorn W, McKnight A, Heeney JL, Sallusto F, Sattentau QJ, Weiss RA, Lanzavecchia A. 2010. Analysis of memory B cell responses and isolation of novel monoclonal antibodies with neutralizing breadth from HIV-1-infected individuals. PLoS One5:e8805. doi:10.1371/journal.pone.0008805. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Nakamura GR, Fonseca DP, O'Rourke SM, Vollrath AL, Berman PW. 2012. Monoclonal antibodies to the V2 domain of MN-rgp120: fine mapping of epitopes and inhibition of alpha4beta7 binding. PLoS One7:e39045. doi:10.1371/journal.pone.0039045. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Liao HX, Bonsignori M, Alam SM, McLellan JS, Tomaras GD, Moody MA, Kozink DM, Hwang KK, Chen X, Tsao CY, Liu P, Lu X, Parks RJ, Montefiori DC, Ferrari G, Pollara J, Rao M, Peachman KK, Santra S, Letvin NL, Karasavvas N, Yang ZY, Dai K, Pancera M, Gorman J, Wiehe K, Nicely NI, Rerks-Ngarm S, Nitayaphan S, Kaewkungwal J, Pitisuttithum P, Tartaglia J, Sinangil F, Kim JH, Michael NL, Kepler TB, Kwong PD, Mascola JR, Nabel GJ, Pinter A, Zolla-Pazner S, Haynes BF. 2013. Vaccine induction of antibodies against a structurally heterogeneous site of immune pressure within HIV-1 envelope protein variable regions 1 and 2. Immunity38:176–186. doi:10.1016/j.immuni.2012.11.011. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Mayr LM, Cohen S, Spurrier B, Kong XP, Zolla-Pazner S. 2013. Epitope mapping of conformational V2-specific anti-HIV human monoclonal antibodies reveals an immunodominant site in V2. PLoS One8:e70859. doi:10.1371/journal.pone.0070859. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Tassaneetrithep B, Tivon D, Swetnam J, Karasavvas N, Michael NL, Kim JH, Marovich M, Cardozo T. 2014. Cryptic determinant of alpha4beta7 binding in the V2 loop of HIV-1 gp120. PLoS One9:e108446. doi:10.1371/journal.pone.0108446. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Perez LG, Chen H, Liao HX, Montefiori DC. 2014. Envelope glycoprotein binding to the integrin alpha4beta7 is not a general property of most HIV-1 strains. J Virol88:10767–10777. doi:10.1128/JVI.03296-13. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Nawaz F, Cicala C, Van Ryk D, Block KE, Jelicic K, McNally JP, Ogundare O, Pascuccio M, Patel N, Wei D, Fauci AS, Arthos J. 2011. The genotype of early-transmitting HIV gp120s promotes alphabeta-reactivity, revealing alphabetaCD4+ T cells as key targets in mucosal transmission. PLoS Pathog7:e1001301. doi:10.1371/journal.ppat.1001301. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Forthal DN, Landucci G, Cole KS, Marthas M, Becerra JC, Van Rompay K. 2006. Rhesus macaque polyclonal and monoclonal antibodies inhibit simian immunodeficiency virus in the presence of human or autologous rhesus effector cells. J Virol80:9217–9225. doi:10.1128/JVI.02746-05. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Forthal DN, Gilbert PB, Landucci G, Phan T. 2007. Recombinant gp120 vaccine-induced antibodies inhibit clinical strains of HIV-1 in the presence of fc receptor-bearing effector cells and correlate inversely with HIV infection rate. J Immunol178:6596–6603. doi:10.4049/jimmunol.178.10.6596. [PubMed] [CrossRef] [Google Scholar]
Hidajat R, Xiao P, Zhou Q, Venzon D, Summers LE, Kalyanaraman VS, Montefiori DC, Robert-Guroff M. 2009. Correlation of vaccine-elicited systemic and mucosal nonneutralizing antibody activities with reduced acute viremia following intrarectal simian immunodeficiency virus SIVmac251 challenge of rhesus macaques. J Virol83:791–801. doi:10.1128/JVI.01672-08. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Florese RH, Demberg T, Xiao P, Kuller L, Larsen K, Summers LE, Venzon D, Cafaro A, Ensoli B, Robert-Guroff M. 2009. Contribution of nonneutralizing vaccine-elicited antibody activities to improved protective efficacy in rhesus macaques immunized with Tat/Env compared with multigenic vaccines. J Immunol182:3718–3727. doi:10.4049/jimmunol.0803115. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Xiao P, Zhao J, Patterson LJ, Brocca-Cofano E, Venzon D, Kozlowski PA, Hidajat R, Demberg T, Robert-Guroff M. 2010. Multiple vaccine-elicited nonneutralizing antienvelope antibody activities contribute to protective efficacy by reducing both acute and chronic viremia following simian/human immunodeficiency virus SHIV89.6P challenge in rhesus macaques. J Virol84:7161–7173. doi:10.1128/JVI.00410-10. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gómez-Román VR, Patterson LJ, Venzon D, Liewehr D, Aldrich K, Florese R, Robert-Guroff M. 2005. Vaccine-elicited antibodies mediate antibody-dependent cellular cytotoxicity correlated with significantly reduced acute viremia in rhesus macaques challenged with SIVmac251. J Immunol174:2185–2189. doi:10.4049/jimmunol.174.4.2185. [PubMed] [CrossRef] [Google Scholar]
Alpert MD, Harvey JD, Lauer WA, Reeves RK, Piatak M Jr, Carville A, Mansfield KG, Lifson JD, Li W, Desrosiers RC, Johnson RP, 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 Pathog8:e1002890. doi:10.1371/journal.ppat.1002890. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Xiao P, Patterson LJ, Kuate S, Brocca-Cofano E, Thomas MA, Venzon D, Zhao J, DiPasquale J, Fenizia C, Lee EM, Kalisz I, Kalyanaraman VS, Pal R, Montefiori D, Keele BF, Robert-Guroff M. 2012. Replicating adenovirus-simian immunodeficiency virus (SIV) recombinant priming and envelope protein boosting elicits localized, mucosal IgA immunity in rhesus macaques correlated with delayed acquisition following a repeated low-dose rectal SIV(mac251) challenge. J Virol86:4644–4657. doi:10.1128/JVI.06812-11. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Wren LH, Chung AW, Isitman G, Kelleher AD, Parsons MS, Amin J, Cooper DA; ADCC study collaboration investigators, Stratov I, Navis M, Kent SJ. 2013. Specific antibody-dependent cellular cytotoxicity responses associated with slow progression of HIV infection. Immunology138:116–123. doi:10.1111/imm.12016. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Barouch DH, Stephenson KE, Borducchi EN, Smith K, Stanley K, McNally AG, Liu J, Abbink P, Maxfield LF, Seaman MS, Dugast AS, Alter G, Ferguson M, Li W, Earl PL, Moss B, Giorgi EE, Szinger JJ, Eller LA, Billings EA, Rao M, Tovanabutra S, Sanders-Buell E, Weijtens M, Pau MG, Schuitemaker H, Robb ML, Kim JH, Korber BT, Michael NL. 2013. Protective efficacy of a global HIV-1 mosaic vaccine against heterologous SHIV challenges in rhesus monkeys. Cell155:531–539. doi:10.1016/j.cell.2013.09.061. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Ackerman ME, Mikhailova A, Brown EP, Dowell KG, Walker BD, Bailey-Kellogg C, Suscovich TJ, Alter G. 2016. Polyfunctional HIV-specific antibody responses are associated with spontaneous HIV control. PLoS Pathog12:e1005315. doi:10.1371/journal.ppat.1005315. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Chung AW, Crispin M, Pritchard L, Robinson H, Gorny MK, Yu X, Bailey-Kellogg C, Ackerman ME, Scanlan C, Zolla-Pazner S, Alter G. 2014. Identification of antibody glycosylation structures that predict monoclonal antibody Fc-effector function. AIDS28:2523–2530. doi:10.1097/QAD.0000000000000444. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Veillette M, Coutu M, Richard J, Batraville LA, Dagher O, Bernard N, Tremblay C, Kaufmann DE, Roger M, 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 Virol89:545–551. doi:10.1128/JVI.02868-14. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Sips M, Krykbaeva M, Diefenbach TJ, Ghebremichael M, Bowman BA, Dugast AS, Boesch AW, Streeck H, Kwon DS, Ackerman ME, Suscovich TJ, Brouckaert P, Schacker TW, Alter G. 2016. Fc receptor-mediated phagocytosis in tissues as a potent mechanism for preventive and therapeutic HIV vaccine strategies. Mucosal Immunol9:1584–1595. doi:10.1038/mi.2016.12. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Chung AW, Ghebremichael M, Robinson H, Brown E, Choi I, Lane S, Dugast AS, Schoen MK, Rolland M, Suscovich TJ, Mahan AE, Liao L, Streeck H, Andrews C, Rerks-Ngarm S, Nitayaphan S, de Souza MS, Kaewkungwal J, Pitisuttithum P, Francis D, Michael NL, Kim JH, Bailey-Kellogg C, Ackerman ME, Alter G. 2014. Polyfunctional Fc-effector profiles mediated by IgG subclass selection distinguish RV144 and VAX003 vaccines. Sci Transl Med6:228ra38. doi:10.1126/scitranslmed.3007736. [PubMed] [CrossRef] [Google Scholar]
Nyambi PN, Mbah HA, Burda S, Williams C, Gorny MK, Nadas A, Zolla-Pazner S. 2000. Conserved and exposed epitopes on intact, native, primary human immunodeficiency virus type 1 virions of group M. J Virol74:7096–7107. doi:10.1128/JVI.74.15.7096-7107.2000. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, VanCott TC, Williams C, Revesz K, Zolla-Pazner S. 2000. Effects of oligomerization on the epitopes of the human immunodeficiency virus type 1 envelope glycoproteins. Virology267:220–228. doi:10.1006/viro.1999.0095. [PubMed] [CrossRef] [Google Scholar]
Pinter A, Honnen WJ, He Y, Gorny MK, Zolla-Pazner S, Kayman SC. 2004. The V1/V2 domain of gp120 is a global regulator of sensitivity of primary human immunodeficiency virus type 1 isolates to neutralization by antibodies commonly induced upon infection. J Virol78:5205–5215. doi:10.1128/JVI.78.10.5205-5215.2004. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, Xu J-Y, Karwowska S, Buchbinder A, Zolla-Pazner S. 1993. Repertoire of neutralizing human monoclonal antibodies specific for the V3 domain of HIV-1 gp120. J Immunol150:635–643. [PubMed] [Google Scholar]
Gorny MK, Williams C, Volsky B, Revesz K, Cohen S, Polonis VR, Honnen WJ, Kayman SC, Krachmarov CP, Pinter A, Zolla-Pazner S. 2002. Human monoclonal antibodies specific for conformation-sensitive epitopes of V3 neutralize HIV-1 primary isolates from various clades. J Virol76:9035–9045. doi:10.1128/JVI.76.18.9035-9045.2002. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, Revesz K, Williams C, Volsky B, Louder MK, Anyangwe CA, Krachmarov CP, Kayman SC, Pinter A, Nadas A, Nyambi PN, Mascola JR, Zolla-Pazner S. 2004. The V3 loop is accessible on the surface of most human immunodeficiency virus type 1 primary isolates and serves as a neutralization epitope. J Virol78:2394–2404. doi:10.1128/JVI.78.5.2394-2404.2004. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, Williams C, Volsky B, Revesz K, Wang XH, Burda S, Kimura T, Koning FA, Nadas A, Anyangwe C, Nyambi P, Krachmarov C, Pinter A, Zolla-Pazner S. 2006. Cross-clade neutralizing activity of human anti-V3 monoclonal antibodies derived from the cells of individuals infected with non-B clades of HIV-1. J Virol80:6865–6872. doi:10.1128/JVI.02202-05. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, Wang XH, Williams C, Volsky B, Revesz K, Witover B, Burda S, Urbanski M, Nyambi P, Krachmarov C, Pinter A, Zolla-Pazner S, Nadas A. 2009. Preferential use of the VH5-51 gene segment by the human immune response to code for antibodies against the V3 domain of HIV-1. Mol Immunol46:917–926. doi:10.1016/j.molimm.2008.09.005. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Li L, Wang XH, Williams C, Volsky B, Steczko O, Seaman MS, Luthra K, Nyambi P, Nadas A, Giudicelli V, Lefranc MP, Zolla-Pazner S, Gorny MK. 2015. A broad range of mutations in HIV-1 neutralizing human monoclonal antibodies specific for V2, V3, and the CD4 binding site. Mol Immunol66:364–374. doi:10.1016/j.molimm.2015.04.011. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Karwowska S, Gorny MK, Buchbinder A, Gianakakos V, Williams C, Fuerst T, Zolla-Pazner S. 1992. Production of human monoclonal antibodies specific for conformational and linear non-V3 epitopes of gp120. AIDS Res Hum Retroviruses8:1099–1106. doi:10.1089/aid.1992.8.1099. [PubMed] [CrossRef] [Google Scholar]
Nyambi PN, Gorny MK, Bastiani L, van der Groen G, Williams C, Zolla-Pazner S. 1998. Mapping of epitopes exposed on intact human immunodeficiency virus type 1 (HIV-1) virions: a new strategy for studying the immunologic relatedness of HIV-1. J Virol72:9384–9391. [PMC free article] [PubMed] [Google Scholar]
Zolla-Pazner S, O'Leary J, Burda S, Gorny MK, Kim M, Mascola J, McCutchan FE. 1995. Serotyping of primary human immunodeficiency virus type 1 isolates from diverse geographic locations by flow cytometry. J Virol69:3807–3815. [PMC free article] [PubMed] [Google Scholar]
Jeffs SA, Gorny MK, Williams C, Revesz K, Volsky B, Burda S, Wang XH, Bandres J, Zolla-Pazner S, Holmes H. 2001. Characterization of human monoclonal antibodies selected with a hypervariable loop-deleted recombinant HIV-1(IIIB) gp120. Immunol Lett79:209–213. doi:10.1016/S0165-2478(01)00289-9. [PubMed] [CrossRef] [Google Scholar]
Xu J-Y, Gorny MK, Palker T, Karwowska S, Zolla-Pazner S. 1991. Epitope mapping of two immunodominant domains of gp41, the transmembrane protein of human immunodeficiency virus type 1, using ten human monoclonal antibodies. J Virol65:4832–4838. [PMC free article] [PubMed] [Google Scholar]
Gigler A, Dorsch S, Hemauer A, Williams C, Kim S, Young NS, Zolla-Pazner S, Wolf H, Gorny MK, Modrow S. 1999. Generation of neutralizing human monoclonal antibodies against parvovirus B19 proteins. J Virol73:1974–1979. [PMC free article] [PubMed] [Google Scholar]
Kwon YD, Pancera M, Acharya P, Georgiev IS, Crooks ET, Gorman J, Joyce MG, Guttman M, Ma X, Narpala S, Soto C, Terry DS, Yang Y, Zhou T, Ahlsen G, Bailer RT, Chambers M, Chuang GY, Doria-Rose NA, Druz A, Hallen MA, Harned A, Kirys T, Louder MK, O'Dell S, Ofek G, Osawa K, Prabhakaran M, Sastry M, Stewart-Jones GB, Stuckey J, Thomas PV, Tittley T, Williams C, Zhang B, Zhao H, Zhou Z, Donald BR, Lee LK, Zolla-Pazner S, Baxa U, Schon A, Freire E, Shapiro L, Lee KK, Arthos J, Munro JB, Blanchard SC, Mothes W, Binley JM, et al. 2015. Crystal structure, conformational fixation and entry-related interactions of mature ligand-free HIV-1 Env. Nat Struct Mol Biol22:522–531. doi:10.1038/nsmb.3051. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Tudor D, Yu H, Maupetit J, Drillet AS, Bouceba T, Schwartz-Cornil I, Lopalco L, Tuffery P, Bomsel M. 2012. Isotype modulates epitope specificity, affinity, and antiviral activities of anti-HIV-1 human broadly neutralizing 2F5 antibody. Proc Natl Acad Sci U S A109:12680–12685. doi:10.1073/pnas.1200024109. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Vidarsson G, Dekkers G, Rispens T. 2014. IgG subclasses and allotypes: from structure to effector functions. Front Immunol5:520. doi:10.3389/fimmu.2014.00520. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Yasmeen A, Ringe R, Derking R, Cupo A, Julien JP, Burton DR, Ward AB, Wilson IA, Sanders RW, Moore JP, Klasse PJ. 2014. Differential binding of neutralizing and non-neutralizing antibodies to native-like soluble HIV-1 Env trimers, uncleaved Env proteins, and monomeric subunits. Retrovirology11:41. doi:10.1186/1742-4690-11-41. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Dugast AS, Chan Y, Hoffner M, Licht A, Nkolola J, Li H, Streeck H, Suscovich TJ, Ghebremichael M, Ackerman ME, Barouch DH, Alter G. 2014. Lack of protection following passive transfer of polyclonal highly functional low-dose non-neutralizing antibodies. PLoS One9:e97229. doi:10.1371/journal.pone.0097229. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Bruhns P. 2012. Properties of mouse and human IgG receptors and their contribution to disease models. Blood119:5640–5649. doi:10.1182/blood-2012-01-380121. [PubMed] [CrossRef] [Google Scholar]
Huber VC, Lynch JM, Bucher DJ, Le J, Metzger DW. 2001. Fc receptor-mediated phagocytosis makes a significant contribution to clearance of influenza virus infections. J Immunol166:7381–7388. doi:10.4049/jimmunol.166.12.7381. [PubMed] [CrossRef] [Google Scholar]
Chung KM, Thompson BS, Fremont DH, Diamond MS. 2007. Antibody recognition of cell surface-associated NS1 triggers Fc-gamma receptor-mediated phagocytosis and clearance of West Nile Virus-infected cells. J Virol81:9551–9555. doi:10.1128/JVI.00879-07. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Huber M, Trkola A. 2007. Humoral immunity to HIV-1: neutralization and beyond. J Intern Med262:5–25. doi:10.1111/j.1365-2796.2007.01819.x. [PubMed] [CrossRef] [Google Scholar]
Bonsignori M, Pollara J, Moody MA, Alpert MD, Chen X, Hwang KK, Gilbert PB, Huang Y, Gurley TC, Kozink DM, Marshall DJ, Whitesides JF, Tsao CY, Kaewkungwal J, Nitayaphan S, Pitisuttithum P, Rerks-Ngarm S, Kim JH, Michael NL, Tomaras GD, Montefiori DC, Lewis GK, DeVico A, Evans DT, Ferrari G, Liao HX, 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 Virol86:11521–11532. doi:10.1128/JVI.01023-12. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Ana-Sosa-Batiz F, Johnston AP, Liu H, Center RJ, Rerks-Ngarm S, Pitisuttithum P, Nitayaphan S, Kaewkungwal J, Kim JH, Michael NL, Kelleher AD, Stratov I, Kent SJ, Kramski M. 2014. HIV-specific antibody-dependent phagocytosis matures during HIV infection. Immunol Cell Biol92:679–687. doi:10.1038/icb.2014.42. [PubMed] [CrossRef] [Google Scholar]
Dugast AS, Tonelli A, Berger CT, Ackerman ME, Sciaranghella G, Liu Q, Sips M, Toth I, Piechocka-Trocha A, Ghebremichael M, Alter G. 2011. Decreased Fc receptor expression on innate immune cells is associated with impaired antibody-mediated cellular phagocytic activity in chronically HIV-1 infected individuals. Virology415:160–167. doi:10.1016/j.virol.2011.03.012. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, Xu J-Y, Gianakakos V, Karwowska S, Williams C, Sheppard HW, Hanson CV, Zolla-Pazner S. 1991. Production of site-selected neutralizing human monoclonal antibodies against the third variable domain of the HIV-1 envelope glycoprotein. Proc Natl Acad Sci U S A88:3238–3242. doi:10.1073/pnas.88.8.3238. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Sanders RW, Derking R, Cupo A, Julien JP, Yasmeen A, de Val N, Kim HJ, Blattner C, de la Pena AT, Korzun J, Golabek M, de Los Reyes K, Ketas TJ, van Gils MJ, King CR, Wilson IA, Ward AB, Klasse PJ, Moore JP. 2013. A next-generation cleaved, soluble HIV-1 Env trimer, BG505 SOSIP.664 gp140, expresses multiple epitopes for broadly neutralizing but not non-neutralizing antibodies. PLoS Pathog9:e1003618. doi:10.1371/journal.ppat.1003618. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Ackerman ME, Moldt B, Wyatt RT, Dugast AS, McAndrew E, Tsoukas S, Jost S, Berger CT, Sciaranghella G, Liu Q, Irvine DJ, Burton DR, Alter G. 2011. A robust, high-throughput assay to determine the phagocytic activity of clinical antibody samples. J Immunol Methods366:8–19. doi:10.1016/j.jim.2010.12.016. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Li L, Wang X-H, Banerjee S, Volsky B, Williams C, Virland D, Nadas A, Seaman MS, Chen X, Spearman P, Zolla-Pazner S, Gorny MK. 2012. Different pattern of immunoglobulin gene usage by HIV-1 compared to non-HIV-1 antibodies derived from the same infected subject. PLoS One7:e39534. doi:10.1371/journal.pone.0039534. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Li L, Wang XH, Banerjee S, Volsky B, Williams C, Moody MA, Zolla-Pazner S, Gorny MK. 2012. Clonal analysis of human anti-V3 monoclonal antibodies selected by a V3 tetramer. Hum Antibodies21:65–73. doi:10.3233/HAB-130264. [PMC free article] [PubMed] [CrossRef] [Google Scholar]
Gorny MK, Gianakakos V, Sharpe S, Zolla-Pazner S. 1989. Generation of human monoclonal antibodies to HIV. Proc Natl Acad Sci U S A86:1624–1628. doi:10.1073/pnas.86.5.1624. [PMC free article] [PubMed] [CrossRef] [Google Scholar]

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Monoclonal Antibodies Specific for the V2, V3, CD4-Binding Site, and gp41 of HIV-1 Mediate Phagocytosis in a Dose-Dependent Manner.

Products Related to gp41, gp120, gp160

Product# 1091 Recombinant HIV-1 MN gp41 Envelope Protein (E.coli)

Product#1001 Recombinant HIV-1 IIIB Envelope Glycoprotein gp120 (Baculovirus)

Product#1024 Recombinant HIV-1 MN Envelope Glycoprotein gp160 (Baculovirus)

INTRODUCTION

RESULTS