Rogier W Sanders

Weill Cornell Medical College, New York, New York, United States

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Publications (117)800.9 Total impact

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    ABSTRACT: When HIV-1 vaccine candidates that include soluble envelope glycoproteins (Env) are tested in humans and other species, the resulting antibody responses to Env are sifted for correlates of protection or risk. One frequently used assay measures the reduction in antibody binding to Env antigens by an added chaotrope (such as thiocyanate). Based on that assay, an avidity index was devised for assessing the affinity maturation of antibodies of unknown concentration in polyclonal sera. Since a high avidity index was linked to protection in animal models of HIV-1 infection, it has become a criterion for evaluating antibody responses to vaccine candidates. But what does the assay measure and what does an avidity index mean? Here, we have used a panel of monoclonal antibodies to well-defined epitopes on Env (gp120, gp41, and SOSIP.664 trimers) to explore how the chaotrope acts. We conclude that the chaotrope sensitivity of antibody binding to Env depends on several properties of the epitopes (continuity vs. tertiary- and quaternary-structural dependence), and that the avidity index has no simple relationship to antibody affinity for functional Env spikes on virions. We show that the binding of broadly neutralizing antibodies against quaternary-structural epitopes is particularly sensitive to chaotrope treatment, whereas antibody binding to epitopes in variable loops and to non-neutralization epitopes in gp41 is generally resistant. As a result of such biases, the avidity index may at best be a mere surrogate for undefined antibody or other immune responses that correlate weakly with protection. An effective HIV-1 vaccine is an important goal. Such a vaccine will probably need to induce antibodies that neutralize typically transmitted variants of HIV-1, i.e., preventing them from infecting target cells. Vaccine candidates have so far failed to induce such antibody responses, although some do protect weakly against infection in animals and possibly humans. In the search for responses associated with protection, an avidity assay based on chemical disruption is often used to measure the strength of antibody binding. We have analyzed this assay mechanistically and found that the epitope specificity of an antibody has a greater influence on the outcome than does its affinity. As a result, the avidity assay is biased towards the detection of some antibody specificities while disfavoring others. We conclude that the assay may yield merely indirect correlations with weak protection, specifically when Env vaccination has failed to induce broad neutralizing responses. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Journal of Virology 03/2015; DOI:10.1128/JVI.00320-15 · 4.65 Impact Factor
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    ABSTRACT: Antibodies with the ability to block the interaction of HIV-1 envelope glycoprotein (Env) gp120 with CD4, including those overlapping the CD4 binding site (CD4bs antibodies), can protect from infection by HIV-1, and their elicitation may be an interesting goal for any vaccination strategy. To identify gp120/CD4 blocking antibodies in plasma samples from HIV-1 infected individuals we have developed a competitive flow cytometry-based functional assay. In a cohort of treatment-naïve chronically infected patients, we showed that gp120/ CD4 blocking antibodies were frequently elicited (detected in 97% plasma samples) and correlated with binding to trimeric HIV-1 envelope glycoproteins. However, no correlation was observed between functional CD4 binding blockade data and titer of CD4bs antibodies determined by ELISA using resurfaced gp120 proteins. Consistently, plasma samples lacking CD4bs antibodies were able to block the interaction between gp120 and its receptor, indicating that antibodies recognizing other epitopes, such as PGT126 and PG16, can also play the same role. Antibodies blocking CD4 binding increased over time and correlated positively with the capacity of plasma samples to neutralize the laboratory-adapted NL4.3 and BaL virus isolates, suggesting their potential contribution to the neutralizing workforce of plasma in vivo. Determining whether this response can be boosted to achieve broadly neutralizing antibodies may provide valuable information for the design of new strategies aimed to improve the anti-HIV-1 humoral response and to develop a successful HIV- 1 vaccine.
    PLoS ONE 03/2015; 10(3):e0120648. DOI:10.1371/journal.pone.0120648 · 3.53 Impact Factor
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    ABSTRACT: Latent HIV type I (HIV-1) infections can frequently occur in short-lived proliferating effector T-lymphocytes. These latently infected cells could revert into resting T-lymphocytes and thereby contribute to the establishment of the long-lived viral reservoir. Monocyte-derived dendritic cells can revert latency in effector T-cells in vitro. Here we investigated the latency activation properties of tissue-specific immune cells, including a large panel of dendritic cell subsets, to explore in which body compartments effector T-cells are most likely to maintain latent HIV-1 provirus and thus potentially contribute to the long-lived reservoir. Our results demonstrate that blood or genital tract dendritic cells do not activate latent provirus in effector T-cells, whereas gut or lymphoid dendritic cells induce virus production from latently infected effector T-cells in our in-vitro model for latency. Toll-like receptor 3-induced interferon production by myeloid dendritic cells abolished the dendritic cells' ability to induce viral gene expression. In this study, we show that HIV-1 provirus residing in the effector T-cells is activated from latency by tissue-specific dendritic cell subsets and other immune cells with remarkably different efficiencies.Our new assay system points to an important, neglected aspect of HIV-1 research: the ability of other immune cells, especially dendritic cells, to differentially affect latency establishment as well as virus reactivation.
    AIDS (London, England) 03/2015; DOI:10.1097/QAD.0000000000000637 · 6.56 Impact Factor
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    ABSTRACT: The trimeric envelope (Env) spike is the focus of vaccine design efforts aimed at generating broadly neutralizing antibodies (bNAbs) to protect against HIV-1 infection. Three recent developments have facilitated a thorough investigation of the antigenic structure of the Env trimer: 1) the isolation of many bNAbs against multiple different epitopes; 2) the generation of a soluble trimer mimic, BG505 SOSIP.664 gp140, that expresses most bNAb epitopes; 3) facile binding assays involving the oriented immobilization of tagged trimers. Using these tools, we generated an antigenic map of the trimer by antibody cross-competition. Our analysis delineates three well-defined epitope clusters (CD4 binding site, quaternary V1V2 and Asn332-centered oligomannose patch) and new epitopes at the gp120-gp41 interface. It also identifies the relationships among these clusters. In addition to epitope overlap, we defined three more ways in which antibodies can cross-compete: steric competition from binding to proximal but non-overlapping epitopes (e.g., PGT151 inhibition of 8ANC195 binding); allosteric inhibition (e.g., PGT145 inhibition of 1NC9, 8ANC195, PGT151 and CD4 binding); and competition by reorientation of glycans (e.g., PGT135 inhibition of CD4bs bNAbs, and CD4bs bNAb inhibition of 8ANC195). We further demonstrate that bNAb binding can be complex, often affecting several other areas of the trimer surface beyond the epitope. This extensive analysis of the antigenic structure and the epitope interrelationships of the Env trimer should aid in design of both bNAb-based therapies and vaccines intended to induce bNAbs.
    PLoS Pathogens 03/2015; 11(3):e1004767. DOI:10.1371/journal.ppat.1004767 · 8.06 Impact Factor
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    ABSTRACT: HIV's envelope glycoprotein (Env) is the sole target for neutralizing antibodies. The structures of many broadly neutralizing antibodies (bNAbs) in complex with truncated Env subunits or components have been reported. However, their interaction with the intact Env trimer, and the structural determinants that underlie neutralization resistance in this more native context are less well understood. Here we use hydrogen/deuterium exchange to examine the interactions between a panel of bNAbs and native-like Env trimers (SOSIP.664 trimers). Highly potent bNAbs cause only localized effects at their binding interface, while the binding of less potent antibodies is associated with elaborate changes throughout the trimer. In conjunction with binding kinetics, our results suggest that poorly neutralizing antibodies can only bind when the trimer transiently samples an open state. We propose that the kinetics of such opening motions varies among isolates, with Env from neutralization-sensitive viruses opening more frequently than Env from resistant viruses.
    Nature Communications 02/2015; 6:6144. DOI:10.1038/ncomms7144 · 10.74 Impact Factor
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    ABSTRACT: Many therapeutic proteins and protein subunit vaccines contain heterologous trimerization domains, such as the widely used GCN4-based isoleucine zipper (IZ) and the T4 bacteriophage fibritin foldon (Fd) trimerization domains. We found that these domains induced potent anti-IZ or anti-Fd antibody responses in animals when fused to an HIV-1 envelope glycoprotein (Env) immunogen. To dampen IZ-induced responses, we constructed an IZ domain containing four N-linked glycans (IZN4) to shield the underlying protein surface. When fused to two different vaccine antigens, HIV-1 Env and influenza hemagglutinin (HA), IZN4 strongly reduced the antibody responses against the IZ, but did not affect the antibody titers against Env or HA. Silencing of immunogenic multimerization domains with glycans might be relevant for therapeutic proteins and protein vaccines. Copyright © 2015, The American Society for Biochemistry and Molecular Biology.
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    ABSTRACT: Recombinant trimeric mimics of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) spike should expose as many epitopes as possible for broadly neutralizing antibodies (bNAbs), but few if any for non-neutralizing antibodies (non-NAbs). Soluble, cleaved SOSIP.664 gp140 trimers based on the subtype A strain, BG505, approach this ideal and are therefore plausible vaccine candidates. Here, we report on the production and in vitro properties of a new SOSIP.664 trimer derived from a subtype B env gene, B41, including how to make this protein in low-serum media without proteolytic damage (clipping) to the V3 region. We also show that non-clipped trimers can be purified successfully via a positive selection affinity column using the bNAb PGT145, which recognizes a quaternary epitope at the trimer apex. Negative-stain electron microscopy imaging shows that the purified, non-clipped, native-like B41 SOSIP.664 trimers contain two sub-populations, which we propose represent an equilibrium between the fully closed and a more open conformation. The latter is different from the fully open, CD4 receptor-bound conformation and may represent an intermediate state of the trimer. This new subtype B trimer adds to the repertoire of native-like Env proteins that are suitable for immunogenicity and structural studies. The cleaved, trimeric envelope protein complex is the only neutralizing antibody target on the HIV-1 surface. Many vaccine strategies are based on inducing neutralizing antibodies. For HIV-1, one approach involves using recombinant, soluble protein mimics of the native trimer. At present, the only reliable way to make native-like, soluble trimers in practical amounts is via the introduction of specific sequence changes that confer stability to the cleaved form of Env. The resulting trimers are known as SOSIP.664 gp140s, and the current paradigm is based on the BG505 subtype A env gene. Here, we describe the production and characterization of a SOSIP.664 trimer derived from a subtype B gene (B41), together with a simple, one-step method to purify native-like trimers by affinity chromatography with a quaternary epitope-specific bNAb, PGT145. The resulting trimers will be useful for structural and immunogenicity experiments aimed at devising ways to make an effective HIV-1 vaccine. Copyright © 2015, American Society for Microbiology. All Rights Reserved.
    Journal of Virology 01/2015; DOI:10.1128/JVI.03473-14 · 4.65 Impact Factor
  • Rogier W Sanders, John P Moore
    Nature 10/2014; 514(7523). DOI:10.1038/nature13926 · 42.35 Impact Factor
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    ABSTRACT: Broadly neutralizing antibodies (bnAbs) targeting the trimer apex of HIV envelope are favored candidates for vaccine design and immunotherapy because of their great neutralization breadth and potency. However, methods of isolating bnAbs against this site have been limited by the quaternary nature of the epitope region. Here we report the use of a recombinant HIV envelope trimer, BG505 SOSIP.664 gp140, as an affinity reagent to isolate quaternary-dependent bnAbs from the peripheral blood mononuclear cells of a chronically infected donor. The newly isolated bnAbs, named "PGDM1400-1412," show a wide range of neutralization breadth and potency. One of these variants, PGDM1400, is exceptionally broad and potent with cross-clade neutralization coverage of 83% at a median IC50 of 0.003 µg/mL. Overall, our results highlight the utility of BG505 SOSIP.664 gp140 as a tool for the isolation of quaternary-dependent antibodies and reveal a mosaic of antibody responses against the trimer apex within a clonal family.
    Proceedings of the National Academy of Sciences 10/2014; 30 Suppl 1(49):A7-8. DOI:10.1089/aid.2014.5002.abstract · 9.81 Impact Factor
  • Marit J. van Gils, Rogier W. Sanders
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    ABSTRACT: Passive immunization studies, including a recent one by Pegu et al., have repeatedly shown that HIV-specific broadly neutralizing antibodies (bnAbs) protect rhesus macaques from HIV acquisition. In vitro neutralization potency and in vivo protection correlate very strongly, supporting the quest for an HIV vaccine that induces potent bnAbs.
    Trends in Microbiology 10/2014; 22(10). DOI:10.1016/j.tim.2014.08.006 · 9.81 Impact Factor
  • AIDS Research and Human Retroviruses 10/2014; 30 Suppl 1:A66-7. DOI:10.1089/aid.2014.5121.abstract · 2.46 Impact Factor
  • AIDS Research and Human Retroviruses 10/2014; 30 Suppl 1:A49. DOI:10.1089/aid.2014.5084.abstract · 2.46 Impact Factor
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    ABSTRACT: Current HIV-1 vaccines based on the HIV-1 envelope glycoprotein spike (Env), the only relevant target for broadly neutralizing antibodies, are unable to induce protective immunity. Env immunogenicity can be enhanced by fusion to costimulatory molecules involved in B cell activation, such as APRIL and CD40L. Here, we found that Env-APRIL signaled through the two receptors, BCMA and TACI. In rabbits, Env-APRIL induced significantly higher antibody responses against Env compared to unconjugated Env, while the antibody responses against the APRIL component were negligible. To extend this finding, we tested Env-APRIL in mice and found minimal antibody responses against APRIL. Furthermore, Env-CD40L did not induce significant anti-CD40L responses. Thus, in contrast to the 4-helix cytokines IL-21 and GM-CSF, the TNF-superfamily members CD40L and APRIL induced negligible autoantibodies. This study confirms and extends previous work and shows that fusion of Env-based immunogens to APRIL can improve Env immunogenicity and might help in designing HIV vaccines that induce protective humoral immunity.
    PLoS ONE 09/2014; 9(9):e107683. DOI:10.1371/journal.pone.0107683 · 3.53 Impact Factor
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    ABSTRACT: The isolation of human monoclonal antibodies is providing important insights into the specificities that underlie broad neutralization of HIV-1 (reviewed in ref. 1). Here we report a broad and extremely potent HIV-specific monoclonal antibody, termed 35O22, which binds a novel HIV-1 envelope glycoprotein (Env) epitope. 35O22 neutralized 62% of 181 pseudoviruses with a half-maximum inhibitory concentration (IC50) <50 μg ml(-1). The median IC50 of neutralized viruses was 0.033 μg ml(-1), among the most potent thus far described. 35O22 did not bind monomeric forms of Env tested, but did bind the trimeric BG505 SOSIP.664. Mutagenesis and a reconstruction by negative-stain electron microscopy of the Fab in complex with trimer revealed that it bound to a conserved epitope, which stretched across gp120 and gp41. The specificity of 35O22 represents a novel site of vulnerability on HIV Env, which serum analysis indicates to be commonly elicited by natural infection. Binding to this new site of vulnerability may thus be an important complement to current monoclonal-antibody-based approaches to immunotherapies, prophylaxis and vaccine design.
    Nature 09/2014; 515(7525). DOI:10.1038/nature13601 · 42.35 Impact Factor
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    ABSTRACT: The HIV envelope glycoprotein (Env) trimer undergoes receptor-induced conformational changes that drive fusion of the viral and cellular membranes. Env conformational changes have been observed using low-resolution electron microscopy, but only large-scale rearrangements have been visible. Here, we use hydrogen-deuterium exchange and oxidative labeling to gain a more precise understanding of the unliganded and CD4-bound forms of soluble Env trimers (SOSIP.664), including their glycan composition. CD4 activation induces the reorganization of bridging sheet elements, V1/V2 and V3, much of the gp120 inner domain, and the gp41 fusion subunit. Two CD4 binding site-targeted inhibitors have substantially different effects: NBD-556 partially mimics CD4-induced destabilization of the V1/V2 and V3 crown, whereas BMS-806 only affects regions around the gp120/gp41 interface. The structural information presented here increases our knowledge of CD4- and small molecule-induced conformational changes in Env and the allosteric pathways that lead to membrane fusion.
    Structure 06/2014; 22(7). DOI:10.1016/j.str.2014.05.001 · 6.79 Impact Factor
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    ABSTRACT: The trimeric envelope glycoproteins (Env) on the surface of HIV-1 virions are the targets for neutralizing antibodies (NAbs). No candidate HIV-1 immunogen has yet induced potent, broadly active NAbs (bNAbs). Part of the explanation may be that previously tested Env proteins inadequately mimic the functional, native Env complex. Trimerization and the proteolytic processing of Env precursors into gp120 and gp41 profoundly alter antigenicity, but soluble cleaved trimers are too unstable to serve as immunogens. By introducing stabilizing mutations (SOSIP), we constructed soluble, cleaved Env trimers derived from the HIV-1 subtype A isolate BG505 that resemble native Env spikes on virions both structurally and antigenically.
    Retrovirology 05/2014; 11(1):41. DOI:10.1186/1742-4690-11-41 · 4.77 Impact Factor
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    ABSTRACT: Recombinant soluble, cleaved HIV-1 envelope glycoprotein SOSIP.664 gp140 trimers based on the subtype A BG505 sequence are being studied structurally and tested as immunogens in animals. For these trimers to become a vaccine candidate for human trials, they would need to be made in appropriate amounts at an acceptable quality. Accomplishing such tasks by transient transfection is likely to be challenging. The traditional way to express recombinant proteins in large amounts is via a permanent cell line, usually of mammalian origin. Making cell lines that produce BG505 SOSIP.664 trimers requires the co-expression of the Furin protease to ensure that the cleavage site between the gp120 and gp41 subunits is fully utilized. We designed a vector capable of expressing Env and Furin, and used it to create Stable 293 T and CHO Flp-InTMcell lines through site-specific recombination. Both lines produce high quality, cleaved trimers at yields of up to 12-15 mg per 1 x 109 cells. Trimer expression at such levels was maintained for up to 30 days (10 passages) after initial seeding and was consistently superior to what could be achieved by transient transfection. Electron microscopy studies confirm that the purified trimers have the same native-like appearance as those derived by transient transfection and used to generate high-resolution structures. They also have appropriate antigenic properties, including the presentation of the quaternary epitope for the broadly neutralizing antibody PGT145. The BG505 SOSIP.664 trimer-expressing cell lines yield proteins of an appropriate quality for structural studies and animal immunogenicity experiments. The methodology is suitable for making similar lines under Good Manufacturing Practice conditions, to produce trimers for human clinical trials. Moreover, any env gene can be incorporated into this vector system, allowing the manufacture of SOSIP trimers from multiple genotypes, either by transient transfection or from stable cell lines.
    Retrovirology 04/2014; 11(1):33. DOI:10.1186/1742-4690-11-33 · 4.77 Impact Factor
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    ABSTRACT: Broadly neutralizing HIV antibodies are much sought after (a) to guide vaccine design, both as templates and as indicators of the authenticity of vaccine candidates, (b) to assist in structural studies, and (c) to serve as potential therapeutics. However, the number of targets on the viral envelope spike for such antibodies has been limited. Here, we describe a set of human monoclonal antibodies that define what is, to the best of our knowledge, a previously undefined target on HIV Env. The antibodies recognize a glycan-dependent epitope on the prefusion conformation of gp41 and unambiguously distinguish cleaved from uncleaved Env trimers, an important property given increasing evidence that cleavage is required for vaccine candidates that seek to mimic the functional HIV envelope spike. The availability of this set of antibodies expands the number of vaccine targets on HIV and provides reagents to characterize the native envelope spike.
    Immunity 04/2014; DOI:10.1016/j.immuni.2014.04.009 · 19.75 Impact Factor
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    ABSTRACT: All previously characterized broadly neutralizing antibodies to the HIV-1 envelope glycoprotein (Env) target one of four major sites of vulnerability. Here, we define and structurally characterize a unique epitope on Env that is recognized by a recently discovered family of human monoclonal antibodies (PGT151-PGT158). The PGT151 epitope is comprised of residues and glycans at the interface of gp41 and gp120 within a single protomer and glycans from both subunits of a second protomer and represents a neutralizing epitope that is dependent on both gp120 and gp41. Because PGT151 binds only to properly formed, cleaved trimers, this distinctive property, and its ability to stabilize Env trimers, has enabled the successful purification of mature, cleaved Env trimers from the cell surface as a complex with PGT151. Here we compare the structural and functional properties of membrane-extracted Env trimers from several clades with those of the soluble, cleaved SOSIP gp140 trimer.
    Immunity 04/2014; DOI:10.1016/j.immuni.2014.04.008 · 19.75 Impact Factor

Publication Stats

3k Citations
800.90 Total Impact Points

Institutions

  • 2002–2015
    • Weill Cornell Medical College
      • • Department of Microbiology and Immunology
      • • Department of Biochemistry
      New York, New York, United States
    • Progenics Pharmaceuticals, Inc.
      태리타운, New York, United States
  • 2009–2014
    • Academisch Medisch Centrum Universiteit van Amsterdam
      • Department of Medical Microbiology
      Amsterdamo, North Holland, Netherlands
    • Cornell University
      • Department of Microbiology and Immunology
      Итак, New York, United States
  • 2001–2014
    • University of Amsterdam
      • Department of Medical Microbiology
      Amsterdamo, North Holland, Netherlands
  • 2011
    • Duke University Medical Center
      • Department of Surgery
      Durham, North Carolina, United States
  • 2008
    • Utrecht University
      • Division of Cellular Protein Chemistry
      Utrecht, Utrecht, Netherlands
  • 2006
    • Academisch Centrum Tandheelkunde Amsterdam
      • Field of Oral Biochemistry
      Amsterdamo, North Holland, Netherlands