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Gold nanoparticles (GNPs) decorated with glycans ameliorate dendritic cells (DC) uptake, antigen-presentation and T-cells cross-talk, which are important aspects in vaccine design. GNPs allow for high antigen loading, DC targeting, lack of toxicity and are straightforward prepared and easy to handle. The present study aimed to assess the capacity of DC to process and present HIV-1-peptides loaded onto GNPs bearing high-mannoside-type oligosaccharides (P1@HM) to autologous T-cells from HIV-1 patients. The results showed that P1@HM increased HIV-specific CD4 + and CD8 + T-cell proliferation and induced highly functional cytokine secretion compared with HIV-peptides alone. P1@HM elicits a highly efficient secretion of pro-T H 1 cytokines and chemokines, a moderate production of pro-T H 2 and significant higher secretion of pro-inflammatory cytokines such as TNF-α and IL-1β. Thus, co-delivery of HIV-1 antigens and HM by GNPs is an excellent vaccine delivery system inducing HIV-specific cellular immune responses in HIV+ patients, being a promising approach to improve anti-HIV-1 vaccines., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Improved detection of anti-carbohydrate antibodies is a need in clinical identification of biomarkers for cancer cells or pathogens. Here, we report a new ELISA approach for the detection of specific immunoglobulins (IgGs) against carbohydrates. Two nanometer gold glyconanoparticles bearing oligosaccharide epitopes of HIV or Streptococcus pneumoniae were used as antigens to coat ELISA-plates. A ~3,000-fold improved detection of specific IgGs in mice immunized against S. pneumoniae respect to the well known BSA-glycoconjugate ELISA was achieved. Moreover, these multivalent glyconanoparticles have been employed in solid phase assays to detect the carbohydrate-dependent binding of human dendritic cells and the lectin DC-SIGN. Multivalent glyconanoparticles in ELISA provide a versatile, easy and highly sensitive method to detect and quantify the binding of glycan to proteins and to facilitate the identification of biomarkers.

Broadly neutralizing anti-HIV-1 monoclonal antibody 2G12 exclusively targets a conserved cluster of high-mannose oligosaccharides present on outer viral envelope glycoprotein gp120. This characteristic makes the otherwise immunogenically "silent" glycan shield of gp120 a tempting target for drug and vaccine design. However, immune responses against carbohydrate-based mimics of gp120 have failed to provide immunization against HIV-1 infection, highlighting the need to understand the molecular events that determine immunogenicity better. In this work, the unbinding kinetics of the gp120-2G12 (k(0) = 0.002 ± 0.09 s(-1), x(++) = 1.5 ± 1.2 nm), Man(4)-2G12 (k(0) = 0.35 ± 0.32 s(-1), x(++) = 0.6 ± 0.2 nm), and Man(5)-2G12 interactions were measured by single-molecule force spectroscopy. To our knowledge, this is the first single-molecule study aimed at dissecting the carbohydrate-antibody recognition of the gp120-2G12 interaction. We were able to confirm crystallographic models that show both the binding of the linear Man(4) arm to 2G12 and also the multivalent gp120 glycan binding to 2G12. These results demonstrate that single-molecule force spectroscopy can be successfully used to dissect the molecular mechanisms underlying immunity.

After nearly three decades since the discovery of human immunodeficiency virus (HIV) (1983), no effective vaccine or microbicide is available, and the virus continues to infect millions of people worldwide each year. HIV antiretroviral drugs reduce the death rate and improve the quality of life in infected patients, but they are not able to completely remove HIV from the body. The glycoprotein gp120, part of the envelope glycoprotein (Env) of HIV, is responsible for virus entry and infection of host cells. High-mannose type glycans that decorate gp120 are involved in different carbohydrate-mediated HIV binding. We have demonstrated that oligomannoside-coated gold nanoparticles (manno-GNPs) are able to interfere with HIV high-mannose glycan-mediated processes. In this chapter, we describe the methods for the preparation and characterization of manno-GNPs and the experiments performed by means of SPR and STD-NMR techniques to evaluate the ability of manno-GNPs to inhibit 2G12 antibody binding to gp120. The antibody 2G12-mediated HIV neutralization and the lectin DC-SIGN-mediated HIV trans-infection in cellular systems are also described., (Copyright © 2012 Elsevier Inc. All rights reserved.)

The structural and affinity details of the interactions of synthetic oligomannosides, linear (di-, tri-, and tetra-) and branched (penta- and hepta-), with the broadly neutralizing anti-HIV-1 antibody 2G12 (HIV=human immunodeficiency virus) have been investigated in solution by using ligand-based NMR techniques, specifically saturation transfer difference (STD) NMR spectroscopy and transferred NOE experiments. Linear oligomannosides show similar binding modes to the antibody, with the nonreducing terminal disaccharide Manα(1→2)Man (Man=mannose) making the closest protein/ligand contacts in the bound state. In contrast, the branched pentamannoside shows two alternate binding modes, involving both ligand arms (D2- and D3-like), a dual binding description of the molecular recognition of this ligand by 2G12 in solution that differs from the single binding mode deduced from X-ray studies. On the contrary, the antibody shows an unexpected selectivity for one arm (D1-like) of the other branched ligand (heptamannoside). This result explains the previously reported lack of affinity enhancement relative to that of the D1-like tetramannoside. Single-ligand STD NMR titration experiments revealed noticeable differences in binding affinities among the linear and branched ligands in solution, with the latter showing decreased affinity. Among the analyzed series of ligands, the strongest 2G12 binders were the linear tri- and tetramannosides because both show similar affinity for the antibody. These results demonstrate that NMR spectroscopic techniques can deliver abundant structural, dynamics, and affinity information for the characterization of oligomannose-2G12 binding in solution, thus complementing, and, as in the case of the pentamannoside, extending, the structural view from X-ray crystallography. This information is of key importance for the development of multivalent synthetic gp120 high-mannose glycoconjugate mimics in the context of vaccine development., (Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Gold nanoparticles (GNPs) decorated with glycans ameliorate dendritic cells (DC) uptake, antigen-presentation and T-cells cross-talk, which are important aspects in vaccine design. GNPs allow for high antigen loading, DC targeting, lack of toxicity and are straightforward prepared and easy to handle. The present study aimed to assess the capacity of DC to process and present HIV-1-peptides loaded onto GNPs bearing high-mannoside-type oligosaccharides (P1@HM) to autologous T-cells from HIV-1 patients. The results showed that P1@HM increased HIV-specific CD4 + and CD8 + T-cell proliferation and induced highly functional cytokine secretion compared with HIV-peptides alone. P1@HM elicits a highly efficient secretion of pro-T H 1 cytokines and chemokines, a moderate production of pro-T H 2 and significant higher secretion of pro-inflammatory cytokines such as TNF‐α and IL-1β. Thus, co-delivery of HIV-1 antigens and HM by GNPs is an excellent vaccine delivery system inducing HIV-specific cellular immune responses in HIV+ patients, being a promising approach to improve anti-HIV-1 vaccines. [ABSTRACT FROM AUTHOR]