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Journal of Virology Mar 2015The isolation of broadly neutralizing HIV-1 monoclonal antibodies (MAbs) to distinct epitopes on the viral envelope glycoprotein (Env) provides the potential to use...
UNLABELLED
The isolation of broadly neutralizing HIV-1 monoclonal antibodies (MAbs) to distinct epitopes on the viral envelope glycoprotein (Env) provides the potential to use combinations of MAbs for prevention and treatment of HIV-1 infection. Since many of these MAbs have been isolated in the last few years, the potency and breadth of MAb combinations have not been well characterized. In two parallel experiments, we examined the in vitro neutralizing activities of double-, triple-, and quadruple-MAb combinations targeting four distinct epitopes, including the CD4-binding site, the V1V2-glycan region, the V3-glycan supersite, and the gp41 membrane-proximal external region (MPER), using a panel of 125 Env-pseudotyped viruses. All MAb combinations showed substantially improved neutralization breadth compared to the corresponding single MAbs, while the neutralization potency of individual MAbs was maintained. At a 50% inhibitory concentration (IC50) cutoff of 1 μg/ml per antibody, double-MAb combinations neutralized 89 to 98% of viruses, and triple combinations neutralized 98 to 100%. Overall, the improvement of neutralization breadth was closely predicted by an additive-effect model and explained by complementary neutralization profiles of antibodies recognizing distinct epitopes. Subtle but consistent favorable interactions were observed in some MAb combinations, whereas less favorable interactions were observed on a small subset of viruses that are highly sensitive to V3-glycan MAbs. These data demonstrate favorable in vitro combinations of broadly neutralizing HIV-1 MAbs and suggest that such combinations could have utility for HIV-1 prevention and treatment.
IMPORTANCE
Over the last 5 years, numerous broadly reactive HIV-1-neutralizing MAbs have been isolated from B cells of HIV-1-infected donors. Each of these MAbs binds to one of the major vulnerable sites (epitopes) on the surface of the viral envelope glycoprotein. Since antibodies to distinct viral epitopes could theoretically act together to provide greater potency and breadth of virus neutralization, we tested physical mixtures of double, triple, and quadruple combinations of neutralizing MAbs targeting four major epitopes on HIV-1 Env. When tested together, antibody combinations showed substantially improved neutralization breadth compared to single MAbs. This improvement could be explained by the complementary neutralization profiles of individual MAbs. We further demonstrated that each antibody maintained its full neutralization potency when used in combination with other MAbs. These data provide a rationale for clinical use of antibody-based combinations for HIV-1 prevention and therapy.
Topics: Anti-HIV Agents; Antibodies, Neutralizing; Drug Interactions; Epitopes; HIV Antibodies; HIV-1; Humans; Inhibitory Concentration 50; Neutralization Tests
PubMed: 25520506
DOI: 10.1128/JVI.03136-14 -
Journal of Internal Medicine Jul 2007
Review
Topics: AIDS Vaccines; HIV Antibodies; HIV Infections; HIV-1; Humans; Immune Tolerance; Immunity, Innate
PubMed: 17598811
DOI: 10.1111/j.1365-2796.2007.01835.x -
Journal of Immunological Methods Apr 2020The recent identification of human monoclonal antibodies with broad and potent neutralizing activity against HIV-1 (bnAbs) has resulted in substantial efforts to develop...
The recent identification of human monoclonal antibodies with broad and potent neutralizing activity against HIV-1 (bnAbs) has resulted in substantial efforts to develop these molecules for clinical use in the prevention and treatment of HIV-1 infection. As with any protein therapeutic drug product, it is imperative to have qualified assays that can accurately detect and quantify anti-drug antibodies (ADA) that may develop in patients receiving passive administration of HIV-1 bnAbs. Here, we have optimized and qualified a functional assay to assess the potential of ADA to inhibit the neutralizing function of HIV-1 bnAbs. Using a modified version of the validated TZM-bl HIV-1 neutralization assay, murine anti-idiotype antibodies were utilized to optimize and evaluate parameters of linearity, range, limit of detection, specificity, and precision for measuring inhibitory ADA activity against multiple HIV-1 bnAbs that are in clinical development. We further demonstrate the utility of this assay for detecting naturally occurring ADA responses in non-human primates receiving passive administration of human bnAbs. This functional assay format complements binding-antibody ADA strategies being developed for HIV-1 bnAbs, and when utilized together, will support a multi-tiered approach for ADA testing that is compliant with Good Clinical Laboratory Practice (GCLP) procedures and FDA guidance.
Topics: Animals; Antibodies, Anti-Idiotypic; Antibodies, Monoclonal, Murine-Derived; Broadly Neutralizing Antibodies; HIV Antibodies; HIV Infections; HIV-1; Humans; Mice; Neutralization Tests
PubMed: 31917969
DOI: 10.1016/j.jim.2020.112736 -
PloS One 2014One strategy for isolating or eliciting antibodies against a specific target region on the envelope glycoprotein trimer (Env) of the human immunodeficiency virus type 1...
One strategy for isolating or eliciting antibodies against a specific target region on the envelope glycoprotein trimer (Env) of the human immunodeficiency virus type 1 (HIV-1) involves the creation of site transplants, which present the target region on a heterologous protein scaffold with preserved antibody-binding properties. If the target region is a supersite of HIV-1 vulnerability, recognized by a collection of broadly neutralizing antibodies, this strategy affords the creation of "supersite transplants", capable of binding (and potentially eliciting) antibodies similar to the template collection of effective antibodies. Here we transplant three supersites of HIV-1 vulnerability, each targeted by effective neutralizing antibodies from multiple donors. To implement our strategy, we chose a single representative antibody against each of the target supersites: antibody 10E8, which recognizes the membrane-proximal external region (MPER) on the HIV-1 gp41 glycoprotein; antibody PG9, which recognizes variable regions one and two (V1V2) on the HIV-1 gp120 glycoprotein; and antibody PGT128 which recognizes a glycopeptide supersite in variable region 3 (glycan V3) on gp120. We used a structural alignment algorithm to identify suitable acceptor proteins, and then designed, expressed, and tested antigenically over 100-supersite transplants in a 96-well microtiter-plate format. The majority of the supersite transplants failed to maintain the antigenic properties of their respective template supersite. However, seven of the glycan V3-supersite transplants exhibited nanomolar affinity to effective neutralizing antibodies from at least three donors and recapitulated the mannose9-N-linked glycan requirement of the template supersite. The binding of these transplants could be further enhanced by placement into self-assembling nanoparticles. Essential elements of the glycan V3 supersite, embodied by as few as 3 N-linked glycans and ∼ 25 Env residues, can be segregated into acceptor scaffolds away from the immune-evading capabilities of the rest of HIV-1 Env, thereby providing a means to focus the immune response on the scaffolded supersite.
Topics: Antibodies, Neutralizing; HIV Antibodies; HIV Envelope Protein gp120; HIV Envelope Protein gp41; HIV-1; Humans; Models, Molecular
PubMed: 24992528
DOI: 10.1371/journal.pone.0099881 -
Frontiers in Immunology 2021Accumulation of somatic hypermutation (SHM) is the primary mechanism to enhance the binding affinity of antibodies to antigens . However, the structural basis of the...
Accumulation of somatic hypermutation (SHM) is the primary mechanism to enhance the binding affinity of antibodies to antigens . However, the structural basis of the effects of many SHMs remains elusive. Here, we integrated atomistic molecular dynamics (MD) simulation and data mining to build a high-throughput structural bioinformatics pipeline to study the effects of individual and combination SHMs on antibody conformation, flexibility, stability, and affinity. By applying this pipeline, we characterized a common mechanism of modulation of heavy-light pairing orientation by frequent SHMs at framework positions 39, 91, 38, and 87 through disruption of a conserved hydrogen-bond network. Q39L alone and in combination with light chain framework 4 (FWR4) insertions further modulated the elbow angle between variable and constant domains of many antibodies, resulting in improved binding affinity for a subset of anti-HIV-1 antibodies. Q39L also alleviated aggregation induced by FWR4 insertion, suggesting remote epistasis between these SHMs. Altogether, this study provides tools and insights for understanding antibody affinity maturation and for engineering functionally improved antibodies.
Topics: Animals; Antibodies; Antibody Affinity; HIV Antibodies; HIV-1; Humans; Molecular Conformation; Molecular Dynamics Simulation; Somatic Hypermutation, Immunoglobulin
PubMed: 35046963
DOI: 10.3389/fimmu.2021.811632 -
Transgenic Research Aug 2009The first evidence that plants represent a valid, safe and cost-effective alternative to traditional expression systems for large-scale production of antigens and... (Review)
Review
The first evidence that plants represent a valid, safe and cost-effective alternative to traditional expression systems for large-scale production of antigens and antibodies was described more than 10 years ago. Since then, considerable improvements have been made to increase the yield of plant-produced proteins. These include the use of signal sequences to target proteins to different cellular compartments, plastid transformation to achieve high transgene dosage, codon usage optimization to boost gene expression, and protein fusions to improve recombinant protein stability and accumulation. Thus, several HIV/SIV antigens and neutralizing anti-HIV antibodies have recently been successfully expressed in plants by stable nuclear or plastid transformation, and by transient expression systems based on plant virus vectors or Agrobacterium-mediated infection. The current article gives an overview of plant expressed HIV antigens and antibodies and provides an account of the use of different strategies aimed at increasing the expression of the accessory multifunctional HIV-1 Nef protein in transgenic plants.
Topics: Genetic Vectors; HIV Antibodies; HIV Antigens; Humans; Neutralization Tests; Plants, Genetically Modified; Protein Stability; Rhizobium; nef Gene Products, Human Immunodeficiency Virus
PubMed: 19169897
DOI: 10.1007/s11248-009-9244-5 -
Journal of Molecular Biology Apr 2017Direct calculation of relative binding affinities between antibodies and antigens is a long-sought goal. However, despite substantial efforts, no generally applicable...
Direct calculation of relative binding affinities between antibodies and antigens is a long-sought goal. However, despite substantial efforts, no generally applicable computational method has been described. Here, we describe a systematic free energy perturbation (FEP) protocol and calculate the binding affinities between the gp120 envelope glycoprotein of HIV-1 and three broadly neutralizing antibodies (bNAbs) of the VRC01 class. The protocol has been adapted from successful studies of small molecules to address the challenges associated with modeling protein-protein interactions. Specifically, we built homology models of the three antibody-gp120 complexes, extended the sampling times for large bulky residues, incorporated the modeling of glycans on the surface of gp120, and utilized continuum solvent-based loop prediction protocols to improve sampling. We present three experimental surface plasmon resonance data sets, in which antibody residues in the antibody/gp120 interface were systematically mutated to alanine. The RMS error in the large set (55 total cases) of FEP tests as compared to these experiments, 0.68kcal/mol, is near experimental accuracy, and it compares favorably with the results obtained from a simpler, empirical methodology. The correlation coefficient for the combined data set including residues with glycan contacts, R=0.49, should be sufficient to guide the choice of residues for antibody optimization projects, assuming that this level of accuracy can be realized in prospective prediction. More generally, these results are encouraging with regard to the possibility of using an FEP approach to calculate the magnitude of protein-protein binding affinities.
Topics: Antibodies, Neutralizing; Computational Biology; HIV Antibodies; HIV Envelope Protein gp120; Protein Binding; Surface Plasmon Resonance; Thermodynamics
PubMed: 27908641
DOI: 10.1016/j.jmb.2016.11.021 -
Proceedings of the National Academy of... Oct 2005HIV has evolved many strategies to avoid neutralizing antibody responses, particularly to conserved regions on the external glycoprotein spikes of the virus.... (Review)
Review
HIV has evolved many strategies to avoid neutralizing antibody responses, particularly to conserved regions on the external glycoprotein spikes of the virus. Nevertheless, a small number of antibodies have been evolved by the human immune system to recognize conserved parts of the glycoproteins, and therefore, have broadly neutralizing cross-strain activities. These antibodies constitute important tools in the quest to design immunogens that can elicit broadly neutralizing antibodies in humans and hence contribute to an effective HIV vaccine. Crystallographic analyses of the antibodies, in many cases in an antigen-complexed form, have revealed novel and, in some instances, remarkable structural adaptations to attain virus recognition. Antibodies, like HIV, can evolve relatively rapidly through mutation and selection. It seems that the structures of these broadly neutralizing antibodies bear witness to a heroic struggle between two titans of rapid evolution.
Topics: AIDS Vaccines; Antigens, Viral; Binding Sites; Evolution, Molecular; Glycoproteins; HIV; HIV Antibodies; HIV Envelope Protein gp120; Humans; Neutralization Tests; Protein Conformation
PubMed: 16219699
DOI: 10.1073/pnas.0505126102 -
Immunity Sep 2017Apex broadly neutralizing HIV antibodies (bnAbs) recognize glycans and protein surface close to the 3-fold axis of the envelope (Env) trimer and are among the most...
Apex broadly neutralizing HIV antibodies (bnAbs) recognize glycans and protein surface close to the 3-fold axis of the envelope (Env) trimer and are among the most potent and broad Abs described. The evolution of apex bnAbs from one donor (CAP256) has been studied in detail and many Abs at different stages of maturation have been described. Using diverse engineering tools, we investigated the involvement of glycan recognition in the development of the CAP256.VRC26 Ab lineage. We found that sialic acid-bearing glycans were recognized by germline-encoded and somatically mutated residues on the Ab heavy chain. This recognition provided an "anchor" for the Abs as the core protein epitope varies, prevented complete neutralization escape, and eventually led to broadening of the response. These findings illustrate how glycan-specific maturation enables a human Ab to cope with pathogen escape mechanisms and will aid in optimization of immunization strategies to induce V2 apex bnAb responses.
Topics: Amino Acid Sequence; Antibodies, Neutralizing; Antibody Affinity; Antibody Formation; Binding Sites; Epitopes; HIV Antibodies; HIV Envelope Protein gp120; HIV Infections; HIV-1; Humans; Immunoglobulin Heavy Chains; Models, Molecular; N-Acetylneuraminic Acid; Neutralization Tests; Peptide Fragments; Phylogeny; Polysaccharides; Protein Binding; Protein Conformation; Protein Multimerization
PubMed: 28916265
DOI: 10.1016/j.immuni.2017.08.006 -
ELife Jan 2021A prerequisite for the design of an HIV vaccine that elicits protective antibodies is understanding the developmental pathways that result in desirable antibody...
A prerequisite for the design of an HIV vaccine that elicits protective antibodies is understanding the developmental pathways that result in desirable antibody features. The development of antibodies that mediate antibody-dependent cellular cytotoxicity (ADCC) is particularly relevant because such antibodies have been associated with HIV protection in humans. We reconstructed the developmental pathways of six human HIV-specific ADCC antibodies using longitudinal antibody sequencing data. Most of the inferred naive antibodies did not mediate detectable ADCC. Gain of antigen binding and ADCC function typically required mutations in complementarity determining regions of one or both chains. Enhancement of ADCC potency often required additional mutations in framework regions. Antigen binding affinity and ADCC activity were correlated, but affinity alone was not sufficient to predict ADCC potency. Thus, elicitation of broadly active ADCC antibodies may require mutations that enable high-affinity antigen recognition along with mutations that optimize factors contributing to functional ADCC activity.
Topics: AIDS Vaccines; Antibody-Dependent Cell Cytotoxicity; Cell Line; HIV Antibodies; HIV-1; Humans
PubMed: 33427196
DOI: 10.7554/eLife.63444