scholarly journals Antibody Recognition of the Dengue Virus Proteome and Implications for Development of Vaccines

2011 ◽  
Vol 18 (4) ◽  
pp. 523-532 ◽  
Author(s):  
Stefan Fernandez ◽  
Emily D. Cisney ◽  
Alexander P. Tikhonov ◽  
Barry Schweitzer ◽  
Robert J. Putnak ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Vaccine Development ◽  
Rhesus Macaques ◽  
Protein Microarray ◽  
Vaccination Strategy ◽  
Antibody Responses ◽  
Wild Type ◽  
Antibody Recognition ◽  
Immune Correlates ◽  
The Tropics

ABSTRACTDengue is a mosquito-borne infection caused by four distinct serotypes of dengue virus, each appearing cyclically in the tropics and subtropics along the equator. Although vaccines are currently under development, none are available to the general population. One of the main impediments to the successful advancement of these vaccines is the lack of well-defined immune correlates of protection. Here, we describe a protein microarray approach for measuring antibody responses to the complete viral proteome comprised of the structural (capsid, membrane, and envelope) and nonstructural (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5) components of all four dengue virus serotypes (1 to 4). We examined rhesus macaques vaccinated with tetravalent vaccines consisting of live-attenuated virus (LAV) or purified inactivated virus (PIV), followed by boosting with LAV and challenging with wild-type dengue virus. We detected temporal increases in antibodies against envelope proteins in response to either vaccine, while only the PIV/LAV vaccination strategy resulted in anticapsid antibodies. In contrast to results from vaccination, naïve macaques challenged with wild-type viruses of each serotype demonstrated a balanced response to nonstructural and structural components, including responses against the membrane protein. Our results demonstrate discriminating details concerning the nature of antibody responses to dengue virus at the proteomic level and suggest the usefulness of this information for vaccine development.

scholarly journals Determination of Specific Antibody Responses to the Six Species of Ebola and Marburg Viruses by Multiplexed Protein Microarrays

2014 ◽  
Vol 21 (12) ◽  
pp. 1605-1612 ◽  
Author(s):  
Teddy Kamata ◽  
Mohan Natesan ◽  
Kelly Warfield ◽  
M. Javad Aman ◽  
Robert G. Ulrich
Keyword(s):  
Rhesus Macaques ◽  
Protein Microarray ◽  
Serum Antibody ◽  
Protein Microarrays ◽  
Marburg Virus ◽  
Antibody Responses ◽  
Diagnostic Methods ◽  
Virion Protein ◽  
Content Type ◽  
Antibody Recognition

ABSTRACTInfectious hemorrhagic fevers caused by the Marburg and Ebola filoviruses result in human mortality rates of up to 90%, and there are no effective vaccines or therapeutics available for clinical use. The highly infectious and lethal nature of these viruses highlights the need for reliable and sensitive diagnostic methods. We assembled a protein microarray displaying nucleoprotein (NP), virion protein 40 (VP40), and glycoprotein (GP) antigens from isolates representing the six species of filoviruses for use as a surveillance and diagnostic platform. Using the microarrays, we examined serum antibody responses of rhesus macaques vaccinated with trivalent (GP, NP, and VP40) virus-like particles (VLP) prior to infection with the Marburg virus (MARV) (i.e.,Marburg marburgvirus) or the Zaire virus (ZEBOV) (i.e.,Zaire ebolavirus). The microarray-based assay detected a significant increase in antigen-specific IgG resulting from immunization, while a greater level of antibody responses resulted from challenge of the vaccinated animals with ZEBOV or MARV. Further, while antibody cross-reactivities were observed among NPs and VP40s of Ebola viruses, antibody recognition of GPs was very specific. The performance of mucin-like domain fragments of GP (GP mucin) expressed inEscherichia coliwas compared to that of GP ectodomains produced in eukaryotic cells. Based on results with ZEBOV and MARV proteins, antibody recognition of GP mucins that were deficient in posttranslational modifications was comparable to that of the eukaryotic cell-expressed GP ectodomains in assay performance. We conclude that the described protein microarray may translate into a sensitive assay for diagnosis and serological surveillance of infections caused by multiple species of filoviruses.

scholarly journals Polyclonal antibody responses to HIV Env immunogens resolved using cryoEM

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Aleksandar Antanasijevic ◽  
Leigh M. Sewall ◽  
Christopher A. Cottrell ◽  
Diane G. Carnathan ◽  
Luis E. Jimenez ◽  
...  
Keyword(s):  
High Resolution ◽  
Polyclonal Antibody ◽  
Vaccine Development ◽  
Mutational Analysis ◽  
Rhesus Macaques ◽  
Antibody Responses ◽  
Glycosylation Sites ◽  
High Resolution Analysis ◽  
Resolution Analysis ◽  
Resolution Mapping

AbstractEngineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal antibody responses without the need for monoclonal antibody isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic.

scholarly journals Immunoprofiles associated with controlled human malaria infection and naturally acquired immunity identify a shared IgA pre-erythrocytic immunoproteome

npj Vaccines ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Andrea A. Berry ◽  
Joshua M. Obiero ◽  
Mark A. Travassos ◽  
Amed Ouattara ◽  
Drissa Coulibaly ◽  
...  
Keyword(s):  
Malaria Infection ◽  
Vaccine Development ◽  
Protein Microarray ◽  
Antibody Responses ◽  
Liver Stage ◽  
Human Malaria ◽  
Controlled Human Malaria Infection ◽  
Iga Antibody ◽  
Potential Vaccine ◽  
Antigen Protein

AbstractKnowledge of the Plasmodium falciparum antigens that comprise the human liver stage immunoproteome is important for pre-erythrocytic vaccine development, but, compared with the erythrocytic stage immunoproteome, more challenging to classify. Previous studies of P. falciparum antibody responses report IgG and rarely IgA responses. We assessed IgG and IgA antibody responses in adult sera collected during two controlled human malaria infection (CHMI) studies in malaria-naïve volunteers and in 1- to 6-year-old malaria-exposed Malian children on a 251 P. falciparum antigen protein microarray. IgG profiles in the two CHMI groups were equivalent and differed from Malian children. IgA profiles were robust in the CHMI groups and a subset of Malian children. We describe immunoproteome differences in naïve vs. exposed individuals and report pre-erythrocytic proteins recognized by the immune system. IgA responses detected in this study expand the list of pre-erythrocytic antigens for further characterization as potential vaccine candidates.

scholarly journals A Universal Dengue Vaccine Elicits Neutralizing Antibodies Against Strains from All Four Dengue Serotypes

2020 ◽  
Author(s):  
Naoko Uno ◽  
Ted M. Ross
Keyword(s):  
Dengue Virus ◽  
Neutralizing Antibodies ◽  
Protective Immunity ◽  
Rhesus Macaques ◽  
Effective Vaccine ◽  
Wild Type ◽  
Monovalent Vaccine ◽  
Viral Particles ◽  
Tropical Areas

Any potential dengue virus (DENV) vaccine needs to elicit protective immunity against strains from all four serotypes to avoid potential antibody dependent enhancement (ADE). In this study, four independent DENV envelope (E) glycoproteins were generated using wild-type E sequences from viruses isolated between 1943 to 2006 using computationally-optimized broadly reactive antigen (COBRA) methodology. COBRA and wild-type E antigens were expressed on the surface of subvirion viral particles (SVPs). Four separate wild-type E antigens were used for each serotype. Mice vaccinated with wild-type DENV SVPs had anti-E IgG antibodies that neutralized serotype specific viruses. COBRA DENV SVPs elicited a broader breadth of antibodies that neutralized strains across all four serotypes. Two COBRA DENV vaccine candidates that elicited the broadest breadth of neutralizing antibodies in mice were used to vaccinate rhesus macaques (Macca mulata) that were either immunologically naïve to any DENV serotype or were had pre-existing antibodies to DENV. Antibodies elicited by COBRA DENV E immunogens neutralized all 12 strains of DENV in vitro, which was comparable to antibodies elicited by a tetravalent wild-type E SVP vaccination mixture. Therefore, using a single DENV COBRA E protein can elicit neutralizing antibodies against strains representing all four serotypes of DENV in both naïve and dengue pre-immune populations. Importance Dengue virus infects millions of people living in the tropical areas of the world. Dengue induced diseases can range from mild to severe with death. An effective vaccine will need to neutralize viruses from all four serotypes of dengue without induced enhanced disease. A dengue E vaccine candidate generated by computationally optimized broadly reactive antigen algorithms elicits broadly neutralizing protection for current circulating strains from all four serotypes regardless of immune status. Most Dengue vaccines in development formulate four separate components based on prM-E from a wild type strain representing each serotype. Designing a monovalent vaccine that elicits protective immunity against all four serotypes is an effective and economical strategy

scholarly journals Cross-Linking of a CD4-Mimetic Miniprotein with HIV-1 Env gp140 Alters Kinetics and Specificities of Antibody Responses against HIV-1 Env in Macaques

2017 ◽  
Vol 91 (19) ◽  
Author(s):  
Xiaoying Shen ◽  
Willy M. Bogers ◽  
Nicole L. Yates ◽  
Guido Ferrari ◽  
Antu K. Dey ◽  
...  
Keyword(s):  
Vaccine Development ◽  
Rhesus Macaques ◽  
Early Time ◽  
Antibody Responses ◽  
Cross Linking ◽  
Effector Functions ◽  
Igg Binding ◽  
The Cross ◽  
Hiv 1 ◽  
Time Point

ABSTRACT Evaluation of the epitope specificities, locations (systemic or mucosal), and effector functions of antibodies elicited by novel HIV-1 immunogens engineered to improve exposure of specific epitopes is critical for HIV-1 vaccine development. Utilizing an array of humoral assays, we evaluated the magnitudes, epitope specificities, avidities, and functions of systemic and mucosal immune responses elicited by a vaccine regimen containing Env cross-linked to a CD4-mimetic miniprotein (gp140-M64U1) in rhesus macaques. Cross-linking of gp140 Env to M64U1 resulted in earlier increases of both the magnitude and avidity of the IgG binding response than those with Env protein alone. Notably, IgG binding responses at an early time point correlated with antibody-dependent cellular cytotoxicity (ADCC) function at the peak immunity time point, which was higher for the cross-linked Env group than for the Env group. In addition, the cross-linked Env group developed higher IgG responses against a linear epitope in the gp120 C1 region of the HIV-1 envelope glycoprotein. These data demonstrate that structural modification of the HIV-1 envelope immunogen by cross-linking of gp140 with the CD4-mimetic M64U1 elicited an earlier increase of binding antibody responses and altered the specificity of the IgG responses, correlating with the rise of subsequent antibody-mediated antiviral functions. IMPORTANCE The development of an efficacious HIV-1 vaccine remains a global priority to prevent new cases of HIV-1 infection. Of the six HIV-1 efficacy trials to date, only one has demonstrated partial efficacy, and immune correlate analysis of that trial revealed a role for binding antibodies and antibody Fc-mediated effector functions. New HIV-1 envelope immunogens are being engineered to selectively expose the most vulnerable and conserved sites on the HIV-1 envelope, with the goal of eliciting antiviral antibodies. Evaluation of the humoral responses elicited by these novel immunogen designs in nonhuman primates is critical for understanding how to improve upon immunogen design to inform further testing in human clinical trials. Our results demonstrate that structural modifications of Env that aim to mimic the CD4-bound conformation can result in earlier antibody elicitation, altered epitope specificity, and increased antiviral function postimmunization.

scholarly journals Polyclonal antibody responses to HIV Env immunogens resolved using cryoEM

2021 ◽  
Author(s):  
Aleksandar Antanasijevic ◽  
Leigh M. Sewall ◽  
Christopher A. Cottrell ◽  
Diane G. Carnathan ◽  
Luis E. Jimenez ◽  
...  
Keyword(s):  
High Resolution ◽  
Polyclonal Antibody ◽  
Vaccine Development ◽  
Mutational Analysis ◽  
Rhesus Macaques ◽  
Antibody Responses ◽  
Binding Modes ◽  
High Resolution Mapping ◽  
High Resolution Analysis ◽  
Resolution Mapping

In BriefHerein, we evaluated the immunogenicity of several BG505 SOSIP-based HIV Env immunogens in the rhesus macaque animal model using a combination of serology and biophysical approaches. We applied electron cryo-microscopy for high-resolution mapping of elicited polyclonal antibody responses, which provided detailed insights into the binding modes of the most common classes of antibodies elicited by BG505 SOSIP immunogens as well as the critical differences in immunogenicity that can occur as a consequence of engineered stabilizing mutations and partial glycan occupancy at different sites.SummaryEngineered ectodomain trimer immunogens based on BG505 envelope glycoprotein are widely utilized as components of HIV vaccine development platforms. In this study, we used rhesus macaques to evaluate the immunogenicity of several stabilized BG505 SOSIP constructs both as free trimers and presented on a nanoparticle. We applied a cryoEM-based method for high-resolution mapping of polyclonal antibody responses elicited in immunized animals (cryoEMPEM). Mutational analysis coupled with neutralization assays were used to probe the neutralization potential at each epitope. We demonstrate that cryoEMPEM data can be used for rapid, high-resolution analysis of polyclonal antibody responses without the need for monoclonal antibody isolation. This approach allowed to resolve structurally distinct classes of antibodies that bind overlapping sites. In addition to comprehensive mapping of commonly targeted neutralizing and non-neutralizing epitopes in BG505 SOSIP immunogens, our analysis revealed that epitopes comprising engineered stabilizing mutations and of partially occupied glycosylation sites can be immunogenic.Graphical abstract

scholarly journals Optimization of Flow-Cytometry Based Assay for Measuring Neutralizing Antibody Responses against Each of the Four Dengue Virus Serotypes

Vaccines ◽  
2021 ◽  
Vol 9 (11) ◽  
pp. 1339
Author(s):  
Pragati Sharma ◽  
Kaustuv Nayak ◽  
Elluri Seetharami Reddy ◽  
Humaira Farooqi ◽  
Kaja Murali-Krishna ◽  
...  
Keyword(s):  
Public Health ◽  
Flow Cytometry ◽  
Dengue Virus ◽  
Vaccine Development ◽  
Neutralizing Antibody ◽  
Neutralization Assay ◽  
Antibody Responses ◽  
Health Concern ◽  
Dengue Virus Serotypes ◽  
Focus Reduction

Dengue is an important public health problem worldwide, with India contributing nearly a third of global dengue disease burden. The measurement of neutralizing antibody responses is critical for understanding dengue pathophysiology, vaccine development and evaluation. Historically, dengue virus neutralization titers were measured using plaque reduction neutralization tests (PRNTs), which were later adapted to focus reduction neutralization tests (FRNTs). Given the slow and laborious nature of both these assays, there has been interest in adapting a high-throughput flow cytometry based neutralization assay. However, flow cytometry based assays typically underestimate neutralization titers, and in situations where the titers are low they can even fail to detect neutralization activity. In this study, by evaluating graded numbers of input Vero cell numbers and viral inoculum, we optimized the flow cytometry based neutralization assay in such a way that it is sensitive and scores titers that are in concordance with focus reduction neutralization tests for each of the four dengue virus serotypes (p < 0.0001). Given that dengue is a global public health concern, and several research groups are making efforts to understand its pathophysiology and accelerate vaccine development and evaluation both in India and worldwide, our findings have timely significance for facilitating these efforts.

scholarly journals Multistage Multiantigen Heterologous Prime Boost Vaccine forPlasmodium knowlesi Malaria Provides Partial Protection in Rhesus Macaques

2001 ◽  
Vol 69 (9) ◽  
pp. 5565-5572 ◽  
Author(s):  
William O. Rogers ◽  
J. Kevin Baird ◽  
Anita Kumar ◽  
John A. Tine ◽  
Walter Weiss ◽  
...  
Keyword(s):  
Dna Vaccine ◽  
Vaccine Development ◽  
Plasmodium Knowlesi ◽  
Rhesus Macaques ◽  
Surface Protein ◽  
Merozoite Surface Protein ◽  
Antibody Responses ◽  
Primate Model ◽  
Boost Vaccine

ABSTRACT A nonhuman primate model for malaria vaccine development allowing reliable, stringent sporozoite challenge and evaluation of both cellular and antibody responses is needed. We therefore constructed a multicomponent, multistage DNA vaccine for the simian malaria species Plasmodium knowlesi including two preerythrocytic-stage antigens, the circumsporozoite protein (PkCSP) and sporozoite surface protein 2 (PkSSP2), and two blood stage antigens, apical merozoite antigen 1 (PkAMA1) and merozoite surface protein 1 (PkMSP1p42), as well as recombinant canarypox viruses encoding the four antigens (ALVAC-4). The DNA vaccine plasmids expressed the corresponding antigens in vitro and induced antiparasite antibodies in mice. Groups of four rhesus monkeys received three doses of a mixture of the four DNA vaccine plasmids and a plasmid encoding rhesus granulocyte-monocyte colony-stimulating factor, followed by boosting with a single dose of ALVAC-4. Three groups received the priming DNA doses by different routes, either by intramuscular needle injection, by intramuscular injection with a needleless injection device, the Biojector, or by a combination of intramuscular and intradermal routes by Biojector. Animals immunized by any route developed antibody responses against sporozoites and infected erythrocytes and against a recombinant PkCSP protein, as well as gamma interferon-secreting T-cell responses against peptides from PkCSP. Following challenge with 100 P. knowlesi sporozoites, 1 of 12 experimental monkeys was completely protected and the mean parasitemia in the remaining monkeys was significantly lower than that in 4 control monkeys. This model will be important in preclinical vaccine development.

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