Evaluation of protection induced by a dengue virus serotype 2 envelope domain III protein scaffold/DNA vaccine in non-human primates

AbstractWe describe the preclinical development of a dengue virus vaccine targeting the dengue virus serotype 2 (DENV2) envelope domain III (EDIII). This study provides proof-of-principle that a dengue EDIII protein scaffold/DNA vaccine can protect against dengue challenge. The dengue vaccine (EDIII-E2) is composed of both a protein particle and a DNA expression plasmid delivered simultaneously via intramuscular injection (protein) and gene gun (DNA) into rhesus macaques. The protein component can contain a maximum of 60 copies of EDIII presented on a multimeric scaffold of Geobacillus stearothermophilus E2 proteins. The DNA component is composed of the EDIII portion of the envelope gene cloned into an expression plasmid. The EDIII-E2 vaccine elicited robust antibody responses to DENV2, with neutralizing antibody responses detectable following the first boost and reaching titers of greater than 1:100,000 following the second and final boost. Vaccinated and naïve groups of macaques were challenged with DENV2. All vaccinated macaques were protected from detectable viremia by infectious assay, while naïve animals had detectable viremia for 2–7 days post-challenge. All naïve macaques had detectable viral RNA from day 2–10 post-challenge. In the EDIII-E2 group, three macaques were negative for viral RNA and three were found to have detectable viral RNA post challenge. Viremia onset was delayed and the duration was shortened relative to naïve controls. The presence of viral RNA post-challenge corresponded to a 10–30-fold boost in neutralization titers 28 days post challenge, whereas no boost was observed in the fully protected animals. Based on these results, we determine that pre-challenge 50% neutralization titers of >1:6000 correlated with sterilizing protection against DENV2 challenge in EDIII-E2 vaccinated macaques. Identification of the critical correlate of protection for the EDIII-E2 platform in the robust non-human primate model lays the groundwork for further development of a tetravalent EDIII-E2 dengue vaccine

Prime-boost immunization of rabbits with HIV-1 gp120 elicits potent neutralization activity against a primary viral isolate

<div><p>Development of a vaccine for HIV-1 requires a detailed understanding of the neutralizing antibody responses that can be experimentally elicited to difficult-to-neutralize primary isolates. Rabbits were immunized with the gp120 subunit of HIV-1 JR-CSF envelope (Env) using a DNA-prime protein-boost regimen. We analyzed five sera that showed potent autologous neutralizing activity (IC50s at ∼10³ to 10⁴ serum dilution) against pseudoviruses containing Env from the primary isolate JR-CSF but not from the related isolate JR-FL. Pseudoviruses were created by exchanging each variable and constant domain of JR-CSF gp120 with that of JR-FL or with mutations in putative N-glycosylation sites. The sera contained different neutralizing activities dependent on C3 and V5, C3 and V4, or V4 regions located on the glycan-rich outer domain of gp120. All sera showed enhanced neutralizing activity toward an Env variant that lacked a glycosylation site in V4. The JR-CSF gp120 epitopes recognized by the sera are generally distinct from those of several well characterized mAbs (targeting conserved sites on Env) or other type-specific responses (targeting V1, V2, or V3 variable regions). The activity of one serum requires specific glycans that are also important for 2G12 neutralization and this serum blocked the binding of 2G12 to gp120. Our findings show that different fine specificities can achieve potent neutralization of HIV-1, yet this strong activity does not result in improved breadth.</p> </div

Evaluation of Heterologous Vaginal SHIV SF162p4 Infection Following Vaccination with a Polyvalent Clade B Virus-Like Particle Vaccine

AbstractThe vast diversity of HIV-1 infections has greatly impeded the development of a successful HIV-1/AIDS vaccine. Previous vaccine work has demonstrated limited levels of protection against SHIV/SIV infection, but protection was observed only when the challenge virus was directly matched to the vaccine strain. As it is likely impossible to directly match the vaccine strain to all infecting strains in nature, it is necessary to develop an HIV-1 vaccine that can protect against a heterologous viral challenge. In this study we investigated the ability of polyvalent and consensus vaccines to protect against a heterologous clade B challenge. Rhesus macaques were vaccinated with ConB or PolyB virus-like particle vaccines. All vaccines were highly immunogenic with high titers of antibody found in all vaccinated groups against SIV Gag. Antibody responses were also observed against a diverse panel of clade B envelopes. Following vaccination nonhuman primates (NHPs) were challenged via the vaginal route with SHIVSF162p4. The PolyB vaccine induced a 66.7% reduction in the rate of infection as well as causing a two log reduction in viral burden if infection was not blocked. ConB vaccination had no effect on either the infection rate or viral burden. These results indicate that a polyvalent clade-matched vaccine is better able to protect against a heterologous challenge as compared to a consensus vaccine

Evaluation of Heterologous Vaginal SHIV SF162p4 Infection Following Vaccination with a Polyvalent Clade B Virus-Like Particle Vaccine

The vast diversity of HIV-1 infections has greatly impeded the development of a successful HIV-1/AIDS vaccine. Previous vaccine work has demonstrated limited levels of protection against SHIV/SIV infection, but protection was observed only when the challenge virus was directly matched to the vaccine strain. As it is likely impossible to directly match the vaccine strain to all infecting strains in nature, it is necessary to develop an HIV-1 vaccine that can protect against a heterologous viral challenge. In this study we investigated the ability of polyvalent and consensus vaccines to protect against a heterologous clade B challenge. Rhesus macaques were vaccinated with ConB or PolyB virus-like particle vaccines. All vaccines were highly immunogenic with high titers of antibody found in all vaccinated groups against SIV Gag. Antibody responses were also observed against a diverse panel of clade B envelopes. Following vaccination nonhuman primates (NHPs) were challenged via the vaginal route with SHIV(SF162p4). The PolyB vaccine induced a 66.7% reduction in the rate of infection as well as causing a two log reduction in viral burden if infection was not blocked. ConB vaccination had no effect on either the infection rate or viral burden. These results indicate that a polyvalent clade-matched vaccine is better able to protect against a heterologous challenge as compared to a consensus vaccine

C1q binding to dengue virus decreases levels of infection and inflammatory molecules transcription in THP-1 cells

Submitted by Kamylla Nascimento ([email protected]) on 2017-12-11T12:51:12Z No. of bitstreams: 1 art. C1q binding - douradinha.pdf: 356297 bytes, checksum: 40c2f62c2a58ba25cf310346ca16f238 (MD5)Approved for entry into archive by Kamylla Nascimento ([email protected]) on 2017-12-11T13:34:26Z (GMT) No. of bitstreams: 1 art. C1q binding - douradinha.pdf: 356297 bytes, checksum: 40c2f62c2a58ba25cf310346ca16f238 (MD5)Made available in DSpace on 2017-12-11T13:34:26Z (GMT). No. of bitstreams: 1 art. C1q binding - douradinha.pdf: 356297 bytes, checksum: 40c2f62c2a58ba25cf310346ca16f238 (MD5) Previous issue date: 2014Fondazione Ri.MED. Palermo, Italy / University of Pittsburgh. Center for Vaccine Research. Pittsburgh, USA.University of Pittsburgh. Center for Vaccine Research. Pittsburgh, USA.University of Pittsburgh. Center for Vaccine Research. Pittsburgh, USA / Fundação Oswaldo Cruz. Instituto Aggeu Magalhães. Laboratório de Virologia e Terapia Experimental. Recife, PE, Brasil.University of Pittsburgh. Center for Vaccine Research. Pittsburgh, USA.Eastern Virginia Medical School. Department of Microbiology and Molecular Cell Biology. Norfolk, Virginia, USA.University of Pittsburgh. Center for Vaccine Research. Pittsburgh, USA / University of Pittsburgh. Graduate School of Public Health. Department of Infectious Diseases and Microbiology. Pittsburgh, USA.University of Pittsburgh. Center for Vaccine Research. Pittsburgh, USA / University of Pittsburgh. School of Medicine. Department of Microbiology and Molecular Genetics. Pittsburgh, USA.University of Pittsburgh. Center for Vaccine Research. Pittsburgh, USA / University of Pittsburgh. Graduate School of Public Health. Department of Infectious Diseases and Microbiology. Pittsburgh, USA.University of Pittsburgh. Center for Vaccine Research. Pittsburgh, USA / Fundação Oswaldo Cruz. Instituto Aggeu Magalhães. Laboratório de Virologia e Terapia Experimental. Recife, PE, Brasil / University of Pittsburgh. Graduate School of Public Health. Department of Infectious Diseases and Microbiology. Pittsburgh, USA.Dengue virus infection elicits a spectrum of clinical presentations ranging from asymptomatic to severe disease. The mechanisms leading to severe dengue are not known, however it has been reported that the complement system is hyper-activated in severe dengue. Screening of complement proteins demonstrated that C1q, a pattern recognition molecule, can bind directly to dengue virus envelope protein and to whole dengue virus serotype 2. Incubation of dengue virus serotype 2 with C1q prior to infection of THP-1 cells led to decreased virus infectivity and modulation of mRNA expression of immunoregulatory molecules suggesting reduced inflammatory responses

Multimeric Scaffolds Displaying the HIV-1 Envelope MPER Induce MPER-Specific Antibodies and Cross-Neutralizing Antibodies when Co-Immunized with gp160 DNA

<div><p>Developing a vaccine that overcomes the diversity of HIV-1 is likely to require a strategy that directs antibody (Ab) responses toward conserved regions of the viral Envelope (Env). However, the generation of neutralizing Abs (NAbs) targeting these regions through vaccination has proven to be difficult. One conserved region of particular interest is the membrane proximal external region (MPER) of Env located within the gp41 ectodomain. In order to direct the immune response to this region, the MPER and gp41 ectodomain were expressed separately as N-terminal fusions to the E2 protein of <i>Geobacillus stearothermophilus</i>. The E2 protein acts as a scaffold by self-assembling into 60-mer particles, displaying up to 60 copies of the fused target on the surface. Rabbits were immunized with E2 particles displaying MPER and/or the gp41 ectodomain in conjunction with DNA encoding full-length gp160. Only vaccines including E2 particles displaying MPER elicited MPER-specific Ab responses. NAbs were elicited after two immunizations that largely targeted the V3 loop. To overcome V3 immunodominance in the DNA component, E2 particles displaying MPER were used in conjunction with gp160 DNA lacking hypervariable regions V2, V3, or combined V1V2V3. All rabbits had HIV binding Ab responses and NAbs following the second vaccination. Using HIV-2/HIV-1 MPER chimeric viruses as targets, NAbs were detected in 12/16 rabbits after three immunizations. Low levels of NAbs specific for Tier 1 and 2 viruses were observed in all groups. This study provides evidence that co-immunizing E2 particles displaying MPER and gp160 DNA can focus Ab responses toward conserved regions of Env.</p></div

Neutralization Activity Against HIV-2/HIV-1 MPER Chimera Viruses elicited in rabbits co-immunized with Env(MPER)-E2 and either SF162 gp160full-length, ΔV3, ΔV2, or ΔV123 DNA.

<p>(A) Neutralization titers against the autologous SF162 pseudovirus, parental HIV-2 virus 7312A wt, and the HIV-2/HIV-1 MPER chimera viruses 7312A C1, C3, and C4 were determined using the TZM-bl assay. Values stated are the IC₅₀ values for individual rabbits in each of the four groups at week 14 following the third vaccination. Pre-bleed samples as well as sera from a Env(V3)-E2+SF162 gp160 DNA vaccinated rabbit were included as controls. Darker colors represent an increase in NAb titer.</p