Protective immune responses against West Nile virus are primed by distinct complement activation pathways

West Nile virus (WNV) causes a severe infection of the central nervous system in several vertebrate animals including humans. Prior studies have shown that complement plays a critical role in controlling WNV infection in complement (C) 3−/− and complement receptor 1/2−/− mice. Here, we dissect the contributions of the individual complement activation pathways to the protection from WNV disease. Genetic deficiencies in C1q, C4, factor B, or factor D all resulted in increased mortality in mice, suggesting that all activation pathways function together to limit WNV spread. In the absence of alternative pathway complement activation, WNV disseminated into the central nervous system at earlier times and was associated with reduced CD8+ T cell responses yet near normal anti-WNV antibody profiles. Animals lacking the classical and lectin pathways had deficits in both B and T cell responses to WNV. Finally, and somewhat surprisingly, C1q was required for productive infection in the spleen but not for development of adaptive immune responses after WNV infection. Our results suggest that individual pathways of complement activation control WNV infection by priming adaptive immune responses through distinct mechanisms

Enhancement of anti-DIII antibodies by the C3d derivative P28 results in lower viral titers and augments protection in mice

Antibodies generated against West Nile virus (WNV) during infection are essential for controlling dissemination. Recent studies have demonstrated that epitopes in all three domains of the flavivirus envelope protein (E) are targets for neutralizing antibodies, with determinants in domain III (DIII) eliciting antibodies with strong inhibitory properties. In order to increase the magnitude and quality of the antibody response against the WNV E protein, DNA vaccines with derivatives of the WNV E gene (full length E, truncated E, or DIII region, some in the context of the pre-membrane [prM] gene) were conjugated to the molecular adjuvant P28. The P28 region of the complement protein C3d is the minimum CR2-binding domain necessary for the adjuvant activity of C3d. Delivery of DNA-based vaccines by gene gun and intramuscular routes stimulated production of IgG antibodies against the WNV DIII region of the E protein. With the exception of the vaccine expressing prM/E given intramuscularly, only mice that received DNA vaccines by gene gun produced protective neutralizing antibody titers (FRNT80 titer >1/40). Correspondingly, mice vaccinated by the gene gun route were protected to a greater level from lethal WNV challenge. In general, mice vaccinated with P28-adjuvated vaccines produced higher IgG titers than mice vaccinated with non-adjuvanted vaccines

Lethal Antibody Enhancement of Dengue Disease in Mice Is Prevented by Fc Modification

Immunity to one of the four dengue virus (DV) serotypes can increase disease severity in humans upon subsequent infection with another DV serotype. Serotype cross-reactive antibodies facilitate DV infection of myeloid cells in vitro by promoting virus entry via Fcγ receptors (FcγR), a process known as antibody-dependent enhancement (ADE). However, despite decades of investigation, no in vivo model for antibody enhancement of dengue disease severity has been described. Analogous to human infants who receive anti-DV antibodies by transplacental transfer and develop severe dengue disease during primary infection, we show here that passive administration of anti-DV antibodies is sufficient to enhance DV infection and disease in mice using both mouse-adapted and clinical DV isolates. Antibody-enhanced lethal disease featured many of the hallmarks of severe dengue disease in humans, including thrombocytopenia, vascular leakage, elevated serum cytokine levels, and increased systemic viral burden in serum and tissue phagocytes. Passive transfer of a high dose of serotype-specific antibodies eliminated viremia, but lower doses of these antibodies or cross-reactive polyclonal or monoclonal antibodies all enhanced disease in vivo even when antibody levels were neutralizing in vitro. In contrast, a genetically engineered antibody variant (E60-N297Q) that cannot bind FcγR exhibited prophylactic and therapeutic efficacy against ADE-induced lethal challenge. These observations provide insight into the pathogenesis of antibody-enhanced dengue disease and identify a novel strategy for the design of therapeutic antibodies against dengue

Complement Protein C1q Inhibits Antibody-Dependent Enhancement of Flavivirus Infection in an IgG Subclass-Specific Manner

SummarySevere dengue virus infection can occur in humans with pre-existing antibodies against the virus. This observation led to the hypothesis that a subneutralizing antibody level in vivo can increase viral burden and cause more severe disease. Indeed, antibody-dependent enhancement of infection (ADE) in vitro has been described for multiple viruses, including the flaviviruses dengue virus and West Nile virus. Here, we demonstrate that the complement component C1q restricts ADE by anti-flavivirus IgG antibodies in an IgG subclass-specific manner in cell culture and in mice. IgG subclasses that avidly bind C1q induced minimal ADE in the presence of C1q. These findings add a layer of complexity for the analysis of humoral immunity and flavivirus infection

Presentation of exogenous whole inactivated simian immunodeficiency virus by mature dendritic cells induces CD4\u3csup\u3e+\u3c/sup\u3e and CD8\u3csup\u3e+\u3c/sup\u3e T-cell responses

Interactions between HIV-1 and dendritic cells (DCs) play an important role in the initial establishment and spread of infection and development of antiviral immunity. We used chemically inactivated aldrithiol-2 (AT-2) simian immunodeficiency virus (SIV) with functional envelope glycoproteins to study virus interactions with DCs and developed an in vitro system to evaluate the quality of SIV antigen (Ag) presentation by DCs to T cells. AT-2 SIV interacts authentically with T cells and DCs and thus allows assessment of natural SIV-specific responses. CD4+ and CD8+ T cells from blood or lymph nodes of SIV-infected macaques released interferon-γ (IFNγ) and proliferated in response to a variety of AT-2 SIV isolates. Responses did not vary significantly as a function of the quantitative envelope glycoprotein content of the virions. Presentation of Ags derived from AT-2 SIV by DCs was more potent than presentation by comparably Ag-loaded monocytes. Interestingly, SIV-pulsed mature DCs stimulated both CD4+ and CD8+ T-cell responses, whereas immature DCs primarily stimulated CD4+ T cells. Further studies using AT-2 inactivated virus may help to define better the details of the virus-DC interactions critical for infection versus induction of antiviral immune responses

Lethal Antibody Enhancement of Dengue Disease in Mice Is Prevented by Fc Modification

Immunity to one of the four dengue virus (DV) serotypes can increase disease severity in humans upon subsequent infection with another DV serotype. Serotype cross-reactive antibodies facilitate DV infection of myeloid cells in vitro by promoting virus entry via Fcγ receptors (FcγR), a process known as antibody-dependent enhancement (ADE). However, despite decades of investigation, no in vivo model for antibody enhancement of dengue disease severity has been described. Analogous to human infants who receive anti-DV antibodies by transplacental transfer and develop severe dengue disease during primary infection, we show here that passive administration of anti-DV antibodies is sufficient to enhance DV infection and disease in mice using both mouse-adapted and clinical DV isolates. Antibody-enhanced lethal disease featured many of the hallmarks of severe dengue disease in humans, including thrombocytopenia, vascular leakage, elevated serum cytokine levels, and increased systemic viral burden in serum and tissue phagocytes. Passive transfer of a high dose of serotype-specific antibodies eliminated viremia, but lower doses of these antibodies or cross-reactive polyclonal or monoclonal antibodies all enhanced disease in vivo even when antibody levels were neutralizing in vitro. In contrast, a genetically engineered antibody variant (E60-N297Q) that cannot bind FcγR exhibited prophylactic and therapeutic efficacy against ADE-induced lethal challenge. These observations provide insight into the pathogenesis of antibody-enhanced dengue disease and identify a novel strategy for the design of therapeutic antibodies against dengue