We propose to develop a novel approach to vaccination based on the long-standing collaborations between the Steinman, Nussenzweig and Ravetch laboratories to directly target dendrific cells with candidate antigens and provide maturation stimuli to insure effective activation of T cells and B cells. In this subproject, we will focus on the role of the IgG Fc domain of the targeting vectors and the IgG Fc receptors expressed on immature dendritic cells and build on our previous observafions that signaling though the activation versions of these receptors will provide a maturation signal for DCs. We will also investigate the role of FcyR activation on effector cells by the IgG Fc domain of broadly neutralizing antibodies anti-HIV anfibodies to their in vivo efficacy at reducing viral load. We have developed both mouse and human IgG Fc variants that display preferential binding to the specific activation receptors expressed on DCs, macrophages, NK cells, mast cells or neutrophils, thereby overriding the normal inhibitory constraints provided by the inhibitory FcR. We will test the hypothesis that modification ofthe Fc domain of DC targefing vectors to preferentially engage activation FcRs expressed on DCs will simultaneously deliver both antigen and a maturation signal to immature DCs, thereby accomplishing in a single step the dual goals of effecfive DC mediated vaccination.
Aim 1 will characterize the in vitro properties of Fc modified targeting vectors on antigen presentation, DC maturafion and Tcell sfimulation for a model antigen, OVA, and forthe HIV antigens described by Steinman (gag) and Nussenzweig (gp140) in their sub-projects.
Aim 2 will extend these studies to in vivo systems to determine if an Fc modified DC targeting vector introduced into a mouse will result in efficient antigen presentation and DC maturation to result in sustained T cell activation and anfibody responses. Proof of concept studies will be performed using Fc modified murine DC targefing vectors and extended to modified human Fc's coupled to anti-human DEC. These later constructs will be tested in an FcR humanized mouse we have generated, that will be modified to express human DEC.
Aim 3 will focus on the role ofthe Fc domain in HIV neutralizing anfibodies, like b12, by generafing Fc modificafions that preferentially engage activation FcRs on effector cells such as macrophages. These modified b12 anfibodies will be tested in vivo in the NOG mouse reconstituted with human hematopoeific progenitors, provide to us as part ofthe user group consortium in place at The Rockefeller University. These reconstituted NOGs will be infected with HIV and passively treated with the modified neutralizing anfibodies.
The generafion of effective anti-HIV immunity remains a formidable challenge to global health. Through the studies proposed in this subproject we will investigate mechanisms to enhance immunogenicity of potenfial target anfigens to generate neutralizing anfibodies and to engineer neutralizing anfibodies to augment their in vivo activity.
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|DiLillo, David J; Tan, Gene S; Palese, Peter et al. (2014) Broadly neutralizing hemagglutinin stalk-specific antibodies require FcÎ³R interactions for protection against influenza virus in vivo. Nat Med 20:143-51|
|Mollah, Shamim A; Dobrin, Joseph S; Feder, Rachel E et al. (2014) Flt3L dependence helps define an uncharacterized subset of murine cutaneous dendritic cells. J Invest Dermatol 134:1265-75|
|Gruell, Henning; Bournazos, Stylianos; Ravetch, Jeffrey V et al. (2013) Antibody and antiretroviral preexposure prophylaxis prevent cervicovaginal HIV-1 infection in a transgenic mouse model. J Virol 87:8535-44|
|Klein, Florian; Mouquet, Hugo; Dosenovic, Pia et al. (2013) Antibodies in HIV-1 vaccine development and therapy. Science 341:1199-204|
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