Christine Trumpfheller

Dendritic cells (DCs) process and present self and foreign antigens to induce tolerance or immunity. In vitro models suggest that induction of immunity is controlled by regulating the presentation of antigen, but little is known about how DCs control antigen presentation in vivo. To examine antigen processing and presentation in vivo, we specifically targeted antigens to two major subsets of DCs by using chimeric monoclonal antibodies. Unlike CD8+ DCs that express the cell surface protein CD205, CD8- DCs, which are positive for the 33D1 antigen, are specialized for presentation on major histocompatibility complex (MHC) class II. This difference in antigen processing is intrinsic to the DC subsets and is associated with increased expression of proteins involved in MHC processing.

Dengue virus is a single-stranded, enveloped RNA virus that productively infects human dendritic cells (DCs) primarily at the immature stage of their differentiation. We now find that all four serotypes of dengue use DC-SIGN (CD209), a C-type lectin, to infect dendritic cells. THP-1 cells become susceptible to dengue infection after transfection of DC-specific ICAM-3 grabbing nonintegrin (DC-SIGN), or its homologue L-SIGN, whereas the infection of dendritic cells is blocked by anti-DC-SIGN antibodies and not by antibodies to other molecules on these cells. Viruses produced by dendritic cells are infectious for DC-SIGN- and L-SIGN-bearing THP-1 cells and other permissive cell lines. Therefore, DC-SIGN may be considered as a new target for designing therapies that block dengue infection.

Relative to several other toll-like receptor (TLR) agonists, we found polyinosinic:polycytidylic acid (poly IC) to be the most effective adjuvant for Th1 CD4(+) T cell responses to a dendritic cell (DC)-targeted HIV gag protein vaccine in mice. To identify mechanisms for adjuvant action in the intact animal and the polyclonal T cell repertoire, we found poly IC to be the most effective inducer of type I interferon (IFN), which was produced by DEC-205(+) DCs, monocytes, and stromal cells. Antibody blocking or deletion of type I IFN receptor showed that IFN was essential for DC maturation and development of CD4(+) immunity. The IFN-AR receptor was directly required for DCs to respond to poly IC. STAT 1 was also essential, in keeping with the type I IFN requirement, but not type II IFN or IL-12 p40. Induction of type I IFN was mda5 dependent, but DCs additionally used TLR3. In bone marrow chimeras, radioresistant and, likely, nonhematopoietic cells were the main source of IFN, but mda5 was required in both marrow-derived and radioresistant host cells for adaptive responses. Therefore, the adjuvant action of poly IC requires a widespread innate type I IFN response that directly links antigen presentation by DCs to adaptive immunity.

CD4(+) Th1 type immunity is implicated in resistance to global infectious diseases. To improve the efficacy of T cell immunity induced by human immunodeficiency virus (HIV) vaccines, we are developing a protein-based approach that directly harnesses the function of dendritic cells (DCs) in intact lymphoid tissues. Vaccine proteins are selectively delivered to DCs by antibodies to DEC-205/CD205, a receptor for antigen presentation. We find that polyriboinosinic:polyribocytidylic acid (poly IC) independently serves as an adjuvant to allow a DC-targeted protein to induce protective CD4(+) T cell responses at a mucosal surface, the airway. After two doses of DEC-targeted, HIV gag p24 along with poly IC, responder CD4(+) T cells have qualitative features that have been correlated with protective function. The T cells simultaneously make IFN-gamma, tumor necrosis factor (TNF)-alpha, and IL-2, and in high amounts for prolonged periods. The T cells also proliferate and continue to secrete IFN-gamma in response to HIV gag p24. The adjuvant role of poly IC requires Toll-like receptor (TLR) 3 and melanoma differentiation-associated gene-5 (MDA5) receptors, but its analog poly IC(12)U requires only TLR3. We suggest that poly IC be tested as an adjuvant with DC-targeted vaccines to induce numerous multifunctional CD4(+) Th1 cells with proliferative capacity.

Improved protein-based vaccines should facilitate the goal of effective vaccines against HIV and other pathogens. With respect to T cells, the efficiency of immunization, or "immunogenicity," is improved by targeting vaccine proteins to maturing dendritic cells (DCs) within mAbs to DC receptors. Here, we compared the capacity of Langerin/CD207, DEC205/CD205, and Clec9A receptors, each expressed on the CD8(+) DC subset in mice, to bring about immunization of microbial-specific T cells from the polyclonal repertoire, using HIV gag-p24 protein as an antigen. α-Langerin mAb targeted splenic CD8(+) DCs selectively in vivo, whereas α-DEC205 and α-Clec9A mAbs targeted additional cell types. When the mAb heavy chains were engineered to express gag-p24, the α-Langerin, α-DEC205, and α-Clec9A fusion mAbs given along with a maturation stimulus induced comparable levels of gag-specific T helper 1 (Th1) and CD8(+) T cells in BALB/c × C57BL/6 F1 mice. These immune T cells were more numerous than targeting the CD8(-) DC subset with α-DCIR2-gag-p24. In an in vivo assay in which gag-primed T cells were used to report the early stages of T-cell responses, α-Langerin, α-DEC205, and α-Clec9A also mediated cross-presentation to primed CD8(+) T cells if, in parallel to antigen uptake, the DCs were stimulated with α-CD40. α-Langerin, α-DEC205, and α-Clec9A targeting greatly enhanced T-cell immunization relative to nonbinding control mAb or nontargeted HIV gag-p24 protein. Therefore, when the appropriate subset of DCs is targeted with a vaccine protein, several different receptors expressed by that subset are able to initiate combined Th1 and CD8(+) immunity.