425. Translation of Enhanced Dendritic Cell Vaccines Using a CID-Inducible CD40 Receptor

Top of pageAbstract Dendritic cell (DC)-based vaccination is a promising strategy to induce cancer immunity. Nevertheless, clinical applications have been almost universally disappointing. One potential limitation of current strategies is the use of maturation cocktails (MC) that include PGE2, which upregulate chemotaxis to lymph nodes, but may inhibit the ultimate activation state of the resulting DC product. Herein, we describe a novel DC-based vaccine that circumvents the use of standard MC and instead utilizes a genetically engineered inducible CD40 (iCD40) receptor 1. Our platform contains human monocyte-derived DCs (MoDCs) transduced with adenovector Ad5/f35-ihCD40 (iCD40-MoDC) and incubated for 2 days in the presence of monophosphoryl lipid A (MPL) and dimerizing drug, AP20187. Here we show that MoDC transduction with Ad5/f35-iCD40 effectively enhances co- stimulatory and migratory functions, including upregulation of IL-6, activation marker CD83, chemokine receptor CCR7 and augmentation of chemotaxis to CCL19/MIP3|[beta]|. Furthermore, the migratory abilities of iCD40-MoDCs were comparable to MoDCs matured with standard maturation cocktail (i.e. TNF-|[alpha]|, IL-1|[beta]|, IL-6 and PGE2). Unlike murine bone marrow-derived DCs expressing iCD40, IL-12 production by human MoDCs required additional TLR-4-ligand agonists, such as LPS or clinical analog MPL, which led to high (ng) levels of IL-12p70 in ihCD40-MoDCs, but not MC-stimulated or control MoDCs. Moreover, iCD40 and TLR4 agonists additively enhanced activation marker expression. We further show that iCD40 and MPL enhanced the immunostimulatory functions of MoDC, such as improving the stimulation and tumor-specific killing activity of MAGE-3 tumor antigen-specific CTL (shown by IFN-|[gamma]| ELISPOT and 51Cr-release assays) and facilitated Th1 polarization of na|[iuml]|ve CD4+ T-helper cells in vitro. Moreover, we have developed a novel in vivo systemof tracking human DCs transduced with adenovirus expressing red-shifted luciferase. MPL-treated iCD40-MoDCs readily migrated to the lymph nodes of SCID mice and were detectable for over 1 week after inoculation. In addition, we appraised the systemic toxicity of the iCD40-DC vaccine in na|[iuml]|ve C57BL/6 male mice. Mouse DCs were pulsed with TRP2 melanoma antigen, transduced with Ad- iCD40 or control Ad-GFP and injected 3 times bi-weekly. Mice were monitored for 3 months and routine CBC and serum chemistry (for liver and kidney functions) was tested regularly. We detected no significant toxicity of the iCD40-DCs vaccine and highly significant long-lived elevation of tumor antigen specific CTLs. This data suggest the potency and safety of this immunotherapeutic approach that combines non-inflammatory TLR stimulation with synthetic activation of iCD40 in human MoDCs. Unlike standard DC maturation approaches, this combination of reagents leads to high IL-12p70 levels, enhanced chemotactic response to lymph node-derived chemokines and augmented tumor antigen-specific CTL and Th1 responses. Inducible CD40-enhanced DCs represent a promising new strategy for cancer immunotherapy.