Dendritic cell (DC)-based immunotherapy has yielded encouraging evidence of providing clinical benefit for the treatment of a broad range of malignancies. Several strategies are being developed to isolate autologous DC, and load them with antigen or peptides ex vivo. Advances in the understanding of immune mechanisms have, in addition to efficient antigen loading, highlighted the importance of the activation and maturation state of DC used for vaccination. Whereas immature DC are more effective in the uptake and processing of antigen, activated/mature DC lose this capacity, and efficiently present antigen to naive T lymphocytes in the context of MHC molecules. Mature DC have been found to be potent antigen presenting cells (APC) to induce primary T lymphocyte responses, overcoming peripheral T cell tolerance and enhance anti-tumor immunity. There are not standard efficient and cost effective methods for combining antigen loading with DC activation and maturation. We propose to develop a novel potent and cost effective approach utilizing proprietary attenuated strains of the intracellular bacterium Listeria monocytogenes (Listeria), engineered to express defined tumor antigens. Listeria is rapidly phagocytosed by DC and transported into the phagolysosomal compartment. This encounter results in the phenotypic maturation of DC and subsequent secretion of a broad profile of immunostimulatory cytokines, including IFN-gamma, IL-12, and TNF-alpha. We have demonstrated that infection of immature DC with recombinant Listeria results in rapid DC activation/maturation, together with MHC class I-restricted presentation of an encoded heterologous antigen. We have also engineered Listeria to be exquisitely sensitive to inactivation by psoralens, a group of compounds that form irreversible cross-links in the genomes of bacteria after illumination with ultraviolet A (UVA) light, so that the applicants are non-viable. The psoralen S-59 is one of a number of Cerus-proprietary compounds known as Helinx. Helinx is approved for sale in the EU as part of the INTERCEPT pathogen inactivation system for platelets. This technology will enable us to generate a safe vaccine for malignant colon cancer based on DC infected with Listeria-CEA vaccines that are genetically inactivated, yet retain the capacity to efficiently program CEA presentation as well as initiating the activation/maturation of the infected DC. The inability of the Listeria vaccine strain to propagate and cause disease ensures the safety of this proprietary immunotherapeutic platform. We believe that this represents a major technological breakthrough for the Listeria vaccine platform that will dramatically facilitate its advancement into clinical trials through eventual approval by regulatory agencies. We propose to generate S-59/UVA inactivated CEA-expressing attenuated Listeria strains. Listeria-CEA vaccines will be used to identify optimal conditions for antigen loading and inducing activation/maturation of human DC.
