Effective immunotherapy of pre-existing tumor in a stringent rat mammary tumor model by targeting the rat neu, “self”, tumor-associated antigen using an alphavirus replicon based immunotherapy with in-vivo DC tropism

6042 Introduction: The majority of human tumor-associated antigens (TAAs) are either minimally altered or malexpressed “self” antigens. Effective anti-tumor immunity must overcome intrinsic tolerance to TAAs and barriers derived from host:tumor interactions to mount a sufficiently potent anti-tumor immune response for tumor cell destruction. The immune system dendritic cell (DC), the most potent antigen-presenting cell, is a logical target for strategies designed to elicit robust immune responses. However, adoptive transfer of ex vivo manipulated DCs must overcome biological and regulatory hurdles. The Venezuelan Equine Encephalitis (VEE)-derived replicon particle (VRP) has in vivo tropism for murine DCs and may be advantageous for eliciting larger magnitude and more efficacious anti-tumor immune responses. Methods: These studies used VRPs encoding the reporter protein dsRed (VRP-dsRed), rat neu target antigen (VRP-rNeu) or an irrelevant antigen influenza hemagglutinin (VRP-HA) and a stringent animal model of a non-mutated “self” TAA in an effort to recapitulate the human clinical state. We used Fisher 344 rats and the syngeneic moderately immunogenic 13762 mammary tumor line, with moderate expression of rat neu. Immunizations, SC or IM administration, q3 weeks x 3 were used to establish anti-tumor immunity for challenge and mechanistic studies. For treatment studies, tumor was established 48 hr prior to initiation of immunizations q10 d. VRPs encoding dsRed were administered SC or in the footpad to evaluate the extent of in-vivo tropism for rat DCs, extending the report of in-vivo murine DC tropism (McDonald & Johnston, J Virol 2000). Data: Administration of VRP-dsRed revealed transfected DCs trafficking to draining lymph nodes DCs within 12 hr and persisting in excess of 84 hr. Immunization with VRP-rNeu resulted in significant antigen-specific, tumor challenge protection (p=0.002), the induction of immunologic memory, low titer antigen-specific antibody responses that were exclusively IgG2 supporting the induction of Th1 biased immune responses, and the ability for cellular transfer immunity by Winn assay. This immunotherapeutic strategy also demonstrated efficacy against pre-established tumor with delayed tumor outgrowth and cure of 20% of animals (p=0.022) by targeting a single TAA. Conclusions: Our findings demonstrate that the VRP vector system has in-vivo tropism for rat DCs that both traffic and remain in secondary lymphoid tissues appropriately. This VRP based, single TAA antigen-specific, immunotherapeutic strategy overcomes tolerance to the “self” antigen neu and elicits very active Th1 anti-tumor immunity. These results support both further evaluation and rapid translation into clinical evaluations of this potent DC tropic vector-based immunotherapy.