Retargeting of oncolytic adenoviruses by bispecific adapter molecules reduces viral liver load and improves therapy by triggering antigen-specific CD8 T-cell responses

Polysialic acid (PolySia) represents an excellent tumor-selective target for oncolytic adenovirus-based therapies since it is selectively expressed on several malignant tumors. In this study, we used a strategy to retarget oncolytic Adenoviruses (OAV) to PolySia-expressing tumors using the bispecific adapter protein CARsc-pSia consisting of the soluble CAR ectodomain and a PolySia-binding scFv domain. By infection experiments in cells of defined PolySia status using CARsc-pSia coated OAV particles, we could show that viral cell entry occurred in a highly PolySia-selective manner. Furthermore, only CARsc-pSia treatment allowed efficient adenoviral infection of normally refractory PolySia-expressing human tumor cells. After systemic administration we could show improved adenoviral infection of s.c. grown PolySia-expressing human tumor xenografts. Masking of the fiberknob by the molecular adapter led to significant reduction of the natural tropism since we could observe a significantly reduced adenoviral liver load and hepatotoxicity in mice. When we investigated the involvement of antitumoral immune responses, therapeutic investigations were therefore performed in an orthotopic animal model in both immunocompetent C67/Bl6 and immunodeficient nude mice. In these models, only CARsc-pSia-complexing of viruses allowed for significant transduction. Interestingly, improved transduction led to improved local tumor regression and prolonged survival in immunocompetent mice but not in T-cell deficient nude mice. Consistently, large infiltrations of CD45-pos. cells in virus-affected, lytic areas were only detectable in mice which received retargeted OAV. Most importantly we could show that retargeted OAV induced an epitope-specific immune response against Gsta-2, a mutated tumor-specific antigen that we could identify by exome sequencing, epitope prediction and in vivo validation. In conclusion, our results show that specific retargeting of OAV represents a promising hepatotoxocity-reduced viroimmuntherapeutic strategy of clinically relevant tumor entities.