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Abstract

Cancer remains a major public health challenge and entities such as glioblastoma (GBM) and pancreatic ductal adenocarcinoma (PDAC) are particularly difficult to treat with the currently available therapeutics: Immune privileged locations, a challenging tumor microenvironment and often-occurring treatment resistances underline the need for novel therapeutic strategies. Radiotherapy is a cornerstone of cancer management and has benefited immensely from technological advances. Nevertheless, dose-limiting toxicities and relapse due to treatment resistance are frequent. A promising approach to increase treatment efficacy is the combination with immunotherapies. Oncolytic viruses, such as the vaccine strain of measles virus (MeV), are one such immunotherapeutic approach: MeV has a natural cancer tropism, lyses tumor cells and induces an anti-tumor immune response. However, MeV monotherapies have shown limited therapeutic efficacy in solid tumors. Preclinical data indicates that combining MeV with radiotherapy can produce synergistic effects and a favorable (innate) immune activation, although interferon (IFN)-mediated antiviral responses can also restrict MeV replication. I thus hypothesized that the addition of a second oncolytic vector, parvovirus, known to suppress the IFN response, could further enhance treatment efficacy. Through RNA sequencing, I characterized the combination of MeV and radiotherapy and observed distinct immune induction patterns in the combination. I then identified candidate cell lines from a panel of GBM and PDAC cell lines with the desired intact IFN signaling capacity, showing attenuation of MeV replication. In these models, I assessed cytotoxicity, synergy and the potential mechanisms of dual (PV + MeV) and triple (radiation + PV + MeV) therapy. Dual virotherapy produced additive cytotoxic effects alongside PV-mediated IFN suppression. While MeV replication was unaffected, its transgene expression was markedly reduced during co-infection. Triple radiovirotherapy demonstrated enhanced cytotoxicity and synergy in GBM cells at specific dose combinations, accompanied by modest IFN dampening and an increase in the immunogenic cell death (ICD) marker calreticulin. I additionally employed a heterotypic spheroid model, where the MeV-mediated IFN response was reduced when combined with certain PV doses, but cytotoxicity was not enhanced. On the contrary, triple combinations showed an antagonistic pattern. Finally, I generated and characterized murine cell lines expressing the MeV entry receptor for future in vivo evaluation. Overall, I performed a comprehensive analysis of (triple) radiovirotherapy. The variable treatment efficacy reflects the complexity of analyzing advanced combination approaches in vitro. Nevertheless, synergistic combinations were identified, suggesting a potential therapeutic benefit for selected cancer patients suffering from refractory cancers such as PDAC and GBM.