We hypothesized that the combination of oncolytic virotherapy with immune checkpoint modulators would reduce tumor burden by direct cell lysis and stimulate antitumor immunity. In this study, we have generated attenuated Measles virus (MV) vectors encoding antibodies against CTLA-4 and PD-L1 (MV-aCTLA-4 and MV-aPD-L1). We characterized the vectors in terms of growth kinetics, antibody expression, and cytotoxicity in vitro. Immunotherapeutic effects were assessed in a newly established, fully immunocompetent murine model of malignant melanoma, B16-CD20. Analyses of tumor-infiltrating lymphocytes and restimulation experiments indicated a favorable immune profile after MV-mediated checkpoint modulation. Therapeutic benefits in terms of delayed tumor progression and prolonged median overall survival were observed for animals treated with vectors encoding anti-CTLA-4 and anti-PD-L1, respectively. Combining systemic administration of antibodies with MV treatment also improved therapeutic outcome. In vivo oncolytic efficacy against human tumors was studied in melanoma xenografts. MV-aCTLA-4 and MV-aPD-L1 were equally efficient as parental MV in this model, with high rates of complete tumor remission (> 80%). Furthermore, we could demonstrate lysis of tumor cells and transgene expression in primary tissue from melanoma patients. The current results suggest rapid translation of combining immune checkpoint modulation with oncolytic viruses into clinical application.

Purpose: Immunotherapy with bispecific T-cell engagers has achieved striking success against hematologic malignancies, but efficacy against solid tumors has been limited. We hypothesized that oncolytic measles viruses encoding bispecific T-cell engagers (MV-BiTEs) represent a safe and effective treatment against solid tumors through local BiTE expression, direct tumor cell lysis and in situ tumor vaccination. Experimental Design: To test this hypothesis, we generated MV-BiTEs from the Edmonston B vaccine strain to target two model antigens. Replicative and oncolytic potential were assessed by infection and cell viability assays, respectively. Functionality of virus-derived BiTEs was tested in vitro by complementary binding and cytotoxicity assays. In vivo efficacy of MV-BiTE was investigated using both syngeneic and xenograft mouse models of solid cancers. Results: We verified secretion of functional BiTE antibodies by MV-BiTE-infected cells. Further, we demonstrated therapeutic efficacy of MV-BiTE against established tumors in fully immunocompetent mice. MV-BiTE efficacy was associated with increased intratumoral T-cell infiltration and induction of protective antitumor immunity. In addition, we showed therapeutic efficacy of MV-BiTE in xenograft models of patient-derived primary colorectal carcinoma spheroids with transfer of peripheral blood mononuclear cells. Conclusions: MV-BiTE treatment was effective in two distinct models of solid tumors without signs of toxicity. This provides strong evidence for therapeutic benefits of tumor-targeted BiTE expression by oncolytic MV. Thus, this study represents proof of concept for an effective strategy to treat solid tumors with BiTEs. Clin Cancer Res; 24(9); 2128–37. ©2018 AACR.

Bispecific T cell engagers (BiTEs) are an innovative class of immunotherapeutics that redirect T cells to tumor surface antigens. While efficacious against certain hematological malignancies, limited bioavailability and severe toxicities have so far hampered broader clinical application, especially against solid tumors. Another emerging cancer immunotherapy are oncolytic viruses (OVs) which selectively infect and replicate in malignant cells, thereby mediating tumor vaccination effects. These oncotropic viruses can serve as vectors for tumor-targeted immunomodulation and synergize with other immunotherapies. In this article, we discuss the use of OVs to overcome challenges in BiTE therapy. We review the current state of the field, covering published preclinical studies as well as ongoing clinical investigations. We systematically introduce OV-BiTE vector design and characteristics as well as evidence for immune-stimulating and anti-tumor effects. Moreover, we address additional combination regimens, including CAR T cells and immune checkpoint inhibitors, and further strategies to modulate the tumor microenvironment using OV-BiTEs. The inherent complexity of these novel therapeutics highlights the importance of translational research including correlative studies in early-phase clinical trials. More broadly, OV-BiTEs can serve as a blueprint for diverse OV-based cancer immunotherapies.

Measles virus (MeV) preferentially replicates in malignant cells, leading to tumor lysis and priming of antitumor immunity. Live attenuated MeV vaccine strains are therefore under investigation as cancer therapeutics. The versatile MeV reverse genetics systems allows for engineering of advanced targeted, armed, and shielded oncolytic viral vectors. Therapeutic efficacy can further be enhanced by combination treatments. An emerging focus in this regard is combination immunotherapy, especially with immune checkpoint blockade. Despite challenges arising from antiviral immunity, availability of preclinical models, and GMP production, early clinical trials have demonstrated safety of oncolytic MeV and yielded promising efficacy data. Future clinical trials with engineered viruses, rational combination regimens, and comprehensive translational research programs will realize the potential of oncolytic immunotherapy.

Oncolytic measles viruses (MV) derived from the live attenuated vaccine strain have been engineered for increased antitumor activity, and are currently under investigation in clinical phase 1 trials. Approaches with other viral vectors have shown that insertion of immunomodulatory transgenes enhances the therapeutic potency. In this study, we engineered MV for expression of the cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF). For the first time, therapeutic efficacy and adaptive immune response in the context of MV oncolysis could be evaluated in the previously established immunocompetent murine colon adenocarcinoma model MC38cea. MC38cea cells express the human carcinoembryonic antigen (CEA), allowing for infection with retargeted MV. Intratumoral application of MV-GMCSF significantly delayed tumor progression and prolonged median overall survival compared with control virus-treated mice. Importantly, more than one-third of mice treated with MV-GMCSF showed complete tumor remission and rejected successive tumor reengraftment, demonstrating robust long-term protection. An enhanced cell-mediated tumor-specific immune response could be detected by lactate dehydrogenase assay and interferon-γ enzyme-linked immunospot assay. Furthermore, MV-GMCSF treatment correlated with increased abundance of tumor-infiltrating CD3⁺ lymphocytes analyzed by quantitative microscopy of tumor sections. These findings underline the potential of oncolytic, GM-CSF-expressing MV as an effective therapeutic cancer vaccine actively recruiting adaptive immune responses for enhanced therapeutic impact and tumor elimination. Thus, the treatment benefit of this combined immunovirotherapy approach has direct implications for future clinical trials.

Combination of oncolytic virotherapy with immunomodulators is emerging as a promising therapeutic strategy for numerous tumor entities. In this study, we developed measles Schwarz vaccine strain vectors encoding immunomodulators to support different phases in the establishment of antitumor immune responses. Therapeutic efficacy of the novel vectors was evaluated in the immunocompetent MC38cea tumor model. We identified vectors encoding an IL-12 fusion protein (MeVac FmIL-12) and an antibody against PD-L1 (MeVac anti-PD-L1), respectively, as the most effective. Treatment of established tumors with MeVac FmIL-12 achieved 90% complete remissions. Profiling of the tumor immune microenvironment revealed activation of a type 1 T helper cell-directed response, with MeVac FmIL-12 ensuring potent early natural killer and effector T cell activation as well as upregulation of the effector cytokines IFN-γ and TNF-α. CD8⁺ T cells were found to be essential for the therapeutic efficacy of MeVac FmIL-12. Results of this study present MeVac FmIL-12 as a novel approach for targeted IL-12 delivery and elucidate mechanisms of successful immunovirotherapy.