Authors: Tiffany T Montellano; Sophie Viaud, PhD; Daniel Mendoza; Maria Rodriguez-Aguirre; Harry Gruber, MD; Douglas Jolly, PhD; Derek Ostertag, PhD (San Diego, CA)

Introduction:  TVocimagene amiretrorepvec is an investigational, conditionally lytic, retroviral replicating vector that selectively infects cancer cells due to their high mitotic index and immune suppressed tumor microenvironment. Vocimagene amiretrorepvec stably delivers optimized yeast cytosine deaminase (CD) that converts subsequent extended release version of 5 fluorocytosine [5 FC] into 5 fluorouracil (5 FU). 5 FU kills infected dividing cancer cells and, in preclinical models, local immunosuppressive myeloid cells, stimulating anti tumor immunity. Patients with recurrent high grade gliomas treated with a local injection of vocimagene amiretrorepvec followed by treatment with 5 fluorocytosine have shown prolonged survival and durable complete responses (median duration of follow up for response: 37.4+ months); responses were delayed in onset, consistent with an immunological mechanism. These data led to a phase III trial (NCT02414165). To improve therapy, we implemented a novel preclinical system to model submaximal infections and investigate combination treatments such as with metronomic cyclophosphamide for its immuno stimulatory and antiangiogenic properties. Methods: Naive B6C3F1 mice were implanted in the flank with a mouse glioma cell line Tu 2449SQ that has been adapted to grow subcutaneously. In order to control vocimagene amiretrorepvec spread, tumor cells previously transduced with vocimagene amiretrorepvec or a sister vector that expresses GFP (Green Fluorescent Protein) instead of CD, were admixed at various percentages as vocimagene amiretrorepvec does not readily infect cells already infected with the GFP virus. Results:  We show a tumor control benefit when metronomic cyclophosphamide is combined with 5 FC with only 10% of tumor cells infected with vocimagene amiretrorepvec, which is associated with a significant depletion of peripheral Treg and increase in CD8+ T cells in the peripheral blood. Conclusion:  These data demonstrate that vocimagene amiretrorepvec and 5 FC therapy can be combined with metronomic chemotherapeutics like cyclophosphamide to enhance efficacy in preclinical models and may help design future clinical development.