Temporal changes in tumor-associated immune cell populations in subcutaneous Tu-2449SC tumors transduced with Toca 511 following treatment with 5-fluorocytosine

Abstract Toca 511 (vocimagene amiretrorepvec), a gamma retroviral replicating vector encoding cytosine deaminase, with 5-fluorocytosine (5-FC), kills tumor by local production of 5-fluorouracil (5-FU), and induces local and systemic immunotherapeutic response resulting in long-term survival after cessation of 5-FC. This work identified the immune cell populations that change over time following administration of 5-FC to mice bearing Tu-2449SC tumors expressing cytosine deaminase. Tu-2449SC cells (2% pretransduced with Toca 511) were injected subcutaneously in B6C3F1 mice. 5-FC or PBS treatment was initiated once tumors were palpable. Tumor burden was significantly reduced in mice that received 5-FC vs. PBS control. At 3, 6, 9, and 14 days after 5-FC start, spleens and tumors were harvested for immunophenotyping. 3 days after treatment initiation, B cells and CD4+ T cells in the tumor were significantly reduced with 5-FC. Of the CD4+ T cells that remained in 5-FC treated tumors, a greater number expressed markers of activation (41BB, CD40L, and PD-1), were defined as effector T cells, and fewer were naïve. At day 6 post 5-FC initiation, CD4+ T cell percentages were no longer reduced with 5-FC vs. PBS. Tumor associated macrophages (TAM), myeloid derived suppressor cells (MDSC), and monocyte populations were significantly reduced in tumors 6 days after 5-FC start. Myeloid cell depletion correlates with previous work with systemic 5-FU. At 14 days post 5-FC, TAM and MDSC remained reduced in tumors of treated animals, and both CD4+ and CD8+ T cells were significantly increased. Toca 511+5-FC treatment results in reduced tumor burden and creates a tumor microenvironment that is more permissive to immune activation and anti-tumor immune response.