Intratumoral dsRNA Sensor Activation Redirects Radiation-Associated Myeloid Cells to Ignite Local and Systemic Anti-Tumor Immunity in Undifferentiated Pleomorphic Sarcoma

Abstract

<h3>Purpose/Objective(s)</h3> Undifferentiated pleomorphic sarcoma (UPS) responds more frequently to immune checkpoint blockade (ICB) than other sarcoma subtypes, but most patients do not respond. To elicit de novo immune responses, we targeted the myeloid-rich tumor microenvironment of UPS. Myeloid cells are highly plastic cells that accumulate in tumors after radiation therapy and can be directed to an antigen-presenting state to generate tumor-specific T cell responses. We hypothesized that intratumoral administration of BO-112, a nanoplexed double-stranded RNA (dsRNA) that activates innate immune sensors, would redirect RT-associated myeloid cells to generate local and systemic anti-tumor immunity. <h3>Materials/Methods</h3> We used an ICB-resistant, KrasG12D/-P53-/- mouse model of UPS to establish large tumors. Tumor-bearing mice were treated with RT (8 Gy x 3 fractions) ± BO-112 (2 doses intratumorally). Tumor volume was tracked between groups (combination, RT alone, BO-112 alone, and mock). To conduct temporospatial analysis of immune cells changes in response to treatment, high-dimensional flow cytometry was performed in tumor and lymphoid tissues. To determine the myeloid subset targeted by therapy, we used fluorescent BO-112 and flow cytometry-based cell imaging. To evaluate the fate of targeted cells, eGFP monocytes were intratumorally transferred and tracked. To dissect the role of lymphocytes, we used lymphocyte-deficient RAG-/-gc-/- mice and antibody-mediated depletion of T cells (CD4 and CD8) and NK cells. <h3>Results</h3> Combination therapy with RT and BO-112 led to maximal tumor control compared to mock and monotherapy groups in vivo, even in large, established tumors (p<0.0001). Intratumoral BO-112 preferentially targeted monocytic antigen presenting cells (Ly6Chi MHCII+CD11c+), which led to an egress of these cells from the tumor into the draining lymph node (p<0.05), confirmed by tracing eGFP+ monocytes adoptively transferred into the tumor. Combination therapy was accompanied by an increase in polyfunctional T cells within the tumor (p<0.05) and systemically (p<0.005) that was sustained over time. Genetic deletion of lymphocytes in RAG-/-gc-/- negated the effect of combination therapy and antibody-mediated subset depletion isolated this effect to CD8 T cells. <h3>Conclusion</h3> Intratumoral BO-112 preferentially targets myeloid cells and augments the response to radiation in an ICB-resistant model of UPS. Reprogramming RT-induced intratumoral myeloid cells toward an antigen-presenting fate and their redirection to the lymph node may bridge an adaptive T cell response and lead to enhanced tumor control.