Background Cytokine-based drugs are being explored as alternative cancer immunotherapies.1 While the cytokine interleukin-18 (IL-18) has immunostimulatory effects, it is negatively regulated by a secreted high-affinity binding protein, IL-18BP, that functions as an immune checkpoint that limits IL-18’s therapeutic efficacy.2–5 A modified version of IL-18, termed ‘decoy-resistant’ or DR-18, that can avoid trapping by IL-18BP while maintaining immune signaling potential, has recently been developed.6 Here, we aim to test the efficacy and determine the cellular mechanism of action of DR-18 in combination with immune checkpoint inhibitors (ICIs) in immunocompetent preclinical models of renal cell carcinoma (RCC).
Methods We engrafted tumors subcutaneously using two different syngeneic, immunocompetent murine RCC models: Renca and RAG. Mice were treated with single-agent DR-18 and combinations of DR-18 with single- and dual-agent anti-PD-1 and anti-CTLA-4. Tumor growth and survival were monitored. In Renca, plasma was collected post-treatment and cytokine/chemokine levels were profiled using a 31-plex discovery assay. Single-cell RNA and TCR sequencing was also performed, and immune cell depletion studies were conducted.
Results In Renca, DR-18 monotherapy modestly inhibited tumor growth and prolonged survival (figure 1). The effects were comparable to single- and dual-agent ICIs. Adding PD-1 blockade to DR-18 did not enhance efficacy whereas the addition of anti-CTLA-4 to DR-18 significantly increased anti-tumor effects. Triple-therapy (DR-18 plus anti-PD-1 plus anti-CTLA-4) did not further inhibit tumor growth or prolong survival compared to the doublet (DR-18 plus anti-CTLA-4). The RAG model produced similar results, showing modest anti-tumor activity of single-agent DR-18 and enhanced benefit of combining with anti-CTLA-4 but not anti-PD-1. Cytokine/chemokine profiling revealed significantly elevated levels of IP-10 (CXCL10) and MIG (CXCL9) after one cycle of DR-18 plus anti-CTLA-4 compared to controls (figure 2). Single-cell transcriptomics demonstrated changes in intra-tumoral T cells, macrophages, and granulocytes with DR-18 plus anti-CTLA-4 relative to other regimens, including enrichment of CD8+ precursor and terminally exhausted T cells and a neutrophil population associated with interferon signaling (figure 3). Immune cell depletion studies identified CD8+ T cells, NK cells, and interferon-gamma as equally required for efficacy of DR-18 plus anti-CTLA-4 (figure 4).
Conclusions We identify DR-18, a ‘decoy-resistant’ IL-18, in combination with anti-CTLA-4 as having enhanced anti-tumor activity in preclinical models of RCC. This regimen was associated with a more pro-inflammatory immune microenvironment. Investigation is ongoing to further elucidate the cellular mechanism of action of this regimen and lay the groundwork for clinical testing of DR-18-based combination therapy in RCC.
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