Abstract PR009: Spatial single-cell transcriptomics reveal targetable macrophage subsets associated with antiangiogenic therapy resistance in renal cell carcinoma

Abstract Purpose: Antiangiogenic tyrosine kinase inhibitors (TKIs) have become a cornerstone in the management of clear cell renal cell carcinoma (ccRCC) as monotherapy or in combination with immune checkpoint blockades (ICB). However, TKIs produce limited long-term durability even in combination with ICBs, indicating that prolonged TKI treatment fosters an immunosuppressive microenvironment contributing to therapy resistance. Method: In this study, we performed Xenium in situ transcriptomic profiling with a 5, 000-gene panel on tissue microarrays comprising treatment-naïve or sunitinib-treated tumors from RCC patients. Renca murine RCC model was used to evaluate therapeutic responses in vivo. Results: From over 220, 000 analyzed cells, we identified two major tumor cell populations. The predominant population exhibited pronounced hypoxia and glycolytic activity, whereas a smaller subset of RCC cells showed reduced metabolic activity but increased metastatic potential and stem-like features. In sunitinib-treated tumors, both tumor and immune cells displayed a highly inflammatory activity, marked by upregulation of type I and type II interferon response pathways. Sunitinib treatment reduced the abundance of endothelial cells and cancer-associated fibroblasts and increased the infiltration of T cells and B cells. Although sunitinib did not alter the overall abundance of tumor-associated macrophages (TAMs), it markedly shifted their spatial distribution: in treatment-naïve tumors, TAMs were sequestered by endothelial cells, whereas in sunitinib-treated tumors, they were released and engaged to interaction with T cells. We further identified two TAM subsets—interferon-primed TAMs (IFN-TAMs) and proliferating TAMs (Prolif-TAMs) —that were expanded in sunitinib-treated tumors and associated with poorer progression-free survival. IFN-TAMs exhibited strong IFN-γ signaling activity and high expression of the immune checkpoint ligands PD-L1 and PD-L2. Prolif-TAMs were defined by elevated proliferation markers and by increased expression of tumor-promoting proteins, such as CSF1R, MSR1, VSIG4, and TREM2. In sunitinib-treated tumors, IFN-TAMs also showed enhanced interactions with multiple immune cell subsets, including proliferating T cells and cytotoxic T lymphocytes, both of which express PD-1. These findings suggest that IFN-TAMs facilitate tumor immune evasion through the PD-1/PD-L1/2 axis, consistent with our observation that PD-1 blockade synergizes with axitinib to enhance tumor suppression. Notably, both IFN- and Prolif-TAMs expressed the A2A adenosine receptor (A2AR), and we found that the A2AR antagonist CPI-444 further potentiated the therapeutic response to axitinib treatment. Conclusion: These results show that sunitinib treatment induces an immunosuppressive tumor microenvironment driven by macrophage reprogramming and altered spatial organization. Targeting these macrophage subsets markedly enhances the therapeutic efficacy of TKIs. Disclosure: AI assistance (ChatGPT) was used to polish the wording of this abstract. Citation Format: Xiande Liu, Hongchao He, Jingjing Liu, Xuesong Zhang, Anh Hoang, Tatiana Karpinets, Kanishka Sircar, Eric Jonasch. Spatial single-cell transcriptomics reveal targetable macrophage subsets associated with antiangiogenic therapy resistance in renal cell carcinoma abstract. In: Proceedings of the AACR Special Conference in Cancer Research: Innovations in Kidney Cancer Research: From Molecular Insights to Therapeutic Breakthroughs; 2026 Mar 13-16; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2026;86 (5Suppl₂): Abstract nr PR009.