Abstract PR003: A feed-forward loop in the adenosine signaling pathway drives myeloid-mediated resistance to immune checkpoint inhibition in prostate cancer
Authors: Lyu Aram , Fan Zenghua , Clark Matthew , Lea Averey , Luong Diamond , Setayesh Ali , Starzinski Alec , Wolters Rachel , Arias-Badia Marcel , Allaire Kate , Wu Kai , Gurunathan Vibha , Valderrábano Laura , Wei Xiao , Miller Richard , Van Allen Eliezer , Fong Lawrence 
Published: February 2025

Background: Patients with metastatic castration-resistant prostate cancer (mCRPC) are generally resistant to immune checkpoint inhibitors (ICIs), partly due to the presence of immunosuppressive myeloid cells within tumors. However, broad targeting strategies, such as CSF1R antagonism, have faced clinical limitations given the heterogeneity of these cells. We hypothesize that a more precise understanding of specific immunosuppressive myeloid subsets and their molecular mechanisms at the single-cell level will be essential in the development of effective immunotherapies. Methods: We performed single-cell profiling on tumor biopsies across patients with localized (n=13), metastatic hormone-sensitive (n=24), and metastatic castration-resistant (n=6) prostate cancer. Results from these analyses were reverse-translated into multi syngeneic mouse models of prostate cancer, where we conducted multi-omic single-cell assessments and mechanistic studies. Findings were subsequently correlated with clinical outcomes. Results: Single-cell analysis of patient biopsies identified a distinct subset of tumor-associated macrophages with high expression of SPP1 (SPP1 hi-TAMs), encoding the osteogenic protein osteopontin (OPN), which were more prevalent in mCRPC than in earlier disease stages. These macrophages showed elevated immunosuppressive molecular programs and significantly lower CSF1R expression than other macrophages, potentially explaining the limited efficacy of CSF1R blockade in mCRPC patients. Multi-omic profiling of syngeneic mouse models of prostate cancer (MyC-CaP and TRAMP-C2) identified an analogous macrophage subset resistant to CSF1R blockade, which strongly suppressed CD8+ T cell activity in co-culture. Adoptive transfer of Spp1 hi-TAMs into CRPC increased the frequency of exhausted CD8+ T cells, promoted ICI resistance and worsened survival outcomes. Pathway analysis implicated adenosine signaling as a possible mechanism. Blocking adenosine pathways, either with the A2AR inhibitor ciforadenant or a CD73-blocking antibody, significantly reversed CD8+ T cell suppression by Spp1 hi-TAMs in vitro and enhanced the sensitivity of CRPC cells to PD-1 blockade in vivo. Moreover, we observed a reduction in Spp1 hi-TAMs following A2AR blockade, consistent with the high expression of Adora2a transcripts (encoding A2AR) and activation of downstream pathways, suggesting a feed-forward loop of adenosine pathways contribute to ICI resistance mediated by SPP1 hi-TAMs. Clinical correlation showed that a subset of mCRPC patients treated with combined adenosine receptor and PD-L1 blockade had improved anti-tumor efficacy compared to those on monotherapy, with SPP1 hi-TAMs potentially serving as predictive biomarkers for treatment response. Conclusions: Our studies reveal a significant increase in SPP1 hi-TAMs abundance with disease progression, contributing to ICI resistance through adenosine pathways. These findings support the therapeutic targeting of SPP1 hi-TAMs and their associated pathways as promising strategies to overcome resistance.

Citation Format: Aram Lyu, Zenghua Fan, Matthew Clark, Averey Lea, Diamond Luong, Ali Setayesh, Alec Starzinski, Rachel Wolters, Marcel Arias-Badia, Kate Allaire, Kai Wu, Vibha Gurunathan, Laura Valderrábano, Xiao Wei, Richard Miller, Eliezer Van Allen, Lawrence Fong. A feed-forward loop in the adenosine signaling pathway drives myeloid-mediated resistance to immune checkpoint inhibition in prostate cancer [abstract]. In: Proceedings of the AACR IO Conference: Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2025 Feb 23-26; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2025;13(2 Suppl):Abstract nr PR003.