Abstract 637: Single-cell analysis of systemic immune response to sequential radiation and immune checkpoint blockade in metastatic colorectal cancers

Abstract Immune checkpoint blockade (ICB) has shown significant clinical efficacy in mismatch repair deficient colorectal cancers (CRCs). However, an effective means to use these therapies for mismatch repair proficient (pMMR) CRC has yet to be developed. To enhance the efficacy of ICB for patients with oligometastatic pMMR CRC, we performed a clinical trial evaluating the sequential use of stereotactic body radiotherapy (SBRT), pembrolizumab, surgical resection, and then adjuvant pembrolizumab (NCT02837263). The primary endpoint of 1-year recurrence-free survival (1 yr RFS) was met in 60% of trial participants compared to a historical control of 50%. From this trial, 13 subjects could have blood samples collected for peripheral blood mononuclear cell isolation pretreatment and after operative management. We hypothesize that the systemic immune response changes upon combined treatment and is distinct between responders and non-responders. We performed scRNA-Seq on these samples to compare the circulating immune cell populations with clinical outcomes. Despite a small sample size, integrated bioinformatics analysis identified (i) pre-existing CD8+ T cells were observed with treatment response (p = 0.031) (ii) combined RT and ICB induces changes in myeloid cell phenotypes, including reduced pro-inflammatory monocytes (p = 0.038) and conventional cDC2 (p = 0.030) in subjects without recurrence and increasing pDCs (p = 0.024) in most patients. CD8+ T cells from those recurrence-free subjects showed terminally differentiated effector phenotypes, expressing CX3CR1, GZMB, and KLRG1 before treatment. cDC2s expressed interferon activity markers, including STAT1, IFI44, and IRF7, that presented a more activated state. CD8+ T cell abundance remained stable, with no significant changes before and after treatment. In contrast, myeloid cells showed greater variability, reflecting unique polarization states through differences in cytokine (CCL3, CXCL8), interferon (ISG15, IFI44), and complement (C1QA, C1QB) associated genes. Future work will increase sample size in independent patient cohorts to elucidate the extent to which circulating phenotypes are also identified in tumors. Overall, this study demonstrates pre-existing T cell abundance and changes in myeloid cell phenotypes may be associated with clinical outcomes from combined immunotherapy, which have the potential to serve as predictive biomarkers for therapeutic response. Citation Format: Joshua Brand, Sean G. Kraus, Michael Bassetti, Dustin A. Deming, Huy Q. Dinh. Single-cell analysis of systemic immune response to sequential radiation and immune checkpoint blockade in metastatic colorectal cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 637.