Spatial transcriptomics analysis of Merkel cell carcinoma reveals novel immune cell subsets and hypoxia as correlates of immunotherapy response

Summary: Immune checkpoint blockade remains the standard of care for advanced Merkel cell carcinoma (MCC); however, approximately 50% of patients develop either primary or acquired resistance. Although the tumor microenvironment (TME) is implicated in mediating therapeutic outcomes, the specific cell types involved, and the impact of their spatial organization remain largely undefined. To address this gap, we applied Xenium spatial transcriptomics (ST) to 27 treatment-naive MCC tumors from a clinical trial of retifanlimab in advanced MCC, with the goal of uncovering TME correlates of immunotherapy response. Our analysis revealed the presence of CXCL9 and SPP1 macrophages that have not been previously reported in MCC. These macrophages were not detectable in several single cell RNA-sequencing (scRNA-seq) MCC datasets, suggesting ST can uncover cell populations missed by traditional scRNA-Seq. Other immune subsets were also observed, including CD4 and CD8 T cells expressing hallmarks of tumor reactivity such as CXCL13, NK cells, B cells, and plasma cells. The tumor compartment contained distinct subpopulations of hypoxic and proliferating tumor cells with unique spatial organization. Cellular neighborhood analysis revealed 3 types of tissue domains: immune rich, tumor rich, and stromal rich regions. Higher levels of hypoxic and proliferating tumor cells were associated with poor outcomes. Interestingly, overall immune cell composition was relatively homogeneous between responders and non-responders, suggesting that immune cell spatial relationships, rather than abundance, may be more strongly associated with immunotherapy response. Tomas Bencomo^{1, 2}, Carina Morningstar¹, Haroldo J. Rodriguez Chevez¹, Dominik Otto², Michael Smith³, Evan Newell², Manu Setty², Paul Nghiem^{1, 2} 1. Dermatology, University of Washington, Seattle, WA, United States. 2. Fred Hutchinson Cancer Center, Seattle, WA, United States. 3. Incyte Corporation, Wilmington, DE, United States. Translational Studies: Cell and Molecular Biology