Longitudinal single-cell and TCR repertoire profiling characterizes clonal entrapment in patients with pMMR/MSS locally advanced rectal cancer

Abstract Elevated intratumoral immune inflammation prior to treatment is typically associated with better outcomes in hot tumors treated with immune checkpoint inhibitors (ICIs). However, we observed a paradox in pMMR/MSS locally advanced rectal cancer (LARC) patients, where a subset with elevated baseline immune inflammation exhibited worse outcomes after combined radiotherapy and ICI treatment compared with patients with minimal immune inflammation. To investigate this counterintuitive finding, we performed paired scRNA-seq and scTCR-seq on longitudinally collected samples, including tumor biopsies (pre-treatment, post-radiotherapy, and post-immunotherapy) and peripheral blood mononuclear cells (pre-treatment and post-immunotherapy), from 20 pMMR/MSS LARC patients treated with sequential radiotherapy and ICI therapy (NCT06493240). We propose the concept of clonal entrapment to explain this phenomenon. Specifically, our profiling results reveal that increased HLA-DQA2 expression in dendritic cells and upregulated GDF15 expression in treatment-resistant tumor cells correlate with the restricted expansion of novel tumor-reactive TCR clonotypes. Consequently, the immune response is limited primarily by pre-existing TCR clonotypes within the tumor, especially those partially expanded under chronic inflammation, leading to the expansion of TCR clonotypes derived mainly from pre-treatment CD8 + T cell pools following ICI therapy. By identifying this feature of the pMMR/MSS LARC microenvironment, our study provides a high-resolution framework for understanding resistance to sequential radiotherapy and ICI therapy.