Multi-omics characterization of response to tyrosine kinase inhibitor plus immunotherapy in MSS/pMMR metastatic colorectal cancer: A multicenter biomarker study.

206 Background: Patients with refractory microsatellite-stable metastatic colorectal cancer (MSS mCRC), representing the vast majority of mCRC cases, derive minimal benefit from immune checkpoint inhibitor (ICI) monotherapy. Combining anti-angiogenic tyrosine kinase inhibitors (TKIs) with ICIs has emerged as a promising strategy to overcome resistance. Methods: This multicenter retrospective study compared the efficacy and safety of anlotinib plus tislelizumab (n=65) versus anlotinib monotherapy (n=59) in patients with refractory MSS mCRC. Integrative whole-exome sequencing (WES), RNA sequencing, and multiplex immunohistochemistry (mIHC) were performed on pre-treatment tumor samples (n=29 from the combination group) to identify biomarkers of response. Immunotherapy response was defined as complete response (CR), partial response (PR), or stable disease (SD) lasting ≥6 months. Results: Combination therapy significantly improved median progression-free survival (196 vs 122 days, P =0.028) and median overall survival (453 vs 245 days, P <0.001) versus monotherapy. Genomic analysis linked TP53 pathway ( P =0.0022) and FLG2 ( P =0.0421) mutations to non-response, and identified mutations in the nucleotide excision repair (NER) ( P =0.0169) sub-pathway of DNA damage response (DDR) showed a trend toward association with response to combination therapy. RNA sequencing suggested that increased immune and metabolic pathways correlated with a better response. Crucially, responders exhibited significantly higher pre-treatment intratumoral infiltration of plasma cells (CD138+) ( P = 0.0237), while CD8+ T-cell infiltration did not differ. A predictive model integrating TP53/NER pathway status, FLG2 mutation, and plasma cell signatures achieved an AUC of 0.962 for response prediction. Conclusions: Anlotinib-tislelizumab combination therapy demonstrates modest antitumor efficacy in refractory MSS mCRC. NER pathway mutations, alongside plasma cell infiltration and TP53-pathway/FLG2 mutations, constitute a novel multi-omics biomarker framework for predicting therapeutic response, supporting contribution to the development of immune combination therapy and the prediction of its efficacy. Clinical trial information: NCT06573424 .