Neoadjuvant treatment is associated with fibrotic and transcriptional remodeling of tumor-draining lymph nodes in rectal cancer

Background Neoadjuvant treatment is associated with reduced lymph-node yield in rectal cancer, but the accompanying changes in tumor-draining lymph nodes (TDLNs) remain incompletely characterized. Methods We analyzed Surveillance, Epidemiology, and End Results data, a 178-patient institutional cohort, quantitative histopathology of 447 histologically negative TDLNs from 159 patients, and bulk RNA sequencing of 30 nodes from 24 patients. Institutional comparisons distinguished a radiotherapy-containing treatment pathway (RT) from management without preoperative radiotherapy (NRT); in the institutional clinical cohort, all 105 RT patients and 10 of 73 NRT patients received neoadjuvant chemotherapy. Pathology models accounted for patient clustering, and RNA counts were aggregated within patients. Results Recorded preoperative radiotherapy was associated with lower examined-node counts in Surveillance, Epidemiology, and End Results (SEER). In the institutional cohort, the expanded pretreatment-adjusted incidence-rate ratio was 0.83 [95% confidence interval (CI), 0.67–1.02]; pretreatment-covariate overlap weighting yielded 0.80 (95% CI, 0.66–0.96). RT-group nodes were more fibrotic and smaller, with lower germinal-center and lymphoid-tissue area fractions. Patient-level RNA-seq identified 4,446 differentially expressed genes, with higher extracellular-matrix and angiogenic programs and lower antigen-receptor and B-cell-receptor signaling. Selected pathway directions were retained across sensitivity analyses. Computational deconvolution provided supportive estimates of relative cell composition. Conclusion A radiotherapy-containing neoadjuvant treatment pathway, including chemoradiotherapy in the institutional clinical, pathology, and RNA-seq cohorts, was associated with lower node yield and structural and transcriptional differences in TDLNs. These findings characterize the nodal structural and transcriptional phenotype associated with neoadjuvant treatment.