Pretreatment stromal CD4⁺ T-cell density predicts favorable response to total neoadjuvant therapy in locally advanced...

Total neoadjuvant therapy (TNT) has become an important treatment strategy for locally advanced rectal cancer (LARC), increasing the need for pretreatment biomarkers that may predict major tumor response and support organ-preserving approaches. This study aimed to evaluate whether pretreatment stromal CD4⁺ and CD8⁺ T-cell densities, together with stromal tumor-infiltrating lymphocytes (TILs), are associated with response to TNT in patients with LARC. In this retrospective study, 45 patients with LARC who completed TNT were analyzed; pretreatment colonoscopic biopsy specimens were available for all patients, and paired post-treatment resection specimens were available for 40 surgically treated patients. CD4⁺ and CD8⁺ T-cell densities were assessed immunohistochemically in stromal hotspot areas and expressed as cells/mm², while stromal TILs were evaluated on hematoxylin-eosin sections. A favorable response was defined as tumor regression grade (TRG) 0-1 in surgically treated patients or clinical complete response in patients managed with a watch-and-wait strategy. Median pretreatment CD4⁺ T-cell density, CD8⁺ T-cell density, and stromal TIL percentage were 320 cells/mm², 64 cells/mm², and 30%, respectively. Receiver operating characteristic (ROC) analysis identified a pretreatment stromal CD4⁺ T-cell density cutoff of 410 cells/mm² for favorable response, with an area under the curve (AUC) of 0.68 (p=0.041). CD4⁺ T-cell density >410 cells/mm² was significantly associated with favorable response and remained independently associated with response in multivariable logistic regression analysis (odds ratio [OR]=5.05, 95% confidence interval [CI]: 1.21-20.83, p=0.026). In contrast, pretreatment CD8⁺ T-cell density and stromal TIL percentage were not significantly associated with response. In paired specimens, CD8⁺ T-cell density increased significantly after treatment (p<0.001), whereas CD4⁺ T-cell density and stromal TIL percentage did not change significantly. Lymphovascular invasion (LVI) and perineural invasion (PNI) were associated with non-favorable response. High pretreatment stromal CD4⁺ T-cell density may represent a candidate biomarker of favorable response to TNT in LARC, while the post-treatment increase in CD8⁺ T-cell density suggests a potential immunomodulatory effect of TNT; however, prospective validation in larger cohorts is required.