Associations between tumor immune response and prognosis in node-negative breast cancer patients in the randomized DBCG HYPO trial

Moderately hypofractionated radiotherapy is increasingly replacing conventional schedules (2 Gy/fraction) in adjuvant breast cancer treatment. Tumor-infiltrating lymphocytes (TILs) have emerged as a prognostic biomarker, particularly in aggressive subtypes, but their role in node-negative patients and their relation to radiotherapy fractionation remain unclear. Immune infiltration in primary tumors was examined in the randomized Danish Breast Cancer Group HYPO trial, including 1329 node-negative breast cancer patients treated with breast-conserving surgery and randomized to standard fractionated (50 Gy/25 fractions) versus hypofractionated (40 Gy/15 fractions) radiotherapy. A case-cohort design included 349/1329 patients for histopathological analyses of TILs and immune cell subsets (CD8, CD4, FOXP3, CD68, CD11c). Associations with the primary endpoint, overall mortality (OM), were evaluated using multivariable flexible parametric survival models. Predictive effects of immune markers on fractionation were assessed through interaction tests. Median follow-up was 7.9 years. High TILs (cut-off ≥ 30%) were observed in 18% of tumors and showed a trend toward improved outcomes, most pronounced in patients with estrogen receptor (ER)-negative tumors (hazard ratio (HR) 0.43, 95% CI (0.09–2.03)). For ER-negative tumors, high CD8 + T-cell infiltration corresponded to an estimated absolute reduction in OM-risk of 43% compared with low CD8 + T-cell infiltration (HR 0.06 (0.01–0.47), test for interaction p = 0.13). In contrast, no consistent prognostic effect was observed in ER-positive disease. No predictive interaction was identified between immune markers and fractionation. Immune infiltration showed trends consistent with a prognostic association in node-negative breast cancer, with the strongest association in ER-negative tumors, in line with findings in node-positive cohorts. No predictive value of immune markers for fractionation was observed. These exploratory findings suggest that immune composition may hold prognostic information across risk groups.