2795 - Longitudinal Multi-omic Immune Profiling and Persistent Systemic Dysregulation and Baseline Predictors of Recurrence after Breast Cancer Radiotherapy

Purpose/Objective(s):

Identification of subsets of breast cancer patients undergoing adjuvant radiotherapy (RT) at elevated recurrence risk andmay benefit from novel combinatorial approaches remains an unmet clinical need. This study aimed to characterize longitudinal changes in circulating immune cells and proteins during and after breast/chest wall plus regional nodal irradiation.

Materials/Methods: A cohort of 121 patients with baseline samples was analyzed; 31 also had serial samples collected at RT completion and post-treatment follow-up. CyTOF-based immunophenotyping was used to characterize circulating immune cell subsets, and Olink proteomic assays quantified circulating cytokines and chemokines. Linear mixed-effects models assessed differences in longitudinal immune trajectories. The prognostic value of baseline biomarkers was assessed using log-rank testing for recurrence-free survival (RFS).

Results:

Median age was 53 years, with tumor subtypes including Luminal A/B (39.5%), triple-negative breast cancer (TNBC; 48.7%), and HER2-amplified disease (11.8%). Over a median follow-up of 47 months, 49 patients (40.5%) experienced recurrence; median RFS was not reached, with a 3-year RFS rate of 57.3% (95% CI, 48.2%–68.1%). Baseline characteristics were well balanced between the recurrence and non-recurrence groups.

Longitudinal CyTOF profiling (n=31) revealed that most immune cell populations, including CD8⁺ T cells, NK cells, and B cells, declined during RT and recovered after treatment completion. NK and B cell frequencies were significantly lower in patients who recurred at all time points (Group P < 0.05). In contrast, recurrent patients exhibited a significant and persistent accumulation of CD8+ aß T cells and Th1-like cells (Group P < 0.05).

Longitudinal OLINK preotomic analysis (n=30) identified CXCL13 as a key differentiating protein among 92 analytes (P = 0.017), with sustained suppression in recurrent patients. Recurrent patients also demonstrated numerically lower FASLG and higher IL-6 levels, although these differences did not reach statistical significance. FASLG levels were strongly positively correlated with NK and B cell frequencies, whereas IL-6 levels were negatively correlated with CD8? aß T cells and Th1-like cells.

In the full cohort (n=121), high baseline IL-6 and low baseline FASLG were significantly associated with inferior RFS (P < 0.01).

Conclusion:

Breast cancer patients who recur after radiotherapy exhibit a pre-existing and persistent systemic immune dysregulation characterized by NK and B cell suppression, accumulation of CD8? aß T and Th1-like cells, and sustained CXCL13 downregulation. Coordinated alterations in immune cell populations and circulating inflammatory mediators, particularly involving FASLG and IL-6, suggest biologically linked pathways underlying recurrence risk. Baseline IL-6 and FASLG may enable early risk stratification and inform the development of novel radio-immunotherapeutic strategies.