A predictive 14-gene signature and FLI1 inhibition overcome tumor-infiltrating lymphocyte dysfunction in triple-negative breast cancer.

Triple-negative breast cancer (TNBC) is an aggressive subtype lacking effective targeted therapies. Although immune checkpoint inhibitors such as pembrolizumab have improved clinical outcomes in a subset of patients, limited response rates and durability highlight the need for alternative strategies. Tumor-infiltrating lymphocyte (TIL)-based adoptive cell therapy represents a promising approach; however, the absence of reliable biomarkers for TIL functionality remains a major obstacle. In this study, we isolated TILs and autologous tumor cells from nine TNBC patients and classified TILs as reactive or non-reactive based on cytotoxic activity. Transcriptomic profiling combined with weighted gene co-expression network analysis and machine learning approaches identified a 14-gene signature that robustly distinguished reactive from non-reactive TILs. Transcription factor analysis revealed that Friend leukemia integration 1 (FLI1) regulates a large proportion of genes upregulated in non-reactive TILs. Notably, pharmacological inhibition of FLI1 using TK216 restored the cytotoxic function of non-reactive TILs and reduced expression of immunosuppressive genes. These findings identify a functional gene signature associated with TIL reactivity and suggest that targeting FLI1 may represent a strategy to enhance TIL-based immunotherapy in TNBC.