Integrated BATF transcriptional network regulates suppressive intratumoral regulatory T cells

Human regulatory T cells (T_regs) are crucial regulators of tissue repair, autoimmune diseases, and cancer. However, it is challenging to inhibit the suppressive function of T_regs for cancer therapy without affecting immune homeostasis. Identifying pathways that may distinguish tumor-restricted T_regs is important, yet the transcriptional programs that control intratumoral T_reg gene expression, and that are distinct from T_regs in healthy tissues, remain largely unknown. We profiled single-cell transcriptomes of CD4⁺ T cells in tumors and peripheral blood from patients with head and neck squamous cell carcinomas (HNSCC) and those in nontumor tonsil tissues and peripheral blood from healthy donors. We identified a subpopulation of activated T_regs expressing multiple tumor necrosis factor receptor (TNFR) genes (TNFR⁺ T_regs) that is highly enriched in the tumor microenvironment (TME) compared with nontumor tissue and the periphery. TNFR⁺ T_regs are associated with worse prognosis in HNSCC and across multiple solid tumor types. Mechanistically, the transcription factor BATF is a central component of a gene regulatory network that governs key aspects of TNFR⁺ T_regs. CRISPR-Cas9-mediated BATF knockout in human activated T_regs in conjunction with bulk RNA sequencing, immunophenotyping, and in vitro functional assays corroborated the central role of BATF in limiting excessive activation and promoting the survival of human activated T_regs. Last, we identified a suite of surface molecules reflective of the BATF-driven transcriptional network on intratumoral T_regs in patients with HNSCC. These findings uncover a primary transcriptional regulator of highly suppressive intratumoral T_regs, highlighting potential opportunities for therapeutic intervention in cancer without affecting immune homeostasis.