CCR2⁻ T Peripheral Helper Cells as Potential Coordinators of Local Immune Architecture in Human Cancer

Abstract Introduction Immune checkpoint inhibitors (ICIs) have revolutionized cancer treatment, yet tumor progression remains a challenge, necessitating improved patient stratification and therapeutic strategies. Tumor growth releases neoantigens that activate adaptive immunity, promoting tertiary lymphoid structure (TLS) formation through B and T cell interactions. T follicular helper (Tfh) cells are key in coordinating B cell responses and are linked to favorable outcomes. A newer subset, T peripheral helper (Tph) cells, shares functional traits with Tfh cells but differs in transcriptional and migratory profiles. Though observed in various cancers, their distribution and role in tumor immunity are not fully understood. Methods To investigate this, a multiplex cyclic immunofluorescence (Cyc-IF) assay was developed to detect and spatially analyze Tph cells in tumors. Results This revealed three CXCL13-expressing CD4⁺ T cell subsets: Tfh, Tph, and a “CCR2⁻ Tph-like” population which, which, in contrast to the classically CCR2-enriched Tph phenotype described in inflamed tissues, shows markedly reduced CCR2 expression. Downregulation of CXCR5 and CCR2 near CCR2⁻ Tph-like cells suggested a shift toward local immune residency, forming immune niches. These niches were enriched with pro-inflammatory cells, including Th1, Th17, CD4⁺, CD8⁺ T cells, and B cells. Spatial profiling showed CCR2⁻ Tph-like cells embedded in an immunoregulatory network, marked by CD69 and inhibitory checkpoints B7-H3 and PD-L1 on surrounding cells. Conclusions This dual signaling suggests CCR2⁻ Tph-like cells may modulate tumor immunity by balancing activation and suppression, with potential implications for checkpoint blockade therapy.