TLR8 agonists remodel the tumor immune microenvironment through PF4-dependent T cell recruitment and ancillary mechanisms

Pattern recognition receptors (PRRs) are crucial modulators of the tumor immune microenvironment (TIME); yet, their comparative efficacy remains poorly characterized. Platelet factor 4 (PF4/CXCL4) exerts paradoxical effects in cancer, and its regulation by PRR signaling is unclear. Here, we systematically screened PRR agonists in murine tumor models and identified TLR8 agonists as the most potent inducers of a pro-immunogenic TIME, superior to agonists targeting TLR3/7/9 or NOD1/2. TLR8 activation drove CD45+ leukocyte infiltration, potentiated conventional dendritic cell (cDC) and macrophage phagocytosis and migration, amplified recruitment of CD8+ and conventional CD4+ (cCD4+) T cells, lowered Treg proportions, elicited pro-inflammatory and tumoricidal phenotypes in innate immune cells and CD8+ T cells. Notably, TLR8 agonism suppressed tumor growth in both immunocompetent and T cell-deficient mice, indicating the involvement of both innate and adaptive immunity. Mechanistically, TLR8 agonists upregulated PF4 expression in macrophages, cDC2, and CD8+ T cells via the NF-κB pathway. PF4 in turn recruited cCD4+ T cells via CXCR3, and its local overexpression mimicked the antitumor effect of TLR8 activation. Beyond PF4, TLR8 signaling mediated PF4-independent effects, including Treg suppression via IFN-γ and enhanced macrophage phagocytosis. Combination of a TLR8 agonist with anti-PD-1 therapy markedly and synergistically improved survival of tumor-bearing mice. Thus, TLR8 agonists optimally remodel the TIME through PF4-dependent T cell recruitment and PF4-independent ancillary mechanisms. Our finding that the antitumor activity of locally induced PF4 contrasts with its reported protumor effects when expressed systemically clarifies the context-dependent duality of PF4 in cancer. These results position TLR8 agonists as promising candidates for combination immunotherapy.