Anti-PD-L1-IFN-α-adjuvanted HBsAg vaccine overcomes HBV immune tolerance through targeting both DCs and macrophages.

Recombinant hepatitis B surface antigen (rHBsAg) vaccine with various adjuvants fails to break T and B cell tolerance in hosts with chronic hepatitis B (CHB). This study aims to explore the mechanisms to break immune tolerance that allows the host to respond to rHBsAg, achieving a cure for CHB. We engineered an anti-PD-L1-IFN-α (aPD-L1-IFN-α) heterodimeric fusion protein to allow rHBsAg to rejuvenate T and B cell responses in hepatitis B virus-tolerant (HBV-tolerant) mice. S.c. coimmunization with aPD-L1-IFN-α and rHBsAg significantly enhanced antigen uptake and maturation of both macrophage and dendritic cell (DC) subsets in draining lymph nodes. Macrophages drove early B cell activation, while cDC1s primed CD8+ T cells, breaking tolerance and leading to both B cell and cytotoxic T lymphocyte (CTL) differentiation. This strategy elicited not only anti-HBsAg neutralizing antibodies but also HBsAg-specific CD8+ T cell responses, achieving a functional cure without systemic toxicity. The efficacy of the aPD-L1-IFN-α adjuvant depended on both PD-L1 cis-targeting and IFN-α receptor signaling in antigen-presenting cells. These findings establish aPD-L1-IFN-α as a translatable adjuvant to break the strong tolerance induced by CHB, providing a dual-pathway strategy to induce HBV-specific T and B cell responses.