Single-cell transcriptomics unveils the immunologic landscape of anti-PD-1-associated indirect drug-induced liver injury.

[BACKGROUND] Immune-related adverse events (irAEs) caused by immune checkpoint inhibitors (ICIs) have become a bottleneck limiting their widespread use in anti-cancer therapy. Among them, immune checkpoint inhibitor-associated liver injury (ILICI) is a unique entity of drug-induced liver injury (DILI) whose clinical management has become a notable emerging challenge in cancer therapy. Nonetheless, the exact pathological mechanisms of ILICI are still poorly understood.

[METHODS] We established a Lewis lung adenocarcinoma-bearing mouse model in C57BL/6 J mice treated with anti-PD-1 (αPD-1). Subsequently, liver tissues from the two mouse groups were collected for single-cell RNA sequencing (scRNA-seq), and the core pathogenic mechanisms of αPD-1-induced liver injury were subsequently validated using both animal tissues and patient specimens.

[RESULTS] Our model revealed that αPD-1 treatment induced a hepatic phenotype characterized by focal inflammatory infiltration and fibrosis, concomitant with elevated serum aminotransferase (ALT) and pro-inflammatory cytokine levels. Further scRNA-seq analysis demonstrated that hepatocytes in the αPD-1 group exhibited features of disrupted lipid metabolism and enhanced NETosis, along with expanded CD8⁺ T cells displaying increased cytotoxicity and expansion of pro-inflammatory Ly6C monocytes. These Ly6C monocyte-derived macrophages robustly recruited and activated neutrophils via the Cxcl2-Cxcr2 and C3-(Itgam/Itgb2) signaling axes. Corroborated by validation in both animal and patient tissues, we identified neutrophil extracellular traps (NETs) as a pivotal event in αPD-1-associated liver injury.

[CONCLUSION] This study provides the first single-cell atlas of the hepatic microenvironment in a mouse model of αPD-1-induced liver injury under tumor-bearing conditions, revealing the concomitant metabolic reprogramming and profibrotic phenotype. Furthermore, our findings propose that the activation of NETosis is closely associated with the progression of liver injury, suggesting it may play a significant role in this pathological process.