Abstract

Background To elucidate the mechanism by which icaritin—a novel agent for hepatocellular carcinoma (HCC)—remodels the tumor microenvironment (TME) by inhibiting HCC cell metabolism-mediated M2 polarization of tumor-associated macrophages (TAMs). Methods Integrative approaches spanning in vitro Transwell cocultures, RNA-seq, LC3-based autophagy tracing, STX16 gene edition, and orthotopic xenografts mechanistically dissected the affective and mechanism of icaritin in remodeling TME. Results Our results indicate that icaritin transcriptionally suppresses ALDOB in HCC cells to reduce lactate production. Consequently, the lactylation of histone H3 at lysine 9 and lysine 18 (H3K9/H3K18la) on the STX16 promoter is diminished, thereby ablating STX16 transcription. STX16 deficiency blocks autophagolysosome biogenesis, leading to the accumulation of autophagosomes in HCC cells. These autophagosomes are subsequently delivered to macrophages via extracellular vesicle (EVs) and then triggers autophagic cell death and p62-guided STAT3 destruction within the macrophages, thereby eliminating M2 TAMs and reprograming the tumor immune microenvironment. In vivo validation confirmed icaritin suppressed tumor growth and M2 macrophage infiltration via the ALDOB/STX16/autophagy/STAT3 axis. Conclusion These results indicate that by orchestrating a novel "metabolism-epigenetics-autophagy-EVs" cascade to eliminate TAMs, icaritin targets ALDOB, STX16, and STAT3, revealing key nodes for therapeutic intervention. Graphical The alternative text for this image may have been generated using AI.