Jianpi-huayu Decotion regulates TREM1/DAP12 pathway to improve the immunosuppressive tumor microenvironment and enhance the anti-hepatocellular carcinoma effect of PD-1 inhibitors.

[ETHNOPHARMACOLOGICAL RELEVANCE] Jianpi-Huayu Decoction (JHD), derived from the traditional Sijunzi decoction, is a traditional Chinese medicine compound that has been clinically proven to have anti-hepatocellular carcinoma (HCC) effects. According to our years of clinical application, JHD showed promising potential in enhancing the therapeutic effect of immune checkpoint inhibitors. However, despite its clinical promise, the immunoregulatory mechanisms by which JHD influences the tumor microenvironment (TME) remain largely unexplored.

[AIM OF THE STUDY] This study aimed to investigate how JHD modulates immune components within the TME and to elucidate its underlying mechanisms of immunosuppression in the context of HCC.

[MATERIALS AND METHODS] JHD was prepared as a spray-dried powder using a double decoction process and applied in both in vivo and in vitro experiments. Subcutaneous and orthotopic HCC models in mice were used to evaluate JHD's effects on immune modulation and tumor suppression. Transcriptomic sequencing, qRT-PCR, immunohistochemistry, and western blotting were employed to assess immune-related pathways. In vitro, the effects of JHD on HCC cell proliferation and molecular mechanisms were analyzed using co-culture systems, MTT and LDH assays, ELISA, qRT-PCR, and western blotting.

[RESULTS] Our results showed that JHD could inhibit the growth of liver cancer in immunocompetent mice, and enhanced immune function may be one of the mechanisms of JHD anti-liver cancer effect by comparative study with immunodeficient mice. Sequencing analysis further found its regulatory effect on DAP12 pathway. The results of bioinformatics analysis of HCC immunity reveal that increased infiltration of immunosuppressive tumor-associated macrophages (TAMs) and decreased natural killer (NK) cells activity are hallmark features of HCC. Notably, TREM1 is significantly upregulated during HCC progression and has emerged as a key regulator of the tumor immune microenvironment. Silencing TREM1 reduced the immunosuppressive phenotype of TAMs, enhanced NK cell-mediated cytotoxicity, and improved the antitumor immune response. JHD exerted its effects by modulating the TREM1/DAP12 signaling axis, thereby diminishing TAMs immunosuppression and promoting NK cell activity. Furthermore, JHD synergistically enhanced the therapeutic efficacy of anti-PD-1 antibodies in vivo, with a favorable safety profile.

[CONCLUSIONS] TAMs and NK cells play a pivotal role in driving immunosuppression in HCC. By targeting TREM1/DAP12 pathway, JHD improves immunosuppression TME and augments the antitumor efficacy of PD-1 inhibitors.