BACKGROUND: Hepatocellular carcinoma (HCC) poses ongoing difficulties for public health systems due to its high incidence and poor prognosis. Huqi formula (HQF), a well-known prescription in traditional Chinese medicine, has demonstrated notable clinical effectiveness in the treatment of HCC. However, the mechanisms underlying its therapeutic effects have yet to be completely elucidated. PURPOSE: This study aimed to investigate the anti-HCC effects of HQF and its underlying mechanisms. METHODS: Chemical profiling and quantification of HQF were conducted by LC-MS and HPLC. Orthotopic and subcutaneous tumor models were established through hydrodynamic injection of Akt/Nras plasmids and subcutaneous injection of c-Met/sgPten cells, respectively, to evaluate the therapeutic effects of HQF on HCC. Network pharmacology, RNA-Seq, molecular docking, Western blot, and flow cytometry were employed to assess the anti-HCC mechanisms. RESULTS: LC-MS analysis identified 41 components, with HPLC quantification showing salvianolic acid B as the most abundant compound (0.303%). In Akt/Nras and c-Met/sgPten-induced HCC models, HQF significantly reduced tissue damage, improved liver function, and inhibited HCC progression. Mechanistic studies revealed that HQF induced apoptosis in HCC cells by downregulating p-PI3K, p-AKT, and p-mTOR expression, with molecular docking indicating the strongest binding affinity between salvianolic acid B and PI3K. HQF further enhanced CD4 CONCLUSION: HQF demonstrated significant anti-HCC effects and a synergistic effect with PD-1, could be used as an alternative therapeutic agent for HCC.