Cell metabolomics study on synergistic anti‐hepatocellular carcinoma effect of Aidi injection combined with doxorubicin

Hepatocellular carcinoma (HCC) is the sixth most common cancer worldwide and the second most common cause of cancer deaths. This study aimed to explore the inhibitory effect and mechanism of Aidi injection (ADI) combined with doxorubicin (DOX) in HCC treatment. The drug concentrations in the combined therapy was determined by investigating the effect of various concentrations of ADI and DOX on the viability of H22 cells. The combination index was also calculated. A metabolomic strategy based on a ultrahigh-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) platform was established to analyze the metabolites. As a result, the combination index values were less than 1, indicating that the combination of ADI and DOX exerted a synergistic effect on HCC treatment. The combination of 40‰ ADI and 1 μmol/l DOX had the strongest inhibitory effect and was used for subsequent metabolomic analysis. A total of 19 metabolic markers were obtained in metabolomic analysis, including amino acids (l-glutamic acid, l-arginine and l-tyrosine), organic acids (succinic acid and citric acid), adenosine and hypoxanthine. Compared with the treatment using DOX or ADI alone, the combined therapy further regulated the levels of metabolic markers in HCC, which may be the reason for the synergistic effect. Seven metabolic pathways were significantly enriched, including phenylalanine, tyrosine and tryptophan biosynthesis, d-glutamine and d-glutamate metabolism, alanine, aspartate and glutamate metabolism, phenylalanine metabolism, arginine biosynthesis, the tricarboxylic acid cycle and purine metabolism. These findings demonstrated that ADI combined with DOX can effectively inhibit the viability of H22 cells, which may exert a synergistic antitumor effect by balancing the metabolism of amino acids and energy-related substances.