Modulation of gut microbiota and its metabolite Equol by Huaier granule suppresses hepatocellular carcinoma via the gut-liver axis

Hepatocellular carcinoma (HCC) is a frequently seen malignant tumor globally. Huaier is the dried fruiting body of the fungus Trametes robiniophila Murr. Huaier granule (HEG), formulated from the Huaier extract, is a Class I innovative anti-cancer drug in China and exhibits significant anti-HCC effects in clinical settings. Nevertheless, the specific mechanisms underlying its efficacy remain incompletely understood. This research demonstrated that HEG effectively suppressed tumor development in the orthotopic HCC mouse model in a gut microbiota-dependent manner and modified the gut microbiota composition. Notably, the primary differential bacterial genera between the Model group and the HEG group included Adlercreutzia. HEG exerted anti-HCC effects by repairing the intestinal barrier, improving colon immunity, and ameliorating the immune microenvironment by suppressing the MAPK signaling pathway via the gut microbiota-gut-liver axis. By integrating 16S rRNA sequencing with metabolomics data, supplemented by literature mining and in vitro validation, Equol, produced by specific gut microbiota Adlercreutzia, was identified as a key metabolite through which HEG exerted its anti-HCC effects by modulating gut microbiota. Moreover, Equol was essential for the anti-HCC effects of HEG. Additionally, Equol ameliorated the immune microenvironment through inhibiting the MAPK signaling pathway, while concurrently inhibiting the growth of HCC cells by inducing the G0/G1 phase blockade through suppression of Cyclin E1-CDK2/Rb signaling pathway. This study provided a robust scientific foundation for the clinical use of HEG, with Equol emerging as a promising candidate for HCC treatment.