Cell-free supernatant of <i>Clostridium leptum</i> inhibits breast cancer cell proliferation

Breast cancer has emerged as the leading cause of global cancer incidence, surpassing lung cancer. Accumulating evidence suggests that probiotics exhibit inhibitory effect on breast cancer progression, highlighting the need to identify gut flora-derived probiotics with potential anti-breast cancer properties. Here, we investigated the effect of the cell-free supernatant of Clostridium leptum (ClCFS) on breast cancer cells by methyl thiazolyl tetrazolium (MTT) assay. Untargeted metabolomics analysis was employed to characterize metabolite alterations in ClCFS. Furthermore, the core targets were predicted by the protein-protein interaction network and the signaling pathways were enriched by the Kyoto Encyclopedia of Genes and Genomes analysis. Our findings demonstrated that ClCFS inhibited the proliferation of breast cancer cells and that the main metabolite of ClCFS might be acetylcarnitine. Utilizing network pharmacological analysis, we identified apoptosis-related signaling pathways as the principal mechanisms underlying ClCFS activity. Furthermore, five core targets of STAT3, IL-1β, BCL2, CASP3, and ESR1 were identified. This study elucidates the main bioactive constituent and the potential targets of ClCFS against breast cancer. It provides a new understanding of the pharmacological activity of ClCFS in breast cancer treatment.