Abstract
PURPOSE: Ursolic Acid (UA), a triterpenoid extracted from Hedyotis Diffusa Willd. (HDW), is known for its anti-inflammatory, antioxidant, and antitumor effects. Nevertheless, the mechanisms underlying UA's anti-colorectal cancer (CRC) effects remain insufficiently understood. This study aimed to identify the key target proteins of UA and investigate their functions in CRC development. METHODS: The cytotoxicity of three active components of HDW (UA, oleanolic acid (OA), and quercetin) on CRC cells was evaluated using the CCK-8 assay. Tandem mass tag (TMT)-based proteomics was employed to detect differentially expressed proteins (DEPs) in CRC cells after UA treatment. Bioinformatics analysis and high-content screening were used to identify UA's key protein targets. The expression and role of RPLP1 in CRC cells were investigated, including the effects of RPLP1 knockdown and its combination with UA treatment on cell proliferation, migration, invasion, and apoptosis. RESULTS: UA demonstrated superior inhibitory effects on CRC cells compared to OA and quercetin, highlighting it as a principal active ingredient of HDW. TMT-based proteomic analysis identified 438 upregulated and 366 downregulated proteins after UA intervention. Among these, RPLP1 was identified as a critical target. UA inhibited RPLP1 expression in CRC cells, resulting in decreased cell proliferation, migration, and invasion. Furthermore, the combination of UA treatment and RPLP1 knockdown exhibited synergistic effects in inhibiting CRC cell growth and migration, as well as promoting apoptosis. CONCLUSIONS: UA, a bioactive triterpenoid of HDW, inhibits CRC development by targeting and suppressing RPLP1 expression. These findings provide novel insights into the therapeutic of UA for CRC and highlight RPLP1 as a promising target for intervention.