Target validation and drug discovery for TNBC: Targeting the Lin28B/Let-7/PBK with the ponicidin.

Triple-negative breast cancer (TNBC), characterized by high aggressiveness and limited targeted therapeutic options, urgently requires novel therapeutic targets and drugs. Although the Lin28/Let-7 axis has shown great therapeutic potential in refractory tumors, its role and specific regulatory mechanisms in TNBC remain unclear. This study first demonstrated through multi-dimensional bioinformatic analyses that Lin28B is preferentially overexpressed in TNBC, closely associated with poor patient prognosis, and exhibits a distinct expression pattern compared to other breast cancer subtypes, establishing its potential as a TNBC-specific therapeutic target. Subsequently, through tiered computational screening of a natural compound library, ponicidin was identified as a potential Lin28B inhibitor. Molecular docking, molecular dynamics simulations, and surface plasmon resonance (SPR) assays suggested that ponicidin binds to the cold-shock domain (CSD) of Lin28B with high affinity, which may competitively block the interaction between Lin28B and Let-7 and potentially promote Lin28B degradation via the ubiquitin-proteasome pathway, thereby relieving Let-7 suppression. Functional experiments demonstrated that ponicidin dose-dependently inhibits proliferation, invasion, and induces apoptosis in TNBC cells (MDA-MB-231, 4T1), with low toxicity to normal mammary epithelial cells. Through bioinformatic analyses including Weighted Gene Co-expression Network Analysis (WGCNA) and Protein-Protein Interaction (PPI) network analysis, PDZ-binding kinase (PBK) was predicted as a core downstream hub gene of the Lin28B/Let-7 axis. Experimental validation confirmed that ponicidin upregulates the expression of Let-7 family members, thereby downregulating multiple oncoproteins including PBK, C-MYC, RAS, and HMGA2. Further verification using a 4T1 cell orthotopic tumor mouse model showed that ponicidin dose-dependently inhibits in vivo tumor growth, reduces Lin28B expression in tumor tissues, increases apoptotic regions. This study is the first to tentatively explore the potential mechanism by which ponicidin inhibits TNBC by targeting the Lin28B/Let-7/PBK axis, clarifying its dual mode of action ("binding inhibition + protein degradation"). It provides a promising candidate compound with both efficacy and safety for TNBC, while laying a theoretical and experimental foundation for the development of Lin28B-targeted drugs.