Shiyiwei Shenqi Pian-derived quercetin reactivates apoptosis in clear cell renal cell carcinoma by destabilizing survivin and x-linked inhibitor of apoptosis protein.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) strongly resists apoptosis via baculoviral inhibitor of apoptosis repeat-containing 5 (BIRC5) and X-linked inhibitor of apoptosis protein (XIAP). Meanwhile, the specific anti-ccRCC mechanisms of the pro-apoptotic herbal mixture Shiyiwei Shenqi Pian (SSP) remain undefined. PURPOSE: To systematically discover the key bioactive compound within SSP and characterize its precise mechanism of action in uncoupling the oncogenic BIRC5-XIAP complex. STUDY DESIGN: An advanced integrative strategy combining network pharmacology, transcriptomic profiling, machine-learning, and structural dynamic simulations was utilized to screen and predict therapeutic targets. The in silico findings were then rigorously and sequentially validated across multiple experimental platforms, progressing from in vitro cell models to in vivo CDX and PDX settings. METHODS: A multi-omics framework was applied to investigate the mechanisms of SSP in ccRCC. Active compounds and disease-related targets were identified through database mining and UPLC-Q-TOF-MS profiling, followed by machine-learning-based prioritization. Molecular interactions were analyzed using docking, molecular dynamics simulations, and single-cell transcriptomics. Protein interactions and stability were validated by co-immunoprecipitation (Co-IP), microscale thermophoresis (MST), cycloheximide (CHX)-chase, and ubiquitination assays. Cellular effects were assessed by proliferation and apoptosis assays, while in vivo efficacy was evaluated using CDX and PDX models. RESULTS: Shared SSP-ccRCC targets were enriched in apoptosis-related pathways, and intact SSP extract suppressed ccRCC cell proliferation and clonogenic growth in vitro. Multi-cohort machine-learning modeling identified BIRC5 as a key prognostic target. Among the major SSP-derived compounds, quercetin showed favorable binding to BIRC5 and XIAP and was selected for mechanistic validation. Single-cell and bulk transcriptomic analyses confirmed co-enrichment and positive correlation of BIRC5 and XIAP in ccRCC. Mechanistically, quercetin disrupted the BIRC5-XIAP complex, weakened their direct protein interaction, and accelerated ubiquitin-mediated degradation of both proteins. This process restored caspase activation and induced caspase-dependent apoptosis, leading to reduced cell proliferation and colony formation. In CDX and PDX models, quercetin suppressed tumor growth and reduced BIRC5/XIAP pathway activity. CONCLUSION: Quercetin was identified as a major mechanistically validated active constituent of SSP. It suppresses ccRCC growth by destabilizing the BIRC5-XIAP anti-apoptotic complex and reactivating caspase-dependent apoptosis, supporting its potential as a therapeutic lead for ccRCC.