Triptolide Increases the Risk of Nephrotoxicity via CASP8: An Integrative Multi-Omics Bioinformatics Analysis

Context: Triptolide (TP) is a highly potent natural diterpenoid, but its clinical application is severely limited by pronounced nephrotoxicity. Understanding the precise molecular targets and dynamic mechanisms underlying TP-induced renal injury remains a major challenge. In this study, we established an integrative computational paradigm to decode this toxicological mechanism. Network screening identified 14 hub targets, and subsequent two-stage Mendelian randomization explicitly pinpointed CASP8 as the core causal risk factor for acute kidney injury. Single-cell RNA sequencing mapped pronounced CASP8 upregulation specifically within vulnerable proximal tubular cells rather than immune infiltrates. Molecular simulations demonstrated a strong binding affinity (− 8.6 kcal/mol) between TP and the CASP8 catalytic pocket. Extended trajectory analyses and energetic decompositions revealed that a highly stable hydrogen bond at the Glu396 residue drives the TP-CASP8 interaction, inducing profound allosteric conformational locking and disrupting the global dynamic network of the protein. Methods TP systemic toxicity was evaluated using ProTox 3.0 and ADMETlab 2.0. Two-stage Mendelian randomization utilized large-scale systemic eQTLs and kidney-specific cis-eQTLs. Spatial expression was mapped via single-cell RNA sequencing datasets. Molecular docking was conducted using AutoDock Vina 1.2.3. Classical explicit-solvent molecular dynamics simulations (500 ns) were performed using GROMACS 2024.2. The AMBER99SB-ILDN force field was applied for the CASP8 protein, and the General AMBER Force Field 2 (GAFF2) was used for the TP ligand. Trajectory analyses, including Principal Component Analysis (PCA), Free Energy Landscape (FEL), and Dynamic Cross-Correlation Matrix (DCCM), were executed to evaluate structural dynamics. Binding free energies and per-residue thermodynamic decompositions were calculated using the MM/PBSA method via gmx_MMPBSA 1.6.4.