Wang_2026_Phytomedicine_157_158302

BACKGROUND: Chronic Obstructive Pulmonary Disease (COPD) remains a major global health challenge due to the lack of therapies that modify its disease progression. Qingfei Tongluo Formula (QFTLF), a classic TCM formula, has shown efficacy in COPD. However, its specific molecular mechanisms and active compounds remain undefined. PURPOSE: This study aimed to decipher the multi-target mechanism of QFTLF against COPD, and to systematically identify its key bioactive serum compounds. METHODS: Based on pharmacodynamic evaluation in a COPD rat model and UPLC-MS/MS-based identification of QFTLF prototype serum compounds, we performed integrative multi-modal analysis, combining transcriptomics, proteomics, PSV2Nbased complex network, and singlecell RNA sequencing, to identify core targets, pathways and cell types regulated by prototype serum compounds. Key findings were further validated through in vivo and in vitro experiments. To identify key bioactive serum compounds, we employed a computeraided drug design (CADD) pipeline, including induced fit docking, molecular dynamics simulations, and binding free energy calculations, with subsequent experimental confirmation via surface plasmon resonance (SPR). RESULTS: Pharmacodynamic evaluation demonstrated that QFTLF significantly improved pulmonary function, lung histopathology and systemic inflammation in COPD rats, with efficacy comparable to the classic therapy aminophylline. UPLC-MS/MS analysis identified 58 prototype serum compounds. Multi-modal analysis and verification revealed that QFTLF downregulates prostaglandin-endoperoxide synthase 2 (PTGS2) and soluble epoxide hydrolase 2(EPHX2) while upregulating CYP2J2 in both COPD rat lung tissues and CSE-induced MLE-12 cells. This regulation restored arachidonic acid metabolic balance, increasing the anti-inflammatory and anti-oxidation metabolite 11,12-EET and decreasing the pro-inflammatory and pro-oxidation metabolite 11,12-DHET. Thereby, QFTLF suppressed downstream pathogenic indicators, including pro-inflammatory cytokine levels (TNF-alpha, IL-1beta, IL-6), lipid peroxidation indicators (reduced GSH and T-SOD, and elevated MDA), and ferroptosis indicators (Fe^2 accumulation, HMGB1 release, and GPX4 downregulation). This effect is mediated through inhibition of the PTGS2-PPARgamma-CYP2J2 and the EPHX2 mediated pathway. Notably, the protective profiles of specific PTGS2 and EPHX2 inhibitors in CSE-induced MLE-12 cells closely paralleled those of QFTLF, providing further support for the involvement of these targets. Furthermore, CADD and SPR identified and validated 7 key serum compounds with high binding affinity: 4 for PTGS2 (Mudanpioside C, Baicalin, Vitexin, Aloe-emodin-8-O-beta-d-glucopyranoside) and 5 for EPHX2 (Hamamelitannin, Choerospondin, Vitexin, Nepitrin, Aloe-emodin-8-O-beta-d-glucopyranoside). CONCLUSION: Our results identify QFTLF as a novel naturally occurring dual inhibitor of PTGS2/EPHX2, which attenuates COPD-associated ferroptosis in lung epithelial cells (the primary site of COPD injury) by restoring arachidonic acid metabolic balance, thereby addressing a critical unmet clinical need in COPD by positioning the QFTLF and its key compounds as promising disease modifying candidates.