Combined treatment of polygonatum and Scutellaria baicalensis suppresses lung cancer cell proliferation through inducing ferroptosis

Objective: To investigate the effects of combined treatment of Scutellaria baicalensis (SB) and polygonatum (HH) for lung cancer and explore the underlying mechanism. Purpose: The study was designed to investigate the anti-tumor effect of HH and reveal its mechanisms. Methods: Based on the Traditional Chinese Medicine (TCM) theory and network pharmacological analysis. The inhibitory effect of HH on the proliferation of A549 and LLC lung cancer cells was predicted by CCK8 assay in a subcutaneous tumor model in vivo. Tumor tissues were collected for proteomics, qPCR and immunohistochemistry assays. A549 and LLC cell proliferation was detected by CCK8 assay after treatment with HH combined with ferroptosis inhibitors and ferroptosis inducer Erastin for 48 h respectively. The expression of Transferrin Receptor (TFRC) was detected by qPCR and Western Blot. Changes in free iron content and lipid peroxide in cytoplasm and mitochondria were observed by laser confocal microscopy. Changes in Mitochondrial Membrane Potential (MMP, Δψm) and ROS content were detected by flow cytometry. Results: Search of Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) database demonstrated that HH played a role in lung cancer treatment. CCK8 assay showed that lung cancer cell proliferation was inhibited by HH in a dose-dependent manner. The animal experiment results showed that the tumor volume in HH-treated group was significantly smaller than that in Control group in the LLC allograft model (P<0.01). The expression level of TFRC in HH-treated group was higher than that in Control group, which was verified by proteomics, qPCR assay and immunohistochemistry. Two ferroptosis specific inhibitors could block HH-induced ferroptosis (P<0.05). Compared with Control group, free iron and lipid peroxide contents were significantly increased in cytoplasm and mitochondria while MMP was decreased and the level of ROS was augmented in HH group (P<0.01). Conclusion: HH increased the content of free iron and lipid peroxide in cytoplasm and mitochondria by upregulating the expression of TFRC in lung cancer cells, which further led to the accumulation of lipid peroxide, caused mitochondrial dysfunction, and finally induced ferroptosis in lung cancer cells.