Network Pharmacology and In vitro Experimental Verification to Explore the Mechanism of Chaiqin Qingning Capsule in the Treatment of Pain.

BACKGROUND: Chaiqin Qingning capsule (CQQNC) has been used to relieve pain in practice. However, the active components, pain targets, and molecular mechanisms for pain control are unclear. OBJECTIVE: To explore the active components and potential mechanisms of the analgesic effect of CQQNC through network pharmacology and in vitro experiments. METHODS: The main active components and the corresponding targets of CQQNC were screened from the TCMSP and the SwissTargetPrediction databases. Pain-related targets were selected in the OMIM, Gene- Cards, and DrugBank databases. These targets were intersected to obtain potential analgesic targets. The analgesic targets were imported into the STRING and DAVID databases for protein-protein interaction (PPI), gene ontology (GO) function enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Cytoscape software (V3.7.1) was used to construct an active component-intersection network. Finally, the key components were docked with the core targets. The analgesic mechanism of CQQNC was verified by RAW264.7 cell experiment. RESULTS: 30 active CQQNC components, 617 corresponding targets, and 3,214 pain-related target genes were found. The main active components were quercetin, kaempferol, and chenodeoxycholic acid etc. The key targets were ALB, AKT1, TNF, IL6, TP53, IL1B, and SRC. CQQNC can exert an analgesic effect through PI3K-Akt, MAPK signaling pathways, etc. Molecular docking showed that these active components had good binding activities with key targets. The results of in vitro experiments showed that CQQNC could exert antiinflammatory and analgesic effects through MAPK/AKT/NF-kB signaling pathways. CONCLUSION: CQQNC exerts pain control through inhibiting MAPK/AKT/NF-kB signaling pathways.

Network Pharmacological Analysis and Experimental Study of the Antipharyngitis Mechanism of the Chaiqin Qingning Capsule.

Objective: The study aimed to explore the active composition and mechanism of the Chaiqin Qingning capsule (CQQN) against pharyngitis based on the network pharmacology and through using a pharyngitis rat model. Methods: The active ingredients and targets of CQQN were queried using the TCMSP database. Disease-related target genes were queried in the DrugBank, GeneCards, OMIM, and DisGeNEt databases using "pharyngitis" as the search term. The STRING database was used to establish a protein-protein interaction (PPI) network. GO function enrichment and KEGG pathway enrichment analyses were performed to identify active components and key targets. Cytoscape software (version 3.7.2) was used to construct an active component/target gene/enrichment pathway network. AutoDock software was used to select the best binding target for molecular docking. The effect of CQQN was verified in the pharyngitis rats. Results: Network pharmacology showed 30 active compounds in CQQN with 240 targets, including 54 for the treatment of pharyngitis. Potential active ingredients included quercetin, kaempferol, stigmasterol, saikosaponin D, and isorhamnetin. The key targets involved were AKT1, TNF, IL-6, and IL-1ß. Signaling pathways included virus infection, TNF, IL-17, and cancer pathways. The molecular docking results showed that the critical components in CQQN had good potential for binding to key target genes. Animal experiments showed that CQQN could significantly reduce the expression of TNF-α, IL-1ß, IL-6, and IL-17 in the serum of rats with pharyngitis (P < 0.05). Hematoxylin and eosin staining showed that the inflammatory state of pharyngeal tissue in rats was significantly reduced compared to that in the model group. Conclusion: CQQN can improve pharyngitis by regulating the TNF and IL-17 signaling pathways. The study makes a positive exploration and provides a new idea for a more comprehensive and in-depth excavation of CQQN with an intervention effect on pharyngitis and other upper respiratory diseases in the future.

Cholesterol promotes EGFR-TKIs resistance in NSCLC by inducing EGFR/Src/Erk/SP1 signaling-mediated ERRα re-expression.

BACKGROUND: The use of epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) brings remarkable benefits for the survival of patients with advanced NSCLC harboring EGFR mutations. Unfortunately, acquired resistance seems to be inevitable and limits the application of EGFR-TKIs in clinical practice. This study reported a common molecular mechanism sustaining resistance and potential treatment options to overcome EGFR-TKIs resistance. METHODS: EGFR-TKIs resistant NSCLC cells were established and confirmed by MTT assay. Cholesterol content was detected and the promotional function of cholesterol on NSCLC growth was determined in vivo. Then, we identified ERRα expression as the downstream factor of cholesterol-mediated drug resistance. To dissect the regulatory mechanism, we conducted experiments, including immunofluorescence, co-immunoprecipitation, luciferase reporter assay and chromatin immunoprecipitation assay. RESULTS: Long-term exposure to EGFR-TKIs generate drug resistance with the characteristic of cholesterol accumulation in lipid rafts, which promotes EGFR and Src to interact and lead EGFR/Src/Erk signaling reactivation-mediated SP1 nuclear translocation and ERRα re-expression. Further investigation identifies ERRα as a target gene of SP1. Functionally, re-expression of ERRα sustains cell proliferation by regulating ROS detoxification process. Lovastatin, a drug used to decrease cholesterol level, and XCT790, an inverse agonist of ERRα, overcome gefitinib and osimertinib resistance both in vitro and in vivo. CONCLUSIONS: Our study indicates that cholesterol/EGFR/Src/Erk/SP1 axis-induced ERRα re-expression promotes survival of gefitinib and osimertinib-resistant cancer cells. Besides, we demonstrate the potential of lowing cholesterol and downregulation of ERRα as effective adjuvant treatment of NSCLC.