Active constituents and mechanisms of Respiratory Detox Shot, a traditional Chinese medicine prescription, for COVID-19 control and prevention: Network-molecular docking-LC-MS analysis / 中西医结合学报

OBJECTIVE@#Lung-toxin Dispelling Formula No. 1, referred to as Respiratory Detox Shot (RDS), was developed based on a classical prescription of traditional Chinese medicine (TCM) and the theoretical understanding of herbal properties within TCM. Therapeutic benefits of using RDS for both disease control and prevention, in the effort to contain the coronavirus disease 2019 (COVID-19), have been shown. However, the biochemically active constituents of RDS and their mechanisms of action are still unclear. The goal of the present study is to clarify the material foundation and action mechanism of RDS.@*METHODS@#To conduct an analysis of RDS, an integrative analytical platform was constructed, including target prediction, protein-protein interaction (PPI) network, and cluster analysis; further, the hub genes involved in the disease-related pathways were identified, and the their corresponding compounds were used for in vitro validation of molecular docking predictions. The presence of these validated compounds was also measured in samples of the RDS formula to quantify the abundance of the biochemically active constituents. In our network pharmacological study, a total of 26 bioinformatic programs and databases were used, and six networks, covering the entire Zang-fu viscera, were constructed to comprehensively analyze the intricate connections among the compounds-targets-disease pathways-meridians of RDS.@*RESULTS@#For all 1071 known chemical constituents of the nine ingredients in RDS, identified from established TCM databases, 157 passed drug-likeness screening and led to 339 predicted targets in the constituent-target network. Forty-two hub genes with core regulatory effects were extracted from the PPI network, and 134 compounds and 29 crucial disease pathways were implicated in the target-constituent-disease network. Twelve disease pathways attributed to the Lung-Large Intestine meridians, with six and five attributed to the Kidney-Urinary Bladder and Stomach-Spleen meridians, respectively. One-hundred and eighteen candidate constituents showed a high binding affinity with SARS-coronavirus-2 3-chymotrypsin-like protease (3CL), as indicated by molecular docking using computational pattern recognition. The in vitro activity of 22 chemical constituents of RDS was validated using the 3CL inhibition assay. Finally, using liquid chromatography mass spectrometry in data-independent analysis mode, the presence of seven out of these 22 constituents was confirmed and validated in an aqueous decoction of RDS, using reference standards in both non-targeted and targeted approaches.@*CONCLUSION@#RDS acts primarily in the Lung-Large Intestine, Kidney-Urinary Bladder and Stomach-Spleen meridians, with other Zang-fu viscera strategically covered by all nine ingredients. In the context of TCM meridian theory, the multiple components and targets of RDS contribute to RDS's dual effects of health-strengthening and pathogen-eliminating. This results in general therapeutic effects for early COVID-19 control and prevention.

Prolactin Inhibits BCL6 Expression in Breast Cancer Cells through a MicroRNA-339-5p-Dependent Pathway / 한국유방암학회지

PURPOSE: Prolactin (PRL) plays a critical role in breast cancer progression by activating its cognate receptor and promotes the growth and differentiation of breast cancer cells. Studies have shown that B-cell lymphoma 6 (BCL6) is the target gene of microRNA-339-5p (miR-339-5p) and that BCL6 expression contributes to breast cancer progression. Herein, we identified PRL as a potent suppressor of BCL6 expression in human breast cancer cells. METHODS: Western blotting and quantitative reverse transcription-polymerase chain reaction were used to investigate molecular mechanisms underlying miR-339-5p expression and BCL6 manipulation in MCF-7, T47D, and SKBR3 breast cancer cells. Phenotypic changes in these breast cancer cell lines were assessed by performing cell viability (MTT), colony formation, migration, and invasion assays. RESULTS: PRL suppressed BCL6 protein and mRNA expression and upregulated miR-339-5p expression in MCF-7 and T47D breast cancer cells. Selective downregulation of miR-339-5p expression significantly reversed PRL-induced suppression of BCL6 mRNA and protein expression. Exogenous PRL stimulation significantly decreased the proliferation, colony formation, migration, and invasion of breast cancer cells, and suppression of miR-339-5p expression reversed these processes in vitro. CONCLUSION: These results indicated that PRL inhibited BCL6 expression and regulated breast cancer progression through a miR-339-5p-dependent pathway.