The cardenolide ouabain suppresses coronaviral replication via augmenting a Na+/K+-ATPase-dependent PI3K_PDK1 axis signaling

• Published in: Toxicology and applied pharmacology 2018-10, Vol.356, p.90-97
• Main Authors: Yang, Cheng-Wei, Chang, Hsin-Yu, Lee, Yue-Zhi, Hsu, Hsing-Yu, Lee, Shiow-Ju
• Format: Article
• Language: English
• Subjects: Animals; Antiviral Agents - pharmacology; Cardenolide; Cell Line; Chromones - pharmacology; Coronavirus; Coronavirus - drug effects; DNA Replication - drug effects; Dose-Response Relationship, Drug; Gene Silencing; Interleukin-6 - antagonists & inhibitors; Interleukin-6 - biosynthesis; Mice; Morpholines - pharmacology; Na+/K+-ATPase; Ouabain; Ouabain - pharmacology; PDK1; Phosphatidylinositol 3-Kinases - drug effects; Phosphoinositide-3 Kinase Inhibitors; PI3K; Protein-Serine-Threonine Kinases - biosynthesis; Protein-Serine-Threonine Kinases - genetics; Pyrimidines - pharmacology; Pyruvate Dehydrogenase Acetyl-Transferring Kinase; Signal Transduction - drug effects; Sodium-Potassium-Exchanging ATPase - drug effects; Thiophenes - pharmacology; Virus Replication - drug effects
• ISSN: 0041-008X; 1096-0333
• DOI: 10.1016/j.taap.2018.07.028

Cardenolides are plant-derived toxic substances. Their cytotoxicity and the underlying mechanistic signaling axes have been extensively documented, but only a few anti-viral activities of cardenolides and the associated signaling pathways have been reported. Previously, we reported that a variety of cardenolides impart anti-transmissible gastroenteritis coronavirus (TGEV) activity in swine testicular (ST) cells, through targeting of the cell membrane sodium/potassium pump, Na+/K+-ATPase. Herein, we further explore the potential signaling cascades associated with this anti-TGEV activity in ST cells. Ouabain, a representative cardenolide, was found to potently diminish TGEV titers and inhibit the TGEV-induced production of IL-6 in a dose dependent manner, with 50% inhibitory concentrations of 37 nM and 23 nM respectively. By pharmacological inhibition and gene silencing, we demonstrated that PI3K_PDK1_RSK2 signaling was induced in TGEV-infected ST cells, and ouabain imparted a degree of anti-TGEV activity via further augmentation of this existing PI3K_PDK1 axis signaling, in a manner dependent upon its association with the Na+/K+-ATPase. Finally, inhibition of PI3K by LY294002 or PDK1 by BX795 antagonized the anti-viral activity of ouabain and restored the TGEV virus titer and yields. This finding is the first report of a PI3K_PDK1 signaling axis further induced by ouabain and implicated in the suppression of TGEV activity and replication; greatly illuminates the underlying mechanism of cardenolide toxicity; and is expected to result in one or more anti-viral applications for the cardenolides in the future. [Display omitted] •Ouabain eliminated TGEV titers and inhibited viral replication.•Ouabain diminished TGEV induced IL-6 production.•Ouabain enhanced PI3K or PDK1 activation induced by TGEV via Na+/K+-ATPase.•PI3K or PDK1 inhibition antagonized the anti-TGEV activity of ouabain.•Ouabain augmented the PI3K_PDK1 axis signaling that inhibited TGEV activity.