PMC:7075884 / 24328-30254 JSONTXT

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{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/7075884","sourcedb":"PMC","sourceid":"7075884","source_url":"https://www.ncbi.nlm.nih.gov/pmc/7075884","text":"Discussion\nThe Ginkgolic acids C13:0, C15:1 and C17:1 are commercially available compounds of Ginkgo leaves. The antioxidative activity of GA has been reported to have multiple therapeutic effects including the treatment of cardiovascular disease, HIV infection, bacterial infections such as Escherichia coli and Staphylococcus aureus, and some tumors7,8,19,20. It has been suggested that GA may operate by several other pathways including: inhibition of fatty acid synthase10; non-specific SIRT inhibition11; activation of protein phosphatase type-2C12; suppression of STAT3 activation through induction of PTEN and SHP-1 Tyrosine Phosphatase21, and protection against Aβ-induced synaptic dysfunction in the hippocampus6.\nIn this study, we are the first to report the fusion inhibitory effect of GA on enveloped viruses representing the three classes of fusion proteins and the inhibition of a non-enveloped human adenovirus. We also report a potential secondary mechanism of action involving viral DNA and protein synthesis. Our results are consistent with previous reports of the inhibitory effect of GA on DNA and protein synthesis22. The mechanism of action through which GA affects DNA and protein synthesis is not yet understood. It may bind to the host cell receptors and activate different cell signaling pathways and/or cause cell cycle arrest, which may explain the inhibitory effect of GA on rapidly dividing cancer cells. It may also enter the cells and work directly on DNA and protein synthesis. Experiments to address these questions are ongoing.\nTo assess the effect of GA on infectious viruses, we performed GA dose-response experiments. The cells were treated with different GA concentrations ranging between 1 µM and 20 µM. We demonstrated a dose dependent effect of GA on HCMV, HSV-1, and ZIKV. The effect of GA was tested in several cell types including HEp-2 (human epithelial carcinoma), 293T (human embryonic kidney), HFF and NHA (normal human astrocytes). GA was shown to have a viral inhibitory effect in all of the tested cells with no cytotoxicity within the active inhibitory range.\nBerg et al. evaluated the cytotoxicity and mutagenicity of GA in male Chinese hamster lung fibroblasts (V79 cells)23. Their results showed toxicity on cells grown in DMEM supplemented with 10% FBS after 24 hours in GA concentrations above 50 µM, with no mutagenic effect. Ahlemeyer et al. reported that 500 µM GA induced neuronal death and activated protein phosphatase type-2C in chick embryonic neurons growing in DMEM with 20% FBS12. B.M Hausen, evaluated the sensitizing capacity of GA in guinea pigs, determining 1000 ppm (2.886 mM) GA as safe to avoid inducing an allergic reaction24. Viral infections of permissive cells are regularly performed in 199 or MEM medium supplemented with 1% or 2% FBS. However, when we tested the activity and toxicity of GA on the cells used in our research, we also incubated the cells with GA in the cells’ recommended growth medium (see results and Fig. S1). The results indicated that the activity and toxicity of GA is affected by the serum concentration in the medium. We concluded that GA interacts with serum factors, which lowers its antiviral activity, and researchers using GA should address this issue in future experiments. In vivo experiments in an animal model are needed to assess the actual therapeutic antiviral effect and cytotoxicity of GA.\nGA’s universal inhibition of viral protein-mediated cell-cell fusion indicates that its inhibitory effect is by a common fusion mechanism. LPC also universally blocks membrane fusion; it does so by conferring spontaneous positive curvature, which prevents hemifusion. This block can be relieved, regardless of fusion protein, by the addition of the negative spontaneous curvature agent OA25,26. The finding that OA relieves the GA-induced inhibition of EBOV GP-mediated fusion implies that, similar to LPC, GA acts by producing positive spontaneous curvature and this prevents hemifusion. A number of rigid amphipathic fusion inhibitors (RAFI) with positive spontaneous curvature have been shown to inhibit fusion induced by unrelated viral fusion proteins27. In the future, it would be interesting to measure the values of the spontaneous curvatures of GA and RAFI and to relate them to the concentration of OA necessary to relieve fusion inhibition. For the inhibition of non-enveloped adenovirus, we suggest that since GA affects lipid bilayer curvature, it would be predicted to affect the endocytic entry of a non-enveloped virus such as adenovirus. In addition, as we report here, GA appears to have potential secondary mechanisms of viral DNA and protein synthesis inhibition, and these would be predicted to be targeted in both enveloped and non-enveloped viruses.\nIn conclusion, we have shown a consistent inhibitory effect of GA on the fusion of a variety of enveloped viruses, including important pathogens such as EBOV, HIV, ZIKA, HSV-1, HCMV, EBV and IAV. We also have shown inhibition of a non-enveloped human adenovirus, which suggests a potential inhibitory effect on other non-enveloped viruses. Furthermore, we found that GA might possibly inhibit HCMV viral DNA and HSV-1 protein synthesis by a secondary mechanism.\nThus, in light of the antiviral effect of GA on established viral infections of permissive cells, GA potentially could be used to treat acute viral infections (e.g. Coronavirus (COVID-19), EBOV, ZIKV, IAV and measles), and it might be determined to be useful in topical application for the successful treatment of active lesions (e.g. HSV-1, HSV-2 and VZV). Finally, our approach for GA usage to inhibit enveloped virus infection is fundamentally different from traditional microbicidal strategies that target virus genome replication. We anticipate that it could complement other direct antiviral agents and offer a new class of inhibitors of enveloped and non-enveloped 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