In this research, we have established a drug screening method based on the autophagy signal pathway using the bimolecular fluorescence complementation - fluorescence resonance energy transfer (BiFC-FRET) technique to develop novel anti-influenza A virus (IAV) drugs. We selected Evodia rutaecarpa Benth out of 83 examples of traditional Chinese medicine and explored the mechanisms of evodiamine, the major active component of Evodia rutaecarpa Benth, on anti-IAV activity. Our results showed that evodiamine could significantly inhibit IAV replication, as determined by a plaque inhibition assay, an IAV vRNA promoter luciferase reporter assay and the Sulforhodamine B method using cytopathic effect (CPE) reduction. Additionally, evodiamine could significantly inhibit the accumulation of LC3-II and p62, and the dot-like aggregation of EGFP-LC3. This compound also inhibited the formation of the Atg5-Atg12/Atg16 heterotrimer, the expressions of Atg5, Atg7 and Atg12, and the cytokine release of TNF-α, IL-1β, IL-6 and IL-8 after IAV infection. Evodiamine inhibited IAV-induced autophagy was also dependent on its action on the AMPK/TSC2/mTOR signal pathway. In conclusion, we have established a new drug screening method, and selected evodiamine as a promising anti-IAV compound.
Lei H, Sheng Z, Ding Q, Chen J, Wei X, et al. (2011) Evaluation of Oral Immunization with Recombinant Avian Influenza Virus HA1 Displayed on the Lactococcus lactis Surface and Combined with the Mucosal Adjuvant Cholera Toxin Subunit B. Clin Vaccine Immunol. 18: 1046–1051.
Meng S, Liu Z, Xu L, Li L, Mei S, et al. (2011) Intranasal immunization with recombinant HA and mast cell activator C48/80 elicits protective immunity against 2009 pandemic H1N1 influenza in mice. PLoS One 6: e19863.
Wang S, Hackett A, Jia N, Zhang C, Zhang L, et al. (2011) Polyvalent DNA vaccines expressing HA antigens of H5N1 influenza viruses with an optimized leader sequence elicit cross-protective antibody responses. PLoS One 6: e28757.
Lee H, Kim IK, Park TG (2010) Intracellular trafficking and unpacking of siRNA/quantum dot-PEI complexes modified with and without cell penetrating peptide: confocal and flow cytometric FRET analysis. Bioconjug Chem 21: 289–295.
Matarrese P, Nencioni L, Checconi P, Ciarlo L, Gambardella L, et al. (2011) Pepstatin A alters host cell autophagic machinery and leads to a decrease in influenza A virus production. J Cell Physiol 226: 3368–3377.
Su CY, Cheng TJ, Lin MI, Wang SY, Huang WI, et al. (2010) High-throughput identification of compounds targeting influenza RNA-dependent RNA polymerase activity. Proc Natl Acad Sci U S A 107: 19151–19156.
Noah JW, Severson W, Noah DL, Rasmussen L, White EL, et al. (2007) A cell-based luminescence assay is effective for high-throughput screening of potential influenza antivirals. Antiviral Res 73: 50–59.