Screening of
chemical libraries with 2,000 synthetic compounds identified
salinomycin as a hit against
influenza A and B viruses, with 50% effective concentrations ranging from 0.4 to 4.3 μM in cells. This compound is a carboxylic polyether
ionophore that exchanges monovalent
ions for
protons across
lipid bilayer membranes. Monitoring the time course of
viral infection showed that
salinomycin blocked nuclear migration of viral
nuclear protein (NP), the most abundant component of the viral
ribonucleoprotein (vRNP) complex. It caused cytoplasmic accumulation of NP, particularly within perinuclear endosomes, during virus entry. This was primarily associated with failure to acidify the endosomal-lysosomal compartments. Similar to the case with
amantadine (AMT),
proton channel activity of viral matrix
protein 2 (M2) was blocked by
salinomycin. Using purified retroviral Gag-based virus-like particles (VLPs) with M2, it was proved that
salinomycin directly affects the kinetics of a
proton influx into the particles but in a manner different from that of AMT. Notably,
oral administration of
salinomycin together with the
neuraminidase inhibitor
oseltamivir phosphate (OSV-P) led to enhanced
antiviral effect over that with either compound used alone in influenza A virus-infected mouse models. These results provide a new paradigm for developing
antivirals and their combination
therapy that control both host and viral factors.IMPORTANCE Influenza virus is a main cause of viral respiratory
infection in humans as well as animals, occasionally with high mortality. Circulation of influenza viruses resistant to the matrix
protein 2 (M2) inhibitor,
amantadine, is highly prevalent. Moreover, the frequency of detection of viruses resistant to the
neuraminidase inhibitors, including
oseltamivir phosphate (OSV-P) or
zanamivir, is also increasing. These issues highlight the need for discovery of new
antiviral agents with different mechanisms.
Salinomycin as the
monovalent cation-
proton antiporter exhibited consistent inhibitory effects against
influenza A and B viruses. It plays multifunctional roles by blocking endosomal acidification and by inactivating the
proton transport function of M2, the key steps for influenza virus uncoating. Notably,
salinomycin resulted in marked
therapeutic effects in influenza virus-infected mice when combined with OSV-P, suggesting that its chemical derivatives could be developed as an adjuvant
antiviral therapy to treat
influenza infections resistant or less sensitive to existing drugs.