Abstract
Influenza virus (IV) infections cause severe respiratory illnesses that can be complicated by bacterial super-infections. Previously, we identified the cellular Raf-MEK-ERK cascade as a promising antiviral target. Inhibitors of MEK, such as CI-1040, showed potent antiviral activity. However, it remained unclear if this inhibitor and its active form, ATR-002, might sensitize host cells to either IV or secondary bacterial infections. To address these questions, we studied the anti-pathogen activity of ATR-002 in comparison to CI-1040, particularly, its impact on Staphylococcus aureus (S. aureus), which is a major cause of IV super-infections. We analysed IV and S. aureus titres in vitro during super-infection in the presence and absence of the drugs and characterized the direct impact of ATR-002 on bacterial growth and phenotypic changes. Importantly, neither CI-1040 nor ATR-002 treatment led to increased bacterial titres during super-infection, indicating that the drug does not sensitize cells for bacterial infection. In contrast, we rather observed reduced bacterial titres in presence of ATR-002. Surprisingly, ATR-002 also led to reduced bacterial growth in suspension cultures, reduced stress- and antibiotic tolerance without resistance induction. Our data identified for the first time that a particular MEK-inhibitor metabolite exhibits direct antibacterial activity, which is likely due to interference with the bacterial PknB kinase/Stp phosphatase signalling system.
Highlights
Influenza viruses cause infections of the respiratory tract resulting in severe diseases, especially in high-risk patients
After 30 min the virus dilution was removed, cells were rinsed with phosphate-buffered saline (PBS) and supplemented with invasion medium with or without S. aureus 6850 (6850) (MOI 0.1) in the presence of 10 μM CI-1040, ATR002 or solvent control. 3 h post bacterial infection cells were treated with lysostaphin (2 μg/mL) for 20 min to remove extracellular bacteria
Microscopic examination revealed that super-infection with both pathogens resulted in a highly increased cytopathic effect (CPE) compared to singular infections (Figure S1, upper panel)
Summary
Influenza viruses cause infections of the respiratory tract resulting in severe diseases, especially in high-risk patients. By directing novel antibacterial treatments towards inhibition of bacterial virulence factors expressed during infection, these compounds may exhibit a lower potential to induce resistance. Prior studies have shown that compounds inhibiting this pathway exhibit significant anti-influenza activity in vitro and in vivo[16,17]. The inhibition of this pathway by use of specific MEK-inhibitors like U0126 and CI-1040 resulted in a reduction of viral titres without induction of www.nature.com/scientificreports/. After 30 min the virus dilution was removed, cells were rinsed with PBS and supplemented with invasion medium with or without S. aureus 6850 (6850) (MOI 0.1) in the presence of 10 μM CI-1040, ATR002 or solvent control. The anti-pathogen activity of ATR-002 has not previously been tested
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