Antiviral Activity of Low-Molecular-Weight Fluorinated Compounds Against Influenza A (H1N1) Virus

Abstract

Influenza A virus takes a stable position in annual acute respiratory disease outbreaks during the autumn-winter period. Antiviral drugs for influenza infection help significantly ease the course of the disease and prevent complications, but there is still no drug to which none of the circulating strains would be resistant. Therefore, the search for new effective drugs against influenza A virus is constantly being conducted, in particular among fluorine-containing organic compounds, taking into account the special properties of fluorine. The goal of the study is to investigate the activity of a group of low-molecular-weight fluorinated compounds against the influenza A (H1N1) virus and to determine the potential mechanism of their action using in silico methods. Methods. Five fluorinated compounds designated as 10S-45, 10S-46, 10S-47, 10S-48 and 10S-49 were studied. In vitro experiments were performed using MDCK cell culture, A/FM/1/47 strain of influenza A (H1N1) virus, and oseltamivir phosphate as a reference drug. The cytotoxic effect on cell culture was determined using the MTT test. Antiviral effect was investigated by post-exposure incubation of compound solutions with cells, and visualization of results was performed using crystal violet dye. The possibility of the interaction of compounds with the cap-binding domain (CBD) of the PB2 subunit of the RNA-dependent RNA polymerase (RdRp) of the virus was evaluated using molecular dynamics simulation. Results. It was established that the studied compounds have a moderate cytotoxic effect on MDCK cell culture: the calculated CC50 values are in the range of 471─577 μg/mL for 10S-45, 10S-46, 10S-47 and 10S-48. Substance 10S-49 is almost 2 times less toxic compared to the rest, but it also has no effect on the influenza virus. Compounds 10S-45, 10S-46 and 10S-48 demonstrate antiviral activity in the range of 12─78 % inhibition of viral reproduction with an inverse dose-dependent effect. Substance 10S-47 showed a relatively stable inhibition of viral reproduction in the range of 63─69 % at all tested concentrations, which is similar to the results of the reference drug. According to the results of the selectivity index calculation, compound 10S-48 has the highest value, and 10S-47 has the lowest one, for the rest of the substances, this indicator is acceptable, but low, compared to the reference drug. For in silico studies, all possible spatial forms of the compounds presented as a mixture of diastereoisomers were generated: 4 stereoisomers each for 10S-45 and 10S-46, as well as 3 for 10S-47. According to the results of molecular docking of substances to the active center of CBD, 10S-45 has the lowest calculated affinity, and 10S-47 has the highest. Accordingly, compound 10S-45 showed no tendency to form ordered interactions with CBD. The 10S-47/CBD complex was not characterized by the presence of stable orientational bonds between receptor and ligand, similar to 10S-48/CBD. However, compound 10S-46 showed a generally stable interaction with CBD, albeit with a period of conformational instability. Conclusions. All studied compounds (except for 10S-49) demonstrate an antiviral effect in vitro against the influenza A (H1N1) virus. Based on the results of the in silico analysis of their interactions with CBD PB2, compound 10S-46 is highly likely to be an inhibitor of the cap-binding activity of RdRp.