Suppression of respiratory syncytial virus infection in vitro by small interfering RNAs complexed with bifunctional peptide carrier
AbstractIntroduction. Respiratory syncytial virus (RSV) is one of the most common pathogens affecting the upper and lower respiratory tract. One of the promising approaches to the creation of antiviral drugs is the use of small interfering RNA (siRNA) molecules that can specifically block the expression of virus genes. Another approach is the use of natural and synthetic peptides as antiviral agents. Notably, peptides can possess several biological properties. For example, we showed that the cationic dendrimeric peptide LTP not only exhibited antiviral properties against RSV, but was also able to transport nucleic acids into epithelial cells.
The aims of this study were to create a complex of bifunctional peptide LTP and siRNA molecules directed against vital RSV genes and to study its antiviral properties in vitro.
Material and methods. The optimal ratio of the component in the siRNA/peptide complex was determined by eukaryotic cell transfection. The toxic effect of the complex on cells was studied by the standard in vitro method (MTT-test). The antiviral properties of the complex were evaluated in the in vitro model of RSV infection. Viral load was assessed by titration on a monolayer of sensitive cells and by quantitative PCR analysis.
Results. We have shown that the optimal ratio of components in the siRNA/LTP complex is 1/12.5 by weight. With this ratio siRNA and peptide molecules form nanostructures with a diameter of 714 ± 125 nm, which have a positive charge of 16.9 ± 6.71 mV. Such physicochemical characteristics allow the complex to penetrate into cells. It was demonstrated that the complex and its individual components are non-cytotoxic in concentrations that have an antiviral effect. We showed that the siRNA/peptide complex provided a more pronounced suppression of virus replication compared to the free peptide. The enhanced antiviral effect of the complex is achieved due to the fact that the peptide has its own antiviral properties and additionally transports siRNA molecules blocking the replication of the virus genome into infected cells.
Conclusion. The complex of bifunctional LTP peptide and siRNA molecules directed against the vital RSV genes provides a more pronounced antiviral effect compared to peptide alone.
Keywords:respiratory syncytial virus; cationic peptides; antiviral peptides; transfection; siRNA; RNA-interference
For citation: Timotievich E.D., Shilovskiy I.P., Yumashev K.V., Kovchina V.I., Nikolskii A.A., Kaganova M.M., Rusak T.E., Andreev S.M., Smirnov V.V., Sergeev I.V., Maerle A.V., Kofiadi I.A., Shatilov A.A., Shatilova A.V., Kudlay D.A., Khaitov M.R. Suppression of respiratory syncytial virus infection in vitro by small interfering RNAs complexed with bifunctional peptide carrier. Immunologiya. 2023; 44 (5): 545–56. DOI: https://doi.org/10.33029/1816-2134-2023-44-5-545-556 (in Russian)
Funding. The study was supported by Russian Science Foundation grant No. 22-25-00182, https://rscf.ru/project/22-25-00182.
Conflict of interests. The authors declare no conflict of interests.
Authors contribution. Study conception and design – Shilovskiy I.P., Smirnov V.V., Khaitov M.R.; material collection and processing – Timotievich E.D., Nikolskii A.A., Kovchina V.I., Yumashev K.V., Kaganova M.M., Rusak T.E., Shatilov A.A., Shatilova A.V.; statistical processing – Andreev S.M., Kofiadi I.A.; manuscript preparation – Timotievich E.D., Shilovskiy I.P.; editing – Kudlay D.A., Sergeev I.V., Maerle A.V.
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