Influenza A virus is the cause of considerable human morbidity and mortality worldwide every year. There are antiviral small-molecular compounds affected the viral M2 ion channel or the viral enzyme neuraminidase. However, virus evolves very fast and becomes resistant to drugs. To solve this problem attempts are made to designed novel anti-influenza preparations. One type of these promising compounds is antisense oligonucleotides (AS ON). AS ONs are targeted to viral RNA: they bind directly to viral RNA and block either transcription of virus genome or expression of viral proteins, thus leading to inhibition of virus propagation.
Based on literature data we selected sequences of AS ONs PB1-AUG and NPv3’ targeting region of AUG codon of PB1 mRNA and 3’-end of viral NP RNA, respectively. Concentrations of AS ONs, multiplicity of infection (MOI) and time of incubation were varied to find optimal conditions to evaluate AS ONs effects on virus propagation. We found that at MOI 0.001 efficient inhibition of influenza virus A A/WSN/33 (H1N1) with non-modified AS ON delivered into MDCK cells using Lipofectamine 2000 was observed. Under these conditions AS ON causes significant decrease of virus titer (2-3 log10) in a concentration dependent manner.
To increase the efficiency of cellular accumulation of AS ONs TiO2-PL (TiO2 – titanium dioxide nanoparticles, PL – polylysin) composite was used. Early, it was shown by confocal microscopy that TiO2-PL composite deliver DNA into HeLa cells [1]. Flow cytometry show that this composite do deliver AS ONs and their analogs into MDCK cells.
1. Levina A.S., Ismagilov Z.R., Repkova M.N., Shikina N.V., Baiborodin S.I., Shatskaya N.V., Zagrebelnyi S.N., Zarytova V.F. Design of TiO2~DNA nanocomposites for penetration into cells// Russian Journal of Bioorganic Chemistry. 2013. V. 39. N 1. P. 77-86.