[Composites of Peptide Nucleic Acids with Titanium Dioxide Nanoparticles. IV+. Antiviral Activity of Nanocomposites Containing DNA/PNA Duplexes].

Antiviral activity of TiO2 · PL · DNA/PNA nanobiocomposites was studied on the MDCK cell culture infected with influenza A virus (subtype H3N2). PNA fragment in nanocomposites as a DNA/PNA heteroduplex is electrostatically bound to titanium dioxide nanoparticles precovered with polylysine (TiO2 · PL). It was shown that TiO2 · PL · DNA1/PNA1 nanobiocomposit bearing PNA1 fragment targeted to the 3'-end of the noncoding region of segment 5 of viral RNA specifically inhibited the virus reproduction with the efficiency of 99.8%. It was determined that the 50% cytotoxic concentration (TC50) of the TiO2 · PL · DNA1/PNA1 nanocomposite is more than 1200 mg/mL. And 50% effective inhibitory concentration (IC50) is less than 0.003 mg/mL. Based on these data, the selectivity index (SI) for TiO2 · PL · DNA1/PNA1 nanobiocomposite defined as the ratio TC50/LC50, is more than 400. Thus TiO2 · PL · DNA/PNA nanobiocomposites can not only penatrate through cell membrane, but and are able to exhibit a high specific antisense activity, without causing toxic effects on the living cells.

[Composites of peptide nucleic acids with titanium dioxide nanoparticles. III. Kinetics of PNA dissociation from nanocomposites containing DNA/PNA duplexes].

When delivering peptide nucleic acids (PNA) to the cells in the nanocomposites TiO2 · PL · DNA/PNA, containing titanium dioxide nanoparticles coated with polylysine (PL) and immobilized DNA/PNA duplexes, it is important not only to transport them to the cell, but also ability to control the release rate of the PNA-drug from the carrier. PNA desorption from TiO2 · PL · DNA/PNA nanocomposite in time has been shown. Desorption is caused by dissociation of immobilized DNA/PNA duplex while the DNA remains on the carrier and PNA goes away in solution. It has been found that the half-retention times of PNA on TiO2 · PL · DNA/PNA nanocomposites containing DNA/PNA duplexes with overlapping complementary base pairs equal to 10, 12, 14, and 16 are 10, 14, 22 and 70 minutes, respectively. Thus, it has been shown that the release rate of the PNA-drug from nanocomposites can be adjusted by varying the overlap of complementary base pairs in the immobilized DNA/PNA duplex. This method of PNA immobilization may be used for designing of nanocomposites with optimum release time of the PNA-drugs. Created TiO2 · PL · DNA/PNA nanocomposites can be used to efficiently deliver therapeutically significant drug PNA and their selective effect on the pathogenic nucleic acid in the cell.

[Affinity capture of specific DNA fragments with the use of short synthetic sequences].

The ability of short peptide nucleic acid (PNA) oligomers and oligonucleotides containing modified residues of 5-methylcitidine, 2-aminoadenosine and 5-propynyl-2'-deoxyuridine (strong binding oligonucleotides, SBO) to affinity capture the target double-stranded DNA fragment from mixture by means of the end invasion was compared. Both types of probes were highly effective at the conditions used. The SBO-based probes may represent a handy and easily prepared alternative to PNA for selection of target DNA fragments from mixtures.

[Composites of peptide nucleic acids with titanium dioxide nanoparticles. II+. Dissociation of DNA/PNA duplexes within TiO2 x polylysine x DNA/PNA nanocomposites and in solution. Effect of polylysine].

When creating effective drugs it is important not only to transport them into the cells, but also important to have the possibility of release them from the "transporter" after delivery into the cell. It was shown that peptide nucleic acids (PNA) in nanocomposite TiO2 x PL x DNA/PNA dissociate with typical shape of the thermal denaturation curve, and polylysine (PL) in the nanocomposite has practically no effect on the dissociation of the DNA/PNA duplexes. These data suggest that the PNA in the nanocomposite TiO2 x PL x DNA/PNA have been immobilized reversible and able to dissociate and be released from TiO2-carrier into solution. In contrast that, the dissociation of DNA/DNA and DNA/PNA duplexes in physiological solution at the presence of PL--was not observed. PL in solution abnormally strong influences on the nature of the optical density dependence on temperature and time for D-duplexes and in a less degree--for P-duplexes. It has been suggested, that PL with DNA/DNA duplexes in physiological solution forms triple polycomplexes (-DNA/DNA x PL)m, consisting of several (m) chains of PL connected with DNA/DNA duplexes. And such polycomplexes able to aggregate and precipitate. PL in solution can interact with DNA/PNA duplexes to form monocomplexes PL x (DNA/PNA)n consisting of one chain PL and one or more (n) DNA/PNA duplexes that do not precipitate, however the dissociation of DNA/PNA duplexes from such monocomplexes is difficult.

[Composites of peptide nucleic acids with titanium dioxide nanoparticles. I. Construction of nanocomposites containing DNA/PNA duplexes and their delivery into HeLa cells].

In order to investigate the possibility of using titanium dioxide (TiO2) nanoparticles to transport peptide nucleic acids (PNA) in eukaryotic cells, a PNA oligomer has been synthesized, and method of PNA immobilization in the form of hybrid DNA/PNA duplexes on the surface of TiO2 nanoparticles covered with polylysine (PL) has been designed. Attaching of DNA/PNA duplex on TiO2 x PL nanoparticles occurred due to electrostatic interactions between the negatively charged DNA chain and the positively charged amino groups of PL. Binding of the PNA with the nanocomposite achieved through noncovalent Watson-Crick interactions between the PNA and complementary DNA. The capacity of obtained TiO2 x PL x DNA/PNA nanocomposites depending on immobilization conditions was 10-30 nmol PNA per 1 mg of TiO2 particles, which corresponds to -1-3 PNA molecules per one TiO2 particle with size of 4-6 nm. By method ofconfocal laser scanning microscopy on the example of the fluorescein labeled PNA oligomer (Flu)PNA it has been shown that the PNA molecules in composition of TiO2 x PL x DNA/(Flu)PNA nanocomposites effectively penetrate and accumulate in HeLa cells without the use oftransfection agents, electroporation, or other auxiliary procedures has been shown.