Potential for Protein Kinase Pharmacological Regulation in Flaviviridae Infections

Protein kinases (PKs) are enzymes that catalyze the transfer of the terminal phosphate group from ATP to a protein acceptor, mainly to serine, threonine, and tyrosine residues. PK catalyzed phosphorylation is critical to the regulation of cellular signaling pathways that affect crucial cell processes, such as growth, differentiation, and metabolism. PKs represent attractive targets for drugs against a wide spectrum of diseases, including viral infections. Two different approaches are being applied in the search for antivirals: compounds directed against viral targets (direct-acting antivirals, DAAs), or against cellular components essential for the viral life cycle (host-directed antivirals, HDAs). One of the main drawbacks of DAAs is the rapid emergence of drug-resistant viruses. In contrast, HDAs present a higher barrier to resistance development. This work reviews the use of chemicals that target cellular PKs as HDAs against virus of the Flaviviridae family (Flavivirus and Hepacivirus), thus being potentially valuable therapeutic targets in the control of these pathogens.

Nucleoside diphosphate and triphosphate prodrugs – An unsolvable task?

In this review, our recent advances in the development of nucleoside di- and nucleoside triphosphate prodrugs is summarized. Previously, we had developed a successful membrane-permeable pronucleotide system for the intracellular delivery of nucleoside monophosphates as well, the so-called cycloSal-approach. In contrast to that work in which the delivery is initiated by a chemically driven hydrolysis reaction, for the di- and triphosphate delivery, an enzymatic trigger mechanism involving (carboxy)esterases had to be used. The other features of the new pronucleotide approaches are: (i) lipophilic modification was restricted to the terminal phosphate group leaving charges at the internal phosphate moieties and (ii) appropriate lipophilicity is introduced by long aliphatic residues within the bipartite prodrug moiety. The conceptional design of the di- and triphosphate prodrug systems will be described and the chemical synthesis, the hydrolysis properties, a structure–activity relationship and antiviral activity data will be discussed as well. The advantage of these new approaches is that all phosphorylation steps from the nucleoside analogue into the bioactive nucleoside triphosphate form can be bypassed in the case of the triphosphate prodrugs. Moreover, enzymatic processes like the deamination of nucleosides or nucleoside monophosphates which lead to catabolic clearance of the potential antivirally active compound can be avoided by the delivery of the higher phosphorylated nucleotides.

Terminal phosphate group influence on DNA - TiO2 nanoparticle interactions

Immobilization of DNA/RNA, onto various metal and metal oxide surfaces is of great importance for the development of future microarray, gene mapping, DNA sequencing, nanoparticle targeting, and sensor applications. Attachment of DNA to solid interfaces typically occurs through either electrostatic interactions or covalent bonds to functional groups introduced to nucleic acid termini. Previously, we and others have demonstrated that alkanephosphates and terminal phosphate groups present on nucleic acids play an important role in the interaction with group IV metal oxides such as zirconium and hafnium, providing a stable linkage to the surface. Titanium dioxide (TiO2), which is frequently employed in various nanoscale applications, belongs to the same group and similar interactions with phosphate are expected. Various adsorption studies have demonstrated binding of nucleic acids to TiO2 surfaces, although the influence of terminal phosphate versus electrostatic interaction (via the DNA/RNA backbone) on the surface interaction is unclear. The research presented here investigates the effect of nucleic acid length, presence of terminal phosphates, and differences between dsDNA and ssDNA on their binding to TiO2 nanoparticles. TiO2 nanoparticles (20 nm) were used to study the adsorption of Lambda DNA (˜48 kbp), and shorter (21 bp) ssDNA and dsDNA oligonucleotides with and without a 5’ phosphate group. Initial adsorption of DNA to nanoparticles was calculated via UV absorption. Results showed that all types of nucleic acids (Lamda DNA, ssDNA and dsDNA) initially bind to nanoparticles, independent of molecular weight single/double strandedness, or phosphorylation state. The total amount of DNA initially adsorbed to nanoparticles (ng/particle) differs between ssDNA and dsDNA, as well as the length of the DNA used. These results show that nucleic acid interactions with TiO2 nanoparticles are not dependent upon the presence of a terminal phosphate group. These results provide valuable data for future applications based on DNA-nanoparticle constructs including nanoelectronics, photovoltaics, and biotemplated synthesis of semiconducting materials.

A New Linker for Solid-Phase Synthesis of Oligonucleotides with Terminal Phosphate Group

Synthesis of a novel cyanoethyl-type linker suitable for the solid-phase synthesis of oligodeoxynucleotides possessing terminal 3'-phosphate group is described. Since the linker is a 2-substituted 2-cyanoethanol, the release of the synthesized oligonucleotide from the solid support is accomplished by β-elimination in the ammonia deprotection step.