Nucleotides as Antiviral Compounds: on the Feasibility of an Esterase-dependent Prodrug Strategy for 2-5a

UNIVERSITY OF TURKU Department of Chemistry/ Faculty of Mathematics and Natural Sciences KIURU, EMILIA: Nucleotides as Antiviral Compounds: On the Feasibility of an Esterase-dependent Prodrug Strategy for 2-5A Doctoral thesis, 172 p. Laboratory of Organic Chemistry and Chemical Biology June 2015 Analogues of nucleotides and oligonucleotides have a significant role in the antiviral therapy. These compounds may inhibit specifically viral proteins or promote the innate immune system, where short 2 ́,5 ́-linked oligomers, called 25A, are the key players. The efficiency of nucleotide and oligonucleotide based drugs is largely dependent on the prodrug strategy used to enhance their cellular uptake. A common prodrug strategy includes masking of the negatively charged phosphate groups with biodegradable lipophilic protecting groups in order to facilitate permeation of the molecule through cell membrane. Inside the cell, upon unmasking by cellular enzymes, such as esterases, the prodrug is transformed into active drug. In the present thesis, the feasibility of an esterase-triggered prodrug strategy for 2-5A trimer has been evaluated by synthesizing two different 2-5A prodrug candidates and studying the release of 2-5A by carboxyesterase. The protecting group scheme was based on esterase-labile 2,2-disubstituted acyloxypropyl groups for the phosphate protection and acyloxymethyl groups for the 3 ́hydroxyl protection. The results demonstrated that deprotection of 2-5A, bearing esterase-labile protecting groups, became significantly slower upon accumulation of negative charge. Additionally, decomposition of the protecting groups produced electrophilic alkylating agents, which have been associated with potential toxicity. For these reasons, six different 2,2-disubstituted 4-acylthio-3oxobutyl groups were developed as protecting groups for phosphodiesters. These groups are cleaved by esterases, but in addition they exhibit a novel feature being removed thermally, which is an advantage when the affinity of the enzyme to a negatively charged substrate is reduced. The hydrolytic and enzymatic stability of the protecting groups is easily adjustable to optimize the deprotection rate. The released protecting groups are not markedly alkylating, since no alkylation with glutathione was observed.