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Antisense oligonucleotides and siRNAs are potential therapeutic agents and their chemical modifications play an important role to improve the properties and activities of oligonucleotides. Isonucleoside is a type of nucleoside analogue, in which the nucleobase is moved from C-1 to other positions of ribose. In this report, a novel isonucleoside 5 containing a 5'-CH(2)-extended chain at the sugar moiety was synthesized, thus isoadenosine 5a and isothymidine 5b were incorporated into a DNA single strand and siRNA. It was found that isonucleoside 5 modified oligonucleotides can form stable double helical structures with their complementary DNA and RNA and the stability towards nuclease and ability to activate RNase H are more promising compared with the unmodified, natural analogues. In siRNA, passenger strand modified with isonucleoside (5a/b) at 3' or 5' terminal can retain the silencing activity and minimize the passenger strand specific off-target effect.

Four types of beta-carboline-nucleoside conjugates were synthesized. The binding affinities of these beta-carboline-nucleoside conjugates , and to TAR RNA were evaluated by affinity capillary electrophoresis. The data of binding affinities to TAR RNA show that conjugates and are stronger binders than the parent compound . Computer modeling indicates that the beta-carboline-nucleoside conjugate can fit to the UCU three-nucleotide bulge region of TAR RNA.

Two types of neamine derivatives, neamine-dinucleotide conjugates 8a-g and neamine-PNA conjugates 12a-c and 14a-d, were synthesized. Compound 8a-g were synthesized by the condensation of azido-neamine with dinucleotide-5'-carboxylic acids, followed by reduction and deprotection. Compound 12a-c and 14a-d were synthesized by the similar strategy. The binding affinities of conjugates 8a-g, 12a-c, and 14a-d towards 16S RNA, 18S RNA, and TAR RNA were evaluated by SPR. It indicates that conjugates 12a-c and 14a-d interact with 16S, 18S RNA at the same level as that of neamine, 14a and 14d show about twofold binding affinities to TAR RNA compared to that of neamine. However, the neamine-dinucleotide conjugates 8a-g exhibit very weak binding affinities to 16S, 18S, and TAR RNA, computer modelling results that negative-negative electrostatic repulsion of phosphate group in compound 8a-g and RNA leads to a sharp decrease of the binding affinities compared with that of neamine, neamine-nucleoside and neamine-PNA conjugates.

A novel class of aminoisonucleoside was synthesized and incorporated into a luciferase gene-targeting siRNA. Structural and functional analyses of such a kind of siRNAs indicated that sense strand modifications with aminoisonucleoside at the 3' or 5' terminal, such as ssIso-1 and ssIso-2, have less effect on RNA duplex thermal and serum stabilities, and their functional activities are also comparable to their native siRNAs. In contrast, antisense strand modifications with aminoisonucleoside at the corresponding positions, such as asIso-2 or asIso-1, bring a striking negative effect on RNA duplex stability but still maintain around 40-50% of gene knockdown.

Two series of 6′-halovinyl-adenosine stereoisomers including 5′-ethylenic and acetylenic substituted l-adenosine, 5′-ethylenic and acetylenic substituted isonucleosides were synthesized. In the l-nucleoside series, compounds 6b, 8b, 10b and 13b showed modest inhibition of SAH hydrolase (21, 44, 50 and 26% respectively) at 100μM. The l-isomers of 5′-ethylenic and acetylenic modified isonucleoside 23, 24 exhibited no activity for the inhibition of SAH hydrolase, however, the d-isomers 30 and 31 showed some activities in the same test (35 and 21%). It indicated clearly the strict stereochemical requirement for the substrate of SAH hydrolase. Compounds 6b, 8b, 8c, 11b exhibited modest to good inhibition effects on the growth of HeLa cells or Bel-7420 cells at 1μM (64, 44, 53 and 82% respectively). [Copyright &y& Elsevier]

Abstract: Gemcitabine (GEM) is a nucleoside analog agent against a wide variety of tumors. To overcome its limitation of rapid metabolism in vivo that results in short circulation time and poor antitumor efficacy, a novel prodrug (CLA–GEM conjugate) has been developed through the covalent coupling of conjugated linoleic acid (CLA) to N4-amino group of GEM. The chemical structure of CLA–GEM conjugate was identified by NMR, FTIR and other methods. From in vitro tests, it was demonstrated that the linkage with CLA increased the plasma stability of GEM as well as the antitumor activity against human breast tumor cells (MCF-7). Importantly, it also altered the transport pattern of GEM across cell membrane (MCF-7 and MDA-MB-231), evidenced by the little effect of nucleoside transporter inhibitors (NBMPR and dipyridamole) on the IC50 values of CLA–GEM, instead of the great effect on that of unmodified GEM. In vivo pharmacokinetic study showed that the CLA–GEM conjugate had a longer plasma half-life and a higher bioavailability compared to that of unmodified GEM. Significant stronger antitumor activity was observed in the nude mice xenografted MCF-7 breast tumor after treated with CLA–GEM than that of unmodified GEM, while no significant body weight loss was found in all treatments. In conclusion, the novel CLA–GEM conjugate prepared in this study would be a promising prodrug of gemcitabine for future clinical use. [Copyright &y& Elsevier]

Abstract: Two types of aminodisaccharide–nucleoside conjugates were synthesized by the condensation of azidodisaccharide and nucleoside using aliphatic diamine as a linker. The corresponding azidodisaccharides could be yielded from neamine in good yield. The binding properties to 16S RNA of these conjugates were evaluated by SPR. It was found that the nucleobase played a significant role in the binding of these conjugates to 16S RNA and a shorter linker between the aminodisaccharide and nucleoside was favorable for 16S RNA binding. [Copyright &y& Elsevier]