Impact of the Guanidinium Group on Hybridization and Cellular Uptake of Cationic Oligonucleotides

Abstract

The grafting of cationic groups to synthetic oligonucleotides (ONs) in order to reduce the charge repulsion between the negatively charged strands of a duplex or triplex, and consequently to increase a complex's stability, has been extensively studied. Guanidinium groups, which are highly basic and positively charged over a wide pH range, could be an efficient ON modification to enhance their affinity for nucleic acid targets and to improve cellular uptake. A straightforward post-synthesis method to convert amino functions attached to ONs (on sugar, nucleobase or backbone) into guanidinium tethers has been perfected. In comparison to amino groups, such cationic groups anchored to alpha-oligonucleotide phosphoramidate backbones play important roles in duplex stability, particularly with RNA targets. This high affinity could be explained by dual recognition resulting from Watson-Crick or Hoogsteen base pairing combined with cationic/anionic backbone recognition between strands involving H-bond formation and salt bridging. Molecular-dynamics simulations corroborate interactions between the cationic backbones of the alpha-ONs and the anionic backbones of the nucleic acid targets. Moreover, ONs with guanidinium modification increased cellular uptake relative to negatively charged ONs. The cellular localization of these new cationic phosphoramidate ONs is mainly cytoplasmic. The uptake of these ON analogues might occur through endocytosis.

The design of guanidinium-rich transporters and their internalization mechanisms

“…Lebleu and coworkers recently reported the cellular uptake of new fluorescently labeled guanidinylated DNA analogs (38) [44]. The results of their study are consistent with previous work, showing that the guanidinylated 12-mer of 38 is taken up into HeLa cells six times more efficiently than its unmodified analog.…”

Chemical Modifications to Improve the Cellular Uptake of Oligonucleotides

“…The cellular uptake of these fully backbone-modified ON was studied recently [61]. Here again, the guanidinium significantly improved the cellular uptake of DNA analogues.…”

Synthesis of novel cationic spermine-conjugated phosphotriester oligonucleotide for improvement of cell membrane permeability

“…8,9 Cationic ONs, which are conjugates of natural phosphodiester (PDE) ON with polycation moieties, such as peptides 10 and polyamines, 11 have been reported. Recently, it was reported that spermine-conjugated PDE ON has higher affinity to complementary strands than natural PDE ON due to the decrease of electrostatic repulsion, 12,13 and also exhibits effective cell penetrating potency by electrostatically anchoring to the cell surface.…”