Nucleic Acids Research, 2003, Vol. 31, No. 18 5282-5290
© 2003 Oxford University Press
Cationic phosphoramidate 
-oligonucleotides efficiently target single-stranded DNA and RNA and inhibit hepatitis C virus IRES-mediated translation
Laboratoire de Chimie Organique Biomoléculaire de Synthèse, UMR 5625 CNRS-UMII, CC 008, Université Montpellier II, Place Eugène Bataillon, 34095 Montpellier Cedex 05, France and 1 UMR 5124 CNRS-UMII, IGMM, Centre National de la Recherche Scientifique, 1919 Route de Mende, 34293 Montpellier Cedex 05, France
*To whom correspondence should be addressed. Tel: +33 4 67 14 38 98; Fax: +33 4 67 04 20 29; Email: debart{at}univ-montp2.fr
The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors
A potential means to improve the efficacy of steric-blocking antisense oligonucleotides (ON) is to increase their affinity for a target RNA. The grafting of cationic amino groups to the backbone of the ON is one way to achieve this, as it reduces the electrostatic repulsion between the ON and its target. We have examined the duplex stabilising effects of introducing cationic phosphoramidate internucleoside linkages into ON with a non-natural 
-anomeric configuration. Cationic -ON bound with high affinity to single-stranded DNA and RNA targets. Duplex stabilisation was proportional to the number of cationic modifications, with fully cationic ON having particularly high thermal stability. The average stabilisation was greatly increased at low ionic strength. The duplex formed between cationic -ON and their RNA targets were not substrates for RNase H. The penalty in Tm inflicted by a single mismatch, however, was high; suggesting that they are well suited as sequence-specific, steric-blocking, antisense agents. Using a well-described target sequence in the internal ribosome entry site of the human hepatitis C virus, we have confirmed this potential in a cell-free translation assay as well as in a whole cell assay. Interestingly, no vectorisation was necessary for the cationic -ON in cell culture.
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