title: Optimizing anti-gene oligonucleotide 'Zorro-LNA' for improved strand invasion into duplex DNA.
abstract: Zorro-LNA (Zorro) is a newly developed, oligonucleotide (ON)-based, Z-shaped construct with the potential of specific binding to each strand of duplex DNA. The first-generation Zorros are formed by two hybridized LNA/DNA mixmers (2-ON Zorros) and was hypothesized to strand invade. We have now established a method, which conclusively demonstrates that an LNA ON can strand invade into duplex DNA. To make Zorros smaller in size and easier to design, we synthesized 3'-5'-5'-3' single-stranded Zorro-LNA (ssZorro) by using both 3'- and 5'-phosphoramidites. With ssZorro, a significantly greater extent and rate of double-strand invasion (DSI) was obtained than with conventional 2-ON Zorros. Introducing hydrophilic PEG-linkers connecting the two strands did not significantly change the rate or extent of DSI as compared to ssZorro with a nucleotide-based linker, while the longest alkyl-chain linker tested (36 carbons) resulted in a very slow DSI. The shortest alkyl-chain linker (3 carbons) did not reduce the extent of DSI of ssZorro, but significantly decreased the DSI rate. Collectively, ssZorro is smaller in size, easier to design and more efficient than conventional 2-ON Zorro in inducing DSI. Analysis of the chemical composition of the linker suggests that it could be of importance for future therapeutic considerations.
authors.affiliation: Department of Laboratory Medicine, Karolinska Institutet, 141 86 Huddinge, Stockholm, Sweden. eman.zaghloul@ki.se
authors.forename: Eman M
authors.initials: EM
authors.lastname: Zaghloul
authors.forename: Andreas S
authors.initials: AS
authors.lastname: Madsen
authors.forename: Pedro M D
authors.initials: PM
authors.lastname: Moreno
authors.forename: Iulian I
authors.initials: II
authors.lastname: Oprea
authors.forename: Samir
authors.initials: S
authors.lastname: El-Andaloussi
authors.forename: Burcu
authors.initials: B
authors.lastname: Bestas
authors.forename: Pankaj
authors.initials: P
authors.lastname: Gupta
authors.forename: Erik B
authors.initials: EB
authors.lastname: Pedersen
authors.forename: Karin E
authors.initials: KE
authors.lastname: Lundin
authors.forename: Jesper
authors.initials: J
authors.lastname: Wengel
authors.forename: C I Edvard
authors.initials: CI
authors.lastname: Smith
mesh_terms: D004247:DNA / Q000737:chemistry*; D020868:Gene Silencing; D009693:Nucleic Acid Hybridization; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D009841:Oligonucleotides / Q000737:chemistry*; D010957:Plasmids / Q000737:chemistry
chemical_list: D009838:Oligodeoxyribonucleotides; D009841:Oligonucleotides; C477371:locked nucleic acid; C051802:triplex DNA; D004247:DNA