A novel concept for ligand attachment to oligonucleotides via a 2'-succinyl linker

Abstract

Conjugation of ligands to antisense oligonucleotides is a promising approach for enhancing their effects. In this report, a new method for synthesizing oligonucleotide conjugates is described. 2'-Amino-2'-deoxy-5'-dimethoxytrityl-uridine was select ively acylated with a succinic acid linker at the 2' position. This compound was incorporated at the 3' end of an oligonucleotide corresponding to the sequence of Oblimersen. The carboxyl group was protected for oligonucleotide synthesis as a benzyl ester, which could be selectively cleaved at the solid phase by a catalytic phase transfer reaction using palladium nanoparticles as catalyst. An oligonucleotide-fluorescein conjugate was prepared by condensation of aminofluorescein. Circular dichroism spectroscopic experiments showed a B-DNA type structure. The melting temperature of the duplex was only slightly lower than that of Oblimersen. Biological activity measured by western blotting resulted in a Bcl-2 target downregulation nearly identical to that of control Oblimersen on human melanoma cells, proving that this method is attractive for the binding of ligands located in the minor groove.

Fluorogenic affinity label for the facile, rapid imaging of proteins in live cells

“…Briefly, known t -Boc–rhodamine 4 12 was subjected to reaction with an in situ -generated isocyanate from protected succinate 5 20 to generate t -Boc–rhodamine–urea 6 . Deprotection with trifluoroacetic acid furnished the urea–rhodamine 7 that underwent carbodiimide-mediated coupling with trimethyl lock acid 8 21 to give benzyl-protected 9 .…”

“…Compound 5 was synthesized according to a published procedure 20. The white crystalline material afforded in the published procedure was dissolved in a minimal amount of 1:1 hexanes/EtOAc and cooled to 4 °C.…”

Synthesis of Functionalised Nucleosides for Incorporation into Nucleic Acid-Based Serine Protease Mimics

“…As for our envisaged carboxylic acid building block, shown in Scheme 2, a derivative similar to 11, only differing in the carboxylic acid protecting group, has been synthesized before. However, this modified nucleoside was incorporated only at the end of oligonucleotide sequences [44]. The carboxyl group was protected as a benzyl ester, which can be selectively cleaved by hydrogenolysis.…”

Off‐Target Effects Related to the Phosphorothioate Modification of Nucleic Acids

“…All oligonucleotides were prepared on a Polygen 10 column DNA/RNA synthesizer (Polygen, Langen, Germany) using standard protocols for each chemical modification24, 25 with DCI as activator. Nucleoside phosphoramidites and reagents were supplied by Proligo‐SAFC (Hamburg, Germany).…”