Uniformly modified 2'-deoxy-2'-fluoro phosphorothioate oligonucleotides as nuclease-resistant antisense compounds with high affinity and specificity for RNA targets.
"Uniformly" modified phosphodiester or phosphorothioate oligonucleotides incorporating 2'-deoxy-2'-fluoroadenosine, -guanosine, -uridine, and -cytidine, reported herein for the first time, when hybridized with RNA afforded consistent additive enhancement of duplex stability without compromising base-pair specificity. CD spectra of the 2'-deoxy-2'-fluoro-modified oligonucleotides hybridized with RNA indicated that the duplex adopts a fully A-form conformation. The 2'-deoxy-2'-fluoro-modified oligonucleotides in phosphodiester form were not resistant to nucleases; however, the modified phosphorothioate oligonucleotides were highly nuclease resistant and retained exceptional binding affinity to the RNA targets. The stabilizing effects of the 2'-deoxy-2'-fluoro modifications on RNA-DNA duplexes were shown to be superior to those of the 2'-O-methylribo substitutions. RNA hybrid duplexes with uniformly 2'-deoxy-2'-fluoro-modified oligonucleotides did not support HeLa RNase H activity; however, incorporation of the modifications into "chimeric" oligonucleotides has been shown to activate mammalian RNase H. "Uniformly" modified 2'-deoxy-2'-fluoro phosphorothioate oligonucleotides afforded antisense molecules with (1) high binding affinity and selectivity for the RNA target and (2) stability toward nucleases.
Kawasaki
A M
AM
ISIS Pharmaceuticals, Carlsbad, California 92008.
Casper
M D
MD
Freier
S M
SM
Lesnik
E A
EA
Zounes
M C
MC
Cummins
L L
LL
Gonzalez
C
C
Cook
P D
PD
D001483:Base Sequence; D003851:Deoxyribonucleases / Q000187:drug effects; D006868:Hydrolysis; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000138:chemical synthesis* / Q000494:pharmacology; D012260:Ribonucleases / Q000187:drug effects; D013329:Structure-Activity Relationship; D013816:Thermodynamics; D013873:Thionucleotides / Q000138:chemical synthesis* / Q000494:pharmacology
D016376:Oligonucleotides, Antisense; D013873:Thionucleotides; D003851:Deoxyribonucleases; D012260:Ribonucleases