Inhibition of Ras p21 synthesis by antisense undecamers with uniform and specifically arranged phosphorothioate linkages

Citation
A. Rait et al., Inhibition of Ras p21 synthesis by antisense undecamers with uniform and specifically arranged phosphorothioate linkages, ANTI-CANC D, 11(3), 2000, pp. 181-191
Citations number
47
Categorie Soggetti
Pharmacology,"Onconogenesis & Cancer Research
Journal title
ANTI-CANCER DRUGS
ISSN journal
09594973 → ACNP
Volume
11
Issue
3
Year of publication
2000
Pages
181 - 191
Database
ISI
SICI code
0959-4973(200003)11:3<181:IORPSB>2.0.ZU;2-Y
Abstract
The design of chimeric oligadeoxynucleotides (ODNs) in which certain phosph odiester linkages are replaced by phosphorothioate (PS) aims to decrease no n-sequence-specific effects of uniform PS ODNs and to preserve the PS-provi ded protection against exo- and endonucleases. This study has, for the fist time, directly compared the differences in nuclease resistance, cellular u ptake, antisense potency and sequence specificity of PS and end-capped, pyr imidine-protected (PPS) undecamer ODNs, that are complementary to the initi ation codon region of human Hares mRNA. At concentrations above 5 mu M, bot h PS and PPS undecamers were moderately and equally stable for over 48 h in complete medium with RS485 cells overexpressing Ha-ras, They were complete ly stable at 0.4 mu M when complexed with Lipofectin reagent that enhanced cellular uptake up to 9-fold, Both the antisense PPS and PS undecamers prod uced well-defined inhibition of Res p21 synthesis in both cell-free and cel l-based assays. However, non-sequence-specific effects of the uniform phosp horothioates were still significant. In contrast, the antisense PPS undecam er, when delivered to RS485 cells with Lipofectin reagent, inhibits human R as p21 synthesis by more than 90% at a concentration of 3.2 mu M, while the effect of controls with inverted, mismatched or scrambled sequence was min imal (5% or less) on p21 synthesis and RS485 cell growth, [(C) 2000 Lippinc ott Williams & Wilkins.].