Minimally modified phosphodiester antisense oligodeoxyribonucleotide directed against the multidrug resistance gene mdr1.
In the perspective of reversing multidrug resistance through antisense strategy while avoiding non-antisense effects of all-phosphorothioate oligonucleotides which non-specifically bind to proteins, a minimally modified antisense phosphodiester oligodeoxyribonucleotide has been designed against mdr1, one of the multidrug resistance genes. Its stability in lysates prepared from NIH/3T3 cells transfected with the human mdr1 gene has already been demonstrated. Confocal microspectrofluorometry using a fluorescence resonance energy transfer technique allowed its stability inside living cells to be proven. Its internalization into the cells was achieved with different delivery agents (addition of a cholesteryl group, Superfect or an amphotericin B cationic derivative) and has been followed by fluorescence imaging. For each of the delivery systems, Western blotting allowed its antisense efficiency to be compared to that of an all-phosphorothioate antisense oligonucleotide. No antisense efficiency was demonstrated for the minimally modified ODN when internalized with Superfect. In both other cases, the best extinction of the P-glycoprotein expression has always been achieved with the all-phosphorothioate antisense. While the difference was significant in the case the amphotericin B derivative was used as delivery agent (20% remaining protein expression with the all-phosphorothioate vs. 40% with the minimally modified antisense), it was negligible for the cholesterol conjugates (2% vs. 6%). It is of great interest to prove that an almost all-phosphodiester oligonucleotide can be an efficient antisense against an overexpressed gene. The reduction of non-antisense effects as non-specific binding to proteins are of importance in the case relatively high ODN concentrations are used, which can prove to be necessary in the case of overexpressed genes.
Brigui
Imane
I
Laboratoire de Physicochimie Biomoléculaire et cellulaire, CNRS (URA 7033) et Université P. et M. Curie, 4 place Jussieu, case 138, 75005 Paris, France.
Djavanbakht-Samani
Taraneh
T
Jollès
Béatrice
B
Pigaglio
Sophie
S
Laigle
Alain
A
D016475:3T3 Cells; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics; D000818:Animals; D001692:Biological Transport; D004355:Drug Stability; D019450:Genes, MDR / Q000187:drug effects*; D051379:Mice; D020319:Oligodeoxyribonucleotides, Antisense / Q000737:chemistry / Q000494:pharmacology*; D010710:Phosphates / Q000737:chemistry
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D020319:Oligodeoxyribonucleotides, Antisense; D010710:Phosphates