Extensive Sugar Modification Improves Triple Helix Forming Oligonucleotide Activity in Vitro but Reduces Activity in Vivo
详细信息 查看全文
- 作者：Md. Rowshon Alam ; Alokes Majumdar ; Arun Kalliat Thazhathveetil ; Su-Ting Liu ; Ji-Lan Liu ; Nitin Puri ; Bernard Cuenoud ; Shigeki Sasaki ; Paul S. Miller ; Michael M. Seidman
- 刊名：Biochemistry
- 出版年：2007
- 出版时间：September 4, 2007
- 年：2007
- 卷：46
- 期：35
- 页码：10222 - 10233
- 全文大小：572K
- 年卷期：v.46,no.35(September 4, 2007)
- ISSN：1520-4995
文摘
We are developing triple helix forming oligonucleotides (TFOs) for gene targeting. Previously,we synthesized bioactive TFOs containing 2'-O-methylribose (2'-OMe) and 2'-O-aminoethylribose (2'-AE) residues. Active TFOs contained four contiguous 2'-AE residues and formed triplexes with highthermal stability and rapid association kinetics. In an effort to further improve bioactivity, we synthesizedthree series of TFOs containing the 2'-AE patch and additional ribose modifications distributed throughoutthe remainder of the oligonucleotide. These were either additional 2'-AE residues, the conformationallylocked BNA/LNA ribose with a 2'-O,4'-C-methylene bridge, or the 2'-O,4'-C-ethylene analogue (ENA).The additionally modified TFOs formed triplexes with greater thermal stability than the reference TFO,and some had improved association kinetics. However, the most active TFOs in the biochemical andbiophysical assays were the least active in the bioassay. We measured the thermal stability of triplexesformed by the TFOs in each series on duplex targets containing a change in sequence at a single position.The Tm value of the variant sequence triplexes increased as the number of all additional modificationsincreased. A simple explanation for the failure of the improved TFOs in the bioassay was that the increasedaffinity for nonspecific targets lowered the effective nuclear concentration. Enhancement of TFO bioactivitywill require chemical modifications that improve interaction with the specific targets while retainingselectivity against mismatched sequences.