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Post-SELEX chemical optimization of a trypanosome-specific RNA aptamer.

Adler, Annette and Forster, Nicole and Homann, Matthias and Göringer, H. Ulrich (2008):
Post-SELEX chemical optimization of a trypanosome-specific RNA aptamer.
In: Combinatorial chemistry & high throughput screening, 11 (1), pp. 16-23, ISSN 1386-2073,
[Article]

Abstract

African trypanosomes are the causative agent of sleeping sickness. The therapeutics used to control and treat the disease are very ineffective and thus, the development of improved drugs is urgently needed. Recently, new strategies for the design of novel trypanocidals have been put forward. Among them are techniques that rely on parasite-specific RNA aptamers. One approach involves the aptamer-directed transport of lytic compounds to the lysosome of the parasite. The aptamer has been termed 2-16 RNA and here we report the optimization of the RNA for its applications in vivo. To convert aptamer 2-16 into a serum-stable reagent 2'-deoxy-2'-F- and/or 2'-deoxy-2'-NH(2)-uridine- and cytidine-substituted RNAs were generated. While 2'-NH(2)-dC/dU-modified RNAs were RNase-resistant, they were functionally inactive. By contrast, 2'-F-dC/dU-substituted 2-16 RNA retained its ability to bind to live trypanosomes (K(d)=45 nM) and was routed to the lysosome identically to unmodified RNA. 2'-F-dC/dU-substituted 2-16 RNA is thermostable (T(m)=75 degrees C) and has a serum half-life of 3.4 days. Furthermore, aptamer 2-16 was site-specifically PEGylated to increase its serum retention time. Conjugation with PEG polymers < or = 10 kDa only marginally impacted the binding characteristics of the RNA, while the addition of higher molecular mass PEG molecules resulted in non-functional aptamers. Together, the data provide optimized conjugation chemistries for the large-scale production of substituted aptamer 2-16 preparations with improved in vivo functionality.

Item Type: Article
Erschienen: 2008
Creators: Adler, Annette and Forster, Nicole and Homann, Matthias and Göringer, H. Ulrich
Title: Post-SELEX chemical optimization of a trypanosome-specific RNA aptamer.
Language: English
Abstract:

African trypanosomes are the causative agent of sleeping sickness. The therapeutics used to control and treat the disease are very ineffective and thus, the development of improved drugs is urgently needed. Recently, new strategies for the design of novel trypanocidals have been put forward. Among them are techniques that rely on parasite-specific RNA aptamers. One approach involves the aptamer-directed transport of lytic compounds to the lysosome of the parasite. The aptamer has been termed 2-16 RNA and here we report the optimization of the RNA for its applications in vivo. To convert aptamer 2-16 into a serum-stable reagent 2'-deoxy-2'-F- and/or 2'-deoxy-2'-NH(2)-uridine- and cytidine-substituted RNAs were generated. While 2'-NH(2)-dC/dU-modified RNAs were RNase-resistant, they were functionally inactive. By contrast, 2'-F-dC/dU-substituted 2-16 RNA retained its ability to bind to live trypanosomes (K(d)=45 nM) and was routed to the lysosome identically to unmodified RNA. 2'-F-dC/dU-substituted 2-16 RNA is thermostable (T(m)=75 degrees C) and has a serum half-life of 3.4 days. Furthermore, aptamer 2-16 was site-specifically PEGylated to increase its serum retention time. Conjugation with PEG polymers < or = 10 kDa only marginally impacted the binding characteristics of the RNA, while the addition of higher molecular mass PEG molecules resulted in non-functional aptamers. Together, the data provide optimized conjugation chemistries for the large-scale production of substituted aptamer 2-16 preparations with improved in vivo functionality.

Journal or Publication Title: Combinatorial chemistry & high throughput screening
Volume: 11
Number: 1
Divisions: 10 Department of Biology > Postranscriptional Gene Regulation and RNA Therapeutics
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10 Department of Biology
Date Deposited: 03 Nov 2011 12:59
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