Brief communication: stability and catalytic activity of novel circular DNAzymes.
DNAzymes represent a new generation of catalytic nucleic acids for specific RNA targeting in order to inhibit protein translation from the specifically cleaved mRNA. The 10-23 DNAzyme was found to hydrolyze RNA in a sequence-specific manner both in vitro and in vivo. Although single-stranded DNAzymes may represent the most effective nucleic acid drug to date, they are nevertheless sensitive to nuclease degradation and require modifications for in vivo application. However, previously used stabilization of DNAzymes by site-specific phosphorothioate (PT) modifications reduces the catalytic activity, and the PTO displays toxic side effects when applied in vivo. Thus, improving the stability of DNAzymes without reducing their catalytic activity is essential if the potential of these compounds should be realized in vivo.


RESULTS
The Circozyme was tested targeting the mRNA of the most common genetic rearrangement in pediatric acute lymphoblastic leukemia TEL/AML1 (ETV6/RUNX1). The Circozyme exhibits a stability comparable to PTO-modified DNAzymes without reduction of catalytic activity and specificity and may represent a promising tool for DNAzyme in vivo applications.


CONCLUSIONS
The inclusion of the catalytic site and the specific mRNA binding sequence of the DNAzyme into a circular loop-stem-loop structure (Circozyme) of approximately 70 bases presented here represents a new effective possibility of DNAzyme stabilization.
Seifert
Georg
G
Charité Universitätsmedizin Berlin, Otto-Heubner Center for Pediatric and Adolescent Medicine (OHC), Department of Pediatric Oncology/Hematology, Berlin, Germany.
Taube
Tillmann
T
Paal
Kriztina
K
von Einsiedel
Hagen Graf
HG
Wellmann
Sven
S
Henze
Günter
G
Seeger
Karl
K
Schroff
Matthias
M
Wittig
Burghardt
B
D001483:Base Sequence; D002384:Catalysis; D050676:Core Binding Factor Alpha 2 Subunit / Q000235:genetics*; D021881:DNA, Catalytic / Q000737:chemistry* / Q000378:metabolism; D004270:DNA, Circular / Q000737:chemistry* / Q000378:metabolism; D004795:Enzyme Stability; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D015514:Oncogene Proteins, Fusion / Q000235:genetics*; D012333:RNA, Messenger / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D000097764:ETS Translocation Variant 6 Protein
D050676:Core Binding Factor Alpha 2 Subunit; D021881:DNA, Catalytic; D004270:DNA, Circular; D015514:Oncogene Proteins, Fusion; D012333:RNA, Messenger; C102421:TEL-AML1 fusion protein; D000097764:ETS Translocation Variant 6 Protein