Albany 2015:Book of Abstracts
June 9-13 2015
©Adenine Press (2012)
Developing a new generation of peptidyl-oligonucleotide conjugates with desired biocatalytic properties against biologically relevant RNA
Sequence-specific RNA cleavage is an attractive approach for targeting specific messenger RNA sequences encoding pathogenic proteins or micro-RNAs (e.g. miR-21) up-regulated in a number of cancer pathologies (Si and Zhu, 2006). Such therapeutic interventions would be particularly beneficial to disease states that are not amenable to small-molecule drugs due to the lack of suitable binding sites or antibody inhibition due to their intracellular location.
We developed here peptidyl-oligonucleotide conjugates (POCs) consisting of a covalently-linked, cationic-peptide and antisense oligonucleotide (ASO) (Figure 1a). These constructs act as artificial ribonucleases (aRNase) against RNA molecules. Specific peptide sequences are able to induce RNA hydrolysis under physiological conditions, whereas oligodeoxynucleotide components are able to enhance, and modulate their catalytic activity towards an RNA target. Previously designed POCs have hydrolysed RNA in a non-sequence specific manner, analogous to natural RNase T1 (Mironova and Pyshnyi, 2007). However, we have discovered a novel POC structure which is able to target (with high-affinity) a specific RNA sequence within tRNAPhe and effectively cleave it at a predetermined location (94% total RNA cleavage-Figure 1b). Additionally, through novel incorporation of a cationic peptide with multiple ASO's we have been able to cleave the tRNAPhe phosphodiester backbone within reinforced RNA bulges. In this case the POC acts like a real RNase turning over multiple RNA substrates, similar to those designed by Niittymaki and Lonnberg (2004), however without the need for exogenous metal ions. Selective targeting of specific mRNA sequences encoding disease-relevant proteins or different micro-RNAs associated with diverse types of cancer may potentially expand the therapeutic window for drug discovery and allow new selective therapies to be developed.
Figure 1: (a) Schematic representation of POC hybridised to tRNAPhe target and phosphodiester cleavage mechanism; (b) Concentration-dependent cleavage of tRNAPhe by novel POC.
This research has been supported by the EPSRC and the Collaborative Research Framework Agreement with SOLVAY (Work Program 2).
Mironova, N. L., Pyshnyi, D. V. et al. (2007). RNase T1 mimicking artificial ribonuclease. Nucleic Acids Res. 35(7), 2356-2367.
Niittymaki, T. and Lonnberg, H. (2004). Sequence-Selective Cleavage of Oligoribonucleotides by 3d Transition Metal Complexes of 1,5,9-Triazacyclododecane-Functionalized 2' O-Methyl Oligoribonucleotides. Bioconj. Chem. 15(6), 1275-1280.
1Manchester Pharmacy School