When Zamecnik and Stephenson first proposed synthetic oligodeoxyribo-nucleotides as reagents to bind to Rous sarcoma virus RNA to inhibit viral replication, they designed an oligonucleotide (ON) to target the initiation site for protein translation in the expectation that it would form an RNA-DNA duplex and physically block the RNA, thus preventing protein synthesis.1 Several years later it was found that an alternative mechanism probably operates when DNA oligomers bind to RNA targets inside cells, that of recognition of the hybrid by the cellular enzyme RNase H and subsequent RNA cleavage. This second attribute of DNA ONs, which extends to their phosphorothioate-(PS-) modified counterparts, became established as the predominant 'antisense' mechanism of action and led to the first industrial development of this class of therapeutic ONs. The original concept of the steric block mechanism of inhibition of protein translation continued to be studied by scientists,2,3 but only recently have steric block ONs been considered seriously as potential therapeutics. In addition, we now know that through duplex
RSC Biomolecular Sciences Therapeutic Oligonucleotides Edited by Jens Kurreck © Royal Society of Chemistry 2008
formation a number of other RNA-processing events can also be inhibited sterically, for example nuclear splicing, which is required for the processing of most mammalian gene transcripts, and which involves numerous steps of RNA-protein recognition.4 Another novel example is the inhibition of endogenous microRNAs by complementary synthetic ON constructs, which has recently been used in vivo.5 Note that there is often confusion in the use of the word 'antisense', which is sometimes used interchangeably for both RNase H and steric block mechanisms. In this chapter, the word antisense is used strictly to imply that the oligomer is likely to work predominantly by an RNase H mechanism of action.
An advantage of the steric block approach is the ability to use a considerably wider range of synthetic analogues (see Section 4.2) than is possible with conventional antisense, since there is no requirement for recognition of the RNA-DNA hybrid by a cellular enzyme. Instead, the issue is to obtain tight binding to the RNA target by the analogue while also showing good resistance to nuclease degradation. This allows a greater ability to manipulate ON chemistry and composition to obtain improved pharmacological parameters. In principle, a steric block oligomer might be expected to show a greater specificity (lower off-target effects) compared to antisense, since the binding to a wrong sequence will often not have any biological consequence, because most parts of the RNA (e.g. coding) are not affected by such oligomer binding. One disadvantage is the need for at least a stoichiometric amount of the oligomer on the RNA target for full inhibition, whereas antisense and small interfering RNAs (siRNAs) are catalytic in action. In practice however, the amounts of many messenger RNA (mRNA) targets inside cells are small and the issue therefore becomes simply a question of delivering sufficient oligomer into cells to obtain the required biological effect.
Was this article helpful?
Learning About 10 Ways Fight Off Cancer Can Have Amazing Benefits For Your Life The Best Tips On How To Keep This Killer At Bay Discovering that you or a loved one has cancer can be utterly terrifying. All the same, once you comprehend the causes of cancer and learn how to reverse those causes, you or your loved one may have more than a fighting chance of beating out cancer.