RNA interference: Targeted delivery by an RNA double

RNA interference: Targeted delivery by an RNA double
August 2006)
by Charlotte Harrison
Nature Reviews: Drug Delivery
One of the major hurdles facing the development and therapeutic application of small interfering RNA (siRNA) is the problem of cell-specific delivery of siRNAs across the plasma membrane. Two papers now describe a novel approach to this problem. By coupling siRNA to RNA aptamers ? oligonucleotides selected to have high affinity and specificity for a macromolecular target ? siRNAs can be specifically delivered to cancer cells that express a particular cell-surface marker, in this case prostate-specific membrane antigen (PSMA).
In their study published in Nucleic Acids Research, Chu and colleagues generated aptamer?siRNA conjugates in which the RNAs were first biotinylated and then non-covalently joined via the biotin-binding protein streptavidin. siRNAs were designed to target either lamin A/C or glyceraldehyde 3-phosphate dehydrogenase (GAPDH), whereas the aptamer targeted PMSA. Aptamer?siRNA conjugates were able to deliver functional siRNA molecules to cells expressing PSMA and rapidly inhibit expression of lamin A/C and GADPH. The presence of the streptavidin molecule had no effect on the function of the attached siRNAs. Importantly, because siRNAs can potentially activate nonspecific inflammatory responses that can lead to cellular toxicity, exposure to the aptamer?siRNA conjugates did not result in increase of interferon- mRNA.
In the second study, reported in Nature Biotechnology, McNamara and colleagues used a sequence-extension method to generate aptamer?siRNA chimeras, in which the aptamer portion targeted PMSA and the siRNA portion targeted one of two survival genes overexpressed in most human tumours: polo-like kinase 1 (PLK1) and B-cell lymphoma 2 (BCL2). Using flow-cytometry and RT-PCR, it was shown that both chimeras specifically bound PSMA on prostate cancer cells and silenced either PLK1 or BCL2 gene expression. In addition, aptamer?siRNA chimeras reduced cellular proliferation and induced apoptosis in PSMA-expressing cells.
McNamara et al. then sought to determine whether the mechanism by which aptamer?siRNA chimeras silence gene expression was dependent on the activity of the endonuclease Dicer. Silencing of PLK1 expression was inhibited by co-transfection of Dicer siRNA, suggesting that aptamer?siRNA chimera-mediated gene silencing is indeed dependent on Dicer and occurs via the RNA interference pathway. Again, treatment of PSMA-expressing cells with aptamer?siRNA chimeras did not induce production of interferon-, suggesting that delivery of aptamer?siRNA chimeras did not trigger a type I interferon response.
Finally, the authors assessed the efficiency and specificity of the Plk1 aptamer?siRNA chimera for its capacity to limit tumour growth in a mouse xenograft model of prostate cancer. PSMA-expressing tumours injected with Plk1 aptamer?siRNA showed a greater than twofold reduction in tumour volume, compared with nearly a fourfold volume increase in controls. Notably, no morbidity or mortality was observed following 20 days treatment with the aptamer?siRNA, suggesting it was not toxic to the animals.
In summary, aptamer?siRNAs can specifically delivery siRNAs, and might provide benefits with regard to specificity and lack of side effects compared with other siRNA-targeting strategies that have been developed so far, such as the use of lipids, peptides and antibodies. Issues such as pharmacokinetics and biodistribution of aptamer?siRNA conjugates and chimeras need to be addressed before they can be used therapeutically, but in principle this approach could have potential for treating prostate cancer and other diseases.
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