Cell-Specific Induction of Apoptosis by Rationally Designed Bivalent Aptamer-siRNA Transcripts Silencing Eukaryotic Elongation Factor 2

Abstract

New strategies for cell type-specific delivery need to be developed if RNA interference is to realize its full therapeutic potential. One possible approach is the use of aptamers to deliver siRNAs selectively to tumor cells with appropriate antigens displayed on the surface. We used an aptamer that binds specifically to PSMA, a cell surface glycoprotein found in abundance on prostate cancer cells, and joined its 3' end to a siRNA specific for Eukaryotic Elongation Factor 2 mRNA (EEF2). This is an attractive target for cancer therapy because inhibiting EEF2 causes the rapid arrest of protein synthesis, inducing apoptosis and leading ultimately to cell death. In order to enhance the therapeutic efficacy of the aptamer-siRNA, we increased the valency of the construct by rational design. Two anti-PSMA aptamers were designed such that each binding sequence could fold independently into its active conformation. Here we show specific cytotoxicity resulting from siRNA-induced silencing of EEF2, as well as specific delivery to PSMA-expressing prostate cancer cells. Increasing the valency of the aptamer resulted in enhanced cytotoxicity compared with the monovalent constructs. The results presented here demonstrate the usefulness of multivalent aptamer-based delivery vehicles for siRNA therapeutics.

Prostate-targeted radiosensitization via aptamer-shRNA chimeras in human tumor xenografts

“…We have previously developed PSMA-targeting aptamers as a means to selectively deliver therapeutic and imaging agent to PCa cells (13). Several groups have independently used these aptamers to target therapeutics, including siRNAs and shRNAs (19)(20)(21)23). Here, we have also extended on the initial work by generating 2′ fluoro-modified pyrimidine aptamer-shRNA chimeras as selective radiosensitizing agents.…”

“…We have previously developed prostate-specific membrane antigen-targeted (PSMA-targeted) RNA aptamers (13), which are capable of targeting drugs, nanoparticles, and toxins to PSMA-expressing PCa cells and tumors (14)(15)(16)(17)(18). When conjugated to siRNAs and shRNAs, these PSMA aptamers are also capable of delivering cell-selective gene knockdown (19)(20)(21)(22)(23). Because PSMA is highly expressed in nearly all localized prostate tumors (24,25), we hypothesized that PSMA-targeted aptamer-shRNA chimeras could be used to inhibit DNA repair pathways in prostatic cells for enhanced radiation therapy of locally advanced PCa.…”

Bivalent aptamer-dual siRNA chimera is emerging as a new combination therapy

“…To overcome short circulation half-life, PEG has been conjugated with aptamer -siRNA chimera to increase circulation time in vivo [11] . In our new chimera, the size of PSEP is bout 59Kd, which is much larger than current PSMA aptamer-siRNA chimeras [11,29] (around 29Kd). We envision that chimera with a bivalent aptamer and dual siRNAs should possess increased circulating half-life and reduced renal excretion.…”

Cell-Specific Aptamers for Nano-medical Applications

“…Addition of a two-nucleotide overhang at the 3 0 end or swap of the guide and passenger strands results in an optimized chimera with improved silencing and therapeutic efficacy (Dassie et al 2009). Wullner et al (2008) designed a bivalent aptamer-eukaryotic elongation factor 2 (EEF2) targeting siRNA chimera. The design of a chimera with two anti-PSMA aptamers resulted in an increased cellular uptake of siRNAs and an induced EEF2-siRNA-mediated cytotoxicity to prostate cancer cells.…”