Aptamer directed delivery via cell surface receptors
Kelly, Linsley A.
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Directly targeting chemotherapeutics has potential for reducing adverse side effects as well as possibly increasing the tolerable dose of some drugs. Ideal targeting agents should bind with high affinity and specificity, be easily modifiable for cargo conjugation, and be easily synthesized and purified. For these reasons, aptamers are an attractive option as delivery agents. Aptamers have high affinity and specificity for their target that rival that of an antibody. An advantage of aptamers is that they can be readily chemically synthesized with reactive species for chemical conjugation to therapeutic and imaging cargoes.;To advance aptamers as targeting agents, I demonstrated the use of an aptamer to deliver toxin drugs specifically to cells. I utilized a mutagenized or "doped" selection method to determine a variant (A9min) of the PSMA binding aptamer A9. While the full length A9 has been shown to be highly specific for its target and effective at delivering cargoes to PSMA expressing cells, its length is unwieldy for synthesis. The minimized variant retained the original affinity and specificity of A9 while only being 43 nucleotides in length, ideal for efficient chemical synthesis. I was able to stably and reducibly link the aptamer to the ribosomal toxin Gelonin (rGel). rGel is highly toxic once it has entered the cytosol, but is unable to cross the cellular membrane unassisted. Utilizing a single cysteine in our recombinant variant, we successfully delivered gelonin to cells expressing PSMA, while showing minimal toxicity to cells not expressing PSMA.;Despite advantages of aptamers and my success with delivering drugs to cells, there are surprisingly few aptamers that have made it to the clinic even though a number of aptamers have been reported to bind therapeutic targets. Characterization methods vary greatly within the field making it difficult to assess which aptamers could potentially be used for delivery applications. I systematically assessed a number of aptamers for binding and delivery in vitro and tumor localization in vivo. A surprising number of these aptamers failed to hit targets in this model therapeutic delivery approach, suggesting that more robust examinations must be applied to ensure therapeutically applicable aptamers.
- Theses and Dissertations