Aptamers as theranostic agents: modifications, serum stability and functionalisation

Abstract

Aptamers, and the selection process known as Systematic Evolution of Ligands by Exponential Enrichment (SELEX) used to generate them, were first described more than twenty years ago. Since then, there have been numerous modifications to the selection procedures. This review discusses the use of modified bases as a means of enhancing serum stability and producing effective therapeutic tools, as well as functionalising these nucleic acids to be used as potential diagnostic agents.

Figure 1.

Schematic of SELEX for DNA (black arrows) or RNA (grey arrows) selection. An initial pool of randomised DNA is amplified by PCR. This pool can be transcribed into RNA prior to incubation with the target of choice. Non-binding species are removed and the binding species are reverse transcribed (RNA aptamers) prior to PCR amplification. This is repeated 8–12 times to produce an enriched pool of binding species that are highly specific for the target. Modified from [18].

Figure 2.

Representative structures of modified bases. (A): RNA bases can be modified at the 2′-OH (R) group with the substitution of fluoro, methoxy, thio or amino groups; The structure of locked nucleic acids is locked in place through a methylene bridge; The structure of unlocked nucleic acids is flexible due to the loss of the C2′-C3′ bond; (B): The sugar ring forms an “S” or “N” type conformation, depending on the functional group (R). Modified from [27,28].

Figure 3.

The insertion of modified bases into the structure of an aptamer can have profound effects on binding affinity. The DNA aptamer TD.05 was modified with locked nucleic acids in the stem region. (A) unmodified DNA aptamer; (B) the last four bases of the aptamer in the stem region were replaced with locked nucleic acids; (C) only the last four bases on the 3′-end of the aptamer were replaced with locked nucleic acids; (D) the first four bases on the 5′-end were modified with locked nucleic acids. Red boxes indicated substituted bases. Modified from [43].