Exploring the conformational behaviour and aggregation properties of lipid-conjugated AS1411 aptamers

AS1411 is a nucleolin-binding aptamer which attracted great interest as active targeting ligand for the selective delivery of therapeutic agents to tumour cells. In this work we selected three AS1411 derivatives 5′-conjugated with lipophilic tails and studied their properties in view of their application in liposomial formulations and/or lipid coated-nanoparticles for targeted therapies. The conformational behaviour of these AS1411 analogs has been investigated in comparison with the unmodified aptamer by CD, UV, PAGE, SEC-HPLC, DLS and thioflavin T (ThT) fluorescence assays to get insight in their secondary structure and aggregation properties. This study has been performed in pseudo-physiological buffers mimicking the extra- and intracellular environments, and at different concentrations in the μM range, paying special attention to the effects of the lipophilic tail on the overall aptamer conformation. The 5′-lipidated AS1411 derivatives proved to fold into stable, parallel unimolecular G-quadruplex structures, forming large aggregates, mainly micelles, at conc. >10 μM. Preliminary bioscreenings on selected cancer cells showed that these derivatives are less cytotoxic than AS1411, but maintain a similar biological behaviour. This study demonstrated that lipophilic tails dramatically favour the formation of AS1411 aggregates, however not impairing the formation and thermal stability of its peculiar G4 motifs.