AS1411 Aptamer Linked to DNA Nanostructures Diverts Its Traffic Inside Cancer Cells and Improves Its Therapeutic Efficacy

Abstract

The nucleolin-binding G-quadruplex AS1411 aptamer has been widely used for cancer therapy and diagnosis and linked to nanoparticles for its selective targeting activity. We applied a computational and experimental integrated approach to study the effect of engineering AS1411 aptamer on an octahedral truncated DNA nanocage to obtain a nanostructure able to combine selective cancer-targeting and anti-tumor activity. The nanocages functionalized with one aptamer molecule (Apt-NC) displayed high stability in serum, were rapidly and selectively internalized in cancer cells through an AS1411-dependent mechanism, and showed over 200-fold increase in anti-cancer activity when compared with the free aptamer. Comparison of Apt-NCs and free AS1411 intracellular distribution showed that they traffic differently inside cells: Apt-NCs distributed through the endo-lysosomal pathway and were never found in the nuclei, while the free AS1411 was mostly found in the perinuclear region and in nucleoli. Molecular dynamics simulations indicated that the aptamer, when linked to the nanocage, sampled a limited conformational space, more confined than in the free state, which is characterized by a large number of metastable conformations. A different intracellular trafficking of Apt-NCs compared with free aptamer and the confined aptamer conformations induced by the nanocage were likely correlated with the high cytotoxic enhancement, suggesting a structure–function relationship for the AS1411 aptamer activity.

Aptamers: Features, Synthesis and Applications

“…The comparative studies of Apt‐NCs with free aptamer showed that Apt‐NCs have high serum stability and a 200‐fold increase in anti‐tumor activity. The results show that aptamer is constrained by the nanocage to a more defined conformation which is probably related to the different intracellular localization and enhanced therapeutic effect of AS1411 aptamer [81] …”

“…The results show that aptamer is constrained by the nanocage to a more defined conformation which is probably related to the different intracellular localization and enhanced therapeutic effect of AS1411 aptamer. [81] Zhang et al developed an AS1411 aptamer containing ProteolysisTargetingChimeras (PROTACs) against breast cancer cells. PROTACs are small molecules with heterobifunctional properties and they are highly efficient for protein degradation.…”

Current Status of Oligonucleotide-Based Protein Degraders

“…However, as evidenced previously, breast-cancer-cell-specific aptamer AS1411 can be conjugated with a PROTAC molecule (as inspired by antibody–drug conjugates) as a delivery means to selectively target cancer cells over normal cells [ 21 ]; this constitutes the third layer of precision targeting. One could use structurally modified AS1411 as an alternative [ 96 , 97 , 98 , 99 , 100 , 101 , 102 ], because AS1411 was withdrawn from clinical trials [ 103 ]. The main reason for the withdrawal of AS1411 from clinical trials was its insufficient anti-cancer activity; meanwhile, its safety issue was found to be good [ 44 ].…”

Insights into the binding mode of AS1411 aptamer to nucleolin

“…Aptamers are oligonucleotide sequences with a length of around 20-80 bases, including short single-strand DNA (ssDNA) or RNA molecules, first introduced in the 1990s (Debra L. Robertson, 1990;Doug Irvine, 1991), that bind to their specific targets with high affinity and specificity thanks to their stable three-dimensional folding. Due to their lower cost, smaller size (about 10-fold smaller), and easier modification compared to antibodies, aptamers have become ideal recognition candidates for diagnostic and therapeutic agents, targeted drug delivery systems (Jéssica Lopes-Nunes et al, 2020), biosensing probe (Giulia Vindigni et al, 2021;Fuze Jiang et al, 2022;Tong Mo et al, 2022), etc. Aptamers are usually developed via systematic evolution of ligands by exponential enrichment (SELEX), while AS1411 is the first non-SELEX anticancer aptamer discovered serendipitously by Paula J.…”