Programming DNA Aptamer Arrays of Prescribed Spatial Features with Enhanced Bioavailability and Cell Growth Modulation

Abstract

Epithelial cell adhesion molecules (EpCAMs) play pivotal roles in tumorigenesis in many cancer types, which is reported to reside within nano- to microscale membrane domains, forming small clusters. We propose that building multivalent ligands that spatially patch to EpCAM clusters may largely enhance their targeting capability. Herein, we assembled EpCAM aptamers into nanoscale arrays of prescribed valency and spatial arrangements by using a rectangular DNA pegboard. Our results revealed that EpCAM aptamer arrays exhibited significantly higher binding avidity to MCF-7 cells than free monovalent aptamers, which was affected by both valency and spatial arrangement of aptamers. Furthermore, EpCAM aptamer arrays showed improved tolerance against competing targets in an extracellular environment and potent bioavailability and targeting specificity in a xenograft tumor model in mice. This work may shed light on rationally designing multivalent ligand complexes of defined parameters with optimized binding avidity and targeting capability toward various applications in the biomedical fields.

Engineering Circular Aptamer Assemblies with Tunable Selectivity to Cell Membrane Antigens In Vitro and In Vivo

“…We next employed the classic cooperativity parameter and enhancement parameter, corresponding to α and β, respectively, to quantitatively decipher the cooperativity of the multivalent aptamer assemblies, as indicated in eqs and . In these classic equations, N stands for the valency of the high-affinity aptamer. K normald , N = false( K normald , 1 false) α N K normald , N = β K normald , 1 …”

“…We next employed the classic cooperativity parameter and enhancement parameter, corresponding to α and β, respectively, to quantitatively decipher the cooperativity of the multivalent aptamer assemblies, as indicated in eqs and . In these classic equations, N stands for the valency of the high-affinity aptamer. …”

“…DNA-based materials offer another platform for engineering highly controllable and addressable scaffolds and binding agents. − For instance, DNA nanotechnology allows for constructions of various nanostructures for precise modification of different binding agents. − System evolution of ligands by exponential enrichment (SELEX) technology facilitates the discovery of aptamer panels that can recognize a wide range of targets, especially the cell surface antigens. , In addition, it is generally believed that these pure DNA-based delivery platforms are with low immunogenicity and therefore have been explored as tools for targeted drug delivery in vivo . However, most reported DNA-based targeting platforms were focusing on improving the binding affinity by functionalizing multiple aptamers on the scaffolds. − Limited studies were reported to investigate the facets of avidity optimization such as tuning the binding selectivity based on antigen densities, ligand valency, and antigen/ligand affinity . Specifically, there is still lack of theoretical models and experimental data that elaborate the performance (i.e., affinity and specificity) of multivalent high/low-affinity interactions of DNA aptamers to their antigens on cell membranes in vitro and in vivo .…”

Multivalent DNA Flowers for High-Performance Isolation, Detection, and Release of Tumor-Derived Extracellular Vesicles

“…26 Inspired by nature, multivalent ligand−receptor systems 27,28 are used to increase the binding affinity because integration of multiple binding sites in a restricted area can improve the relative proximity and local concentrations. 29 The rapid development of DNA nanotechnology provides an opportunity for multivalent ligand−receptor binding. Various DNA nanostructures have been designed such as DNA tetrahedrons, 30−32 DNA dendrimers, 33 DNA origami, 18,29 and others 34−36 during the past decades.…”

“…The rapid development of DNA nanotechnology provides an opportunity for multivalent ligand–receptor binding. Various DNA nanostructures have been designed such as DNA tetrahedrons, − DNA dendrimers, DNA origami, , and others − during the past decades. Some of them have aptamer nanoarrays with specific pricing and spatial arrangement, showing significantly higher binding affinity. ,, But after the identification of EVs, due to the small size of these DNA materials, EVs cannot be separated from the biological matrix by simple centrifugation.…”