Biophysical characterisation of DNA origami nanostructures reveals inaccessibility to intercalation binding sites

Helen L Miller,Mark C Leake,Deborah O’Connell,Katherine E Dunn,Adam J M Wollman,Sonia Contera,Sandra Schröter,Adam Hirst

doi:10.1088/1361-6528/ab7a2b

Helen L Miller, Mark C Leake + Show 6 more

Open Access

https://doi.org/10.1088/1361-6528/ab7a2b

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Journal: Nanotechnology	Publication Date: Mar 20, 2020
Citations: 27	License type: cc-by

Affiliation: University of York, University of Oxford

Abstract

Intercalation of drug molecules into synthetic DNA nanostructures formed through self-assembled origami has been postulated as a valuable future method for targeted drug delivery. This is due to the excellent biocompatibility of synthetic DNA nanostructures, and high potential for flexible programmability including facile drug release into or near to target cells. Such favourable properties may enable high initial loading and efficient release for a predictable number of drug molecules per nanostructure carrier, important for efficient delivery of safe and effective drug doses to minimise non-specific release away from target cells. However, basic questions remain as to how intercalation-mediated loading depends on the DNA carrier structure. Here we use the interaction of dyes YOYO-1 and acridine orange with a tightly-packed 2D DNA origami tile as a simple model system to investigate intercalation-mediated loading. We employed multiple biophysical techniques including single-molecule fluorescence microscopy, atomic force microscopy, gel electrophoresis and controllable damage using low temperature plasma on synthetic DNA origami samples. Our results indicate that not all potential DNA binding sites are accessible for dye intercalation, which has implications for future DNA nanostructures designed for targeted drug delivery.

Highlights

Over the past decade important progress has been made in the practical use of DNA nanostructures for targeted therapeutic drug delivery [1,2,3,4,5], in particular for intercalating molecules [6,7,8,9], including the anti-cancer drug doxorubicin (DOX)
Our results indicate that not all potential DNA binding sites are accessible for dye intercalation, which has implications for future DNA nanostructures designed for targeted drug delivery
M13mp18ss is single stranded and appeared red in the merged fluorescence image, producing a red to green fluorescence intensity ratio (RG ratio, see methods section 1.7) of 21.54 ± 4.53 (± standard deviation; figure 2b) when imaged in red and green fluorescence using a gel imager (ChemiDoc MP Imaging System, BioRad Laboratories)

Summary

Introduction

Over the past decade important progress has been made in the practical use of DNA nanostructures for targeted therapeutic drug delivery [1,2,3,4,5], in particular for intercalating molecules [6,7,8,9], including the anti-cancer drug doxorubicin (DOX). Many previous studies have used DNA origami with cage-like structures, but the use of compact structures with intercalated drug molecules could enable higher drug loading, leading to more efficient delivery of the active agent to the target cells. A recent study showed that tetrahedral DNA nanostructures did not cause significant impairment to native physiology [11], triggering interest in DNA structures with an internal column for drug transportation. Another recent work [12] has shown that large (~MDa) DNA nanoparticles with a high external surface area-to-volume ratio are preferentially taken up in mammalian cell lines relevant to therapeutic drug delivery. Zeng et al [9] found that rigid 3D DNA origami shapes are more readily taken up by cells, and exhibit sustained drug release, compared to more flexible 2D DNA structures

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