Time-lapse live cell imaging to monitor doxorubicin release from DNA origami nanostructures

Abstract

Self-assembled DNA nanostructures have attracted significant research interest in biomedical applications because of their excellent programmability and biocompatibility. To develop multifunctional drug delivery from DNA nanostructures, considerable key information is still needed for clinical application. Traditional fixed endpoint assays do not reflect the dynamic and heterogeneous responses of cells with regard to drugs, and may lead to the misinterpretation of experimental results. For the first time, an integrated time-lapse live cell imaging system was used to study the cellular internalization and controlled drug release profile of three different shaped DNA origami/doxorubicin (DOX) complexes for three days. Our results demonstrated the dependence of DNA nanostructures on shape for drug delivery efficiency, while the rigid 3D DNA origami triangle frame exhibited enhanced cellular uptake capability, as compared with flexible 2D DNA structures. In addition, the translocation of released DOX into the nucleus was proved by fluorescence microscopy, in which a DOX-loaded 3D DNA triangle frame displayed a stronger accumulation of DOX in nuclei. Moreover, given the facile drug loading and auto fluorescence of the anti-cancer drug, DOX, our results suggest that the DNA nanostructure is a promising candidate, as a label-free nanocarrier, for DOX delivery, with great potential for anticancer therapy as well.

Progress toward Understanding the Interactions between DNA Nanostructures and the Cell

“…PBS: phosphate‐buffered saline. Reproduced with permission . Copyright 2018, Royal Society of Chemistry.…”

“…Recent years have witnessed structure–function studies on the roles of size and shape on the cellular uptake of origami structures. In 2018, Zeng et al reported the uptake of 2D nanocrosses (with a thickness of ≈2 nm), 2D nanorectangles (with a thickness of ≈2 nm), and 3D nanotriangles (with a thickness of 6.2 nm) of similar hydrodynamic sizes of ≈50 nm by MDA‐MB‐231 breast cancer cells . Despite statistically insignificant differences in cellular uptake between the three doxorubicin (DOX)‐loaded nanostructures during the first 2 days of incubation, 3D nanotriangles enter the cells more abundantly than the other two structures on day 3, based on the intracellular fluorescence levels of doxorubicin (Figure D).…”

Advances in DNA Origami–Cell Interfaces

“…It is also important, however, to find correlations between the structures, and the functions in which we are interested, such as the actual release of the drug in the target cells. Zeng and co‐workers used time‐lapse live‐cell imaging to investigate the internalisation of three different DOX‐loaded DNA origami nanostructures, including 3D triangles and flexible 2D rectangles and crosses (Figure C), and tracked drug release for up to 3 days . They found that the 3D triangle showed a significantly enhanced cellular uptake and sustained drug release compared with that of the 2D structures.…”

“…Zeng and co-workersu sed timelapse live-cell imaging to investigate the internalisationo f three different DOX-loaded DNA origami nanostructures, including 3D trianglesa nd flexible 2D rectangles and crosses ( Figure 1C), and tracked drug releasef or up to 3days. [14] They found that the 3D triangles howedas ignificantly enhanced cellular uptake and sustainedd rug release compared with that of the 2D structures. It wass imultaneously demonstratedt hat these nanostructures could translocate into the nucleus where the 3D triangled isplayed greater accumulation.…”

“…C) Schematic illustration of different shapes of DNA origamin anocarriers for DOX delivery.Reproduced with permission from ref. [14].Copyright: 2018, Royal SocietyofC hemistry.D)Simulations chematic depicting the mechanism of like-charge attraction during the cellular entry of framework nucleic acids. Adapted with permission from ref.…”