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tumor growth.

      1.13.2 Drug Release Using the Properties Characteristic for DNA Origami

      The twist of the bundled double‐helices can be controlled by changing the pitches which can be adjusted by the number of base‐pairs between crossovers [29]. Högberg and coworkers developed a drug delivery system using a pitch‐controlled 3D origami with Dox (Figure 1.15b) [109]. The 3D origami structure was designed using double helices with different pitches. By designing the DNA origami structures with a pitch of the usual 10.5 base pairs and a looser one of 12 base pairs, the amount of Dox binding to the loose structure increased by 33% compared with the usual type. The amount of incorporated drug can be adjusted, and it can be released slowly over time. The drug was efficiently taken up into cells, confirming that the DNA origami structure is effective for the efficient delivery of Dox. The structure was retained in the cells and apoptosis of cancer cells was effectively induced. This study shows the characteristics of DNA origami that can regulate drug release by designing the structure using different degrees of twist.

      1.13.3 DNA Origami Structures Coated with Lipids and Polymers

      To use a DNA origami structure in vivo, the instability of the DNA structure and the activation of the immune system are obstacles to further application. Therefore, it is necessary to suppress both degradation of the DNA structure against the nuclease and the suppression of immune activation in vivo. Natural biological particles such as viruses have a mechanism to avoid immune recognition during infection by covering the structure with lipids.

      Shih and coworker prepared an octahedral frame‐type DNA origami (c. 50 nm in diameter) covered with a lipid bilayer to prevent degradation and circulate in the blood stream of mice (Figure 1.15c) [110]. By coating with a PEG (polyethylene glycol)‐modified lipid bilayer, the DNA origami device showed resistance to degradation by nucleases. Immune activation was significantly reduced compared to uncoated structures. When these structures were injected into mice, the noncoated DNA origami was rapidly excreted (half‐life = 38 minutes), whereas the lipid‐coated DNA origami was retained for much longer (half‐life = 370 minutes). Furthermore, the researchers confirmed that the DNA origami structures were stable in low salt concentrations and in the medium by covering a barrel‐shaped DNA origami structure with cationic polymers (Figure 1.15d) [111]. In particular, it was found that by covering with a PEG‐conjugated cationic polymer, the DNA origami structure can stably exist in the body of a mouse up to 24 hours. In this way, by establishing a method suitable for biological applications, a medically applicable DNA nanodevices have also been developed.

      1.13.4 Nanorobot with Dynamic Mechanism

      One of the goals of the development of DNA molecular machines is their application to control cellular functions. DNA nanostructures are also as a container for the molecules, by incorporating a dynamic open/close system to release or expose target molecules. The first example of a dynamic nanostructure with open/close system is a DNA box, whose lid opening is controlled by strand displacement with toehold‐containing DNAs [26]. An octahedral structure with a photoresponsive open/close system was constructed to include and release AuNPs [114].

Douglas and co‐workers created a nanorobot for transporting molecular payloads to target biomolecules on the cell surface, which induces a 3D structure change and selectively controls cellular functions. (Figure 1.16a) [115]. The DNA nanorobot with a hexagonal barrel shape opened into two domains, and one end of the structure had hinges to allow the opening and closing of the structure. When the target molecule bound to the connecting dsDNAs (aptamer locks) in the closed structure, it opened by dissociation of the connecting dsDNA. The barrel structure was designed to be opened by recognition and binding to the target molecules on the cell surface. When the barrel structure was designed to open with two types of target molecules, it could be opened only when both target molecules were present on the cell surface. A specific antibody was also incorporated into the structure. The opening of the tubular structure by recognition of the target biomolecules on the cell surface led to the binding of this antibody to the receptor on the cell surface and subsequently activated a signaling pathway to induce either cell death or cell division, as instructed. Using this system, cancer cells can be killed based on the recognition of multiple cellular markers. The nanorobot could be an innovative medical molecular machine that targets various biomolecules and induces regulation of cellular functions.

      1.13.5 Nanorobot Targeting Tumor In Vivo

DNA nanorobots with dynamic mechanisms also have great potential for use in intelligent drug delivery systems that respond to target molecules [116]. Programmed DNA origami structures that respond to specific molecules were designed to be effective for tumor recognition [117]. A nanorobot was functionalized with an exterior DNA aptamer that binds to a protein specifically expressed on tumor‐associated endothelial cells and the blood coagulation protease thrombin on the interior. The aptamer functions as a lock for the DNA nanorobot to open mechanically. When the nanorobot opens, the thrombin inside is exposed and promotes blood clotting at the tumor site. Using a mouse model, the DNA nanorobot was introduced into the blood and delivered thrombin specifically to the tumor‐associated blood vessel, induced tumor necrosis, and inhibited tumor growth. In addition, the nanorobot was immunologically inactive in vivo. This demonstrates that the strategy to use target selective DNA nanorobots is promising for accurate drug delivery and cancer therapy.

Figure 1.16 A DNA nanorobot that recognizes cells. Schematic drawings of the closed nanorobot in hexagonal barrel shape loaded with protein payloads inside in front view. Antibodies are introduced into the tubular shape and is closed by the “key” DNA strand (dashed square). Mechanism for opening the “key.” The target molecule (circle) binds to the upper DNA strand (aptamer DNA) and the initial dsDNA is dissociated. Schematic view of the open state of the nanorobot by protein key displacement of aptamer locks. When the nanorobot is in the open state, antibodies inside bind to cell‐specific antigens.

      Source: Douglas et al. [115]/with permission of American Association for the Advancement of Science.

1.14 Conclusions

The emergence of DNA origami technology has enabled the construction of various 2D and 3D nanostructures and a wide variety of applications. Since DNA origami was first reported in 2006, this technology has relegalized target‐oriented research in the field nanotechnology related to computer science, chemistry, physics, materials science, biology, and medicine. Compared with the use of small DNA assemblies, the DNA origami method improves the flexibility of the structure design and reduces experimental uncertainties. For chemical applications, various methods have been developed to construct synthetic nanosystems and functionalized nanodevices. For physical applications, various single‐molecule studies have been conducted at the nanoscale using DNA origami. For biological applications, cellular and tumor‐targeting studies have been performed and progressed using dynamic DNA origami with various target functions. These achievements show that it is possible to assemble functional DNA origamis as a module to express higher level functionalities in a programmed fashion. Functionalized DNA origami has already been combined with top‐down nanotechnology including semiconductor processing techniques. This technology also opens a way to express the complex functionality of the programmed organization of many different modules seen in living systems.

References

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