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Designing and monitoring DNA origami particles for single-cell precision delivery
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Title:
Designing and monitoring DNA origami particles for single-cell precision delivery
Author:
Molbay, Müge
Subjects:
ddc:600
;
ddc:610
;
Medizinische Fakultät
Description:
Nanomedicine is a rapidly developing field that holds significant potential for targeted drug delivery and treatment of various diseases. Among the emerging technologies, DNA nanotechnology has gained attention for its versatility, biocompatibility, and programmability in constructing structures for targeted drug delivery. However, despite significant investments in these technologies, most are currently limited to in vitro evaluation. Moreover, the lack of tools for investigating the biodistribution of these structures at the cellular level hinders the ability to track them throughout the body. Recently, tissue-clearing methods have been developed to overcome these limitations and enable the exploration of in-tact biological specimens by rendering tissues transparent and imaging them using laser scanning fluorescence microscopy. Advancements in tissue clearing and light sheet microscopy have made it possible to investigate large volumes, such as whole mouse bodies, at cellular resolution. These new techniques provide a powerful tool for studying the bio-distribution of DNA nanotechnology-based drug delivery systems in vivo, ultimately advancing the development of nanomedicine as a promising approach for treating a wide range of diseases. In pursuit of developing an efficient and safe DNA nanotechnology-based drug delivery system, we generated DNA origami rods (8 nm x 80 nm) carrying fluorescent dyes. Prior to in vivo evaluation, we assessed the production efficiency and stability of the DNA origami in vitro, followed by evaluating its immune compatibility, half-life, targeting efficiency, and biodistribution in CD1 mice (n=5) after intravenous (iv) or oral administration. Results showed that the DNA origami treatment did not alter immune cell counts up to 7 days post-injection, indicating the immune safety of the treatment. In vivo imaging of the brain surface using intravital 2-photon microscopy revealed the clearance of the DNA origami from the meningeal blood circulation within 20 minutes post-injection, without any leakage into the brain parenchyma. In the periphery, the DNA origami was gradually cleared from the liver until 24 hours post-injection. Further, we investigated the biodistribution and targeting potential of DNA origami coupled with antibodies after IV injection. After 20 minutes of circulation, mouse bodies were subjected to vDISCO clearing and imaged using light sheet microscopy. Results showed that DNA origami coupled with CX3CR1 antibody was observable in lymph nodes, spleen, and Kupffer cells in the liver while coupling with human carbonic anhydrase (CA) XII-specific antibody (6A10) directed the DNA origami to the lung where it co-localized with metastases in mice injected with MDA-MB-231 metastatic cancer cells. These findings demonstrate the potential of DNA origami-based drug delivery systems for targeted drug delivery, highlighting their potential for use in treating a range of diseases. Thus, this approach can significantly help develop DNA nanotechnology for in vivo applications including drug delivery and gene editing. So far, our approach enables the assessment of biodistribution in the intact body with a sensitivity down to the single-cell level, revealing DNA origami's feasibility for drug targeting.
Publisher:
Ludwig-Maximilians-Universität München
Creation Date:
2024
Language:
German
Source:
Digital Dissertations (Universitätsbibliothek der LMU Muenchen)
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