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Title: Design, synthesis and properties of multivalent DNA nanomaterials for cancer diagnostics and therapeutics
Author: Chen, Jinfeng
ISNI:       0000 0004 8507 4120
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 2019
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Aptamer-functionalised DNA nanostructures have been developed as drug nanocarriers for targeted cancer therapy and other in vivo applications. Among them, hierarchical DNA nanoflowers (DNA-NFs) with densely packed DNA that can self-assemble using rolling circle amplification (RCA) hold promise for cancer therapy. To expand the functionality of DNA-NFs, the inorganic core of the nanoflowers was chemically modified through substitution of Mg2+ with either Mn2+ or Co2+ during the RCA process, resulting in formation of nanomaterials in a range of morphologies. These nanoconstructs retain the stability against enzymatic degradation of their magnesium counterparts and the Mn2+ and Co2+ DNA-NFs were endowed with magnetic properties. Cyclic nucleic acid templates are essential substrates in RCA reactions, and are most commonly obtained via enzymatic cyclisation. However, the cyclisation efficiency mainly depends upon the length of the linear precursor and is inefficient when the linear oligonucleotide has complex secondary structures. To circumvent the limitations of enzyme-catalysed DNA cyclisation, three different chemical cyclisation approaches were explored for the preparation of single-stranded cyclic DNA constructs. These chemical cyclisation methods are cheaper and easier to carry out in a large scale than their ligase-mediated counterpart. They can also be carried out under denaturing conditions and are therefore particular valuable for cyclic DNA templates containing complicated secondary structure. The resulting cyclic DNA templates contain a single unnatural backbone linkage at the ligation site (triazole, phosphoramidate or amide). They were compared to unmodified cyclic DNA templates in RCA reactions using Ф-29 and Bst 2.0 DNA polymerases. The phosphoramidate-modified cyclic templates were particularly well tolerated by Ф-29 DNA polymerase, consistently performing as well as the unmodified natural backbone in RCA. In contrast, the cyclic templates containing a triazole backbone linkage generally showed lower and more variable yields of RCA products, a significant proportion of which were double-stranded DNA side-products. The performance of the amide-modified cyclic templates fell in between those of the triazole- and phosphoramidate-modified cyclic templates. Moreover, all cyclic templates containing modified backbone linkage could be used to produce DNA-NFs during the RCA process.
Supervisor: Brown, Tom Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available