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|Title:||Intracellular Microenvironment Triggered Co-delivery of Anticancer Drugs and Genes|
|School/Discipline:||School of Chemical Engineering and Advanced Materials|
|Abstract:||Currently, infallible cancer treatment has not been achieved. Doxorubicin (DOX), an active anticancer drug, initiates the apoptotic signaling pathway by intercalating topoisomerase-II, resulting in DNA replication disorder and producing reactive oxygen species that can damage DNA and activate cell death receptors. However, long-term DOX administration is always accompanied with serious multidrug resistance (MDR) and adverse side-effects. Therefore, delivery systems for DOX and gene drug, such as miRNA-21 inhibitor (miR-21i) which mitigates MDR, have been developed to reduce side-effects caused by non-selective cellular uptake of co-drugs in the absence of suitable delivery systems. However, difficulties in designing delivery systems for different types of drugs to achieve both high therapeutic efficacy and low side-effects have been reported. In this thesis, various biodegradable and biocompatible smart microgels are developed for DOX and gene co-delivery to achieve maximal anticancer performance and minimal side-effects. The microgel delivery carriers are prepared by crosslinking biocompatible low molecular weight polyethyleneimine (PEI800) using glutathione-cleavable diselenide crosslinkers. To the carriers, DOX is conjugated via various intracellular microenvironment sensitive linkers, and genes are electrostatically loaded to glutathione-cleavable polyethyleneimine. Finally, anionic hyaluronic acid (HA) surface coating is able to inhibit serum protein adsorption during blood circulation and promote HA receptor-mediated endocytosis for selective metastatic cancer cells. The hydrodynamic sizes of resulting nano-drugs at 100–200 nm allow prolonged circulation. The developed nano-drugs degrade into less than 10 nm fragments in the glutathione-rich cytosol of cancer cells by cleaving diselenide crosslinkers, allowing intensive gene release and complete urinary excretion. Real-time tracks of release profiles are developed using fluorescence quenching technique. On the other hand, to eliminate premature release, DOX is conjugated to microgel carriers through various linkers for efficiently controlled spatiotemporal release in cancer lysosomes, such as acid-responsive hydrazone linkers, glutathione-cleavable diselenide linkers and enzyme-cleavable peptide linkers. The system comprising of DOX with a hydrazone bond demonstrates 4.4-fold higher in vitro anticancer performance on MDA-MB-231 cell line than free DOX. The nano-drug system of miR-21i and conjugated DOX via hydrazone and diselenide dual bonds completely surpasses DOX premature leakage in physiological condition and results in a high survival rate of over 95 % for HEK293T kidney cell line at a DOX concentration of 2.5 μg mL−1, but it reveals 3.2-fold increase in therapeutic effect on the multidrug-resistant cancer cell line of MDA-MB-231-R12w compared with free DOX. To simplify DOX loading process, diselenide crosslinkers can be used for both microgel synthesis and DOX conjugation to produce a simultaneous release system of gene and DOX in cytosol. The introduction of ATP aptamer to the miR-21i and DOX co-delivery system further increases intracellular DOX accumulation in MDA-MD-231-R12w cells with 4.2-fold higher anticancer performance than free DOX but with low cytotoxicity to healthy cells. To achieve sequential release, the nano-drug system of miR-21i and conjugated DOX with a caspase-3-cleavable DEVDC peptide linker enables the step release of miR-21i followed by DOX. Compared with free DOX, this systems demonstrates a 7.3-fold increase in anticancer effect on MDA-MB-231-R24w and negligible side effects. In summary, we have developed various smart nano-drugs with outstanding advantages of low cytotoxicity, complete biodegradability, targeting and selective delivery and high anticancer performance to MDR cells. Our approach suppresses traditional gene and drug delivery concepts, and advances knowledge for successive design of delivery systems in clinical applications.|
|Dissertation Note:||Thesis (Ph.D.) -- University of Adelaide, School of Chemical Engineering and Advanced Materials, 2019|
|Provenance:||This electronic version is made publicly available by the University of Adelaide in accordance with its open access policy for student theses. Copyright in this thesis remains with the author. This thesis may incorporate third party material which has been used by the author pursuant to Fair Dealing exceptions. If you are the owner of any included third party copyright material you wish to be removed from this electronic version, please complete the take down form located at: http://www.adelaide.edu.au/legals|
|Appears in Collections:||Research Theses|
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