Potts, Jordan Cameron
(2023)
Light-Induced Cytochrome c Redox Mediated Cell Killing.
PhD thesis, University of Nottingham.
Abstract
This research presents a novel cancer treatment strategy employing gold nanoparticles as Cytochrome c carriers. Leveraging the protein's inherent apoptotic capabilities, this approach activates the therapeutic potential of Cytochrome c using a porphyrin "trigger" when exposed to monochromatic visible light. This research meticulously details the synthesis, functionalisation, and characterisation of AuNPs with diameters of 20, 50, and 100 nm, utilising many analytical techniques. The AuNPs were functionalised with Cyt c and Zn Porph through two distinct ligands and conjugation strategies, preserving their stability. Our novel spectral deconvolution approach critically ascertains the precise concentration and number of conjugated molecules on AuNPs, setting the stage for an in-depth analysis of their impact, primarily on the redox capabilities of Cyt c. The core objective was to determine if conjugated Cyt c and Zn Porph can instigate apoptosis in cancer cells upon exposure to light.
We examined the electrochemical properties of Cyt c and Zn Porph before and after their conjugation to ascertain any functional alterations. The conjugation process instigated noticeable changes in the structure of Cyt c, dependent on the AuNP size and conjugation methods. ToF-SIMS measurements pinpointed the orientation of Cyt c post-conjugation. Strikingly, while Cyt c showed reduced electron transfer rates post-conjugation, Zn Porph maintained its electron transfer capabilities. Notably, the potential of light-triggered Cyt c activation via Zn Porph remained intact, paving the way for evaluating its anti-cancer efficacy.
Finally, our investigations into the therapeutic application of multifunctionalised AuNPs for cancer therapy revealed that these nanoparticles are proficiently internalised by MCF-7 cells. Their therapeutic potential was realised upon exposure to amber light, leading to significant cellular death. Intriguingly, the size of the AuNPs played a crucial role in enhancing the peroxidase activity of Cyt c. However, certain measurements, such as the PrestoBlue assay, were hindered by fluorescent overlaps, suggesting a need for more apt methodologies. Our findings conclude that AuNPs, when multifunctionalized with Cyt c and Zn Porph and activated by amber light, possess remarkable potential in initiating apoptosis in cancer cells. This pioneering approach holds transformative potential for targeted cancer therapies, marking a pivotal advancement in nanoparticle-based medical intervention.
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