Developing Ligation Strategies for Preparing Antibody-Gold Nanocluster Conjugates
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Authors
Golds, Alexandra
Date
2024-10-04
Type
thesis
Language
eng
Keyword
antibody , gold nanocluster , antibody remodeling , cysteine-maleimide ligations , glyco-engineering , bioconjugation , click chemistry
Alternative Title
Abstract
Radiation therapy is one of the most effective and widely used treatments for solid cancers, however, typical therapeutic doses can severely damage surrounding healthy tissues. Recent work has shown that coupling radio-sensitizing agents to targeting molecules for targeted photodynamic therapies (PDTs) can greatly improve the precision and selectivity of radiation treatment. As such, targeted PDTs have emerged as a highly effective alternative treatment for hard-to-treat solid cancers. Gold nanoclusters (AuNCs) show promise as radio- and photo-sensitizers; gold is highly biocompatible and non-toxic, yet has photo-physical properties that aid in tumour killing. While AuNCs accumulate in tumours by entry through their leaky vasculature, the lack of site-directed uptake leaves room for off target events. To improve the targeting and toxicity of nanomedicines, tumour-targeting ligands, like antibodies, can be attached as a drug delivery system.
This thesis describes the development of two ligation strategies to conjugate antibodies to AuNCs using strain-promoted azide-alkyne cycloadditions (SPAAC). SPAAC is a simple and robust bio-compatible conjugation approach, whereby a dibenzocyclooctyne (DBCO) bio-orthogonal functional group installed onto antibodies and can be ligated to azidated (N3)-AuNCs. Cysteine-maleimide ligation was used as a quick and effective means to install DBCO onto antibodies. N3-AuNCs were then ligated, and conjugation results were analyzed using SDS-PAGE, western blot, HPLC-SEC, flow-cytometry, and excitation-emission matrix measurements. Ongoing work includes a second strategy where DBCO is installed onto pre-made N-glycans to enable glyco-engineering by ENGase trans-glycosylation to antibodies. This work demonstrates the feasibility of conjugating antibodies to AuNCs and describes methods to characterize these products, thus enabling future work in examining Ab-AuNCs as cancer therapeutics.
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Queen's University's Thesis/Dissertation Non-Exclusive License for Deposit to QSpace and Library and Archives Canada
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Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.
Attribution-NonCommercial-NoDerivatives 4.0 International
ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.
Attribution-NonCommercial-NoDerivatives 4.0 International