Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.747204
Title: Pyridazinediones : versatile scaffolds for site-selective protein modification
Author: Lee, Maximillian
ISNI:       0000 0004 7229 022X
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2018
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Abstract:
Disulfide bonds represent an important target for site-selective protein modification, particularly via the strategy of functional re-bridging. Reduction of interchain disulfide bonds, followed by their re-bridging allows proteins to be functionalised in a site-selective manner whilst retaining the stability and integrity offered by the original bridge. This work describes the design and development of two distinct pyridazinedione-based technologies that, through the conduit of functional disulfide re-bridging, enable the synthesis of antibody – drug conjugates with hitherto unmet levels of control and homogeneity. As proteins often contain multiple disulfide bonds that are critical to conformation and stability, reagents that allow functional disulfide re-bridging without disulfide scrambling (non-native disulfide re-bridging) in multiple disulfide containing systems are critical for the success of this method. The first presented technology is a molecule that is capable of both reducing and re-bridging disulfide bonds, enabling a rapid and efficient one-reagent protocol for the functionalisation of disulfide containing proteins, moreover, it does so in such a way that native disulfide configuration is retained via a high local concentration effect. This novel pyridazinedione scaffold has been shown to functionalise a variety of therapeutically relevant proteins, including the widely used mAb HerceptinTM, enabling the synthesis of homogenous antibody – drug conjugates from a native mAb. Shifting focus from homogeneity to control over drug loading, the second presented technology is a single pyridazinedione-based molecule that contains four cysteine reactive centres and only one bioorthogonal reactive handle, which enables the generation of antibody conjugates with a loading of two modules. A loading of two is desirable for many reasons, especially in the context of large, hydrophobic payloads, which are increasingly popular for use in antibody-drug conjugates. A loading of two drugs per antibody has been shown to provide an optimal balance between efficacy and biophysical properties in many cases. A reliable method based on a native antibody scaffold without the use of enzymes or harsh oxidative conditions has hitherto not been achieved. The use of native antibodies has several advantages in terms of cost, practicality, accessibility and time. Thus, a novel, reliable method of furnishing antibody conjugates with a loading of two modules starting from a native antibody scaffold was developed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.747204  DOI: Not available
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