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Development of a Novel Bioconjugation Platform for the Generation of Homogenous Antibody-Drug Conjugates


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Type

Thesis

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Authors

Dannheim, Friederike  ORCID logo  https://orcid.org/0000-0003-4651-0850

Abstract

Antibody-drug conjugates (ADCs) are therapeutic entities which leverage the specificity of antibodies to selectively deliver potent cytotoxins to specific cell types, such as cancer cells. The pharmacology of an ADC is critically dependent on its stability, homogeneity, and drug-to-antibody ratio (DAR) – all of which are controlled by the chemistry used to attach the drug to the antibody. Thus, to optimise these ADC characteristics, significant effort has been invested in the development of bioconjugation methods which yield homogenous ADCs with stable linkages and well-defined modification sites. However, despite many recent advances in the field, significant limitations remain. For instance, many methods necessitate the use of antibody engineering which typically requires laborious case-by-case optimisation. Additionally, efficient modulation of DAR in integer increments while maintaining homogeneity and stability remains exceptionally challenging. The primary aim of this work was the establishment of a linker technology that yields homogenous, stable, and functional ADCs from native antibodies. The secondary aim of this work was the derivatisation of these linkers to allow for the facile modulation of DAR in integer increments. Both of these goals could be achieved by the development of a novel class of disulfide rebridging linkers, based on the previously reported divinylpyrimidine (DVP) motif. This report details the development of a novel set of disulfide rebridging linkers, which contain four DVP motifs (termed ‘TetraDVPs’). The TetraDVP motif can conjugate to eight distinct cysteine residues, allowing simultaneous rebridging of all four interchain disulfides in an IgG1 antibody with a single linker molecule, a strategy which generates antibody conjugates with excellent homogeneity. Derivatisation of the initial TetraDVP linker with varying numbers of payload attachment handles enabled facile modulation of drug loading. Thus, antibody conjugates were functionalised with varying numbers of biologically relevant moieties (e.g. fluorophores, cytotoxins) through bioorthogonal chemistry. Assessment of the biological activity of the resultant conjugates demonstrated exceptional stability in human plasma along with potent and selective cytotoxicity in a series of cell-based assays.

Description

Date

2021-08-13

Advisors

Spring, David

Keywords

antibody-drug conjugates, bioconjugation

Qualification

Doctor of Philosophy (PhD)

Awarding Institution

University of Cambridge
Sponsorship
BBSRC (1942966)