The construction of Anglo-Indian spaces in middlebrow works of Raj fiction, 1880-1914

Although bestsellers in their time, works of Raj fiction produced by women such as Maud Diver, Sara Jeannette Duncan, Alice Perrin, and Bithia May Croker have since fallen into obscurity with the general reading public and as a source of academic inquiry. This thesis readdresses this oversight through a reading of women-authored works of Raj fiction, a field of colonial literature that this thesis identifies as being fiction produced between 1857-1949 that in some way is related to the British Raj. The thesis suggests that in their depiction of the spaces of Anglo-Indian life these works shaped impressions of the Raj, but also contributed towards the maintenance of the Empire. Analysis of the imaginative organisation of space reveals both the limitations and nuances of Anglo-Indian women’s lives and the ideological expectations of Anglo-Indian femininity which is encoded through these spaces. Through an acknowledgement that these texts are part of the conservative middlebrow genre, the thesis argues that Raj fiction functioned as a form of anxiety management, containing and resolving anxieties around about the place of European women in the Raj and of a broader imperial decline. Each chapter of the thesis focuses on key spatial locations in this fiction. The Anglo-Indian home of the bungalow was considered as to be the responsibility for Anglo-Indian women and a space where fears of transgression may be depicted and then assuaged by the narrative. The traditionally masculine club space is decentralised from its emblematic position in Anglo-Indian life and is instead read as one that must be carefully navigated by female characters. The summer capital of Simla has similar connotations to the club social scene, but the feminine reputation of that city allows a more stable imagined space to be constructed. The picnic emerges as a space from which anxieties of decline emerge; the temporary nature of the space allows for more permanent spaces to function as sites of resolution. The imperial centre of England is read as a space that can accommodate and possibly resolve anxieties that are disruptive in the less stable space of the Raj. An analysis of these specific, reoccurring spaces allows for both an alternate conception of the Raj to emerge as well as offering a method for analysing the anxiety-management function of the Raj fiction genre

Bioconjugation of Green Fluorescent Protein via an Unexpectedly Stable Cyclic Sulfonium Intermediate

Bioconjugation of superfolder GFP involving the formation of an unusually stable, and unprecedented, cyclic sulfonium species is described. This sulfonium can undergo smooth reaction with a range of nucleophiles to give sulfur-, selenium- and azide-modified GFP derivatives in high conversions

A novel synthetic chemistry approach to linkage-specific ubiquitin conjugation.

Ubiquitination is of great importance as the post-translational modification of proteins with ubiquitin, or ubiquitin chains, facilitates a number of vital cellular processes. Herein we present a facile method of preparing various ubiquitin conjugates under mild conditions using michael acceptors based on dibromo-maleimides and dibromo-pyridazinediones

A plug-and-play approach to antibody-based therapeutics via a chemoselective dual click strategy.

Although recent methods for the engineering of antibody-drug conjugates (ADCs) have gone some way to addressing the challenging issues of ADC construction, significant hurdles still remain. There is clear demand for the construction of novel ADC platforms that offer greater stability, homogeneity and flexibility. Here we describe a significant step towards a platform for next-generation antibody-based therapeutics by providing constructs that combine site-specific modification, exceptional versatility and high stability, with retention of antibody binding and structure post-modification. The relevance of the work in a biological context is also demonstrated in a cytotoxicity assay and a cell internalization study with HER2-positive and -negative breast cancer cell lines

A novel approach to the site-selective dual labelling of a protein via chemoselective cysteine modification

Local protein microenvironment is used to control the outcome of reaction between cysteine residues and 2,5-dibromohexanediamide. The differential reactivity is exploited to introduce two orthogonal reactive handles onto the surface of a double cysteine mutant of superfolder green fluorescent protein in a regioselective manner. Subsequent elaboration with commonly used thiol and alkyne containing reagents affects site-selective protein dual labelling

A mild TCEP-based para-azidobenzyl cleavage strategy to transform reversible cysteine thiol labelling reagents into irreversible conjugates

It has recently emerged that the succinimide linkage of a maleimide thiol addition product is fragile, which is a major issue in fields where thiol functionalisation needs to be robust. Herein we deliver a strategy that generates selective cysteine thiol labelling reagents, which are stable to hydrolysis and thiol exchange

Identification of an active metabolite of PAR-1 antagonist RWJ-58259 and synthesis of analogues to enhance its metabolic stability

The discontinuation of PAR-1 antagonist RWJ-58259 beyond use as a biological probe is most likely due to it's short half-life in vivo. However, retention of significant in vivo activity beyond the point where most of the RWJ-58259 had been consumed implies the generation of an active metabolite. Herein we describe the biological activity of a predicted metabolite of RWJ-58259 and the synthesis of analogues designed to enhance the metabolic stability of RWJ-58259

A Plug-and-Play Approach for the De Novo Generation of Dually Functionalized Bispecifics

Diseases are multifactorial, with redundancies and synergies between various pathways. However, most of the antibody-based therapeutics on the market interact with only one target, thus limiting their efficacy. The targeting of multiple epitopes could improve the therapeutic index of treatment and counteract mechanisms of resistance. To this effect, a new class of therapeutics has emerged: bispecific antibodies. Bispecific formation using chemical methods is rare and low-yielding and/or requires a large excess of one of the two proteins to avoid homodimerization and heterogeneity. In order for chemically prepared bispecifics to deliver their full potential, high-yielding, modular, and reliable cross-linking technologies are required. Herein, we describe a novel approach not only for the rapid and high-yielding chemical generation of bispecific antibodies from native antibody fragments, but also for the site-specific dual functionalization of the resulting bioconjugates. Based on orthogonal clickable functional groups, this strategy enables the assembly of functionalized bispecifics with controlled loading in a modular and convergent manner

Recent advances in the construction of antibody–drug conjugates

Antibody-drug conjugates (ADCs) are derived from antibodies covalently attached to highly potent drugs using a variety of conjugation and linker technologies. This class of therapeutic conceptually combines the exquisite specificity of antibodies (i.e. enabling discrimination between healthy and diseased tissue) with the cell-killing ability of potent cytotoxic drugs. This powerful and exciting class of targeted therapy has shown considerable promise in the treatment of various cancers with two US Food and Drug Administration (FDA) approved ADCs currently on the market (i.e. Adcetris™ and Kadcyla™) and ca. 40 ADCs currently undergoing clinical evaluation. However, in order for ADCs to deliver their full potential, sophisticated site-specific conjugation technologies to connect the drug to the antibody are vital. This perspective discusses the strategies currently used for the site-specific construction of ADCs and appraises their merits and disadvantages