Synthesis and biological evaluation of mycobacterium tuberculosis derived glyconjugates and glycosylated nanoparticles

Abstract

This thesis is entitled 'Synthesis and biological evaluation of Mycobacterium tuberculosis derived glycoconjugates and glycosylated nanoparticles' and is composed of 6 chapters. Chapter 1 begins with an overview of the biology of Mycobacterium tuberculosis (M. tb), the causative agent of the disease Tuberculosis (TB). Its effect on the host immune system and the current treatments available for the disease are covered. The next section details the role of phenolic glycolipids (PGLs) and the related glycans, the para-hydroxybenzoic acid derivatives (p-HBADs) in M. tb infection, followed by their biosynthesis and previously reported syntheses of these glycans. Following this, the successful application of carbohydrate-based vaccines is covered, along with significant advancements in the development of carbohydrate-based nanoparticle vaccine candidates. The next section introduces the trehalose disaccharide and its role in M. tb infection. Particular emphasis is placed on the synthesis of unnatural trehalose derivatives for employment in numerous biological applications. The final section provides a summary of the concept of carbohydrate based enzymatic probes for the detection of glycosidase enzymes. Following this, some recent examples of nanoparticle based probes are outlined, prior to a brief discussion of the detection of the trehalase enzyme. This chapter concludes with an overview of the work presented within this thesis. The first part of this work, comprising of Chapters 2 and 3, details the development of p-HBAD functionalised gold nanoparticles (AuNPs) as potential vaccine candidates for M. tb. Chapter 2 begins with a discussion of key concepts in the chemical synthesis of carbohydrates. Subsequently, this chapter focuses on the synthesis of the p-HBAD glycans and related glycoconjugates suitable for AuNP conjugation. The synthetic approach utilised for the preparation of these glycoconjugates involves the use of multiple protecting group manipulations and optimisation of glycosylation conditions to achieve the native methylation patterns and stereochemistry of the anomeric positions of the p-HBAD glycans. Chapter 3 begins with a summary of previous work reported on the immunomodulatory activities of the p-HBAD glycans, followed by a brief introduction into the employment of AuNPs for biological applications. The investigation into the immunomodulatory effect of the p-HBAD glycoconjugates, synthesised in Chapter 2, on murine cytokine production is also presented. Subsequently, the synthesis of p-HBAD functionalised AuNPs, which was carried out by Dr. Finn Purcell-Milton and Prof. Yurii Gun?ko in the School of Chemistry, TCD is briefly discussed. Finally, preliminary investigations into the biological evaluation of the p-HBAD functionalised AuNPs is detailed. The second part of this work, covered in Chapter 4, outlines the use of trehalose functionalised quantum dots (QDs) for the development of a fluorescent enzyme sensor for detection of trehalase activity. This chapter begins with a brief overview of QDs and their utilisation as nanoprobes for the detection of enzymatic activity. Following this, the design, synthesis, evolution and optimisation of a library of trehalose derivatives suitable for QD functionalised is discussed. Subsequently, the QD functionalisation, carried out by Vera Kuznetsova, Anastasia Visheratina and Prof. Yurii Gun?ko in the School of Chemistry, TCD, is briefly discussed. This chapter concludes with the preliminary evaluation of the ability of the trehalose functionalised QDs to serve as fluorescent probes for observing trehalase activity. Chapter 5 concludes the work described in this thesis and outlines future work. Lastly, Chapter 6 details the procedures used for the cellular studies, experimental procedures and characterisation of all compounds prepared within this thesis.