Expanding oligosaccharide synthesis to progress the targeted delivery of therapeutics to the lungs

O'Sullivan, Joeseph (2023) Expanding oligosaccharide synthesis to progress the targeted delivery of therapeutics to the lungs. Doctoral thesis, Northumbria University.

Text (Doctoral thesis) osullivan.joseph_phd(18033752).pdf - Submitted Version Restricted to Repository staff only until 22 June 2025. Download (32MB) | Request a copy

Abstract

For over 30 years, nucleic acid therapeutics (NATs) promised a revolution in the field of medicine, offering unique and personalised therapeutics to otherwise undruggable targets; their clinical use, however, has been restricted by their inability to access target cell cytosols. A breakthrough came in attaching these therapies to a glycoside-based targeting ligand, N-acetylgalactosamine (GalNAc), which invokes receptor-mediated endocytosis into hepatocyte cytosols. This successfully translated into several FDA-approved drugs, saturating the market for hepatic targeting, and revealing the potential in glycoside-based targeting in the body. Delivery to alternative target cell types, such as lung epithelia, is still a bottleneck with current systems unable to avoid the immune response and reach target cytosols.

Schistosoma mansoni (S. mansoni) is a parasite that uses humans as hosts. During its lifecycle, it resides in the lungs, externally presenting glycoconjugates with an affinity for the surfactant protein D (SP-D), found throughout the pulmonary system. The two schistosomal glycoside-ligands with the greatest binding profile for SP-D are F-Gn (fucosylated N-acetyl-D glucosamine) and F-LDN (fucosylated-di-N-acetylated lactosamine). Seeking to expand the GalNAc paradigm for the liver, the aim of this project was to evaluate if these terminal glycosides may act as targeting ligands for lung epithelial cells, to mimic the parasite entry mechanism, and deliver therapeutics to epithelial cell cytosols, thereby resolving targeting for the lungs.

The synthetic assembly of the ligands resulted in the preparation of several isomerically pure substrates required to form the ligands. In pursuit of the biocatalytic assembly of the ligands, the expression of a microbial transmembrane glycosyltransferase (GT) was achieved in Escherichia coli (E. coli). Its incorporation into outer membrane vesicles (OMVs) yielded competitive substrate depletion indicating substrate binding by the allogenic GT, identifying this bioproduction method as a novel solution for enabling laboratory use of membrane-embedded proteins. This technology could unlock the potential use of numerous currently inaccessible GTs and other membrane-embedded proteins, for their scalable production and use in biocatalysis.

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