Projekte

The dipteran glycan array

Interdisciplinary project (chemistry - glycobiology) with Prof. Iain Wilson

The project combines analytical and chemical synthesis expertise to identify functional glycomotifs for important model organisms or pathogen vectors, including fruit flies, tsetse flies and mosquitoes. The aim of the project is to develop a new microarray reflecting the glycome of dipteran insects based on a large set of natural, redesigned and chemically synthesised oligosaccharide probes.

To investigate whether N-, O- and lipid-linked insect glycans with anionic and/or zwitterionic antennal modifications are bound by mammalian immune lectins that recognise glycans on mosquito-derived viruses and are involved in host-pathogen interactions and thus have specific innate immune functions, we will chemically synthesise a series of complex biomolecules comprising di-, tri- and tetrasaccharides substituted with anionic (sulphates) or zwitterionic (such as phosphoethanolamine) modifications. The oligosaccharides will be chemically linked to an alkylamino spacer group to allow printing onto NHS- functionalised glass slides.

Synthetic glycans for exploration of lipopolysaccharide (LPS) recognition by the proteins of innate immune system

The Toll-like receptor 4 signaling pathway plays a central role in the prompt defence against pathogens, providing an immediate response to infectious challenge. The TLR4/MD-2 complex can recognise and respond to various pathogen-associated molecular patterns (PAMPs) with bacterial lipopolysaccharide (LPS) being the most potent and commonly encountered activator of the TLR4-mediated inflammation. TLR4 is considered to be both a friend and foe, as improperly regulated TLR4 signaling can lead to over-activation of the immune response, resulting in sepsis, acute lung injury or pathological chronic inflammation implicated in cancer and autoimmune diseases. The dual action of the TLR4 complex justifies efforts to develop both TLR4 antagonists as anti-sepsis drug candidates or remedies for chronic inflammatory diseases and TLR4 agonists as vaccine adjuvants or immunotherapeutics.

Using diversity-oriented synthesis, we will assemble a library of LPS mimicking molecules, although structurally simpler than the microheterogenic bacterial lipopolysaccharide. Synthetic molecules will be diversified by specific chemical modifications, such as the attachment  of differently substituted phosphate groups and long-chain β-hydroxyacyl residues, to achieve better affinity for the respective innate immune receptor and to improve immunobiological activities. The structure-activity relationships generated by biological screening of synthetic probes will refine and improve our understanding of the molecular basis of host-pathogen interaction and would pave the way for the development of new immunotherapeutic agents.

Synthetic glycolipids for studying inflammation and sepsis

Therapeutic immunomodulation through Pattern Recognition Receptors (PRRs) is a promising strategy for the treatment of inflammatory diseases ranging from chronic inflammation, allergy and asthma to acute inflammatory conditions such as antibiotic-resistant infections and sepsis syndrome.

The aim of the project is to design and chemically synthesise a library of carbohydrate-based biomolecules capable of interacting with the innate immune receptors specific for Gram-negative bacterial lipopolysaccharide. The synthetic glycans and glycolipids will be designed to target both the transmembrane and cytosolic LPS receptors, TLR4 and caspase-4/11 respectively, with particular emphasis on the 3-D molecular shape of the protein-bound ligands. Synthetic TLR4 ligands will be chemically assembled based on the rigidified disaccharide scaffolds, which will be chemically modified by attachment of various functional groups. In addition to competitive TLR4 and caspase-4/11 antagonists (which can inhibit LPS-induced inflammation), partial TLR4 agonists will also be developed, which can bind to and activate TLR4, but cannot elicit the maximal response induced by LPS. In the presence of a natural TLR4 agonist, LPS, the partial agonist will act as an antagonist, competing with LPS for the same binding pocket on MD-2, thereby abolishing TLR4 activation and the overwhelming inflammatory response. At the same time, exposure of the TLR4/MD-2 complex to the partial agonist will ensure a constant, low level of activity, resulting in a moderate immune stimulation, potentially overcoming the consequences of sepsis-induced immunosuppression and endotoxin tolerance.

Hunting and exploiting enzymes which synthesise bacterial and parasite
immunomodulatory zwitterionic glycans

Interdisciplinary project (chemistry - glycobiology) with Prof. Iain Wilson

The 'sugar' coating of any organism, consisting of a variety of glycoconjugates, is the first point of interaction between bacteria, viruses or parasites and host cells. Bacterial surface polysaccharides contain many unusual modifications, including non-sugar moieties such as phosphorylcholine (PC) - which is also found on the glycan chains of glycoproteins and glycolipids of helminth parasites, as well as glycans produced by insect cell lines used as cell factories for recombinant proteins, including vaccines. Despite many reports that PC is immunomodulatory, the biosynthesis and interactions of PC-modified glycans are poorly understood. This project will generate chemical tools to fill this knowledge gap.

Potential PC-transferring enzymes will be expressed (Wilson group) and new substrates and their PC-modified products will be chemically synthesised (Zamyatina group) to study them. Based on this new knowledge, we will use these enzymes to create new array-based probes for testing interactions with immune system proteins, including pentraxins and antibodies.

The proposed combination of chemical and enzymatic synthesis and glycan array technology will enable us to gain new insights into how and why PC modifications of pathogens, whether bacteria or parasites, are recognised by the host immune system. We can also begin to explore the role of PC in modulating the immune system, including responses to vaccines.