Chemical intervention in heparan sulphate-dependent growth factor signalling systems using engineered heparin saccharides

Our labs have expertise in studying the structure and activities of the complex polysaccharide heparan sulphate, which is found on the surface of all cells, and in studying biological processes in cell and developmental biology. These sugars have considerable structural diversity, and bind to many specialised proteins, such as growth factors, responsible for controlling cell behaviour in all tissues. Our labs are interested in finding out more about the molecular basis for their actions and exploiting their exciting natural properties by using them as tools to chemically interfere in biological systems. In this project we are planning to exploit a new approach we have developed for creating libraries of HS sugars with diverse structures, to study the relationships between structure and activity for a number of growth factors. We will examine how differences in structure affect the ability of these sugars to alter growth factor activities in biological assays using test tube, cell and whole animal assays. This will allow us to carry out experiments in a biological system in which targeted intervention of a known pathway and its biological outcome will be conducted. We will collect and compare the data from screening assays to both determine structural specificity amongst diverse growth factors, and identify activating and inhibitory saccharide molecules for application as selective chemical intervention tools in biological systems. We will also use the information to design selected structures for full chemical synthesis and testing, with the aim of providing a supply of defined molecules for future intervention experiments. The ultimate goal will be to establish the application of engineered heparin saccharide as a powerful tool for revealing novel insights into the functional roles of specific HS sequences in regulation of biological systems. These studies will provide new information which will help us to understand how these sugar-protein interactions regulate the functions of cells. They fit into a number of areas within the broad Council remit of increasing understanding of how living organisms function (in this case, molecular interactions relevant to cell signaling and regulation) and providing knowledge which can be used to develop new technologies and products for medicine and industry.

Specific sequences in heparan sulfate (HS) determine selective protein binding and regulatory properties and are thus ideally designed as one of Nature's tools for chemical intervention. However, structure-activity studies have been hampered by limited amounts of active sequences available from natural HS. We have developed novel approaches for creating libraries of heparin-based structures 'engineered' to produce a high level of structural diversity, combined with new techniques for their further separation and analysis. This approach allows large numbers of sequences to be searched for activity for the first time, providing an opportunity to develop a generic 'chemical genetics' approach for rapid exploration of structure-activity relationships in biological systems. We will exploit this opportunity by generating a library of highly diverse HS analogue saccharides by chemical modification of poly- and oligo-saccharide heparin templates. Their activity will be screened using established in vitro binding and activity assays for 3 types of growth factors / FGFs, GDNF, and VEGF / and also using relevant ex vivo and in vivo assays in model systems (angiogenesis; kidney development) and whole organisms (C elegans and zebrafish). This will allow us to assess their potential use for targeted intervention of known signaling pathways and examination of the resulting biological outcomes. The data will be collated to determine both the level of structural specificity amongst these different growth factors and to identify activating and inhibitory saccharide molecules for application as selective chemical intervention tools in biological systems. The information gained will be used to prepare saccharide-nanoprobes for testing in biological assays, and also for the design of a limited number of relevant oligosaccharide structures for direct chemical synthesis using newly developed methods (in collaboration with Prof. Geert-Jan Boons, University of Georgia). The latter would provide highly defined chemical tools for intervention in biological systems. The ultimate goal will be to establish the application of engineered heparin saccharides as a powerful tool for revealing novel insights into the functional roles of specific HS sequences in regulation of biological systems.