Superselective cell targeting through multivalent lectin-glycan interactions

All cells are covered in a forest of diverse carbohydrates structures known as the glycocalyx. There are many types of cell-surface carbohydrates (glycans) - some are long linear polymer chains, while others have short highly branched architectures like small trees. The composition of the glycocalyx is different for different types of cells - while the range of glycan structures present can be similar, the relative quantities of each glycan can vary considerably from one type of cell to another. Glycocalyx composition also changes if a cell becomes cancerous, and so measuring the composition of a glycocalyx presents opportunities for cancer diagnosis. Currently, the only way to measure how much of each type of glycan is present on a cell is to chop them off the cell, and weigh them individually in a mass spectrometer.

The aim of this research project is to develop molecular tools that can be used to quantify how much of a particular glycan is present on an intact cell surface, and to bind with high selectivity to cells that have a particular glycan composition. These probes will have applications in understanding biological processes, and could ultimately be used as medical diagnostics and for targeted delivery of drugs to specific cell types.

So how do you differentiate between two cells that have the same set of cell surface molecules, and differ only in the relative abundance of those molecules? Traditional probes like antibodies usually bind with high affinity to only one or two copies of their target molecule. They can be used to tell if a specific type of molecule is present on a cell surface, but not to bind selectively in response to a specific density of their target molecules.

Density-dependent 'superselective' binding requires a different strategy that is inspired by glycobiology - the biology of carbohydrates. Carbohydrate-binding proteins often interact relatively weakly with their target glycan and strong interactions are achieved by having many copies of the glycans and glycan-binding proteins interacting with one another in concert - so-called multivalent binding. In this way, many weak interactions come together to enhance binding strength, but it also greatly enhances the selectivity of binding.

Here we will develop multivalent probes that can bind in a density-dependent manner to cell surface glycans. We will develop probes that can distinguish between cancerous and healthy cells, and probes that can be used to map out complex net-like glycocalyces that regulate the function of neuronal cells. The methods developed will have much broader application for highly specific binding to target cells in both biology and medicine.

The cellular glycan coat, the glycocalyx, is essential for multicellular life and instrumental for cells to communicate with their environment. Rapid growth in the understanding of how glycan profiles change through development and oncogenesis provide an unprecedented opportunity to target cells by their glycan density profile and to visualise glycocalyces with high contrast for their roles in biological processes. This project aims to develop 'superselective' glycocalyx probes that are not only able to recognise a specific glycan epitope, but additionally are also exquisitely sensitive to the density of glycans expressed on the cell surface, as a broadly applicable technology for the biosciences and bioimaging. Carbohydrate binding proteins (lectins) will serve as glycan recognition units, and multivalent lectin-on-polymer probes will be assembled from a library of clickable lectin modules, polymer scaffolds and imaging labels. Bio-orthogonal conjugation chemistry will enable a modular 'plug and play' approach to prepare probes with quantitatively tuneable valency and size, and glycan specificity, and emerging concepts from soft matter physics tools will facilitate the rational design of probes with the desired superselective glycan binding properties. We aim to demonstrate the feasibility of superselective glycan targeting, and the benefits over conventional targeting, in selected applications of relevance in the biosciences, namely, the sharp discrimination of cancer cells over healthy cells, high-contrast visualisation of neuronal glycocalyces, and spatial mapping of glycan epitope distributions in the endothelial glycocalyx. The project will provide a new 'toolbox' for glycobiologists to analyse glycocalyces, and for cell biologists to target and discriminate cells by their glycan density profiles, with potential long-term impact for the development of new diagnostic and therapeutic probes in biomedical applications that require superselective cell recognition.