Hetero-bivalent GLP-1/glibenclamide for targeting pancreatic β-cells.

G protein-coupled receptor (GPCR) cell signalling cascades are initiated upon binding of a specific agonist ligand to its cell surface receptor. Linking multiple heterologous ligands that simultaneously bind and potentially link different receptors on the cell surface is a unique approach to modulate cell responses. Moreover, if the target receptors are selected based on analysis of cell-specific expression of a receptor combination, then the linked binding elements might provide enhanced specificity of targeting the cell type of interest, that is, only to cells that express the complementary receptors. Two receptors whose expression is relatively specific (in combination) to insulin-secreting pancreatic β-cells are the sulfonylurea-1 (SUR1) and the glucagon-like peptide-1 (GLP-1) receptors. A heterobivalent ligand was assembled from the active fragment of GLP-1 (7-36 GLP-1) and glibenclamide, a small organic ligand for SUR1. The synthetic construct was labelled with Cy5 or europium chelated in DTPA to evaluate binding to β-cells, by using fluorescence microscopy or time-resolved saturation and competition binding assays, respectively. Once the ligand binds to β-cells, it is rapidly capped and presumably removed from the cell surface by endocytosis. The bivalent ligand had an affinity approximately fivefold higher than monomeric europium-labelled GLP-1, likely a result of cooperative binding to the complementary receptors on the βTC3 cells. The high-affinity binding was lost in the presence of either unlabelled monomer, thus demonstrating that interaction with both receptors is required for the enhanced binding at low concentrations. Importantly, bivalent enhancement was accomplished in a cell system with physiological levels of expression of the complementary receptors, thus indicating that this approach might be applicable for β-cell targeting in vivo.