TARGETING DIABETES: HIGH SPECIFICITY BINDING OF A YOHIMBINE LINKED GLP‐1 DIVALENT LIGAND TO PANCREATIC B‐CELLS

Abstract

Pancreatic β-cell dysfunction, and subsequent loss of β-cell mass, underlies the development of Diabetes Mellitus. A primary limitation in the analysis of β-cell mass is the lack of specificity of β-cell targeting agents. We propose that multivalent ligands (MVL), composed of two or more unique receptor binding moieties against β-cell targets, would increase β-cell specificity. To test this, we synthesized a ligand composed of Yohimbine (Yhb, antagonist to α2 adrenergic receptor) linked to Glucagon-like Peptide 1(GLP-1). Receptors to both ligands are expressed on βTC3 cells (β-cell model). Analysis by fluorescence microscopy showed rapid binding with ligand internalization in 2–3 min. From population based competition assays, the binding affinity for monovalent GLP-1 was ~30 nM, whereas GLP-1/Yhb exhibited two distinct binding sites, one of high affinity (6 pM) and another lower affinity 70 nM site. The high affinity binding is likely related to MVL crosslinking of the complimentary receptors causing a decrease in ligand off rate. The low affinity site is consistent with weak binding of the MVL GLP-1 moiety to spare GLP-1 receptors that are expressed at higher levels than α2-AR in βTC3 cells. Our findings suggest that GLP-1/Yhb will provide enhanced targeting to β-cells due to decreased binding to cells that express only one receptor of the complimentary pair coupled with enhanced binding and retention to β-cells. Supported by: Juvenile Diabetes Research Foundation, and the Arizona Biomedical Research Commission