Development of proteolytically stable N-methylated peptide inhibitors of aggregation of the amylin peptide implicated in type 2 diabetes

Abstract

Islet amyloid polypeptide, also known as amylin, is the main component of the amyloid deposits present in approximately 90% of people with type 2 diabetes mellitus (T2DM). In this disease, amylin aggregates into multimeric β-pleated sheet structures which cause damage to pancreatic islet β-cells. Inhibitors of early-stage amylin aggregation could therefore provide a disease-modifying treatment for T2DM. In this study, overlapping peptides were designed to target the ‘binding’ region (RLANFLVHSS, residues 11–20) of human amylin, and their effects on amyloid fibril formation were determined by thioflavin-T assay. The first generation peptides showed less than 50% inhibition of aggregation, but a second generation peptide (H 2 N-RGANFLVHGR-CONH 2 ) showed strong inhibitory effects on amylin aggregation, and this was confirmed by negative stain electron microscopy. Cytotoxicity studies revealed that this peptide protected human pancreatic 1.4E7 (ECACC 10070102) insulin-secreting cells from the toxic effects of human amylin. Unlike the retro-inverso version of this peptide, which stimulated aggregation, two N-methylated peptides (H 2 N-RGAmNFmLVmHGR-CONH 2 and H 2 N-RGANmFLmVHmR-CONH 2 ) gave very clear dose-dependent inhibition of fibril formation. These two peptides were also stable against a range of different proteolytic enzymes, and in human plasma. These N-methylated peptides could provide a novel treatment for slowing progression of T2DM.