Abstract
Type 2 Diabetes Mellitus (T2DM) is characterized by pancreatic amyloid aggregates composed of islet amyloid polypeptide (IAPP; also called amylin), a small, intrinsically disordered peptide hormone. Pancreatic β-cell death due to membrane disruption mediated by non-amyloid or pre-amyloid oligomers of IAPP is thought to be central to the progression of T2DM. However, the rational design and screening of small-molecule and peptide ligands targeting these oligomeric states of IAPP is challenging using current pharmacological and biophysical methods. We recently used intermolecular single-pair Forster resonance energy transfer (spFRET) combined with constrained Monte Carlo simulations implemented in Rosetta to probe the heterogeneous, dynamic and transient ensemble of IAPP oligomers and define structural models for a family of membrane-bound IAPP dimers [Nath, Miranker & Rhoades. (2011) Angew. Chemie. Intl. Ed. 50(46):10859]. Here we describe efforts, based on this novel structural insight, to design and identify small molecules and peptides that bind selectively to particular configurations of IAPP dimers, and thereby bias the oligomeric ensemble so as to alter membrane-binding and subsequent toxicity mediated by higher-order states. Small molecules were identified from large commercial libraries (N > 100000) using a virtual screening protocol incorporating computational docking and statistical inference, while short, selective peptides were generated using RosettaDesign. The effects of these compounds on IAPP membrane-binding, conformation and cytotoxicity were then characterized using fluorescence correlation spectroscopy (FCS), spFRET and cell viability assays. Active modulators could serve as valuable tools in mechanistic studies of membrane disruption mediated by IAPP, and might also function as leads for new therapeutics targeting T2DM. In addition, the strategies developed here could serve as a framework for efforts targeting other clinically-relevant amyloidogenic intrinsically disordered proteins such as α-synuclein, amyloid-β and tau.
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