Design of a mimic of nonamyloidogenic and bioactive human islet amyloid polypeptide (IAPP) as nanomolar affinity inhibitor of IAPP cytotoxic fibrillogenesis

doi:10.1073/pnas.0507471103

Design of a mimic of nonamyloidogenic and bioactive human islet amyloid polypeptide (IAPP) as nanomolar affinity inhibitor of IAPP cytotoxic fibrillogenesis

Laboratory of Bioorganic and Medicinal Chemistry, Institute of Biochemistry, University Hospital of the Rheinisch–Westfälische Technische Hochschule Aachen, Pauwelstrasse 30, D-52074 Aachen, Germany

Edited by William F. DeGrado, University of Pennsylvania School of Medicine, Philadelphia, PA, and approved December 15, 2005 (received for review August 26, 2005)

Abstract

Protein aggregation into cytotoxic oligomers and fibrils in vivo is linked to cell degeneration and the pathogenesis of >25 uncurable diseases, whereas the high aggregation propensity and insolubility of several bioactive polypeptides and proteins in vitro prevent their therapeutic use. Aggregation of human islet amyloid polypeptide (IAPP) into pancreatic amyloid is strongly associated with the pathogenesis of type II diabetes. IAPP is a 37-residue polypeptide that acts as a neuroendocrine regulator of glucose homeostasis. However, IAPP misfolds and self-associates into cytotoxic aggregates and fibrils even at nanomolar concentrations. Because IAPP aggregation causes β-cell death and prohibits therapeutic application of IAPP in diabetes, we pursued a minimalistic chemical design approach to generate a molecular mimic of a nonamyloidogenic and bioactive IAPP conformation that would still be able to associate with IAPP and thus inhibit its fibrillogenesis and cytotoxicity. We show that the double N-methylated full length IAPP analog [(N-Me)G24, (N-Me)I26]-IAPP (IAPP-GI) is a highly soluble, nonamyloidogenic, and noncytotoxic IAPP molecular mimic and an IAPP receptor agonist. Moreover, IAPP-GI binds IAPP with low nanomolar affinity and completely blocks IAPP cytotoxic self-assembly and fibrillogenesis with activity in the low nanomolar concentration range. Importantly, IAPP-GI dissociates cytotoxic IAPP oligomers and fibrils and is able to reverse their cytotoxicity. Bifunctional soluble IAPP mimics that combine bioactivity with the ability to block and reverse IAPP cytotoxic self-assembly are promising candidates for the treatment of diabetes. Moreover, our amyloid disease inhibitor design concept may be applicable to other protein aggregation diseases.

Footnotes

*To whom correspondence should be addressed. E-mail: akapurniotu{at}ukaachen.de
Author contributions: A. Kapurniotu designed research; L.-M.Y., M.T.-N., A.V., A. Kazantzis, and A. Kapurniotu performed research; L.-M.Y., M.T.-N., A.V., A. Kazantzis, and A. Kapurniotu analyzed data; and A. Kapurniotu wrote the paper.
Conflict of interest statement: No conflicts declared.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations:

Abbreviations:

IAPP,

human islet amyloid polypeptide;

IAPP-GI,

[(N-Me)G24, N-Me)I26]-IAPP;

ThT,

thioflavin T;

MTT,

3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide;

TEM,

transmission EM;

Fluos-IAPP,

Nα-amino-terminal fluorescein-labeled IAPP;

Fluos-IAPP-GI,

Nα-amino-terminal fluorescein-labeled IAPP-GI;

RIN 5fm,

cultured pancreatic rat insulinoma cells;

Dig-IAPP-GI,

amino-terminal digoxigenin-labeled IAPP-GI;

Biotin-IAPP,

Na-amino-terminal biotin-labeled IAPP.