Angewandte
Chemie
DOI: 10.1002/anie.200705372
Small-Molecule Inhibitors
Small-Molecule Inhibitors of Islet Amyloid Polypeptide Fibril
Formation**
Rajesh Mishra, Bruno Bulic, Daniel Sellin, Suman Jha, Herbert Waldmann, and Roland Winter*
Newly synthesized proteins in the cell adopt a functional
folded state resulting from a highly regulated process. Failure
to form this functional state leads to the degradation of
proteins inside the cell. However, under certain conditions,
some proteins can also adopt an alternative state by the
assembly of unfolded or partially folded monomers or protein
fragments into a b-sheet structure called amyloid fibril.[1] In
spite of arising from diverse amino acid sequences, they have
a similar fibrillar structure that binds the dye Congo red.[2]
These amyloids are involved in a number of devastating
diseases including Alzheimerꢀs disease, prion diseases, and
type-II diabetes mellitus.[3]
In the type-II diabetes, deposition of extracellular amyloid
plaques in pancreatic beta cells has been observed in humans.
Biochemical analysis of the plaques revealed the presence of
a 37-residue peptide called islet amyloid polypeptide (IAPP)
or amylin, which is co-secreted with insulin. It has a disulfide
bond between residues 2–7 and the C-terminus is ami-
dated.[4–6] IAPP is also known to interact with lipid mem-
branes which are able to induce and foster the fibril
formation.[7] Recently, a possible mechanism of IAPP fibril
formation at anionic lipid interfaces has been proposed in
which it has been shown that IAPP forms b-sheet-rich
amyloid fibrils via an intermediate a-helical state.[8] The
presence of IAPP amyloid finally leads to the apoptosis of
pancreatic beta cells.[9] However, it is still not clear whether
the fibrils themselves or their intermediate states are respon-
sible for the cell death.[10] In nature, IAPP amyloid fibril
formation can be prevented by altering the primary amino
acid sequence, such as in rat IAPP where three proline
residues, which are absent in the human IAPP, are thought to
prevent the amyloid fibril formation.[11] Recently, Kapurnio-
tuꢀs group succeeded in the synthesis of conformationally
constrained analogues of IAPP, which are methylated at
amide bonds and do not fibrillize.[12,13]
Inhibition of amyloid fibril formation is considered to be a
potentially key therapeutic approach towards diabetes and
other amyloid-related diseases.[14] Surprisingly, very little
attempt has been made to inhibit IAPP fibril formation by
small-molecule inhibitors.[15] Small-molecule inhibitors have
advantages over peptide inhibitors because they could more
easily cross the blood brain barrier, avoid immunological
response, and are more stable in biological fluids and
tissues.[16] In addition, the high flexibility of peptide inhibitors
may, for entropic reasons, prevent efficient binding. This
problem may be overcome by synthesis of conformationally
restricted peptides.[12,13] The bottleneck in the discovery of
small-molecule inhibitors of amyloid fibril formation is the
lack of structural information about amyloids. However, this
did not prevent the discovery of small-molecule inhibitors for
other amyloid fibrils such as, Ab[17,18] and tau,[19,20] which are
involved in Alzheimerꢀs disease.
In a recent study on a cellular model of tau aggregate
inhibition, two rhodanine-scaffold (2-thioxothiazolidin-4-
one) based inhibitors have been identified which have very
low cell toxicity.[21] These compounds were chosen because of
the presence of a rhodanine heterocyclic core, which is
biocompatible, non-mutagenic,[22] and has a drug-like profile.
Inspired by these results, we wanted to explore whether these
compounds will also inhibit amyloid fibril formation of IAPP
which has a completely different amino acid sequence but
shares a similar fibrillar morphology with the tau aggregate.
To our knowledge, this is the first study on such small-
molecule inhibitors of IAPP amyloid formation.
The compounds 1 and 2 (Figure 1) were synthesized as
described earlier.[21] Amyloid fibril formation was carried out
in 10 mm sodium phosphate buffer at pH 7.5 for 96 h. To
reveal the effect of the two potential inhibitors, different
concentrations of the compounds were added to the buffer
solution. Fibril formation was quantified by measuring the
fluorescence intensity of the amyloid-specific dye thioflavin T
(ThT) at a wavelength of 480 nm. The fluorescence intensity
of amyloid fibrils increases upon binding to ThT. The
efficiency of inhibition was monitored by measuring the
ThT fluorescence intensity with respect to that of pure IAPP
aggregate without inhibitor (100%). It is evident from
Figure 1b that both compounds have a marked inhibitory
effect. The concentration at which half of the fibril formation
is inhibited (IC50) is 1.23 mm for compound 1, and 0.45 mm for
compound 2. A similar trend has been observed for the
aggregation of tau, with IC50 values of 0.67 and 0.26 mm for
compounds 1 and 2, respectively.[21] From the results on tau
and IAPP aggregation it is clear that compound 2 is a more
[*] Dr. R. Mishra, D. Sellin, S. Jha, Prof. Dr. R. Winter
Faculty of Chemistry
Physical Chemistry I—Biophysical Chemistry
Technical University Dortmund
Otto-Hahn-Strasse 6, 44227 Dortmund (Germany)
Fax. (+49)231-755-3901
E-mail: roland.winter@uni-dortmund.de
Dr. B. Bulic, Prof. Dr. H. Waldmann
Max-Planck-Institute for Molecular Physiology
Department of Chemical Biology, and
Center for Applied Chemical Genomics
Otto-Hahn-Strasse 11, 44227 Dortmund (Germany)
[**] Financial support from the DFG, the Fonds der Chemischen
Industrie, the country NRWand the EU (Europäischer Fonds für
regionale Entwicklung) is gratefully acknowledged.
Supporting information for this article is available on the WWW
Angew. Chem. Int. Ed. 2008, 47, 4679 –4682
ꢀ 2008 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
4679