Bioorganic & Medicinal Chemistry Letters
Structure–activity relationships of oxysterol-derived
pharmacological chaperones for Niemann–Pick type C1 protein
a
a
b
a
Kenji Ohgane a,b, , Fumika Karaki , Tomomi Noguchi-Yachide , Kosuke Dodo , Yuichi Hashimoto
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a Institute of Molecular and Cellular Biosciences, The University of Tokyo, 1-1-1, Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan
b RIKEN, 2-1, Hirosawa, Wako-shi, Saitama 351-0198, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Niemann–Pick disease type C is a fatal neurodegenerative disease, and its major cause is mutations in
NPC1 gene. This gene encodes NPC1 protein, a late endosomal polytopic membrane protein required
for intracellular cholesterol trafficking. One prevalent mutation (I1061T) has been shown to cause a fold-
ing defect, which results in failure of endosomal localization of the protein, leading to loss-of-function
phenotype. We have previously demonstrated that several oxysterols and their derivatives act as phar-
macological chaperones; binding of these compounds to NPC1I1061T mutant protein corrects the localiza-
tion/maturation defect of the mutant protein. Here, we disclose detailed structure–activity relationships
of oxysterol derivatives as pharmacological chaperones for NPC1I1061T mutant.
Received 24 April 2014
Revised 15 May 2014
Accepted 17 May 2014
Available online xxxx
Keywords:
Niemann–Pick disease type C
NPC1
Pharmacological chaperone
Oxysterol
Ó 2014 Elsevier Ltd. All rights reserved.
Structure–activity relationships
Niemann–Pick disease type C is a fatal, heritable neurological
disorder characterized by massive accumulation of cholesterol
and other lipids in the late endosomal compartment.1 Mutation
in the NPC1 gene,2 which encodes late endosomal membrane pro-
tein NPC1, is carried by about 95% of patients.3–5 This protein binds
cholesterol6–9 and is essential for intracellular cholesterol traffick-
ing; thus, loss of function of the protein leads to late endosomal
accumulation of cholesterol derived from endocytosed low-density
lipoprotein (LDL). A major and well-characterized mutation in
NPC1, I1061T (Ile1061 to Thr),10 causes loss of function due to fold-
ing defect and instability, though the intrinsic function of the
protein is retained.11,12 In general, folding-defective membrane
proteins are picked up by the cellular quality control system at
the endoplasmic reticulum (ER), and are retained in the ER and
degraded by proteasome through a process called ER-associated
degradation (ERAD). I1061T mutant proteins are indeed retained
and rapidly degraded in the ER, resulting in loss of functional
NPC1 in late endosomes.11
degradation via ERAD, and assists their exit from the ER. As rescued
mutant proteins are, in most cases, at least partially functional
at the correct location, overall cellular function can be restored
by correction of mislocalization, mediated by pharmacological
chaperones.
We previously reported that 25-hydroxycholesterol (1, 25HC,
Fig. 1), mo56HC (2), and some other oxysterol derivatives including
mo56CFA (3) act as pharmacological chaperones for NPC1 protein.
These compounds corrected the folding-defective phenotypes of
NPC1I1061T mutant proteins, including localization/maturation
defect, instability, and endosomal cholesterol accumulation in
patient-derived fibroblasts, via direct binding to a putative second
sterol-binding site24,25 distinct from the N-terminal domain,7,8,26,27
which is a well-characterized sterol-binding site on NPC1. Here, we
disclose detailed structure–activity relationships of oxysterol
derivatives as pharmacological chaperones for NPC1I1061T mutant
protein.
In our previous report,24 we evaluated the relative efficacy of
the sterol derivatives by quantitatively analyzing the altered local-
ization of NPC1 proteins (Fig. 2B), by means of colocalization
analysis with late-endosomal marker LAMP1. However, the
colocalization analysis requires time-consuming immunocyto-
chemical staining of LAMP1, and is not suitable for structure–
activity relationship studies, where large numbers of derivatives
need to be assayed. In order to increase throughput in the present
work, we used a phenotypic analysis instead of colocalization anal-
ysis.23 As shown in Figure 2A, the localization pattern of NPC1I1061T
mutant protein was classified on a cell-by-cell basis into three
Since the late 1990s, accumulating evidence has shown that
loss-of-function phenotypes of folding-defective mutants can be
corrected by using selective, small-molecular ligands.13–23 These
molecules, called pharmacological chaperones, are considered to
rescue folding-defective mutant proteins via direct binding to the
folding intermediates in the ER.13 Binding of the pharmacological
chaperone stabilizes mutant proteins, thus preventing their
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Corresponding author. Tel.: +81 3 5841 7848; fax: +81 3 5841 8495.
0960-894X/Ó 2014 Elsevier Ltd. All rights reserved.