An orally available BACE1 inhibitor that affords robust CNS Aβ reduction without cardiovascular liabilities

Article abstract of DOI:10.1021/ml500458t

BACE1 inhibition to prevent Aβ peptide formation is considered to be a potential route to a disease-modifying treatment for Alzheimer's disease. Previous efforts in our laboratory using a combined structure- and property-based approach have resulted in th

Full text of DOI:10.1021/ml500458t

Letter
pubs.acs.org/acsmedchemlett
An Orally Available BACE1 Inhibitor That Affords Robust CNS Aβ
Reduction without Cardiovascular Liabilities
Yuan Cheng,*,† James Brown,† Ted C. Judd,† Patricia Lopez,† Wenyuan Qian,† Timothy S.∇Powers,†,△
Jian Jeffrey Chen,† Michael D. Bartberger,‡ Kui Chen,∥ Robert T. Dunn, II,# Oleg Epstein,
Robert T. Fremeau, Jr.,§ Scott Harried,†,□ Dean Hickman,⊥ Stephen A. Hitchcock,†,◆ Yi Luo,§
∇,¶
∇
Ana Elena Minatti,† Vinod F. Patel,
Hugo M. Vargas,# Robert C. Wahl,∥ Matthew M. Weiss,
Paul H. Wen,§ Ryan D. White, Douglas A. Whittington,○ Xiao Mei Zheng, and Stephen Wood§
∇
∇
†Department of Medicinal Chemistry, ‡Department of Molecular Structure, §Department of Neuroscience, ∥Department of HTS and
#
Molecular Pharmacology, and ⊥Department of Pharmacokinetics and Drug Metabolism, Comparative Biology and Safety Sciences,
Amgen Inc., One Amgen Center Drive, Thousand Oaks, California 91320, United States
∇Department of Medicinal Chemistry and ○Department of Molecular Structure, Amgen Inc., 360 Binney Street, Cambridge,
Massachusetts 02142, United States
S
* Supporting Information
ABSTRACT: BACE1 inhibition to prevent Aβ peptide
formation is considered to be a potential route to a disease-
modifying treatment for Alzheimer’s disease. Previous efforts
in our laboratory using a combined structure- and property-
based approach have resulted in the identification of
aminooxazoline xanthenes as potent BACE1 inhibitors.
Herein, we report further optimization leading to the discovery
of inhibitor 15 as an orally available and highly efficacious
BACE1 inhibitor that robustly reduces CSF and brain Aβ
levels in both rats and nonhuman primates. In addition, compound 15 exhibited low activity on the hERG ion channel and was
well tolerated in an integrated cardiovascular safety model.
KEYWORDS: β-site amyloid precursor protein cleaving enzyme 1 (BACE1), Alzheimer’s disease (AD), Aβ, aminooxazoline, xanthene
lzheimer’s disease (AD) is the most common neuro-
Despite extensive research efforts for over a decade, the
Adegenerative disorder and accounts for 50−80% of identification of brain penetrant and efficacious BACE1
inhibitors remains a major challenge. Early work focused on
peptidic transition state mimetics that met with limited
success.9 In recent years, a major advance in the design of
BACE1 inhibitors was the discovery of 2-amino heterocycles
that engage both active site aspartyl moieties of BACE1.10−15
These ligands possessed more rigid structural characteristics
and lower molecular weight, offering a promising opportunity
to develop potent, brain penetrant inhibitors. This approach
was supported by encouraging results in a recent report of a
nonpeptidic BACE1 inhibitor that exhibited a significant
reduction in central Aβ levels in preclinical species and healthy
volunteers in early stage clinical trials.16
Previously, we reported the discovery of potent aminooxazo-
line xanthene BACE1 inhibitors rationally designed using a
structure- and property-based approach.12 In the course of lead
optimization, compound 1 was identified as a potent BACE1
inhibitor.17 Compound 1 acutely lowered CSF and brain Aβ40
by approximately 80% and 61%, respectively, 4 h postdose
dementia cases identified each year. Current therapeutics for
AD only provide temporary symptomatic relief and do not
address the underlying neuropathology. A therapeutic treat-
ment that directly modifies the progression of the disease
remains one of the largest unmet medical needs in neuro-
biology.
The most widely held hypothesis for the underlying
pathogenesis of AD posits that aggregation of β-amyloid
peptides (Aβ) and deposition into amyloid plaques in the
neural parenchyma play an essential role. Aβ is produced by
sequential endoproteolytic cleavage of amyloid precursor
protein (APP) by the aspartyl protease β-site APP cleaving
enzyme-1 (BACE1) and γ-secretase.1,2 Inhibition of BACE1
decreases the production of all major forms of Aβ including the
most pathogenic species Aβ42,3,4 indicating that BACE1 is a
prime therapeutic target for the development of disease-
modifying therapies for AD.5−7 Genetic evidence published
recently has provided additional support for the amyloid
hypothesis by demonstrating that even modest reductions in
the β-cleavage of APP (resulting from a mutation in the APP
gene) could significantly protect against the development of
AD.8
Received: November 6, 2014
Accepted: December 29, 2014
Published: December 29, 2014
© 2014 American Chemical Society
210
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ACS Medicinal Chemistry Letters
Letter
when administered orally to rats at 30 mg/kg. However, further
advancement of this compound was hampered due to off-target
activity at the hERG cardiac ion channel, which translated into
an induced prolongation of the QTc interval in an integrated
dog in vivo cardiovascular model. Activity on the hERG
channel has been observed for other BACE1 inhibitors
containing a 2-amino heterocyclic warhead.14,17−19 Concurrent
efforts focused on modification of the xanthene core and
optimization of the peripheral groups to identify BACE1
inhibitors with minimized hERG activity, improved BACE1
potency, and CNS exposure.17−19 We report herein the design
and optimization of a series of BACE1 inhibitors based on the
3-aza-4-fluoro-xanthene scaffold 2 that evolved from the lead
xanthene compound 1 (Chart 1).
xanthene core, resulting in a significant improvement in BACE1
potency, as demonstrated by compound 4 that showed
significant Aβ reduction in our PD assay.17 Given the
encouraging results obtained with the 3-aza and 4-fluoro
xanthenes, we sought to combine the 3-aza and 4-fluoro
moieties in one molecule (2). This resultant combination could
potentially further improve the BACE1 potency and benefit
from the 3-aza that was shown to mitigate hERG activity with
the 4-fluoro group tempering the basicity of the 3-aza nitrogen
with the expectation of moderating Pgp efflux.
The 3-aza-4-fluoroxanthene analogues (5−23) were designed
to keep the overall physicochemical properties (molecular
weight, PSA, and clogP) within a range expected to be favorable
for CNS penetration.20 The synthesis of these compounds is
discussed in the Supporting Information. The in vitro results
are provided in Table 1. All the compounds (5−23) showed
subnanomolar range BACE1 potency in the enzymatic assay
and hERG potency >5 μM. Notably, compound 12 was more
potent in the BACE1 enzymatic assay and the cell assay than its
3-aza-xanthene counterpart 3, while 12 exhibited higher passive
permeability than 3 and retained a low efflux ratio.
Correspondingly, compound 16 showed enzymatic and cell
potency comparable to its 4-fluoro-xanthene counterpart 4 and
significantly improved passive permeability due to reduced
lipophilicity. These results demonstrated that the 3-aza-4-
fluoro-xanthene was a promising core structure with a profile
superior to other xanthene cores.
Chart 1. Early Aminooxazoline Xanthene Analogues
As previously observed, the P2′ region of the molecule was
found to be fairly accommodating, accepting saturated and
unsaturated ring systems including cyclic amines. The
unsubstituted phenyl (5) and 3- and 4-pyridyl (6 and 7) all
displayed single-digit nanomolar cellular activity. 2-Fluoro-4-
pyridyl (12) retained the same level of potency, and the o-
methyl substituent in both 4-pyridyl and 3-pyridyl groups (8
and 9) were also well tolerated, although 9 appeared to be
more labile in human liver microsomes. Introducing an o-fluoro
atom to the 3-pyridyl group, 2-fluoropyridin-3-yl (10) was 4-
and 8-fold less potent than 6 in the BACE1 enzymatic and cell
assays, respectively, and 2-fluoropyridin-5-yl (11) showed
comparable potency to 6 in the enzymatic assay but was 3-
fold less potent in the cell assay. All the P2′ dihydropyran
analogues (13−16) exhibited single-digit nanomolar potency in
both BACE1 enzymatic and cell assays. The permeability and
efflux properties suggested exposure in the CNS should be
easily achieved. The 3,6-dihydro-2H-pyrans (15−16) were
more metabolically stable than the 3,4-dihydro-2H-pyrans
(13−14) in both human and rat liver microsomal studies.
The fully saturated tetrahydropyrans (17−18) showed
improved metabolic stability but potency in the cell assay was
decreased (5-fold 18 vs 16, 25-fold 17 vs 13 and 15). Replacing
tetrahydro-2H-pyran with a morpholino group (19) improved
both BACE1 enzymatic and cell potency by 2- and 4-fold,
respectively. Compound 19 was metabolically stable in both
human and rat liver microsomes and had good permeability and
efflux properties. Further cyclic amine P2′ groups, 4,4-
difluoropiperidyl (20), 3,3-difluoropyrrolidinyl (21), and 3-
fluoropyrrolidinyl (22), were found to be well tolerated and
exhibited excellent overall profiles. The 3-methylisoxazole (23)
showed good BACE1 potency in the enzymatic assay but
suffered from a high cell shift.
From the X-ray cocrystal structure of 1 in BACE1, it is
observed that the isobutyl group extends into the P2′ pocket.12
Structure−activity relationship (SAR) efforts involving P2′
substitution showed that introduction of polar P2′ groups could
moderate the hERG activity, but typically with the consequence
of increased Pgp efflux that resulted in reduced CNS
penetration.17 Further SAR studies on the xanthene core
indicated that replacement of carbon with nitrogen at the 3-
position decreased hERG activity and increased BACE1
potency. The gain in BACE1 potency was consistent with
models based on the cocrystal structure of 1 suggesting that the
3-aza nitrogen could form a water-mediated hydrogen bond
with the Trp76 side chain. Optimization of the 3-azaxanthene
series produced 3 with robust reduction of CSF and brain Aβ
levels in a rat pharmacodynamic (PD) model.19 The Trp76
residue was also found to engage in a hydrogen bonding
interaction with a fluorine substituent at the 4-position of the
The cocrystal structure of compound 15 in BACE1 is shown
in Figure 1.21 As observed in previous structures,12,19−21 the
aminooxazoline moiety interacts with two catalytic aspartic
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Table 1. 3-Aza-4-fluoroxanthenes
a
b
c
IC50 values were averaged values determined by at least two independent experiments, n ≥ 2. Human embryonic kidney cell, n ≥ 2. Human liver
microsomal (HLM) and rat liver microsomal (RLM) clearance, n = 1. Compound concentration = 1 mM. Microsomal protein concentration = 250
d
mg/mL. Apparent permeability measured in parental LLC-PK1 cells. Values are an average of apical to basolateral (A to B) and basolateral to apical
(B to A) velocities and are reported as 10−6 cm/s, n = 1. Efflux measured in LLC-PK1 cells transfected with either rat MDR1A/1B or human
MDR1 and are reported as a ratio of (B to A)/(A to B), n = 1. hERG filter binding assay using radiolabeled dofetilide as ligand, n = 1. Racemic.
e
f
g
acids and Tyr71 engages in a π-stacking interaction with the
xanthene core. The nitrogen atom of the 2-fluoropyridin-3-yl
group interacts with Ser229 via a bridging water molecule in the
P3 pocket. As anticipated, the 4-F substitution of xanthene-
thene engages the indole NH of Trp76 in a hydrogen bonding
interaction, with the nitrogen atom in the xanthene moiety
involved in a water-mediated hydrogen bond to the Trp76 side
chain. The oxygen atom in the P2′ dihydropyranyl group
engages Tyr198 via a bridging water molecule.
On the basis of in vitro profiles, a number of compounds
were profiled in a rat PD assay to study the effect on CNS Aβ
reduction. Male Sprague−Dawley rats were administrated a
single oral dose and the Aβ levels were measured in
cerebrospinal fluid (CSF) and brain after 4 h. The unbound
plasma EC50 was estimated for Aβ reduction in CSF and brain
(Table 2). The aromatic P2′ analogues (5−7, 12) showed 3- to
6-fold shifts of the unbound plasma EC50 for brain Aβ
reduction compared to the cell IC50 (Table 1). The unbound
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Figure 2. Time course of brain and CSF Aβ lowering in rat following a
3 mg/kg dose of 15.
Figure 1. Co-crystal structure of BACE1 and compound 15. Red
spheres denote water molecules, and dashed lines indicate hydrogen
bonds. The flap covering the active site is colored yellow.
Table 2. Reduction of CSF and Brain Aβ40 in Wild-Type
Sprague−Dawley Rats
CSF
brain
Aβ40
reduction
(%)
Aβ40
reduction
(%)
dose
Cpds (mg/kg)
unbound
EC50 (mM)
unbound
EC50 (mM)
5
10
3
75
56
36
58
74
77
77
69
66
78
77
0.026
0.01
62
44
23
46
75
72
68
58
56
73
81
0.048
0.016
0.031
0.018
0.002
0.005
0.022
0.015
0.02
Figure 3. PK−PD relationship of 15 in Rhesus monkey following a 5
mg/kg po dose.
6
7
3
0.017
0.012
0.002
0.002
0.014
0.009
0.013
0.002
0.002
12
15
16
18
19
20
21
22
3
3
The pharmacokinetic profile of compound 15 was evaluated
in rat, dog, and Cynomolgus monkey (Table 3). Compound 15
demonstrated a good pharmacokinetic profile in all species.
Consistent with the in vitro microsomal stability data, 15
showed the lowest in vivo clearance and the longest half-life in
dog. When dosed orally, the bioavailability was 50%, 121%, and
43% in rat, dog, and Cynomolgus monkey, respectively.
3
10
3
3
3
0.003
0.005
3
Table 3. Pharmacokinetic Properties of 15
plasma EC50 for brain Aβ reduction of the dihydropyran
analogous (15−16) and the tetrahydropyran 18 were
consistent with their corresponding cell potencies. In particular,
the dihydropyran analogues 15 and 16 demonstrated
substantial Aβ40 reduction at the 3 mg/kg oral dose resulting
in single-digit nanomolar EC50 values in both CSF and brain.
We believe that these are the most orally efficacious BACE1
inhibitors in a rodent PD model published to date. Interestingly
the plasma unbound EC50 of compound 15 was consistent in
both brain and CSF while the other compounds exhibited
about 1.5- to 2.5-fold shift between brain EC50 and CSF EC50.
The cyclic amine analogues (19−22) displayed robust Aβ40
reduction in CSF and brain and a 2- to 3-fold shift of the brain
EC50 relative to the cell IC50.
Compound 15 was further evaluated in a time-course PD
assay in rat using a 3 mg/kg oral dose (Figure 2). Maximum Aβ
reduction in both CSF and brain was observed at 4 h postdose.
An indirect response model was used to model the PK−PD
relationship (Figure 3). The generated in vivo unbound plasma
IC50 values for CSF and brain Aβ reduction (3.7 and 4.3 nM,
respectively) were in agreement with the estimated plasma
EC50 obtained from the single dose PD study. Compound 15
was subsequently studied in a Rhesus monkey time-course PD
assay. A robust Aβ40 reduction in CSF was observed at a 5 mg/
kg dose. The generated in vivo unbound plasma IC50 values for
CSF was at 0.5 nM, 7-fold more potent than in rats.
iv
Vdss
po
dose iv/po
(mg/kg)
CL
(L/h/kg) (L/kg)
T1/2
F
species
(h) AUC (μM·h) (%)
rat
2.0/2.0
1.0/2.0
1.0/2.0
0.68
0.26
0.9
1.8
4
3.49
23.9
2.18
50
dog
cyno
2.50
2.1
8.9
4.3
121
43
To evaluate the cardiovascular safety profile, 15 was
administered intravenously to anesthetized Beagle dogs in a
series of three consecutive 30 min infusions at doses of 1, 3, and
12 mg/kg. No significant effect on the QTc interval was
observed at these doses. The safety margin over the PD effect
(IC50) in rats was determined to be 223-fold for electrocardio-
graphic effects and 51-fold for hemodynamic effects. The
margins observed in this study represent a significant
improvement over compound 1.
In conclusion, application of structure-based design with
simultaneous balance of physicochemical parameters enabled
the discovery of a series of orally highly potent 3-aza-4-fluoro-
xanthene BACE1 inhibitors. Compound 15 emerged as an
optimized lead that demonstrated robust reduction of CNS Aβ
levels in both rats and monkeys and provided significant safety
margins in a canine cardiovascular safety model.
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(9) Stachel, S. Progress Toward the Development of a Viable BACE-
1 Inhibitor. Drug Dev. Res. 2009, 9, 101−110.
ASSOCIATED CONTENT
■
S
* Supporting Information
(10) Probst, G.; Xu, Y. Small-molecule BACE1 Inhibitors: a Patent
Literature Review (2006−2011). Expert Opin. Ther. Pat. 2012, 22,
511−540.
Synthesis and characterization data for new compounds, and
methods for in vitro, in vivo, and pharmacokinetic assays. This
material is available free of charge via the Internet at http://
pubs.acs.org.
(11) Cheng, Y.; Judd, T. C.; Bartberger, M. D.; Brown, J.; Chen, K.;
Fremeau, R. T., Jr.; Hickman, D.; Hitchcock, S. A.; Jordan, B.; Li, V.;
Lopez, P.; Louie, S. W.; Luo, Y.; Michelsen, K.; Nixey, T.; Powers, T.
S.; Rattan, C.; Sickmier, E. A.; St. Jean, D. J., Jr.; Wahl, R. C.; Wen, P.
H.; Wood, S. From Fragment Screening to In Vivo Efficacy:
Optimization of a Series of 2-Aminoquinolines as Potent Inhibitors
of Beta-Site Amyloid Presursor Protein Cleaving Enzyme 1 (BACE1).
J. Med. Chem. 2011, 54, 5836−5857.
(12) Huang, H.; La, D. S.; Cheng, A. C.; Whittington, D. A.; Patel, V.
F.; Chen, K.; Dineen, T. A.; Epstein, O.; Graceffa, R.; Hickman, D.;
Kiang, Y.-H.; Louie, S.; Luo, Y.; Wahl, R. C.; Wen, P. H.; Wood, S.;
Fremeau, R. T., Jr. Stucture- and Property-Based Design of
Aminooxazoline Xanthenes as Selective, Orally Efficacious, and CNS
Penetrable BACE Inhibitors for the Treatment of Alzheimer’s Disease.
J. Med. Chem. 2012, 55, 9156−9169.
(13) Mandal, M.; Zhu, Z.; Cumming, J. N.; Liu, X.; Stickland, C.;
Mazzola, R. D.; Caldwell, J. P.; Leach, P.; Grzelak, M.; Hyde, L.;
Zhang, Q.; Terracina, G.; Zhang, L.; Chen, X.; Kuvelkar, R.; Kennedy,
M. E.; Favreau, L.; Cox, K.; Orth, P.; Buevich, A.; Voigt, J.; Wang, H.;
Kazakevich, I.; McKittrick, B. A.; Greenlee, W.; Parker, E. M.;
Stamford, A. W. Design and Validation of Bicyclic Iminopyrimidinones
as Beta-Amyloid Cleaving Enzyme-1 (BACE1) Inhibitors: Conforma-
tional Constraint to Favor a Bioactive Conformation. J. Med. Chem.
2012, 55, 9331−9345.
(14) Ginman, T.; Viklund, J.; Malmstrom, J. B.; Emond, R.;
Forsblom, R.; Johansson, A.; Kers, A.; Lake, F.; Sehgelmeble, F.;
Sterky, K. J.; Bergh, M.; Lindgren, A.; Johansson, P.; Jeppsson, F.;
Falting, J.; Gravenfors, Y.; Rahm, F. Gore Refinement toward
Permeable β-Secretase (BACE-1) Inhibitors with Low hERG Activity.
J. Med. Chem. 2013, 56, 4181−5205.
(15) Thomas, A. A.; Hunt, K. W.; Volgraf, M.; Watts, R. J.; Liu, X.;
Vigers, G. P. A.; Smith, D.; Sammond, D.; Tang, T. P.; Rhodes, S. P.;
Metcalf, A. T.; Brown, K. D.; Otten, J. N.; Burkard, M.; Cox, A. A.;
Geck Do, M. K.; Dutcher, D.; Rana, S.; KeLisle, R. K.; Regal, K.;
Wright, A. D.; Groneberg, R.; Scearce-Levie, K.; Siu, M.; Purkey, H. E.;
Lyssikatos, J. P.; Gunawardana, I. W. Discovery of 7-Tetrahydropyran-
2-yl Chromans: Beta-Site Amyloid Precursor Protein Cleaving Enzyme
1 (BACE1) Inhibitors that Reduce Amyloid β-Protein (Aβ) in the
Central Nervous System. J. Med. Chem. 2014, 57, 878−902.
(16) May, P. C.; Dean, R. A.; Lowe, S. L.; Martenyi, F.; Sheehan, S.
M.; Boggs, L. N.; Monk, S. A.; Mathes, B. M.; Mergott, D. J.; Watson,
B. M.; Stout, S. L.; Timm, D. E.; Smith Labell, E.; Gonzales, C. R.;
Nakano, M.; Jhee, S. S.; Yen, M.; Ershefsky, L.; Lindstrom, T. D.;
Calligaro, D. O.; Cocke, P. J.; Greg Hall, D.; Friedrich, S.; Citron, M.;
Audia, J. E. Robust Central Reduction of Amyloid-β in Humans with a
Orally Available, Non-Peptidic β-Secretase Inhibitor. J. Neurosci. 2011,
31, 6507−651.
(17) Epstein, O.; Bryan, M. C.; Cheng, A. C.; Derakhchan, K.;
Dineen, T. A.; Hickman, D.; Hua, Z.; Human, J. B.; Kreiman, C. E.;
Marx, I.; Weiss, M. M.; Qu, Y.; Wahl, R. C.; Wen, P. H.; Whittington,
D. A.; Wood, S.; Zheng, X.; Fremeau, R. T., Jr.; White, R. D.; Patel, V.
F. Lead Optimization and Modulation of hERG Activity in a Series of
Aminooxazoline Xanthene BACE Inhibitors. J. Med. Chem. Accepted.
(18) Dineen, T. A.; Chen, K.; Cheng, A. C.; Epstein, O.; Esmay, J.;
Hickman, D.; Kreiman, C. E.; Marx, I. E.; Wahl, R. C.; Wen, P. H.;
Weiss, M. M.; Whittington, D. A.; Wood, S.; Fremeau, Jr., R. T.;
White, R. D.; Vinod, F. P. Design and Optimization of 2-
Aminooxazoline 4-Azaxanthene Inhibitors of BACE1. J. Med. Chem.
Submitted.
AUTHOR INFORMATION
■
Corresponding Author
*Tel: 805-447-3543. E-mail: yuanc@amgen.com.
Present Addresses
◆Takeda California Inc., 10410 Science Center Drive, San
Diego, California 92121, United States.
¶Sanofi, 153 Second Avenue, Waltham, Massachusetts 02451,
United States.
□Knopp Biosciences LLC, 2100 Wharton Street, Suite 615,
Pittsburgh, Pennsylvania 15203, United States.
△31230 Bailard Road, Malibu, California 90265, United States.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We thank Kyung Gahm, Samuel Thomas, and Wesley Barnhart
for chiral separation and Alexander Long for protein
purification. We gratefully acknowledge Shamrock Structures
LLC for data collection at the Advanced Photon Source (APS).
Use of the APS was supported by the U.S. Department of
Energy, Office of Science, Office of Basic Energy Sciences,
under Contract No. W-31-109-Eng-38.
REFERENCES
■
(1) Vassar, R.; Bennet, B. D.; Babu-Khan, S.; Kahn, S.; Mendiaz, E.
A.; Denis, P.; Teplow, D. B.; Ross, S.; Amarante, P.; Loeloff, R.; Luo,
Y.; Fisher, S.; Fuller, J.; Edenson, S.; Lile, J.; Jarosinski, M. A.; Biere, A.
L.; Curran, E.; Burgess, T.; Louis, J. C. Beta-secretase Cleavage of
Alzheimer’s Amyloid Precursor Protein by the Transmembrane
Aspartic Protease BACE. Science 1999, 286, 735−741.
(2) Vassar, R.; Kovacs, D. M.; Yan, R.; Wong, P. C. The β-secretase
Enxyme in Health and Alzheimer’s Disease: Regulation, Cell Biology,
Function, and Therapeutic Potential. J. Neurosci. 2009, 29, 12787−
12794.
(3) Younkin, S. G. The Role of A beta 42 in Alzheimer’s Disease. J.
Physiol. 1998, 92, 289−29w.
(4) Lambert, M. P.; Barlow, A. K.; Chromy, B. A.; Edwards, C.;
Freed, R.; Liosatos, M.; Morgan, T. E.; Rozovsky, I.; Trommer, B.;
Viola, K. L.; Wals, P.; Zhang, C.; Finch, C. E.; Krafft, G. A.; Klein, W.
L. Diffusible, Nonfibrillar Ligands Derived from Aβ1−42 are Potent
Central Nervous System Neurotoxins. Proc. Natl. Acad. Sci. U.S.A.
1998, 95, 6448−6453.
(5) Citron, M. Alzheimer’s Disease: Strategies for Disease
Modification. Nat. Rev. Neurology 2010, 9, 387−398.
(6) Hardy, J.; Selkoe, D. J. The Amyloid Hypothesis of Alzheimer’s
Disease: Progress and Problems on the Road to Therapeutics. Science
2002, 297, 353−356.
(7) De Strooper, B.; Vasar, R.; Golde, T. The Secretases: Enzymes
with Therapeutic Potential in Alzheimer’s Disease. Nature Rev. Neurol.
2010, 6, 99−107.
(8) Jonsson, T.; Atwal, J. K.; Steinberg, S.; Snaedal, J.; Josson, P. V.;
Bjornsson, S.; Stefansson, H.; Sulem, P.; Gudbjartsson, D.; Maloney,
J.; Hoyte, K.; Gustafson, A.; Liu, Y.; Lu, Y.; Bhangale, T.; Graham, R.
R.; Huttenlocher, J.; Bjornsdottir, G.; Andreassen, O. A.; Jonsson, E.
G.; Palotie, A.; Behrens, T. W.; Magnusson, O. T.; Kong, A.;
Thorsteinsdottir, U. A Mutation in APP Protects against Alzheimer’s
Disease and Age-related Cognitive Decline. Nature 2012, 488, 96−99.
(19) Chen, J. J.; Liu, Q.; Yuan, C.; Gore, V.; Lopez, P.; Ma, V.; Judd,
T.; Minatti, A. E.; Brown, J.; Cheng, Y.; Hitchcock, S. A.; Patel, V. F.;
Powers, T. S.; White, R. D.; Wahl, R. C.; Wen, P.; Whittington, D. A.;
Cheng, A. C. ; Bartberger, M.; Hickman, D.; Dunn, B.; Wood, S.;
Fremeau, R. T., Jr. Developememt of 2-Aminooxazoline 3-
214
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Azaxanthenes as Orally Active β-Secretase Inhibitors for the
Treatments of Alzheimer’s Disease. Manuscript in preparation.
(20) Hitchcock, S. A.; Pennington, L. D. Structure-Brain Exposure
Relationships. J. Med. Chem. 2006, 49, 7559−7583.
(21) The cocrystal structure of BACE1 + 15 was determined at 1.85
Å resolution. Coordinates and structure factors have been deposited in
the PDB with accession code 4WPU.
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