Rational design and synthesis of potent dibenzazepine motifs as β-secretase inhibitors

Article abstract of DOI:10.1021/jm9008482

We have identified small-molecule dibenzazepine inhibitors of β-secretase (BACE1). These BACE1 inhibitors possess two key salient features. The first is a seven-membered heterocyclic ring fused to two aromatic rings representing the P3-P2 residues. The se

Full text of DOI:10.1021/jm9008482

6484 J. Med. Chem. 2009, 52, 6484–6488
DOI: 10.1021/jm9008482
Rational Design and Synthesis of Potent Dibenzazepine Motifs as β-Secretase Inhibitors
Taleb H. Al-Tel,*,† Raed A. Al-Qawasmeh,‡ Marco F. Schmidt, Amal Al-Aboudi,‡ Shashidhar N. Rao,^ Salim S. Sabri,† and
Wolfgang Voelter§
†College of Pharmacy, University of Sharjah, P.O. Box 27272, Sharjah, UAE, ‡Department of Chemistry, University of Jordan, Amman 1194,
Jordan, Leibniz-Institute of Molecular Pharmacology (FMP), Robert-Rossle-Strasse 10, 13125 Berlin, Germany, ^Schrodinger, 120 W. 45th
Street, New York, New York 10036, and Interfakultares Institut fur Biochemie, Eberhard-Karls-Universitat Tuebingen, Hoppe-Seyler-Strasse,
72076 Tuebingen, Germany
§
Received June 2, 2009
We have identified small-molecule dibenzazepine inhibitors of β-secretase (BACE1). These BACE1
inhibitors possess two key salient features. The first is a seven-membered heterocyclic ring fused to two
aromatic rings representing the P3-P2 residues. The second is an amide and/or amide bioisostere
representing the P10 residue. Rational optimization led to the identification of potent analogues, such as
10 (KI = 211 nM).
Introduction
(representing P3 residue) fused to a phthalic acid derivative
(representing P2 residue) might retain the pharmacodynamic
properties and improve the pharmacokinetic characteristics
found in isophthalic acid derivatives against BACE1
(Figure 2).8 Furthermore, P10-residue could be manipulated
to include amides and/or amide bioisosters that might fit the
S10 pocket of the enzyme active site.
To verify this hypothesis, we have designed low molecular
weight nonpeptidic BACE1 inhibitors that incorporate a
dibenzazepine core. Although the hydroxyethylamine
(HEA) derivatives of isophthalic acid published earlier are
highly potent inhibitors of BACE1, these compounds suffered
from poor pharmacokinetic stability.8 Hence, we replaced
both of the amides in isophthalic acid derivatives, represented
in compound 1, with dibenzazepine and imidazole isosters
to fit in the S3 and S10 subpockets (I, II), respectively. The
synthetic route used to prepare these motifs to connect in
tandem the probable blocks corresponding to P3-P10 resi-
dues is outlined in Scheme 1.
Synthesis. Compound 4 was prepared in 85% yield
(Scheme 2) through the nucleophilic aromatic substitution
carried out between 2 and 3. The nitro group in the diphe-
nylamine 4 was subjected to Pd/C reduction using HCONH4
as a source of H2 to deliver the amine 5 in 75% yields after
column chromatography. Amide formation using ethylfor-
myl chloride followed by electrophilic aromatic substitution
catalyzed by POCl3 furnished the dibenzazepine framework
7. Saponification of 7 produced acid 8. At this stage, we took
advantage of the flexibility present in the carboxylic acid
appendage and the possibility of functionalizing this group
with various residues.
Therefore, three different classes in relation to P10 residue
were synthesized. These included simple amides, substituted
imidazole and benzimidazole derivatives as outlined in
Schemes 3 and 4. The acid 8 was converted to the amides 9
and 10 through standard coupling protocol using TBTU/
DIPEA (Scheme 3) in 85% and 87% yields, respectively.
For the synthesis of the second class, conventional synthetic
Alzheimer0s disease (ADa) is a progressive, neurodegen-
erative disorder with no current cure and is the leading cause
of dementia and death in elderly people.1-3 A key hallmark of
AD is deposition of aggregated β-amyloid peptides (Aβ-40,
-42) as plaques in the brain.4 Two proteases, β- and
γ-secretase, have been identified to be involved in the sequen-
tial proteolysis of membrane-anchored amyloid precursor
protein (APP).5
β-Secretase (BACE1) has been shown to be a membrane-
bound aspartyl protease and is considered to be involved in the
rate-limiting step in the processing of APP to Aβ peptide
responsible for the production of the membrane-bound
β-C-terminal fragment, which is then further cleaved to Aβ
by γ-secretase (Figure 1).6 BACE1 knockout homozygote mice
show a complete absence of the plaques resulting from the
production of Aβ in the brain, and the lack of BACE1 has been
reported to have no side effects.7 Thus, BACE1, which is highly
expressed in the CNS, is deemed to be an attractive target for
drug discovery through inhibition of Aβ production.8
Different classes of nonpeptidic potent BACE1 inhibitors,
including acylguanidines, isophthalic acid derivatives, amino
aromatic heterocyclic motifs, and arylpiperazines, have been
reported.8 In particular, the isophthalic acid derivatives were
shown to demonstrate potent inhibitory activity in the sub-
nanomolar range.8c,d
Results and Discussion
In the present study, it was anticipated that the isophtha-
lic acid motifs reported by many groups could be envisioned
to be superimposed on the more rigid dibenzazepine
motifs (Figure 2).8 Specifically, we hypothesized that the
incorporation of a seven-membered diazepine ring system
*To whom correspondence should be addressed. Phone: (971) 50
1732950. Fax: þ971 6 5585812. E-mail: taltal@sharjah.ac.ae.
a Abbreviations: AD, Alzheimer’s disease; APP, amyloid precursor
protein; BACE1, β-site amyloid precursor protein cleaving enzyme 1.
r
pubs.acs.org/jmc
Published on Web 09/29/2009
2009 American Chemical Society

Brief Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 20 6485
procedures were followed to achieve our scaffolds. There-
fore, the imidazole isosters 11 and 12 were synthesized
through coupling between acid 8 with the desired R-bromo
ketone followed by refluxing the produced esters in
AcONH4/AcOH (Scheme 3). The third types of motifs were
benzimidazole analogues 13 and 14 (Scheme 4) and were
made from acid 8 utilizing standard coupling with the desired
diamine using TBTU/DIPEA followed by refluxing the
amide products in AcOH.
Biology. BACE1 inhibitory activity of the synthesized
motifs was determined using a fluorescence resonance energy
transfer assay (FRET).9 Analogues of 9 were then synthe-
sized and allowed for optimization of the P10 substituent.
The activity of these novel derivatives is summarized in
Table 1. In general, the fluorophenylamide motif 10 led to
a compound that is a relatively more potent BACE1 inhi-
bitor (KI = 211 nM) than the corresponding pyridylamide
analogue 9 (KI = 295 nM). On the other hand, the aryl-fused
imidazole derivatives (e.g., 14, KI = 244 nM) are more
potent than the substituted imidazole systems (e.g., 12,
KI = 293 nM). The fluorophenyl analog (10) was consis-
tently the most potent in the BACE inhibition assay. Succes-
sively rigidifying P10 substituents (11 and 13) eroded activity
of the pyridyl motifs in a systematic fashion. The lack of a
potency difference between amide 10 and its imidazole
isoster 14 (Table 1) was striking, especially considering the
small conformational difference observed in both analogues.
In conclusion, these SAR findings suggested that the fluor-
ophenyl analogues in the three derivatives (10, 12, and 14) are
more active than their pyridyl analogues (9, 11, and 13),
perhaps because of the similarity in the nature of interaction
forces between these motifs and BACE1 active site. The
absolute inhibition constants and the ligand efficiency in-
dexes of our new diabenzazepine analogues are summarized
in Table 1 and were found to be consistent with KI values.
Furthermore, in comparison with the known BACE1 inhi-
bitors 1 and the peptide-based inhibitor OM99-2 (H-Glu-
Val-Asn-ψ-Leu-Ala-Glu-Phe-OH; ψ denotes replacement of
CONH by (S)-CH(OH)CH2) (Figure 3), our designed scaf-
folds indicated their potential inhibition properties toward
BACE1. At this junction it is worthwhile mentioning that 1
and the known peptide inhibitor OM99-2 showed obviously
a 100-fold better activity than our developed compounds.
But both are not applicable for clinical use because of their
poor pharmacokinetic.8 On the other hand, benzazepine
derivatives are known to have appreciable pharmacokinetic
profile and able to cross the blood-brain barrier.10 Second,
by calculation of the ligand efficiency (Δg = free binding
energy/number of non-hydrogen atoms), it could also be
Figure 1. Schematic of the APP and its metabolites (not drawn
to scale). APP can be processed along two major pathways, the
R-secretase pathway and the amyloid forming β-secretase pathway.
In the nonamyloidogenic pathway, R-secretase cleaves in the middle
of the Aβ region to release a large soluble APP fragment, R-APPs.
The C-terminal C83 peptide is metabolized to p3 and AICD (APP
intracellular domain) by γ-secretase. In the amyloid forming
β-secretase pathway, β-secretase releases a large soluble fragment,
β-APPs. The C-terminal C99 peptide is then metabolized to Aβ and
AICD by γ-secretase. β-Secretase inhibitors block the formation of
β-APPs and C99.
Figure 2. Two dimensional overlay of compound 14 (blue) and 1
(red) indicating the possible interaction modes with the BACE1
pockets.
Scheme 1. Rational Design of Dibenzazepine BACE1 Inhibitors Based on Isophthalic Acid Derivatives

6486 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 20
Scheme 2. Synthesis of the Benzodiazepam Starting Materiala
Al-Tel et al.
a (a) K2CO2, CuI, DMF, 60 ꢀC, 12 h (85%); (b) 10 mol % of 10% Pd/C, methanol, HCONH4, 50 ꢀC, 5 h (75%); (c) ethyl carbonochloridate, pyridine,
0 ꢀC, 6 h (93%); (d) POCl3, P2O5, reflux, 6 h (62%); (e) NaOH, methanol, H2O, 25 ꢀC, 12 h (95%).
Scheme 3. Synthesis of BACE1 Inhibitorsa
Scheme 4. Synthesis of BACE1 Inhibitorsa
a (a) 13: 3,4-diaminopyridine, TBTU, DIPEA, DMF, 0 ꢀC, 6 h (82%);
then AcOH, reflux, 6 h (64%). 14: 1,2-diamino-4-fluorobenezene,
TBTU, DIPEA, DMF, 0 ꢀC, 6 h (86%); then AcOH, reflux, 6 h (58%).
Silica gel 60 (Merck 40-63 μm) was used for flash column chro-
matography. 1H NMR and 13C NMR spectra were recorded with
Bruker 500 and Bruker AMX-400 spectrometers using CDCl3.Data
are presented as follows: chemical shift (parts per million, ppm),
multiplicity ((s) singlet, (d) doublet, (t) triplet, (q) quartet, (m)
multiplet, (br) broad), coupling constant J (in hertz), and integra-
tion. Carbon magnetic resonance (13C NMR) spectra were recorded
at 75 or 125 MHz. Data for 13C NMR are reported in terms of
chemical shifts (ppm). The EI and FD mass spectra were recorded
on a Finnigan MAT 312 mass spectrometer connected to a PDO
11/34 (DEC) computer system. Elemental analysis was performed
on a Perkin-Elmer elemental analyzer, model 240.
All compounds were determined to be >95% pure by high-
performance liquid chromatography (HPLC). Purity of com-
pounds were determined on a Phenomenex Luna C18-(2), 3 mm
column, 4.6 mm i.d. ꢀ 30 mm length, with 30-75% acetonitrile/
water/0.1% trifluoroacetic acid, 1.0 mL/min elution at room
temperature using 210, 254, or 280 nm wavelength.
BACE1 Enzyme Assay. BACE1 assay was carried out accord-
ing to the manufacturer prescribed protocol available from
Invitrogen (http://tools.invitrogen.com/content/sfs/manuals/
L0724.pdf). Briefly, the in vitro assay was carried out by
fluorescence resonance energy transfer (FRET). An APP-based
peptide substrate (rhodamine-EVNLDAEFK-DABCYL,
KM = 20 μM) carrying the Swedish mutation and containing
a rhodamine as a fluorescence donor and a quencher acceptor at
each end was used. The intact substrate is weakly fluorescent
and becomes highly fluorescent upon enzymatic cleavage. The
assay was conducted in 50 mM sodium acetate buffer, pH 4.5, in
a final enzyme reaction mixture of enzyme (1 U/mL), inhibitor
(10, 3, 1, and 0.3 μM) and substrate (750 nM). Inhibitor
compounds were diluted from stock solutions to result in
3.3% DMSO final concentration. The reaction was incubated
for 60 min at 25 ꢀC under dark conditions and then stopped with
2.5 M sodium acetate. Fluorescence was monitored with a
a Reagents and conditions. (a) 9: 4-aminopyridine, TBTU, DIPEA,
DMF, 0 ꢀC, 6 h (85%). 10: p-fluoroaniline, TBTU, DIPEA, DMF, 0 ꢀC,
6 h (87%). (b) 11: R-bromomethyl pyridyl ketone, DMF, DIPEA, 0 ꢀC
to room temp, 6 h (95%); then AcOH, AcONH4, reflux, 8 h (55%). 12:
R-bromomethyl (4- fluorophenyl) ketone, DMF, DIPEA, 0 ꢀC, 6 h
(95%); then AcOH/AcONH4, reflux, 8 h (60%).
shown that our low molecular weight derivatives bind more
efficiently to the target compared to the peptidic inhibitor
OM99-2 (Δg ≈ 1.4 kJ/mol for our described dibenzazepine
inhibitors vs Δg = 0.79 kJ/mol for OM99-2; see Table 1).
These findings support our assumption that the described
concept of dibenzazepine ligands as BACE1 inhibitors might
be the starting point of a new generation of anti-AD drugs.
In summary, the combination of dibenzazepine motif with
optimized replacements for the P2/P3 residues and P10 ap-
pendage resulted in compounds, 10 and 14, that represent
promising new series of BACE-1 inhibitors. These new com-
pounds displayed good enzymatic inhibitory potency and a
molecular weight below 350. Further improvement of these
parameters and the incorporation of other druglike properties
into these inhibitors are the subjects of ongoing efforts.
Experimental Section
Chemistry. Materials and General Methods. Reagents and
solvents were purchased from commercial sources and were used
as received. Reaction progress was monitored by thin-layer chro-
matography on Merck silica gel 60 F-254 with detection by UV.

Brief Article
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 20 6487
Table 1. SAR of BACE1 Inhibitors
Figure 3. Structures of compounds 1 and OM99-2, their inhibition
constants, and ligand efficiency indexes.
137.5, 145.3, 146.7, 147.9, 148.1, 148.4, 163.2, 165.1. FD-MS
(M þ 1) = 331. Anal. Calcd for C19H14N4O2: C, 69.08; H, 4.27;
N, 16.96. Found: C, 69.05; H, 4.21; N, 16.89.
N-(4-Fluorophenyl)-11-oxo-10,11-dihydro-5H-dibenzo[b,e]-
[1,4]diazepine-3-carboxamide (10). 1H NMR (CDCl3, 400
MHz): δ = 4.52 (1H, bs), 7.26-7.01 (5H, m), 7.90-7.58 (3H,
m), 8.00 (3H, m), 8.34 (1H, s), 9.01 (1H, bs). 13C NMR:
δ = 114.9, 115.1, 117.8, 119.5, 120.6, 121.9, 123.7, 123.8,
124.3, 127.3, 129.4, 133.6, 135.9, 137.0, 142.7, 151.3, 158.2,
163.4, 165.1. FD-MS (M þ 1) = 348. Anal. Calcd for
C20H14FN3O2: C, 69.16; H, 4.06; F, 5.47; N, 12.10. Found: C,
69.09; H, 4.13; F, 5.51; N, 12.09.
spectrofluorometer at 545 nm excitation and 585 nm emission.
The assay kit was validated by manufacturer. The obtained
values are the mean values of three different experiments. IC50
values were calculated by plotting the obtained relative fluores-
cence unit per hour (RFU/h) against the logarithm of inhibitor
concentration. The measured inhibition data were analyzed in
GraphPad Prism 4 for Windows (GraphPad Software Inc., La
Jolla, CA) by nonlinear regression (curve fitting). Based on
obtained IC50 values, KI values were calculated (detailed de-
scription is in the Supporting Information).
Methyl 11-Oxo-N-(pyridin-4-yl)-10,11-dihydro-5H-dibenzo-
[b,e][1,4]diazepine-3-carboxamide (9) and N-(4-Fluorophenyl)-
11-oxo-10,11-dihydro-5H-dibenzo[b,e][1,4]diazepine-3-carbox-
amide (10). To a solution of methyl 11-oxo-10,11-dihydro-5H-
dibenzo[b,e][1,4]diazepine-3-carboxylic acid (8) (254 mg, 1.0
mmol, 1 equiv) in DMF (8 mL) at 0 ꢀC was added DIPEA
(0.21 mL, 1.2 mmol, 1.2 equiv). After 10 min TBTU (551 mg, 1.2
mmol, 1.2 equiv) was added and the resulting mixture stirred at
the same temperature for 30 min. Then 4-aminopyridine or
4-flouroaniline (1.1 mmol, 1.1 equiv) was added. The resulting
mixture was stirred at 0 ꢀC for 6 h and then quenched with
ice-water. The precipitated solid was filtered, washed with
water, and dissolved in EtOAc. The organic phase was washed
with a 1 N HCl aqueous solution, then with a saturated
NaHCO3 aqueous solution, and finally with H2O, dried over
MgSO4, and concentrated in vacuo. The crude product was
purified by flash chromatography on silica gel (DCM/AcOEt,
9/1 to 7/3) to give methyl 11-oxo-N-(pyridin-4-yl)-10,11-di-
hydro-5H-dibenzo[b,e][1,4]diazepine-3-carboxamide (9) (85%)
or N-(4-fluorophenyl)-11-oxo-10,11-dihydro-5H-dibenzo[b,e]-
[1,4]diazepine-3-carboxamide (10) (87%).
3-(5-(Pyridin-4-yl)-1H-imidazol-2-yl)-5H dibenzo[b,e][1,4]-
diazepin-11(10H)-one (11) and 3-(5-(4-Fluorophenyl)-1H-imida-
zol-2-yl)-5H dibenzo[b,e][1,4]diazepin-11(10H)-one (12). To a
solution of methyl 11-oxo-10,11-dihydro-5H-dibenzo[b,e][1,4]-
diazepine-3-carboxylic acid (8) (254 mg, 1.0 mmol, 1 equiv) in
DMF (10 mL) at 0 ꢀC was added DIPEA (0.21 mL, 1.2 mmol,
1.2 equiv). The resulting mixture was stirred at 0 ꢀC for 30 min
followed by dropwise addition of the appropriate addition of the
R-bromoketone (1.1 mmol, 1.1 equiv) in DMF. The resulting
mixture was stirred at 0 ꢀC for 6 h, then quenched with ice-
water. The precipitated solid was filtered, washed with water,
and dissolved in EtOAc. The organic phase was washed with a 1
N HCl aqueous solution and then a saturated NaHCO3 aqueous
solution, then H2O, dried over MgSO4, and concentrated in
vacuo, which was used in the next stage without further pur-
ification. To the appropriate R-keto ester were added AcOH
(25 mL) and AcONH4 (924 mg, 12 mmol, 12 equiv), and the
resulting suspension was refluxed for 8 h, cooled to room
temperature, concentrated in vacuo, and diluted with crushed
ice. The brown solid was filtered, washed thoroughly with water.
The crude cake was dissolved in EtOAc washed with a saturated
NaHCO3 aqueous solution and with H2O, dried over MgSO4,
and concentrated in vacuo. The crude product was purified by
flash chromatography on silica gel (DCM/AcOEt, 8/2 to 7/3) to
give 3-(5-(pyridin-4-yl)-1H-imidazol-2-yl)-5H dibenzo[b,e]-
[1,4]diazepin-11(10H)-one (11) (55%) or 3-(5-(4-fluorophenyl)-
1H-imidazol-2-yl)-5H dibenzo[b,e][1,4]diazepin-11(10H)-one
(12) (60%).
3-(5-(Pyridin-4-yl)-1H-imidazol-2-yl)-5H-dibenzo[b,e][1,4]di-
azepin-11(10H)-one (11). 1H NMR (CDCl3, 400 MHz): δ = 5.34
(1H, s), 7.20-7.03 (2H, m), 7.28 (1 h, dd, J = 2.4, 8.3 Hz), 7.37
(1H, s), 7.53 (1H, m), 7.73-7.70 (2H, m), 9.92 (1 h, s), 7.99 (1 h,
d, J = 1.82 Hz), 8.42 (2H, m), 8.49 (1H, dd, J = 2.3, 8.2 Hz),
9.21 (1H, s), 9.84 (1H, bs). 13C NMR: δ = 117.8, 117.9, 118.4,
120.9, 121.5, 122.7, 124.0, 125.6, 127.6, 129.1, 129.3, 131.0,
11-Oxo-N-(pyridin-4-yl)-10,11-dihydro-5H-dibenzo[b,e][1,4]-
diazepine-3-carboxamide (9). 1H NMR (CDCl3, 300 MHz): δ =
4.34 (1H, bs), 7.24-7.02 (3H, m), 7.54 (1H, bs), 7.80 (2H, m),
7.92 (1H, bs), 8.00 (1H, d, J = 2.3 Hz), 8.63 (1H, dd, J = 2.1, 8.2
Hz), 9.13 (1H, bs), 9.51 (1H, bs). 13C NMR: δ = 114.2, 114.8,
115.0, 115.9, 119.9, 122.3, 123.9, 125.8, 127.9, 129.2, 132.0,

6488 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 20
Al-Tel et al.
133.1, 135.3, 136.5, 141.8, 142.8, 149.7, 151.0, 163.4. FD-MS
(M þ 1) = 354. Anal. Calcd for C21H15N5O: C, 71.38; H, 4.28;
N, 19.82. Found: C, 71.44; H, 4.32; N, 19.78.
Supporting Information Available: Experimental procedures
for the synthesis of 9-14, in vitro assays and protocols
for the determination of KI. This material is available free of
charge via the Internet at http://pubs.acs.org.
3-(5-(4-Fluorophenyl)-1H-imidazol-2-yl)-5H-dibenzo[b,e][1,4]-
1
diazepin-11(10H)-one (12). H NMR (CDCl3, 400 MHz): δ =
5.31 (1H, bs), 7.20-7.01 (5H, m), 7.29 (1H, dd, J = 2.2, 8.3 Hz),
7.55 (1H, m), 8.09-7.91 (4H, m), 8.61 (1H, dd, J = 2.2, 8.2 Hz),
9.24 (1H, bs), 10.1 (1H, s). 13C NMR: δ = 115.9, 116.9, 117.5,
117.8, 122.3, 124.5, 124.5, 125.7, 125.4, 126.7, 127.5, 128.9,
129.8, 130.7, 130.9, 131.3, 131.6, 138.1, 153.4, 151.1, 161.8,
164.0. FD-MS (M þ 1) = 371. Anal. Calcd for C22H15FN4O:
C, 71.34; H, 4.08; F, 5.13; N, 15.13. Found: C, 71.36; H, 4.11; F,
5.17; N, 15.20.
References
(1) (a) Cummings, J. L. Alzheimer’s Disease. N. Engl. J. Med. 2004,
351, 56–67. (b) Nguyen, J.; Yamani, A.; Kiso, Y. Views on Amyloid
Hypothesis and Secretase Inhibitors for Treating Alzheimer's Disease:
Progress and Problems. Curr. Pharm. Des. 2006, 12, 4295–4312.
(c) For online resources and figures about AD, see www.alz.org and
www.ahaf.org.
(2) Celsis, P. Age-related cognitive decline, mild cognitive impairment
or pre-clinical Alzheimer0s disease. Ann. Med. 2000, 32, 6–14.
(3) (a) Stavrovskaya, I. G.; Kristal, B. S. The Powerhouse Takes
Control of the Cell: Is the Mitochondrial Permeability Transition
a Viable Therapeutic Target against Neuronal Dysfunction and
Death? Free Radical Biol. Med. 2005, 38, 687–697. (b) Milligan, C.
Caspase Cleavage of APP Results in a Cytotoxic Proteolytic Peptide.
Nat. Med. 2000, 6, 385–386.
(4) Yan, R.; Bienkowski, M. J.; Shuck, M. E.; Miao, H.; Tory, M. C.;
Pauley, A. M.; Brashier, J. R.; Stratman, N. C.; Mathews, R. W.;
Buhl, A. E.; Carter, D. B.; Tomasselli, A. G.; Parodi, L. A.;
Heinrikson, R. L.; Gurney, M. Membrane-Anchored Aspartyl
Protease with Alzheimer’s Disease Beta-Secretase Activity. Nature
1999, 402, 533–537.
(5) (a) Iwatsubo, T.; Odaka, A.; Suzuki, N.; Mizusawa, H.; Nukina,
N.; Ihara, Y. Visualization of Aβ42(43) and Aβ40 in Senile
Plaques with End-Specific Aβ Monoclonals: Evidence That an
Initially Deposited Species Is Aβ42(43). Neuron 1994, 13, 45–53.
(b) St George-Hyslop, P. H.; Morris, J. C. Will Anti-Amyloid Thera-
pies Work for Alzheimer's Disease? Lancet 2008, 372, 180–182. (c)
Ghosh, A. K.; Bilcer, G.; Hong, L.; Koelsch, G.; Tang, J. Memapsin 2
(Beta-Secretase) Inhibitor Drug, between Fantasy and Reality. Curr.
Alzheimer Res. 2007, 4, 418–422. (d) Durham, T. B.; Shepherd, T. A.
Progress toward the Discovery and Development of Efficacious
BACE Inhibitors. Curr. Opin. Drug Discovery Dev. 2006, 9,
776–791.
3-(3H-Imidazo[4,5-c]pyridin-2-yl)-5H-dibenzo[b,e][1,4]diaze-
pin-11(10H)-one (13) and 3-(6-Fluoro-1H-benzo[d]imidazol-2-yl)-
5H-dibenzo[b,e][1,4]diazepin-11(10H)-one (14). To a solution of
methyl 11-oxo-10,11-dihydro-5H-dibenzo[b,e][1,4]diazepine-3-
carboxylic acid (8) (254 mg, 1.0 mmol, 1 equiv) in DMF (8 mL)
at 0 ꢀC was added DIPEA (0.21 mL, 1.2 mmol, 1.2 equiv). After
10 min, TBTU (551 mg, 1.2 mmol, 1.2 equiv) was added and the
resulting mixture stirred at the same temperature for 30 min.
Then diaminopyridine or diaminobenzene (1.1 mmol, 1.1 equiv)
was added. The resulting mixture was stirred at 0 ꢀC for 6 h and
then quenched with ice-water. The precipitated solid was
filtered, washed with water, and dissolved in EtOAc. The
organic phase was washed with a 1 N HCl aqueous solution,
then with a saturated NaHCO3 aqueous solution, and finally
with H2O, dried over MgSO4, concentrated in vacuo, and then
used in the next stage without further purification. To the
appropriate amide was added AcOH (30 mL), and the resulting
suspension was refluxed for 6 h, cooled to room temperature,
concentrated in vacuo, and diluted with crushed ice. The brown
solid was filtered and washed thoroughly with water. The
crude was dissolved in EtOAc, washed with a saturated NaH-
CO3 aqueous solution and with H2O, dried over MgSO4, and
concentrated in vacuo. The crude product was purified by
flash chromatography on silica gel (DCM/AcOEt, 8/2 to 7/3)
to give 3-(3H-imidazo[4,5-c]pyridin-2-yl)-5H-dibenzo[b,e][1,4]-
diazepin-11(10H)-one (13) (64%) or 3-(6-fluoro-1H-benzo-
[d]imidazol-2-yl)-5H-dibenzo[b,e][1,4]diazepin-11(10H)-one
(14) (58%).
(6) Wolfe, M. S. Secretase Targets for Alzheimer’s Disease: Identifi-
cation and Therapeutic Potential. J. Med. Chem. 2001, 44,
2039–2060.
(7) (a) Willem, M.; Garratt, A. N.; Novak, B.; Citron, M.; Kaufmann,
S.; Rittger, A.; DeStrooper, B.; Saftig, P.; Birchmeier, C.; Haass, C.
Control of Peripheral Nerve Myelination by the β-Secretase
BACE1. Science 2006, 314, 664–666.
(8) (a) Stanton, M. G.; Stauffer, S. R.; Gregro, A. R.; Selnick, H.
Discovery of Isonicotinamide Derived β-Secretase Inhibitors: In
Vivo Reduction of β-Amyloid. J. Med. Chem. 2007, 50, 3431–3433.
(b) Charrier, N.; Clarke, B.; Cutler, L.; Demont, E.; Dingwall, C.;
Dunsdon, R.; East, P.; Hawkins, J.; Howes, C.; Hussain, I.; Jeffrey, P.;
Maile, G.; Matico, R.; Mosley, J.; Naylor, A.; O'Brien, A.; Redshaw, S.;
Rowland, P.; Soleil, V.; Smith, K. J.; Sweitzer, S.; Theobald, P.; Vesey,
D.; Walter, D. S.; Wayne, G. Second Generation of Hydroxyethylamine
BACE-1 Inhibitors: Optimizing Potency and Oral Bioavailability. J.
Med. Chem. 2008, 51, 3313–3317. (c) Cumming, J. N.; Le, T. X.;
Babu, S.; Carroll, C.; Chen, X.; Favreau, L.; Gaspari, P.; Guo, T.; Hobbs,
D. W.; Huang, Y.; Iserloh, U.; Kennedy, M. E.; Kuvelkar, R.; Li, G.;
Lowrie, J.; McHugh, N. A.; Ozgur, L.; Pan, J.; Parker, E. M.; Saionz, K.;
Stamford, A. W.; Strickland, C.; Tadesse, D.; Voigt, J.; Wang, L.; Wu,
Y.; Zhang, L.; Zhang, Q. Rational Design of Novel, Potent Piperazinone
and Imidazolidinone BACE1 Inhibitors. Bioorg. Med. Chem. Lett.
2008, 18, 3236–3241. (d) Kortum, S. W.; Benson, T. E.; Bielnkowski,
M. J.; Emmons, T. L.; Prince, D. B.; Paddock, D. J.; Tomasselli, A. J.;
Moon, J. B.; LaBorde, A.; TenBrink, R. E. Potent and Selective
Isophthalamide S2 Hydroxyethylamine Inhibitors of BACE1. Bioorg.
Med. Chem. Lett. 2007, 17, 3378–3383. (e) Iserloh, U.; Pan, J.;
Samford, A. W.; Kennedy, M. E.; Zhang, Q.; Zhang, L.; Parker, E.
M.; McHugh, N. A.; Favreu, L.; Strickland, C.; Voigt, J. Discovery of an
Orally Efficacious 4-Phenoxypyrrolidine-Based BACE-1 Inhibitor.
Bioorg. Med. Chem. Lett. 2008, 18, 418–422.
3-(3H-Imidazo[4,5-c]pyridin-2-yl)-5H-dibenzo[b,e][1,4]diaze-
pin-11(10H)-one (13). H NMR (CDCl3, 400 MHz): δ = 4.84
1
(1H, bs), 7.1-7.01 (2H, m), 7.29 (1H, bd, J = 2.5 Hz), 7.55 (2H,
m), 7.90 (1H, bs), 8.00 (1H, dd, J = 2.2, 8.3 Hz), 8.30 (1H, d, J =
1.9 Hz), 8.60 (1H, dd, J = 2.4, 8.3 Hz), 8.90 (1H, s), 9.42 (1H, s),
9.81 (1H, s). 13C NMR: δ = 105.5, 108.6, 117.8, 119.5, 120.9,
124.1, 125.3, 129.0, 130.1, 131.8, 133.4, 138.8, 139.1, 140.2,
140.7, 141.0, 145.7, 153.5, 164.0. FD-MS (M þ 1) = 328. Anal.
Calcd for C19H13N5O: C, 69.71; H, 4.00; N, 21.39. Found: C,
69.68; H, 4.07; N, 21.43.
3-(6-Fluoro-1H-benzo[d]imidazol-2-yl)-5H-dibenzo[b,e][1,4]-
1
diazepin-11(10H)-one (14). H NMR (CDCl3, 400 MHz): δ =
4.85 (1H, s), 7.20-7.05 (3H, m), 7.29 (1H, dd, J = 2.4, 8.3 Hz),
7.48-7.39 (2H, m), 7.84 (1H, s), 7.95 (1H, s), 8.12 (1H, dd, J =
2.2, 8.1 Hz), 8.50 (1H, d, J = 8.4 Hz), 9.30 (1H, bs), 9.88 (1H, s).
13C NMR: δ = 105.1, 106.1, 116.4, 117.8, 118.6, 120.9, 121.7,
122.3, 125.7, 129.1, 130.2, 131.3, 136.5, 137.0, 137.2, 140.7,
148.1, 152.2, 1157.2, 167.5. FD-MS (M þ 1) = 345. Anal. Calcd
for C20H13FN4O: C, 69.76; H, 3.81; F, 5.52; N, 16.27. Found: C,
69.71; H, 3.78; F, 5.59; N, 16.200.
(9) Choi, S.-H; Hur, J.-M.; Yang, E.-J.; Jun, M.; Park, H.-J.; Lee, K.-B.;
Moon, E.; Song, K.-S. β-Secretase (BACE1) Inhibitors from Perilla
frutescens var acuta. Arch Pharm Res. 2008, 31, 183–187. (b) http://
tools.invitrogen.com/content/sfs/manuals/L0724.pdf.
(10) Albert, U.; Aguglia, E.; Maina, G.; Bogetto, F. Venlafaxine versus
Clomipramine in the Treatment of Obsessive-Compulsive Disor-
der: A Preliminary Single-Blind, 12-Week, Controlled Study.
J. Clin. Psychiatry 2002, 63, 1004–1009.
Acknowledgment. We thank Dr. Thomas Nittoli (Wyeth
Research, NY) for fruitful discussions and suggestions. The
authors are also grateful to DAAD and the College of
Graduate Studies and Research, University of Sharjah;
UAE for supporting this project.