Pyridine-derived γ-secretase modulators

Article abstract of DOI:10.1016/j.bmcl.2011.06.042

SAR of a novel series of pyridine-derived γ-secretase modulators is described. Compound 5 was found to be a potent modulator in vitro, which on further profiling, was found to decrease Aβ42 and Aβ40, and maintain (or increase) the levels of total Aβ. Furthermore, representative compounds 1 and 5 demonstrated in vivo efficacy to lower Aβ42 in the brain without altering Notch processing in the peripheral.

Full text of DOI:10.1016/j.bmcl.2011.06.042

Bioorganic & Medicinal Chemistry Letters 21 (2011) 4832–4835
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Pyridine-derived c-secretase modulators
⇑
Zehong Wan , Adrian Hall, Yingxia Sang, Jia-Ning Xiang, Eric Yang, Beverley Smith, David C. Harrison,
Guang Yang, Haihua Yu, Helen S. Price, Jian Wang, Julie Hawkins, Lit-Fui Lau, Matthew R. Johnson, Tina Li,
Wei Zhao, William L. Mitchell, Xiaobo Su, Xiaomin Zhang, Yanjiao Zhou, Yun Jin, Zhaolong Tong,
Ziqiang Cheng, Ishrut Hussain, John D. Elliott, Yasuji Matsuoka
Research and Development, GlaxoSmithKline Pharmaceuticals, 898 Halei Road, Zhangjiang Hi-tech Park, Pudong, Shanghai 201023, China
a r t i c l e i n f o
a b s t r a c t
Article history:
SAR of a novel series of pyridine-derived c-secretase modulators is described. Compound 5 was found to
Received 1 May 2011
Revised 7 June 2011
Accepted 10 June 2011
Available online 17 June 2011
be a potent modulator in vitro, which on further profiling, was found to decrease Ab42 and Ab40, and
maintain (or increase) the levels of total Ab. Furthermore, representative compounds 1 and 5 demon-
strated in vivo efficacy to lower Ab42 in the brain without altering Notch processing in the peripheral.
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
c
c
-Secretase inhibitor
-Secretase modulator
SAR
Pyridine
Alzheimer’s disease (AD) is a chronic and relentlessly progres-
transmembrane receptor involved in the regulation of cell differen-
tiation, leads to abnormalities in the gastrointestinal tract and the
immune system.8 To avoid adverse reactions mediated by inhibi-
tion of Notch processing, research focus has been shifted to devel-
sive neurologic disorder, affecting over 30 million people world-
wide, and is characterized by specific patterns of neuronal loss
leading to significant cognitive and other functional impairments.1
The deposition of fibril Tau and Ab within the CNS are the defining
neuropathological hallmarks of AD. The latter are thought to be
causative of the disease and consist of amyloid-beta (Ab) peptides
of 40–42 amino acids (Ab40 and Ab42, respectively).2 These pep-
tides are produced from amyloid precursor protein (APP) by the
O
F
N
N
N
N
H
N
1
sequential action of b-secretase and
effort has been invested to identify inhibitors of either enzyme to
reduce Ab production.3
-secretase inhibitors (GSIs) have been
c-secretase, thus a significant
c
Figure 1. Pyridazine-derived GSM.
shown to successfully reduce brain Ab dose-dependently in pre-
clinical models.4 Several GSIs have been progressed to clinical
development with LY-450139 as the most advanced. However, its
Phase III clinical trial was recently halted due to lack of cognitive
improvement in AD patients and higher risk of developing skin
cancer upon treatment (Eli Lilly News Release, 2010).
F
N
H2N
200 oC, 2 h
N
F
I
Cl
I
NH
Consistent with the unexpected outcomes of LY-450139, some
microwave
40%
2
preclinical studies showed that inhibition of c-secretase may im-
3
pair learning and memory5 and induce skin cancer.6 Causes of
these adverse outcomes are not completely understood. However,
MeO
N
OH
MeO
N
it is known that many other substrates are cleaved by c-secretase
N
B
N
F
in addition to APP.7 Toxicity may arise through inhibition of the
processing of these other substrates. For example, inhibition of
4
OH
N
NH
90 oC,1.5 h, microwave
cleavage of Notch, which is a known c-secretase substrate and a
Pd(PPh3)4, K2CO3, Toluene
40%
5
⇑
Corresponding author.
E-mail address: Zehong.2.Wan@gsk.com (Z. Wan).
Scheme 1. Synthetic strategy for analogs 5 and 11–24.
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.06.042

Z. Wan et al. / Bioorg. Med. Chem. Lett. 21 (2011) 4832–4835
4833
Table 1
In vitro SAR for pyridine derivatives
Me
HET2
HET1
NH
F
R1
Compd
HET1
R1
HET2
Ab42a pIC50
Ab40a pIC50
Notcha pIC50
N
N
N
N
N
1
–OCH3
7.2
6.7
5.0
N
N
N
N
N
11
12
–OCH3
–OCH3
7.2
6.7
6.7
6.0
<4.7
5.0
N
N
N
N
N
N
13
–OCH3
6.6
5.9
5.0
N
N
N
N
N
N
N
5
–OCH3
–OCH3
6.9
6.2
6.1
5.0
5.0
5.2
N
14
N
15
16
17
–OCH3
–OCH3
–OCH3
6.1
7.1
6.5
4.3
6.2
4.5
NA
NA
NA
S
N
N
N
N
N
N
N
S
O
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
N
18
19
20
21
22
23
24
10
–CONHCH3
–N(CH3)2
–Ph
<4
<4
NA
NA
NA
NA
NA
NA
NA
4.8
N
N
N
N
N
N
N
4.3
4.8
5.2
6.4
6.3
5.5
6.6
4.4
5.0
4.8
5.7
5.3
5.1
5.7
–CH3
–Cl
–OCH2CH3
–OCH(CH3)2
–OCH3
S
N
N
N
N
N
25
26
–OCH3
6.4
6.4
5.7
5.3
4.9
5.0
N
N
–OCH3
–OCH3
S
N
27
6.3
5.2
5.2
a
Ab42, Ab40 and Notch were assayed in the same way as reported previously.10
oping
of APP by
c
-secretase modulators (GSMs). Heterogeneous proteolysis
-secretase leads to the production of Ab peptides in var-
intact. Therefore, GSMs are believed to be a better tolerated
Ab-lowering approach than GSIs.10,11
c
ious lengths, ranging from 37 to 43 amino acid residues. While GSIs
inhibit production of all Ab peptide species, GSMs shift the
cleavage site towards production of shorter nonamyloidogenic
Our research program was aimed at discovering novel, potent
and brain penetrant GSMs for AD. The previous letter described a
novel series of pyridazine-derived GSMs (1, Fig. 1).12 Herein, we
describe the subsequent optimization of this series leading to the
identification of a novel pyridine series. Proof of in vivo activity
peptides.9 Because GSMs selectively inhibit
c-secretase activity,
Notch processing is spared and its signaling function is largely

4834
Z. Wan et al. / Bioorg. Med. Chem. Lett. 21 (2011) 4832–4835
will be disclosed for both pyridazine and pyridine series in this
Letter.
were profiled to evaluate their potential as time-dependent CYP
inhibitors. Based on these data, analog 5 emerged as the most
interesting compound for studying mechanism of action (Fig. 2).
SHSY5Y-APPswe cells were thus pre-incubated with 5 for 24 h.
Medium was harvested and subject to analysis of Ab42, Ab40, total
Ab and cell viability (determined via WST-1). Total Ab in this assay
Synthesis: Pyridine derivative 5 was prepared from 2-chloro-5-
iodopyridine (2, Scheme 1). Displacement of chlorine with (S)-1-
(4-fluorophenyl)-ethanamine provided intermediate 3. Suzuki
coupling with boronic acid 4 then afforded compound 5. This syn-
thetic route was utilized to prepare compounds 11–24 (Table 1) by
replacing 2 and 4 with appropriate reactants. Compound 10 was
synthesized from 4-bromo-3-methoxyphenyl 4-methylbenzene-
sulfonate (6, Scheme 2). Formation of boronic acid 7 followed by
Suzuki coupling with 5-bromo-3-methylisothiazole provided 8.
Another round of acid (or boronate) formation and Suzuki coupling
then furnished 10. This synthetic sequence was applied to prepare
pyridine derivatives 25–27 (Table 1) by selecting appropriate start-
ing materials to replace the isothiazole.
system includes all Ab generated by
c-secretase, such as Ab36,
Ab37, Ab38 and Ab40. Since levels of Ab42, Ab40 and total Ab were
determined using different assays, the magnitude of reduction of
Ab42 and Ab40 and the magnitude of increase of total Ab were
not completely matched. The levels of Ab42 and Ab40 were de-
creased in a concentration-dependent manner after treatment
(0.01–3 lM), while total Ab levels were maintained or increased
indicating the rise in shorter Ab species compensating for the de-
crease in Ab42 and Ab40. This is in contrast to the profile of inhib-
itors such as LY-450139 which decreases levels of all Ab species
and thus the total Ab. Furthermore, 5 was found not to be cytotoxic
Results and discussion: After discovering pyridazine analog 1 as a
potent and brain penetrant c-secretase modulator, we then turned
our attention to exploring the SAR profile of the two heterocycles
(HET1 and HET2) and substituent R1 in order to identify another
potent GSM from a different chemical class for in vivo evaluation.
Replacement of the pyridazine (HET2) with other 6-/5-membered
heterocycles was explored first. Pyrimidine analogs (12 and 13)
are slightly less potent. Regio-isomers of pyridine exhibit different
potency and 2-pyridine (5) is more potent than 3-pyridine (14).
Thiadiazole (16) is the most potent among the three 5-membered
heteroaryl groups we studied and its potency is similar to that of 1
and 11.
and it had no effect on cellular viability up to 3 lM.
The in vivo pharmacokinetic profile of 5 is listed in Table 2.
Compound 5 demonstrated low clearance and excellent oral bio-
availability in the rat.
Compound 5 (Table 3) also demonstrated good brain penetra-
tion and showed that a 30 mg/kg oral dose could deliver brain con-
centrations of 8.7
l
M which is significantly higher than the in vitro
M at Ab40) values indicating that
IC50 (0.126 M at Ab42 and 0.8
l
l
compound 5 has the potential to deliver in vivo efficacy at low
doses.
Having identified several heteroaryl groups which afforded
good levels of in vitro potency, we selected 2-pyridine due to its
similarity to pyridazine and fixed this while we varied R1 and
HET1. R1 was investigated next, but very tight SAR was observed.
Methyl amide group (18), electron rich dimethyl-amino group
(19) and the bulky phenyl group (20) were detrimental to potency.
Methyl (21), chlorine (22), ethoxy (23) and isopropoxy (24) also re-
sulted in reduced potency when compared with methoxy (5). HET1
was studied last. Isothiazole (10), pyridazine (25), thiazole (26) and
pyridine (27) are tolerated, but less potent than imidazole (5).
Compound 5 also contains the fragment of 1-(2-methoxyphenyl)-
With two brain penetrant and orally bioavailable c-secretase
modulators 1 and 5 in hand, we then evaluated their in vivo effi-
cacy at lowering Ab42 upon sub-chronic oral dosing. Both com-
pounds were dosed to naive SD rats for 5 days consecutively
4-methyl-1H-imidazole, a common component of known GSMs11c
,
variation of the R1 substitution position (e.g., 2-OCH3) was not
studied at this stage.
Following in vitro potency assessment compounds with a pIC50
value P6 were assessed in terms of in vitro metabolic stability in
rat and human liver microsomes and for inhibition of five human
CYP isoforms (CYP1A2, 2C9, 2C19, 2D6 and 3A4). Selected analogs
Figure 2. Effect of 5 on Ab levels and cell viability in SHSY5Y-APPswe cells.
N
S
nBuLi, B(iPrO)3
HCl, 87%
HO
HO
Br
MeO
OTs
B
OTs
37%
Br
Table 2
Rat PK (iv  po) of 5a
MeO
Pd(dppf)2Cl2
K2CO3
6
7
CLb (mL/min/kg)
12.2
Vdss (L/kg)
3.7
t½ (h)
3.3
Fpo (%)
100
NaOH, 76%
N(Tf)2
S
N
a
S
O
B
O
100%
Crystalline HCl salts were dosed to SD rats at 1 mg/kg (iv) and 2 mg/kg (po) with
OTs
N
vehicle of 1% (w/v) methylcellulose (po) and DMSO solution with 10% HP-b-CD (w/
v) (iv).
O
O
O
MeO
B B
MeO
8
O
Pd(dppf)2Cl2
52%
9
Table 3
Rat CNS penetration data of 5a
N
F
S
3
N
NH
Blood conc (
l
M)
Brain conc (
lM)
Br:Bl
Pd(PPh3)4
4.983 1.381
8.745 1.802
1.74 0.33
MeO
10
a
Compound was dosed orally in (30 mg/kg) 1% (w/v) methylcellulose aq. Values
are the mean from three rats. Samples were taken 6 h post-dose.
Scheme 2. Synthetic strategy for analogs 10 and 25–27.

Z. Wan et al. / Bioorg. Med. Chem. Lett. 21 (2011) 4832–4835
4835
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(30 mg/kg/day, oral). Animals were sacrificed 6 h after the last
dose. Brain samples were collected and homogenized to analyze
the levels of Ab42 and compound concentration. Reduction of
45.7% (p <0.01) and 27.6% (p <0.01) at Ab42 and brain concentra-
tion of 10.4 and 8.86 l
M were achieved by compounds 1 and 5,13
respectively. At the end of the same study, measurement of Hes-
1 mRNA levels from the ileum and flow cytometric assessment of
CD4 and CD8 markers in thymocytes were also carried out. No sig-
nificant difference from the vehicle control group was observed,
suggesting a lack of effect for both compounds on Notch process-
ing, which is consistent with their in vitro Notch inhibition data
as shown in Table 1.
In summary, a novel series of pyridines was discovered as po-
tent c-secretase modulator. Representative analog 5 from the ser-
ies demonstrated the mechanism of action consistent with that of
GSMs. Both pyridine derivative 5 and pyridazine derivative 1 dis-
played in vivo efficacy to inhibit Ab42 without altering Notch
processing.
10. Hall, A.; Elliott, R. L.; Giblin, G. M.; Hussain, I.; Musgrave, J.; Naylor, A.; Sasse, R.;
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Acknowledgment
11. (a) Kounnas, M. Z.; Danks, A. M.; Cheng, S.; Tyree, C.; Ackerman, E.; Zhang, X.;
Ahn, K.; Nguyen, P.; Comer, D.; Mao, L.; Yu, C.; Pleynet, D.; Digregorio, P. J.;
Velicelebi, G.; Stauderman, K. A.; Comer, W. T.; Mobley, W. C.; Li, Y.; Sisodia, S.
S.; Tanzi, R. E.; Wagner, S. L. Neuron 2010, 67, 769; (b) Kushida, I.; Doi, E.; Ito, K.;
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Helpful discussion with Dr. Jeffrey K. Kerns during manuscript
preparation is acknowledged.
References and notes
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d 9.26 (d,
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J = 1.5 Hz, 1H), 8.43 (s, 1H), 7.88 (s, 1H), 7.77 (s, 2H), 7.67 (s, 1H), 7.55–7.50
(m, 2H), 7.14 (t, J = 8.8 Hz, 2H), 5.25 (dd, J = 7.0, 13.8 Hz, 1H), 4.05 (s, 3H), 2.61
(s, 3H), 2.46 (s, 3H), 1.77 (d, J = 7.0 Hz, 3H) LC–MS m/z 418 [M+H]+; compound
5: 1H NMR (400 MHz, MeOD) d 8.98 (s, 1H), 8.25 (d, 1H), 7.96 (dd, J1 = 2.4 Hz,
J2 = 9.2 Hz, 1H), 7.56–7.52 (m, 2H), 7.46–7.41 (m, 3H), 7.41 (m, 1H), 7.31 (dd,
J1 = 2.2 Hz, J2 = 8.4 Hz, 1H), 7.06 (t, J = 7.8 Hz, 2H), 6.81(d, J = 8.8 Hz, 1H), 5.03
(q, J = 6.8 Hz, 1H), 3.98 (s, 3H), 2.42 (s, 3H), 1.58 (d, J = 6.8 Hz, 3H) LC–MS m/z
403 [M+H]+.
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