Fluorinated piperidine acetic acids as γ-secretase modulators

Article abstract of DOI:10.1016/j.bmcl.2009.11.034

We report herein a novel series of difluoropiperidine acetic acids as modulators of γ-secretase. Synthesis of 2-aryl-3,3-difluoropiperidine analogs was facilitated by a unique and selective β-difluorination with Selectfluor. Compounds 1f and 2c were selec

Full text of DOI:10.1016/j.bmcl.2009.11.034

Bioorganic & Medicinal Chemistry Letters 20 (2010) 755–758
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Fluorinated piperidine acetic acids as
c-secretase modulators
a
a
a
c
Matthew G. Stanton a, , Jed Hubbs , David Sloman , Christopher Hamblett , Paula Andrade ,
Minilik Angagaw c, Grace Bi a, Regina M. Black f, Jamie Crispino a, Jonathan C. Cruz b, Eric Fan d,
Georgia Farris e, Bethany L. Hughes a, Candia M. Kenific b, Richard E. Middleton a, George Nikov a,
Peter Sajonz f, Sanjiv Shah b, Nirah Shomer c, Alexander A. Szewczak a, Flobert Tanga b, Matthew T. Tudge d,
Mark Shearman b, Benito Munoz a
*
a Department of Drug Design and Optimization, Merck Research Laboratories, 33 Avenue Louis Pasteur, Boston, MA 02115, USA
b Department of Neuroscience Drug Discovery, Merck Research Laboratories, 33 Avenue Louis Pasteur, Boston, MA 02115, USA
c Department of Laboratory Animal Resources, Merck Research Laboratories, 33 Avenue Louis Pasteur, Boston, MA 02115, USA
d Department of Process Research, Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065, USA
e Department of Safety Assessment, 770 Sumneytown Pike, PO Box 4, West Point, PA 19486, USA
f Department of Preparative Chemistry and Separations, PO Box 2000, Rahway, NJ 07065, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
We report herein a novel series of difluoropiperidine acetic acids as modulators of c-secretase. Synthesis
Received 18 September 2009
Revised 9 November 2009
Accepted 10 November 2009
Available online 13 November 2009
of 2-aryl-3,3-difluoropiperidine analogs was facilitated by a unique and selective b-difluorination with
SelectfluorÒ. Compounds 1f and 2c were selected for in vivo assessment and demonstrated selective low-
ering of Ab42 in a genetically engineered mouse model of APP processing. Moreover, in a 7-day safety
study, rats treated orally with compound 1f (250 mg/kg per day, AUC0–24 = 2100 lM h) did not exhibit
Notch-related effects.
Keywords:
Ó 2009 Elsevier Ltd. All rights reserved.
Alzheimer’s disease
c
-Secretase
Alzheimer’s disease (AD) is a progressive, chronic neurodegen-
Although inhibiting
c
-secretase is an attractive strategy for treat-
erative disorder characterized by impairments in memory and cog-
nition.1 In the United States, estimates suggest that an individual
develops AD every 72 seconds. By 2050, this rate is projected to in-
crease to one person developing the disease every 33 seconds2 and
the number of people worldwide with the disease is expected to
triple by the year 2050, intensifying what is already a serious
worldwide public health problem.3
Accumulation of amyloid-b (Fb) peptides and formation of
amyloid plaques is a central event in sporadic and familial AD
pathology.4 Proteolytic cleavage of amyloid precursor protein
(APP) by two membrane-bound aspartyl proteases, b-secretase
ing AD, it has led to undesirable adverse events in clinical trials, most
notably gastrointestinal toxicity.5 These side effects are associated
with blocking the processing of Notch, a transmembrane receptor
signaling protein which is also a
of -secretase was introduced as a strategy to avoid Notch-related
c
-secretase substrate.6 Modulation
c
toxicity when certain non-steroidal anti-inflammatory drugs were
found to selectively inhibit the production of Ab42 while not affect-
ing the production of Ab40 or Notch processing.7,8
Piperidine acetic acid
c-secretase modulators have been the
subject of a recent patent application by our group.9 In this Letter,
we describe modulators within this series where fluorine is incor-
porated into the piperidine ring as a part of our continuing effort to
identify compounds with desirable pharmacokinetic (PK) proper-
ties (Fig. 1).
(BACE) and
c-secretase, leads to the formation of Ab. Among the
two major Fb peptides produced by
c-secretase cleavage, Fb40
and Ab42, the less common Ab42 fragment is thought to play the
most important role in AD pathology.4 Pharmacological interven-
tion by inhibiting the function of these two enzymes with a small
molecule, and thus reducing Ab levels, has been and remains an
attractive strategy for developing disease modifying AD treatment.
The synthetic strategy employed to prepare 4,4-difluoropiperi-
dine acetic acids with the general formula 1 is outlined in
Scheme 1. Enone 4 was prepared by treating 4-methoxypyridine
(3) with benzyl chloroformate and (p-trifluoromethylphenyl)mag-
nesium bromide using the method previously reported by Comins
et al.10,11 Treatment of 4 with LiHMDS in THF followed by quench-
ing with methyl bromoacetate and reduction with L-SelectrideÒ
afforded trans-ketopiperidine 5. Subsequent treatment of ketopi-
peridine 5 with Deoxo-FluorTM followed by hydrogenation afforded
* Corresponding author at present address: Merck Research Laboratories, WP14-
3, 770 Sumneytown Pike, PO Box 4, West Point, PA 19486, USA. Tel.: +1 215 652
6975; fax: +1 215 652 3971.
E-mail address: matthew_stanton@merck.com (M.G. Stanton).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.11.034

756
M. G. Stanton et al. / Bioorg. Med. Chem. Lett. 20 (2010) 755–758
HOOC
CO2Me
Ar
CO2Me
Ar
COOH
F
Me
F
F
F
a, b
c, d
N
N
N
H
N
N
Cl
R3
CF3
9
R1
R2
CF3
8
10
R4
CF3
Ar =
1
2
e, f
Figure 1. Novel class of difluoropiperidine acetic acids.
HO2C
MeO2C
MeO2C
F
F
F
F
O
O
CO2Me
OMe
g
h, i
N
Ar
N
Ar
N
Ar
a
b, c
OH
R1
R1
R1
N
Cbz
4
Ar
N
Cbz
5
Ar
N
R2
R2
R2
3
2
d-f
12
11
Scheme 2. Synthetic strategy for the preparation of 3,3-difluoropiperidine acetic
acids (2). Reagents and conditions: (a) 4-trifluoromethylbenzeneboronic acid,
Pd(PPh3)4, Na2CO3, DME, H2O; (b) Me2CO3, LDA, THF, À78 °C to 0 °C; (c) H2
(20 psi), HCl, Pt2O, MeOH; (d) L-(+)-mandelic acid, i-PrOH; (e) AuBr3 (5 mol %),
R1CHO, R2CCH, H2O, 75 °C; (f) H2 (50 psi), Pd/C (10%), EtOH; (g) SelectfluorÒ, DMF
then H2O; (h) BH3ÁTHF, THF; (i) LiOH, H2O, THF, MeOH.
COOH
Ar
CO2Me
F
F
F
F
CO2Me
Ar
F
F
g
h, i
N
N
Ar
N
H
R2
R2
R1
R1
1e-h
7
6
i
MeO2C
MeO2C
MeO2C
CO2H
F
F
Selectfluor®
Ar
- HF
N
Ar
N
Ar =
N
Ar
F
N
Ar
R1
R1
R1
R2
CF3
R2
R2
R2
R1
1b-d
14
11
13
Scheme 1. Synthetic strategy for the preparation of 4,4-difluoropiperidine acetic
acids (1). Reagents and conditions: (a) Cbz-Cl, (p-trifluoromethylphenyl)magne-
sium bromide, THF, À78 °C to 0 °C; (b) LiHMDS, methyl bromoacetate, THF, À78 °C;
(c) L-SelectrideÒ, THF, À78 °C; (d) Deoxo-FluorTM, DCM; (e) Pd/C, H2, MeOH; (f) chiral
resolution, HPLC (ChiracelÒ-AD 10% i-PrOH/heptane); (g) alkyne, aldehyde, AuBr3,
2 equiv
Selectfluor®
MeO2C
H20, 75 °C, lW; (h) Raney-Nickel, MeOH, H2, 60 psi; (i) 1 M KOH, MeOH.
Cl
2 BF4
F
N
N
F
H2O
the methyl 4,4-difluoropiperidineacetic acid methyl ester 6, which
was resolved using a Chiral pakÒ AD-H column on multi-gram
scale. Enantiomerically pure amine 6 could be alkylated with
methyl iodide and the ester hydrolyzed to give compound 1a
(not shown). For the preparation of more complex, N-branched car-
boxylic acids, the key step to afford the 4,4-difluoropiperidineacetic
acids with the general structure 1 was a three-component gold(III)
bromide catalyzed Mannich reaction with the piperidine 6.12 The
acetylene analogs 7 were reduced with Raney-Nickel at 60 psi H2
in methanol from 24 to 48 h to afford the fully saturated products.
Methyl esters were treated with potassium hydroxide in methanol
to afford the desired 4,4-difluoropiperidine acetic acids 1.
The synthesis of the 3,3-difluoropiperidine acetic acids with the
general structure 2 proceeded as outlined in Scheme 2. Pyridine 8
was coupled to 4-trifluoromethylbenzeneboronic acid under Suzu-
ki reaction conditions13 to afford the biaryl pyridine, which was
subsequently acylated with dimethylcarbonate to afford methyl
ester 9. Ester 9 was reduced with Adam’s catalyst in methanolic
HCl to give the corresponding piperidine, which could then be re-
solved by recrystallization as its mandelate salt. Enantiomerically
pure piperidine 10 was subjected to the three-component gold(III)
bromide catalyzed Mannich reaction described above, followed by
palladium-catalyzed hydrogenation to afford piperidine 11. The
key transformation in this synthetic sequence was difluorination
12
F
N
Ar
Selectfluor®
R1
R2
15
Figure 2. Putative piperidine fluorination mechanism.
of 11 with SelectfluorÒ, which provided the difluorohemiaminal
12. The desired -secretase modulators of the general structure 2
were obtained by first reducing the hemiaminal to the amine with
borane THF complex, followed by hydrolysis of the ester with lith-
ium hydroxide.14
A possible mechanism for the difluorination reaction is proposed
in Figure 2. Piperidine 11 is fluorinated on nitrogen to give ammo-
nium intermediate 13, which subsequently eliminates HF to give
an iminium ion that can tautomerize to afford enamine 14. This en-
amine is difluorinated by SelectfluorÒ to give iminium ion 15 that
can be hydrated upon workup to provide hemiaminal 12.
The required structural features for potency in these series is
the presence of the carboxylic acid at either the 3 or 4 position,
the aryl substituent at the 2 position, and functionalization of the
piperidine nitrogen. The carboxylic acid moiety at the 3 or 4
c

M. G. Stanton et al. / Bioorg. Med. Chem. Lett. 20 (2010) 755–758
757
Table 3
Rodent PK for 1f and 2c
position was ideally suited for incorporation of fluorine and led to
an attenuation of the pKa of the nitrogen. The initial SAR utilized
racemic material, however, select compounds were further evalu-
ated using the enantiomerically pure amine 6. The SAR is summa-
rized in Table 1.
1f
2c
Dose (iv; po) (mg/kg)
Clp (mL/min/kg)
Vdss (L/Kg)
t1/2 (iv) (h)
%F
1; 2
3.8
1.9
5.1
160
13
1; 2
9.1
5.1
6.4
20
Increasing alkyl substitution beyond the methyl group of 1a led
to an enhancement in potency against Ab42. The preferred stereo-
chemical requirements of the 4,4-difluoropiperidine acetic acid
scaffold was determined based on the results of 1b and the subse-
quent enantiomerically pure isomers 1c and 1d, indicating that the
activity resided in the enantiomerically pure 1d. Reduction of the
alkyne led to compound 1f. Compound 1f exhibited an IC50 against
Notch activity of greater than 10,000 nM (data not shown).16
The optimal N-alkyl substituents identified in Table 1 were
incorporated into the examples of the 3,3-difluoropiperidineacetic
acids of structure 2. The results are summarized in Table 2. The
data indicate that 3,3-difluoro- and 4,4-difluoropiperidine analogs
AUCN0–24 po (
l
M h kg/mg)
1.2
Table 4
Relative levels of Ab42 and Ab40 inhibition in APP-YAC transgenic mouse model for 1f
and 2c
Ab42 (%)
Ab40
Brain (
lM)
Plasma (lM)
1f
2c
À84
À64
N.S.
N.S.
0.58
N.A.
1.1
N.A.
have similar activity against c-secretase.
Compounds 1f and 2c exhibited favorable rodent PK (Table 3)
and were tested in APP-YAC transgenic mice and non-transgenic
rats.
N.S. = not significant.
Initial efficacy screening in mice (10 mg/kg po, 7 h, Table 4)
demonstrated that 1f and 2c led to a selective inhibition of brain
Ab42 levels relative to Ab40 (Table 4). Exposure levels of 1f in
Further evaluation of 1f in rats (1, 3, 10, and 30 mg/kg po, 7 h)
demonstrated a dose-dependent lowering of Ab42 (ED50 = 5 mg/
kg, brain EC50 = 1 lM, and plasma EC50 = 3.7 lM). Maximum inhi-
mouse brain/plasma were 0.58/1.14 lM.
bition of brain Ab42 levels after treatment with 1f was observed
at 7 h (data not shown).
In summary, a novel series difluoropiperidine acetic acids were
discovered that showed moderate potency against the c-secretase
complex in vitro and demonstrated robust PK/PD activity in estab-
lished rodent models. The selective lowering of Ab42 without
effecting Ab40 or Notch activity is consistent with the mechanism
Table 1
In vitro activity of 4,4-difluoropiperidine acetic acids (1) against Ab42 and Ab40
CO2H
F
F
of
study with 1f (250 mg/kg per day, dosing 7 days, five animals, fe-
males, AUC0–24 = 2100 M h) no adverse Notch effects were ob-
served. These results further validate that modulation may prove
to be a more formidable approach to targeting -secretase with a
c-secretase modulation. Moreover, in a 7-day oral rat safety
N
l
R1
R2
CF3
c
Compound
R1
R2
Ab42
Ab40
IC5015(nM)
IC50145(nM)
small molecule in order to circumvent the undesired affects asso-
ciated with Notch.
1ac
1ba
1cb
1dc
1eb
1fc
H
H
>10,000
800
1500
390
1400
600
640
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
>10,000
CF3CH2CH2
CF3CH2CH2
CF3CH2CH2
CF3CH2CH2
CF3CH2CH2
CF3CH2CH2
i-PrCH2CH2
t-BuCC
t-BuCC
t-BuCC
t-BuCH2CH2
t-BuCH2CH2
Me3SiCH2CH2
i-PrCH2CH2
References and notes
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Ther. 2008, 12, 704.
1gc
1hc
880
a
b
c
Racemic.
From piperidine enantiomer 1.
From piperidine enantiomer 2.
4. (a) Dickson, D. W. J. Neuropathol. Exp. Neurol. 1997, 56, 321; (b) Hardy, J.;
Selkoe, D. J. Science 2002, 297, 353.
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Z.; Datar, R.; Murtaught, L. C.; Melton, D. A. Proc. Natl. Acad. Sci. U.S.A. 2005, 102,
12443.
Table 2
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In vitro activity of 3,3-difluoropiperidine acetic acids (2) against Ab42 and Ab40
7. (a) In’t Veld, B. A.; Ruitenberg, A.; Hofman, A.; Launer, L. J.; Van Duijn, C. M.;
Stijnen, T.; Breteler, M. M. B.; Stricker, B. H. C. N. Eng. J. Med 2001, 345, 1515; (b)
Weggen, S.; Eriksen, J. L.; Das, P.; Sagi, S. A.; Wang, R.; Pietrzik, C. U.; Findlay, K.
A.; Smith, T. E.; Murphy, M. P.; Butler, T.; Kang, D. E.; Marquez-Sterling, N.;
Golde, T. E.; Koo, E. H. Nature 2001, 414, 212; (c) Lim, G. P.; Yang, F.; Chu, T.;
Chem, P.; Beech, W.; Teter, B.; Tran, T.; Ubeda, O.; Hsaio Ahse, K.; Reautschy, S.
A.; Cole, G. M. J. Neurosci. 2000, 20, 5709; (d) Gasparini, L.; Ongini, E.; Wenk, G.
L. J. Neurochem. 2004, 91, 521.
O
HO
F
F
N
8. (a) Imbimbino, B. P. Curr. Top. Med. Chem. 2008, 8, 54; (b) Peretto, I.; La Porta, E.
Curr. Top. Med. Chem. 2008, 8, 38.
R1
CF3
9. (a) Hannam, J. C.; Kulagowski, J. J.; Madin, A.; Ridgill, M. P.; Seward, E. M.
WO20060430064, 2006; (b) Hannam, J. C.; Hartmann, S.; Madin, A.; Ridgill, M.
P. WO2007110667, 2007; (c) Madin, A.; Ridgill, M. P.; Kulagowski, J.J.
WO2007116228, 2007; (d) a recent publication describes similar piperdine
carboxylic acid gamma-secretase modulators inspired by our patent sited
above: Hall, A.; Elliot, R. L.; Giblin, G. M. P; Hussain, I.; Musgrave, J.; Naylor, A.;
Sasse, R.; Smith, B. Bioorg. Med. Chem. Lett., in press.
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Foley, M. A.; Comins, D. L. J. Am. Chem. Soc. 1988, 100, 7445; (c) Comins, D. L.;
LaMunyon, D. H. Tetrahedron Lett. 1989, 30, 5053; (d) Comins, D. L.; Zeller, E.
R2
Compound
R1
R2
Ab42
IC50 (nM)
Ab40
IC50 (nM)
15
15
2a
2b
2c
4-CF3Ph
CF3CH2CH2
CF3CH2CH2
i-Pr
t-Bu
710
490
230
>10,000
>10,000
>10,000
Si(CH3)3

758
M. G. Stanton et al. / Bioorg. Med. Chem. Lett. 20 (2010) 755–758
Tetrahedron Lett. 1991, 32, 5889; (e) Comins, D. L.; Joseph, S. P.; Goehring, R. R. J.
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14. Reduction of the hemiaminal provided a 4:1 ratio of C2 stereoisomers, favoring
the isomer as shown. The selectivity is thought to arise from the reduction of
the intermediate imine via hydride delivery to the less hindered face (governed
by the 4-substituent).
Lewis, H. D.; Pattison, C.; Reilly, M.; Harrison, T.; Shearman, M. S.; Williamson,
T. L.; Atack, J. R. J. Pharmacol. Exp. Ther. 2005, 313, 902; (b) Clarke, E. E.;
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16. HeLa cells were made to co-express nonfunctional halves of luciferase, one
fused to Notch
Notch E results in release of Notch intracellular domain (NICD)/N-terminal
luciferase, which translocates to the nucleus and binds the RBP-J /C-terminal
luciferase to form functional luciferase enzyme. This split-luciferase
DE and the other to RBP-Jj. c-Secretase mediated cleavage of
D
j
a
15. Ab40 and Ab42 IC50 measured using electrochemiluminescent detection of
peptides secreted by SH-SY5Y cells stably overexpressing the b-APP C-terminal
fragment SPA4CT (a) Best, J. D.; Jay, M. T.; Otu, F.; Ma, J.; Nadin, A.; Ellis, S.;
complementation system is used to detect NICD levels by measuring total
luminescence upon addition of luciferin to lysed cells. Paulmurugan, R.;
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