4-Substituted cyclohexyl sulfones as potent, orally active γ-secretase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2005.10.009

The protease γ-secretase plays a pivotal role in the synthesis of pathogenic amyloid-β in Alzheimer's disease (AD). Here, we report a further extension to a series of cyclohexyl sulfone-based γ-secretase inhibitors which has allowed the preparation of hig

Full text of DOI:10.1016/j.bmcl.2005.10.009

Bioorganic & Medicinal Chemistry Letters 16 (2006) 280–284
4-Substituted cyclohexyl sulfones as potent, orally active
c-secretase inhibitors
b
c
a
b
Ian Churcher,a, Dirk Beher, Jonathan D. Best, Jose L. Castro, Earl E. Clarke,
*
´
Amy Gentry,a Timothy Harrison,a Laure Hitzel,a Euan Kay,a Sonia Kerrad,a
Huw D. Lewis,b Pablo Morentin-Gutierrez,a Russell Mortishire-Smith,a Paul J. Oakley,a
Michael Reilly,a Duncan E. Shaw,a Mark S. Shearman,b Martin R. Teall,a
Susie Williamsa and Jonathan D. J. Wrigleyb
aDepartment of Medicinal Chemistry, The Neuroscience Research Centre, Merck Sharp and Dohme Research Laboratories,
Terlings Park, Eastwick Road, Harlow, Essex CM20 2QR, UK
bDepartment of Molecular and Cellular Neuroscience, The Neuroscience Research Centre, Merck Sharp and Dohme
Research Laboratories, Terlings Park, Eastwick Road, Harlow, Essex CM20 2QR, UK
cDepartment of In Vivo Neuroscience, The Neuroscience Research Centre, Merck Sharp and Dohme Research Laboratories,
Terlings Park, Eastwick Road, Harlow, Essex CM20 2QR, UK
Received 28 September 2005; revised 3 October 2005; accepted 4 October 2005
Available online 3 November 2005
Abstract—The protease c-secretase plays a pivotal role in the synthesis of pathogenic amyloid-b in AlzheimerÕs disease. Here, we
report a further extension to a series of cyclohexyl sulfone-based c-secretase inhibitors which has allowed the preparation of highly
potent compounds which also demonstrate robust Ab(40) lowering in vivo (e.g., compound 32, MED 1 mg/kg p.o. in APP-YAC
mice).
Ó 2005 Elsevier Ltd. All rights reserved.
AlzheimerÕs disease (AD) is a devastating disorder of the
aged population and with current palliative treatments
being of modest efficacy, the need remains for a therapy
able to alter the underlying pathophysiology of the dis-
ease. The predominantly 40–42 amino acid amyloid-b
(Ab) peptide is the major component of the extracellular
proteinaceous plaques seen in AlzheimerÕs disease (Ab)
and much evidence suggests a pivotal role for Ab in the
disease process.1 In particular, individuals possessing
autosomal dominant mutations in the genes encoding
for amyloid-b precursor protein (bAPP) or the mem-
brane-bound protein homologs presenilin 1 and 2 have
elevated Ab levels and suffer from aggressive forms of
early onset AD.2,3 These observations have led to the
hypothesis that Ab, either in its soluble form or when
aggregated into oligomers, fibrils, and subsequently pla-
ques, is responsible for neuronal toxicity and cell death.4
Ab is derived by processing of the 695–770 residue,
type I transmembrane protein bAPP.5 The major met-
abolic pathway of bAPP involves sequential cleavage
by the proteases a-secretase and c-secretase leading
to non-amyloidogenic fragments. Alternative process-
ing by stepwise cleavage mediated by b-secretase and
c-secretase leads to the production of Ab and it is
inhibitors of the latter enzyme that were targeted in
the current work.6,7
c-Secretase is a novel membrane-bound aspartyl prote-
ase complex formed from presenilin-1/2, nicastrin,
Aph-1, and pen-2 whose catalytic center is unusual in
likely being within transmembrane helices of the prese-
nilin unit. As the role of c-secretase has become better
characterized, a variety of substrates8 including the
Notch receptor have been identified leading to the pos-
sibility of mechanism-based effects9 or alternatively to
Keywords: AlzheimerÕs disease; c-Secretase; Protease inhibitor;
Amyloid; In vivo.
*
Corresponding author. Tel.: +44 1279 440000; fax: +44 1279
440390; e-mail: ian_churcher@merck.com
10
additional therapeutic uses for c-secretase inhibitors.
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.10.009

I. Churcher et al. / Bioorg. Med. Chem. Lett. 16 (2006) 280–284
281
rangement or iodide 8 by Hunsdiecker-type radical
iodination.16
These intermediates could be further elaborated to a
wide range of derivatives, a representative number of
which are summarized in Table 1. Whilst the hydroxy-
methyl analog 5 showed activity similar to that of
unsubstituted 1 only, formation of a carbamate (e.g.,
compound 9) gave an increase in potency. Similarly,
the poor activity of primary amine 7 could be improved
markedly by acylation or sulfonylation with ureas, car-
bamates, and sulfamides all being well tolerated. Addi-
tionally, several heterocycles including succinimide
provided compounds with good activity.
A whole-cell c-secretase inhibition assay using SH-
SY5Y neuroblastoma cells in which human c-secretase
catalyzes the breakdown of the overexpressed exogenous
substrate A4CTF has been developed.11 We have previ-
ously disclosed12–15 a series of cyclohexyl sulfones which
have demonstrated in vitro inhibition of Ab(40) secre-
tion in this assay. In this letter, we describe the further
optimization of these leads resulting in the identification
of highly potent analogs demonstrating in vivo reduc-
tion of brain Ab(40) in a mouse model.
The iodide 8 also allowed ready access to either sulfon-
amides (via KNO3/SO2Cl2 oxidation17 of the intermedi-
ate thiol) or sulfones (via SN2 displacement of the iodide
and subsequent oxidation) as shown in Scheme 2. The
in vitro potencies of a representative range of sulfonyl-
containing analogs are summarized in Table 2.
Based upon the 1,1-disubstituted cyclohexane 1, which
has previously been shown to be a potent c-secretase
inhibitor (IC50 3 nM),12 we chose to probe the 4-position
on the cyclohexane ring in order to improve both poten-
cy at c-secretase and also ADME properties.
The series of simple alkyl and aryl sulfones gave mostly
modest inhibitory activity (data not shown) although
cyclopropyl analog 16 was shown to be potent. Howev-
er, the utilization of an aromatic substituent bearing a
hydrogen bond acceptor at the ortho position proved
to be advantageous; anisyl derivative 20, in particular,
showing high potency.
Investigation of substitution at the 4-position was facil-
itated by the ready availability, on scale, of the cyclohex-
anone derivative 2.12 This could be elaborated to
provide a range of useful intermediates as shown in
Scheme 1. Reduction of 2 gave predominantly the
anti-alcohol 4 which underwent smooth cyanide dis-
placement via a mesylate to give, after hydrolysis and
reduction, syn hydroxymethyl 5. Alternatively, Wads-
worth–Horner–Emmons reaction followed by stereo-
selective reduction and hydrolysis gave the carboxylate
6, which was elaborated to amine 7 via Curtius rear-
Sulfonamides were also well tolerated with azetidine
derivative 24 being optimal in this series.
Whilst many of the 4-substituted cyclohexyl analogs
prepared showed excellent in vitro potency, their in vivo
efficacy profiles were often compromised by poor meta-
bolic stability prompting us to investigate the cause of
this issue. It was postulated that the activated methylene
exo to the cyclohexyl core may be a site of metabolism
and this was confirmed in the case of methyl analog 25
with the isolation of acid 26 as a major metabolite fol-
lowing in vitro incubation with rat liver microsomes
(Fig. 1).
To address this potential route of metabolism, com-
pounds lacking this labile methylene unit were next pre-
pared. Initially, a wide range of functional groups were
introduced directly linked to the cyclohexyl ring at the 4-
position with the potency of a representative number
summarized in Table 3. These compounds were avail-
able from the syn alcohol 3 (Scheme 1) or syn amine
27 (Scheme 3).
In most cases, the presence of a polar group directly at-
tached to the cyclohexyl core was attended with inferior
in vitro potency (note, in particular, the parent alcohol
3 and amine 27). Acylation of 3 or 27 was also not produc-
tive, except in the case of carbamate 28 where potency was
at best improved only to the level of unsubstituted 1.
Scheme 1. Reagents and conditions: (i) NaBH4, EtOH, 10 °C (anti:syn
6:1); (ii) MsCl, Et3N, DCM, À50 °C; (iii) Bu4NCN, PhMe, 75 °C; (iv)
i
AcOH/concd HCl, 110 °C; (v) BuOCOCl, Et3N, THF then NaBH4,
H2O; (vi) (EtO)2P(O)CH2CO2Et, NaH, THF; (vii) L-Selectride, THF,
À40 °C; (viii) LiOH, aq MeOH; (ix) (COCl)2, DMF, THF then NaN3,
Bu4NBr, PhH then BnOH, reflux; (x) HBr/AcOH then NaOH; (xi)
PhI(OAc)2, I2, PhH, reflux, ht.
In marked contrast to acylated amino analogs, extend-
ing this work to a series of sulfonamides and sulfamates
gave compounds with increased potency.

282
I. Churcher et al. / Bioorg. Med. Chem. Lett. 16 (2006) 280–284
Table 1. In vitro c-secretase inhibition of compounds 5–14
Compound
Preparation
R
IC50 (nM)
5
6
Scheme 1
Scheme 1
OH
CO2H
4.7 1.7
21.2 1.5
148 28
14.3 7.2
0.51 0.15
1.3 0.2
1.0 0.1
2.2 0.1
1.7 0.3
7
Scheme 1
Scheme 1
NH2
I
8
9
5, 4-Nitrophenyl chloroformate, pyr, THF then MeNH2/EtOH
7, Ac2O, DMAP, Et3N, DCM
6, DPPA, Et3N, PhMe, 110 °C then NH3/dioxan, rt
7, MeO2CCl, Et3N, DCM, 0 °C
7, catechol sulfate, THF then ethylamine/dioxan
OCONHMe
NHAc
10
11
12
13
NHCONH2
NHCO2Me
NHSO2NHEt
O
N
14
5, MsCl, Et3N, DCM then succinimide, NaH, DMF, 80 °C
1.0 0.4
O
Table 2. In vitro c-secretase inhibition of compounds 16–24
Compound
R
IC50 (nM)
1.4 0.3
16
cPr
17
18
19
20
21
22
23
24
CF3
Ph
21.5
5
8.8 1.7
0.65 0.1
0.51 0.02
2.1 0.2
Scheme 2. Reagents and conditions: (i) KSAc, DMF; (ii) NaOH, aq
MeOH; (iii) KNO3, SO2Cl2, MeCN; (iv) RSH, KOH, EtOH, reflux; (v)
cat. RuO2, NaIO4, EtOAc/H2O; (vi) R1R2NH, THF.
OH
OMe
N
In the series of sulfonamides, alkyl substituents were
well tolerated with n-propyl (31) and trifluoromethyl
(32) substituents preferred. A series of aryl sulfonamides
was also prepared and whilst the observation noted ear-
lier of the favorable effect of introduction of ortho, elec-
tronegative substituents appeared not to translate to this
series, several potent heterocyclic analogs (e.g., 33)
could be prepared. With some parallel to the earlier ser-
ies of sulfamides (Table 2), this functional group was
further utilized to give potent compounds with the azeti-
dine derivative 35 giving extremely high levels of
activity.
NH2
4.2 1.4
NMe2
1.1 0.3
0.36 0.02
N
very similar IC50s18 and inhibited the cleavage of Notch
receptor19 at concentrations similar to those required for
inhibition of APP processing,20 consistent with the mod-
el that c-secretase is required for both of these processes.
On consideration of in vitro potency and ADME pro-
files of the potent inhibitors, the trifluoromethane sul-
fonamide 32 was selected for further work. Compound
32 inhibited the generation of Ab(40) and Ab(42) with

I. Churcher et al. / Bioorg. Med. Chem. Lett. 16 (2006) 280–284
283
Figure 1. In vitro metabolism of methyl analog 25.
Scheme 3. Preparation of amine 27. Reagents and conditions: (i)
MsCl, Et3N, DCM, À50 °C; (ii) NaN3, DMF, 90 °C; (iii) PPh3, aq
THF.
This compound was profiled in vivo in the APP-YAC
mouse model21 and demonstrated a robust decrease of
DEA-extractable brain Ab(40) levels22,23 4 h after oral
dosing with a minimum effective dose of 1 mg/kg
(Fig. 2). A time-course study at 10 mg/kg showed excel-
lent duration of action with a significant lowering of
brain Ab(40) levels throughout a 24 h period.
inhibitors, such as 32, which was shown to have excel-
lent efficacy in reducing central Ab(40) levels in APP-
YAC mice with a minimum effective dose of 1 mg/kg
p.o.
In summary, we have developed a series of 4-substituted
cyclohexyl sulfones which inhibit c-secretase in vitro
in the low to sub-nanomolar range. This high level of
in vitro potency could be coupled with good pharmaco-
kinetics to produce potent, orally active c-secretase
Acknowledgments
The authors thank Beth Oxley, Ian Gowers, and Robert
Newman for assistance in screening.
Table 3. In vitro c-secretase inhibition of compounds 27–35
Compound
Preparation
Scheme 1
R
IC50 (nM)
10.7 1.7
3
OH
28
27
29
30
31
32
33
34
35
3, ClSO2NCO, THF, 0 °C
Scheme 3
OCONH2
NH2
1.9 0.5
510 10
27, Ac2O, Et3N, DCM
NHAc
35
3
27, MsCl, Et3N, DCM
NHSO2Me
NHSO2Pr
NHSO2CF3
4.2 1.2
0.82 0.22
0.65 0.13
0.25 0.07
0.66 0.01
0.15 0.07
27, PrSO2 Cl, Et3N, DCM
27, Tf2O, Et3N, DCM
H
N
27, ArSO2Cl, Et3N, DCM
27, ArSO2Cl, Et3N, DCM
27, [reagent from azetidine/SO2Cl2], HunigÕs base, MeCN
S
S
O2
H
N
S
O2
N
H
N
N
S
O2

284
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