The discovery of MK-0674, an orally bioavailable cathepsin K inhibitor

Article abstract of DOI:10.1016/j.bmcl.2009.12.083

MK-0674 is a potent and selective cathepsin K inhibitor from the same structural class as odanacatib with a comparable inhibitory potency profile against Cat K. It is orally bioavailable and exhibits long half-life in pre-clinical species. In vivo studies

Full text of DOI:10.1016/j.bmcl.2009.12.083

Bioorganic & Medicinal Chemistry Letters 20 (2010) 887–892
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
The discovery of MK-0674, an orally bioavailable cathepsin K inhibitor
a
a
a
a
b
Elise Isabel a, , Kevin P. Bateman , Nathalie Chauret , Wanda Cromlish , Sylvie Desmarais , Le T. Duong ,
Jean-Pierre Falgueyret a, Jacques Yves Gauthier a, Sonia Lamontagne a, Cheuk K. Lau a, Serge Léger a,
Tammy LeRiche a, Jean-François Lévesque a, Chun Sing Li a, Frédéric Massé a, Daniel J. McKay a,
Christophe Mellon a, Deborah A. Nicoll-Griffith a, Renata M. Oballa a, M. David Percival a, Denis Riendeau a,
Joël Robichaud a, Gideon A. Rodan b, Sevgi B. Rodan b, Carmai Seto a, Michel Thérien a, Vouy Linh Truong a,
Gregg Wesolowski b, Robert N. Young a, Robert Zamboni a, W. Cameron Black a
*
a Merck Frosst Centre for Therapeutic Research, 16711 TransCanada Hwy, Kirkland, Québec, Canada H9H 3L1
b Department of Bone Biology and Osteoporosis, Merck Research Laboratories, West Point, PA 19846, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
MK-0674 is a potent and selective cathepsin K inhibitor from the same structural class as odanacatib with
a comparable inhibitory potency profile against Cat K. It is orally bioavailable and exhibits long half-life in
pre-clinical species. In vivo studies using deuterated MK-0674 show stereoselective epimerization of the
alcohol stereocenter via an oxidation/reduction cycle. From in vitro incubations, two metabolites could
be identified: the hydroxyleucine and the glucuronide conjugate which were confirmed using authentic
synthetic standards.
Received 3 November 2009
Revised 16 December 2009
Accepted 18 December 2009
Available online 28 December 2009
Keywords:
Cathepsin K
Ó 2009 Published by Elsevier Ltd.
MK-0674
Glucuronide conjugate
Deuteration experiment
Osteoporosis is a disease characterized by low bone mass and
structural deterioration of bone tissues. This leads to bone fragility
and an increased susceptibility to fractures, especially of the hip,
spine and wrist. Significant risk has been reported in aging popula-
tions, particularly post-menopausal women being most suscepti-
ble. Indeed, 80% of people affected by this disease are women.
Age is the highest risk factor in both men and women: up to 55%
of the people over 50 are suspected to have low bone mass density
or osteopenia. Because bone loss is asymptomatic, osteoporosis is
often known as ‘the silent thief’. As the population continues to
age, the burden of osteoporosis on medical care systems will
increase. According to the National Osteoporosis Foundation, in
2005 osteoporosis-related fractures were responsible for an esti-
mated $17 billion costs in the USA. By 2025, these costs could rise
as high as $25 billion.1 Bisphosphonates, estrogen replacement
therapy and parathyroid hormone are the current standard care
for osteoporosis.2,3 Because these treatments do not suit all
patients and reduce approximately 50% of fracture risk, there is
an urgent need for new therapies.
Approximately 15 years ago, cathepsin K (Cat K) was isolated
and characterized.4 There is strong evidence that Cat K plays a ma-
jor role in bone degradation.5,6 Many Cat K inhibitors have been re-
ported in the literature7 since then, including balicatib,8 relacatib9
and odanacatib, currently in Phase III fracture prevention trial in
post-menopausal women with osteoporosis.10 Herein, we report
on the successful identification of MK-0674, an orally bioavailable
inhibitor from the same structural class as odanacatib with a dif-
ferent metabolic profile.
Odanacatib (Fig 1), in human hepatocyte incubations, is pre-
dominantly metabolized by CYP3A4.11 The purpose of this work
was to maintain the beneficial features of odanacatib in P1 (cyclo-
propane) and P2 (fluoroleucine) to maintain high metabolic stabil-
ity while introducing another group in P3 to allow a different
F
F
F
F
H
N
N
N
H
O
* Corresponding author. Address: Merck Frosst Canada, PO Box 1005, Pointe-
Claire-Dorval, Québec, Canada H9R 4P8. Tel.: +1 514 428 3655; fax: +1 514 428
2624.
S
O
O
E-mail address: elise_isabel@merck.com (E. Isabel).
Figure 1. Odanacatib.
0960-894X/$ - see front matter Ó 2009 Published by Elsevier Ltd.
doi:10.1016/j.bmcl.2009.12.083

888
E. Isabel et al. / Bioorg. Med. Chem. Lett. 20 (2010) 887–892
metabolism pathway without compromising its high intrinsic po-
tency and selectivity. To do so, the biaryl moiety was substituted
by a variety of functional groups: heterocycles, amides and alco-
hols. Several reports from our laboratories12–15 have described effi-
cient methods to enantiomerically synthesize large quantities of
molecules in this class. Compound 1 was an intermediate from
the synthesis of odanacatib and therefore, considerable amount
of material was available to study SAR in the P3 region.
N
N
a
NH2
Br
S
17
S
18
Br
b
Br
c
Br
O
O
N
HO
N
H2N
N
19
20
23
21
Cl
d
Cl
Cl
Cl
e
All analogs were synthesized using the chemistry described in
Scheme 1. The required intermediates were synthesized by meth-
ods shown in Scheme 2. Compound 3 was obtained by a palladium
cross-coupling between 2 and the commercially available 5-bro-
mo-2-methanesulfonylpyridine. The same conditions were used
to synthesize 4 from 6-bromoquinoline. Compound 5 was obtained
from a coupling between 2 and 2-bromo-4-methylthiazole 18
which was synthesized following a well-established literature pro-
cedure16 from the commercially available 2-amino-4-methylthia-
zole 17. The syntheses of compounds 6 and 7 were initiated with
1-(4-bromo-phenyl)-cyclopropanecarbonitrile 19 and 2-chloro-5-
hydroxymethylpyridine 22, respectively. The alcohol was first
mesylated and further displaced by NaCN to yield 23. The cyclo-
propane 24 was obtained by a double alkylation of 23. Then, com-
pounds 19 and 24 were submitted to a basic hydrolysis to generate
carboxylic acids 20 and 25. The primary amides 21 and 26 were ob-
tained after treatment with ammonia of the in situ formed mixed
anhydrides. Building blocks 21 and 26 were finally reacted with
2 to provide 6 and 7. Compound 8 was synthesized in a few steps
from 4-bromophenyl acetic acid methyl ester 27. The benzylic po-
sition was methylated with iodomethane in the presence of DMPU
and LiHMDS. The carboxylic acid 28 obtained by hydrolysis of the
methyl ester was then reacted with oxalyl chloride and catalytic
HO
N
N
N
N
N
22
24
b
Cl
O
O
c
H2N
HO
26
25
Br
Br
O
O
f, g
O
HO
27
28
h
Br
Br
Br
O
O
i, j
O
O
H2N
O
30
29
Br
k
Br
Cl
l
O
O
HO
TBSO
TBSO
O
31
32
35
33
36
Cl
Cl
l
k
N
HO
N
N
amount of DMF. The ketene formed in situ was reacted with L-lac-
O
tic acid to provide a single diastereomeric ester 29. The ester was
carefully hydrolyzed with LiOH and hydrogen peroxide to give
the carboxylic acid in high enantiomeric excess. The corresponding
primary amide 30 was obtained without racemization.17 Com-
pound 8 was obtained by reacting 30 with 2 under standard condi-
tions. Cyclopropanols 32 and 35 were synthesized from the
corresponding commercially available methyl esters 31 and 34.18
Palladium-catalyzed cross-coupling reactions were giving low
yields when the coupling was performed on the free alcohol.
Acceptable yields and cleaner material were obtained following a
simple silyl ether protection with TBS. Cross-coupling between 2
and reagents 35 or 36 followed by deprotection using HFÁpyridine
afforded compounds 9 and 10. Methyl Grignard addition to methyl
4-bromobenzoate 31 provided the necessary tertiary alcohol 37 to
yield compound 11.19 Difluoroacetophenone12 38 was reacted neat
34
Br
Br
Br
m
n
O
F
HO
O
31
37
Br
F
F
F
F
O
OH
39
o
38
Br
F
OH
40
Scheme 2. Reagents and conditions: (a) 85% H3PO4, 65% HNO3, NaNO2 (1.3 equiv),
CuSO4Á5H2O (1.3 equiv), H2O, 10%; (b) 25% NaOH, EtOH, 100 °C, 90%; (c) isobutyl
chloroformate (1.2 equiv), Et3N (1.3 equiv), CHCl3, NH3(g), 0 °C; (d) (i) MsCl
(1.1 equiv), Et3N (1.2 equiv), CH2Cl2, À78 °C; (ii) NaCN (1.1 equiv), DMF, 80%; (e)
1-bromo-2-chloro-ethane (1.1 equiv), benzyltriethylammonium chloride (0.03
equiv), 50% NaOH (3.1 equiv), 60 °C, 88%; (f) LDA (1.1 equiv), CH3I (4.0 equiv),
À78 °C, 100%; (g) 1 M LiOH (3.5 equiv), THF/MeOH (2:1), 0 °C , 96%; (h) oxalyl
chloride (1.2 equiv), DMF (0.03 equiv), toluene, N,N-dimethylethylamine (3.0 equiv),
F
F
F
F
F
F
F
F
H
N
H
N
N
N
a
N
H
N
H
ethyl L-lactate (1.2 equiv), 56%; (i) 4 N LiOH (1.1 equiv), 30% H2O2 (2.7 equiv), MeOH,
O
O
O
0 °C, 67%; (j) HOBT (1.5 equiv), HATU (1.5 equiv), NH4Cl (2.6 equiv), Hunig’s base
(3.9 equiv), 90%; (k) EtMgBr (3.2 equiv), Ti(OiPr)4 (1.0 equiv), Et2O, À78 °C; (l)
imidazole (2.0 equiv), DMAP (0.1 equiv), TBSCl (1.1 equiv), 95%; (m) MeMgBr
(3.0 equiv), THF, 0 °C, 75%; (n) S-alpine borane (1.2 equiv), 70%; (o) R-alpine borane
(1.2 equiv), 90%.
Br
B
1
2
O
b
F
F
F
F
H
N
N
N
with either S-alpine borane or R-alpine borane20 to afford the
enantiomerically pure reactants 39 or 40, respectively. These were
coupled under standard conditions to provide 12 and 13.
H
O
R
3-13
Subtle changes to the P3 portion of odanacatib had a major im-
pact on the potency, selectivity or pharmacokinetics of the new
compounds (Table 1). By comparing compound 3 to odanacatib,
it was clear that the introduction of the 3-pyridyl ring was detri-
Scheme 1. Reagents and conditions: (a) Diboron pinacol ester (1.1 equiv), KOAc
(3.1 equiv), PdCl2dppf (5 mol %), dioxane, 80 °C, 80%; (b) ArBr (1.2 equiv), 2 M
Na2CO3 (3 equiv), PdCl2dppf (3 mol %), 80 °C or ArCl (1.2 equiv), 2 M Na2CO3
(2.5 equiv), Pd(OAc)2ÁPPh3 (5 mol %), 1-propanol/DMF 4:1, 80 °C.

E. Isabel et al. / Bioorg. Med. Chem. Lett. 20 (2010) 887–892
889
Table 1
In vitro activity of odanacatib analogs with P3 substitution
F
F
F
F
H
N
N
N
H
O
R
Compound
Odanacatib
3
R
Hrab Cat K23
IC50 (nM)
Cat B/K
Cat F/K
3232
n/a
Cat L/K
14,975
1303
Cat S/K
300
‘Corrected’ Bone
Res24 IC50 (nM)
Comments
0.2
2.6
5170
725
5
8
S
S
O
O
O
O
28
10Â loss in potency
N
Lack of selectivity
against Cat S
4
5
0.4
0.3
2997
4044
n/a
6021
44
8
N
N
1985
18,753
777
14
Poor%F
Poor%F
S
O
6
0.2
1555
2379
11,120
766
12
H2N
H2N
O
O
7
8
0.5
0.2
3924
1440
3442
3592
3695
849
693
35
35
Circulating metabolite
N
Complicated
metabolic profile
21,332
H2N
HO
Lack of selectivity
against Cat S
9
0.2
0.3
0.2
1420
3643
2197
1147
2759
2410
11,005
18,022
28,916
106
210
715
13
10
11
n.a.
10
Poor%F
HO
HO
N
Short half-life
F
F
12
13
0.4
0.4
1474
1156
1600
1465
14,119
11,857
197
243
7
Good candidate
Good candidate
OH
OH
F
F
10
mental: there was a 10-fold loss in potency. However, by replacing
it with the quinoline of compound 4, most of the potency was
recovered but the selectivity against Cat S was greatly reduced.
Heterocycles like the methylthiazole 5 provided a more interesting
in vitro profile. However, the oral bioavailability was disappointing
(10% in rats and 5% in the squirrel monkeys). We then turned our
Table 2
Correlation between in vitro hepatocyte incubations and pharmacokinetics of 12 and 13 (1% methocel as oral vehicle)
Species
12
13
In vivo
In vitro
In vivo
In vitro
%F
86
132
139
T1/2 (h)
% parent remaining
%F
T1/2 (h)
% parent remaining
Rat
Rabbit
Rhesus monkey
10
8
6
94
70
88
122
70
64
7
14
11
95
93
93

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E. Isabel et al. / Bioorg. Med. Chem. Lett. 20 (2010) 887–892
attention to benzylic-substituted primary amides as an isostere of
the methylsulfone. Compound 6 bearing a cyclopropane amide had
a similar profile to 5. Here again, the compound failed because of
poor bioavailability in rats (8%) and squirrel monkeys (5%). It was
surprising that the introduction of a pyridine to give Compound
7 did not impair the potency or the selectivity. Compound 7, when
formulated as a tosylate salt, provided 50% bioavailability in rats.
However, further evaluation led to the identification of an abun-
dant circulating metabolite in rats so it was abandoned. Similarly,
compound 8 suffered from poor oral bioavailability and a compli-
cated metabolic pathway: in pre-clinical species, circulating
metabolites that were observed had not been observed in micro-
some or hepatocyte incubations. This poor in vitro–in vivo correla-
tion would have led to significant developmental challenges. Since
amide moieties demonstrated unsatisfactory ADME properties, we
began to study benzylic alcohols in P3. All compounds incorporat-
ing an alcohol moiety at the benzylic position exhibited high
potency against Cat K. Compound 9 showed poor selectivity
against Cat S. Introduction of a pyridine improved this selectivity
but compound 10 suffered from low bioavailability in rats (7%).
The dimethyl carbinol analogue 11 was highly selective against
Cat S with moderate bioavailability in rats of 24%. However 11 suf-
fered from a half-life in rats of less than 1 h. Then, a pair of enan-
tiomerically pure alcohols 12 and 13 was evaluated. These two
compounds were potent and highly selective in in vitro assays.
As well, they were potent in a functional bone resorption assay
in rabbit osteoclasts.21 Both 12 and 13 exhibited high oral bioava-
ilabilities22 and long half-lives in rats, rabbits and rhesus monkeys
(Table 2). These results were consistent with the low levels of
metabolism observed in hepatocyte incubations.
12D/12H+13H or 13D/12H+13H Ratio in Plasma over
Time when Dosing with 12D or 13D
18
16
14
12
10
8
12
13
6
4
2
0
0
5
10
15
20
Time (h)
Chart 1. D/H ratio over 24 h measured in rat (n = 2) plasma when either 12D or 13D
was administered P.O. Both 12D and 13D lose their deuterium at a similar rate.
24h Rat Plasma Concentrations when Dosing with 12D
or 13D
4.5
4.0
13H+13D
3.5
12H+12D
3.0
2.5
2.0
1.5
1.0
0.5
0.0
In order to better understand the metabolism of the secondary
alcohol, deuterated 12D and 13D were prepared as a racemate and
separated on
a semi-preparative AD normal-phase column
(Scheme 3). Rats were dosed P.O. (25 mg/kg, n = 2) with either
12D or 13D. Plasma samples were analyzed by LC–MS both on a
standard C18 column to determine the isotopic ratio and on an
OJ-R reverse-phase column to assess the diastereomeric ratio of
12 and 13. Results of this experiment are shown in Charts 1 and
2. First of all, we observed that 12D and 13D25 were losing their
deuterium at a similar rate over time (Chart 1). We also noticed
Dosing with 12D
Dosing with 13D
Chart 2. Concentrations of 12 and 13 in rat plasma 24 h post-dosing of either 12D
or 13D. As 12D loses its deuterium, it is converted to 13H. When 13D loses its
deuterium according to the same mechanism, it is converted to 13H as well,
indicating selective reduction of an intermediate ketone.
F
F
F
F
H
N
N
Br
Br
F
F
N
H
a
b
O
F
F
F
OH
D
O
38
F
41
42
OH
D
c
F
F
F
F
F
F
F
F
H
N
H
N
N
+
N
N
H
N
H
O
O
F
F
F
F
12D
13D
OH
OH
D
D
F
F
F
F
H
N
N
Br
N
H
F
b
O
F
F
O
F
38
14
O
Scheme 3. Reagents and conditions: (a) NaBD4 (1.0 equiv), EtOD, 100%; (b) 2 (1.0 equiv), ArBr (1.2 equiv), 2 M Na2CO3 (3.0 equiv), PdCl2dppf (3 mol %), 80 °C, 87%; (c)
chiralcel AD normal phase semi-preparative HPLC1:1 hexanes/EtOH isocratic, tR = 5.09 and 6.53 min.

E. Isabel et al. / Bioorg. Med. Chem. Lett. 20 (2010) 887–892
891
Table 3
incubations. The in vivo dog data was generated by dosing with
pure 13D and showed partial conversion to 12H that was consis-
tent with the in vitro results (P.O., t = 24 h).
In vitro and in vivo 13/12 ratio following ketone reduction
Species
In vitro
13/12 ratio
In vivo
13/12 ratio
Since 12 epimerized to 13 in vivo, 12 was abandoned and fur-
ther profiling was done on 13. Two metabolites were identified
from incubations: hydroxyleucine 15 and the glucuronide conju-
gate 16. Authentic standards were synthesized in order to confirm
the structure (Scheme 4). Compound 15 was obtained after a mul-
ti-step synthesis. Compound 1 was treated with BF3ÁEt2O to afford
the alkene 43. It was dihydroxylated with catalytic osmium tetrox-
ide to give the diol 44. The primary alcohol was acylated, whereas
the tertiary alcohol was treated with DAST to yield 45. The aryl
bromide 40 was turned into the boronic ester 46 following the pro-
cedure described above. A Suzuki cross-coupling was conducted
between 45 and 46 followed by a basic hydrolysis to afford the
metabolite 15.
Compound 16 was obtained from dog liver microsome incuba-
tion26 and isolated from the incubate according to a published
method.27 Incubations with human liver microsomes revealed that
the oxidative metabolism was mainly due to CYP3A4. However,
incubations with 12 different rUGTs (recombinant uridine 5’-phos-
phate-glucuronosyl transferase) indicated that multiple UGTs were
involved in the glucuronidation process.
Rat
Dog
Rabbit
Human
24:1
5:1
48:1
>200:1
>10:1
5:1 to 7:1
n.a.
n.a.
that 12D was converted into 13H in vivo (Chart 2). When 13D was
administered to rats, we observed that 13D was converted to 13H,
but very little 12H was formed. These results suggested that 12D or
13D were both oxidized in vivo to the corresponding ketone, fol-
lowed by stereoselective reduction to 13H. To verify this hypothe-
sis, the ketone 14 was synthesized (Scheme 3) and dosed P.O.
(25 mg/kg) and I.V. (5 mg/kg). Samples were analyzed as described
above. Ketone 14 itself was not detected in any samples (earliest
time point = 5 min) but both 12 and 13 were formed with a
13:12 ratio greater than 10:1 at all time points. Ketone 14 was
incubated with hepatocytes from four species: extensive and selec-
tive reduction to 13 was observed. This reduction was most selec-
tive in human and least selective in dog (Table 3). Comparison to
PK data from rat and dog showed a good correlation with in vitro
In conclusion, by replacing the methyl sulfone of odanacatib
with a difluoromethylcarbinol, MK-0674 was identified. It fulfilled
our requirements in terms of potency, selectivity, pharmacokinet-
ics and metabolism to provide a valuable back-up. It differentiates
from odanacatib since it is partially excreted via a glucuronide
conjugate.
F
F
F
F
F
F
F
H
N
H
N
a
N
N
N
H
N
H
O
O
Br
Br
References and notes
1
43
b
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O
OH
OH
F
F
F
F
F
F
F
O
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H
N
H
N
N
N
c, d
N
N
H
H
O
O
Br
Br
45
44
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OH
F
e, f
F
F
F
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H
N
N
N
H
O
B
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O
F
O
F
15
46 =
F
F
g
OH
OH
F
F
F
F
H
N
N
N
H
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O
F
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F
F
13
F
F
F
H
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N
OH
N
N
H
O
OH
O
O
O
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Asymmetry 1994, 5, 1075.
21. Assay conditions described in: Falgueyret, J.-P.; Oballa, R. M.; Okamoto, O.;
Wesolowski, G.; Aubin, Y.; Rydzewki, R. M.; Prasit, P.; Riendeau, D.; Rodan, S.
V.; Percival, M. D. J. Med. Chem. 2001, 44, 94–104.
16
F
HO
OH
F
OH
Scheme 4. Reagents and conditions: (a) BF3ÁEt2O (5.0 equiv), CH2Cl2, 0 °C, 53%; (b)
OsO4 (0.01 equiv), NMO (1.1 equiv), acetone/H2O 1.4:1, 90%; (c) acetic anhydride
(1.1 equiv), DMAP 0.1 equiv), Et3N (2.0 equiv), CH2Cl2, 0 °C, 65%; (d) DAST
(1.2 equiv), CH2Cl2, À78 °C, 25%; (e) 46 (2.0 equiv), 2 M Na2CO3 (4.0 equiv),
PdCl2dppf (5 mol %), DMF, 80 °C, 44%; (f) K2CO3 (1.5 equiv), MeOH, 65%; (g) dog
liver microsomes incubation, 37 °C, 3.5 h, SPE purification.

892
E. Isabel et al. / Bioorg. Med. Chem. Lett. 20 (2010) 887–892
22. The fact that oral bioavailabilities exceeding 100% were observed most likely
means that the excretion pathway was saturated. This should not be observed
if lower doses were administered.
26. MK-0674 was incubated at 100
(60 mM, pH 7.4) containing 1 mg/mL of dog liver microsomes, 12.5 mM MgCl2,
21 mM -saccharic acid 1–4 lactone and 13 mM of UDPGA for 3.5 h at 37 °C
lM final concentration in phosphate buffer
D
23. Human Cat B, F, K, L and S enzymes. Screening conditions: Desmarais, S.;
Massé, F.; Percival, M. D. Biol. Chem. 2009, 390, 941. and references cited
therein.
24. Bone res IC50 x Hrab Cat K IC50/rabbit Cat K IC50. Corrects for differences
between rabbit and human potency: see Ref. 10.
under 95/5 O2:CO2 atmosphere. Incubation broth (ꢀ80 mL) was centrifuged
5 min at 4300 rcf, the supernatant collected, and then the pellet was washed
with 1 mL of acetonitrile. Aqueous and organic fractions were combined and an
Oasis HLB 6 cc cartridge from Waters was used to isolate the glucuronide
conjugate of MK-0674.
25. Compounds 12D, 13D and 14 were stable in plasma over a 24–h period: no
deuterium loss or reduction observed, respectively.
27. Gagné, S.; Laterreur, J.; Mahrouche, L.; Sørensen, D.; Gauthier, J.-Y.; Truong, V.
L.; Chauret, N.; Lévesque, J.-F. J. Chromatogr., B 2008, 863, 242.