2-(2-Aminothiazol-4-yl)pyrrolidine-based tartrate diamides as potent, selective and orally bioavailable TACE inhibitors

Article abstract of DOI:10.1016/j.bmcl.2011.01.002

TNF-α converting enzyme (TACE) inhibitors are promising agents to treat inflammatory disorders and cancer. We have investigated novel tartrate diamide TACE inhibitors where the tartrate core binds to zinc in a unique tridentate fashion. Incorporating (R)-

Full text of DOI:10.1016/j.bmcl.2011.01.002

Bioorganic & Medicinal Chemistry Letters 21 (2011) 3172–3176
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
2-(2-Aminothiazol-4-yl)pyrrolidine-based tartrate diamides as potent, selective
and orally bioavailable TACE inhibitors
Chaoyang Dai a, , Dansu Li a, Janeta Popovici-Muller a, Lianyun Zhao a, Vinay M. Girijavallabhan b,
⇑
Kristin E. Rosner a, Brian J. Lavey b, Razia Rizvi b, Bandarpalle B. Shankar b, Michael K. C. Wong b,
Zhuyan Guo b, Peter Orth b, Corey O. Strickland b, Jing Sun c, Xiaoda Niu c, Shiying Chen d,
Joseph A. Kozlowski b, Daniel J. Lundell c, John J. Piwinski a, Neng-Yang Shih a, M. Arshad Siddiqui a
a Department of Chemistry, Merck Research Laboratories, 320 Bent Street, Cambridge, MA 02141, USA
b Department of Chemistry, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, NJ 07033-0539, USA
c Department of Inflammation, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, NJ 07033-0539, USA
d Department of Exploratory Drug Metabolism and Pharmacokinetics, Merck Research Laboratories, 2015 Galloping Hill Road, Kenilworth, NJ 07033-0539, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
TNF-
a converting enzyme (TACE) inhibitors are promising agents to treat inflammatory disorders and
Received 18 October 2010
Revised 23 December 2010
Accepted 3 January 2011
Available online 6 January 2011
cancer. We have investigated novel tartrate diamide TACE inhibitors where the tartrate core binds to zinc
in a unique tridentate fashion. Incorporating (R)-2-(2-N-alkylaminothiazol-4-yl)pyrrolidines into the left
hand side amide of the tartrate scaffold led to the discovery of potent and selective TACE inhibitors, some
of which exhibited good rat oral bioavailability.
Ó 2011 Published by Elsevier Ltd.
Keywords:
TNF-a converting enzyme
TACE tartrate
Zinc binding group (R)-2-(2-N-
alkylaminothiazol-4-yl)pyrrolidines
Tumor necrosis factor-
a
(TNF-
a
) is a major mediator of inflam-
acids and hydroxylamine, which is toxic. Therefore, there is a con-
siderable interest in identifying non-hydroxamate TACE inhibitors.
Hydantoins,7 barbiturates,8 and thiols9 have been reported as zinc
binding groups.
Recently, we have discovered a novel series of tartrate diamide
TACE inhibitors, which display a unique tridendate zinc binding
mode with the tartrate scaffold.10,11 The combination of a tertiary
amide at the left hand side (LHS) and a secondary amide at the
right hand side (RHS) of the tartrate core appears to be essential
for their TACE activities. Through extensive structure–activity
matory responses in various autoimmune disorders such as rheu-
matoid arthritis (RA), Crohn’s disease, and psoriasis.1 The success
of anti-TNF-
has demonstrated that inhibition of TNF-
a
biologics such as EnbrelÒ, RemicadeÒ, and HumiraÒ
a
could result in effective
control of these inflammatory responses and autoimmune dis-
eases. The search for small molecule, orally administered inhibitors
of TNF-
TNF-
ing enzyme (TACE) mediated cleavage of its membrane bound
26 kDa precursor form, pro-TNF- to the 17 kDa soluble compo-
nent.3 Inhibition of TACE would decrease the level of TNF-
re-
a
production has been an area of intensive research.2
a
is released into the bloodstream via the TNF-a convert-
a
a
leased into circulation and could hold promise for treating
related inflammatory diseases.4 Recently, studies have also shown
that TACE inhibitors may be useful for inhibition of pathogenic
EGFR signaling in cancer.5
The majority of known TACE inhibitors rely on hydroxamic
acids as the zinc binding groups (ZBG).6 However, hydroxamic
acids are often poorly absorbed because of high renal clearance.
They also carry potential metabolic liabilities such as O-glucuron-
idation and hydrolysis in vivo to give the corresponding carboxylic
Cl
N
N
N
N
O
OH
O
OH
Z
H
N
H
N
N
N
OH O
OH O
n
1
LHS
n = 1–3;
RHS
TACE Ki = 0.8 nM
Cell IC50 = 886 nM
AUC = 2,949 nM.h
Z = aryl, heteroaryl
C
6h = 96 nM
⇑
Corresponding author.
E-mail addresses: chaoyang.dai@merck.com, chaoyangdai@hotmail.com (C. Dai).
Figure 1. Replacements of the 3-chlorophenyl group and pyrrolidine ring modifi-
cations on inhibitor 1.
0960-894X/$ - see front matter Ó 2011 Published by Elsevier Ltd.
doi:10.1016/j.bmcl.2011.01.002

C. Dai et al. / Bioorg. Med. Chem. Lett. 21 (2011) 3172–3176
3173
Table 1
Table 2
TACE Ki and rapid rat PK of LHS 2-phenylpyrrolidine analogs
TACE Ki and rapid rat PK of LHS pyrrolidine-based tartrate diamides
N
N
N
N
O
OH
O
OH
R
H
N
H
N
R
N
OH O
OH O
a
Compound
R
TACE Ki (nM)
AUCa (nM h)
2949
Compound
R
TACE Ki (nM)
AUC (nM h)
C6h (nM)
96
Cl
Cl
1
0.8
2949
578
1
0.8
N
N
NH
N
12
5
0
2
3
0.4
0.5
1465
461
N
N
13
14
15
4
2921
nd
86
nd
nd
Cl
S
N
S
10
10
N
nd
N
Cl
NH2
N
( )-4
( )-5
6.7
7.6
4765
3908
S
HO
16
6
2276
247
N
Cl
a
Compound concentration in plasma at the 6th hour after dosing.
N
HO
Cl
then turned our attention to optimizing the LHS amine building
blocks in the hope of improving their oral PK profiles while main-
taining or further improving its biochemical potencies. Our results
are reported herein.
Cl
( )-6
87
nd
N
Initial efforts focused on the structural variation of the 2-(3-
chlorophenyl)pyrrolidyl unit of 1 as a whole. Introduction of a qua-
ternary methyl group at C-2 of the pyrrolidine ring (compound 2,
Table 1) improves potency slightly, but also leads to a reduced
AUC relative to compound 1. Likewise, the 3-pyrroline analog (3)
shows similar activity, but the AUC is significantly eroded. Hydrox-
ylation of the pyrrolidine ring at C-3 or C-4 (4 and 5, respectively)
results in a slight improvement in AUC, but is accompanied by a
10-fold drop in activity. Ring expansion of the pyrrolidine to a
piperidine ring while preserving the 3-chlorophenyl substituent
(6) results in a 100-fold drop in TACE activity relative to 1. The
4-chlorophenyl and unsubstituted phenyl analogs (7 and 8, respec-
tively) do not suffer as great a reduction in activity, but are still less
potent than 1. The benzannulated analogs 9 and 10 both show a
10-fold improvement in activity over their monocyclic congeners
7 and 8, respectively. The activity of 10 is comparable to that of
compound 1 but disappointingly, the pharmacokinetic profile of
10 is not improved over that of compound 1. Further ring expan-
sion to yield the benzazepine 11 results in a significant loss of
activity over 1 and 10. The preparation of compounds 2–11 fol-
lowed previously described procedures.13
At this juncture, we decided to revise our optimization strategy
by preserving the pyrrolidine ring of 1 and explore the effect of
replacing the 3-chlorophenyl group of 1 with heteroaromatic
groups. Among the heterocycles that were investigated, pyrazoles
(12 and 13) and thiazoles (14–16) have emerged as promising aryl
group replacements to maintain TACE potencies in the low nano-
molar range (Table 2). Compound 16 is interesting in that its
AUC (10 mpk, po) is only moderate, but the plasma concentration
at the 6th hour still remains relatively high (C6h = 247 nM) com-
pared to the value for compound 1 (C6h = 96 nM).12 Moreover,
the 2-aminothiazole moiety provides a good handle for further
structural modifications to fine-tune the hydrophobicity and other
properties of this series of compounds, hence to optimize their
overall profiles.
( )-7
( )-8
22
14
nd
nd
N
N
Cl
N
9
2.6
nd
MeO
MeO
( )-10
0.9
38
1239
nd
MeO
MeO
N
MeO
MeO
11
N
a
Sprague–Dawley rats were dosed orally with 10 mpk compounds in 0.4% MC;
data were collected within 6 h after dosing and not determined.
relationship (SAR) investigations, we have identified several series
of preferred RHS amine building blocks.11 In particular, (R)-1-(4-
(1H-pyrazol-1-yl)phenyl)ethanamine has emerged as an preferred
RHS amine when combined with various 2-phenylpyrrolidine LHS
pieces such as compound 1 which showed very good binding affin-
ity and modest cellular activity and oral PK in the rat12 (Fig. 1). We

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C. Dai et al. / Bioorg. Med. Chem. Lett. 21 (2011) 3172–3176
R2
R2
R1
N
R1
N
Cl
S
S
O
OMe
Boc
O
c
a
b
N
N
1)
2)
Boc
N
N
Boc
N
NH
17
18
19
20
O
O
O
O
O
O
MeO
NH
N
OH
d
MeO
N
O
O
22
21
R2
R1
O
N
O
O
O
N
N
S
NH
N
N
HO
e
N
f
OH
H
O
N
g
N
OH O
16, 24-33
23
Scheme 1. Reagents and conditions: (a) LDA, CH2ICl, THF, À78 °C, 74%; (b) thioureas, DMF, rt; (c) HCl/dioxane, MeOH; (d) HATU, DIEA, DMF, 78%; (e) LiOH, THF, 93%; (f) 20,
HATU, DIEA, DMF, rt; (g) HCl/dioxane, water.
A series of N-substituted 2-aminothiazole-based tartrate inhib-
itors was readily synthesized (Scheme 1). Various 2-(2-N-alkylthia-
compound 32 resulted in a loss of activity (Ki = 65 vs 6 nM in com-
pound 16), and the cyclic piperidine compound 33 is also less
active.
zol-4-yl)pyrrolidines (20) were obtained from
methyl ester (17) through initial conversion to the corresponding
-chloroketone (18)14 with chloroiodomethane and LDA. Subse-
D-Boc-proline
The pharmacokinetic (PK) properties of these TACE inhibitors
were studied through rapid rat PK analysis.12 As shown in Table 3,
mono N-alkylation on 2-aminothiazole dramatically improves its
oral PK. For instance, simply adding an N-methyl group (24 vs
16) brings rapid rat AUC to 30,706 from 2278 nM h. Ethyl (25)
and n-propyl (26) substitutions have similar effects. The isopropyl
(27) and cyclopropyl (28) compounds show remarkably high AUCs
and drug concentrations at the 6th hour remain high. On the other
hand, N-aromatic substitution is not favored in terms of PK as seen
in compounds 30 and 31. Compared to the initial lead compound 1,
the thiazole analogs 24–29 have much improved oral exposure.
However, they generally have weaker cellular activities (cell
IC50 = 3016 nM for compound 28 vs IC50 = 886 nM for 1).
In order to improve the cellular activity of the thiazole series, it
was decided to further explore the SAR around the 2-(2-amin-
othiazol-4-yl)pyrrolidine moiety. Therefore, the effect of a 5-sub-
stituent was investigated (Table 4). Starting from Boc-protected
a
quent condensation/cyclization with various thioureas in DMF
afforded the corresponding 2-aminothiazoles, and a final acid-
mediated deprotection gave the desired pyrrolidines (20). Sepa-
rately, the acetonide-protected tartrate half acid (21) was firstly
coupled with (R)-1-(4-(1H-pyrazol-1-yl)phenyl)ethanamine to
form the mono-amide (22), which was subsequently saponified
to acid (23). Amide coupling of 23 with the various LHS amines
(20) afforded the corresponding tartrate diamide compounds (16,
24–33) after acetonide deprotection.
As shown in Table 3, 2-(2-aminothiazol-4-yl)pyrroline amide 16
is a potent TACE inhibitor. The high binding affinities are largely
maintained in compounds with mono-alkyl N-substitutions (24–
29). Mono N-substitution with 2-fluorophenyl (30) or 3-pyridyl
(31) groups also results in potent compounds. On the other hand,
N,N-disubstitution is not well tolerated. For example, dimethyl
Table 3
TACE inhibition potencies and rapid rat PK of 2-aminothiazole-based tartrate diamides
R2
R1
N
N
N
S
N
O
OH
H
N
N
OH O
Compound
R1
R2
TACE Ki (nM)
Cell IC50 (nM)
AUC (nM h)
C6h (nM)
16
24
25
26
27
28
29
30
31
32
H
H
H
H
H
H
H
H
H
Me
H
Me
Et
n-Pr
i-Pr
c-Pr
c-Bu
2-Fluorophenyl
3-Pyridyl
Me
6
7
7
4
17
5
4
1
2
65
4286
5799
5107
4481
6677
3016
2153
1143
5552
nd
2276
247
30,706
51,978
48,348
114,792
134,267
31,390
130
2535
5967
5403
12,305
17,646
3542
0
0
nd
0
nd
33
39
nd
1959
0
N

C. Dai et al. / Bioorg. Med. Chem. Lett. 21 (2011) 3172–3176
3175
Table 4
Table 5
TACE inhibition potencies and rat PK of 5-substituted-2-aminothiazole-based tartrate
diamides
TACE inhibition potencies and rapid rat PK of 2-aminothiazole-based tartrate
diamides
HN
N
N
N
S
O
OH
H
N
N
N
O
OH
R
H
N
R
N
OH O
OH O
Compound
R
TACE Ki (nM) Cell IC50 (nM) AUC (nM h)
Compound
R
TACE Ki (nM) Cell IC50 (nM) AUC (nM h)
NH
24
34
35
36
37
38
H
Me
c-Pr
4-Fluorophenyl
CN
SO2Me
7
2
2
1
2
0.4
5799
2217
2574
3245
3036
720
30,706
5438
11,423
nd
154
0
S
N
( )-39
9
6
13,811
4040
2439
5618
N
NH
S
N
40
41
N
NBoc
S
N
c
NH
Boc
S
N
N
4
2769
1417
35a
N
NBoc
NBoc
NH
N
F
S
S
S
NBoc
N
F
N
N
Br
S
a
b
HN
N
d
e
Boc
Boc
Boc
N
S
S
S
N
N
Ar
Boc
42
43
0.6
1338
707
1809
925
N
19c
19a
19b
N
36a
H
N
NBoc
N
g
f
S
N
1
NC
Boc
N
N
NBoc
NBoc
NBoc
N
S
h
S
S
N
HN
N
N
37a
S
S
O
Boc
O
Boc
N
Boc
N
N
44
531
nd
nd
N
34a
38a
38b
HN
N
S
Scheme 2. Reagents and conditions: (a) Boc2O, DMAP, THF, rt, 95%; (b) NBS, CHCl3,
50 °C, 95%; (c) cyclopropylboronic acid, PdCl2(dppf), K3PO4, 80 °C, 85%; (d)
4-fluorophenylboronic acid PdCl2(dppf), K3PO4, 80 °C, 88%; (e) Zn(CN)2, Pd2(dba)3,
dppf, 85 °C, 85%; (f) n-BuLi, MeI, À78 °C, 84%; (g) n-BuLi, MeSSMe, À78 °C, 64%; (h)
m-CPBA, DCM, 95%.
45
46
1
416
384
1133
237
N
HN
N
S
16
N
F
F
HN
S
N
N
47
48
2
288
257
nd
NH
S
N
0.5
0
N
N
N
H2N
Figure 2. Crystal structure of 38 (cyan) bound to TACE (yellow). Only residues that
involved in zinc binding and hydrogen bonding with the inhibitor (dashed yellow
lines) are shown.
2-(2-N-methylaminothiazol-4-yl)pyrrolidine (19a), the protected
amine building blocks 35a–37a were obtained through Boc-protec-
tion, NBS bromination, and palladium catalyzed cross-couplings.
Alternatively, deprotonation of 19b with n-BuLi followed by MeI

3176
C. Dai et al. / Bioorg. Med. Chem. Lett. 21 (2011) 3172–3176
Table 6
ring (i.e., 48) give rise to compounds with significantly improved
cellular activities.
Selectivity profile of inhibitor 1, 28, and 45
Ki (nM)
1
28
45
References and notes
TACE
0.8
5
1
MMP1
MMP2
MMP3
MMP7
MMP9
MMP13
MMP14
ADAM10
>100 000
43,660
—
3769
88,420
3300
1148
166
>40,000
20,500
>40,000
18,100
>40,000
>40,000
>40,000
1350
>40,000
>40,000
>40,000
13,500
10,100
14,100
16,400
120
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or MeSSMe quenching afforded compounds 34a and 38a, respec-
tively. Further oxidation of 38a with m-CPBA gave methyl sulfone
38b (Scheme 2). These building blocks were then readily deprotec-
ted and coupled with tartrate half amide/acid 23 to provide 5-
substituted-2-N-methylaminothiazole based tartrate diamides
34–38 (Table 4) by following the same procedure as in Scheme 1.
As shown in Table 4, introducing methyl (34), cyclopropyl (35),
aryl (36), or cyano (37) groups at the 5-position of the thiazole ring
slightly increases the potencies of the inhibitors versus 24. The
methylsulfonyl group at 5-position in compound 38 is the most
effective one, showing approximately 10-fold improvement in
both the biochemical and cellular activities (0.4 and 720 nM,
respectively) versus compound 24 (7 and 5799 nM, respectively).
X-ray crystallography15 shows that the methylsulfonyl group
makes hydrogen bonding interactions with a water molecule
which in turn, makes hydrogen bonding interactions with Ser355
and Pro356 of the protein (Fig. 2). However, incorporation of the
5-methanesulfonyl group results in loss of oral exposure.
The effect of pyrrolidine ring modifications was also investi-
gated. As shown in Table 5, simply adding a methyl group at
2- or 4-position (39 and 40) or 4,4-difluoro substitutions (41) on
pyrrolidine does not have much effect on the TACE activities of
the resulting tartrate diamide compound. However, when the
pyrrolidine ring is flattened with a double bond (42), its potency
is significantly improved (Ki = 0.6 nM). Therefore, cyclopropyl and
phenyl fusions to the pyrrolidine ring were subsequently explored.
It is interesting that the cis fusion compound (43) is highly potent
(Ki = 1 nM) but the corresponding trans compound (44) loses
activity (Ki = 531 nM). As anticipated, when the pyrrolidine ring
is fused with phenyl (45), the corresponding compound gives rise
to much improved enzymatic and cellular activities (Ki = 1 nM, cell
IC50 = 416 nM) comparing to compound 24. Fluoro substitutions on
the phenyl ring are well tolerated (46, 47). Finally, fusion with a
2-aminopyrimidine ring results in compound 48 with good cellular
activity (cell IC50 = 257 nM). However, these compounds all exhibit
low oral exposures in rat.
12. Description of the rat PK studies: Following an overnight fast, two Male
Sprague–Dawley rats (Charles River, Co.) were dosed orally at a dose of 10 mg/
kg. Blood was collected into heparin-containing tubes serially from each
animal at 0.5, 1, 2, 3, 4 and 6 h post-dosing and centrifuged to generate plasma.
Samples at each time point collected from two rats were pooled for LC/MS/MS
analysis. For further experimental details of the rapid rat assay, see
Korfmacher, W. A.; Cox, K. A.; Ng, K. J.; Veals, J.; Hsieh, Y.; Wainhaus, S.;
Broske, L.; Prelusky, D.; Nomeir, A.; White, R. E. Rapid Commun. Mass Spectrom.
2001, 15, 335.
TACE inhibitors of this class are very selective against related
MMPs, as exemplified by compound 28 and 45 (Table 6). Their
selectivity profiles are similar to that of compound 1.
In summary, we have further optimized the tartrate based TACE
inhibitors by incorporating 2-heteroaryl substituted pyrrolidines
as the left hand side amine groups and by pyrrolidine ring modifi-
cations. In particular, 2-(N-alkylamino)thiazole containing com-
pounds afford potent and selective TACE inhibitors with
improved oral bioavailability (i.e., compounds 24–29 show more
than 10-fold AUC improvement). On the other hand, 5-methylsulf-
one substitution on thiazole (i.e., 38) and flattening the pyrrolidine
13. (a) Guo, Z.; Orth, P.; Zhu, Z.; Mazzola, R. D.; Chan, T.-Y.; Vaccaro, H. A.;
McKittrick, B.; Kozlowski, J. A.; Lavey, B. J.; Zhou, G.; Paliwal, S.; Wong, S.-C.;
Shih, N.-Y; Ting, P. C.; Rosner, K. E.; Shipps, G. W., Jr.; Siddiqui, M. A.; Belanger,
D. B.; Dai, C.; Li, D.; Girijavallabhan, V. M.; Popovici-Müller, J.; Yu, W.; Zhao, L.
U.S. Patent 7652020 B2.; (b) 7,8-Dimethoxy-1-phenyl-3-benzazepine was
prepared via the method of Neumeyer et al., J. Med. Chem. 1991, 34, 3366 and
references cited therein.
14. Chen, P.; Cheng, P. T. W.; Spegel, S. H.; Zahler, R.; Wang, X.; Thottathil, J.;
Barrish, J. C.; Polniaszek, R. P. Tetrahedron Lett. 1997, 38, 3175.
15. RCSB protein data bank (PDB) deposition number 3O64.