5′-Phenyl-3′H-spiro[indoline-3,2′-[1,3,4]thiadiazol]-2-one inhibitors of ADAMTS-5 (Aggrecanase-2)

Article abstract of DOI:10.1016/j.bmcl.2007.07.048

5′-Phenyl-3′H-spiro[indoline-3,2′-[1,3,4]thiadiazol]-2-one inhibitors of ADAMTS-5 (Aggrecanase-2) have been prepared via commercially available starting materials. Selected compounds 23, 33-35 show sub-micromolar ADAMTS-5 potency and strong SAR trends with selectivity over the related metalloproteases ADAMTS-4 (Aggrecanase-1), MMP12, and MMP13. This series of compounds represents progress toward a selective ADAMTS-5 inhibitor as a disease modifying osteoarthritis drug.

Full text of DOI:10.1016/j.bmcl.2007.07.048

Bioorganic & Medicinal Chemistry Letters 17 (2007) 5630–5633
50-Phenyl-30H-spiro[indoline-3,20-[1,3,4]thiadiazol]-2-one
inhibitors of ADAMTS-5 (Aggrecanase-2)
Matthew G. Bursavich,a,* Adam M. Gilbert,a Sabrina Lombardi,a Katy E. Georgiadis,b
Erica Reifenberg,b Carl R. Flanneryb and Elisabeth A. Morrisb
aExploratory Medicinal Chemistry, Chemical and Screening Sciences, Wyeth Research, 401 North Middletown Road, Pearl River,
NY 10965, USA
bWomen’s Health and Musculoskeletal Biology, Wyeth Research, 200 Cambridge Park Drive, Cambridge, MA 02140, USA
Received 22 June 2007; revised 23 July 2007; accepted 23 July 2007
Available online 19 August 2007
Abstract—50-Phenyl-30H-spiro[indoline-3,20-[1,3,4]thiadiazol]-2-one inhibitors of ADAMTS-5 (Aggrecanase-2) have been prepared
via commercially available starting materials. Selected compounds 23, 33–35 show sub-micromolar ADAMTS-5 potency and strong
SAR trends with selectivity over the related metalloproteases ADAMTS-4 (Aggrecanase-1), MMP12, and MMP13. This series of
compounds represents progress toward a selective ADAMTS-5 inhibitor as a disease modifying osteoarthritis drug.
Ó 2007 Elsevier Ltd. All rights reserved.
Osteoarthritis (OA) is a debilitating disease caused by
degradation of aggrecan and collagen in the articular
cartilage matrix leading to progressive and chronic joint
pain and inflammation. Aggrecan is a multidomain pro-
teoglycan that provides the elasticity and compressive
resistance to the articular cartilage which is lost in the
initial phases of OA. This loss of aggrecan fragments
via proteolysis is attributable to aggrecanase activity.1
If this degradation is not halted or reversed, the cartilage
will be subject to further breakdown by additional
metalloproteases resulting in irreversible damage to the
joint. While current OA treatments provide only symp-
tomatic relief (NSAIDs, intra-articular injections of
hyaluronic acid conjugates, and surgical joint replace-
ment), there is no therapy available to halt and/or re-
verse the progression of this debilitating disease.2
demonstrated significantly reduced OA severity for
ADAMTS-5 knockout mice in a surgically induced
instability model.3 ADAMTS-5 has also been shown
to be the major ADAMTS in a mouse model of inflam-
matory arthritis.4 Thus, the inhibition of ADAMTS-5
may therefore protect cartilage from damage and pro-
vide the first potential therapy to halt and/or reverse
the progression of OA.
Recently we reported a series of rhodanine-based inhib-
itors of ADAMTS-5.5,6 In this letter, we report our ini-
tial studies on a series of 50-phenyl-30H-spiro[indoline-
3,20-[1,3,4]thiadiazol]-2-one ADAMTS-5 inhibitors 1
found via high throughput screening7 and show their
potential utility as selective ADAMTS-5 therapeutics
to treat OA. While other general aggrecanase inhibitors
have been disclosed,8–11 to our knowledge the com-
pounds presented herein represent the first disclosure
of submicromolar, selective ADAMTS-5 inhibitors.
Aggrecanases are members of the ADAMTS (A Disinte-
grin And Metalloprotease with Thrombospondin Mo-
tifs) family of zinc metalloproteases. Both ADAMTS-4
(Aggrecanase-1) and ADAMTS-5 (Aggrecanase-2) have
been shown to cleave aggrecan at the physiologically rel-
evant Glu373-Ala374 peptide bond.1 Recent work has
R2
R3
NH
N
Keywords: ADAMTS-5; Aggrecanase-2; Metalloprotease inhibitors;
Osteoarthritis; Thiadiazoles.
N
S
R4
O
R1
*
Corresponding author. Tel.: +1 845 602 8143; fax: +1 845 602
5561; e-mail: Bursavm@wyeth.com
1
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.07.048

M. G. Bursavich et al. / Bioorg. Med. Chem. Lett. 17 (2007) 5630–5633
5631
Table 1. In vitro ADAMTS-5 inhibition data for N–H 50-phenyl-30H-
Spiro[indoline-3,20-[1,3,4]thiadiazol]-2-one (thiadiazole)
ADAMTS-5 inhibitors 1 are prepared according to
Scheme 1.12 Thus 2-(arylcarbonothioylthio)acetic acids
3 are synthesized by quenching the corresponding Grig-
nard reagent of 2 with carbon disulfide and subsequent
alkylation with chloroacetic acid.13 Arylthiohydrazides
4 are prepared by treatment of 3 with hydrazine under
basic conditions.14 The non-commercially available isa-
tins 6 and 7 are prepared using the Sandmeyer isatin
synthesis (treating anilines 5 with 2,2,2-trichloroacetal-
dehyde monohydrate, hydroxylamine hydrochloride,
and concentrated sulfuric acid to form 6).15 Alkylation
of 6 is accomplished using NaH and alkyl/benzyl bro-
mides in THF.16 Conversely, arylation of 6 can be carried
out using Cu and aryl boronic acids.17 The synthesis of
thiadiazoles 1 is completed by condensing arylthiohyd-
razides 4 with isatins 6 or 7 in EtOH at 40 °C.
spiro[indoline-3,20-[1,3,4]thiadiazol]-2-ones
R2
R3
NH
N
N
S
H
O
R1
R1
Compound
R2
R3
ADAMTS-5
a
IC50 (lM)
8
9
H
H
Br
H
24.2
13.6
11.1
9.6
H
H
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
H
Me
Et
H
H
H
H
Me
OCF3
H
H
7.2
3.7
H
Me
Me
Me
Et
H
3.0
1.9
ADAMTS-5 inhibition data for N–H thiadiazoles (8–
25) is shown in Table 1. Compounds 8–12 demonstrate
that isatin substitution at the R2 position plays a key
role in ADAMTS-5 activity. When R1 = R2 = R3 = H
(8), weak ADAMTS-5 activity is seen. The correspond-
ing R2 = Br analog 9 shows a nearly twofold increase
in ADAMTS-5 potency (IC50 = 11.1 lM). Other R2
substitutions also demonstrate enhanced ADAMTS-5
activity (R2 = Me 10: IC50 = 11.1 lM; R2 = Et 11:
IC50 = 9.6 lM). Further improvement is demonstrated
with a OCF3 group in 12 (IC50 = 7.2 lM).
H
Me
H
n-Pr
H
1.4
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
H
13.4
2.5
Me
Et
H
H
1.4
2.0
Cl
Br
H
H
2.4
4.4
NO2
H
H
n-Pr
Me
Cl
0.78
1.4
H
H
1.2
a Values are means of two experiments, standard deviations are 10%.
A series of R1 substituted thiadiazoles 13–25 continue to
show improved ADAMTS-5 potency. The R1 = Me
R2 = R3 = H analog 13 leads to a sixfold potency increase
over 8 (IC50 = 3.7 lM vs 24.2 lM). Analogs where
IC50 = 2.5 lM; R2 = Et 19: IC50 = 1.4 lM) produces ana-
logs with increased ADAMTS-5 potency. Analogs where
R2 = Cl (20: IC50 = 2.0 lM), R2 = Br (21: IC50
=
R2 = Me (14: IC50 = 3.0 lM) or R2 = Et (15: IC50
=
2.4 lM), and R2 = NO2 (22: IC50 = 4.4 lM) also show
low micromolar activity. Substitution of the R3 position
affords analog 23 (R1 = OMe, R2 = H, R3 = n-Pr) that
demonstrates sub-micromolar ADAMTS-5 inhibition
(IC50 = 0.78 lM). Further investigation of the R3
position affords analogs 24 (R3 = Me: IC50 = 1.4 lM)
and 25 (R3 = Cl: IC50 = 1.2 lM) with minimal loss in
ADAMTS-5 activity.
1.9 lM) show similaror slightlygreater potency. Incorpo-
ration of a R3 = n-Pr substituent shows similar low
micromolar ADAMTS-5 potency (16: IC50 = 1.4 lM).
Thiadiazole analogs where R1 = OMe 17–26 were also
pursued to further expand the scope of the R1 substituent.
The R1 = OMe, R2 = R3 = H analog 17 shows a twofold
potency increase over 8 (IC50 = 13.4 lM vs 24.2 lM).
As shown above, R2 substitution (R2 = Me 18:
S
S
Br
OH
R2
a
b
NH2
S
N
H
R3
R1
O
R1
R1
2
NH
N
3
4
f
N
S
R4
O
R1
O
1
R2
R2
R3
c, d
O
R3
N
NH2
R4
6 R4 = H
7 R4 = Bn, Ar
5
e
Scheme 1. Reagents and conditions: (a) i.Mg, Et2O; ii.CS2, Et2O then chloroacetic acid; (b) H2NNH2, 1 N NaOH; (c) chloral hydrate, Na2SO4,
H2NOH–HCl, H2O, HCl; (d) H2SO4; (e) BnBr, NaH, THF or ArB(OH)2, Cu(OAc)2, pyridine, CH2Cl2; (f) EtOH, 40 °C.

5632
M. G. Bursavich et al. / Bioorg. Med. Chem. Lett. 17 (2007) 5630–5633
Table 2. In vitro ADAMTS-5 inhibition data for R4 N-substituted 50-
phenyl-30H-spiro[indoline-3,20-[1,3,4]thiadiazol]-2-ones
sub-micromolar ADAMTS-5 inhibition (IC50
0.61 lM).
=
R2
R3
The most consistently potent series of thiadiazoles are
obtained when R1 = OMe and R4 is substituted with
either an aryl or benzyl group 33–37 (Table 2). While
the R1 = OMe, R4 = Me analog 32 possesses moderate
NH
N
N
S
ADAMTS-5
inhibition
(IC50 = 7.5 lM),
the
R4
R1 = OMe, R4 = benzyl analog shows sub-micromolar
activity (33: IC50 = 0.87 lM). Adding an R2 = Me
substituent to 33 gives a more potent compound
(34: IC50 = 0.64 lM). Other variations at R4 when
R1 = OMe and R2 = Me also give active compounds
(R4 = (o-Me)Ph 35: IC50 = 0.90 lM; R4 = (p-Cl)Ph 36:
IC50 = 1.5 lM). Attempting to combine optimal substit-
uents in all four positions (R1 = OMe; R2 = Me; R3 =
n-Pr; R4 = Bn) affords the sub-micromolar ADAMTS-
5 inhibitor 37 (IC50 = 0.95 lM).
O
R1
R1
Compound
R2
R3
R4
ADAM TS-5
a
IC50 (lM)
26
27
28
29
30
31
32
33
34
35
36
37
H
H
H
Me
i-Pr
Ph
24.1
3.1
H
H
H
H
Me
H
H
2.6
4.6
H
H
Me
Me
Me
Me
Me
H
H
(o-Me)Ph
Bn
Me
1.2
H
0.61
7.5
OMe
OMe
OMe
OMe
OMe
OMe
H
H
H
Bn
Bn
0.87
0.64
0.90
1.5
Given the suggestive mouse ADAMTS-5 knockout data
discussed above,3,4 we assessed ADAMTS-5/ADAMTS-
4 selectivity for several of the more potent thiadiazoles
presented in this manuscript (Table 3). Analogs 23 and
33–35 show high levels of selectivity over ADAMTS-4.
Only compound 35 shows any appreciable activity
against ADAMTS-4, but still maintains 11-fold selectiv-
ity for ADAMTS-5. To follow up the observed ADAM-
TS selectivity we also assessed metalloprotease
selectivity against both MMP-12 and MMP-13.
Whereas none of the analogs showed any activity
against MMP-13, compounds 23 and 33 show modest
activity against MMP-12 (IC50 = 1.7 and 5.6 lM,
respectively). Compound 34 appears to be very selective
over ADAMTS-4, MMP12, and MMP13.
Me
Me
Me
Me
H
H
(o-Me)Ph
(p-Cl)Ph
Bn
H
n-Pr
0.95
a Values are means of two experiments, standard deviations are 10%.
ADAMTS-5 inhibition data for the R4 N-substituted
thiadiazoles 26–37 are shown in Table 2. While incorpo-
ration of the R4 = Me group in 26 affords no potency in-
crease over the R4 = H analog 8 (24: IC50 = 24.1 lM vs
8: IC50 = 24.2 lM), the R4 = i-Pr analog 27 shows nearly
a 10-fold increase in potency (IC50 = 3.1 lM). The
R4 = Ph analog 28 shows similar low micromolar
ADAMTS-5 potency (IC50 = 2.6 lM). Incorporation
of an R1 = Me into the R4 substituted thiadiazole ana-
logs 29–31 shows continued potency improvement over
the R1 = H analogs. While the R1 = Me, R4 = Me
analog 29 does not give a potency increase over 13
(13: IC50 = 3.7 lM vs 29: IC50 = 4.6 lM), combining
R1, R2 = Me and R4 = (o-Me)Ph substituents affords
compound 30 (IC50 = 1.2 lM) with low micromolar
activity. The R1 = R2 = Me, R4 = benzyl analog 31 gives
In conclusion, we have presented a series of 50-phenyl-
30H-spiro[indoline-3,20-[1,3,4]thiadiazol]-2-one ADAM-
TS-5 inhibitors. This series of compounds has tractable
SAR with several analogs of sub-micromolar potency
demonstrating functional selectivity for ADAMTS-5
over ADAMTS-4, MMP-12, and MMP-13. The contin-
ued development of selective ADAMTS-5 inhibitors is
currently ongoing and will be reported in due course.
Table 3. In vitro selectivity data for 50-phenyl-30H-spiro[indoline-3,20-[1,3,4]thiadiazol]-2-ones
R2
Pr
NH
N
NH
N
N
S
N
S
H
R4
O
O
MeO
MeO
23
33: R2 = H, R4 = Bn
34: R2 = Me, R4 = Bn
35: R2 = Me, R4 = o-Me Ph
a
a
a
a
Compound
ADAMTS-5 IC50 (lM)
ADAMTS-4 IC50 (lM)
MMP13 IC50 (lM)
MMP12 IC50 (lM)
23
33
34
35
0.78
0.87
0.64
0.90
>22
>22
>22
10.5
>22
1.7
5.6
>100
>100
>100
>22
>22
a Values are means of two experiments, standard deviations are 10%.

M. G. Bursavich et al. / Bioorg. Med. Chem. Lett. 17 (2007) 5630–5633
5633
to target studies were conducted using fluorescence polar-
ization experiments using a xanthene-based ADAMTS-5
hydroxamic acid inhibitor.
Acknowledgments
We thank Dr. John Ellingboe for support of this work
and Dr. Jeremy I. Levin for useful discussion in prepa-
ration of this manuscript. We thank Dr. Kristina Cunn-
ingham for performing fluorescence polarization
binding to target studies.
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