Reverse hydroxamate-based selective TACE inhibitors

Article abstract of DOI:10.1016/j.bmcl.2004.03.048

Reverse hydroxamate-based selective TACE inhibitors are described. They have potent TACE inhibitory activities and excellent selectivities against MMP-1, 2, 3, 8, 9, 13, 14, and 17. One representative compound, 18 has demonstrated an excellent oral inhibitory activity of the lipopolysaccharide (LPS)-stimulated TNF-α production in rats.

Full text of DOI:10.1016/j.bmcl.2004.03.048

Bioorganic & Medicinal Chemistry Letters 14 (2004) 2897–2900
Reverse hydroxamate-based selective TACE inhibitors
Noriyuki Kamei, Tomohiro Tanaka, Kentaro Kawai, Kyosei Miyawaki,
Akihiko Okuyama, Yoshiko Murakami, Yoshio Arakawa, Makoto Haino,
Tatsuhiro Harada and Masanao Shimano*
Kaken Pharmaceutical Co., Ltd, 14, Shinomiya, Minamikawara-cho, Kyoto, Yamashina-ku 607-8042, Japan
Received 9 February 2004; accepted 12 March 2004
Abstract—Reverse hydroxamate-based selective TACE inhibitors are described. They have potent TACE inhibitory activities and
excellent selectivities against MMP-1, 2, 3, 8, 9, 13, 14, and 17. One representative compound, 18 has demonstrated an excellent oral
inhibitory activity of the lipopolysaccharide (LPS)-stimulated TNF-a production in rats.
Ó 2004 Elsevier Ltd. All rights reserved.
Tumor necrosis factor a (TNF-a) is one of the major
pro-inflammatory cytokines that is primarily or partly
responsible for many kinds of inflammatory events
within the living body.1 The landmark success of cur-
rently marketed biologics targeting TNF-a, etanercept
(EnbrelTM), and infliximab(Remicade TM), has clearly
implicated TNF-a in rheumatoid arthritis,2 and the
other therapeutic indications are aggressively under
exploration.3 However, these two biologics are orally
inactive and are inevitably administered parenterally.
Furthermore, biologics commonly have a long half-life
in the body and the above two biologics are not the
exception. This may untimely cause infection incidents
during the anti-TNF-a therapy, because TNF-a also
plays an important role as a self-defense factor against
pathogen.4 Another concern is allergic reactions to the
biologics and the neutralization antibody productions.5
Based on these points of view, orally active anti-TNF-a
small molecules with appropriate half-lives in the body
are highly desirable.
TACE may be one of the attractive targets in the anti-
TNF-a therapy,7 and as a matter of fact, there have been
many reports on the TACE inhibitors.8;9 The represen-
tative selective TACE inhibitors are shown in Figure
1.8b;d;f
TACE is structurally categorized into the ADAM (a
disintegrin and a metalloprotease) family, but its cata-
lytic site is quite similar to that of matrix metallopro-
teinases (MMPs).10 MMPs are involved in the
degradation and remodeling of connective tissues, and
about 20 MMPs have been uncovered so far.11 Each
MMP should have distinctive or wide-ranging roles in
the body, but the complete clarification of each MMP’s
functions leaves tremendous efforts still to be done.
Some broad-spectrum and partially selective MMP
inhibitors have been reported to cause musculoskeletal
side effects in clinical trials.12 Other experimental evi-
dence that MMP-9 is essential for the liver regenera-
tion13 and that MMP-14-deficiency causes arthritis-like
diseases14 also supports that the inhibition of MMPs as
foreboding difficulties to start a TACE inhibitor project
for the treatment of RA. Accordingly, we tried to seek
TNF-a exists in two forms, the membrane-bound form
(pro-TNF-a) and the soluble form (matured-TNF-a).
Pro-TNF-a is processed into matured-TNF-a b y a
metalloproteinase. This metalloproteinase is called the
TNF-a converting enzyme (TACE),6 and TACE has
been demonstrated to be the enzyme responsible for this
process. More than 90% of matured-TNF-a is catalyti-
cally prepared by TACE and this fact implies that
* Corresponding author. Tel.: +81-75-594-0787; fax: +81-75-594-0790;
e-mail: shimano_masanao@kaken.co.jp
Figure 1. Selective TACE inhibitors.
0960-894X/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2004.03.048

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N. Kamei et al. / Bioorg. Med. Chem. Lett. 14 (2004) 2897–2900
selective TACE inhibitors over MMPs to preclude any
unexpected side effects caused by the inhibition of the
MMPs.
intermediate 6. After the deprotonation of 6 with lith-
ium diisopropylamide, Julia coupling with a variety of
aldehydes17 gave rise to the corresponding alcohols 7c–
d. In the case of using esters17 as an electrophile, the
co-existence of stoichiometric lithium hexamethyldisi-
lazide resulted in the corresponding b-ketosulfones with
much better yields, which were successively reduced to
alcohols 7a–b in a one-pot procedure. Dehydration of
alcohols 7a–d afforded the a,b-unsaturated sulfones 8a–
d. The 1,4-addition of hydroxylamine to compounds 8a–
d afforded the hydroxylamine adducts 9a–d, which were
finally converted into the RH derivatives 10a–d by
the treatment of acetic formic mixed anhydride. The
deprotection of the acetal group of 9b–d afforded the
free ketones, which were converted into the RH deriv-
atives 10e,g, and 10i, respectively, in a similar fashion.
The treatment of 9b–d with hydroxylamine under acidic
conditions provided the corresponding oximes, which
were then converted into 10f,h, and 10j, respectively.
Our TACE program started by exploring the SAR of a
series of sulfonamide hydroxamate derivatives, and at
the same time, by probing their pharmacokinetics, since
the oral bioavailability and metabolic rate of a drug
candidate often become a major concern in the late
development stage. During this pilot study,15 we
observed that many of the prepared sulfonamide
hydroxamate derivatives have a strong tendency toward
hydrolysis of the hydroxamate moiety, eventually pro-
ducing highly toxic hydroxylamine in the rodent blood
plasma. Another drawback was the oral bioavailability.
The accumulation of the SAR and the pharmacokinetics
of the in-house TACE inhibiting compounds directed us
toward the reverse hydroxamate (RH) derivatives16 with
which, we postulated, the aforementioned problems
would be overcome. In this letter, we would like to
describe the syntheses and biological profiles for a series
of RH derivatives.
The inhibitory activities of compounds 10a–c,e–j against
TACE and MMPs (MMP-1, 2, 3, 8, 9, 13, 14, 17) are
summarized in Table 1. Those against MMP-1, 8, 14
were very weak (>100,000 nM) and were omitted from
the table. Compounds 10 exhibited excellent inhibitory
activities if compared to the structurally similar
RHs 10a–j were prepared as shown in Scheme 1. The
coupling of phenol 4 and 4-chloromethyl-2-methyl-
quinoline 5 under basic conditions gave the common
Scheme 1. Reagents and conditions: (a) 4 M NaOHaq, EtOH; (b) LDA, aldehyde; (c) LDA, LHMDS, ester, AcOH, NaBH4, MeOH; (d) MsCl,
Et3N; (e) 50% NH2OHaq, THF; (f) HCOOH, Ac2O, THF or pyridine, MeOH; (g) 2 M HClaq; (h) 2 M HClaq, 50% NH2OHaq.
Table 1. Inhibitory activities of reverse hydroxamates 10a–c,e–j, 11, 12, 15a,b, and 18 against TACE and MMPsa
Compound
TACEb
MMP-2b
MMP-3b
MMP-9b
MMP-13b
MMP-17b
10a
10b
10c
10e
10f
10g
10h
10i
10j
11
45
15
33,000
7000
6000
6000
280
>100,000
40,000
12,000
28,000
20,000
53,000
40,000
50,000
45,000
53,000
31,000
28,000
>100,000
90,000
7300
3300
2000
220
60
20
1200
2000
1600
1300
550
5000
7000
1100
600
30,000
22,000
15,000
8000
9.0
4.5
3.0
3.5
2.0
2.2
3.0
3.0
6.0
12
7000
3300
500
350
4000
330
41,000
30,000
60,000
55,000
7000
6300
5000
5500
10,000
1500
10,000
1500
22,000
15,000
25,000
25,000
7000
2500
3000
3000
4000
550
12
15a
15b
18
>100,000
5000
40,000
4000
4000
350
a Inhibitory activities against MMP-1, 8, 14 were weak (IC50 > 100,000 nM) and were not shown at the table.
b IC50, nM.

N. Kamei et al. / Bioorg. Med. Chem. Lett. 14 (2004) 2897–2900
2899
Scheme 2. Reagents and conditions: (a) 6, LDA, aldehyde; (b) MsCl, Et3N; (c) 50% NH2OHaq, THF; (d) HCOOH, Ac2O, THF or pyridine, MeOH;
(e) 6, LDA, LHMDS, ester, AcOH, NaBH4, MeOH; (f) 2 M HCl; (g) PivCl, Et3N; (h)iPrCHO, NaBH(OAc)3, AcOH.
hydroxamate derivatives,15 though the RH group is
known to have a less potent zinc binding capacity than
that of the hydroxamate group.18 This fact suggests that
the character of the RH group is favorable for the
binding to the catalytic site of TACE. The TACE
inhibitory activities among 10a,e, and 10f showed that
the capacities of the hydrogen bond acceptor and/or
hydrogen bond receptor might contribute for the
enhancement of the inhibitory activity. The one meth-
ylene carbon insertion between the cyclohexyl ring and
the carbon directly attached to the RH group increased
the TACE inhibitory activity shown by the comparison
between 10e,f, and 10g,h, respectively. Another inter-
esting and delightful finding was that the existence of the
double bond definitely increased the TACE selectivity
over MMPs. The RH group itself also slightly increased
the TACE selectivity.
The series of RH derivatives we have explained in this
letter had fairly good to excellent pharmacokinetics. The
human plasma protein binding ratios of the cyclohexyl
compounds 10 were just about 95%. Compound 10a had
an excellent oral bioavailability, 53% in rodent and was
stable in the rodent blood plasma. Heterocyclic com-
pounds 15b and 18 had much better human plasma pro-
tein binding ratios (81% and 74%, respectively) and both
of them have proved satisfactory in terms of solubility.
The in vivo TACE inhibitory activities of compound 18
was measured by the inhibition of the LPS-stimulated
TNF-a production in rats. The compound, orally
administered at 30 mg/kg 4 h prior to the LPS injection,
provided the 92% reduction of the blood TNF-a levels,19
and the blood levels of 18 was well above the IC50
against TACE.
Transformation of the cyclohexyl group into a hetero-
cyclic group was carried out to examine whether there
was a favorable improvement in the activity and the
selectivity, and at the same time, in order to compare
the difference in the pharmacokinetics properties between
the cyclohexyl compounds and the heterocyclic com-
pounds. The heterocyclic compounds were synthesized in
the same fashion as that of the cyclohexyl compounds
(Scheme 2).17 Compound 13 was subjected to deprotec-
tion of the Boc group, followed by the treatment of
pivaloyl chloride and Et3N, to afford the pivalamide 14a.
On the other hand, compounds 13 and 16 were trans-
formed into compounds 14b and 17, respectively, by
deprotection of the Boc group and the following reduc-
tive amination. Compounds 14a–b and 17 were converted
into the corresponding RHs 15a–b and 18, respectively.
In summary, we have created a series of RH derivatives
with excellent TACE inhibitory activities and selectivities
against the tested MMPs. The SAR of our compounds
suggested that the cyclohexyl moiety has a strong influ-
ence on both the TACE inhibitory activities and the
selectivities over MMPs. The pharmacokinetics of our
compounds, especially with the heterocyclic substituent,
was satisfactory as a drug candidate and the stability in
the rodent blood plasma was effectively improved by
changing the zinc binding group from the hydroxamate
moiety to the RH moiety. Since all of the compounds
explained above are racemics, the optically pure forms
would have the preferable characteristics.
Acknowledgements
We thank Ms. Kazumi Sumita and Ms. Miyako Ohara
for their analytical assistance.
The inhibitory activities of compounds 11, 12, 15a–b, 18
are summarized in Table 1. The inhibitory characteristics
of each individual compound possessing the heterocyclic
group were in much the same with those of cyclohexyl
compounds. The double bond effect remained intact and
the comparison between 15b and 18 clearly demonstrated
that this double bond should have special abilities for the
TACE selectivity, though it is still unclear whether the
characteristics of the double bond itself or the three-
dimensional shape of the six-membered ring restricted by
the double bond accounts for this difference.
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DMSO, 10 mM HCl, 0.5% methylcellulose) (feeding) 4 h
prior to the intravenous injection of LPS (1 mg/kg). After
90 min, the blood was collected from the tail vein and the
plasma was prepared. The plasma concentrations of TNF-
a were measured by ELISA (DIACLONE). The TNF-a
concentrations from an n ¼ 4 rats group receiving the test
compound were compared to those from a control group
receiving vehicle alone (n ¼ 6–7).