Design, synthesis, and evaluation of benzothiadiazepine hydroxamates as selective tumor necrosis factor-α converting enzyme inhibitors

Article abstract of DOI:10.1021/jm020475w

Elevated levels of tumor necrosis factor-α (TNF-α) have been associated with several inflammatory diseases, and therefore, strategies for its suppression have become important targets in drug discovery. Our efforts to suppress TNF-α have centered on the i

Full text of DOI:10.1021/jm020475w

J . Med. Chem. 2003, 46, 1811-1823
1811
Articles
Design , Syn th esis, a n d Eva lu a tion of Ben zoth ia d ia zep in e Hyd r oxa m a tes a s
Selective Tu m or Necr osis F a ctor -r Con ver tin g En zym e In h ibitor s
Robert J . Cherney,* J ames J .-W. Duan,* Matthew E. Voss, Lihua Chen, Li Wang, Dayton T. Meyer,
Zelda R. Wasserman, Karl D. Hardman, Rui-Qin Liu, Maryanne B. Covington, Mingxin Qian,
Sandhya Mandlekar, David D. Christ, J ames M. Trzaskos, Robert C. Newton, Ronald L. Magolda,
Ruth R. Wexler, and Carl P. Decicco
Bristol-Myers Squibb Pharmaceutical Research Institute, Princeton, New J ersey 08543-4000
Received October 24, 2002
Elevated levels of tumor necrosis factor-R (TNF-R) have been associated with several
inflammatory diseases, and therefore, strategies for its suppression have become important
targets in drug discovery. Our efforts to suppress TNF-R have centered on the inhibition of
TNF-R converting enzyme (TACE) through the use of hydroxamate inhibitors. Starting from
broad-spectrum matrix metalloproteinase (MMP) inhibitors, we have designed and synthesized
novel benzothiadiazepines as potent and selective TACE inhibitors. The benzothiadiazepines
were synthesized with variation in P1 and P1′ in order to effect potency and selectivity. The
inhibitors were evaluated versus porcine TACE (pTACE), and the initial selectivity was assessed
with counterscreens of MMP-1, -2, and -9. Several potent and selective inhibitors were
discovered with compound 41 being the most active against pTACE (Ki ) 5 nM) while still
maintaining good selectivity versus the MMP’s (at least 75-fold). Most compounds were assessed
in the human peripheral blood mononuclear cell assay (PBMC) and the human whole blood
assay (WBA) to determine their ability to suppress TNF-R. Compound 32 was the most potent
compound in the PBMC assay (IC50 ) 0.35 µM), while compound 62 was the most active in the
WBA (IC50 ) 1.4 µM).
Sch em e 1. Benzothiadiazepine Design
In tr od u ction
Tumor necrosis factor-R (TNF-R) is a cytokine nor-
mally produced by mononuclear cells in response to
immunostimuli to protect against infection, tissue de-
struction, and tumors.1 However, when overproduced,
it is proinflammatory in nature and initiates a cascade
of other agents including IL-1 and IL-6. The over-
production of TNF-R has been linked to several diseases
including rheumatoid arthritis (RA),2 Crohn’s disease,3
and psoriasis.4 TNF-R is initially produced as a mem-
brane-bound 26 kDa propeptide on the cell surface.
TNF-R converting enzyme (TACE)5,6 is the metallo-
proteinase that processes pro-TNF-R to its soluble 17
kDa form, thus resulting in its release from the cell
surface. TACE (ADAM-17) is a member of the ada-
malysin/reprolysin subfamily contained within the
metzincin superfamily, which also includes the matrix
metalloproteinases (MMPs).7,8 In fact, initially it was
shown that a subset of MMP inhibitors was capable of
inhibiting the release of TNF-R from cells through their
interaction with TACE.9-11 In the intervening time,
TACE has been purified and cloned12,13 as well as
crystallized with a bound inhibitor.14 As the major
sheddase identified in the release of soluble TNF-R,
there has been a great deal of interest in the design of
TACE inhibitors15 in order to suppress the amount of
circulating TNF-R.16 The beneficial effect of TNF-R
suppression was demonstrated clinically via two protein-
based drugs in RA and Crohn’s disease.17-20 A bioavail-
able small molecule that could achieve this level of
effectiveness is highly desirable. As a result, we under-
took the structure-based design of novel TACE inhibi-
tors via MMP inhibitor leads.
Design
An early, non-peptidic lead compound to emerge from
the MMP field was the Novartis compound CGS
27023A.21 Scheme 1 shows the general binding features
of CGS 27023A functioning as an MMP inhibitor that
we utilized in our design.21 The tetrahedral sulfone not
* To whom correspondence should be addressed. For R.J .C.: (phone)
609-252-3066; (fax) 609-252-6601; (e-mail) robert.cherney@bms.com.
For J .J .-W.D.: (phone) 609-252-4199; (fax) 609-252-7199; (e-mail)
james.duan@bms.com.
10.1021/jm020475w CCC: $25.00 © 2003 American Chemical Society
Published on Web 04/02/2003

1812 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10
Cherney et al.
Sch em e 2a
F igu r e 1. Overlay of benzothiadiazepine 2 (white) and CGS
27023A (green).
a
Reagents: (a) 4-methoxy-2-nitrobenzenesulfonyl chloride,
iPr2NEt, CH2Cl2, 0 °C; (b) H2, 10% Pd/C, MeOH; (c) (i) 37% HCHO,
MeOH, (ii) p-TsOH, DMF; (d) NH2OH, KOH, MeOH.
only directs the p-methoxyphenyl group into the S1′
pocket but also acts as a hydrogen bond acceptor from
Leu-164 of the protein backbone. In turn, the pyridyl
group is oriented toward the solvent-exposed area of the
active site and therefore has minimal contact with the
protein. In addition, the sulfonamide orients bond a-b
and bond c-d synplanar to one another, thus placing b
and d in proximity. We felt this was an excellent place
to constrain the system and proposed to connect b to d
while removing the pyridyl ring, since it appeared
solvent exposed. In addition, from docking experiments
it appeared that insertion of a hydrogen bond donor
would be beneficial for interaction with the backbone
amide carbonyl of residue Pro-221. This hydrogen bond
could best be realized with the installation of an N-H
at the ring juncture. The above requirements are best
fulfilled by a six-membered benzothiadiazine 1 or seven-
membered benzothiadiazepine 2 as shown in Scheme
1. In fact, benzothiadiazepine 2 could adopt a conforma-
tion similar to that of CGS 27023A as shown by an
overlay in Figure 1.
Sch em e 3a
Syn th esis
As shown in Scheme 2, the benzothiadiazine 7 was
synthesized from the methyl ester of D-alanine 3.
Commercially available 4-methoxy-2-nitrobenzene-
sulfonyl chloride was coupled with 3 to give sulfonamide
4, which was subsequently reduced. The resulting
aniline 5 was stirred in methanol with formaldehyde
and then cyclized with p-toluenesulfonic acid to give the
six-membered ring 6. Direct conversion of the methyl
ester to the target hydroxamate 7 was accomplished
with a hydroxylamine solution.
a
Reagents: (a) 60% NaH, allyl bromide, DMF; (b) (i) O3, CH2Cl2,
MeOH, (ii) Me2S; (c) Zn, HOAc, ∆; (d) H2NOH, KOH, MeOH.
to the target hydroxamate 11 with a hydroxylamine
solution. Compound 2 and compounds 11-14 were
made in this manner according to Scheme 3.
As illustrated with compound 11 (Scheme 3), the first
benzothiadiazepines were synthesized from the sulfon-
amide 4. Alkylation of the sulfonamide with allyl
bromide gave 8, which was oxidized to aldehyde 9.
Treatment of 9 with zinc/acetic acid at reflux effected
reduction of the nitro group with a concomitant intra-
molecular reductive alkylation to generate benzo-
thiadiazepine 10. Again, the methyl ester was converted
Synthesis of second-generation benzothiadiazepines
required a suitably protected C-7 hydroxyl group. As
illustrated with compound 26 in Scheme 4, the synthesis
was initiated with a chemoselective alkylation of 4-amino-
3-nitrophenol 15 to give the benzyl-protected 16. Diazo-
tization of 16 allowed for its conversion to the sulfonyl
chloride 17. On a large scale, we found that the use of
commercially available sulfurous acid solutions was

Benzothiadiazepine Hydroxamates
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10 1813
Sch em e 4a
Sch em e 5a
a
Reagents: (a) NMM, 2,4-dinitrobenzenesulfonyl chloride, di-
oxane; (b) BrCH2CH2OH, DEAD, PPh3, THF; (c) (i) Fe, AcOH, 60
°C, (ii) NMM, DMF, 80 °C; (d) 3,5-dimethoxybenzaldehyde, NaBH4,
EtOH; (e) (i) TFA, CH2Cl2, (ii) H2NOBn, BOP, NMM, DMF, (iii)
H2, 5% Pd/BaSO4, MeOH; (f) (i) LiOH, THF/H2O, (ii) H2NOBn,
BOP, NMM, DMF, (iii) 3,5-dimethylbenzoic acid, BOP reagent,
NMM, DMF; (g) H2, 5% Pd/BaSO4, MeOH.
a
Reagents: (a) 1 M KOtBu, BnBr, DMF, 0 °C; (b) (i) NaNO2,
TFA, HCl, 0 °C, (ii) H2SO3, CuCl2, CuCl, AcOH; (c) D-Ala-OMe‚HCl,
iPr2NEt, CH2Cl2, 0 °C; (d) BrCH2CH2OH, DEAD, PPh3, THF; (e)
Fe, AcOH, 60 °C; (f) NMM, DMF, 80 °C; (g) H2NOH, KOH, MeOH;
(h) H2, 5% Pd/BaSO4, MeOH; (i) 1 M KOtBu, 3,5-dimethylbenzyl
bromide, DMF, 0 °C; (j) H2NOH, KOH, MeOH (method A) or (i)
TFA, CH2Cl2; (ii) BOP reagent, H2NOH‚HCl, NMM, DMF (method
B).
gave the versatile benzothiadiazepine 47 (47a was
obtained in an analogous manner starting with D-Leu-
OtBu and 2,4-dinitrophenylsulfonyl chloride). Reductive
amination of 47a with 3,5-dimethoxybenzaldehyde gave
48. The ester was converted to the O-benzyl-protected
hydroxamate before hydrogenation afforded the target
hydroxamate 49. Conversion of 47 to the O-benzyl
hydroxamate followed by an acylation with 3,5-di-
methylbenzoic acid gave 50. Hydrogenation of this
material gave hydroxamate 51.
As illustrated with compound 55 (Scheme 6), phenol
24 was also converted to aryl triflate 53 and utilized
for palladium couplings. Again, standard derivatization
of ester 54 gave hydroxamate 55. Compound 56 was also
made by this method. As shown with compound 58,
phenol 24 was directly coupled to 4-(trifluoromethyl)-
benzeneboronic acid23,24 to give phenyl ether 57, which
was in turn processed to hydroxamate 58. Compounds
59-62 were made in analogous fashion. Ether produc-
tion was also accomplished by an SNAr reaction of 24
with 1-fluoro-4-nitrobenzene and cesium carbonate to
yield 57a . Normal processing via method A gave the
hydroxamate 63.
more convenient than the use of gaseous sulfur dioxide
for this transformation. Condensation of 17 with a wide
variety of amino acid esters produced sulfonamide 18.
The sulfonamide was substituted under Mitsunobu
conditions22 to give bromide 19. Reduction of the nitro
gave aniline 20, which was cyclized to benzothiadi-
azepine 21 by heating with 4-methylmorpholine. Treat-
ment of 21 with a hydroxylamine solution gave the
hydroxamate 22, which was debenzylated to 23. The
benzyl group of 21 was also removed to give the versatile
phenol 24. In many cases, phenol 24 was alkylated with
substituted benzyl bromides to give 25. This was
processed to the hydroxamate via two methods depend-
ing on the ester present in 25. For methyl esters, the
aforementioned direct transformation with hydroxyl-
amine was used to afford 26 (method A; compounds 27-
30 were made via this method). If a tert-butyl ester was
present, it was converted to the carboxylate and then
directly coupled with BOP reagent and hydroxylamine
to give the target hydroxamate (method B; compounds
31-40 were made via this method).
Resu lts a n d Discu ssion
As shown in Scheme 5, an amino group was incorpo-
rated at C-7 with the use of commercially available 2,4-
dinitrophenylsulfonyl chloride. Hence, sulfonamide 45
was used in the Mitsunobu reaction to give bromide 46.
Reduction of 46 followed by cyclization in warm DMF
The newly synthesized hydroxamates were evaluated
against porcine TACE (pTACE) to assess their inhibi-
tory potency.25 Porcine TACE was selected as a result
of its availability and its high homology to human

1814 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10
Cherney et al.
Sch em e 6a
a
Reagent: (a) Tf2O, iPr2NEt, CH2Cl2, -78 °C; (b) Pd(OAc)2, C6H5B(OH)2, PPh3, toluene, 110 °C; (c) H2NOH, KOH, MeOH; (d) Cu(OAc)2,
Et3N, 4-trifluoromethylbenzeneboronic acid, 4 Å molecular sieves, CH2Cl2; (e) 1-fluoro-4-nitrobenzene, Cs2CO3, DMF.
TACE. All compounds were then screened for initial
selectivity against MMP-1, -2, and -9.25 Broad-spectrum
MMP inhibitors have displayed musculoskeletal pain
and inflammation as a side effect in clinical trials;26,27
hence, it was our desire to achieve at least 100-fold
selectivity for pTACE over MMP-1, -2, and -9. As shown
in Table 1, we were gratified to observe good pTACE
affinity for benzothiadiazepine 11, thereby validating
our design (Scheme 1). The six-membered benzothia-
diazine 7 was about 10-fold less active versus pTACE
than the seven-membered 11. This was also reflected
in their MMP values, and therefore, we concentrated
on the seven-membered template. With this information
in hand, we examined the effect of substitution at P1
and P1′ of the benzothiadiazepine. Without substitution
at P1 and P1′, the benzothiadiazepine was inactive as
shown with compound 12. However, most of the activity
was restored with a P1 methyl substitution as in 13,
which was about 2-fold less active than 11 in pTACE.
In contrast, methoxy substitution alone at P1′ (see
compound 14) was inactive. Early optimization of P1
indicated that an isopropyl group was beneficial, since
2 was about 4-fold more active than 11.
we needed to go deeper into S1′. In addition, we
surmised that as the main recognition pocket for the
MMPs, S1′ would be an excellent location to achieve
selectivity for pTACE over the MMPs. With a methyl
in P1, we began to probe S1′ with benzyl groups
alkylated on the C-7 phenol. The phenol itself, 23, did
have affinity for pTACE (Ki ) 391 nM); however, it was
virtually devoid of selectivity versus the MMPs with
only a 2-fold margin over MMP-1 relative to pTACE.
The benzyl ether 22 did not improve the pTACE activity;
however, selectivity against MMP-1 was increased to
at least 7-fold. The selectivity of the benzyl substituent
over MMP-1 is probably attributed to the short S1′
pocket of MMP-128-30 relative to the S1′ pocket of TACE.
Selectivity versus MMP-2 and -9 remained unaffected
as a result of their long S1′ pockets, and therefore, other
factors were considered. In this lower section of S1′,
homology modeling of TACE predicted pronounced
curvature31 whereas the MMPs are essentially linear.
Taking advantage of this, it has been our experience32
that an appropriately placed 3,5-disubstituted aromatic
will maintain a good fit in TACE while clashing into
the wall of the MMP S1′pocket. We initially installed
the 3,5-dimethyl substituent in 26 and observed about
a 3-fold increase in pTACE affinity (Ki ) 229 nM)
relative to the parent benzyl 22. In accord with the
model,31 the 3,5-dimethyl substitution of 26 also dra-
matically improved the selectivity over the MMPs with
an MMP-9 selectivity of at least 9-fold. The 3,5-dichloro
substitution of 27 gave a profile similar to that of 26,
whereas the 3,5-dimethoxy 28 provided increases not
only in pTACE activity (Ki ) 27 nM) but also in MMP
selectivity, since 28 was selective over MMP-2 by 18-
fold relative to pTACE. As we progressed to the larger
3,5-diethoxy substitution, 29, the pTACE activity (Ki )
81 nM) decreased 3-fold relative to 28, indicating that
the 3,5-diethoxy substitution was approaching a size
incompatible with the size of the S1′ of pTACE. The
picolyl ether 30 was also investigated but was found to
have diminished (2-fold) pTACE affinity relative to 26.
To optimize the template further, we turned our atten-
tion to the isobutyl substituent in P1.
Ta ble 1. In Vitro Evaluation: Early Inhibitors
a
Ki (nM)
compd
n
R
R1
pTACE MMP-1 MMP-2 MMP-9
7
11
12
13
14
2
0
1
1
1
1
1
Me OMe
Me OMe
3100
274
>4949
1060
>4949
>4949
>4949
732
312
8
1915
75
1497
52
1050
104
H
Me
H
H
H
>1000
>2128
484
567
OMe >1000
>2128
751
iPr OMe
71
a
Ki values are an average from three determinations. Standard
deviations are less than 15% in all cases.
With these early compounds, we were able to validate
our design; however, to attain more affinity for pTACE,

Benzothiadiazepine Hydroxamates
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10 1815
Ta ble 2. In Vitro Evaluation of Benzothiadiazepine Inhibitors
b
IC50 (µM) or
% inhibition @ 10 µM
a
Ki (nM)
compd
R
X
Ar
pTACE
MMP-1 MMP-2
MMP-9
PBMC
WBA
23
22
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
49
51
55
56
58
59
60
61
62
63
Me
Me
Me
Me
Me
Me
Me
iBu
iBu
iBu
iPr
iPr
iPr
OH
391
705
229
153
27
81
544
89
980
>4949
>4949
>4949
>4949
>4949
>4949
479
<2.8
<2.8
136
49
491
690
75
100
3.6
NT
NT
NT
0.94
0.77
4.5
2.9
9.5
0.35
5.4
4.0
8.2
24.5
7.2
6.2
0.9
21%
30%
2%
OCH2
OCH2
OCH2
OCH2
OCH2
OCH2
OH
OCH2
OCH2
OCH2
OCH2
OCH2
OCH2
OCH2
OCH2
OCH2
phenyl
3,5-dimethylphenyl
3,5-dichlorophenyl
3,5-dimethoxyphenyl
3,5-diethoxyphenyl
2,6-dimethyl-4-pyridinyl
>2128
1209
>2128
>2128
1839
80
<2.1
>2128
981
>2128
>2128
>2128
>2128
>2128
>2128
>2128
118
12%
28 µM
26%
37%
20%
39%
16%
11%
17%
0%
0%
0%
31%
24 µM
38 µM
3%
3%
19%
NT
NT
NT
44%
NT
NT
11
phenyl
177
17
10
548
<2.8
154
647
3,5-dimethoxyphenyl
3,5-dimethoxyphenyl
3,5-diethoxyphenyl
4,5-dimethyl-2-thiazolyl
3,5-dimethoxyphenyl
3,5-diethoxyphenyl
3,5-dimethoxyphenyl
3,5-diethoxyphenyl
3,5-dimethoxyphenyl
3,5-dimethoxyphenyl
3,5-dimethoxyphenyl
3,5-dimethoxyphenyl
>4949
>4949
>4949
>4949
>4949
>4949
>4949
>4949
>4949
>4949
>4949
>4949
>4949
272
>4949
>4949
>4949
>4949
2019
>4949
>4949
51
3001
290
177
242
10
13
5
77
31
56
361
1595
960
332
1376
391
32
231
75
94
<2.8
2518
<2.8
<2.8
23
(CH2)2CO2Me
(CH2)2CO2Me
(CH2)3CO2Me
(CH2)3CO2Me
(CH2)4NHSO2Me OCH2
(CH2)4NHBoc
(CH2)4NH2
iBu
Me
Me
Me
Me
Me
Me
Me
Me
Me
1.1
1.9
OCH2
OCH2
NT
NT
NT
NT
NT
NT
0.84
NT
NT
NT
0.47
1.2
>2128
NHCH2
NHC(O) 3,5-dimethylphenyl
phenyl
347
>1000
>1000
>1000
758
621
734
>1000
129
11
993
8
2-(trifluoromethyl)phenyl
>2128
O
O
O
O
O
O
4-(trifluoromethyl)phenyl
4-methoxyphenyl
phenyl
4-biphenyl
3-nitrophenyl
4-nitrophenyl
7
<2.1
3
<2.1
96
<2.8
85
8
NT
1.4 µM
15 µM
19
a
b
Ki values are from three determinations. Standard deviations are less than 15% in all cases. IC50 values are an average from three
donors.
When the rigidity of the template and the lipophilicity
played 18-fold more affinity for pTACE than its corre-
sponding glutamate. In keeping with the trend above,
the 3,5-diethoxy 40 had excellent selectivity over the
MMPs tested (100-fold over MMP-2).
Continuing in the P1 position, several lysine deriva-
tives were also incorporated. The best compound in the
lysine series was the methyl sulfonamide 41, which had
excellent pTACE affinity (Ki ) 5 nM) and excellent
MMP selectivity (70-fold over MMP-2). Lipophilic groups
such as the N-Boc of 42 had a detrimental effect on
MMP selectivity. However, this was reversed by conver-
sion to the parent amine 43.
At this point, we assessed different substitution
patterns at C-7 to probe S1′. The 3,5-dimethoxybenzyl
substituent was incorporated into an amine (compound
49); however, it displayed diminished pTACE affinity
(3-fold) when compared to the corresponding 3,5-
dimethoxybenzyl ether 33. In addition, amine 49 lacked
substantial selectivity over MMP-2 and -9 and only
displayed selectivity over MMP-1. Switching to the
amide 51 resulted in an altered projection into S1′ and
a compound inactive in pTACE.
Other substitution patterns were probed as we in-
vestigated the direct attachment of the distal ring,
which gave the biaryl 55. These data suggest that the
curvature of the pTACE S1′ cannot accommodate the
biaryl of 55; however, 55 was accommodated by the
somewhat linear MMP S1′. The added o-trifluoromethyl
in P1 were increased, the parent isobutyl 31 had a 4-fold
increase in affinity for pTACE over the P1 methyl
compound 23. The same effect was observed for the
benzyl compound 32 compared to the P1 methyl com-
pound 22. To restore the MMP selectivity, the 3,5-
dimethoxy substitution was installed, and 33 was
almost 2-fold more active versus pTACE than the P1
methyl analogue 28. Continuing to probe P1, we in-
stalled the isopropyl group along with the 3,5-dimethoxy-
benzyl to afford 34, which was about 2-fold better than
28 and 33 versus pTACE. Perhaps even more important
was the effect the P1 isopropyl had on the MMP-2
selectivity, since 34 had a selectivity of 60-fold for
pTACE over MMP-2. Trying to capitalize on this, we
again increased the size of the 3,5-substitution to
diethoxy; however, 35 lost pTACE affinity while retain-
ing excellent MMP selectivity. We also found hetero-
cycles that were accommodated in S1′, since the thiazole
36 had good activity for pTACE and selectivity against
MMP-1, -2, and -9.
In an effort to gain more affinity, we extended P1 by
utilizing glutamate and homoglutamate methyl esters
at that position. The glutamate ester was paired with
the 3,5-dimethoxy (37) and 3,5-diethoxy (38) but proved
to be unattractive with only moderate pTACE affinity.
However, the homoglutamate ester of both the 3,5-
dimethoxy (39) and 3,5-diethoxy (40) surprisingly dis-

1816 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10
Cherney et al.
of 56 had poor affinity in pTACE and MMP-1, -2, and
-9. In this case, the 2-substitution probably disrupts the
binding in the MMP S1′ much like the 3,5-disubstitution
discussed earlier.
Exp er im en ta l Section
All reactions were performed under nitrogen with continu-
ous magnetic stirring unless otherwise stated. Anhydrous
solvents from commercial sources were used for all reactions.
All other reagents and solvents were reagent grade and used
as received from commercial sources. Flash chromatography
was performed with E. Merck Kieselgel 60 silica gel (230-
400 mesh). As indicated, final compounds were purified via
reversed-phase high-performance liquid chromatography
(HPLC) on a Dynamax C18 column using gradiant elution
(solvent A, water; solvent B, acetonitrile). All 1H NMR were
obtained on a Varian Inova spectrometer (operating at 300
MHz), and the signals are reported in ppm relative to TMS.
All mass spectra were recorded on a Finnigan Navigator mass
spectrometer using electrospray ionization (ESMS) in the
positive (pos) or negative (neg) mode as indicated. Elemental
analysis was performed by Quantitative Technologies Inc,
Bound Brook, NJ .
To install some curvature back into P1′, the phenyl
ethers were examined. Substitution of the distal ring
with small groups in the para position was allowable
because the trifluoromethyl (58), methoxy (59), and
unsubstituted cases (60) all showed fair pTACE affinity
while lacking MMP-2 and -9 selectivity. Large groups
(P1′ biphenyl 61) unable to negotiate the putative
curvature of the TACE S1′ were inactive while still
showing excellent MMP-2 and -9 activity. Moderate
pTACE activity was observered for mono substitution
at the meta position because the 3-nitro group of 62 was
tolerated. However, the best substitution was the 4-
nitro, as shown by 63 (pTACE Ki ) 11 nM).
Hydroxamic acids were prepared via two methods.
Meth od A (fr om a Meth yl Ester ). A 1.76 M NH2OH stock
solution was prepared by dissolving hydroxylamine hydro-
chloride (2.34 g) in hot MeOH (12 mL). After the mixture was
stirred for 5 min, a solution of KOH (2.81 g) in MeOH (7 mL)
was added dropwise. The resulting solution was cooled to room
temperature and filtered. The filtrate (approximately a 1.76
M NH2OH stock solution) was stored under nitrogen and used
in the next step. The methyl ester (1.17 mmol) was dissolved
in MeOH (5 mL) prior to the addition of the 1.76 M NH2OH
stock solution (2.66 mL, 4.68 mmol). After the mixture was
stirred for 2 h, 1 N HCl was added to adjust the pH to ∼5.
The solution was concentrated, and the residue was parti-
tioned between EtOAc and water. The organic layer was dried
and concentrated. Reverse-phase HPLC (water/acetonitrile) or
flash chromatography of the resulting residue gave the desired
hydroxamate.
Meth od B (fr om a ter t-Bu tyl Ester ). The tert-butyl ester
(0.58 mmol) was dissolved in CH2Cl2 (3 mL), and the mixture
was cooled to 0 °C prior to the addition of TFA (15 mL). The
reaction mixture was warmed to room temperature and was
stirred for 2 h (monitored by TLC for disappearance of tert-
butyl ester). The solution was concentrated and dried to give
the crude carboxylate, which was used directly in the next step.
The crude carboxylate was dissolved in DMF prior to the
addition of N-methylmorpholine (0.26 mL, 2.4 mmol). The
mixture was cooled to 0 °C, and BOP reagent (294 mg, 0.66
mmol) was added. After the mixture was stirred for 20 min at
0 °C, HONH2‚HCl (82 mg, 1.2 mmol) was added. The reaction
mixture was warmed to room temperature and was stirred
overnight. The solvent was removed, and the resulting residue
was partitioned between EtOAc and saturated NH4Cl solution.
The EtOAc was dried and concentrated. Flash chromatography
or reverse-phase HPLC (water/acetonitrile) of the resulting
residue gave the desired hydroxamate.
N-(4-Meth oxy-2-n itr oben zen esu lfon yl)-D-Alan in e Meth -
yl Ester (4). D-Alanine methyl ester hydrochloride 3 (3.9 g,
28.1 mmol) was dissolved in CH2Cl2 prior to the addition of
diisopropylethylamine (12.2 mL, 70.2 mmol). After the mixture
was cooled to 0 °C, 4-methoxy-2-nitrobenzenesulfonyl chloride
(8.5 g, 33.7 mmol) was added dropwise. The reaction mixture
was warmed to room temperature and was stirred overnight.
The reaction mixture was washed with 1 N HCl, saturated
NaHCO3, and brine. The CH2Cl2 was dried, filtered, and con-
centrated. Flash chromatography (35% ethyl acetate/hexane)
of the resulting residue gave the sulfonamide 4 (6.4 g, 20.1
mmol, 71%) as a white foam. 1H NMR (CDCl3): δ 1.47 (d, 3H),
3.56 (s, 3H), 3.85 (s, 3H), 4.2 (m, 1H), 6.01 (br d, 1H), 7.14
(dd, 1H), 7.4 (d, 1H), 7.98 (d, 1H). ESMS (neg), m/z: 317.1 (M
- H).
Cellu la r Assa ys
As shown in Table 2, the best compounds were
assessed in two cellular assays, the human peripheral
blood mononuclear cell assay (PBMC)33 and the human
whole blood assay (WBA),25 for their ability to suppress
the formation of TNF-R brought about by lipopoly-
saccharide (LPS) stimulation. Success in the PBMC
assay relies on the ability of the inhibitor to penetrate
into the cells, thus enabling it to inhibit TACE and
thereby to suppress the release of intracellular, soluble
TNF-R.34,35 In the WBA, compounds must also have the
ability to penetrate cells; however, this is complicated
by several other factors, the foremost being protein
binding. The majority of the benzothiadiazepines had
poor activity in the WBA. Nevertheless, several com-
pounds displayed reasonable PBMC activity, indicating
some ability to penetrate cells. In fact, two phenyl ethers
(58 and 62) and a benzyl ether (32) had PBMC values
only slightly displaced from their pTACE values, indi-
cating good cell penetration. However, for the most part,
the 3,5-disubstituted benzyl ethers had PBMC values
displaced by at least 20-fold from their pTACE values.
An exception to this was 27, which had its PBMC value
displaced by only 6-fold from the pTACE value. Yet,
even this case did not translate well into the WBA
because 27 exhibited poor inhibition (12% inhibition at
10 µM in WBA). Protein binding presumably has an
effect on the WBA value of 27, since the free fraction is
only 0.6% in human whole blood. Contrast this with the
most potent benzothiadiazepine in the WBA compound
62 (IC50 ) 1.4 µM), which has an 8-fold higher free
fraction (5% free) in protein binding.
Con clu sion
Utilizing structure-based design principles, we have
converted a sulfonamide MMP lead structure into a
novel benzothiadiazepine inhibitor. From key structural
differences between the MMPs and TACE, we modified
substitution patterns within P1′ to obtain selectivity and
affinity exclusively for TACE. The majority of these
inhibitors have displayed poor translation into cellular
assays, and we have suggested a combination of vari-
ables to be optimized. However, with their affinity for
TACE and selectivity over the MMPs, the benzothia-
diazepines should aid in the design and development
of future metalloproteinase inhibitors.
N-(4-Meth oxy-2-am in oben zen esu lfon yl)-D-Alan in e Meth -
yl Ester (5). The sulfonamide 4 (3.8 g, 11.9 mmol) was
dissolved in a mixture of MeOH (10 mL) and THF (3 mL) prior
to the addition of 10% Degussa Pd/C (380 mg). The solution
was placed on a Parr at 50 psi for 18 h. The solution was
filtered and concentrated. This gave the crude aniline 5 (3.4

Benzothiadiazepine Hydroxamates
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10 1817
g, 11.8 mmol, 99%) as a white foam of sufficient purity for the
next step. H NMR (CDCl3): δ 1.36 (d, 3H), 3.55 (s, 3H), 3.79
(s, 3H), 3.93 (m, 1H), 4.94 (br s, 2H), 5.42 (br d, 1H), 6.2 (d,
1H), 6.34 (dd, 1H), 7.58 (d, 1H). ESMS (pos), m/z: 289.2 (M +
H).
Compounds 2 and 12-14 were synthesized in a fashion
analogous to that of compound 11.
1
N-H yd r oxy-2-[2,3,4,5-t et r a h yd r ob en zo[1,2,5-f]t h ia d i-
a zep in e-1,1-d ioxid e]eth a n a m id e (12). 1H NMR (CD3OD):
δ 3.35-3.39 (m, 2H), 3.57-3.61 (m, 2H), 3.72 (s, 2H), 6.92-
6.99 (m, 2H), 7.35 (m, 1H), 7.71 (dd, 1H). MS (CI, NH3), m/z:
272 (M + H). HRMS calcd for (C10H14N3O4S + H) 272.0705,
found 272.0711. Anal. (C10H14N3O4S) C, H, N, S.
N -H y d r o x y -2(R )-[2,3,4,5-t e t r a h y d r o b e n zo [1,2,5-f]-
th ia d ia zep in e-1,1-d ioxid e]p r op a n a m id e (13). 1H NMR
(CD3OD): δ 1.33 (d, 3H), 3.36-3.45 (m, 1H), 3.60 (t, 2H), 3.68-
3.77 (m, 1H), 4.47 (q, 1H), 6.81-6.90 (m, 2H), 7.27 (m, 1H),
7.65 (dd, 1H). ESMS (neg), m/z: 283.9 (M - H). HRMS calcd
for (C11H16N3O4S + H) 286.0862, found 286.0863. Anal.
(C11H16N3O4S1‚0.1H2O) C, H, N, S.
N-Hydr oxy-2-[7-m eth oxy-2,3,4,5-tetr ah ydr oben zo[1,2,5-
f]th ia d ia zep in e-1,1-d ioxid e]eth a n a m id e (14). 1H NMR
(CD3OD): δ 3.28-3.35 (m, 2H), 3.54-3.58 (m, 2H), 3.68 (s,
2H), 3.80 (s, 3H), 6.50-6.55 (m, 2H), 7.62 (d, 1H). ESMS (neg)
300.0 (M - H). HRMS calcd for (C11H15N3O5S + H) 302.0811,
found 302.0807. Anal. (C11H15N3O5S1‚0.1H2O) C, H, N, S.
N-Hyd r oxy-2(R)-[7-(m eth oxy)-2,3,4,5-tetr a h yd r oben zo-
[1,2,5-f]t h ia d ia ze p in e -1,1-d io x id e ]-4-m e t h y lp e n t a n -
a m id e (2). 1H NMR (CD3OD): δ 0.88 (d, 3H), 0.98 (d, 3H),
2.13-2.24 (m, 1H), 3.26-3.38 (m, 1H), 3.51-3.61 (m, 2H), 3.76
(s, 3H), 3.76-3.85 (m, 2H), 6.38-6.41 (m, 2H), 7.58 (d, 1H).
ESMS (neg), m/z: 342.0 (M - H). HRMS calcd for (C14H22N3O5S
+ H) 344.1280, found 344.1277. Anal. (C14H22N3O5S) C, H, N,
S.
4-Ben zyloxy-2-n itr oa n ilin e (16). 4-Amino-3-nitrophenol
15 (25 g, 162.2 mmol) was dissolved in DMF and cooled to 0
°C. 1 M potassium tert-butoxide (162.2 mL) was added drop-
wise over 20 min. After the mixture was stirred for 30 min at
0 °C, benzyl bromide (19.8 mL, 162.2 mmol) in DMF was added
dropwise. After 30 min at 0 °C, the reaction was quenched
with NH4Cl solution. 4-Benzyloxy-2-nitroaniline 16 (32 g, 131
mmol, 81%) was isolated by filtration. 1H NMR (CDCl3): δ 5.03
(s, 2H), 5.9 (br s, 2H), 6.76 (d, 1H), 7.14 (dd, 1H), 7.4 (m, 5H),
7.65 (d, 1H). MS (CI, NH3), m/z: 245 (M + H).
2(R)-(6-Met h oxy-1,1-d ioxo-3,4-d ih yd r o-1H -1λ6-b en zo-
[1,2,4]t h ia d ia zin -2-yl)p r op ion ic Acid Met h yl E st er (6).
The aniline 5 (400 mg, 1.4 mmol) was dissolved in MeOH (4
mL) prior to the addition of 37% formaldehyde (1.8 mL). After
being stirred overnight, the solution was concentrated and
then was filtered through silica gel to give a clear oil (360 mg).
A portion of this material (100 mg) was dissolved in DMF (2
mL) prior to the addition of p-TsOH (10 mg). The solution was
stirred overnight before being concentrated. Flash chroma-
tography (30-70% ethyl acetate/hexane) of the resulting
residue gave the thiadiazine 6 (7 mg, 0.02 mmol) as a white
1
foam. H NMR (CDCl3): δ 1.46 (d, 3H), 3.54 (s, 3H), 3.77 (s,
3H), 4.4 (br s, 1H), 4.58 (m, 2H), 5.06 (m, 2H), 6.11 (d, 1H),
6.43 (dd, 1H), 7.57 (d, 1H). ESMS (pos), m/z: 323.0 (M + Na).
N-Hyd r oxy-2(R)-(6-m eth oxy-1,1-d ioxo-3,4-d ih yd r o-1H-
1λ6-ben zo[1,2,4]th iadiazin -2-yl)pr opion am ide (7). The thia-
diazine 6 (60 mg, 0.2 mmol) was incorporated into method A,
and HPLC gave the hydroxamate 7 (15 mg, 0.05 mmol, 25%)
1
as a white powder. H NMR (DMSO): δ 1.17 (br d, 3H), 3.69
(s, 3H), 4.04 (m, 1H), 4.89 (br d, 1H), 5.12 (dd, 1H), 6.19 (br s,
1H), 6.28 (dd, 1H), 7.13 (br s, 1H), 7.35 (d, 1H), 8.91 (br s,
1H), 10.74 (s, 1H). ESMS (neg), m/z: 299.8 (M - H). HRMS
calcd for (C11H15N3O5S + H) 302.081 068, found 302.081 282.
N-(4-Meth oxy-2-n itr oben zen esu lfon yl)-N-(a llyl)-D-Ala -
n in e Meth yl Ester (8). The sulfonamide 4 (4.54 g, 14.26
mmol) was dissolved in DMF prior to the addition of allyl
bromide (2.47 mL, 28.5 mmol) followed by portionwise addition
of 60% NaH (0.86 g, 21.4 mmol). After 20 min, pH 7 phosphate
buffer (200 mL) was added, and the mixture was extracted
with EtOAc. The EtOAc was washed with pH 7 buffer, water,
and brine before being dried and concentrated. The resulting
residue was purified by flash chromatography (30-40% ethyl
1
acetate/hexane) to give 8 (2.85 g, 56%) as a yellow liquid. H
NMR (CDCl3): δ 1.49 (d, 3H), 3.66 (s, 3H), 3.84-3.93 (m, 1H),
3.92 (s, 3H), 4.04-4.12 (m, 1H), 4.79 (q, 1H), 5.06-5.22 (m,
2H), 5.75-5.90 (m, 1H), 7.08-7.13 (m, 2H), 7.98 (d, 1H). ESMS
(pos), m/z: 381.0 (M + Na).
4-Ben zyloxy-2-n it r ob en zen esu lfon yl Ch lor id e (17).
4-Benzyloxy-2-nitroaniline 16 (1.5 g, 6.1 mmol) was dissolved
in TFA (20 mL) and concentrated HCl (2 mL). After the
mixture was cooled to 0 °C, NaNO2 (529.6 mg, 7.7 mmol) in
H2O was added dropwise over 30 min. Once the addition was
complete, the reaction mixture was stirred an additional 5 min.
The resulting solution was poured into AcOH (20 mL), H2SO3
(20 mL), and CuCl2 (412.8 mg, 3.1 mmol) containing a catalytic
amount of CuCl (ca. 25 mg) at 0 °C. After 35 min at room
temperature, the solid material was filtered off and was
washed with water. This provided 4-benzyloxy-2-nitrobenzene-
Meth yl 2(R)-[7-Meth oxy-2,3,4,5-tetr a h yd r oben zo[1,2,5-
f]th ia d ia zep in e-1,1-d ioxid e]p r op a n oa te (10). Ozone was
bubbled into a solution of 8 (2.70 g, 7.53 mmol) in CH2Cl2/
MeOH (1:1, 100 mL) at -78 °C until the reaction mixture
turned green. The reaction mixture was purged with N2 until
it was bright-yellow. Methyl sulfide (3.2 mL) was added, and
the reaction mixture was stirred at ambient temperature
overnight. The mixture was concentrated to yield the aldehyde
9 as yellow oil, which was used immediately. The aldehyde 9
was dissolved in HOAc (100 mL), and the mixture was cooled
with a water bath (15-20 °C) prior to the addition of Zn
powder (13 g) portionwise. The mixture was heated at reflux
for 3 h. After cooling to room temperature, the reaction mixture
was diluted with chloroform (50 mL). The mixture was filtered,
and the filter cake was washed with EtOH/CHCl3 (1:1, 200
mL). The filtrate was concentrated and purified by flash
chromatograph (40-60% ethyl acetate/hexane) to afford the
benzothiadiazepine 10 (0.42 g, 18% for two steps) as a light-
1
sulfonyl chloride 17 (1.5 g, 4.6 mmol, 75%). H NMR (CDCl3):
δ 5.23 (s, 2H), 2.6 (m, 1H), 7.39 (d, 1H), 7.42 (m, 5H), 8.14 (d,
1H). ESMS (neg), m/z: 307.8 (M - Cl + OH - H).
N-(4-Ben zyloxy-2-n itr oben zen esu lfon yl)-D-alan in e Meth -
yl Ester (18). D-Alanine methyl ester hydrochloride (5.7 g,
40.8 mmol) was dissolved in CH2Cl2 and diisopropylethylamine
(14.8 mL, 85.1 mmol). At 0 °C, 4-benzyloxy-2-nitrobenzene-
sulfonyl chloride 17 (11.1 g, 34.0 mmol) was added dropwise
in CH2Cl2. After being stirred overnight, the solution was
washed with 1 N HCl. The CH2Cl2 was dried, filtered, and
concentrated. Flash chromatography (10-40% ethyl acetate/
hexane) of the resulting residue gave the sulfonamide 18 (10.6
1
yellow solid. H NMR (CDCl3): δ 1.38 (d, 3H), 3.50-3.62 (m,
1
7H), 3.78 (s, 3H), 4.73 (q, 1H), 6.21 (d, 1H), 6.43 (dd, 1H), 7.68
g, 26.9 mmol, 66%). H NMR (CDCl3): δ 1.48 (d, 3H), 3.51 (s,
(d, 1H). MS (CI, NH3), m/z: 315 (M + H).
3H), 4.19 (m, 1H), 5.18 (s, 2H), 6.01 (br d, 1H), 7.19 (dd, 1H),
7.41 (m, 5H), 7.49 (d, 1H), 7.97 (d, 1H). ESMS (pos), m/z: 416.9
(M + Na).
N-Hyd r oxy-2(R)-[7-m eth oxy-2,3,4,5-tetr a h yd r oben zo-
[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]p r op a n a m id e (11). The
methyl ester 10 (0.37 g, 1.17 mmol) was incorporated into
method A, and preparative TLC purification (7.5% MeOH/
CHCl3) gave the hydroxamate 11 (90 mg, 24%). 1H NMR
(CD3OD): δ 1.33 (d, 3H), 3.34-3.42 (m, 1H), 3.59 (t, 2H), 3.67-
3.75 (m, 1H), 3.78 (s, 3H), 4.43 (q, 1H), 6.40-6.44 (m, 2H),
7.56 (d, 1H). ESMS (neg), m/z: 314.1 (M - H). HRMS calcd
for (C12H18N3O5S + H) 316.0967, found 316.0954. Anal.
(C12H17N3O5S‚0.7H2O) C, H, N, S.
N -(4-Be n zyloxy-2-n it r ob e n ze n e su lfon yl)-N -(b r om o-
eth ylen e)-D-a la n in e Meth yl Ester (19). The sulfonamide 18
(10.6 g, 26.9 mmol) was dissolved in THF prior to the addition
of triphenylphosphine (14.1 g, 53.8 mmol) and 2-bromoethanol
(3.8 mL, 53.8 mmol). At room temperature, diethyl azodicar-
boxylate (8.5 mL, 53.8 mmol) was added dropwise and the
reaction mixture was stirred overnight. The THF was removed,
and flash chromatography (10-40% ethyl acetate/hexane) of

1818 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10
the resulting residue gave the alkylated sulfonamide 19 (11.5
Cherney et al.
dried, and concentrated. Flash chromatography (20-60% ethyl
acetate/hexane) of the resulting residue afforded the alkylated
phenol 25 (85 mg, 0.20 mmol, 61%). H NMR (CDCl3): δ 1.38
1
g, 22.9 mmol, 85%). H NMR (CDCl3): δ 1.54 (d, 3H), 3.38-
1
3.58 (m, 3H), 3.61 (s, 3H), 3.62-3.78 (m, 1H), 4.2 (m, 1H), 4.75
(q, 1H), 5.16 (s, 2H), 7.16 (d, 1H), 7.2 (dd, 1H), 7.41 (m, 5H),
7.97 (d, 1H). ESMS (pos), m/z: 522.8 (M + Na).
(d, 3H), 2.32 (s, 6H), 3.44-4.56 (m, 2H), 3.54 (s, 3H), 3.62 (m,
2H), 4.3 (m, 1H), 4.73 (q, 1H), 4.97 (s, 2H), 6.25 (d, 1H), 6.51
(dd, 1H), 6.98 (m, 3H), 7.69 (d, 1H). ESMS (pos), m/z: 418.9
(M + H).
N-Hyd r oxy-2(R)-[7-(3,5-d im eth ylben zyloxy)-2,3,4,5-tet-
r a h yd r oben zo[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]p r op a n -
a m id e (26). The ester 25 was incorporated into method A,
and the resulting residue was recrystallized from benzene to
give the hydroxamate 26 (30 mg, 0.07 mmol) as a white
N-(4-Ben zyloxy-2-a m in ob en zen esu lfon yl)-N-(b r om o-
eth ylen e)-D-a la n in e Meth yl Ester (20). The sulfonamide 19
(11.5 g, 22.9 mmol) was dissolved in glacial AcOH and H2O
prior to the addition of Fe dust (12.8 g). This solution was
heated at 60 °C for 1 h. After the mixture was cooled to room
temperature, the Fe was filtered off and the AcOH was
removed. The resulting residue was dissolved in EtOAc and
washed with saturated NaHCO3 solution. The EtOAc was
dried, filtered, and concentrated to give the crude aniline 20
(8.9 g, 8.9 mmol, 39%), which was directly used in the next
procedure. 1H NMR (CDCl3): δ 1.43 (d, 3H), 3.38-3.5 (m, 2H),
3.54 (s, 3H), 3.57-3.77 (m, 2H), 4.2 (m, 1H), 4.62 (q, 1H), 5.01
(br s, 2H), 5.06 (s, 2H), 6.23 (d, 1H), 6.4 (dd, 1H), 7.39 (m, 5H),
7.54 (d, 1H). ESMS (neg), m/z: 470.8 (M - H).
Meth yl 2(R)-[7-Ben zyloxy-2,3,4,5-tetr ah ydr oben zo[1,2,5-
f]th ia d ia zep in e-1,1-d ioxid e]p r op a n oa te (21). The crude
aniline 20 (7.1 g, 15.1 mmol) was dissolved in DMF, and
N-methylmorpholine (5.1 mL, 47.1 mmol) was added. This
mixture was heated at 80 °C for 5 h. After it was cooled, the
DMF was removed and EtOAc was added. The EtOAc was
washed with brine, dried, and concentrated. Flash chroma-
tography (10-50% ethyl acetate/hexane) of the resulting
residue gave the cyclized product 21 (4.4 g, 11.3 mmol, 75%).
1H NMR (CDCl3): δ 1.38 (d, 3H), 3.42-3.6 (m, 2H), 3.53 (s,
3H), 3.62 (m, 2H), 4.31 (m, 1H), 4.73 (q, 1H), 5.05 (s, 2H), 6.26
(d, 1H), 6.51 (dd, 1H), 7.38 (m, 5H), 7.69 (d, 1H). ESMS (pos),
m/z: 413.0 (M + Na). Anal. (C19H22N2O5S) C, H, N, S.
1
powder. H NMR (300 MHz, DMSO): δ 1.13 (t, 3H), 2.24 (s,
6H), 3.2 (m, 1H), 3.4-3.65 (m, 3H), 4.22 (q, 1H), 4.93 (s, 2H),
6.4 (m, 2H), 6.54 (br s, 1H), 6.93 (s, 1H), 6.98 (s, 2H), 7.41 (d,
1H), 8.85 (s, 1H), 10.62 (s, 1H). ESMS (neg), m/z: 417.8 (M -
H). HRMS calcd for (C20H25N3O5S + H) 420.159 318, found
420.158 900. Anal. (C20H25N3O5S) C, H, N.
Compounds 27-43 were synthesized in fashion analogous
to that of compound 26.
N-Hyd r oxy-2(R)-[7-(3,5-d ich lor oben zyloxy)-2,3,4,5-tet-
r a h yd r oben zo[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]p r op a n -
a m id e (27). 1H NMR (DMSO): δ 1.14 (t, 3H), 3.17-3.3 (m,
2H), 3.35-3.7 (m, 2H), 4.22 (q, 1H), 5.05 (s, 2H), 6.4 (m, 2H),
6.58 (br s, 1H), 7.46 (m, 3H), 7.57 (m, 1H), 8.86 (br s, 1H),
10.64 (br s, 1H). ESMS (neg), m/z: 457.6 (M - H). HRMS calcd
for (C18H19N3O5SCl2 + H) 460.050 073, found 460.051 100.
Anal. (C18H19N3O5SCl) C, H, N.
N-H yd r oxy-2(R)-[7-(3,5-d im et h oxyb en zyloxy)-2,3,4,5-
t et r a h yd r oben zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r o-
p a n a m id e (28). 1H NMR (DMSO): δ 1.14 (d, 3H), 3.24 (m,
1H), 3.35-3.7 (m, 3H), 4.23 (q, 1H), 4.97 (s, 2H), 6.4 (m, 3H),
6.55 (m, 3H), 7.41 (d, 1H), 8.85 (s, 1H), 10.63 (s, 1H). ESMS
(neg), m/z: 449.7 (M - H). HRMS calcd for (C20H25N3O7S +
H) 452.149 147, found 452.148 200. Anal. (C20H25N3O7S‚
0.25C6H6) C, H, N.
N-Hyd r oxy-2(R)-[7-ben zyloxy-2,3,4,5-tetr a h yd r oben zo-
[1,2,5-f]th iadiazepin e-1,1-dioxide]pr opan am ide (22). Com-
pound 21 (0.64 mmol) was incorporated into method A and
recrystallized from benzene to give the hydroxamate 22 (160
1
mg, 0.41 mmol, 64%) as a white powder. H NMR (DMSO): δ
N-Hyd r oxy-2(R)-[7-(3,5-d ieth oxyben zyloxy)-2,3,4,5-tet-
r a h yd r oben zo[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]p r op a n -
a m id e (29). 1H NMR (DMSO): δ 1.12 (d, 3H), 1.27 (t, 6H),
3.2 (m, 1H), 3.4-3.65 (m, 3H), 3.96 (q, 4H), 4.21 (q, 1H), 4.94
(s, 2H), 6.38 (m, 3H), 6.5 (m, 3H), 7.4 (d, 1H), 8.85 (d, 1H),
10.62 (br s, 1H). ESMS (pos), m/z: 479.9 (M + H). HRMS calcd
for (C22H29N3O7S + H) 480.180 448, found 480.178 600. Anal.
(C22H29N3O7S‚0.2C6H6) C, H, N.
N-Hydr oxy-2(R)-[7-(2,6-m eth ylpyr idyl-4-m eth ylen eoxy)-
2,3,4,5-tetr ah ydr oben zo[1,2,5-f]th iadiazepin e-1,1-dioxide]-
p r op a n a m id e (30). 1H NMR (DMSO): δ 1.13 (d, 3H), 2.47
(s, 6H), 3.2-3.6 (m, 4H), 4.21 (q, 2H), 5.01 (s, 2H), 6.41 (m,
2H), 6.56 (s, 1H), 7.02 (s, 2H), 7.42 (d, 1H), 8.86 (s, 1H), 10.63
(s, 1H). ESMS (pos), m/z: 421.3 (M + H). HRMS calcd for
(C19H24N4O5S + H) 421.154 567, found 421.155 300.
N-Hyd r oxy-2(R)-[7-ben zyloxy-2,3,4,5-tetr a h yd r oben zo-
[1,2,5-f]t h ia d ia ze p in e -1,1-d io x id e ]-4-m e t h y lp e n t a n -
a m id e (32). The ester 21a (produced in analogy to 21 but
starting from d-Leu-OtBu and 17) (268 mg, 0.56 mmol) was
incorporated into method B, and reverse-phase HPLC gave the
hydroxamate 32 (90 mg, 0.2 mmol) as a white powder. 1H NMR
(DMSO): δ 0.75 (t, 6H), 1.3-1.5 (m, 3H), 3.3 (m, 1H), 3.48
(m, 3H), 4.2 (t, 1H), 5.05 (s, 2H), 6.36-6.5 (m, 3H), 7.3-7.44
(m, 6H), 10.73 (s, 1H). ESMS (neg), m/z: 431.9 (M - H). HRMS
calcd for (C21H27N3O5S + H) 434.174 968, found 434.174 735.
1.14 (t, 3H), 3.2 (m, 2H), 3.35-3.7 (m, 2H), 4.22 (q, 1H), 5.03
(s, 2H), 6.41 (m, 2H), 6.54 (br s, 1H), 7.28-7.44 (m, 6H), 8.85
(br s, 1H), 10.59 (br s, 1H). ESMS (pos), m/z: 413.0 (M + Na).
HRMS calcd for (C19H22N2O5S + H) 391.132 769, found
391.131 503. Anal. (C19H22N2O5S) C, H, N.
N-H yd r oxy-2(R)-[7-h yd r oxy-2,3,4,5-t et r a h yd r ob en zo-
[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]p r op a n a m id e (23). The
hydroxamate 22 (100 mg, 0.25 mmol) was dissolved in MeOH
(10 mL) prior to the addition of 5% Pd/BaSO4 (150 mg). The
solution was placed on a Parr apparatus at 35 psi. After 2.5
h, the solution was filtered and was concentrated to give the
hydroxamate 23 (59.9 mg, 0.2 mmol, 80%) as a white powder.
1H NMR (DMSO): δ 1.12 (t, 3H), 3.16 (m, 1H), 3.3-3.6 (m,
3H), 4.19 (q, 1H), 6.16 (d, 1H), 6.21 (s, 1H), 6.4 (br s, 1H), 7.3
(d, 1H), 8.85 (br s, 1H), 9.87 (s, 1H), 10.61 (s, 1H). ESMS (neg),
m/z: 299.7 (M - H). HRMS calcd for (C11H15N3O5S + H)
302.081 068, found 302.080 845. Anal. (C11H15N3O5S‚0.25C6H6)
C, H, N.
Meth yl 2(R)-[7-Hyd r oxy-2,3,4,5-tetr a h yd r oben zo[1,2,5-
f]th ia d ia zep in e-1,1-d ioxid e]p r op a n oa te (24). The benzo-
thiadiazepine 21 (2.4 g, 6.1 mmol) was dissolved in MeOH,
and 10% Pd/C (240 mg) was added. This mixture was hydro-
genated at 50 psi on a Parr for 4 h. The Pd/C was removed,
and the MeOH was concentrated to give the phenol 24 (2.0 g,
1
5.6 mmol, 92%). H NMR (CDCl3): δ 1.38 (d, 3H), 3.43-3.52
(m, 2H), 3.58 (s, 3H), 3.61 (m, 2H), 4.34 (m, 1H), 4.73 (q, 1H),
6.16 (d, 1H), 6.33 (dd, 1H), 7.64 (d, 1H). ESMS (neg), m/z:
299.7 (M - H).
N-H yd r oxy-2(R)-[7-h yd r oxy-2,3,4,5-t et r a h yd r ob en zo-
[1,2,5-f]th iadiazepin e-1,1-dioxide]-4-m eth ylpen tam ide (31).
The hydroxamate 32 (40 mg, 0.1 mmol) was dissolved in MeOH
(5 mL) prior to the addition of 5% Pd/BaSO4 (120 mg). The
solution was placed on a Parr apparatus at 55 psi. After 2 h,
the solution was filtered and was concentrated. Reverse-phase
HPLC of the resulting residue gave hydroxamate 31 (25.2 mg,
0.07 mmol) as a white powder. 1H NMR (DMSO): δ 0.74 (t,
6H), 1.3-1.5 (m, 3H), 3.25 (m, 1H), 3.37-3.5 (m, 3H), 4.19 (t,
1H), 6.15 (dd, 1H), 6.22 (d, 1H), 7.32 (d, 1H), 10.72 (br s, 1H).
ESMS (neg), m/z: 341.8 (M - H). HRMS calcd for (C14H21N3O5S
+ H) 344.128 018, found 344.125 763.
Meth yl 2(R)-[7-(3,5-Dim eth ylben zyloxy)-2,3,4,5-tetr ah y-
d r oben zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r op a n oa t e
(25). The phenol 24 (100 mg, 0.33 mmol) was dissolved in DMF
and cooled to 0 °C. At 0 °C, 1 M KOtBu (0.36 mL, 0.36 mmol)
in tBuOH was added dropwise. After 20 min, 3,5-dimethyl-
benzyl bromide (66.3 mg, 0.33 mmol) was added and the
reaction mixture was warmed to room temperature. After 15
h, the reaction was quenched with saturated NH4Cl solution
and EtOAc was added. The EtOAc was washed with brine,

Benzothiadiazepine Hydroxamates
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10 1819
N-H yd r oxy-2(R)-[7-(3,5-d im et h oxyb en zyloxy)-2,3,4,5-
t et r a h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]-4-
m eth ylp en ta n a m id e (33). H NMR (DMSO): δ 0.73 (t, 6H),
(br s, 3H), 4.96 (s, 2H), 4.46 (t, 1H), 3.76 (s, 6H), 3.69 (m, 1H),
3.58-3.24 (m, 3H), 3.38 (s, 3H), 2.93 (m, 2H), 1.82-1.57 (m,
4H), 1.37 (m, 2H). ESMS (pos), m/z: 508 (M + H).
1
1.3-1.5 (m, 3H), 3.1 (m, 1H), 3.5 (m, 3H), 3.7 (s, 6H), 4.2 (t,
1H), 4.97 (s, 2H), 6.37-6.54 (m, 6H), 7.42 (d, 1H), 8.84 (s, 1H),
10.73 (s, 1H). ESMS (neg), m/z: 491.7 (M - H). HRMS calcd
for (C23H31N3O7S + H) 494.196 098, found 494.196 400.
N-H yd r oxy-2(R)-[7-(3,5-d im et h oxyb en zyloxy)-2,3,4,5-
t et r a h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]-3-
m eth ylbu ta n a m id e (34). 1H NMR (DMSO): δ 0.76 (dd, 6H),
2.0 (m, 1H), 3.24 (m, 1H), 3.4 (m, 3H), 3.68 (m, 1H), 3.7 (s,
6H), 4.95 (s, 2H), 6.38 (m, 4H), 6.42 (d, 2H), 7.42 (d, 1H), 8.8
(s, 1H), 10.6 (s, 1H). ESMS (neg), m/z: 478.0 (M - H). HRMS
calcd for (C22H29N3O7S + H) 480.180 448, found 480.183 058.
N-Hyd r oxy-2(R)-[7-(3,5-d ieth oxyben zyloxy)-2,3,4,5-tet-
r a h yd r oben zo[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]-3-m eth -
Methanesulfonyl chloride (8 mg, 0.067 mmol) was added to
the previous compound (27 mg, 0.045 mmol) in THF/H2O (4
mL, 3:1) in the presence of lithium hydroxide (9.4 mg, 0.22
mmol) at room temperature. The reaction mixture was stirred
overnight, and then the solvent was removed. The residue was
dissolved in water (10 mL) and extracted with ethyl acetate
(3×). The organic layer was washed with brine, dried, and
filtered. The solvent was removed to provide the product 25c
as a brittle foam (22 mg, 86%). 1H NMR (CDCl3): δ 7.63 (d,
1H), 6.53 (d, 2H), 6.50 (d, 1H), 6.41 (t, 1H), 6.38 (s, 1H), 4.97
(s, 2H), 4.62 (m, 2H), 3.78 (s, 6H), 3.80-3.57 (m, 4H), 3.45 (m,
2H), 3.00 (m, 1H), 2.90 (s, 3H), 1.94 (m, 1H), 1.76 (m, 1H),
1.57-1.24 (m, 4H). ESMS (neg), m/z: 570 (M - H).
1
ylbu ta n a m id e (35). H NMR (DMSO): δ 0.68 (dd, 6H), 1.18
Compound 25c was converted to hydroxamic acid analogue
1
(t, 6H), 1.9 (m, 1H), 3.17 (m, 1H), 3.3 (m, 2H), 3.6 (m, 2H),
3.87 (q, 4H), 4.85 (s, 2H), 6.29 (m, 4H), 6.41 (d, 2H), 7.33 (d,
1H), 8.71 (s, 1H), 10.5 (br s, 1H). ESMS (neg), m/z: 505.8 (M
- H). HRMS calcd for (C24H33N3O7S + H) 508.211 748, found
508.213 100.
41 by method B. H NMR (DMSO): δ 10.69 (s, 1H), 7.41 (d,
1H), 6.86 (br t, 1H), 6.54 (s, 1H), 6.53 (s, 1H), 6.41 (m, 3H),
4.95 (s, 2H), 4.11 (t, 1H), 3.70 (s, 6H), 3.56-3.29 (m, 4H), 2.80
(s, 3H), 2.77 (m, 2H), 1.64-1.03 (m, 6H). ESMS (neg), m/z:
570 (M - H). HRMS calcd for (C24H34N4O9S2 + H) 587.1845,
found 587.1426.
N-Hydr oxy-2(R)-[7-(4,5-dim eth ylth iazolyl-2-m eth ylen e-
oxy)-2,3,4,5-t et r a h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-
d ioxid e]-3-m eth ylbu ta n a m id e (36). 1H NMR (DMSO): δ
0.76 (dd, 6H), 2.03 (m, 1H), 2.22 (s, 3H), 2.29 (s, 3H), 3.25 (m,
1H), 3.4 (m, 2H), 3.71 (m, 2H), 5.23 (s, 2H), 6.42 (m, 3H), 7.44
(d, 1H), 8.8 (s, 1H), 10.59 (s, 1H). ESMS (neg), m/z: 452.9 (M
- H). HRMS calcd for (C19H26N4O5S2 + H) 455.142 289, found
455.142 300.
N-H yd r oxy-2(R)-[7-(3,5-d im et h oxyb en zyloxy)-2,3,4,5-
t et r a h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]-4-
m eth oxyca r bon ylbu ta n a m id e (37). 1H NMR (DMSO): δ
1.75-1.9 (m, 2H), 2.1-2.25 (m, 2H), 3.29-3.45 (m, 4H), 3.49
(s, 3H), 3.7 (s, 6H), 4.96 (s, 2H), 6.42 (m, 4H), 6.54 (d, 2H),
7.42 (d, 1H), 8.88 (s, 1H), 10.66 (br s, 1H). ESMS (neg), m/z:
521.9 (M - H). HRMS calcd for (C23H29N3O9S + H) 524.170 277,
found 524.169 500.
N-Hyd r oxy-2(R)-[7-(3,5-d ieth oxyben zyloxy)-2,3,4,5-tet-
r a h yd r oben zo[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]-4-m eth -
oxyca r bon ylbu ta n a m id e (38). 1H NMR (DMSO): δ 1.26 (t,
6H), 1.75-1.9 (m, 2H), 2.1-2.25 (m, 2H), 3.28-3.49 (m, 4H),
3.49 (s, 3H), 3.96 (q, 4H), 4.15 (t, 1H), 4.94 (s, 2H), 6.36-6.5
(m, 6H), 7.42 (d, 1H), 8.87 (br s, 1H), 10.65 (br s, 1H). ESMS
(neg), m/z: 549.8 (M - H). HRMS calcd for (C25H33N3O9S +
H) 552.201 577, found 552.199 500.
N-H yd r oxy-2(R)-[7-(3,5-d im et h oxyb en zyloxy)-2,3,4,5-
tetr a h yd r oben zo[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]-6-N-
[(1,1-d im eth yleth oxy)ca r bon yl]h exyla m id e (42). 1H NMR
(DMSO): δ 10.66 (s, 1H), 7.41 (d, 1H), 6.69 (br t, 1H), 6.54 (s,
1H), 6.53 (s, 1H), 6.41 (m, 3H), 4.95 (s, 2H), 4.09 (t, 1H), 3.70
(s, 6H), 3.68-3.30 (m, 4H), 2.75 (m, 2H), 1.65-0.98 (m, 6H),
1.32 (s, 9H). ESMS (neg), m/z: 607 (M - H). HRMS calcd for
(C28H40N4O9S + H) 609.2594, found 609.2579.
N-H yd r oxy-2(R)-[7-(3,5-d im et h oxyb en zyloxy)-2,3,4,5-
tetr ah ydr oben zo[1,2,5-f]th iadiazepin e-1,1-dioxide]-6-am i-
n oh exyla m id et r iflu or oa cet ic Acid Sa lt (43). Trifluoro-
acetic acid (1 mL) was added to compound 42 (20 mg, 0.033
mmol) in methylene chloride (1 mL) at room temperature.
After 20 min, the solvent was removed. The resulting residue
was azeotroped with chloroform three times. The final residue
was taken up in water (2 mL) and lyophilized to afford
compound 43 as a white powder (17 mg, 83%). 1H NMR
(DMSO): δ 10.70 (s, 1H), 8.88 (s, 1H), 7.57 (br s, 3H), 7.43 (d,
1H), 6.54 (s, 1H), 6.53 (s, 1H), 6,43 (m, 3H), 4.96 (s, 2H), 4.12
(t, 1H), 3.70 (s, 6H), 3.52-3.26 (m, 4H), 2.65 (m, 2H), 2.47 (m,
1H), 1,65-1.04 (m, 6H). ESMS (pos), m/z: 509 (M + H). HRMS
calcd for (C23H32N4O7S + H) 509.2070, found 509.2066. Anal.
(C25H33F3N4O9S‚2TFA) C, H, N.
N-H yd r oxy-2(R)-[7-(3,5-d im et h oxyb en zyloxy)-2,3,4,5-
t et r a h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]-4-
m eth oxyca r bon ylp en ta n a m id e (39). H NMR (DMSO): δ
Meth yl N-[(2,4-Din itr oph en yl)su lfon yl]-D-alan in ate (45).
D-Alanine methyl ester hydrochloride salt (10 g, 71.6 mmol)
was dissolved in CH2Cl2 (100 mL) and washed with 10%
sodium carbonate (2 × 40 mL). After the mixture was dried
over MgSO4, the solvent was removed to provide the free base
amino acid. N-Methylmorpholine (14.5 g, 143 mmol) was added
to d-Ala-OMe in dioxane (75 mL) followed by dropwise addition
of 2,4-dinitrobenzenesulfonyl chloride (19.1 g, 71.6 mmol) in
dioxane (50 mL). This mixture was stirred for 24 h at room
temperature and then was evaporated. The crude product was
dissolved in ethyl acetate (100 mL) and was washed with
water, 10% citric acid, saturated NaHCO3 (2×), and brine.
After the mixture was dried over MgSO4, the solvent was
removed and the crude product was purified by recrystalliza-
tion (ethyl acetate/hexane) to afford 45 (5.81 g, 24%) as a light-
yellow crystalline solid. 1H NMR (CDCl3): δ 8.75 (d, 1H), 8.29
(dd, 1H), 8.29 (d, 1H), 6.14 (d, 1H), 4.32 (m, 1H), 3.59 (s, 1H),
1.53 (d, 3H). ESMS (neg), m/z: 332 (M - H). HRMS calcd for
1
1.25-1.45 (m, 2H), 1.5-1.68 (m, 2H), 2.21 (t, 2H), 3.26-3.6
(m, 4H), 3.49 (s, 3H), 3.7 (s, 6H), 4.11 (t, 1H), 4.95 (s, 2H),
6.38-6.42 (m, 3H), 6.53 (d, 2H), 7.41 (d, 1H), 10.65 (br s, 1H).
ESMS (pos), m/z: 538.2 (M + H). HRMS calcd for (C24H31N3O9S
+ H) 538.185 927, found 538.185 700. Anal. (C24H31N3O9S‚
0.4TFA) C, H, N.
N-Hyd r oxy-2(R)-[7-(3,5-d ieth oxyben zyloxy)-2,3,4,5-tet-
r a h yd r oben zo[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]-4 m eth -
oxyca r bon ylp en ta n a m id e (40). 1H NMR (DMSO): δ 1.26
(t, 6H), 1.3-1.45 (m, 2H), 1.45-1.68 (m, 2H), 2.21 (t, 2H),
3.25-3.5 (m, 4H), 3.49 (s, 3H), 3.95 (q, 4H), 4.11 (t, 1H), 4.94
(s, 2H), 6.36-6.42 (m, 3H), 6.49 (d, 2H), 7.41 (d, 1H), 10.67
(br s, 1H). ESMS (pos), m/z: 566.2 (M + H). HRMS calcd for
(C26H35N3O9S + H) 566.217 227, found 566.217 500. Anal.
(C26H35N3O9S‚H2O) C, H, N.
(C10H11N3O8S + H) 334.0345, found 334.0349. Anal. (C10H11
-
N-H yd r oxy-2(R)-[7-(3,5-d im et h oxyb en zyloxy)-2,3,4,5-
tetr a h yd r oben zo[1,2,5-f]th ia d ia zep in e 1,1-d ioxid e]-6-N-
(m eth a n esu lfon yl)h exyla m id e (41). Trifluoroacetic acid (1
mL) was added to compound 25b (produced in analogy to 25
but starting from D-Lys(Boc)-OMe and 17) (72 mg, 0.12 mmol)
in methylene chloride (1 mL) at room temperature. After 20
min, the solvent was removed. This was azeotroped with
chloroform three times, and the resulting residue was taken
N3O8S) C, H, N.
Meth yl N-(2-Br om oeth yl)-N-[(2,4-din itr oph en yl)su lfon -
yl]-D-a la n in a te (46). Diethyl azadicarboxylate (16.3 g, 93.6
mmol) was added dropwise at room temperature to 45 (20.8
g, 62.4 mmol), triphenylphosphine (25.2 g, 93.6 mmol), and
2-bromoethanol (11.7 g, 93.6 mmol) in THF (225 mL). After
the mixture was stirred for 24 h, the solvent was removed and
the residue was purified by flash chromatography (20-50%
ethyl acetate/hexane) to provide 46 (25.9 g, 94%) as a viscous
1
forward without additional purification. H NMR (CDCl3): δ
7.59 (d, 1H), 7.58 (br s, 2H), 6.51 (m, 3H), 6.39 (m, 2H), 6.29

1820 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10
Cherney et al.
1
oil. H NMR (CDCl3): δ 8.53 (dd, 1H), 8.43 (d, 1 H), 8.30 (d,
4.73 (d, 1H), 4.61 (d, 1H), 4.40 (q, 1H), 3.52 (m, 4H), 2.34 (s,
6H), 1.31 (d, 3H). ESMS (pos), m/z: 545 (M + Na). HRMS calcd
for (C27H30N4O5S + H) 523.2015, found 523.2013.
1H), 4.81 (q, 1H), 3.82 (m, 1H), 3.68 (m, 1H), 3.65 (s, 3H), 3.43
(m, 2H), 1.60 (d, 3H). ESMS (pos), m/z: 440 (M - H).
Meth yl (2R)-2-(7-Am in o-1,1-d ioxid o-4,5-d ih yd r o-1,2,5-
ben zoth ia d ia zep in -2(3H)-yl)p r op a n oa te (47). Iron dust
(7.65 g, 137 mmol) was added in one portion to 46 (6.03 g,
13.7 mmol) in acetic acid/water (85 mL, 16:1). This solution
was heated at 60 °C for 1.5 h. The reaction mixture was diluted
with ethyl acetate (500 mL), and the solids were filtered. With
stirring, saturated NaHCO3 (200 mL) was added to the ethyl
acetate solution followed by addition of solid NaHCO3 until
no gas evolution was noted. The aqueous solution was ex-
tracted with ethyl acetate. The ethyl acetate layer was washed
with water, saturated NaHCO3, and brine. After it was dried
over MgSO4, the crude product was concentrated [ESMS (pos),
m/z: 404 (M + Na)]. The resulting residue was dissolved in
DMF (250 mL) prior to the addition of N-methylmorpholine
(1.71 g, 16.9 mmol). This solution was heated at 80 °C for 5.5
h. The solvent was removed, and the residue was dissolved in
ethyl acetate. The ethyl acetate was washed with water,
saturated NaHCO3, and brine. After drying over MgSO4 the
crude product was concentrated. The resulting residue was
purified by flash chromatography (30-70% ethyl acetate/
N -H yd r oxy-2(R )-[7-(N -3,5-d im e t h ylb e n zoyla m in o)-
2,3,4,5-tetr ah ydr oben zo[1,2,5-f]th iadiazepin e-1,1-dioxide]-
p r op yla m id e (51). A sample of 5% palladium on barium
sulfate (unreduced, 50 mg) was added to 50 (79 mg, 0.15 mmol)
in methanol (10 mL). A balloon filled with hydrogen was
attached via a three-way stopcock, and the atmosphere above
the reaction mixture was removed and replaced with hydrogen
three times. After 0.45 h, the catalyst was filtered off and the
filtrate was concentrated. The crude product was dissolved in
THF (1 mL) and was crystallized by slow addition of ether (30
mL). The hydroscopic solid was filtered and dried under
vacuum to provide compound 51 (33 mg, 50%). 1H NMR
(DMSO): δ 10.65 (s, 1H), 10.21 (s, 1H), 8.87 (s, 1H), 7.49 (m,
4H), 7.19 (s, 1H), 7.03 (dd, 1H), 6.64 (br s, 1H), 4.24 (q, 1H),
3.62-3.18 (m, 4H), 2.32 (s, 6H), 1.15 (d, 3H). ESMS (pos),
m/z: 455 (M + Na). HRMS calcd for (C20H24N4O5S + H)
433.1546, found 433.1548.
ter t-Bu tyl (2R)-2-[7-[(3,5-Dim eth oxyben zyl)oxy]-1,1-d i-
oxid o-4,5-d ih yd r o-1,2,5-b en zot h ia d ia zep in -2(3H )-yl]-4-
m eth ylp en ta n oa te (48). Ethanol (300 µL) was added to
compound 47a (produced in analogy to 47 but starting from
D-Leu-OtBu and 2,4-dinitrobenzenesulfonyl chloride) (117 mg,
0.31 mmol) and 3,5-dimethoxybenzaldehyde (51 mg, 0.31
mmol). After the mixture was stirred for 5 h at room temper-
ature, sodium borohydride (12 mg, 0.31 mmol) was added in
one portion and the viscous reaction mixture was stirred
overnight at room temperature. The reaction was quenched
with saturated ammonium chloride (3 mL) prior to the addition
of water (10 mL). The aqueous solution was extracted with
ethyl acetate. The organic layer was washed with water,
saturated NaHCO3, and brine. After the mixture was dried
over MgSO4, the solvent was evaporated and the crude product
was purified by flash chromatography (20-70% ethyl acetate/
hexane) to provide 48 (104 mg, 64%). 1H NMR (CDCl3): δ 7.54
(d, 1H), 6.45 (s, 2H), 6.35 (m, 1H), 6.12 (dd, 1H), 5.82 (d, 1H),
4.56 (m, 1H), 4.24 (s, 2H), 3.76 (s, 6H), 3.55 (m, 1H), 3.51 (m,
3H), 3.36 (m, 1H), 1.59 (m, 3H), 1.23 (s, 9H), 0.90 (d, 6H).
ESMS (neg), m/z: 532 (M - H). HRMS calcd for (C27H39N3O6S
+ Na) 556.2457, found 556.2471.
1
hexane) to provide 47 (1.76 g, 43%) as a white solid. H NMR
(CDCl3): δ 7.13 (d, 1H), 6.08 (br t, 1H), 5.94 (dd, 1H), 5.90 (d,
1H), 5.54 (s, 2H), 4.41 (q, 1H), 3.51 (s, 3H), 3.45-3.17 (m, 4H),
1.19 (d, 3H). ESMS (neg), m/z: 524 (M - H). HRMS calcd for
(C12H17N3O4S + H) 300.1018, found 300.1028. Anal. (C12H17
N3O4S) C, H, N.
-
N-(2-{(1R)-2-[(Ben zyloxy)a m in o]-1-m eth yl-2-oxoeth yl}-
1,1-d ioxid o-2,3,4,5-tetr a h yd r o-1,2,5-ben zoth ia d ia zep in -7-
yl)-3,5-d im eth ylben za m id e (50). Lithium hydroxide mono-
hydrate (0.74 g, 17.6 mmol) in water (10 mL) was added to 47
(1.76 g, 5.88 mmol) in tetrahydrofuran (25 mL) at room
temperature. After 1 h, the solvent was removed. The residue
was dissolved in water (30 mL) and was washed with ether
(2×). The ether layer was discarded, and the aqueous layer
was acidified with 1 N HCl. The aqueous layer was then ex-
tracted with ethyl acetate. The organic layer was washed with
brine, dried over MgSO4, and evaporated to provide the
carboxylate intermediate as a brittle foam. 1H NMR (CD3OD):
δ 7.33 (d, 1H), 6.11 (dd, 1H), 6.05 (d, 1H), 4.54 (q, 1H), 3.59
(m, 2H), 3.45 (m, 1H), 3.23 (m, 1H), 1.31 (d, 3H). ESMS (neg),
m/z: 284 (M - H). HRMS calcd for (C11H15N3O4S +H)
286.0862, found 286.0847.
N-Hydr oxy-2(R)-[7-(N-(3,5-dim eth oxym eth ylen eam in o)-
2,3,4,5-tetr ah ydr oben zo[1,2,5-f]th iadiazepin e-1,1-dioxide]-
4-m eth ylp en ta m id e (49). Compound 48 (91 mg, 0.17 mmol)
was dissolved in trifluoroacetic acid (3 mL). After 1.5 h, the
trifluoroacetic acid was removed. The crude product was
azeotroped with chloroform (5 mL) three times. Drying over-
night under vacuum provided the acid intermediate as a brittle
foam that was carried forward without further purification.
1H NMR (CDCl3): δ 7.68 (br s), 7.56 (d, 1H), 6.55 (s, 1H), 6.52-
6.35 (m, 4H), 4.67 (m, 1H), 4.22 (s, 2H), 3.81 (s, 3H), 3.80-
3.55 (m, 4H), 1.65 (m, 3H), 0.93 (d, 6H). ESMS (neg), m/z: 476
(M - H). HRMS calcd for (C23H31N3O6S + H) 478.2012, found
478.2014. Diisopropylethylamine (110 mg, 0.85 mmol) was
added dropwise at 0 °C to the compound from the previous
reaction (0.17 mmol), BOP reagent (83 mg, 0.19 mmol), and
benzylhydroxylamine hydrochloride (54 mg, 0.34 mmol) in
DMF (2 mL). The ice bath was removed after 0.5 h, and the
reaction mixture was stirred overnight at room temperature.
The solvent was removed, and the residue was dissolved in
ethyl acetate. The ethyl acetate was washed with water,
saturated NaHCO3, and brine. After the mixture was dried
over MgSO4, the crude product was purified by flash chro-
matography (1-10% MeOH/CH2Cl2) to provide the pro-
Diisopropylethylamine (3.51 g, 27.2 mmol) was added drop-
wise at 0 °C to the compound from the previous reaction (1.55
g, 5.43 mmol), BOP reagent (2.64 g, 5.98 mmol), and benzyl-
hydroxylamine hydrochloride (1.30 g, 8.15 mmol) in DMF (25
mL). The ice bath was removed after 0.5 h, and the reaction
mixture was stirred overnight at room temperature. The
solvent was removed. The residue was dissolved in ethyl
acetate (100 mL) and was washed with water, saturated
NaHCO3, and brine. After drying over MgSO4, the crude
product was purified by flash chromatography (1-10% MeOH/
CH2Cl2) to provide the protected hydroxamic acid intermediate
1
(1.94 g, 91%) as a white solid. H NMR (CD3OD): δ 7.36 (d,
1H), 7.32 (m, 5H), 6.16 (dd, 1H), 6.05 (d, 1H), 4.73 (d, 1H),
4.63 (d, 1H), 4.36 (q, 1H), 3.47 (m, 3H), 3.34 (m, 1H), 1.29 (d,
3H). ESMS (pos), m/z: 391 (M + H). HRMS calcd for
(C18H22N4O4S + H) 391.1440, found 391.1440. Anal. (C18H22
N4O4S) C, H, N.
-
Diisopropylethylamine (170 mg, 1.32 mmol) was added
dropwise at 0 °C to the compound from the previous reaction
(103 mg, 0.26 mmol), BOP reagent (140 mg, 0.32 mmol), and
3,5-dimethylbenzoic acid (48 mg, 0.32 mmol) in DMF (2 mL).
The ice bath was removed after 0.5 h, and the reaction mixture
was stirred overnight at room temperature. The solvent was
removed, and the residue was dissolved in ethyl acetate. The
ethyl acetate was washed with water, 10% citric acid, satu-
rated NaHCO3, and brine. After drying over MgSO4, the crude
product was purified by flash chromatography (1-10% MeOH/
CH2Cl2) to provide 50 (79 mg, 58%). 1H NMR (CD3OD): δ 7.60
(d, 1H), 7.47 (m, 2H), 7.30 (m, 5H), 7.20 (m, 1H), 7.07 (m, 2H),
1
tected hydroxamic acid intermediate (74 mg, 75%). H NMR
(CD3OD): δ 10.05 (s, 1H), 7.47 (d, 1H), 7.31 (m, 5H), 6.47 (d,
2H), 6.39 (t, 1H), 6.18 (dd, 1H), 5.61 (d, 1H), 4.80 (d, 1H), 4.69
(d, 1H), 4.62 (dd, 1H), 4.49 (br t, 1H), 4.25 (d, 2H), 3.77 (s,
6H), 3.68-3.50 (m, 2H), 3.25 (m, 1H), 3.10 (m, 2H), 1.90 (m,
1H), 1.67 (m, 2H), 0.98 (d, 3H), 0.93 (d, 3H). ESMS (pos), m/z:
583 (M + H). HRMS calcd for (C30H38N4O6S + H) 583.2590,
found 583.2584.

Benzothiadiazepine Hydroxamates
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10 1821
A sample of 5% palladium on barium sulfate (unreduced,
50 mg) was added to the compound from the previous reaction
(74 mg, 0.13 mmol) in methanol (5 mL). A balloon filled with
hydrogen was attached via a three-way stopcock, and the
atmosphere above the reaction mixture was removed and
replaced with hydrogen three times. After 0.45 h, the catalyst
was filtered off and the filtrate was concentrated. The crude
product was dissolved in THF (1 mL) and was crystallized by
slow addition of ether (30 mL). The hydroscopic solid was
filtered and dried under vacuum to provide analogue 49 (17
mg, 27%). 1H NMR (CD3OD): δ 7.37 (d, 1H), 6.48 (d, 2H), 6.31
(t, 1H), 6.14 (dd, 1H), 5.99 (d, 1H), 4.39 (m, 1H), 4.24 (s, 2H),
3.71 (s, 6H), 3.53-3.25 (m, 4H), 1.58 (m, 3H), 0.86 (m, 6H).
ESMS (pos), m/z: 493 (M + H). HRMS calcd for (C23H32N4O6S
+ H) 493.2121, found 493.2134. Anal. (C23H32N4O6S‚1.2H2O)
C, H, N.
and concentrated. Flash chromatography (20-60% ethyl acetate/
hexane) of the resulting residue gave the ester 57 (125 mg,
0.28 mmol, 56%) as a white foam. 1H NMR (CDCl3): δ 1.41
(d, 3H), 3.45-3.6 (m, 2H), 3.61 (s, 3H), 3.62-3.71 (m, 2H), 4.51
(m, 1H), 4.75 (q, 1H), 6.34 (d, 1H), 6.49 (m, 1H), 7.1 (d, 2H),
7.62 (d, 2H), 7.74 (d, 1H). ESMS (pos), m/z: 445.1 (M + H).
N-Hyd r oxy-2(R)-[7-(4-tr iflu or om eth ylp h en oxy)-2,3,4,5-
t et r a h yd r oben zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r o-
p a n a m id e (58). The ester 57 (110 mg, 0.25 mmol) was
incorporated into method A, and flash chromatography (1-
10% MeOH/CH2Cl2) gave the hydroxamate 58 (32 mg, 0.07
1
mmol, 28%) as a white powder. H NMR (DMSO): δ 1.15 (d,
3H), 3.25 (m, 1H), 3.43-3.7 (m, 3H), 4.24 (q, 1H), 6.43 (m, 2H),
6.72 (m, 1H), 7.23 (d, 2H), 7.53 (d, 1H), 7.75 (d, 2H), 8.87 (s,
1H), 10.64 (s, 1H). ESMS (neg), m/z: 444.1 (M - H). HRMS
calcd for (C18H18F3N3O5S + H) 446.099 753, found 446.099 100.
Anal. (C18H18F3N3O5S) C, H, N.
Meth yl (2R)-2-[1,1-Dioxid o-7-{[(tr iflu or om eth yl)su lfon -
yl]oxy}-4,5-dih ydr o-1,2,5-ben zoth iadiazepin -2(3H)-yl]pr o-
p a n oa t e (53). Trifluoromethanesulfonic anhydride (48 mg,
0.17 mmol) was added to 24 (51 mg, 0.17 mmol) and DIEA
(66 mg, 0.51 mmol) in methylene chloride (5 mL) at -78 °C
under nitrogen. The reaction mixture was stirred for 2.5 h,
and the reaction was quenched at -78 °C by the addition of
saturated ammonium chloride. After warming to room tem-
perature, the solution was extracted with methylene chloride.
The organic layer was washed with brine and then was dried
over MgSO4. After filtration, the solvent was removed and the
residue was purified by flash chromatography (10-50% ethyl
Compounds 59-62 were synthesized in a fashion analogous
to that of compound 58.
N-Hyd r oxy-2(R)-[7-(4-m eth oxyp h en oxy)-2,3,4,5-tetr a -
h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r op a n -
a m id e (59). 1H NMR (DMSO): δ 1.13 (d, 3H), 3.2 (m, 1H),
3.35-3.65 (m, 3H), 3.72 (s, 3H), 4.22 (q, 1H), 6.27 (m, 2H), 6.6
(m, 1H), 6.95 (d, 2H), 7.02 (d, 2H), 7.44 (d, 2H), 8.86 (s, 1H),
10.63 (s, 1H). ESMS (neg), m/z: 406.1 (M - H).
N-Hyd r oxy-2(R)-[7-p h en oxy-2,3,4,5-tetr a h yd r oben zo-
1
[1,2,5-f]th ia d ia zep in e-1,1-d ioxid e]p r op a n a m id e (60). H
NMR (DMSO): δ 1.14 (d, 1H), 3.2 (m, 1H), 3.38-3.7 (m, 3H),
4.22 (q, 1H), 6.31 (m, 2H), 6.65 (m, 1H), 7.05 (d, 2H), 7.18 (t,
1H), 7.42 (m, 3H), 8.86 (d, 1H), 10.62 (s, 1H). ESMS (neg),
m/z: 376.1 (M - H). HRMS calcd for (C17H19N3O5S + H)
378.112 368, found 378.113 500.
1
acetate/hexane) to afford 53 (47 mg, 64%). H NMR (CDCl3):
δ 7.83 (d, 1H), 6.74 (dd, 1H), 6.64 (d, 1H), 4.73 (m, 2H), 3.85
(m, 1H), 3.71 (m, 1H), 3.62-3.48 (m, 2H), 3.54 (s, 3H), 1.42
(d, 3H). ESMS (pos), m/z: 433 (M + H).
Meth yl (2R)-2-(1,1-Dioxid o-7-p h en yl-4,5-d ih yd r o-1,2,5-
ben zoth ia d ia zep in -2(3H)-yl)p r op a n oa te (54). Palladium
acetate (5 mg, 0.022 mmol) was added to 53 (47 mg, 0.11
mmol), phenylboronic acid (27 mg, 0.22 mmol), triphenylphos-
phine (28 mg, 0.11 mmol), and potassium carbonate (60 mg,
0.43 mmol) in degassed toluene (10 mL) under nitrogen. The
reaction mixture was heated to reflux for 45 min. After the
mixture was cooled, water (10 mL) was added and the mixture
was extracted with ethyl acetate. The organic layer was
washed with water, 10% citric acid, saturated NaHCO3, and
brine. The organic layer was dried over MgSO4 and was
filtered. The solvent was removed, and the residue was purified
by flash chromatography (20-60% ethyl acetate/hexane) to
afford 54 (25 mg, 64%). 1H NMR (CDCl3): δ 8.25 (dd, 1H), 7.82
(d, 1H), 7.63-7.35 (m, 4H), 7.10 (dd, 1H), 6.91 (d, 1H), 4.77
(q, 1H), 4.50 (bs, 1H), 3.68-3.43 (m, 4H), 3.56 (s, 3H), 1.41 (d,
3H, 7.4). ESMS (pos), m/z: 361 (M + H).
N-Hydr oxy-2(R)-[7-ph en yl-2,3,4,5-tetr ah ydr oben zo[1,2,5-
f]th ia d ia zep in e-1,1-d ioxid e]p r op yla m id e (55). Compound
54 was converted to hydroxamic acid analogue 55 via method
A. 1H NMR (DMSO): δ 10.66 (s, 1H), 7.58-7.34 (m, 6H), 7.13
(d, 1H),7.02 (dd, 1H), 4.27 (q, 1H), 3.70-3.35 (m, 4H), 1.17 (d,
3H). ESMS (pos), m/z: 362 (M + H). HRMS calcd for
(C17H19N3O4S + H) 362.1174, found 362.1180.
N-H yd r oxy-2(R)-[7-(4-p h en ylp h en oxy)-2,3,4,5-t et r a -
h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r op a n -
a m id e (61). H NMR (DMSO): δ 1.15 (d, 3H), 3.22 (m, 1H),
3.4-3.7 (m, 3H), 4.25 (q, 1H), 6.39 (m, 2H), 6.68 (m, 1H), 7.15
(d, 2H), 7.35 (t, 1H), 7.4-7.54 (m, 3H), 7.63 (d, 2H), 7.71 (d,
2H), 8.86 (s, 1H), 10.64 (s, 1H). ESMS (neg), m/z: 452.1 (M -
H). HRMS calcd for (C23H23N3O5S + H) 454.143 668, found
454.144 000.
1
N -H yd r ox y -2(R )-[7-(3-n it r o p h e n ox y)-2,3,4,5-t e t r a -
h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r op a n -
a m id e (62). 1H NMR (DMSO): δ 1.16 (d, 3H), 3.2 (m, 1H),
3.38-3.74 (m, 3H), 4.24 (q, 1H), 6.37-6.48 (m, 2H), 6.71 (m,
1H), 7.54 (m, 2H), 7.69 (t, 1H), 7.83 (t, 1H), 8.04 (dd, 1H), 8.87
(s, 1H), 10.64 (s, 1H). ESMS (neg), m/z: 421.1 (M - H). HRMS
calcd for (C17H18N4O7S + H) 423.097 446, found 423.095 600.
Met h yl 2(R)-[7-(4-Nit r op h en oxy)-2,3,4,5-t et r a h yd r o-
ben zo[1,2,5-f]th iadiazepin e-1,1-dioxide]pr opan oate (57a).
The phenol 24 (200 mg, 0.67 mmol) was dissolved in DMF (5
mL) prior to the addition of 1-fluoro-4-nitrobenzene (0.08 mL,
0.74 mmol) and Cs2CO3 (391 mg, 1.2 mmol). After 2.5 h, the
solution was quenched with aqueous NH4Cl and extracted with
EtOAc. The organic layer was dried, filtered, and concentrated.
Flash chromatography (20-60% ethyl acetate/hexane) of the
resulting residue gave 57a (170 mg, 0.4 mmol, 60%). 1H NMR
(DMSO): δ 1.43 (d, 3H), 4.6-4.6 (m, 2H), 3.62 (s, 3H), 3.72
(m, 2H), 4.46 (m, 1H), 4.77 (q, 1H), 6.4 (d, 1H), 6.54 (dd, 1H),
7.08 (d, 2H), 7.8 (d, 1H), 8.24 (d, 2H). ESMS (neg), m/z: 420.1
(M - 1).
N -H yd r ox y -2(R )-[7-(4-n it r o p h e n ox y)-2,3,4,5-t e t r a -
h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r op a n -
a m id e (63). Ester 57a (165 mg, 0.39 mmol) was incorporated
into method A, and flash chromatography (1-10% MeOH/
CH2Cl2) gave the hydroxamate 63 (120 mg, 0.28 mmol, 72%)
as a white solid. 1H NMR (DMSO): δ 1.17 (d, 3H), 3.3 (m, 1H),
3.4-3.7 (m, 3H), 4.25 (q, 1H), 6.47 (dd, 1H), 6.54 (d, 1H), 6.78
(m, 1H), 7.22 (d, 2H), 7.55 (d, 1H), 8.24 (d, 2H), 8.87 (s, 1H),
10.64 (s, 1H). ESMS (pos), m/z: 423.1 (M + H). HRMS calcd
for (C17H18N4O7S + H) 423.097 446, found 423.098 600.
P r otein Bin d in g in Hu m a n Ser u m . The protein binding
was determined by equilibrium dialysis using the Dianorm
system (Munchen, Germany). This device consists of Teflon
cells and Diachema cellulose based dialysis membrane with a
N-H yd r oxy-2(R)-[7-(2-t r iflu r om et h ylp h en yl)-2,3,4,5-
t et r a h yd r oben zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r o-
p yla m id e (56). Compound 56 was synthesized in a fashion
analogous to that of compound 55. 1H NMR (DMSO): δ 10.67
(s, 1H), 7.79 (d, 1H), 7.70-7.59 (m, 2H), 7.52 (d, 1H), 7.36 (d,
1H), 6.77 (s, 1H), 6.64 (d, 1H), 4.28 (q, 1H), 3.78-3.35 (m, 4H),
1.16 (d, 3H). ESMS (pos), m/z: 430 (M + H). HRMS calcd for
(C18H18F3N4O4S + H) 430.1048, found 430.1063. Anal. (C18H18
F3N3O4S‚0.7TFA) C, H, N.
-
Meth yl 2(R)-[7-(4-(Tr iflu or om eth yl)p h en oxy)-2,3,4,5-
t et r a h yd r ob en zo[1,2,5-f]t h ia d ia zep in e-1,1-d ioxid e]p r o-
p a n oa te (57). The phenol 24 (150 mg, 0.5 mmol) was dissolved
in CH2Cl2 (6 mL) prior to the addition of Cu(OAc)2 (100 mg,
0.55 mmol), 4-(trifluoromethyl)benzeneboronic acid (190 mg,
1.0 mmol), powdered 4 Å molecular sieves (353 mg), and
triethylamine (0.35 mL, 2.5 mmol). After being stirred for 18
h in the open air, the solution was filtered and was washed
with 1 N HCl and 1 N NaOH. The organic layer was dried

1822 J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 10
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Ack n ow led gm en t . The authors thank Dianna L.
Blessington, J ohn V. Giannaras, Sherrill A. Nurnberg,
and Paul J . Strzemienski for in vitro and in vivo results
as well as Li Liang and Bing Lu for protein binding
studies. The authors also thank Drs. Percy H. Carter
and J ames D. Rodgers for a critical review of the
manuscript.
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