Design, synthesis, and structure-activity relationships of macrocyclic hydroxamic acids that inhibit tumor necrosis factor α release in vitro and in vivo

Article abstract of DOI:10.1021/jm010127e

To search for TNF-α (tumor necrosis factor α) converting enzyme (TACE) inhibitors, we designed a new class of macrocyclic hydroxamic acids by linking the P1 and P2′ residues of acyclic anti-succinate-based hydroxamic acids. A variety of residues including amide, carbamate, alkyl, sulfonamido, Boc-amino, and amino were found to be suitable P1 P1-P2′ linkers. With an N-methylamide at P3′, the 13-16-membered macrocycles prepared exhibited low micromolar activities in the inhibition of TNF-α release from LPS-stimulated human whole blood. Further elaboration in the P3′-P4′ area using the cyclophane and cyclic carbamate templates led to the identification of a number of potent analogues with IC₅₀ values of ≤0.2 μM in whole blood assay (WBA). Although the P3′ area can accommodate a broad array of structurally diversified functional groups including polar residues, hydrophobic residues, and amino and carboxylic acid moieties, in both the cyclophane series and the cyclic carbamate series, a glycine residue at P3′ was identified as a critical structural component to achieve both good in vitro potency and good oral activity. With a glycine residue at P3′, an N-methylamide at P4′ provided the best cyclophane analogue, SL422 (WBA IC₅₀ = 0.22 μM, LPS-mouse ED₅₀ = 15 mg/kg, po), whereas a morpholinylamide at P4′ afforded the most potent and most orally active cyclic carbamate analogue, SP057 (WBAIC₅₀ = 0.067 μM, LPS-mouse ED₅₀ = 2.3 mg/kg, po). Further profiling for SL422 and SP057 showed that these macrocyclic compounds are potent TACE inhibitors, with K_i values of 12 and 4.2 nM in the porcine TACE assay, and are broad-spectrum MMP inhibitors. Pharmacokinetic studies in beagle dogs revealed that SL422 and SP057 are orally bioavailable, with oral bioavailabilities of 11% and 23%, respectively.

Full text of DOI:10.1021/jm010127e

2636
J . Med. Chem. 2001, 44, 2636-2660
Design , Syn th esis, a n d Str u ctu r e-Activity Rela tion sh ip s of Ma cr ocyclic
Hyd r oxa m ic Acid s Th a t In h ibit Tu m or Necr osis F a ctor r Relea se in Vitr o a n d
in Vivo
Chu-Biao Xue,* Matthew E. Voss, David J . Nelson, J ames J .-W. Duan, Robert J . Cherney, Irina C. J acobson,
Xiaohua He, J ohn Roderick, Lihua Chen, Ronald L. Corbett, Li Wang, Dayton T. Meyer, Kenneth Kennedy,
William F. DeGrado†, Karl D. Hardman, Christopher A. Teleha, Bruce D. J affee,‡ Rui-Qin Liu,
Robert A. Copeland, Maryanne B. Covington, David D. Christ, J ames M. Trzaskos, Robert C. Newton,
Ronald L. Magolda,§ Ruth R. Wexler, and Carl P. Decicco*
DuPont Pharmaceuticals Company, Experimental Station, P.O. Box 80500, Wilmington, Delaware 19880-0500
Received March 21, 2001
To search for TNF-R (tumor necrosis factor R) converting enzyme (TACE) inhibitors, we designed
a new class of macrocyclic hydroxamic acids by linking the P1 and P2′ residues of acyclic anti-
succinate-based hydroxamic acids. A variety of residues including amide, carbamate, alkyl,
sulfonamido, Boc-amino, and amino were found to be suitable P1-P2′ linkers. With an
N-methylamide at P3′, the 13-16-membered macrocycles prepared exhibited low micromolar
activities in the inhibition of TNF-R release from LPS-stimulated human whole blood. Further
elaboration in the P3′-P4′ area using the cyclophane and cyclic carbamate templates led to
the identification of a number of potent analogues with IC50 values of e0.2 µM in whole blood
assay (WBA). Although the P3′ area can accommodate a broad array of structurally diversified
functional groups including polar residues, hydrophobic residues, and amino and carboxylic
acid moieties, in both the cyclophane series and the cyclic carbamate series, a glycine residue
at P3′ was identified as a critical structural component to achieve both good in vitro potency
and good oral activity. With a glycine residue at P3′, an N-methylamide at P4′ provided the
best cyclophane analogue, SL422 (WBA IC50 ) 0.22 µM, LPS-mouse ED50 ) 15 mg/kg, po),
whereas a morpholinylamide at P4′ afforded the most potent and most orally active cyclic
carbamate analogue, SP057 (WBA IC50 ) 0.067 µM, LPS-mouse ED50 ) 2.3 mg/kg, po). Further
profiling for SL422 and SP057 showed that these macrocyclic compounds are potent TACE
inhibitors, with Ki values of 12 and 4.2 nM in the porcine TACE assay, and are broad-spectrum
MMP inhibitors. Pharmacokinetic studies in beagle dogs revealed that SL422 and SP057 are
orally bioavailable, with oral bioavailabilities of 11% and 23%, respectively.
In tr od u ction
TNF-R is synthesized as a membrane-bound proform
comprising 233 amino acids, with a molecular mass of
26 kDa. The pro-TNF-R is then processed by TNF-R
converting enzyme (TACE)13,14 to yield a monomeric
soluble form of 17 kDa comprising 157 nonglycosylated
amino acids. Under physiological conditions, the soluble
TNF-R forms a noncovalently bound homotrimer.15
Although the biological function of pro-TNF-R has yet
to be understood clearly,16 the mature TNF-R exerts its
multiple biological effects via interaction with two
structurally and functionally distinct high-affinity re-
ceptors: TNFR1 (p55) and TNFR2 (p75). Binding of
TNF-R to these receptors results in the activation of
several signal transduction pathways.
Tumor necrosis factor R (TNF-R),1 a critical proin-
flammatory cytokine produced primarily by activated
monocytes/macrophages in response to a variety of
stimuli, is a key mediator of the biological response to
bacterial infection and inflammation and an inducer of
other proinflammatory cytokines such as IL-1â, IL-6,
and IL-8.2-4 During normal host defense, low levels of
serum TNF-R confer protection against infectious agents,
tumors, and tissue damage, and have an important role
in the development of the humoral immune response.5
However, overproduction of TNF-R can lead to a variety
of infectious, autoimmune, and inflammatory disorders
including rheumatoid arthritis (RA), septic shock, mul-
tiple sclerosis, cachexia, congestive heart failure, peri-
odontal disease, and Crohn’s disease.6-9 For example,
elevated TNF-R concentrations have been demonstrated
in a variety of human inflammatory diseases such as
RA10 and Crohn’s disease.11,12
Given the essential role of this cytokine in inflam-
matory diseases, inhibition of TNF-R production or
reduction of TNF-R levels should be beneficial in treat-
ing these pathological conditions. This has now been
proved by anti-TNF-R drugs currently available on the
market. For instance, the disease-modifying anti-TNF-R
drugs such as the PDE-4 inhibitor pentoxifylline17 and
thalidomide18 are TNF-R synthesis blockers. They act
through interference with transcription, translation, or
mRNA half-life, although their effects are broad and
have been associated with considerable toxicity. Perhaps
* To whom correspondence should be addressed. (C.-B.X.) Phone:
(302) 695-2053. Fax: (302) 695-1173. E-mail: chu-biao.xue@dupont-
pharma.com.
† Current address: Department of Biochemistry and Biophysics,
University of Pennsylvania School of Medicine, Philadelphia, PA 19104.
‡ Current address: Millenium Pharmaceuticals, 215 First St.,
Cambridge, MA 02142.
§ Current address: Boehringer-Ingelheim, 900 Ridgebury Rd., P.O.
Box 368, Ridgefield, CT 06877-0368.
10.1021/jm010127e CCC: $20.00 © 2001 American Chemical Society
Published on Web 07/10/2001

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2637
the benefit in treating autoimmune diseases by inhibi-
tion of TNF-R production has been most clearly dem-
onstrated by the monoclonal TNF-R antibody Remicade
(infliximab)19-21 and the soluble TNF p75 receptor
fusion protein (TNFRp75:Fc) Enbrel (etanercept).22,23
These recently approved biologics have proven to be
effective anti-TNF-R drugs. Remicade was approved for
the treatment of Crohn’s disease in 1998, and in
conjunction with methotrexate was later approved for
the treatment of RA.24 Enbrel received approval for the
treatment of RA in 1998.25 Both drugs eliminate TNF-R
effects by neutralizing excess TNF-R in the body. As
proteins, however, these drugs are expensive and in-
convenient to patients. Given the drawbacks of the
current anti-TNF-R biologics, small-molecule anti-
TNF-R agents are currently being exploited through
multiple approaches.7 One very promising small-mol-
ecule approach is the inhibition of TNF-R processing
through the inhibition of TACE.
F igu r e 1. Design of SC903 and SE205 based on marimastat
TACE (ADAM 17), described independently by two
groups,13,14 is a membrane-anchored multidomain en-
zyme containing extracellular disintegrin and protease
regions. It is a member of the adamalysin/ADAM
subfamily of the metzincin superfamily that also in-
cludes the astacins, serralysins, and MMPs. It has been
shown that TACE is the primary enzyme responsible
for the shedding of the biologically active TNF-R from
its membrane-bound proform and inhibition of TACE
blocks TNF-R release.26 This evidence strongly impli-
cates the therapeutic potential of TACE inhibitors for
the treatment of TNF-R-mediated pathologies. As a
result, TACE has recently emerged as a vigorously
pursued therapeutic target.27 The high wave of interest
in TACE inhibitors is attributed not only to their
potential therapeutic application but also to the obser-
vation that MMP inhibitors inhibit TNF-R release in
vitro and in vivo,28,29 implying that the target is ap-
proachable using MMP leads. Small-molecule MMP
inhibitors have been an extensively explored area.30-32
The large number of MMP inhibitors prepared by many
groups of researchers provide in-house libraries for
screening. Structure-activity relationship (SAR) data
on MMP inhibitors have been extensively reported in
the literature and provide useful information for the
design of TACE inhibitors.
As part of our efforts to search for TACE inhibitors,
we have designed a new class of macrocyclic hydroxamic
acids based on anti-succinate-based hydroxamic ac-
ids.33,34 Our extensive investigations on the cyclic link-
age and further systematic SAR studies in the P3′-P4′
area have led to the identification of a number of
macrocycles that are potent inhibitors of TNF-R release
from cells. In our previous paper, we disclosed the two
test molecules SC903 and SE205.33 This paper will
provide a comprehensive account of the rational design,
synthesis, and systematic SAR studies of these macro-
cyclic inhibitors.
and BB-16.
activity include a hydroxamic acid as zinc chelator, a
P1 residue, a P1′ residue (typically an isobutyl group),
a P2′ residue, and a P3′ residue (typically an N-
methylamide). Conformational analysis using molecular
modeling reveals that the preferred backbone conforma-
tion of this type of inhibitor is an extended conformation,
with the two carboxylates of the succinate moiety
assuming an antiperiplanar arrangement, placing the
P1 residue and the P2′ residue on the same side with a
close relationship spatially. X-ray crystallographic stud-
ies of inhibitor/enzyme complexes provided a clearer
understanding of the inhibitor binding interaction at the
active site. This includes chelation of the hydroxamic
acid to the active site Zn and hydrogen-bonding of both
the P1′-P2′ amide bond and the P2′-P3′ amide bond
with the enzyme. In addition, while the P1′ isobutyl
binds in the S1′ pocket, the P1 and P2′ residues have
little interaction with the enzyme and are directed away
from the active site into the solvent. The close proximity
of the P1 and P2′ residues and their exposure to solvent
prompted us to consider a linkage to hold these two
residues to form a macrocycle. Besides novelty, macro-
cycles are known to impart improved physical properties
such as solubility, metabolic stability, and permeability
due to ring rigidity.35 The prerequisite for this is that
the macrocycle should be able to hold all of the struc-
tural elements critical to binding in positions analogous
to those in the corresponding acyclic molecule. Molecular
modeling suggested that the smallest macrocycle ring
should be a 13-membered ring to maintain the active
extended conformation of the acyclic molecules required
for optimal enzyme-inhibitor interaction. We first chose
marimastat as the template to test our design hypoth-
esis. Replacement of the P2′ L-tert-leucine residue in
marimastat with an L-lysine residue, alkylation at the
hydroxyl group R to the hydroxamic acid with bromoac-
etate, and amide bond formation between the acetate
and the lysine side chain amine provided the first test
molecule, SC903, a 13-membered lactam (Figure 1).
Based on BB-16, another test molecule, SE205, a 14-
membered cyclophane, was constructed by tying up the
methyl group at P1 and the methoxy group at P2′ in
BB-16 with a methylene residue (Figure 1).
Design
Succinate-based hydroxamic acids represent an ex-
tensively studied class of potent MMP inhibitors30
possessing modest cellular activity in the inhibition of
TNF-R release.27 The structural requirements of this
class of inhibitor for MMP activity and anti-TNF-R

2638 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
Sch em e 1a
Sch em e 2a
a
Conditions: (a) NH2CH3‚HCl, BOP, DIEA, DMF, 94%; (b)
CH3I, K2CO3, DMF, 100 °C, 43%; (c) LiOH, THF, 100%; (d) 6, BOP,
DIEA, DMF, 93%; (e) 4 N HCl/dioxane; (f) H2, Pd-C, 2-propanol;
(g) BOP, DIEA, DMF, 38% for three steps; (h) LiOH, THF, 85%;
(i) BnONH2‚HCl, BOP, DIEA, DMF, 79%; (j) H2, Pd-C, MeOH.
a
Conditions: (a) n-BuO2CCHO, LDA/THF, 36%; (b) LiOH,
the resulting carboxylic acid with O-benzylhydroxy-
H2O2, 74%; (c) BnBr, DBU, benzene, 79%; (d) NaH, BrCH2CO2-
t-Bu, THF, 71%; (e) H2, Pd-C, MeOH, 99%; (f) TFA, CH2Cl2; (g)
L-Lys(Cbz)NHMe, BOP, DIEA, DMF, 75%; (h) H2, Pd-C, followed
by 4 N HCl/dioxane; (i) BOP, DIEA, DMF/CHCl3, 50%; (j) 1 N
LiOH, THF, 73%; (k) BnONH2‚HCl, BOP, DIEA, DMF, 35%; (l)
H2, Pd-C, MeOH, 62%.
lamine, and hydrogenolytic removal of the benzyl group.
Unlike the synthesis of 9, the synthesis of its N-
methyl analogue 15 involved an intramolecular cycliza-
tion at the backbone amide bond as shown in Scheme
2. The Nꢀ-methyl-L-lysine derivative 12 was obtained
by subjecting the commercially available NR-Boc-Nꢀ-
(CF3CO)-L-Lys (10) to a coupling with methylamine
using BOP and subsequently to a methylation with
CH3I/K2CO3 at an elevated temperature followed by a
hydrolysis using LiOH to remove the trifluoroacetyl
group. Coupling of 12 with the acid intermediate 6
afforded the amide 13 as a 3:1 mixture of two diaster-
eomers. Intramolecular cyclization was accomplished
using BOP following hydrogenolytic removal of the Cbz
group and acidolytic removal of the Boc group in 13.
The optically pure ester 14, obtained by silica gel
chromatography after cyclization, was then converted
to the hydroxamic acid 15 in a manner similar to that
described for the preparation of 9 in Scheme 1.
The macrocycle 22 was prepared using the protocol
outlined in Scheme 3. The L-cysteine derivative 17 was
obtained by treatment of L-cysteine (16) with o-chlo-
ronitrobenzene followed by Boc protection. After conver-
sion of the carboxylic acid to an N-methylamide (18),
the Boc group was removed using acid. The resulting
amine 19 was coupled with the acid intermediate 5 to
give amide 20. The nitro group in 20 was reduced to an
amine using zinc, and the tert-butyl group was cleaved
with acid. Following intramolecular cyclization using
BOP, we did not try to convert the n-butyl ester to a
hydroxamic acid using the normal procedures described
in Scheme 1 due to the presence of the thioether in the
Ch em istr y
The first macrocyclic compound, 9 (SC903), with an
ether-amide linkage, was prepared using the sequence
depicted in Scheme 1. Evan’s aldol condensation36 of 1
[Xc ) (S)-4-benzyl-2-oxazolidinone] with n-butyl gly-
oxylate provided 2-hydroxy-3(R)-isobutylsuccinate 2 as
a 3:1 mixture of two diastereomers, with the major
diastereomer the desired anti isomer.37 The mixture was
inseparable by silica gel chromatography and was
carried all the way through macrocyclization. Removal
of the chiral auxiliary group in 2 using LiOH/H2O2
followed by benzyl esterification provided the benzyl
ester 3, which was subjected to an alkylation at the
hydroxyl group with tert-butyl bromoacetate to give
compound 4. Hydrogenolytic removal of the benzyl
group and acidolytic removal of the tert-butyl group in
4 afforded the two carboxylic acids 5 and 6, respectively.
Coupling of 5 with Nꢀ-Cbz-L-lysine N-methylamide
afforded the amide 7. Following removal of the tert-butyl
group using acid and of the Cbz group by hydrogenoly-
sis, an intramolecular cyclization was performed using
BOP to afford the macrocycle as a mixture of two
diastereomers. Separation of the two diastereomers by
silica gel chromatography furnished the major diaste-
reomer 8. The hydroxamic acid 9 was then obtained by
sequential hydrolysis of the n-butyl ester, coupling of

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2639
Sch em e 3a
Sch em e 4a
a
Conditions: (a) (1) o-chloronitrobenzene, K2CO3, DMF, 60 °C,
(2) (Boc)2O, 48%; (b) NH2CH3‚HCl, BOP, DIEA, DMF, 82%; (c) 4
N HCl/dioxane; (d) 5, BOP, DIEA, DMF, 83%; (e) Zn, AcOH, H2O;
(f) BOP, DIEA, DMF, 71%; (g) NH2OH‚HCl, DIEA, MeOH, 60 °C,
40%.
a
Conditions: (a) 1,4-dibromo-2-butene, NaH, DMF, 0 °C, 65%;
(b) NR-Boc-L-TrpNHMe, NaH, DMF, 0 °C, 62%; (c) H2, Pd-C,
2-propanol, 91%; (d) 4 N HCl/dioxane; (e) BOP, DIEA, DMF,
CHCl3, 45%; (f) LiOH, THF; (g) BnONH2‚HCl, BOP, DIEA, DMF,
65%; (h) H2, Pd-C, MeOH, 86%.
molecule. Instead, the hydroxamic acid 22 was obtained
by treatment of the n-butyl ester with hydroxylamine
in MeOH at 60 °C, although the yield was low (40%).
The macrocycle 27 was synthesized starting from the
intermediate 3 (Scheme 4). Alkylation of 3 with 1,4-
dibromo-2-butene using NaH gave the ether derivative
23. Following a second alkylation with NR-Boc-L-tryp-
tophan N-methylamide on 23 using NaH, the resulting
intermediate 24 was subjected to a hydrogenolysis to
remove the benzyl group and reduce the olefin and
subsequently to an acid treatment to remove the Boc
group. Macrocyclization using BOP followed by silica gel
chromatography afforded the desired anti diastereomer
25. Conversion of the ester 25 to the hydroxamic acid
27 was accomplished by employing conditions similar
to those used for the preparation of 9 in Scheme 1.
Scheme 5 illustrates the synthesis of two common key
intermediates (31a ,b) and subsequent conversion of 31a
to 38. LDA-promoted alkylation of 28a (R ) isobutyl)
and 28b (R ) n-hexyl)38 with allyl bromide provided the
corresponding succinate derivatives as a 10:1 (syn:anti)
mixture of two diastereomers. Epimerization by eno-
lization using LDA followed by addition of Et2AlCl prior
to quenching with methanol altered the ratio of the two
diastereomers to 1:8 (syn:anti). Without the use of Et2-
AlCl, the two diastereomers were obtained in a 1:1 ratio.
The desired anti diastereomer was isolated by silica gel
chromatography following conversion of the carboxylic
acids 29a ,b to benzyl esters 30a ,b. Hydroboration of the
olefin in 30a ,b using 9-BBN provided alcohols 31a ,b.
Hydrogenation of 31a produced the carboxylic acid 32,
which was subsequently used for the synthesis of the
macrocycle 38. This involved coupling of 32 with Nꢀ-
Cbz-L-Lys-NHMe using BOP, oxidation of the alcohol
in 33 to a carboxylic acid using RuCl3/H5IO6, hydro-
genolytic removal of the Cbz group in 34, and macro-
cyclization of 35 using BOP to give the 13-membered
lactam 36. Cleavage of the tert-butyl ester in 36 with
acid followed by coupling with O-benzylhydroxylamine
using BOP furnished the O-benzyl hydroxamate 37,
from which the hydroxamic acid 38 was obtained by
catalytic hydrogenation.
Starting from the intermediate 32, three macrocycles
(41, 43, and 44) were prepared as shown in Scheme 6.
Coupling of 32 with an L-lysine derivative provided the
amides 39a ,b, which were subjected to an intramolecu-
lar Mitsunobu reaction to furnish the cyclic sulfona-
mides 40a ,b. The phenylsulfonamide intermediate 40a
was then converted to a hydroxamic acid (41) using
procedures analogous to those described for 38. Removal
of the mesitylsulfonyl group in 40b using HBr/AcOH
followed by Boc protection provided the carboxylic acid
42, which was converted to a hydroxamic acid (43) by
coupling with O-benzylhydroxylamine followed by cata-
lytic hydrogenolysis. Removal of the Boc group in 43
using acid afforded target compound 44.
The enantiopure amino acid derivative 48, which was
used for the synthesis of the cyclophane 57, was
prepared using Meyers’ methodology39 as illustrated in
Scheme 7. The alcohol 45 was treated with methane-
sulfonyl chloride, and the resulting mesylate was con-
verted to iodide 46. LDA alkylation of pseudoephedrine
glycinamide with 46 provided the amide 47, which was
subsequently hydrolyzed and esterified to give 48.

2640 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
Sch em e 5a
Sch em e 6a
a
Conditions: (a) L-Lys(RSO2)NHMe, BOP, DIEA, DMF; (b)
DIAD, PPh3, THF; (c) TFA, CH2Cl2, (d) BnONH2‚HCl, BOP, DIEA,
DMF; (e) H2, Pd-C, MeOH; (f) HBr, AcOH; (g) (Boc)2O, DIEA,
DMF; (h) 4 N HCl/dioxane.
a
Conditions: (a) LDA, allyl bromide, THF; (b) (1) LDA, THF,
-78 °C to rt, (2) -78 °C, Et2AlCl, -78 °C to rt, (3) -78 °C, MeOH;
(c) BnBr, DBU, benzene; (d) 9-BBN, H2O2, NaOAc, THF; (e) H2,
Pd-C, MeOH; (f) L-Lys(Cbz)NHMe, BOP, DIEA, DMF, 77%; (g)
RuCl3, H5IO6, 60%; (h) BOP, DIEA, DMF, 70%; (i) TFA, CH2Cl2;
(j) BnONH2‚HCl, BOP, DIEA, DMF, 61%.
Sch em e 7a
Several series of cyclophanes were prepared starting
from the intermediates 31a and 31b (Scheme 8). The
alcohol in 31a ,b was converted to a bromide, and the
benzyl group was subsequently removed by hydrogena-
tion. Coupling of 49a ,b with an L-tyrosine derivative,
an L-homotyrosine derivative, or 48 gave the cyclization
precursors 50a -d . Cyclization was carried out using
Cs2CO3 in a mixed solvent system of DMF/DMSO at an
elevated temperature to provide the cyclophanes 51a -d
in high yields. Removal of the tert-butyl group using
TFA followed by coupling with O-benzylhydroxylamine
provided the benzyl-protected hydroxamates 52a -d .
Hydrogenation of 52b afforded compound 53. Saponi-
fication of the methyl ester in 52a -d , subsequent
coupling of the carboxylic acid with an amino residue,
and hydrogenolysis furnished compounds 54a -v (R )
isobutyl, n ) 1), 55a -g (R ) n-hexyl, n ) 1), 56 (R )
isobutyl, n ) 2), and 57 (R ) isobutyl, n ) 3). Removal
of the Boc group at the lysine side chain in 54v and 55g
afforded compounds 58 and 59.
a
Conditions: (a) MeSO2Cl, NEt3, CH2Cl2, 100%; (b) NaI,
acetone, reflux, 96%; (c) (1S,2S)-pseudoephedrine glycinamide,
LDA, LiCl, THF, 47%; (d) NaOH, MeOH, reflux; (e) MeOH, HCl,
reflux, 99%.
with iodomethane using K2CO3 in DMF at 110 °C
produced the dimethylated product 62, which was then
subjected to treatment with LiOH in THF to hydrolyze
the trifluoroacetyl group. The methyl ester was also
hydrolyzed under these conditions. Subsequent rees-
terification using 4 N HCl in dioxane/MeOH afforded
the lysine methyl ester 63.
The synthesis of two series of cyclic carbamate
analogues started with the intermediate 31a (Scheme
10). Reaction of the alcohol 31a with 4-nitrophenyl
chloroformate gave the activated carbonate 64, which
was converted to a carbamate (65a , R1 ) H; 65b, R1 )
The side chain N-methyl derivative of L-lysine (63),
which was used for the synthesis of a series of cyclic
N-methylcarbamate analogues, was prepared according
to Scheme 9. Reaction of the commercially available NR-
Cbz-L-Lys (60) with ethyl trifluoroacetate gave NR-Cbz-
L-Lys-(CF3CO)OH (61) in high yield. Methylation of 61

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2641
Sch em e 8a
Sch em e 9a
a
Conditions: (a) CF3CO2Et, K2CO3, MeOH, H2O, 100%; (b)
CH3I, K2CO3, DMF, 110 ïC, 90%; (c) LiOH, THF; (d) 4 N HCl/
dioxane, MeOH.
followed by coupling with an aminopyridine or ami-
nothiazole derivative produced the intermediates 71 (R1
) H) and 72a -f (R1 ) Me), which were subjected to an
acidolytic removal of the tert-butyl group and subse-
quently to a coupling with hydroxylamine hydrochloride
to afford hydroxamic acids 73 (R1 ) H) and 74a -f (R1
) Me). Hydrolysis of the methyl ester in 73 (R2 ) 4-(2-
methoxy-2-oxoethyl)thiazol-2-yl) using LiOH afforded
compound 75. The carboxylic acid in 75 was then
coupled with morpholine using BOP to provide com-
pound 76. In this reaction, an excess amount of mor-
pholine (5 equiv) was used to avoid coupling of the
carboxylic acid with the hydroxamic acid in 75.
Resu lts a n d Discu ssion
A cellular assay was used to determine the activity
in the inhibition of TNF-R release from LPS-stimulated
human whole blood.40 Since whole blood activities are
determined and contributed to by many factors includ-
ing serum protein binding and cell penetration, we also
counterscreened with MMP (1, 3, and 9) enzyme assays
in the early stage of study to aid in our design. On the
basis of the assumption that TACE has a high level of
homology with MMPs in the active site, MMP activities
could provide us with a better understanding of the
effect of a linkage on the backbone conformation that
is critical to binding. Compounds that were potent in
whole blood assay (WBA) were selected for further
studies in the LPS-mouse model to assess their oral
activity.
SARs of th e P 1-P 2′ Lin k a ge. Based on marimas-
tat, we designed and synthesized the first test molecule,
9 (SC903), a 13-membered lactam with an ether-amide
linkage. As illustrated in Table 1, SC903 was potent
against MMP-1, MMP-2, and MMP-9 and had an IC50
of 6 µM in WBA, which is slightly less active than
marimastat (IC50 ) 4.1 µM). An X-ray crystallographic
study of SC903 bound to MMP-3 reveals that this
macrocycle effectively maintains the extended confor-
mation with a binding mode very similar to that of the
linear inhibitors,33 confirming our design hypothesis.
Encouraged by these results, several modifications were
made to improve the anti-TNF-R activity. One of the
a
Conditions: (a) CBr4, PPh3, DIEA, THF; (b) H2, Pd-C, MeOH;
(c) L-TyrOMe, TBTU, DIEA, DMF; (d) Cs2CO3, DMF, DMSO, 60
°C; (e) TFA, CH2Cl2; (f) BnONH2‚HCl, TBTU, DIEA, DMF, 80 °C;
(g) H2, Pd-BaSO4, MeOH; (h) LiOH, THF; (i) R′NH2, TBTU, DIEA,
DMF.
Me) by treatment with NR-Cbz-L-Lys-OMe or 63. After
hydrogenolytic removal of the Cbz and the benzyl
groups, the carboxylic acid and the amine generated
were coupled together to form a macrocyclic carbamate
(66a , R1 ) H; 66b, R1 ) Me). The tert-butyl ester was
then converted to a benzyl-protected hydroxamate using
the standard procedures. The final target compounds
68a -z and 68a a -k k (R1 ) H) and 69a -h (R1 ) Me)
were obtained by hydrolysis of the methyl ester, cou-
pling of the generated carboxylic acid with an amine,
and hydrogenation to remove the benzyl group. Removal
of the Boc group at the lysine side chain in compound
68w furnished compound 70.
The macrocyclic carbamate compounds 75 and 76 (R1
) H) and 74a -f (R1 ) Me) containing an N-thiazoly-
lamide residue or an N-pyridylamide residue were
prepared using a sequence different from that described
in Scheme 10. This sequence involves the introduction
of an aminopyridine or aminothiazole group at P3′-P4′
before conversion of the tert-butyl ester to a hydroxam-
ate (Scheme 11). Hydrolysis of the methyl ester in 66a ,b

2642 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
Sch em e 10a
a
Conditions: (a) p-nitrophenyl chloroformate, DIEA, THF, 91%; (b) NR-Cbz-L-Lys(R1)OMe, K2CO3, DMF; (c) H2, Pd-C, MeOH; (d)
BOP, DIEA, DMF, CHCl3; (e) TFA, CH2Cl2; (f) BnONH2‚HCl, BOP, DIEA, DMF; (g) LiOH, THF; (h) R2NH2, BOP, DIEA, DMF; (i) H2,
Pd-BaSO4, MeOH.
Sch em e 11a
Ta ble 1. Cyclic Inhibitors with a Variety of Linkers
Ki, nMb
WBAa
MMP- MMP-
compd
X
IC50, µM MMP-1
3
9
marimastat
BB-16
9 (SC903) -OCH2CONH(CH2)4-
15
22
27
38
44
41
4.1
1.8
6.5
3.8
8.0
<1
<1
2.8
<1
<1
<1
1.4
<1
<1
<1
<1
<1
1.2
5.3
1.8
2.4
<1
<1
2.6
3.9
NTc
NT
3.6
12.8
<1
<1
1.1
<1
1.8
2.1
3.3
1.93
24.1
3
22.3
2.88
NT
NT
3.4
-OCH2CON(Me)(CH2)4-
-OCH2CONHPh-2-SCH2-
-O(CH2)4(1-indol-3-yl)CH2- 3.7
-(CH2)2CONH(CH2)4-
-(CH2)3NH(CH2)4-
-(CH2)3N(PhSO2)(CH2)4-
-(CH2)3N(Boc)(CH2)4-
-(CH2)3OCONH(CH2)4-
-(CH2)3OCONMe(CH2)4-
1.6
2.7
1.3
1.4
1.0
2.3
1.2
1.0
2.4
43
68a
69a
3
NT
NT
32.7
48.0
16.2
54a (SE205) -(CH2)3OPh-4-CH2-
56
57
-(CH2)3OPh-4-(CH2)2-
-(CH2)3OPh-4-(CH2)3-
a
Inhibition of TNF-R release in human WBA was determined
in three donors. Ki values are from a single determination. c NT
b
) not tested.
the better WBA potency of BB-16 (IC50 ) 1.8 µM) than
that of marimastat (IC50 ) 4.1 µM) might be ascribed
to the more hydrophobic 4-methoxybenzyl P2′ group in
BB-16 led us to prepare two analogues with a hydro-
phobic aromatic ring at P2′. Thus, replacement of the
P2′ L-lysine residue in SC903 with an S-(2-aminophe-
nyl)-L-cysteine residue afforded the 13-membered lac-
tam 22, which was more potent against MMP-1 and
equipotent against MMP-3 but slightly less potent in
WBA (IC50 ) 8.0 µM) relative to SC903. The P2′
L-tryptophan analogue 27, a 15-membered macrocycle
with an alkyl linkage, was equipotent to inhibitor 15
in the inhibition of MMP-1, MMP-3, and TNF-R release
(IC50 ) 3.7 µM).
a
Conditions: (a) LiOH, THF; (b) R2NH2, BOP, DIEA, DMF; (c)
TFA, CH2Cl2; (d) HONH2‚HCl, DIEA, BOP, DMF; (e) morpholine,
BOP, DIEA, DMF.
physical properties observed for SC903 is its high
polarity, presumably due to too many amide bonds in
the molecule. This observation prompted us to make an
N-methyl substitution at the linker amide bond, which
was accompanied by significantly improved MMP-1 and
MMP-3 activity and slightly improved whole blood
activity (15, WBA IC50 ) 3.8 µM). The speculation that
We next replaced the oxygen R to the hydroxamate
in SC903 with a methylene, affording the 13-membered

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2643
a
Ta ble 2. Cyclophanes with Isobutyl at P1′
lactam 38. Interestingly, while this replacement re-
sulted in similar MMP-1 and MMP-9 activities, the
WBA potency was improved by approximately 4-fold
(IC50 ) 1.6 µM), suggesting that a less polar group R to
the hydroxamate is beneficial to whole blood activity.
Several analogues were synthesized by variation at the
linkage in 38 while the P2′ lysine residue was kept
constant. Replacement of the carbonyl of the linker
amide in 38 with a methylene furnished the secondary
amino analogue 44, which showed better MMP-1 activ-
ity but reduced MMP-9 activity and slightly diminished
whole blood activity (IC50 ) 2.7 µM) relative to 38.
N-Substitution at the amino in 44 with a phenylsulfonyl
(41) or a Boc (43) led to a 2-fold improvement in WBA
potency and a marked improvement in MMP-9 activity
over 44, suggesting that a lipophilic group is preferred
at this position. Insertion of an oxycarbonyl into the
secondary amino linkage in 44 afforded the 15-mem-
bered macrocyclic carbamate 68a , which demonstrated
better WBA (IC50 ) 1.0 µM) and MMP-9 activities
compared with 44. N-Methyl substitution at the car-
bamate residue in 68a resulted in a 2-fold drop in the
inhibition of TNF-R release (69a , WBA IC50 ) 2.3 µM).
This result is in contrast to that observed for SC903 in
which N-methyl substitution resulted in slightly im-
proved whole blood activity.
LPS-mouse (po)
WBA
% inhib or
compd
R
IC50, µM ED50 (mg/kg)
54a (SE205) NHMe
1.2
0.68 NT
0.70 34% at 10 mg/kg
1.0
0.22 ED50 ) 15
0.24 NT
0.20 31% at 30 mg/kg
0.20 21% at 30 mg/kg
0.20 0% at 10 mg/kg
0.70 NT
ED50 ) 17
54b
54c
54d
NHCH2CF3
NHCH(CH3)2
NH(CH2)2Ph(4-SO2NH2)
NT
54e (SL422) Gly-NHMe
54f
54g
54h
54i
54j
54k
54l
54m
54n
54o
54p
54q
54r
54s
54t
58
Gly-NHCH2CF3
Gly-NH-cyclobutyl
Gly-NH-t-Bu
Gly-NHPh
Gly-NH(2,4-difluoroPh)
Gly-NH-fluoren-1-yl
Gly-NMe2
Gly(piperidin-1-yl)
Gly(4-hydroxypiperidin-1-yl)
0.93 NT
0.40 45% at 30 mg/kg
0.14 49% at 10 mg/kg
0.20 ED50 ) 21
Gly(3,5-dimethylpiperidin-1-yl) 0.27 15% at 10 mg/kg
Gly(3,3-dimethylpiperidin-1-yl) 0.46 NT
L-Ala-NHMe
D-Ala-NHMe
L-Tyr(OMe)NHMe
L-Ser(O-t-Bu)NHMe
L-Lys-NHMe
0.70 ED50 > 30
4.49 NT
0.85 NT
0.95 NT
0.83 NT
Based on BB-16, we designed and synthesized the
second test molecule, 54a (SE205), a 14-membered
cyclophane. While SE205 was less active in the inhibi-
tion of MMPs than BB-16, it was equipotent to BB-16
in the inhibition of TNF-R release (WBA IC50 ) 1.2
µM).41 The crystal structure of SE205 bound to MMP-3
reveals that the 14-membered cyclophane ring holds all
the structural elements (hydroxamate, P1′ isobutyl, and
two amide bonds) in positions analogous to those in BB-
16.34 More importantly, SE205 was found to exhibit
much better water solubility (13 mg/mL) compared with
BB-16 (0.3 mg/mL) and good oral activity in the LPS-
mouse model, with an ED50 of 17 mg/kg, making this
molecule an attractive prototype for further modifica-
tion. To explore the effect of ring size, two more
cyclophanes with larger rings were prepared by replac-
ing the L-tyrosine in SE205 with L-homotyrosine (56)
and L-homo-homotyrosine (57). While the 15-membered
cyclophane 56 maintained the WBA potency (IC50 ) 1.0
µM), the 16-membered cyclophane 57 showed a 2-fold
loss in the inhibition of TNF-R release (IC50 ) 2.4 µM).
Beginning with the design of the first two test
molecules, SC903 and SE205, we have conducted ex-
tensive investigations into the macrocyclic framework
using a variety of linkages or by changing the ring size,
leading to the identification of several inhibitors with
IC50 values of 1-2 µM in the inhibition of TNF-R
release. However, the similar whole blood activities
among the majority of inhibitors illustrated in Table 1
suggest that changing the linkage has little effect on
the cellular anti-TNF-R activity. This observation
prompted us to shift our efforts to the P3′-P4′ area for
further SAR exploration. On the basis of low molecular
weight, whole blood potency, and water solubility, the
cyclophane template SE205 and the cyclic carbamate
template 68a were selected for further structure refine-
ment and potency optimization in the P3′-P4′ area,
54u
L-Val-NHMe
1.51 NT
a
See footnotes a and c in Table 1.
with emphasis on improving WBA potency and oral
anti-TNF-R activity in the LPS-mouse model.
Cyclop h a n e Ser ies. As shown in Table 2, replace-
ment of the P3′ N-methyl group in SE205 with a longer
alkyl (54b, trifluoroethyl; 54c, isopropyl) or arylalkyl
(54d , 4-(aminosulfonyl)phenethyl) led to a relatively
minor improvement in WBA potency. A more remark-
able enhancement in potency in the inhibition of TNF-R
release was achieved when a glycine N-methylamide
residue was incorporated into the P3′ position, as
observed with 54e (SL422; IC50 ) 0.22 µM). Despite its
5-fold improvement in whole blood activity over SE205,
SL422 was found to be equipotent to SE205 in the LPS-
mouse model. Larger, more hydrophobic groups than
methyl at P4′ such as trifluoroethyl (54f), cyclobutyl
(54g), tert-butyl (54h ), and phenyl (54i) maintained the
WBA potency but reduced the oral activity. Further
increasing the lipophilicity at P4′ was found to be
detrimental to whole blood activity as seen in the 2,4-
difluorophenyl analogue 54j and the fluoren-1-yl ana-
logue 54k , possibly due to higher protein binding of
these groups. While N-methyl substitution at the P4′
N-methylamide moiety in SL422 led to a drop in in vitro
and in vivo potencies (54l), the P4′ piperidine amide
analogue 54m showed WBA and oral activities compa-
rable to those of SL422. Substitution with a polar
hydroxyl group on the piperidine ring (54n ) basically
retained the in vitro and in vivo potencies, whereas
hydrophobic substituents on the piperidine ring (54o
and 54p ) reduced the WBA and oral activities.
By keeping the P4′ N-methylamide constant, we next
investigated the SAR at the P3′ position by replacing
the glycine residue in SL422 with several other amino
acid residues. In general, increasing the size of the P3′

2644 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
a
Ta ble 3. Cyclophanes with n-Hexyl at P1′
Ta ble 4. Cyclic Carbamate Analoguesa
WBA
LPS-mouse (po)
compd
R
IC50, µM
inhib at 30 mg/kg
WBA LPS-mouse (po)
IC50
,
inhib at 10 mg/kg
or ED50 (mg/kg)
53
OMe
NHMe
NHBn
>50
2.5
2.2
0.48
0.82
0.55
0.76
0.30
NT
compd
68a
68b
68c
68d
75
R
µM
55a
55b
55c
55d
55e
59
26% at 100 mg/kg
NT
NHMe
NHCH2CF3
NHPh
1.0
NT
2.0
NT
NH(CH2)2Ph(4-SO2NH2)
NH(CH2)3(imidazol-1-yl)
Gly-NHMe
26%
30%
NT
36%
18%
0.28
0.19
NT
NHPh(morpholin-4-yl)
31%
44%
24%
NH(4-carboxymethylthiazol-2-yl) 0.083
NH{[4-(morpholin-4-yl)carbonyl- 0.12
methyl]thiazol-2-yl}
L-Lys-NH2
76
55f
L-Ser(O-t-Bu)NHMe
a
See footnotes in Tables 1 and 2.
68e
68f
68g
68h
68i
68j
68k
68l
NHBn
0.74
0.70
0.50
0.49
0.39
NT
NT
NT
NT
NT
0%
NH(2-picolyl)
NH(3-picolyl)
NH(4-picolyl)
NH[2-(2-pyridyl)ethyl]
residue diminished the WBA potency as indicated by
comparison of 54q, 54s,t, and 58 with SL422 (Table 2).
The similar whole blood activities among 54q, 54s,t, and
58 suggest that the TACE S3′ pocket can accommodate
a variety of structurally diversified groups including an
amino residue (58). However, a sterically congested
group such as an isopropyl (54u , IC50 ) 1.51 µM) at the
R-position of the glycine residue is less tolerated. The
over 6-fold difference in WBA potency between the
D-alanine analogue 54r and the L-alanine analogue 54q
indicates that the stereochemistry at this position is
important to anti-TNF-R activity.
In an SAR study parallel to that of the SE205 series,
we investigated the P3′-P4′ area of the cyclophane
template bearing a longer P1′ alkyl group (n-hexyl) that
replaces the P1′ isobutyl group in the SE205 series. This
study was initiated on the basis of the rationale that a
longer, more hydrophobic P1′ group might confer better
binding to TACE in the S1′ pocket as has been observed
with some MMPs.42 A comparison of the WBA data in
Table 3 with those in Table 2 reveals that there is no
clear trend as to whether the n-hexyl offers any advan-
tage over the isobutyl in the in vitro anti-TNF-R activity.
While the P3′ N-methylamide analogue 55a 41 and the
P3′ glycine N-methylamide analogue 55e are about
2-fold less active than their isobutyl counterparts SE205
and SL422, analogues 55c and 55f are 2-fold and 3-fold
more active than their isobutyl counterparts 54d and
54t, respectively, in WBA. However, the dramatic loss
in oral activity in the LPS-mouse model when the
isobutyl in SE205 was replaced with n-hexyl (55a )
clearly indicates that a longer, hydrophobic P1′ group
results in lower oral activity. Of particular note is the
result from the replacement of the P3′ N-methylamide
in 55a with a methyl ester (53). The complete loss in
whole blood activity in 53 suggests the importance of
the amide NH for binding (as a hydrogen bond donor)
at this position.
NH[2-(6-methoxyindol-3-yl)ethyl] 0.094
NH[2-(5-methoxyindol-3-yl)ethyl] 0.49
4%
NH[2-(5-fluoroindol-3-yl)ethyl]
0.13
39%
NT
21%
33%
68m
68n
68o
NH[2-(5-methylindol-3-yl)ethyl] 1.17
NH(2-hydroxy-2-phenylethyl)
NH(1-ethoxycarbonyl-
piperidin-4-yl)
NH(2(R)-hydroxy-1(S)-indane)
NH(2(S)-hydroxy-1(R)-indane)
0.085
0.13
68p
68q
68r
68s
68t
68u
68v
68w
70
0.11
0.40
27%
NT
NHCH2(2(S)-tetrahydrofuranyl) 0.26
NHCH2(2(R)-tetrahydrofuranyl) 0.98
NT
NT
Gly-NHMe
L-Ala-NHMe
L-Leu-NHMe
0.12
0.27
0.10
0.21
0.69
1.2
45%
NT
0%
0%
NT
L-Lys(Boc)NHMe
L-Lys-NHMe
L-tert-Leu-NHMe
Gly(morpholin-4-yl)
68x
68y
(SP057)
NT
0.067
ED50 ) 2.6
68z
Gly(2,6-dimethylmorpholin-4-yl) 0.15
43%
68a a
68bb
68cc
68d d
Gly(4-methyl-piperazin-1-yl)
Gly(4-phenylpiperazin-1-yl)
0.16
37%
0.070
29%
Gly[4-(2-pyridinyl)piperazin-1-yl] 0.070
ED50 ) 8.7
ED50 ) 7
Gly(4-ethoxycarbonyl-
piperazin-1-yl)
0.13
68ee
68ff
Gly(4-hydroxypiperidin-1-yl)
Gly[4-(piperidin-1-yl)-
piperidin-1-yl]
Gly(4-carboxypiperidin-1-yl)
Gly(3-carboxypiperidin-1-yl)
L-Ala(morpholin-4-yl)
L-Ser(morpholin-4-yl)
b-Ala(morpholin-4-yl)
0.28
0.18
ED50 ) 3.6
ED50 ) 8.1
68gg
68h h
68ii
68jj
68k k
0.22
0.58
0.15
0.10
0.17
NT
NT
ED50 > 10
ED50 > 10
0%
a
See footnotes in Tables 1 and 2.
was replaced with a thiazole bearing a polar substituent
at the 4-position (75 and 76). These modifications
resulted in some improvement in WBA potency over
that of 68c. Unfortunately, these potent analogues
exhibited low oral activity in the LPS-mouse model. The
similar in vitro potencies between 75 and 76 suggest
that a carboxylic acid at this position is tolerated for
whole blood activity.
Several aryl or heteroaryl groups connected to the
P2′-P3′ amide through 1-2 methylene unit(s) were
examined at the P3′ position. Insertion of a methylene
between the amide bond and the phenyl group in 68c
led to a slight loss in WBA as observed with 68e.
Replacement of the phenyl in 68e with more polar
pyridyl residues (68f-h ) did not significantly improve
the WBA potency. One carbon longer between the P2′-
P3′ amide bond and the pyridyl in 68f led to a relatively
Cyclic Ca r ba m a te Ser ies. The SAR data of the
cyclic carbamate series are shown in Table 4. While
replacement of the P3′ methyl group in 68a with a
trifluoroethyl group (68b) resulted in a 2-fold loss in
WBA, a phenyl at this position (68c) enhanced the
activity by over 3-fold. To improve the water solubility
of 68c, a polar morpholinyl group was introduced at the
para position of the phenyl group (68d ) or the phenyl

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2645
Ta ble 5. Cyclic N-Methylcarbamate Analoguesa
minor enhancement in WBA (68i). The in vitro activity
of the four indolyl analogues 68j-m is dependent on
the nature and position of the substituent on the indole
ring, with the 6-methoxyindolyl analogue 68j exhibiting
the best WBA potency (IC50 ) 0.094 µM) but a lack of
oral activity. A 2-hydroxy-2-phenylethyl group at P3′
provided another potent analogue (68n ) in WBA (IC50
) 0.085 µM), which was inactive when administered
orally in the LPS-mouse model.
WBA LPS-mouse (po)
Nonaromatic carbocycles or heterocycles appended to
the P2′-P3′ amide through 0-1 methylene spacer were
also examined. The 1-ethoxycarbonylpiperidin-4-yl ana-
logue 68o, 2(R)-hydroxy-1(S)-indane analogue 68p , and
2(S)-tetrahydrofuran analogue 68r all were potent in
whole blood but lacked oral activity. The almost 4-fold
difference in WBA potency between 68p and 68q and
between 68r and 68s provides further evidence that the
stereochemistry at this position is important to anti-
TNF-R activity.
Our next phase of the SAR investigations at P3′-P4′
was to incorporate an amino acid residue at P3′ with
an N-methylamide at P4′. The glycine N-methylamide
analogue 68t demonstrated good WBA potency with
weak oral activity in the LPS-mouse model. In contrast
to the SAR in the SE205 series (vide supra), increasing
the size of P3′ had a lesser effect on the in vitro anti-
TNF activity as revealed by comparison of 68u , 68v,
and 68w with 68t. Despite the good in vitro potency,
however, the L-leucine analogue and the L-lysine ana-
logue showed no in vivo activity when dosed orally to
mice. In parallel to the SAR seen in the SE205 series,
a sterically hindered group such as a tert-butyl at P3′
is detrimental to whole blood activity (68x, IC50 ) 1.2
µM). The 3-fold loss in WBA (70) by removing the lysine
side chain Boc in 68w suggests that a charge at P3′ is
less preferred in this series.
compd
R
IC50, µM ED50, mg/kg
69a NHMe
69b Gly(morpholin-4-yl)
69c Gly(4-hydroxypiperidin-1-yl)
69d Gly(4-methylpiperazin-1-yl)
2.3
NT
0.20
0.30
0.12
4.9
2.7
11.4
>30
6.3
>30
>30
28
>30
>30
>30
>30
>30
69e Gly(4-ethoxycarbonylpiperazin-1-yl) 0.60
69f Gly-N(Me)2
69g Gly(pyrrolidin-1-yl)
69h Gly-NHMe
74a Gly-NH(pyridin-2-yl)
74b Gly-NH(4-methylthiazol-2-yl)
74c NH(pyridin-2-yl)
74d NH(pyridin-3-yl)
74e NH(pyridin-4-yl)
74f NH(4-methylthiazol-2-yl)
0.30
0.40
0.42
0.20
0.30
0.20
0.50
0.50
0.30
a
See footnotes in Tables 1 and 2.
analogues bearing a morpholino at P4′ were prepared.
Surprisingly, replacing the glycine residue in SP057
with an L-alanine residue (68ii), an L-serine residue
(68jj), or a â-alanine residue (68k k ) resulted only in a
minor loss in WBA potency but in a dramatic loss in
oral activity. These results, taken together with those
from the previous studies in this series and the cyclo-
phane series (vide supra), provide convincing evidence
that a glycine residue at P3′ is a critical structural
component to achieve both good in vitro activity and
good oral activity.
To understand whether N-methylation at the car-
bamate moiety can offer any advantage in whole blood
activity and/or oral potency, we also conducted SAR
studies at the P3′-P4′ area using the N-methylcarbam-
ate template 69a (Table 5). In general, this series of
analogues were less active in WBA as compared with
their NH-carbamate counterparts. For example, the
morpholino analogue 69b, the 4-(ethoxycarbonyl)piper-
azino analogue 69e, and the glycine N-methylamide
analogue 69h were 3-fold, over 4-fold, and over 3-fold
less active than their NH-carbamate counterparts SP057,
68d d , and 68t, respectively, in in vitro anti-TNF-R
activity. More polar residues at P4′ appeared to main-
tain the WBA and oral activities as revealed by com-
parison of 69c and 69d with their counterparts 68ee
and 68a a , respectively. Of particular note in this series
is the dramatic enhanced oral activity of the P4′ N,N-
dimethyl analogue 69f (ED50 ) 6.3 mg/kg) versus the
P4′ N-methyl analogue 69h . This oral activity was
completely lost when the dimethylamino in 69f was
replaced with a pyrrolidino moiety (69g). Analogues
with a pyridine or thiazole directly connected to the
P2′-P3′ amide or through a glycine residue (74a -f)
showed good WBA potency but lacked oral activity.
These results further confirm our previous observations
that arylamide- or heteroarylamide-containing ana-
logues have low oral anti-TNF-R activity.
Keeping the P3′ glycine residue constant, we next
examined a variety of heterocycles including morpho-
line, piperazine, and piperidine that form a tertiary
amide bond with the glycine residue at P4′. With a
glycine residue at P3′, a morpholino residue at P4′
afforded one of the most potent inhibitors, 68y (SP057),
with an IC50 of 67 nM in WBA. More importantly, SP057
was highly potent in the inhibition of TNF-R release
when administered orally to mice, with an ED50 of 2.6
mg/kg. Dimethyl substitution at the 2,6-positions on the
morpholine (68z) decreased the in vitro and in vivo
potencies. Replacement of the morpholine in SP057 with
a piperazine or piperidine derivative (68a a -h h ) re-
sulted in similar or less WBA potency. Although the
N-phenylpiperazino analogue 68bb and the N-2-py-
ridylpiperazino analogue 68cc were equipotent to SP057
in WBA, they were less active orally. However, despite
the 4-fold drop in WBA potency when the morpholine
in SP057 was replaced with 4-hydroxypiperidine, the
resulting analogue 68ee exhibited oral activity (ED50
) 3.6 mg/kg) comparable to that of SP057. The similar
WBA potencies between the 4-carboxypiperidine ana-
logue 68gg and the neutral piperidine analogue 68ee
or the basic piperidine analogue 68ff suggest that a
carboxylic acid at P4′ is tolerated for whole blood
activity.
TACE Activity a n d Selectivity P r ofile of SL422
a n d SP 057. SL422 and SP057, representing the cyclo-
phane series and the cyclic carbamate series, respec-
To further assess the importance of the P3′ glycine
residue to in vitro and in vivo activity, three more

2646 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
Ta ble 6. Inhibitory Profiles for SL422 and SP057
Ki, nMa
compd
pTAC E
MMP-1
MMP-2
MMP-3
MMP-7
MMP-8
MMP-9
MMP-13
MMP-14
MMP-15
MMP-16
SL422
SP057
12
4.2
3.0
13
1.1
1.1
14
7.8
0.8
1.5
0.5
1.1
0.9
1.2
0.45
2.3
94
140
20
71
35
107
a
Ki values are from five determinations for TACE and three determinations for MMPs.
Ta ble 7. Pharmacokinetics of SL422 and SP057 in Beagle Dogs
compd
SL422
dose, mg/kg
T1/2, h
Cl, L/h/kg
0.42
Vss, L/kg
Cmax, ng/mL
Tmax, h
AUC, ng‚h/mL
F, %
0.74 (iv)
0.863 (po)
0.5 (iv)
0.81
1.29
1.0
0.70
121
0.88
2613
342
2080
1910
11
SP057
0.29
0.33
667
1.25
23
2.0 (po)
2.2
tively, were tested in the porcine TACE assay. As seen
in Table 6, SP057 and SL422 are potent against TACE,
with Ki values of 4.2 and 12 nM, respectively. The 3-fold
better TACE potency of SP057 versus SL422 is consis-
tent with what we have observed with these two
compounds in WBA, where SP057 is about 3-fold more
potent than SL422. To gain insight into the selectivity
of TACE over MMPs, SL422 and SP057 were screened
against a panel of 10 MMPs. As shown in Table 6, these
two inhibitors exhibited high potency against MMP-1,
MMP-2, MMP-3, MMP-7, MMP-8, MMP-9, and MMP-
13 and lesser potency against MMP-14, MMP-15, and
MMP-16. These results indicate that SL422 and SP057
are nonselective TACE inhibitors. Since excessive TNF-R
production has been noted in several disease conditions
that are also characterized by MMP-mediated tissue
degradation, it is likely that these nonselective inhibi-
tors that inhibit both TACE and MMPs may provide
some advantage in the treatment of diseases where both
mechanisms are involved.
P h a r m a cok in etics of SL422 a n d SP 057 in Bea gle
Dogs. SL422 and SP057 were selected for pharmaco-
kinetic studies in beagle dogs (Table 7). Both compounds
showed a short half-life following iv or oral dosing, with
T1/2 values of 0.81 h (iv) and 1.29 h (po) for SL422 and
1.0 h (iv) and 2.2 h (po) for SP057. After oral dosing,
both compounds were rapidly absorbed, with Tmax
values of 0.88 and 1.25 h for SL422 and SP057,
respectively. The oral bioavailability (F) was 11% and
23% for SL422 and SP057, respectively. The low bio-
availability of both compounds may reflect an incom-
plete oral absorption rather than presystematic me-
tabolism as indicated by their low clearances.
accommodate a broad array of functional groups includ-
ing polar residues, hydrophobic residues, and amino and
carboxylic acid moieties, suggesting that, like the P1-
P2′ area, the TACE P3′-P4′ area might also be a
solvent-exposed area. While a number of structurally
diversified residues at P3′ are able to confer good in vitro
potency in WBA, in both series, a glycine residue at P3′
was identified as a critical structural component to
achieve both good in vitro potency and good oral activity.
With a glycine residue at P3′, an N-methylamide at P4′
provided the best cyclophane analogue, SL422 (WBA
IC50 ) 0.22 µM, LPS-mouse ED50 ) 15 mg/kg, po),
whereas a morpholinylamide at P4′ afforded the most
potent and most orally active cyclic carbamate analogue,
SP057 (WBA IC50 ) 0.067 µM, LPS-mouse ED50 ) 2.3
mg/kg, po). Although replacement of the P1′ isobutyl in
the cyclophane series with a longer alkyl (n-hexyl) had
positive and negative effects on WBA potency depending
on the P3′-P4′ residues, it was clearly demonstrated
that a longer, hydrophobic group at P1′ resulted in lower
oral activity. N-Methyl substitution at the carbamate
moiety in the cyclic carbamate series offered no advan-
tage in WBA and oral activities.
Further profiling for SL422 and SP057 showed that
these macrocyclic compounds are potent TACE inhibi-
tors, with Ki values of 12 and 4.2 nM in the porcine
TACE assay, and are broad-spectrum MMP inhibitors.
Pharmacokinetic studies in beagle dogs revealed that
SL422 and SP057 are orally bioavailable, with F ) 11%
and 23%, respectively. The antiarthritic efficacy of these
two compounds has been studied in animal models, and
the results will be published in due course.
Exp er im en ta l Section
Con clu sion
Melting points were determined on a Thomas-Hoover melt-
ing point apparatus and are uncorrected. Analytical HPLC was
run on a Vydac C18 column (4.6 × 250 mm) operated at room
temperature, eluting at 1 mL/min using a linear gradient of
water (0.05% TFA)/acetonitrile (0.05% TFA) from 90:10 to 20:
80 over 30 min. Preparative HPLC was performed on a Vydac
C18 column (22 × 250 mm) operated at room temperature,
eluting at 8 mL/min with a linear gradient of water (0.1%
TFA)/acetonitrile (0.1% TFA) from 100:0 to 0:100 over 80 min.
We designed a new class of macrocyclic hydroxamic
acids by linking the P1 and P2′ residues of acyclic anti-
succinate-based hydroxamic acids. A variety of residues
including amide, carbamate, alkyl, sulfonamido, Boc-
amino, and amino were found to be suitable P1-P2′
linkers. With an N-methylamide at P3′, the 13-16-
membered macrocycles exhibited moderate inhibition of
TNF-R release in WBA (IC50 > 1 µM). The similar whole
blood potencies observed among the majority of the
cyclic templates prepared suggest that, like MMPs, the
TACE P1-P2′ area might be a solvent-exposed area.
To further improve the whole blood activity, we
selected the cyclophane and cyclic carbamate templates
for refinement and optimization in the P3′-P4′ area.
Our SAR studies revealed that the P3′-P4′ area can
1
UV detection was at 220 nm. H NMR data were obtained at
300 MHz using a Varian VXR400 spectrometer and were
referenced to TMS. Mass spectral data were obtained on either
a VG 70-VSE (FAB, high-resolution FAB, high-resolution DCI)
or a Finnigan MAT 8230 (DCI) mass spectrometer. Combus-
tion analyses were performed by Quantitative Technologies,
Inc., Bound Brook, NJ . Solvents and reagents were used as
purchased from commercial suppliers unless otherwise indi-
cated. Yields quoted herein are isolated yields.

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2647
(4S)-4-Ben zyl-3-(4-m eth ylp en ta n oyl)-1,3-oxa zolid in -2-
on e (1). To a stirred solution of 4(S)-(phenylmethyl)-2-oxazo-
lidinone (48.3 g, 272 mmol) in THF (500 mL) cooled to -78 °C
was added 2.5 M n-butyllithium (131 mL, 327 mmol) in hexane
over 20 min, and stirring was continued at -78 °C for 45 min.
To it was added 4-methylpentanoyl chloride (44 g, 327 mmol).
The solution was allowed to warm to room temperature, and
stirring was continued for 2.5 h. The volatiles were removed
under reduced pressure, and EtOAc (500 mL) was added. The
resulting solution was washed with 10% citric acid, brine,
NaHCO3, and brine, dried (MgSO4), and concentrated. Puri-
fication by flash chromatography on silica eluting with 10%
EtOAc/hexanes yielded the desired product (68.53 g, 91.5%)
as an oil: MS m/z 276.3 (M + H)+. Anal. (C16H21NO3) C, H, N.
Bu tyl (3R)-3-{[(4S)-4-Ben zyl-2-oxo-1,3-oxa zolid in -3-yl]-
ca r bon yl}-2-h yd r oxy-5-m eth ylh exa n oa te (2). To a stirred
solution of diisopropylamine (3.25 mL, 23.25 mmol) in THF
(20 mL) cooled to -78 °C was added 2.5 M n-butyllithium (9.3
mL, 23.25 mmol) in hexane. The solution was allowed to stir
at 0 °C for 30 min and then cooled back to -78 °C. To it was
added a solution of 1 (5.82 g, 21.13 mmol) in THF (50 mL)
cooled at -78 °C. After the resulting solution was stirred at
-78 °C for 1 h, a solution of n-butyl glyoxalate (4.12 g, 31.69
mmol) in THF (10 mL) was added. The mixture was stirred
at -78 °C for 3 h, and the reaction was quenched with ice
water. Ethyl acetate was added followed by 10% citric acid
solution. The organic layer was separated, washed with water,
NaHCO3, and brine, dried (MgSO4), and concentrated. Puri-
fication on silica gel eluting gradually with 5% EtOAc/hexanes,
10% EtOAc/hexanes, and 20% EtOAc/hexanes provided com-
pound 2 (3.1 g, 36%) as a mixture of two diastereomers (3:1):
MS m/z 406.2 (M + H)+. Anal. (C22H31NO6) C, H, N.
4-Ben zyl 1-Bu tyl (3R)-2-Hyd r oxy-3-isobu tylbu ta n ed io-
a te (3). To a solution of 2 (5.1 g, 12.57 mmol) in THF/H2O
(250 mL, 4:1) cooled in an ice bath was added hydrogen
peroxide (7.84 mL, 50.3 mmol) followed by a solution of LiOH
(791 mg, 18.85 mmol) in water (8 mL). After the resulting
solution was stirred for 1 h, the reaction was quenched with a
solution of Na2SO3 (6.33 g, 50.28 mmol) in water. THF was
removed under reduced pressure, and the solution was ex-
tracted with ethyl acetate twice. The water layer was acidified
with cold concentrated HCl to pH 3 and extracted with CH2-
Cl2 three times. The combined CH2Cl2 layer was washed with
water and brine, dried (MgSO4), and concentrated. Silica gel
chromatography eluting gradually with CHCl3, 5% MeOH/
CHCl3, and 10% MeOH/CHCl3 provided the carboxylic acid
(2.29 g, 74%) as a mixture of two diastereomers (3:1): MS
(NH3, CI) m/z 264.0 (M + NH4)+.
A mixture of the above carboxylic acid (8.33 g, 33.82 mmol),
benzyl bromide (7.0 g, 37.2 mmol), and DBU (6.07 mL, 40.58
mmol) in benzene (100 mL) was stirred at 50 °C for 3 h.
Benzene was removed under reduced pressure. The residue
was taken up in EtOAc. The solution was washed with brine
3×, dried (MgSO4), and concentrated. Flash chromatography
on a silica gel column eluting with 10% EtOAc/hexanes
provided 3 (9 g, 79%) as a mixture of two diastereomers (ratio
3:1): MS (ESI) m/z 337.3 (M + H)+. Anal. (C19H28O5) C, H, N.
catalyst was filtered off, and the solution was concentrated to
give 5 (3.96 g, 99%) as a mixture of two diastereomers (3:1):
MS (ESI) m/z 361.4 (M + H)+.
{[(2R)-2-[(Ben zyloxy)ca r bon yl]-1-(bu toxyca r bon yl)-4-
m eth ylp en tyl]oxy}a cetic Acid (6). To a solution of 4 (2.5
g, 5.55 mmol) in CH2Cl2 (25 mL) was added TFA (25 mL). The
solution was stirred at room temperature for 4 h and concen-
trated to give 6 (2.45 g) as a mixture of two diastereomers (3:
1): MS (ESI) m/z 395.3 (M + H)+.
ter t-Bu t yl (9S,12R)-13-(Bu t oxyca r b on yl)-12-isob u t yl-
9-[(m eth yla m in o)ca r bon yl]-3,11-d ioxo-1-p h en yl-2,14-d i-
oxa -4,10-d ia za h exa d eca n -16-oa te (7). To a solution of NR-
(tert-butoxycarbonyl)-Nꢀ-[(benzyloxy)carbonyl]-L-lysine (12.39
g, 32 mmol) and methylamine hydrochloride (4.4 g, 65 mmol)
in DMF (30 mL) cooled in an ice bath was added BOP (14.16
g, 32 mmol) followed by DIEA (25 mL, 128 mmol). The solution
was allowed to stir at room temperature overnight. EtOAc (250
mL) was added, and the solution was washed with 10% citric
acid, brine, saturated NaHCO3, and brine, dried (MgSO4), and
concentrated. Purification on a silica gel column eluting with
80% EtOAc/hexanes yielded the N-methylamide (12.92 g, 95%)
as a solid: MS (ESI) m/z 394.4 (M + H)+.
The above N-methylamide (6 g, 15.26 mmol) was dissolved
in 4 N HCl/dioxane (50 mL). After being stirred at room
temperature for 1 h, the solution was concentrated under
reduced pressure. The residue was triturated with ether to
give the Boc-deprotected product (5.2 g, 100%) as a solid: MS
(ESI) m/z 294.3 (M + H)+.
Compound 5 (1.76 g, 4.88 mmol) and the above lysine
N-methylamide (1.61 g, 4.88 mmol) were dissolved in DMF
(10 mL), and the solution was cooled in an ice bath. To it was
added BOP (2.16 g, 4.88 mmol) followed by DIEA (3.42 mL,
10.58 mmol). The mixture was allowed to stir at room
temperature for 4 h. EtOAc was added, and the solution was
washed with 10% citric acid, brine, NaHCO3, and brine, dried
(MgSO4), and concentrated. Flash chromatography on a silica
gel column eluting with 10% MeOH/CHCl3 provided 7 (2.32
g, 75%) as a mixture of two diastereomers (3:1): MS (ESI) m/z
636.6 (M + H)+.
Bu tyl (9S,12R,13S)-12-Isobu tyl-9-[(m eth yla m in o)ca r -
bon yl]-3,11-d ioxo-1-oxa -4,10-d ia za cyclotr id eca n e-13-ca r -
boxyla te (8). To a solution of 7 (2.21 g, 3.47 mmol) in
2-propanol (30 mL) and 4 N HCl/dioxane (1 mL) was added
10% Pd/C (0.35 g). The mixture was stirred under H2 (balloon)
for 2 h. The catalyst was filtered off, and the solution was
concentrated under reduced pressure to give a solid. The solid
was dissolved in 4 N HCl/dioxane (30 mL). The solution was
stirred for 2 h and concentrated under reduced pressure. The
residue was taken up in DMF (5 mL). This solution was slowly
added to a solution of BOP (1.64 g, 3.7 mmol) and DIEA (2.6
mL, 14.6 mmol) in CHCl3 (100 mL) cooled in an ice bath over
a period of 2 h. The solution was allowed to stir at room
temperature overnight and concentrated. The residue was
taken up in EtOAc, and the solution was washed with 10%
citric acid, brine, NaHCO3, and brine, dried (MgSO4), and
concentrated to give the crude product as a mixture of two
diastereomers. The two diastereomers were separated by flash
chromatography on a silica gel column eluting with 15%
MeOH/CH2Cl2 to give the major diastereomer 8 (0.8 g, 50%)
as a solid: MS (ESI) m/z 428.3 (M + H)+.
4-Ben zyl 1-Bu tyl (3R)-2-(2-ter t-Bu toxy-2-oxoeth oxy)-3-
isobu tylbu ta n ed ioa te (4). To a solution of 3 (8.95 g, 26.64
mmol) and tert-butyl bromoacetate (4.33 mL, 29.3 mmol) in
THF (50 mL) cooled in an ice bath was added NaH (1.5 g, 60%
oil dispersion, 32 mmol) in batches. The mixture was stirred
in the ice bath for 30 min and at room temperature for 2 h.
The reaction was terminated by quenching with 10% citric acid
solution. EtOAc was added, and the solution was washed with
10% citric acid and brine, dried (MgSO4), and concentrated.
The residue was purified by flash chromatography on a silica
gel column to give 4 (8.6 g, 71%) as a mixture of two
diastereomers (3:1): MS (ESI) m/z 451.4 (M + H)+. Anal.
(C25H38O7) C, H, N.
(9S,12R,13S)-N13-Hyd r oxy-12-isobu tyl-N9-m eth yl-3,11-
d ioxo-1-oxa -4,10-d ia za cyclotr id eca n e-9,13-d ica r boxa m -
id e (9). To a solution of 8 (0.77 g, 1.8 mmol) in THF (10 mL)
was added 1 N LiOH (4 mL). The solution was stirred at room
temperature for 2 h and acidified with 4 N HCl/dioxane to pH
3. The volatiles were removed under reduced pressure, and
tert-butyl alcohol was added. The organic phase was separated,
washed with brine 3×, dried (MgSO4), and concentrated to give
the carboxylic acid (0.49 g, 73%) as a powder: MS (ESI) m/z
372.2 (M + H)+.
(2R)-2-[2-Bu toxy-1-(2-ter t-bu toxy-2-oxoeth oxy)-2-oxo-
eth yl]-4-m eth ylp en ta n oic Acid (5). To a solution of 4 (5 g,
11.11 mmol) in MeOH (50 mL) was added 10% Pd/C (0.5 g).
The mixture was stirred under H2 (balloon) for 3 h. The
To a solution of the above carboxylic acid (0.47 g, 1.27 mmol)
and O-benzylhydroxylamine hydrochloride (0.2 g, 1.27 mmol)
in DMF (5 mL) cooled in an ice bath was added BOP (0.56 g,
1.27 mmol) followed by DIEA (1.0 mL, 5.2 mmol). The solution

2648 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
was allowed to stir at room temperature overnight. EtOAc was
added, and the solution was washed with 10% citric acid, brine,
NaHCO3, and brine, dried (MgSO4), and concentrated. Puri-
fication on a silica gel column eluting with 5% MeOH/CH2Cl2
provided the O-benzylhydroxamate (0.21 g, 35%) as a solid:
MS (ESI) m/z 477.3 (M + H)+.
The O-benzylhydroxamate (100 mg, 0.21 mmol) was dis-
solved in MeOH (10 mL), and 10% Pd/C (15 mg) was added.
The mixture was stirred under H2 (balloon) for 2 h. The
catalyst was filtered off, and the solution was concentrated.
The residue was purified by reversed-phase HPLC to give 9
(50 mg, 62%) as a powder: 1H NMR (DMSO-d6) δ 10.95 (s,
1H), 9.20 (br, 1H), 8.31 (d, 1H), 7.84 (q, 1H), 6.96 (t, 1H), 4.25
(m, 1H), 3.92 (d, 1H), 3.85 (d, 1H), 3.60 (d, 1H), 2.95 (m, 2H),
2.62 (d, 3H), 2.48 (m, 1H), 1.8-1.2 (m, 9H), 0.92 (d, 3H), 0.84
(d, 3H); MS (ESI) m/z 387.3 (M + H)+. Anal. (C17H30N4O6) C,
H, N.
ter t-Bu tyl (1S)-1-[(Meth yla m in o)ca r bon yl]-5-[m eth yl-
(tr iflu or oa cetyl)a m in o]p en tylca r ba m a te (11). To a solu-
tion of NR-[(tert-butyloxy)carbonyl]-Nꢀ-(trifluoroacetyl)-L-lysine
(10) (10.27 g, 30 mmol) and methylamine hydrochloride (4.05
g, 60 mmol) in DMF (30 mL) cooled in an ice bath was added
BOP (13.27 g, 30 mmol) followed by DIEA (23.5 mL, 135
mmol). The mixture was stirred at room temperature over-
night. EtOAc (500 mL) was added. The solution was washed
with 10% citric acid, brine, NaHCO3, and brine, dried (MgSO4),
and concentrated to give a solid. Recrystallization from EtOAc/
Et2O provided the N-methylamide (10.1 g, 94%) as a crystal:
mp 95-98 °C; MS (ESI) m/z 356.3 (M + H)+.
A mixture of the above N-methylamide (9.9 g, 27.88 mmol),
iodomethane (13.88 mL, 223 mmol), and K2CO3 (7.7 g, 55.8
mmol) in DMF (50 mL) was stirred at 100 °C for 24 h. After
the mixture was cooled to room temperature, insoluble materi-
als were filtered off. The filtrate was diluted with EtOAc (500
mL). The resulting solution was washed with brine 3×, dried
(MgSO4), and concentrated. The residue was purified by flash
chromatography on silica eluting with 5% MeOH/CH2Cl2 to
give 11 (4.45 g, 43%) as a solid: MS (ESI) m/z 370.2 (M +
H)+. Anal. (C15H26N3O4F3) C, H, N.
ter t-Bu tyl (1S)-1-[(Meth yla m in o)ca r bon yl]-5-(m eth y-
la m in o)p en tylca r ba m a te (12). To a solution of 11 (4.35 g,
11.78 mmol) in MeOH (20 mL) was added 1 N LiOH (14.5 mL).
The solution was stirred at room temperature for 30 min and
concentrated. The residue was taken up in CHCl3, and
insoluble materials were filtered off. The filtrate was dried
(MgSO4) and concentrated to give 12 (3.65 g, 100%) as an oil:
MS (ESI) m/z 274.5 (M + H)+.
4-Ben zyl 1-Bu tyl (3R)-2-{2-[[(5S)-5-[(ter t-bu toxyca r bo-
n yl)a m in o]-6-(m eth yla m in o)-6-oxoh exyl]m eth yla m in o]-
2-oxoeth oxy}-3-isobu tylbu ta n ed ioa te (13). To a solution
of 6 (3.06 g, 7.77 mmol) and 12 (2.4 g, 7.77 mmol) in DMF (15
mL) cooled in an ice bath was added BOP (3.44 g, 7.77 mmol)
followed by DIEA (4.74 mL, 27 mmol). The mixture was stirred
at room temperature overnight. EtOAc was added, and the
solution was washed with citric acid, brine, NaHCO3, and
brine, dried (MgSO4), and concentrated. Flash chromatography
on a silica gel column eluting with 5% MeOH/CH2Cl2 provided
13 (4.46 g, 93%) as a mixture of two diastereomers (3:1): MS
(ESI) m/z 650.7 (M + H)+. Anal. (C34H55N3O9) C, H, N.
Bu t yl (9S,12R,13S)-12-Isob u t yl-4-m et h yl-9-[(m et h y-
la m in o)ca r bon yl]-3,11-d ioxo-1-oxa -4,10-d ia za cyclotr id e-
ca n e-13-ca r boxyla te (14). Compound 13 (4.31 g, 6.98 mmol)
was dissolved in 4 N HCl in dioxane (50 mL). After being
stirred for 1 h, the solution was concentrated. The residue was
taken up in 2-propanol (60 mL), and 10% Pd-C (0.5 g) was
added. The mixture was stirred under H2 (balloon) for 2 h.
The catalyst was filtered off, and the solvent was removed
under reduced pressure to give a solid. The solid was dissolved
in CHCl3 (20 mL). This solution was slowly added to a solution
of BOP (2.68 g, 6.05 mmol) and DIEA (3.69 mL, 21.2 mmol)
in CHCl3 (200 mL) cooled in an ice bath over 1 h. The mixture
was stirred at room temperature overnight. The volatiles were
removed under reduced pressure. The residue was taken up
in EtOAc, and the solution was washed with citric acid, brine,
NaHCO3, and brine, dried (MgSO4), and concentrated to give
the crude product as a mixture of two diastereomers which
were separated by flash chromatography on silica gel eluting
with 5% MeOH/CH2Cl2, affording the major diastereomer 14
(1 g, 38%) as a solid: MS (ESI) m/z 442.5 (M + H)+.
(9S,12R,13S)-N13-H yd r oxy-12-isob u t yl-N9,4-d im et h yl-
3,11-d ioxo-1-oxa -4,10-d ia za cyclotr id eca n e-9,13-d ica r box-
a m id e (15). To a solution of 14 (0.25 g, 0.566 mmol) in THF
(5 mL) was added 1 N LiOH (0.7 mL). The solution was stirred
at room temperature for 1 h, acidified with 1 N HCl solution,
and concentrated. Purification by reversed-phase HPLC pro-
vided the carboxylic acid (180 mg, 85%) as a powder: MS (ESI)
m/z 386.3 (M + H)+.
To a solution of the above carboxylic acid (0.17 g, 0.48 mmol)
and O-benzylhydroxylamine hydrochloride (91 mg, 0.576
mmol) in DMSO (2 mL) cooled in an ice bath was added BOP
(254 mg, 0.576 mmol) followed by DIEA (0.33 mL, 1.92 mmol).
The solution was stirred at room temperature for 1 h. Purifica-
tion by reversed-phase HPLC provided O-benzylhydroxamate
(170 mg, 79%) as a powder: MS (ESI) m/z 491.6 (M + H)+.
The above O-benzylhydroxamate was hydrogenated follow-
ing the procedure described for 9 to provide the hydroxamic
acid 15: 1H NMR (DMSO-d6) δ 10.90 (s, 1H), 8.08 (d, 1H), 7.68
(q, 1H), 4.50 (m, 1H), 4.40 (m, 1H), 3.80 (dd, 2H), 3.60 (m, 2H),
3.00 (m, 1H), 2.70 (s, 3H), 2.58 (d, 3H), 1.65 (m, 2H), 1.5-1.2
(m, 7H), 0.85-75 (m, 6H); MS (ESI) m/z 401.6 (M + H)+. Anal.
(C18H32N4O6‚0.5H2O‚0.3CF3CO2H) C, H, N.
(2S)-2-[(ter t-Bu toxycar bon yl)am in o]-3-[(2-n itr oph en yl)-
su lfa n yl]p r op a n oic Acid (17). A suspension of 2-chloroni-
trobenzene (7.88 g, 50 mmol), L-cysteine (6.66 g, 55 mmol),
and potassium carbonate (7.6 g, 55 mmol) in DMF (30 mL)
was stirred at 80 °C for 4 h. After the suspension was cooled
to room temperature, water (20 mL) was added and the
solution was cooled in an ice bath. To it was added di-tert-
butyl dicarbonate (10.9 g, 50 mmol). The solution was stirred
for 2 h, and water (100 mL) was added. The resulting solution
was extracted with ether 3×. The water layer was acidified
with 1 N HCl at 0 °C and extracted with ethyl acetate 3×.
The combined organic phase was washed with brine, dried
(MgSO4), and concentrated to give 17 (8.21 g, 48%) as a solid:
MS (ESI) m/z 343.2 (M + H)+.
(2S)-2-[(ter t-Bu t oxyca r b on yl)a m in o]-N-m et h yl-3-[(2-
n itr op h en yl)su lfa n yl]p r op a n a m id e (18). To a solution of
17 (8.1 g, 23.66 mmol) and methylamine hydrochloride (2.03
g, 30 mmol) cooled in an ice bath was added DIEA (16.5 mL,
95 mmol) followed by BOP (10.47 g, 23.66 mmol). After the
resulting solution was stirred for 2 h at room temperature,
EtOAc was added and the solution was washed with citric acid,
brine, NaHCO3, and brine, dried (MgSO4), and concentrated.
Purification on a silica gel column eluting with 5% MeOH/
CH2Cl2 provided 18 (6.24 g, 82%) as a solid: MS (ESI) m/z
356.3 (M + H)+.
(2S)-2-Am in o-N-m eth yl-3-[(2-n itr oph en yl)su lfa n yl]p r o-
p a n a m id e (19). Compound 18 (6.0 g, 17 mmol) was dissolved
in 4 N HCl in dioxane (50 mL). After being stirred for 1 h, the
solution was concentrated. The residue was triturated with
ether to give 19 (3.88 g, 71%) as a solid: MS (ESI) m/z 256.1
(M + H)+.
Bu t yl (3R)-2-(2-ter t-Bu t oxy-2-oxoet h oxy)-5-m et h yl-3-
{[((1S)-2-(m eth ylam in o)-1-{[(2-n itr oph en yl)su lfan yl]m eth -
yl}-2-oxoeth yl)a m in o]ca r bon yl}h exa n oa te (20). To a solu-
tion of 5 (2.36 g, 6.5 mmol) and 19 (1.91 g, 6.5 mmol) in CHCl3
(15 mL) cooled in an ice bath was added DIEA (4.53 mL, 26
mmol) followed by BOP (2.88 g, 6.5 mmol). The solution was
stirred at room temperature overnight and concentrated. The
residue was taken up in EtOAc, and the solution was washed
with citric acid, brine, NaHCO3, and brine, dried (MgSO4), and
concentrated. Flash chromatography on a silica gel column
eluting with 3% MeOH/25% EtOAc/72% CH2Cl2 provided 20
(3.21 g, 83%) as a mixture of two diastereomers (3:1): MS (ESI)
m/z 598.6 (M + H)+.
{[(2R)-2-({[(1S)-1-{[(2-Am in op h en yl)su lfa n yl]m eth yl}-
2-(m eth yla m in o)-2-oxoeth yl]a m in o}ca r bon yl)-1-(bu toxy-
ca r bon yl)-4-m eth ylp en tyl]oxy}a cetic Acid (21). To a so-

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2649
lution of 20 (3.05 g, 5.1 mmol) in acetic acid (15 mL) and water
(0.5 mL) was added zinc (3 g). The mixture was stirred at room
temperature for 30 min. Methanol was added, and insoluble
materials were filtered off. The filtrate was concentrated under
reduced pressure. The residue was taken up in EtOAc. The
solution was washed with NaHCO3 3×, dried (MgSO4), and
concentrated. The residue was taken up in 4 N HCl in dioxane
(30 mL) in the presence of 0.5 mL of water. After the solution
was stirred for 1 h, the solvent was removed under reduced
pressure to give 21 as a mixture of two diastereomers (3:1):
MS (ESI) m/z 512.5 (M + H)+.
(5S,6R,9S)-N5-Hyd r oxy-6-isobu tyl-N9-m eth yl-2,7-d ioxo-
2,3,5,6,7,8,9,10-octa h yd r o-1H-4,11,1,8-ben zoxa th ia d ia za -
cyclotr id ecin e-5,9-d ica r boxa m id e (22). To a solution of
BOP (1.81 g, 4.1 mmol) in CHCl3 (50 mL) cooled in an ice bath
was added a solution of 21 (2.1 g, 4.1 mmol) and DIEA (2.86
mL, 16.4 mmol) in CHCl3 (20 mL) over a period of 2 h. The
solution was allowed to stir at room temperature overnight.
The solvent was removed under reduced pressure. EtOAc was
added, and the solution was washed with citric acid, brine,
NaHCO3, and brine, dried (MgSO4), and concentrated. Flash
(1.65 g, 3.75 mmol) and DIEA (2.1 mL, 12 mmol) in CHCl3
(50 mL) cooled in an ice bath. The mixture was allowed to stir
at room temperature for 2 h. The solvent was removed under
reduced pressure and the residue taken up in EtOAc. The
EtOAc solution was washed with brine 3×, dried (MgSO4), and
concentrated. Flash chromatography on silica eluting with 3%
MeOH/CH2Cl2 provided the major diastereomer 25 (843 mg,
45%) as a solid: mp 225 ïC; MS (ESI) m/z 500.4 (M + H)+.
(7S,8R,11S)-N7-(Ben zyloxy)-8-isob u t yl-N11-m et h yl-9-
oxo-6-oxa -1,10-d ia za tr icyclo[11.6.1.014,19]icosa -13(20),14,-
16,18-tetr a en e-7,11-d ica r boxa m id e (26). To a solution of
25 (450 mg, 1.05 mmol) in THF (10 mL) was added 1 N LiOH
(2 mL). The solution was stirred for 30 min and acidified with
1 N HCl. EtOAc was added. The solution was washed with
brine 3×, dried (MgSO4), and concentrated. The residue was
taken up in DMF (5 mL), and the solution was cooled in an
ice bath. To it were added O-benzylhydroxamine (207 mg, 1.3
mmol), BOP (575 mg, 1.3 mmol), and DIEA (0.95 mL, 5.2
mmol). The mixture was stirred in the ice bath for 1 h. EtOAc
(100 mL) was added. The resulting solution was washed with
citric acid, brine, NaHCO3, and brine, dried (MgSO4), and
concentrated. Purification by reversed-phase HPLC provided
26 (360 mg, 65%) as a solid: MS (ESI) m/z 549.4 (M + H)+.
(7S,8R,11S)-N7-Hyd r oxy-8-isobu tyl-N11-m eth yl-9-oxo-6-
oxa -1,10-d ia za t r icyclo[11.6.1.014,19]icosa -13(20),14,16,18-
tetr a en e-7,11-d ica r boxa m id e (27). To a solution of 26 (260
mg, 0.474 mmol) in MeOH (20 mL) was added 10% Pd on
carbon (50 mg). The mixture was stirred under H2 (balloon)
for 1.5 h. The catalyst was filtered off, and the solvent was
removed under reduced pressure. Purification by reversed-
phase HPLC provided the hydroxamic acid 27 (188 mg, 86%)
as a powder: 1H NMR (DMSO-d6) δ 10.80 (s, 1H), 8.20 (d, 1H),
7.65 (q, 1H), 7.58 (d, 1H), 7.40 (d, 1H), 7.30 (s, 1H), 7.10 (t,
1H), 7.00 (t, 1H), 4.45 (m, 1H), 4.05 (m, 2H), 3.42 (d, 1H), 3.15
(m, 1H), 2.98 (m, 2H), 2.83 (m, 1H), 2.70 (m, 1H), 2.62 (d, 3H),
2.00 (m, 1H), 1.75 (m, 1H), 1.40 (m, 2H), 1.00 (m, 1H), 0.80
(m, 8H); MS (ESI) m/z 459.4 (M + H)+. Anal. (C24H34N4O5) C,
H, N.
chromatography on
a silica gel column eluting with 1%
2-propanol/10% EtOAc/89% CH2Cl2 provided the cyclized
product (1.45 g, 71%) as a mixture of two diastereomers: MS
(ESI) m/z 494.4 (M + H)+.
A portion of the above cyclized product (0.76 g, 1.53 mmol)
was dissolved in MeOH (10 mL). To it were added HONH2‚
HCl (0.43 g, 6.19 mmol) and DIEA (2 mL, 11.5 mmol). The
mixture was stirred at 50 °C overnight and the solvent
removed under reduced pressure. The residue was purified by
reversed-phase HPLC to provide the major diastereomer 22
(276 mg, 40%) as a powder: 1H NMR (DMSO-d6) δ 10.90 (s,
1H), 9.47 (s, 1H), 8.75 (d, 1H), 8.28 (d, 1H), 7.70 (q, 1H), 7.66
(d, 1H), 7.35 (t, 1H), 7.10 (t, 1H), 4.10 (d, 1H), 3.85 (m, 1H),
3.68 (dd, 2H), 3.00 (m, 3H), 2.45 (d, 3H), 1.35 (m, 2H), 1.20
(m, 1H), 0.80 (d, 3H), 0.70 (d, 3H); MS (ESI) m/z 453.4 (M +
H)+. Anal. (C20H28N4O6S‚CH3CN‚2.8H2O) C, H, N.
4-Ben zyl 1-Bu tyl (3R)-2-{[(2E)-4-Br om o-2-bu ten yl]oxy}-
3-isobu tylbu ta n ed ioa te (23). To a solution of 3 (4.03 g, 12
mmol) in DMF (24 mL) cooled in an ice bath was added NaH
(691 mg, 60% dispersion in oil, 14.4 mmol). The mixture was
stirred for 15 min, at which time 1,4-dibromo-2-butene (10.2
g, 48 mmol) was added. After the resulting mixture was stirred
in the ice bath for 20 min, EtOAc (200 mL) was added. The
solution was washed with brine 3×, dried (MgSO4), and
concentrated. Flash chromatography on a silica gel column
eluting with 10% EtOAc/hexanes provided 23 (3.64 g, 65%) as
a mixture of two diastereomers (3:1): MS (ESI) m/z 471.2 (M
+ H)+. Anal. (C23H33O5Br) C, H, N.
4-Ben zyl 1-Bu tyl (3R)-2-[((2E)-4-{3-[(1S)-1-[(ter t-Bu tox-
ycar bon yl)am in o]-2-(m eth ylam in o)-2-oxoeth yl]-1H-in dol-
1-yl}-2-bu ten yl)oxy]-3-isobu tylbu ta n ed ioa te (24). To a
solution of NR-Boc-L-tryptophan N-methylamide (1.96 g, 6.2
mmol) in DMF (14 mL) cooled in an ice bath was added NaH
(446 mg, 9.3 mmol) with vigorous stirring. After the resulting
solution was stirred for 15 min, compound 23 (2.9 g, 6.2 mmol)
was added. The mixture was allowed to stir in the ice bath for
1 h. EtOAc (200 mL) was added. The solution was washed with
citric acid and brine, dried (MgSO4), and concentrated. Puri-
fication on a silica gel column eluting with 5% MeOH/CH2Cl2
provided 24 (2.7 g, 62%) as a mixture of two diastereomers
(3:1): MS (ESI) m/z 706.4 (M + H)+. Anal. (C40H55N3O8) C, H,
N.
Bu tyl (7S,8R,11S)-8-Isobu tyl-11-[(m eth yla m in o)ca r bo-
n yl]-9-oxo-6-oxa -1,10-d ia za tr icyclo[11.6.1.014,19]icosa -13-
(20),14,16,18-tetr a en e-7-ca r boxyla te (25). To a solution of
24 (2.65 g, 3.75 mmol) in 2-propanol (40 mL) was added 10%
Pd on carbon (265 mg). The mixture was stirred under H2
(balloon) for 4 h. The catalyst was filtered off, and the solvent
was removed under reduced pressure to give the carboxylic
acid (2.1 g, 91%) as a solid. The solid was dissolved in 4 N
HCl in dioxane (20 mL). The solution was stirred for 1.5 h
and concentrated. The residue was taken up in CHCl3 (15 mL).
The resulting solution was added dropwise to a solution of BOP
(2R,3S)-3-(ter t-Bu t oxyca r b on yl)-2-isob u t yl-5-h exen o-
ic Acid (29a ). To a stirred solution of (2S)-2-(2-tert-butoxy-
2-oxoethyl)-4-methylpentanoic acid (28a ) (20 g, 87 mmol) in
anhydrous THF (400 mL) cooled at -78 °C was added 1 M
LDA in THF (180 mL) over 20 min. After the resulting solution
was stirred for 1 h, 8.3 mL (96 mmol) of allyl bromide was
added dropwise. The mixture was allowed to slowly warm to
ambient temperature and stirred overnight. The reaction was
quenched with 10% aqueous citric acid followed by removal of
the volatiles under reduced pressure. The remaining material
was taken into EtOAc and the solution washed with water.
The aqueous phase was extracted 3× with EtOAc, and the
combined organic phase was washed with 10% citric acid,
saturated NaHCO3 (2×), H2O (2×), and brine, and dried over
MgSO4. The solvent was removed under reduced pressure,
affording 23.3 g (99%) of the acid that was carried on without
purification. Proton NMR indicated a 1:10 ratio of two dia-
stereomers (anti/syn): MS (ESI) m/z 293 (M + Na)+.
To a stirred solution of the above acid (2 g, 7.4 mmol) in
anhydrous THF (25 mL) cooled at -78 °C was added LDA (16.3
mmol) over 15 min. The reaction was stirred at -78 °C for 15
min and in a water bath (24 °C) for 15 min. After the reaction
was cooled back to -78 °C, a solution of 1 M diethylaluminum
chloride in hexane (25.6 mL) was added. The reaction was
stirred for 10 min at -78 °C and for 15 min in an ambient
temperature water bath. After being cooled back to -78 °C,
the reaction was quenched with the rapid addition of methanol.
The mixture was concentrated to ∼1/4 its original volume
under reduced pressure, and the resulting material was
dissolved in EtOAc (200 mL). The EtOAc solution was washed
with 1 N HCl (70 mL) containing 100 g of ice. The aqueous
phase was extracted 2× with EtOAc. The combined organic
phase was washed with a solution of KF (3.5 g) in water (100
mL) containing 1 N HCl (15 mL), and with brine and dried
over MgSO4, and the volatiles were removed under reduced

2650 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
pressure, affording 1.84 g (92%) of 29a . 1H NMR indicated an
8:1 ratio of two diastereomers (anti:syn): MS (ESI) m/z 293
(M + Na)+.
off and the solution was concentrated to yield the amino acid
35 (32 mg, 90%) as a white foam: MS (ESI) m/z 444.4 (M +
H)+.
ter t-Bu tyl (6S,9R,10S)-9-Isobu tyl-6-[(m eth yla m in o)ca r -
b on yl]-8,13-d ioxo-1,7-d ia za cyclot r id eca n e-10-ca r b oxy-
la te (36). To a solution of HBTU (769 mg, 2.0 mmol) and NMM
(0.15 mL, 6.0 mmol) in DMF (10 mL) at 60 °C was added a
solution of 35 (200.0 mg, 0.4 mmol) in DMF (10 mL) dropwise.
After the addition was complete, the mixture was stirred for
an additional 30 min. The solution was concentrated, and silica
gel chromatography afforded the lactam 36 (135 mg, 70%) as
a light yellow solid: MS (ESI) m/z 426.2 (M + H)+.
4-Ben zyl 1-ter t-Bu tyl (2S,3R)-2-Allyl-3-isobu tylbu ta n e-
d ioa te (30a ). To a stirred solution of 29a (20.6 g, 76 mmol)
in benzene (75 mL) cooled in an ice bath was added DBU (11.4
mL, 76 mmol) followed by benzyl bromide (9.98 g, 84 mmol).
After 10 min the mixture was heated to reflux for 4 h. The
reaction was then cooled and diluted to 3× its original volume
with EtOAc. This was washed 3× with 10% aqueous citric acid.
The combined aqueous phase was extracted 3× with EtOAc.
The combined organic phase was then washed with brine and
dried over MgSO4, and the volatiles were removed under
reduced pressure. The resulting material was chromato-
graphed on silica gel eluting with 2.2% EtOAc/hexanes,
affording 16.9 g (62%) of the benzyl ester 30a : MS (CI, NH3)
m/z 378 (M + NH4)+.
4-Ben zyl 1-ter t-Bu t yl (2S,3R)-2-(3-Hyd r oxyp r op yl)-3-
isobu tylbu ta n ed ioa te (31a ). To a stirred solution of 30a (5.2
g, 14.4 mmol) in anhydrous THF (100 mL) at 0 °C was added
0.5 M 9-BBN in THF (72.2 mL) over 1 h. The reaction was
allowed to warm to room temperature over 12 h. After the
reaction was cooled to 0 °C, H2O (2.9 mL) was added (caution
foaming) dropwise over 5 min, and after the reaction was
stirred for an additional 20 min, H2O (8 mL) containing NaOAc
(3.21 g) was added simultaneously with 30% H2O2 (8 mL) over
5 min. The mixture was stirred for 20 min, followed by removal
of the volatiles under reduced pressure. The remaining mate-
rial was dissolved in EtOAc and the solution washed with
brine. The aqueous layer was extracted twice with EtOAc. The
combined organic phase was washed with water and brine and
then dried over MgSO4. The volatiles were removed under
reduced pressure following filtration. The resulting material
was chromatographed on silica gel eluting gradually with 1:20,
1:10, and 1:5 EtOAc/hexanes, affording 3.5 g (64%) of the
alcohol 31a : MS (ESI) m/z 379 (M + H)+; HRMS m/z
379.250194 [(M + H)+, calcd for C22H35O5 379.248450].
(2S,11S,12R)-N11-(Ben zyloxy)-12-isobu tyl-N2-m eth yl-8,-
13-dioxo-1,7-diazacyclotr idecan e-2,11-dicar boxam ide (37).
The lactam 36 (85 mg, 0.2 mmol) was dissolved in CH2Cl2 (2
mL) and TFA (2 mL). After being stirred overnight, the
solution was concentrated to afford the crude acid. This
material was dissolved in DMF (1.5 mL) along with O-
benzylhydroxylamine (78.8 mg, 0.6 mmol), DIEA (0.07 mL, 0.4
mmol), and BOP (97.3 mg, 0.2 mmol). After the resulting
solution was stirred overnight, the solid product was filtered
from the solution to give the O-benzylhydroxamate 37 (58 mg,
61%): MS (ESI) m/z 497.3 (M + Na)+.
(2S,11S,12R)-N11-Hyd r oxy-12-isobu tyl-N2-m eth yl-8,13-
d ioxo-1,7-d ia za cyclotr id eca n e-2,11-d ica r boxa m id e (38).
The O-benzylhydroxamate 37 (50 mg, 0.1 mmol) was hydro-
genated in a MeOH/CHCl3 mixture (3:1, 40 mL) with 10% Pd/C
(20 mg) under a hydrogen atmosphere (balloon). After the
solution was stirred for 6 h, the catalyst was filtered off and
the solution was concentrated to yield the title hydroxamate
38 (38 mg, 93%) as a white foam: 1H NMR (DMSO-d6) δ 10.51
(s, 1H), 8.71 (s, 1H), 7.95 (d, 1H), 7.70 (m, 1H), 7.51 (t, 1H),
4.33 (m, 1H), 3.21 (m, 1H), 2.78 (m, 1H), 2.57 (d, 3H), 2.26 (m,
2H), 1.80-1.08 (m, 12H), 0.88 (m, 1H), 0.82 (d, 3H), 0.75 (d,
3H); MS (ESI) m/z 407.3 (M + Na)+; HRMS m/z 385.246983
[(M + H)+ calcd for C18H33N4O5 385.245096].
ter t-Bu tyl (2S,3R)-2-(3-Hyd r oxyp r op yl)-5-m eth yl-3-[({-
(1S)-1-[(m eth ylam in o)car bon yl]-5-[(ph en ylsu lfon yl)am in o]-
p en tyl}a m in o)ca r bon yl]h exa n oa te (39a ). To a solution of
the acid 32 (460.0 mg, 1.6 mmol), Nꢀ-(phenylsulfonyl)-L-lysine
N-methylamide (696.5 mg, 2.1 mmol), and DIEA (0.84 mL, 4.8
mmol) in DMF (5 mL) was added BOP (849.6 mg, 1.9 mmol).
After the resulting solution was stirred overnight, EtOAc was
added and the solution was washed with 10% citric acid,
saturated NaHCO3 solution, and brine. The ethyl acetate layer
was dried (MgSO4) and concentrated. The resulting residue
was purified by silica gel chromatography to yield the amide
39a (833 mg, 90%) as a white foam: MS (ESI) m/z 570.3 (M +
H)+.
(2R,3S)-3-(ter t-Bu toxyca r bon yl)-6-h yd r oxy-2-isobu tyl-
h exa n oic Acid (32). Compound 31a was subjected to a
hydrogenation following the procedure described for 5: MS
(ESI) m/z 289.2 (M + H)+.
ter t-Bu tyl (2S,3R)-3-[({(1S)-5-{[(Ben zyloxy)ca r bon yl]-
a m in o }-1-[(m e t h y la m in o )c a r b o n y l]p e n t y l}a m in o )-
ca r bon yl]-2-(3-h yd r oxyp r op yl)-5-m eth ylh exa n oa te (33).
To a solution of the acid 32 (170 mg, 0.6 mmol) and N-
[(benzyloxy)carbonyl]-L-lysine N-methylamide (224.6 mg, 0.8
mol) in DMF (6 mL) were added DIEA (0.26 mL, 1.5 mmol)
and BOP (286.9 mg, 0.6 mmol). After the resulting solution
was stirred overnight, EtOAc was added and the solution was
washed with 10% citric acid, saturated NaHCO3 solution, and
brine. The EtOAc layer was dried (MgSO4) and concentrated.
The resulting residue was purified by silica gel chromatogra-
phy to yield the amide 33 (255 mg, 77%) as a white foam: MS
(ESI) m/z 564.4 (M + H)+.
(4S,5R)-5-[({(1S)-5-{[(Ben zyloxy)ca r b on yl]a m in o}-1-
[(m eth ylam in o)car bon yl]pen tyl}am in o)car bon yl]-4-(ter t-
bu toxyca r bon yl)-7-m eth ylocta n oic Acid (34). To a solution
of 33 (813 mg, 1.4 mmol) in a mixed solvent of CH3CN (3 mL),
CCl4 (3 mL), and H2O (4.5 mL) at room temperature were
added H5IO6 (1.3 g, 5.9 mmol) and RuCl3‚H2O (6 mg, 0.03
mmol). After the resulting solution was stirred for 1.5 h, 10%
citric acid was added and the layers were separated. The
organic layer was dried and concentrated. The resulting
residue was purified by silica gel chromatography to yield the
acid 34 (504 mg, 60%) as a white foam: MS (ESI) m/z 578.5
(M + H)+.
ter t-Bu tyl (6S,9R,10S)-9-Isobu tyl-6-[(m eth yla m in o)ca r -
bon yl]-8-oxo-1-(p h en ylsu lfon yl)-1,7-d ia za cyclotr id eca n e-
10-ca r boxyla te (40a ). The amide 39a (875.0 mg, 1.5 mmol)
was dissolved in THF (27 mL) prior to the addition of PPh3
(1.21 g, 4.5 mmol) followed by DIAD (0.88 mL, 4.5 mmol)
dropwise. After being stirred overnight, the solution was
concentrated, and the residue was purified by silica gel
chromatography to yield the sultam 40a (470 mg, 55%) as a
white solid: MS (ESI) m/z 552.4 (M + H)+.
(2S,11S,12R)-N11-H yd r oxy-12-isob u t yl-N2-m et h yl-13-
oxo-7-(p h en ylsu lfon yl )-1,7-d ia za cyclotr id eca n e-2,11-d i-
ca r boxa m id e (41). The sultam 40a (473.0 mg, 0.86 mmol)
was dissolved in CH2Cl2 (6 mL) and TFA (5 mL). After being
stirred overnight, the solution was concentrated to afford the
crude acid. A portion of this acid (260.0 mg, 0.52 mmol) was
dissolved in DMF prior to the addition of O-benzylhydroxy-
lamine (192.0 mg, 1.6 mmol), DIEA (0.18 mL, 1.0 mmol), and
BOP (278.0 mg, 0.63 mmol). After the resulting solution was
stirred overnight, the solid product was filtered from the
solution to give the crude O-benzylhydroxamate (172 mg, 57%).
A portion of this O-benzylhydroxamate (150.0 mg, 0.25 mmol)
was hydrogenated in a MeOH/CHCl3 mixture (3:1, 50 mL) with
5% Pd/BaSO4 (300 mg) under a hydrogen atmosphere (50 psi).
After the resulting solution was stirred for 3 h, the catalyst
was filtered off and the solution was concentrated to yield the
(4S,5R)-5-[({(1S)-5-Am in o-1-[(m eth yla m in o)ca r bon yl]-
p en t yl}a m in o)ca r b on yl]-4-(ter t-b u t oxyca r b on yl)-7-m e-
th ylocta n oic Acid (35). The N-Cbz amine 34 (45 mg, 0.08
mmol) was hydrogenated in MeOH (5 mL) with 5% Pd/C-
Degussa (15 mg) under a hydrogen atmosphere (50 psi). After
the solution was stirred overnight, the catalyst was filtered

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2651
hydroxamate 41 (52 mg, 41%) as a white solid: 1H NMR
(DMSO-d6) δ 10.44 (s, 1H), 8.78 (br, 1H), 8.26 (d, 1H), 7.72 (d,
2H), 7.67-7.48 (m, 4H), 4.22 (m, 1H), 3.15-2.95 (m, 2H), 2.78-
2.56 (m, 4H), 2.54 (d, 3H), 1.95 (m, 1H), 1.8-1.2 (m, 10H), 0.86
(m, 1H), 0.83 (d, 3H), 0.73 (d, 3H); MS (ESI) m/z 511.3 (M +
H)+; HRMS m/z 511.261183 [(M + H)+ calcd for C24H39N4O6
511.259032].
chloride (1.76 mL, 22.7 mmol). After being stirred for 1 h, the
solution was poured into saturated aqueous NaHCO3. The
aqueous layer was extracted 2× with CH2Cl2. The organic
phase was washed with H2O, 10% aqueous citric acid, H2O,
and brine and dried over MgSO4, and the solvent was removed
under reduced pressure, affording a quantitative yield of the
mesylate as a white solid: MS (ESI) m/z 338 (M + H)+.
To the above mesylate in acetone (100 mL) was added NaI
(3.9 g). After the solution was stirred overnight at ambient
temperature, an additional 3.9 g of NaI was added and the
reaction was refluxed for 1 h. The mixture was filtered, and
the volatiles were removed under reduced pressure. The solid,
which immediately started turning yellow, was dissolved in
hexane, washed with H2O, 5% aqueous sodium thiosulfate,
H2O, and brine, and dried over MgSO4, and the volatiles were
removed under reduced pressure, affording 6.79 g (96%) of
46: MS (ESI) m/z 370 (M + H)+.
(2S)-2-Am in o-5-[4-(ben zyloxy)p h en yl]-N-[(1S,2S)-2-h y-
d r oxy-1-m et h yl-2-p h en ylet h yl]-N-m et h ylp en t a n a m id e
(47). To a stirred slurry of 1.15 g of LiCl (previously flame-
dried in flask under vacuum) and 2-amino-N-[(1S,2S)-2-
hydroxy-1-methyl-2-phenylethyl]-N-methylacetamide39 (0.99 g,
4.48 mmol) in THF (30 mL) cooled to -78 °C was added a
solution of 1 M LDA in THF/hexane (8.7 mL) over 10 min.
The mixture was stirred for 20 min at -78 °C and 30 min at
0 °C, at which time a solution of 46 (1.57 g, 4.25 mmol) in
THF (10 mL) was added dropwise over 10 min. The mixture
was allowed to slowly warm to room temperature while being
stirred overnight. The reaction was quenched with 10%
aqueous citric acid, and the volatiles were removed under
reduced pressure. The remaining material was dissolved in
EtOAc. The resulting solution was washed with H2O, 5%
aqueous sodium thiosulfate, H2O, saturated aqueous NaHCO3,
H2O, and brine and dried over MgSO4, and the volatiles were
removed under reduced pressure. The residue was chromato-
graphed on silica gel eluting with MeOH/CHCl3 (4:100),
affording 0.9 g (47%) of 47: MS (ESI) m/z 447 (M + H)+.
Meth yl (2S)-2-Am in o-5-[4-(ben zyloxy)p h en yl]p en ta n o-
a te (48). To a solution of 47 (3.5 g, 7.8 mmol) in H2O (40 mL)
and MeOH (25 mL) was added 1 N aqueous NaOH (15.7 mL).
The reaction was refluxed for 1 h, at which time more MeOH
(25 mL) was added. The reaction was refluxed for an additional
3 h, and the volatiles were removed under reduced pressure.
The solid was triturated with CH2Cl2 and filtered, affording a
mixture of NaOH and the sodium salt of the product (5.5 g).
The CH2Cl2 filtrate was evaporated, and the solid was tritu-
rated with Et2O, affording an additional 1.1 g of the sodium
salt of the product: MS (ESI) m/z 298 (M + H)+.
To the above mixture of sodium salt and NaOH in MeOH
(150 mL) was added concentrated HCl (3 mL). The solution
was refluxed overnight, and the volatiles were removed under
reduced pressure. The resulting material was taken into
EtOAc, the solution was washed with saturated aqueous
NaHCO3 and brine and dried over MgSO4, and the volatiles
were removed under reduced pressure, affording 2.4 g (99%)
of 48: MS (ESI) m/z 314 (M + H)+.
ter t-Bu tyl (6S,9R,10S)-9-Isobu tyl-6-[(m eth yla m in o)ca r -
bon yl]-8-oxo-1-(m esitylsu lfon yl)-1,7-diazacyclotr idecan e-
10-ca r boxyla te (40b). To a solution of the acid 32 (990 mg,
3.4 mmol) and Ne-(mesitylsulfonyl)-L-lycine N-methylamide
hydrochloride (1.7 g, 4.5 mmol) in DMF were added DIEA (1.8
mL, 10.2 mmol) and BOP (1.8 g, 4.1 mmol). After the resulting
solution was stirred overnight, DMF was removed under
reduced pressure and CH2Cl2 was added. The solution was
washed with 10% citric acid, saturated NaHCO3 solution, and
brine. The CH2Cl2 layer was dried (MgSO4) and concentrated.
The resulting residue was purified by silica gel chromatogra-
phy to yield the crude amide (2 g), which was dissolved in THF
(158 mL). To the THF solution was added PPh3 (2.8 g, 10.6
mmol) followed by DIAD (2 mL, 10.1 mmol) in THF. After
being stirred overnight, the solution was concentrated, and
the residue was purified by silica gel chromatography to yield
the sultam 40b (680 mg, 30%) as a yellow solid: MS (ESI)
m/z 594.5 (M + H)+.
(6S,9R,10S)-1-(ter t-Bu toxyca r bon yl)-9-isobu tyl-6-[(m e-
th yla m in o)ca r bon yl]- 8-oxo-1,7-d ia za cyclotr id eca n e-10-
ca r boxylic Acid (42). The sultam 40b (300 mg, 0.5 mmol)
was dissolved in 33% HBr/AcOH (6.8 mL) containing phenol
(63 mg, 0.67 mmol). After being stirred for 5 h, the solution
was concentrated, and the solid was filtered off with CH2Cl2/
Et2O. This provided the crude amino acid salt (500 mg,
quantitative). A portion of this amino acid (140 mg, 0.32 mmol)
was dissolved in THF (4 mL)/H2O (0.6 mL). To it was added
Et3N (0.38 mL, 2.6 mmol) followed by (Boc)2O (452 mg, 206
mmol) at room temperature. After the resulting solution was
stirred overnight, the solvent was removed and CH2Cl2 was
added. The CH2Cl2 solution was washed with 10% HCl, dried
(MgSO4), and concentrated. The resulting residue was purified
by silica gel chromatography to yield the N-Boc acid 42 (130
mg) as a solid: MS (ESI) m/z 456.5 (M + H)+.
ter t-Bu tyl (6S,9R,10S)-10-[(Hyd r oxya m in o)ca r bon yl]-
9-isobu tyl-6-[(m eth yla m in o)ca r bon yl]-8-oxo-1,7-d ia za cy-
clotr id eca n e-1-ca r boxyla te (43). The N-Boc acid 42 (130
mg) was dissolved in DMF (5 mL) prior to the addition of
O-benzylhydroxylamine (108 mg, 0.87 mmol), DIEA (0.15 mL,
0.82 mmol), and BOP (214 mg, 0.48 mmol). After the solution
was stirred overnight, the solid product was filtered off with
CH2Cl2 to give O-benzylhydroxamate (120 mg, 67%). The
O-benzylhydroxamate (160 mg, 0.29 mmol) was hydrogenated
in MeOH (40 mL) with 5% Pd/BaSO4 (240 mg) under a
hydrogen atmosphere (50 psi). After the resulting solution was
stirred for 3 h, the catalyst was filtered off and the solution
was concentrated to yield the hydroxamate 43 (140 mg
quantitative) as a pale yellow solid: 1H NMR (CD3OD) δ 4.40
(m, 1H), 3.2-2.9 (m, 4H), 2.70 (s, 3H), 2.45 (m, 1H), 2.02 (m,
1H), 1.44 (s, 9H), 1.8-1.2 (m, 12H), 0.99 (m, 1H), 0.91 (d, 3H),
0.83 (d, 3H): MS (ESI) m/z 471.5 (M + H)+; HRMS m/z
471.319240 [(M + H)+ calcd for C23H43N4O6 471.318261].
(2R,3S)-6-Br om o-3-(ter t-b u t oxyca r b on yl)-2-isob u t yl-
h exa n oic Acid (49a ). To a stirred solution of PPh3 (8.32 g,
31.7 mmol), imidazole (2.15 g 31.6 mmol), and CBr4 (10.54 g,
31.8 mmol) in CH2Cl2 (60 mL) at 0 °C was added dropwise
over 15 min a solution of 31a (8.0 g, 21.2 mmol) in CH2Cl2 (60
mL). The reaction was stirred at 0 °C for 30 min followed by
a second addition of PPh3 (4.15 g, 15.8 mmol), imidazole (1.55
g, 22.8 mmol), and CBr4 (5.25 g, 15.8 mmol) in CH2Cl2 (30 mL)
in one portion. The reaction was stirred for an additional 2.5
h at 0 °C and then for 20 min at ambient temperature, followed
by dilution with hexanes (320 mL), filtration through a short
silica gel plug, and rinsing with 25% EtOAc/hexanes. The
volatiles were removed under reduced pressure, and the
resulting material was chromatographed on silica gel eluting
with a 1-10% EtOAc/hexanes gradient, affording 6.1 g (65%)
of the bromide: MS (ESI) m/z 441 (M + H)+.
(2S,11S,12R)-N11-H yd r oxy-12-isob u t yl-N2-m et h yl-13-
oxo-1,7-diazacyclotr idecan e-2,11-dicar boxam ide (44). The
N-Boc hydroxamate 43 (56 mg, 0.12 mmol) was dissolved in 4
N HCl in dioxane (2 mL) at room temperature. After the
solution was stirred for 3 h, the solvent was removed under
reduced pressure to yield the desired hydroxamate 44 (45 mg,
quantitative): 1H NMR (CD3OD) δ 4.19 (m, 1H), 2.99 (m, 4H),
2.72 (s, 3H), 2.15 (m, 1H), 1.88 (m, 1H), 1.78-1.24 (m, 12H),
1.03 (m, 1H), 0.93 (d, 3H), 0.84 (d, 3H); MS (ESI) m/z 371.4
(M + H)+; HRMS m/z 471.267256 [(M + H)+ calcd for
C
18H35N4O4 471.265831].
1-(Ben zyloxy)-4-(3-iodopr opyl)ben zen e (46). To a stirred
solution of 3-[4-(benzyloxy)phenyl]-1-propanol (45) (5.0 g, 20.6
mmol) in THF (100 mL) cooled in an ice bath was added
triethylamine (4.3 mL, 31 mmol) followed by methanesulfonyl
To the above benzyl ester (10.5 g, 23.9 mmol) in MeOH (250
mL) was added 10% Pd-C (1 g). The mixture was stirred

2652 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
under H2 (balloon) for 3 h. The catalyst was removed by
filtration through a 0.45 µM PTFE disposable filter, and the
volatiles were evaporated under reduced pressure, affording
8.3 g (99%) of the acid 49a : MS (ESI) m/z 349 (M - H)-.
2H), 6.73 (m, 1H), 6.29 (d, J ) 9.9 Hz, 1H), 4.92 (m, 1H), 4.70
(s, 2H), 3.92 (m, 2H), 3.40 (m, 1H), 2.45 (m, 1H), 2.05 (m, 2H),
1.72 (m, 1H), 1.02-1.28 (m, 5H), 0.72 (m, 1H), 0.62 (d, J ) 6.6
Hz, 3H), 0.56 (d, J ) 6.6 Hz, 3H), -0.60 (m, 1H); MS (ESI)
m/z 483 (M + H)+.
ter t-Bu tyl (2R,3S)-2-(3-Br om op r op yl)-3-({[(1R)-1-(4-h y-
d r oxyben zyl)-2-m eth oxy-2-oxoeth yl]a m in o}ca r bon yl)-5-
m eth ylh exa n oa te (50a ). To a solution of 49a (8.4 g, 24
mmol), L-tyrosine methyl ester hydrochloride (5.5 g, 24 mmol),
and NMM (9.1 mL, 65 mmol) in DMF (200 mL) was added a
solution of TBTU (9.52 g, 29.6 mmol) in DMF (120 mL) over
30 min. The reaction was stirred for 2 h at ambient temper-
ature followed by removal of the volatiles under reduced
pressure. The resulting material was dissolved in EtOAc. The
solution was washed with cold 1 N HCl. The aqueous phase
was extracted 3× with EtOAc. The combined organic phase
was washed with H2O, saturated NaHCO3, H2O, and brine and
dried over MgSO4, and the volatiles were removed under
reduced pressure. The residue was chromatographed on silica
gel eluting with a gradient of 25-33% EtOAc/hexanes, afford-
ing 9.5 g (75%) of the amide product 50a : MS (ESI) m/z 529
(M + H)+.
6-ter t-Bu tyl 10-Meth yl (6S,7R,10S)-7-Isobu tyl-8-oxo-2-
oxa -9-a za bicyclo[10.2.2]h exa d eca -1(14),12,15-tr ien e-6,10-
d ica r boxyla te (51a ). To a stirred suspension of Cs2CO3 (5.2
g, 16 mmol) in DMF (130 mL) and DMSO (32.5 mL) at 60 °C
was added a solution of 50a (2.35 g, 4.45 mmol) in DMF (25
mL) over 15 min. The reaction was heated at 80 °C for 30 min,
cooled in an ice bath, and quenched with 10% citric acid. The
volatiles were removed under reduced pressure, and the
resulting material was partitioned in EtOAc/H2O. The aqueous
layer was extracted 4× with EtOAc, the combined organic
extracts were washed 4× with H2O and once with brine and
dried over MgSO4, and the volatiles were removed under
reduced pressure. The residue was chromatographed on silica
gel eluting with 1.5% MeOH/CH2Cl2, affording 2.0 g (74%) of
the macrocycle 51a : MS (ESI) m/z 448 (M + H)+.
To a solution of the acid (30 mg) in DMF (2 mL) were added
NMM (0.030 mL), glycine-N-methylamide hydrochloride (15
mg), and TBTU (26 mg). After being stirred for 18 h at ambient
temperature, the reaction was heated to 80 °C for 15 min. The
volatiles were removed under reduced pressure, and the
resulting material was purified by preparative TLC (1 mm
with 0.25 mm concentration zone) eluting with 5% MeOH/
CHCl3, affording 30 mg of the intermediate. This was dissolved
in MeOH (10 mL), and to this was added 5% Pd/BaSO4 (35
mg). The mixture was shaken under H2 (50 psi) for 6 h and
filtered, and the volatiles were removed under reduced pres-
sure, affording 20 mg (70%) of 54e: 1H NMR (CD3OD) δ 7.21
(m, 1H), 7.10 (m, 2H), 6.86 (m, 1H), 4.92 (m, 1H), 4.12 (m,
2H), 3.82 (m, 2H), 3.35 (m, 1H), 2.73 (s, 3H), 2.68 (m, 1H),
2.14 (m, 1H), 1.68 (m, 1H), 1.30 (m, 5H), 0.85 (m, 1H), 0.81 (d,
J ) 6.6 Hz, 3H), 0.75 (d, J ) 6.6 Hz, 3H), -0.55 (m, 1H); MS
(ESI) m/z 463.2 (M + H)+. Anal. (C23H34N4O6) C, H, N.
The following compounds were prepared in
a manner
analogous to that for 54e.
(6S,7R,10S)-N6-Hyd r oxy-7-isobu tyl-N10-m eth yl-8-oxo-2-
oxa -9-a za bicyclo[10.2.2]h exa d eca -1(14),12,15-tr ien e-6,10-
d ica r boxa m id e (54a ): mp 350-352 °C; IR ν 574, 642, 672,
1002, 1054, 1172, 1184, 1194, 1222, 1258, 1298, 1366, 1406,
1
1442, 1516, 1636, 2928, 3314 cm-1; H NMR (CD3OD) δ 8.20
(d, 1H), 7.85 (m, 1H), 7.18 (m, 1H), 7.11 (m, 1H), 7.04 (m, 1H),
6.85 (m, 1H), 4.90 (m, 1H), 4.10 (m, 2H), 3.23 (m, 1H), 2.74
(m, 1H), 2.70 (s, 3H), 2.14 (m, 1H), 1.66 (m, 1H), 1.26-1.37
(m, 5H), 0.88 (m, 1H), 0.78 (m, 6H), -0.55 (m, 1H); MS (ESI)
m/z 404.2 (M - H)-; HRMS m/z 406.234761 [(M + H)+ calcd
for C21H32N3O5 406.234197]. Anal. (C21H31N3O5) C, H, N.
(6S ,7R ,10S )-N 6-H yd r oxy-7-isob u t yl-8-oxo-2-oxa -N 10
-
(2,2,2-tr iflu or oeth yl)-9-a za bicyclo[10.2.2]h exa d eca -1(14),-
12,15-tr ien e-6,10-d ica r boxa m id e (54b): 1H NMR (CD3OD)
δ 8.30 (d, 1H), 7.20 (d, 1H), 7.10 (d, 1H), 7.02 (d, 1H), 6.85 (d,
1H), 4.80 (m, 1H), 4.10 (m, 2H), 3.95 (m, 2H), 2.86 (t, J ) 13.2
Hz, 2H), 2.10 (m, 2H), 1.65 (m, 2H), 1.4-1.2 (m, 4H), 0.90 (m,
1H), 0.79 (d, J ) 6.6 Hz, 3H), 0.74 (d, J ) 6.6 Hz, 3H), -0.55
(m, 1H); MS (ESI) m/z 474.2. Anal. (C22H30F3N3O5‚0.5H2O) C,
H, N.
(6S,7R,10S)-N6-Hydr oxy-7-isobu tyl-N10-isopr opyl-8-oxo-
2-oxa -9-a za bicyclo[10.2.2]h exa d eca -1(14),12,15-tr ien e-6,-
10-d ica r boxa m id e (54c): 1H NMR (CD3OD) δ 7.88 (s, 1H),
7.25-7.00 (m, 3H), 6.85 (d, 1H), 4.60 (m, 1H), 4.10 (m, 2H),
3.30 (m, 1H), 2.80 (m, 2H), 2.10 (m, 2H), 1.60 (m, 2H), 1.4-
0.9 (m, 10H), 0.82 (d, J ) 6.2 Hz, 3H), 0.74 (d, J ) 5.9 Hz,
3H),-0.60(m,1H);MS (ESI)m/z434.2.Anal.(C23H35N3O5‚0.2CF3-
CO2H) C, H, N.
(6S,7R,10S)-N10-{2-[4-(Am in osu lfon yl)p h en yl]et h yl}-
N6-h yd r oxy-7-isobu tyl -8-oxo-2-oxa -9-a za bicyclo[10.2.2]-
h exadeca-1(14),12,15-tr ien e-6,10-dicar boxam ide (54d): 1H
NMR (CD3OD) δ 8.21 (d, J ) 9.5 Hz, 1H), 8.10 (m, 1H), 7.82
(d, J ) 8.3 Hz, 2H), 7.39 (d, J ) 8.3 Hz, 2H), 7.10 (m, 3H),
6.84 (m, 1H), 4.80 (m, 1H), 4.10 (m, 2H), 3.45 (m, 2H), 3.18
(m, 1H), 2.88 (m, 2H), 2.60 (m, 1H), 2.12 (m, 1H), 1.65 (m,
1H), 1.30 (m, 5H), 0.86 (m, 1H), 0.81 (d, J ) 6.4 Hz, 3H), 0.75
(d, J ) 6.6 Hz, 3H), -0.55 (m, 1H); MS (ESI) m/z 573.2 (M -
H)-. Anal. (C28H38N4O7S‚0.5H2O) C, H, N.
Meth yl (6S,7R,10S)-6-{[(Ben zyloxy)a m in o]ca r bon yl}-
7-isobu tyl-8-oxo-2-oxa-9-azabicyclo[10.2.2]h exadeca-1(14),-
12,15-tr ien e-10-ca r boxyla te (52a ). Compound 51a (0.77 g,
1.7 mmol) was dissolved in TFA (25 mL). The solution was
stirred for 1 h at ambient temperature. The volatiles were
removed under reduced pressure, affording 0.67 g (99%) of the
acid: MS (ESI) m/z 392 (M + H)+.
To a solution of the above acid (2.4 g, 6.1 mmol) in DMF
(75 mL) were added NMM (3.37 mL, 24 mmol), HATU (5.24
g, 13.7 mmol), and O-benzylhydroxylamine hydrochloride (3.77
g, 23.6 mmol). After being stirred overnight at ambient
temperature, the reaction was heated to 60 °C for 30 min. After
the reaction was cooled to room temperature, the volatiles were
removed under reduced pressure and the resulting material
was dissolved in EtOAc. The solution was washed with 10%
citric acid. The aqueous layer was extracted 3× with EtOAc.
The combined organic phase was washed 3× with H2O and
brine and dried over MgSO4, and the volatiles were removed
under reduced pressure. The residue was triturated 4× with
a mixture of 1:1:2 EtOAc/hexane/ether to afford 1.4 g of
product. The mother liquor was concentrated and chromato-
graphed on silica gel eluting with a gradient of 25-90% EtOAc/
hexanes, affording an additional 1.05 g of 52a (combined yield
81%).
(6S,7R,10S)-N6-Hyd r oxy-7-isobu tyl-N10-[2-(m eth yla m i-
n o)-2-oxoeth yl]-8-oxo-2-oxa-9-azabicyclo[10.2.2]h exadeca-
1(14),12,15-tr ien e-6,10-d ica r boxa m id e (54e). To a solution
of 52a (0.7 g, 1.4 mmol) in THF (65 mL) and H2O (15 mL) was
added saturated aqueous LiOH (2.23 mL). The reaction was
stirred for 2 h at ambient temperature and quenched with the
addition of 1 N HCl (10 mL). The majority of the volatiles were
removed under reduced pressure. The remaining material was
diluted with EtOAc and washed with H2O and 1 N HCl. The
combined aqueous phase was extracted 4× with EtOAc. The
combined organic phase was washed with H2O and brine and
dried over MgSO4, and the volatiles were removed under
reduced pressure, affording 0.67 g (99%) of the acid: 1H NMR
(CDCl3) δ 9.53 (s, 1H), 7.20 (m, 5H), 7.00 (m, 1H), 6.85 (m,
(6S,7R,10S)-N6-Hyd r oxy-7-isobu tyl-8-oxo-N10-{2-oxo-2-
[(2,2,2-t r iflu or oet h yl)a m in o]et h yl}-2-oxa -9-a za b icyclo-
[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d ica r b oxa m id e
(54f): 1H NMR (CD3OD) δ 8.28 (d, J ) 9.5 Hz, 1H), 7.22 (m,
1H), 7.08 (m, 2H), 6.86 (m, 1H), 4.86 (m, 1H), 4.10 (m, 2H),
3.80 (m, 4H), 3.38 (m, 1H), 2.66 (m, 1H), 2.14 (m, 1H), 1.68
(m, 1H), 1.22-1.39 (m, 5H), 0.86 (m, 1H), 0.80 (d, J ) 6.6 Hz,
3H), 0.74 (d, J ) 6.6 Hz, 3H), -0.56 (m, 1H); MS (ESI) m/z
529.1 (M - H)-. Anal. (C24H33F3N4O6‚0.2CF3CO2H) C, H, N.
(6S,7R,10S)-N10-[2-(Cyclob u t yla m in o)-2-oxoet h yl]-N6-
h yd r oxy-7-isob u t yl-8-oxo-2-oxa -9-a za b icyclo[10.2.2]-
h exadeca-1(14),12,15-tr ien e-6,10-dicar boxam ide (54g): 1H

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2653
NMR (CD3OD) δ 8.30 (d, J ) 9.9 Hz, 1H), 7.21 (m, 1H), 7.10
(m, 2H), 6.80 (m, 1H), 4.90 (m, 1H), 4.28 (m, 1H), 4.12 (m,
2H), 3.80 (m, 2H), 3.36 (m, 1H), 2.66 (m, 1H), 2.12 (m, 3H),
1.92 (m, 2H), 1.70 (m, 3H), 1.20-1.39 (m, 5H), 0.86 (m, 1H),
0.80 (d, J ) 6.2 Hz, 3H), 0.74 (d, J ) 6.6 Hz, 3H), -0.55 (m,
1H); MS (ESI) m/z 503 (M + H)+. Anal. (C26H38N4O6‚0.7H2O)
C, H, N.
(6S,7R,10S)-N10-[2-(ter t-Bu t yla m in o)-2-oxoet h yl]-N6-
h yd r oxy-7-isob u t yl-8-oxo-2-oxa -9-a za b icyclo[10.2.2]-
h exadeca-1(14),12,15-tr ien e-6,10-dicar boxam ide (54h ): 1H
NMR (CD3OD) δ 8.27 (d, J ) 9.9 Hz, 1H), 7.47 (s, 1H), 7.22
(m, 1H), 7.08 (m, 2H), 6.86 (m, 1H), 4.92 (m, 1H), 4.10 (m,
2H), 3.80 (m, 2H), 3.38 (m, 1H), 2.65 (m, 1H), 2.15 (m, 1H),
1.68 (m, 1H), 1.25-1.39 (m, 5H), 1.31 (s, 9H), 0.86 (m, 1H),
0.82 (d, J ) 6.6 Hz, 3H), 0.75 (d, J ) 6.6 Hz, 3H), -0.55 (m,
1H). Anal. (C26H40N4O6‚H2O) C, H, N.
[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d ica r b oxa m id e
(54o): 1H NMR (CD3OD) δ 8.27 (d, J ) 9.5 Hz, 1H), 7.22 (m,
1H), 7.08 (m, 2H), 6.86 (m, 1H), 4.96 (m, 1H), 4.05 (m, 1H),
4.10 (m, 4H), 3.70 (m, 1H), 3.42 (m, 1H), 2.65 (m, 1H), 2.55
(m, 1H), 2.12 (m, 2H), 1.82 (m, 1H), 1.46-1.70 (m, 4H), 1.23-
1.39 (m, 5H), 0.91 (m, 7H), 0.82 (d, J ) 6.6 Hz, 3H), 0.75 (d, J
) 6.6 Hz, 3H), -0.56 (m, 1H); MS (ESI) m/z 543.3 (M - H)-.
Anal. (C29H44N4O6‚0.25CF3CO2H) C, H, N.
(6S,7R,10S)-N10-[2-(3,3-Dim eth yl-1-p ip er id in yl)-2-oxo-
et h yl]-N6-h yd r oxy-7-isob u t yl-8-oxo-2-oxa -9-a za b icyclo-
[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d ica r b oxa m id e
(54p ): 1H NMR (CD3OD) δ 8.27 (d, J ) 9.9 Hz, 1H), 7.23 (m,
1H), 7.08 (m, 2H), 6.86 (m, 1H), 4.96 (m, 1H), 4.10 (m, 4H),
3.40 (m, 3H), 3.20 (m, 2H), 2.65 (m, 1H), 2.15 (m, 1H), 1.60
(m, 3H), 1.23-1.46 (m, 7H), 0.90 (m, 7H), 0.82 (d, J ) 6.2 Hz,
3H), 0.74 (d, J ) 6.7 Hz, 3H), -0.57 (m, 1H); MS (ESI) m/z
543.2 (M - H)-. Anal. (C29H44N4O6‚1.2H2O) C, H, N.
(6S,7R,10S)-N6-Hyd r oxy-7-isobu tyl-N10-[(1S)-1-m eth yl-
2-(m et h yla m in o)-2-oxoet h yl]-8-oxo-2-oxa -9-a za b icyclo-
[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d ica r b oxa m id e
(54q): 1H NMR (CD3OD) δ 8.28 (d, J ) 9.9 Hz, 1H), 7.25 (m,
1H), 7.08 (m, 2H), 6.85 (m, 1H), 4.95 (m, 1H), 4.30 (q, J ) 7.0
Hz, 1H), 4.10 (m, 2H), 3.34 (m, 1H), 2.68 (m, 4H), 2.12 (m,
1H), 1.65 (m, 1H), 1.13-1.38 (m, 8H), 0.87 (m, 1H), 0.80 (d, J
) 6.6 Hz, 3H), 0.75 (d, J ) 6.6 Hz, 3H), -0.56 (m, 1H); MS
(ESI) m/z 475 (M - H)-; HRMS m/z 477.2693 [(M + H)+ calcd
for C24H37N4O6 477.2713].
(6S,7R,10S)-N6-Hyd r oxy-7-isobu tyl-N10-[(1R)-1-m eth yl-
2-(m et h yla m in o)-2-oxoet h yl]-8-oxo-2-oxa -9-a za b icyclo-
[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d ica r b oxa m id e
(54r ): 1H NMR (CD3OD) δ 8.31 (d, J ) 9.5 Hz, 1H), 7.21 (m,
1H), 7.09 (m, 2H), 6.86 (m, 1H), 4.95 (m, 1H), 4.31 (q, J ) 7.3
Hz, 1H), 4.10 (m, 2H), 3.34 (m, 1H), 2.70 (s, 3H), 2.65 (m, 1H),
2.14 (m, 1H), 1.66 (m, 1H), 1.12-1.42 (m, 8H), 0.86 (m, 1H),
0.82 (d, J ) 6.6 Hz, 3H), 0.76 (d, J ) 6.2 Hz, 3H), -0.56 (m,
1H); MS (ESI) m/z 475.2 (M - H)-. Anal. (C24H36N4O6‚H2O)
C, H, N.
(6S,7R,10S)-N10-(2-An ilin o-2-oxoet h yl)-N6-h yd r oxy-7-
isobu tyl-8-oxo-2-oxa -9-a za bicyclo[10.2.2]h exa d eca -1(14),-
12,15-tr ien e-6,10-d ica r boxa m id e (54i): 1H NMR (CD3OD)
δ 8.33 (d, J ) 9.9 Hz, 1H), 7.54 (d, J ) 7.7 Hz, 2H), 7.26 (m,
3H), 7.08 (m, 3H), 6.86 (m, 1H), 4.98 (m, 1H), 4.08 (m, 4H),
3.40 (m, 1H), 2.71 (m, 1H), 2.18 (m, 1H), 1.68 (m, 1H), 1.23-
1.40 (m, 5H), 0.86 (m, 1H), 0.80 (d, J ) 6.6 Hz, 3H), 0.73 (d, J
) 6.2 Hz, 3H), -0.55 (m, 1H); MS (ESI) m/z 523.2 (M - H)-.
Anal. (C28H36N4O6‚0.95H2O) C, H, N.
(6S,7R,10S)-N10-{2-[(2,4-Diflu or oben zyl)a m in o]-2-oxo-
et h yl}-N6-h yd r oxy-7-isob u t yl-8-oxo-2-oxa -9-a za b icyclo-
[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d ica r boxa m id e
(54j): 1H NMR (CD3OD) δ 7.36 (m, 1H), 7.21 (m, 1H), 7.08
(m, 2H), 6.89 (m, 3H), 4.92 (m, 1H), 4.39 (s, 2H), 4.10 (m, 2H),
3.90 (m, 2H), 3.37 (m, 1H), 2.66 (m, 1H), 2.14, (m, 1H), 1.66
(m, 1H), 1.18-1.38 (m, 5H), 0.86 (m, 1H), 0.79 (d, J ) 6.6 Hz,
3H), 0.71 (d, J ) 6.2 Hz, 3H), -0.55 (m, 1H); MS (ESI) m/z
573.1 (M - H)-. Anal. (C29H36F2N4O6‚0.5H2O) C, H, N.
(6S,7R,10S)-N10-[2-(9H-F lu or en -9-yla m in o)-2-oxoeth yl]-
N6-h yd r oxy-7-isob u t yl-8-oxo-2-oxa -9-a za bicyclo[10.2.2]-
h exadeca-1(14),12,15-tr ien e-6,10-dicar boxam ide (54k): 1H
NMR (CD3OD) δ 8.30 (d, J ) 9.5 Hz, 1H), 7.74 (d, J ) 7.3 Hz,
2H), 7.52 (d, J ) 7.3 Hz, 2H), 7.38 (t, J ) 7.3 Hz, 2H), 7.3-7.0
(m, 5H), 6.86 (m, 1H), 6.09 (s, 1H), 4.95 (m, 1H), 4.10 (m, 2H),
3.90 (m, 2H), 3.38 (m, 1H), 2.68 (m, 1H), 2.14 (m, 1H), 1.68
(m, 1H), 1.23-1.38 (m, 5H), 0.86 (m, 1H), 0.82 (d, J ) 6.6 Hz,
3H), 0.73 (d, J ) 6.2 Hz, 3H), -0.55 (m, 1H); MS (ESI) m/z
611.2 (M - H)-. Anal. (C35H40N4O6‚1.1H2O) C, H, N.
(6S,7R,10S)-N10-[2-(Dim eth yla m in o)-2-oxoeth yl]-N6-h y-
d r o x y -7-i s o b u t y l-8-o x o -2-o x a -9-a z a b i c y c lo [10.2.2]-
h exa d eca -1(14),12,15-tr ien e-6,10-d ica r boxa m id e (54l): 1H
NMR (CD3OD) δ 8.28 (d, J ) 9.9 Hz), 7.23 (m, 1H), 7.10 (m,
2H), 6.86 (m, 1H), 4.95 (m, 1H), 4.10 (m, 4H), 3.40 (m, 1H),
3.02 (s, 3H), 2.94 (s, 3H), 2.65 (m, 1H), 2.14 (m, 1H), 1.68 (m,
1H), 1.23-1.39 (m, 5H), 0.86 (m, 1H), 0.82 (d, J ) 6.6 Hz, 3H),
0.75 (d, J ) 6.2 Hz, 3H), -0.56 (m, 1H). Anal. (C24H36N4O6‚
H2O) C, H, N.
(6S,7R,10S)-N6-Hyd r oxy-7-isobu tyl-N10-[(1S)-1-(4-m eth -
oxyb en zyl)-2-(m et h yla m in o)-2-oxoet h yl]-8-oxo-2-oxa -9-
a za bicyclo[10.2.2]h exa d eca -1(14),12,15-tr ien e-6,10-d ica r -
boxa m id e (54s): 1H NMR (CD3OD) δ 7.22 (m, 1H), 7.09 (m,
4H), 6.85 (m, 3H), 4.95 (m, 1H), 4.46 (t, J ) 7.3 Hz, 1H), 4.12
(m, 2H), 3.74 (s, 3H), 3.40 (m, 1H), 2.92 (m, 2H), 2.65 (s, 3H),
2.61 (m, 1H), 2.12 (m, 1H), 1.66 (m, 1H), 1.21-1.39 (m, 5H),
0.86 (m, 1H), 0.81 (d, J ) 6.6 Hz, 3H), 0.75 (d, J ) 6.6 Hz,
3H), -0.56 (m, 1H); MS (ESI) m/z 581.2 (M - H)-. Anal.
(C31H42N4O7‚H2O) C, H, N.
(6S,7R,10S)-N10-[(1S)-1-(ter t-Bu toxym eth yl)-2-(m eth y-
la m in o)-2-oxoeth yl]-N6-h yd r oxy-7-isobu tyl-8-oxo-2-oxa -
9-a za b icyclo[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d i-
ca r boxa m id e (54t): 1H NMR (CD3OD) δ 8.29 (d, J ) 9.9 Hz,
1H), 7.81 (d, J ) 7.7 Hz, 1H), 7.22 (m, 1H), 7.08 (m, 2H), 6,86
(m, 1H), 4.95 (m, 1H), 4.38 (m, 1H), 4.10 (m, 2H), 3.62 (m,
1H), 3.48 (m, 1H), 3.34 (m, 1H), 2.63-2.84 (m, 4H), 2.12 (m,
1H), 1.66 (m, 1H), 1.23-1.40 (m, 5H), 1.15 (s, 9H), 0.88 (m,
1H), 0.81 (d, J ) 6.2 Hz, 3H), 0.76 (d, J ) 6.6 Hz, 3H), -0.56
(m, 1H); MS (ESI) m/z 547.2 (M - H)-. Anal. (C28H44N4O7‚
0.8H2O) C, H, N.
(6S,7R,10S)-N6-Hyd r oxy-7-isobu tyl-N10-{(1S)-2-m eth yl-
1-[(m e t h y la m i n o )c a r b o n y l]p r o p y l}-8-o x o -2-o x a -9-
a za bicyclo[10.2.2]h exa d eca -1(14),12,15-tr ien e-6,10-d ica r -
boxa m id e (54u ): 1H NMR (CD3OD) δ 8.27 (d, J ) 9.9 Hz,
1H), 7.89 (d, J ) 8.8 Hz, 1H), 7.22 (m, 1H), 7.08 (m, 2H), 6.85
(m, 1H), 4.94 (m, 1H), 4.10 (m, 3H), 3.32 (m, 1H), 2.73 (s, 3H),
2.65 (m, 1H), 2.10 (m, 1H), 2.01 (m, 1H), 1.66 (m, 1H), 1.23-
1.39 (m, 5H), 0.86-0.92 (m, 7H), 0.80 (d, J ) 6.6 Hz, 3H), 0.74
(d, J ) 6.6 Hz, 3H), -0.57 (m, 1H); MS (ESI) m/z 503.3 (M -
H)-. Anal. (C26H40N4O6‚0.25CF3CO2H) C, H, N.
(6S,7R,10S)-N6-Hyd r oxy-7-isobu tyl-8-oxo-N10-[2-oxo-2-
(1-piper id in yl)eth yl]-2-oxa -9-a za bicyclo[10.2.2]h exa deca -
1(14),12,15-tr ien e-6,10-d ica r boxa m id e (54m ): 1H NMR
(CD3OD) δ 8.28 (d, J ) 9.5 Hz, 1H), 7.23 (m, 1H), 7.10 (m,
2H), 6.86 (m, 1H), 4.95 (m, 1H), 4.10 (m, 4H), 3.53 (m, 2H),
3.40 (m, 3H), 2.65 (m, 1H), 2.14 (m, 1H), 1.50-1.69 (m, 7H),
1.23-1.39 (m, 5H), 0.86 (m, 1H), 0.82 (d, J ) 6.6 Hz, 3H), 0.74
(d, J ) 6.2 Hz, 3H), -0.57 (m, 1H); MS (ESI) m/z 515.2 (M -
H)-. Anal. (C27H40N4O6‚H2O) C, H, N.
(6S,7R,10S)-N6-Hyd r oxy-N10-[2-(4-h yd r oxy-1-p ip er id i-
n yl)-2-oxoeth yl]-7-isobu tyl-8-oxo-2-oxa-9-azabicyclo[10.2.2]-
h exadeca-1(14),12,15-tr ien e-6,10-dicar boxam ide (54n ): 1H
NMR (CD3OD) δ 8.29 (d, J ) 9.5 Hz, 1H), 7.94 (m, 1H), 7.22
(m, 1H), 7.10 (m, 2H), 6.86 (m, 1H), 4.95 (m, 1H), 4.10 (m,
5H), 3.82 (m, 1H), 3.68 (m, 1H), 3.41 (m, 1H), 3.20 (m, 2H),
2.65 (m, 1H), 2.14 (m, 1H), 1.85 (m, 2H), 1.66 (m, 1H), 1.22-
1.55 (m, 7H), 0.86 (m, 1H), 0.82 (d, J ) 6.2 Hz, 3H), 0.75 (d, J
) 6.2 Hz, 3H), -0.56 (m, 1H). Anal. (C27H40N4O7‚0.2C6H6) C,
H, N.
Meth yl (6S,7R,10S)-7-Hexyl-6-[(h yd r oxya m in o)ca r bo-
n yl]-8-oxo-2-oxa-9-azabicyclo[10.2.2]h exadeca-1(14),12,15-
tr ien e-10-ca r boxyla te (53): 1H NMR (CD3OD) δ 8.38 (d, 1H),
7.20 (m, 1H), 7.07 (m, 2H), 6.87 (m, 1H), 5.00 (dd, 1H), 4.14
(m, 2H), 3.75 (s, 3H), 3.47 (dd, 1H), 2.63 (t, 1H), 2.00 (m, 1H),
1.69 (m, 1H), 1.35-1.10 (m, 12H), 0.86 (t, 3H), 0.78 (m, 1H),
(6S,7R,10S)-N10-[2-(3,5-Dim eth yl-1-p ip er id in yl)-2-oxo-
et h yl]-N6-h yd r oxy-7-isob u t yl-8-oxo-2-oxa -9-a za b icyclo-

2654 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
-0.55 (m, 1H); MS (ESI) m/z 435.3 (M + H)+; HRMS (FAB)
m/z 435.2515 [(M + H)+, calcd for C23H35N2O6 435.2495]. Anal.
(C23H34N2O6) C, H, N.
cyclo[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d ica r b ox-
a m id e (58). Compound 54v (50 mg) was dissolved in 50%
TFA/CH2Cl2 (5 mL). After the solution was stirred at room
temperature for 1 h, the volatiles were removed under reduced
pressure and the residue was triturated with ether to provide
58 as a solid: 1H NMR (CD3OD) δ 7.22 (m, 1H), 7.08 (m, 2H),
6.86 (m, 1H), 4.94 (m, 1H), 4.35 (m, 1H), 4.10 (m, 2H), 3.36
(m, 1H), 2.65 (m, 3H), 2.16 (m, 1H), 1.24-1.87 (m, 12H), 0.87
(m, 1H), 0.80 (d, J ) 6.2 Hz, 3H), 0.76 (d, J ) 6.2 Hz, 3H),
-0.57 (m, 1H); MS (ESI) m/z 520.3 (M + H)+. Anal. (C26H41N5O6‚
0.25CF3CO2H) C, H, N.
(6S,7R,10S)-N10-[(1S)-5-Am in o-1-(a m in oca r bon yl)p en -
tyl]-7-h exyl-N6-h yd r oxy-8-oxo-2-oxa -9-a za bicyclo[10.2.2]-
h exadeca-1(14),12,15-tr ien e-6,10-dicar boxam ide (59). Com-
pound 57 was obtained from 53g following the procedure
described for 56: 1H NMR (CD3OD) δ 7.25 (m, 1H), 7.08 (m,
2H), 6.87 (m, 1H), 4.96 (dd, 1H), 4.37 (dd, 1H), 4.13 (m, 2H),
3.39 (dd, 1H), 2.71 (m, 3H), 2.07 (m, 1H), 1.83 (m, 1H), 1.70
(m, 2H), 1.56 (m, 2H), 1.43 (m, 2H), 1.35-1.08 (m, 12H), 0.85
(t, 3H), 0.77 (m, 1H), -0.56 (m, 1H); MS (ESI) m/z 548.6 (M +
H)+. Anal. (C28H45N5O6‚0.5H2O) C, H, N.
(2S)-2-{[(Ben zyloxy)car bon yl]am in o}-6-[(tr iflu or oacety-
l)a m in o]h exa n oic Acid (61). To a suspension of NR-Cbz-L-
lysine (60) (5.6 g, 20 mmol) in H2O (40 mL) and dioxane (5
mL) cooled in an ice bath was added K2CO3 (5.53 g, 40 mmol)
followed by ethyl trifluoroacetate (7.14 mL, 60 mmol). The
mixture was stirred for 1 h and diluted with water. The
solution was extracted with ether 2×. The aqueous phase was
acidified with 10% citric acid solution and extracted with
EtOAc three times. The combined organic phase was washed
with brine 3×, dried (MgSO4), and concentrated to give 7.6 g
(100%) of the lysine derivative 61: MS (ESI) m/z 377.2 (M +
H)+.
Meth yl (2S)-2-{[(Ben zyloxy)ca r bon yl]a m in o}-6-[m eth -
yl(tr iflu or oa cetyl)a m in o]h exa n oa te (62). A mixture of 61
(20 g, 53.1 mmol), K2CO3 (36.7 g, 265 mmol), and iodomethane
(50 mL, 795 mmol) in DMF (150 mL) was stirred at 110 °C
for 10 h. Insoluble materials were filtered off, and the solution
was concentrated under reduced pressure. The residue was
taken up in EtOAc. The resulting solution was washed with
brine 6×, dried (MgSO4), and concentrated to give 19.45 g
(90%) of the ester 62: MS (ESI) m/z 405.2 (M + H)+. Anal.
(C18H23N2O5F3) C, H, N.
Meth yl (2S)-2-{[(Ben zyloxy)ca r bon yl]a m in o}-6-(m eth -
yla m in o)h exa n oa te (63). To a solution of 62 (2.02 g, 5 mmol)
in THF (15 mL) was added a solution of 1 N LiOH (15 mL).
The mixture was stirred at room temperature for 2 h and
acidified with 1 N HCl (20 mL). The solvents were removed
under reduced pressure, and the residue was taken up in
MeOH (20 mL). To it was added 4 N HCl/dioxane (50 mL).
The solution was stirred at room temperature overnight and
concentrated under reduced pressure, affording the crude 63,
which was used for the next reaction without purification: MS
(CI, NH3) m/z 309.
1-Ben zyl 4-ter t-Bu tyl (2R,3S)-2-Isobu tyl-3-(3-{[(4-n itr o-
p h en oxy)ca r bon yl]oxy}p r op yl)bu ta n ed ioa te (64). To a
solution of 31a (11.4 g, 33.1 mmol) and 4-nitrophenyl chloro-
formate (10.0 g, 50 mmol) in CH2Cl2 (50 mL) cooled in an ice
bath was slowly added NMM (4.4 mL, 40 mmol), and the
mixture was stirred at room temperature overnight. The
solvent was removed under reduced pressure, and the residue
was taken up in EtOAc (200 mL). The solution was washed
with brine 3×, dried (MgSO4), and concentrated. Purification
on a silica gel column eluting with 10% EtOAc/hexanes
provided 64 (15.0 g, 91%) as a pale yellow solid: MS (CI, NH3)
m/z 561 (M + NH4)+. Anal. (C29H37NO9) C, H, N.
(6S,7R,10S)-7-H exyl-N6-h yd r oxy-N10-m et h yl-8-oxo-2-
oxa -9-a za bicyclo[10.2.2]h exa d eca -1(14),12,15-tr ien e-6,10-
d ica r boxa m id e (55a ): 1H NMR (CD3OD) δ 8.17 (d, 1H), 7.20
(dd, 1H), 7.08 (m, 2H), 6.86 (dd, 1H), 4.85 (m, 1H), 4.14 (m,
2H), 3.17 (dd, 1H), 2.77 (s, 3H), 2.66 (t, 1H), 2.02 (m, 1H), 1.69
(m, 1H), 1.3-1.0 (m, 12H), 0.85 (t, 3H), 0.79 (m, 1H), -0.55
(m, 1H); MS (ESI) m/z 434.4 (M + H)+; HRMS (FAB) m/z
434.2667 [(M + H)+, calcd for C23H36N3O5 434.2668]. Anal.
(C23H35N3O5) C, H, N.
(6S,7R,10S)-N10-Ben zyl-7-h exyl-N6-h ydr oxy-8-oxo-2-oxa-
9-a za b icyclo[10.2.2]h exa d eca -1(14),12,15-t r ien e-6,10-d i-
ca r boxa m id e (55b): 1H NMR (CD3OD) δ 7.36-7.20 (m, 6H),
7.07 (m, 2H), 6.86 (m, 1H), 4.93 (m, 1H), 4.40 (m, 2H), 4.13
(m, 2H), 3.32 (m, 1H), 2.69 (t, 1H), 2.02 (m, 1H), 1.72 (m, 1H),
1.32-1.00 (m, 12H), 0.82 (t, 3H), 0.77 (m, 1H), -0.55 (m, 1H);
MS (ESI) m/z 510.5 (M + H)+. Anal. (C29H39N3O5) C, H, N.
(6S,7R,10S)-N10-{2-[4-(Am in osu lfon yl)p h en yl]eth yl}-7-
h e x y l-N 6-h y d r o x y -8-o x o -2-o x a -9-a za b ic y c lo [10.2.2]-
h exadeca-1(14),12,15-tr ien e-6,10-dicar boxam ide (55c): 1H
NMR (CD3OD) δ 8.18 (d, 1H), 7.84 (d, 2H), 7.40 (d, 2H), 7.19
(m, 1H), 7.07 (m, 2H), 6.85 (m, 1H), 4.83 (m, 1H), 4.15 (m,
2H), 3.46 (m, 2H), 3.20 (m, 1H), 2.90 (m, 2H), 2.61 (t, 1H),
2.04 (m, 1H), 1.72 (m, 1H), 1.36-1.08 (m, 12H), 0.83 (t, 3H),
0.77 (m, 1H), -0.55 (m, 1H); MS (ESI) m/z 603.5 (M + H)+;
HRMS (FAB) m/z 603.2867 [(M + H)+, calcd for C30H43N4O7S
603.2866]. Anal. (C30H42N4O7S) C, H, N.
(6S,7R,10S)-7-Hexyl-N6-h yd r oxy-N10-[3-(1H-im id a zol-1-
yl)pr opyl]-8-oxo-2-oxa-9-azabicyclo[10.2.2]h exadeca-1(14),-
12,15-tr ien e-6,10-d ica r boxa m id e (55d ): 1H NMR (CD3OD)
δ 7.68 (s, 1H), 7.22 (m, 1H), 7.15 (s, 1H), 7.08 (m, 2H), 6.96 (s,
1H), 6.87 (m, 1H), 4.86 (m, 1H), 4.14 (m, 2H), 4.06 (t, 2H),
3.24 (m, 3H), 2.70 (t, 1H), 2.00 (m, 3H), 1.74 (m, 1H), 1.35-
1.05 (m, 12H), 0.81 (m, 1H), 0.78 (t, 3H), -0.55 (m, 1H); MS
(ESI) m/z 528.5 (M + H)+; HRMS (FAB) m/z 528.3175 [(M +
H)+, calcd for C28H42N5O5 528.3173]. Anal. (C28H41N5O5‚H2O)
C, H, N.
(6S,7R,10S)-7-Hexyl-N6-h yd r oxy-N10-[2-(m eth yla m in o)-
2-oxoet h yl]-8-oxo-2-oxa -9-a za b icyclo[10.2.2]h exa d eca -
1(14),12,15-tr ien e-6,10-dicar boxam ide (55e): 1H NMR (CD3-
OD) δ 7.23 (m, 1H), 7.08 (m, 2H), 6.87 (m, 1H), 4.95 (m, 1H),
4.13 (m, 2H), 3.86 (m, 2H), 3.40 (m, 1H), 2.75 (s, 3H), 2.68 (t,
1H), 2.06 (m, 1H), 1.73 (m, 1H), 1.3-1.1 (m, 12H), 0.84 (t, 3H),
0.77 (m, 1H), -0.56 (m, 1H); MS (ESI) m/z 491.5 (M + H)+.
Anal. (C25H38N4O6) C, H, N.
(6S,7R,10S)-N10-[(1S)-1-(ter t-Bu toxym eth yl)-2-(m eth y-
la m in o)-2-oxoet h yl]-7-h exyl-N6-h yd r oxy-8-oxo-2-oxa -9-
a za bicyclo[10.2.2]h exa d eca -1(14),12,15-tr ien e-6,10-d ica r -
boxa m id e (55f): 1H NMR (CD3OD) δ 7.23 (m, 1H), 7.10 (m,
2H), 6.88 (m, 1H), 4.94 (dd, 1H), 4.38 (t, 1H), 4.14 (m, 2H),
3.67 (dd, 1H), 3.53 (dd, 1H), 3.37 (dd, 1H), 2.76 (s, 3H), 2.67
(t, 1H), 2.03 (m, 1H), 1.72 (m, 1H), 1.17 (s, 9H), 1.35-1.05 (m,
12H), 0.85 (t, 3H), 0.77 (m, 1H), -0.54 (m, 1H); MS (ESI) m/z
577.6 (M + H)+. Anal. (C30H48N4O7) C, H, N.
(6S,7S,10R)-N6-Hyd r oxy-7-isobu tyl-N10-m eth yl-8-oxo-2-
oxa -9-a za bicyclo[1 1.2.2]h ep ta d eca -1(15),13,16-tr ien e-6,-
10-d ica r boxa m id e (56): 1H NMR (CD3OD) δ 7.88 (m, 1H),
7.74 (m, 1H), 7.19 (m, 1H), 7.08 (m, 2H), 6.78 (m, 1H), 4.18
(m, 3H), 2.96 (m, 1H), 2.67 (m, 3H), 2.55 (m, 1H), 1.85-2.18
(m, 3H), 1.70 (m, 1H), 1.26-1.48 (m, 5H), 0.90 (m, 1H), 0.79
(d, J ) 6.6 Hz, 3H), 0.73 (d, J ) 6.6 Hz, 3H), 0.40 (m, 1H);
HRMS m/z 420.2512 [(M + H)+ calcd for C22H34N3O5 420.2498].
(6S,7S,10R)-N6-Hyd r oxy-7-isobu tyl-N10-m eth yl-8-oxo-2-
oxa -9-a za bicyclo[1 2.2.2]octa d eca -1(16),14,17-tr ien e-6,10-
d ica r boxa m id e (57): 1H NMR (CD3OD) δ 7.05 (m, 3H), 6.78
(m, 1H), 4.30 (m, 1H), 4.12 (m, 1H), 3.90 (m, 1H), 2.70 (m,
4H), 2.48 (m, 1H), 2.28 (m, 1H), 1.88 (m, 1H), 1.04-1.63 (m,
9H), 0.92 (m, 1H), 0.80 (d, J ) 6.2 Hz, 3H), 0.75 (d, J ) 6.2
Hz, 3H), 0.75 (m, 1H); MS (ESI) m/z 434 (M + H)+; HRMS
m/z 434.2655 [(M + H)+ calcd for C23H36N3O5 434.2655].
(6S,7R,10S)-N10-{(1S)-5-a m in o-1-[(m eth yla m in o)ca r bo-
n yl]p en t yl}-N6-h yd r oxy-7-isob u t yl-8-oxo-2-oxa -9-a za b i-
17-Ben zyl 16-ter t-Bu tyl 5-Meth yl (5S,16S,17R)-19-Meth -
yl-3,11-d ioxo-1-p h en yl-2,12-d ioxa -4,10-d ia za icosa n e-5,16,-
17-tr ica r boxyla te (65a ). To a solution of 64 (15.20 g, 27.28
mmol) and 63 (11.22 g, 32.78 mmol) in DMF (50 mL) was
added K2CO3 (15 g, 109 mmol). The mixture was stirred at 50
°C for 2 h. Insoluble materials were filtered off, and the filtrate
was diluted with EtOAc (300 mL). The resulting solution was
washed with 10% citric acid, brine, NaHCO3, and brine, dried

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2655
(MgSO4), and concentrated. Flash chromatography on silica
gel eluting with 15% EtOAc/hexanes provided 65a (17.0 g,
91%) as an oil: MS (ESI) m/z 699.5 (M + H)+. Anal.
(C38H54N2O10) C, H, N.
0.80 (d, 3H), 0.72 (d, 3H); MS (ESI) m/z 505.3 (M + Na)+. Anal.
(C20H33F3N4O6) C, H, N.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-2,10-d ioxo-N8-
p h en yl-1-oxa -3,9-d ia za cyclop en ta d eca n e-8,12-d ica r boxa -
m id e (68c): 1H NMR (DMSO-d6) δ 10.51 (s, 1H), 9.84 (s, 1H),
8.82 (s, 1H), 8.26 (d, 1H), 7.54 (d, 2H), 7.30 (t, 2H), 7.05-6.98
(m, 2H), 4.42 (m, 1H), 4.18 (m, 1H), 3.97 (m, 1H), 3.00 (m,
2H), 2.25 (m, 1H), 1.99 (m, 1H), 1.73-1.22 (m, 12H), 0.98-
0.70 (m, 7H); MS (ESI) m/z 499.3 (M + Na)+.
12-ter t-Bu t yl 8-Met h yl (8S,11R,12S)-11-Isob u t yl-2,10-
d ioxo-1-oxa -3,9-d ia za cyclop en ta d eca n e-8,12-d ica r boxy-
la te (66a ). To a solution of 65a (14.5 g, 20.8 mmol) in MeOH
(100 mL) was added 10% Pd on carbon (1.5 g). The mixture
was stirred under H2 (balloon) for 1 h. The catalyst was filtered
off, and the solvent was removed under reduced pressure. The
residue was taken up in DMF (50 mL). This solution was
dropwise added to a solution of BOP (9.2 g, 20.8 mmol) and
DIEA (12 mL, 70 mmol) in CHCl3 (1000 mL) cooled in an ice
bath. The mixture was allowed to stir at room temperature
overnight. The volatiles were removed under reduced pressure,
and the residue was taken up in EtOAc. The resulting solution
was washed with 5% citric acid, brine, NaHCO3, and brine,
dried (MgSO4), and concentrated. Purification on a silica gel
column eluting with 7% MeOH/CH2Cl2 provided 66a (6.6 g,
70%) as a powder: MS (ESI) m/z 457.4 (M + H)+.
Meth yl (8S,11R,12S)-12-{[(Ben zyloxy)am in o]car bon yl}-
11-isobu tyl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en ta d eca n e-
8-ca r boxyla te (67a ). Compound 66a (2.6 g, 5.5 mmol) was
dissolved in 50% TFA in CH2Cl2 (20 mL). The solution was
stirred for 1 h, and the volatiles were removed under reduced
pressure. The residue was taken up in DMF (10 mL), and the
solution was cooled in an ice bath. To it were added O-
benzylhydroxylamine hydrochloride (0.96 g, 6.15 mmol), DIEA
(4.3 mL, 24.6 mmol), and BOP (2.72 g, 6.15 mmol). The
solution was allowed to stir at room temperature overnight
and diluted with EtOAc. The resulting solution was washed
with 5% citric acid, brine, NaHCO3, and brine, dried (MgSO4),
and concentrated. The residue was triturated with ether to
give 67a (2.9 g, 90%) as a solid: MS (ESI) m/z 506.5 (M +
H)+.
(8S,11R,12S)-N12-Hydr oxy-11-isobu tyl-N8-[2-(4-m or ph oli-
n yl)-2-oxoeth yl]-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en ta d e-
ca n e-8,12-d ica r boxa m id e (68y). To a solution of 67a (1.1 g,
2 mmol) in THF (10 mL) was added 1 N LiOH (4 mL). The
mixture was stirred at room temperature for 1 h and acidified
with TFA. EtOAc was added, and the resulting solution was
washed with brine 3×, dried (MgSO4), and concentrated to give
the acid as a solid. The solid was dissolved in DMF (10 mL),
and the solution was cooled in an ice bath. To it were added
N-glycinylmorpholine (0.55 g, 3 mmol), BOP (0.9 g, 2 mmol),
and DIEA (1.75 mL, 10 mmol). The mixture was stirred at
room temperature overnight and diluted with EtOAc. The
resulting solution was washed with NaHCO3 3× and brine 3×,
dried (MgSO4), and concentrated to give a solid which was
dissolved in MeOH (30 mL). To the solution was added Pd/
BaSO4 (0.5 g). The mixture was stirred under H2 at 50 psi for
5 h. The catalyst was filtered off, and the solvent was removed
under reduced pressure. Purification by reversed-phase HPLC
afforded the hydroxamic acid 68y (81 mg, 82%) as a powder:
1H NMR (DMSO-d6) δ 10.50 (s, 1H), 8.20 (d, 1H), 7.70 (t, 1H),
6.95 (t, 1H), 4.20 (m, 1H), 3.95 (m, 4H), 3.55 (m, 4H), 3.40 (m,
4H), 3.00 (m, 2H), 2.45 (m, 1H), 2.00 (m, 1H), 1.6-1.2 (m, 12H),
0.90-0.75 (m, 7H); MS (ESI) m/z 528.5 (M + H)+. Anal.
(C24H41N5O8) C, H, N.
(8S,11R,12S)-N12-Hydr oxy-11-isobu tyl-N8-[4-(4-m or ph oli-
n yl)ph en yl]-2,10-dioxo-1-oxa -3,9-d ia za cyclopen ta d eca n e-
8,12-d ica r boxa m id e (68d ): 1H NMR (DMSO-d6) δ 10.50 (s,
1H), 9.70 (s, 1H), 8.20 (d, 1H), 7.20 (d, 2H), 6.95 (d, 2H), 6.80
(t, 1H), 4.20 (m, 1H), 3.80 (m, 10H), 3.00 (m, 2H), 2.4-2.0 (m,
2H), 1.6-1.2 (m, 12H), 0.90-0.75 (m, 7H); MS (ESI) m/z 562.3
(M + H)+. Anal. (C28H43N5O7‚CF3CO2H‚0.4H2O) C, H, N.
(8S,11R,12S)-N8-Ben zyl-N12-h yd r oxy-11-isobu tyl-2,10-
d ioxo-1-oxa -3,9-d ia za cyclop en ta d eca n e-8,12-d ica r boxa -
m id e (68e): 1H NMR (DMSO-d6) δ 10.50 (s, 1H), 8.81 (s, 1H),
8.14 (d, 1H), 8.05 (t, 1H), 7.32-7.19 (m, 5H), 6.95 (t, 1H), 4.25
(d, 2H), 4.23 (m, 2H), 3.97 (m, 1H), 3.10-2.88 (m, 2H), 2.43
(m, 1H), 1.95 (m, 1H), 1.65-1.22 (m, 12H), 0.86 (m, 1H), 0.73-
0.68 (m, 6H); MS (ESI) m/z 513.4 (M + Na)+. Anal. (C25H38N4O6‚
0.25H2O) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-2,10-d ioxo-N8-(2-
p yr id in ylm eth yl)-1-oxa -3,9-d ia za cyclop en ta d eca n e-8,12-
d ica r boxa m id e (68f): 1H NMR (DMSO-d6) δ 10.51 (s, 1H),
8.82 (s, 1H), 8.48 (d, 1H), 8.35-8.19 (m, 2H), 7.73 (t, 1H), 7.24
(m, 2H), 6.96 (m, 1H), 4.32 (d, 2H), 4.33-4.08 (m, 2H), 3.93
(m, 1H), 3.15-2.80 (m, 2H), 2.52 (m, 1H), 1.95 (m, 1H), 1.78-
1.11 (m, 12H), 0.86-0.67 (m, 7H); MS (ESI) m/z 492.3 (M +
H)+. Anal. (C24H37N5O6‚0.4H2O) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-2,10-d ioxo-N8-(3-
p yr id in ylm eth yl)-1-oxa -3,9-d ia za cyclop en ta d eca n e-8,12-
d ica r boxa m id e (68g): 1H NMR (DMSO-d6) δ 10.54 (s, 1H),
8.81 (s, 1H), 8.44 (bs, 2H), 8.37-8.11 (m, 2H), 7.59 (d, 1H),
7.31 (dd, 1H), 6.95 (m, 1H), 4.26 (d, 2H), 4.28-4.10 (m, 2H),
3.93 (m, 1H), 3.09-2.82 (m, 2H), 2.51 (m, 1H), 1.94 (m, 1H),
1.69-1.14 (m, 12H), 0.83 (m, 1H), 0.73 (d, 3H), 0.68 (d, 3H);
MS (ESI) m/z 492.3 (M + H)+. Anal. (C24H37N5O6‚0.25H2O) C,
H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-2,10-d ioxo-N8-(4-
p yr id in ylm eth yl)-1-oxa -3,9-d ia za cyclop en ta d eca n e-8,12-
d ica r boxa m id e (68h ): 1H NMR (DMSO-d6) δ 10.50 (s, 1H),
8.82 (s, 1H), 8.46 (dd, 2H), 8.30 (t, 1H), 8.18 (d, 1H), 7.19 (d,
2H), 6.97 (t, 1H), 4.35-4.10 (m, 2H), 4.27 (d, 2H), 3.93 (m,
1H), 3.16-2.85 (m, 2H), 2.49, (m, 1H), 1.95 (m, 1H), 1.72-
1.15 (m, 12H), 0.87 (m, 1H), 0.75 (d, 3H), 0.69 (d, 3H); MS
(ESI) m/z 492.3 (M + H)+. Anal. (C24H37N5O6‚HCl‚0.5MeOH)
C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-2,10-d ioxo-N8-[2-
(2-p yr id in yl)eth yl]-1-oxa -3,9-d ia za cyclop en ta d eca n e-8,-
12-d ica r boxa m id e (68i): 1H NMR (DMSO-d6) δ 10.47 (s, 1H),
8.77 (s, 1H), 8.45 (m, 1H), 8.02 (d, 1H), 7.64 (m, 2H), 7.18 (m,
2H), 6.90 (t, 1H), 4.13 (m, 2H), 3.88 (m, 1H), 3.34 (m, 2H),
3.08-2.87 (m, 2H), 2.78 (t, 2H), 2.42 (m, 1H), 1.90 (m, 1H),
1.60-1.03 (m, 12H), 0.83 (m, 1H), 0.75 (d, 3H), 0.69 (d, 3H);
MS (ESI) m/z 506.4 (M + H)+. Anal. (C25H39N5O6‚HCl‚H2O)
C, H, N.
(8S,11R,12S)-N12-Hydr oxy-11-isobu tyl-N8-[2-(6-m eth oxy-
1H-in d ol-3-yl)eth yl]-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en -
ta d eca n e-8,12-d ica r boxa m id e (68j): 1H NMR (DMSO-d6)
δ 10.60 (s, 1H), 10.46 (s, 1H), 8.75 (s, 1H), 8.06 (m, 1H), 7.57
(m, 1H), 7.21 (d, 1H), 7.07 (s, 1H), 6.99 (s, 1H), 6.70 (m, 1H),
6.68 (d, 1H), 4.21 (m, 1H), 4.19 (m, 2H), 3.91 (m, 1H), 3.73 (s,
3H), 3.30 (m, 2H), 3.01 (m, 1H), 2.92 (m, 1H), 2.71 (m, 2H),
2.01 (m, 1H), 1.39-1.23 (m, 12H), 0.96 (m, 1H), 0.80 (d, 3H),
0.72 (d, 3H); HRMS (FAB) m/z. 574.325412 [(M + H)+ calcd
for C29H44N5O7 574.325924]. Anal. (C29H43N5O7‚0.1CF3CO2H)
C, H, N.
The following compounds were prepared in
a manner
analogous to that described for 68y.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-N8-m eth yl-2,10-
d ioxo-1-oxa -3,9-d ia za cyclop en ta d eca n e-8,12-d ica r boxa -
m id e (68a ): 1H NMR (CD3OD) δ 8.43 (d, 1H), 7.78 (s, 1H),
6.76 (t, 1H), 4.34 (m, 2H), 3.97 (m, 1H), 3.25-3.02 (m, 2H),
2.68 (d, 3H), 2.57 (m, 1H), 2.07 (m, 1H), 1.77-1.23 (m, 12H),
1.01 (m, 1H), 0.87 (d, 3H), 0.81 (d, 3H); MS (ESI) m/z 437.3
(M + Na)+. Anal. (C19H34N4O6‚0.1CH2Cl2) C, H, N.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-2,10-d ioxo-N8-
(2,2,2-t r iflu or oet h yl)-1-oxa -3,9-d ia za cyclop en t a d eca n e-
8,12-d ica r boxa m id e (68b): 1H NMR (DMSO-d6) δ 10.50 (s,
1H), 8.81 (s, 1H), 8.37 (m, 1H), 8.17 (d, 1H), 6.96 (t, 1H), 4.31,
(m, 1H), 4.16, (m, 1H), 4.02-3.73 (m, 3H), 3.10-2.86 (m, 2H),
2.49 (m, 1H), 1.95 (m, 1H), 1.68-1.15 (m, 12H), 0.87 (m, 1H),
(8S,11R,12S)-N12-Hydr oxy-11-isobu tyl-N8-[2-(5-m eth oxy-
1H-in d ol-3-yl)eth yl]-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en -
ta d eca n e-8,12-d ica r boxa m id e (68k ): 1H NMR (DMSO-d6)
δ 10.57 (s, 1H), 10.48 (s, 1H), 8.77 (s, 1H), 8.06 (m, 1H), 7.60

2656 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
(m, 1H), 7.34 (m, 1H), 6.96 (s, 1H), 6.89 (m, 1H), 6.82 (s, 1H),
6.63 (m, 1H), 4.21 (m, 1H), 4.19 (m, 2H), 3.91 (m, 1H), 3.73 (s,
3H), 3.30 (m, 2H), 3.01 (m, 1H), 2.92 (m, 1H), 2.71 (m, 2H),
2.01 (m, 1H), 1.39-1.23 (m, 12H), 0.96 (m, 1H), 0.80 (d, 3H),
0.72 (d, 3H); HRMS (FAB) m/z 574.324074 [(M + H)+ calcd
for C29H44N5O7 574.324279]. Anal. (C29H43N5O7‚0.7H2O) C, H,
N.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-2,10-d ioxo-N8-
[(2R)-tetr a h yd r o-2-fu r a n ylm eth yl]-1-oxa -3,9-d ia za cyclo-
p en ta d eca n e-8,12-d ica r boxa m id e (68s): 1H NMR (DMSO-
d6) δ 10.53 (s, 1H), 8.78 (s, 1H), 8.07 (d, 1H), 7.51 (t, 1H), 6.91
(t, 1H), 4.16 (m, 2H), 3.89 (m, 1H), 3.71 (m, 3H), 3.57 (m, 1H),
3.15-2.80 (m, 4H), 2.45 (m, 1H), 1.94 (m, 1H), 1.76 (m, 3H),
1.60-1.05 (m, 12H), 0.84 (m, 1H), 0.77 (d, 3H), 0.71 (d, 3H);
MS (ESI) m/z 507.4 (M + Na)+. Anal. (C23N40N4O7) C, H, N.
(8S,11R,12S)-N8-[2-(5-F lu or o-1H-in d ol-3-yl)et h yl]-N12
-
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-N8-[2-(m et h y-
la m in o)-2-oxoeth yl]-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en -
ta d eca n e-8,12-d ica r boxa m id e (68t): 1H NMR (DMSO-d6)
δ 10.51 (s, 1H), 8.82 (s, 1H), 8.23 (d, 1H), 7.93 (t, 1H), 7.73 (m,
1H), 6.97 (t, 1H), 4.19 (m, 2H), 3.94 (m, 1H), 3.61 (d, 2H), 3.11-
2.88 (m, 2H), 2.56 (d, 3H), 2.52 (m, 1H), 1.98 (m, 1H), 1.64-
1.16 (m, 12H), 0.86 (m 1H), 0.81 (d, 3H), 0.74 (d, 3H); MS (ESI)
m/z 494.3 (M + Na)+. Anal. (C21H37N5O7) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-N8-[(1S)-1-m eth -
yl-2-(m et h yla m in o)-2-oxoet h yl]-2,10-d ioxo-1-oxa -3,9-d i-
azacyclopen tadecan e-8,12-dicar boxam ide (68u ): 1H NMR
(DMSO-d6) δ 10.51 (s, 1H), 8.82 (s, 1H), 8.17 (d, 1H), 7.84 (m,
1H), 7.54 (m, 1H), 6.95 (t, 1H), 4.16 (m, 2H), 4.01-3.79 (m,
2H), 3.11-2.84 (m, 2H), 2.56 (d, 3H), 2.49 (m, 1H), 1.95 (m,
1H), 1.64-1.15 (m, 12H), 1.13 (d, 3H), 0.88 (m 1H), 0.81 (d,
3H), 0.74 (d, 3H); MS (ESI) m/z 508.4 (M + Na)+. Anal.
(C22H39N5O7) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-N8-{(1S)-3-m eth -
yl-1-[(m eth yla m in o)ca r bon yl]bu tyl}-2,10-d ioxo-1-oxa -3,9-
d ia za cyclop en ta d eca n e-8,12-d ica r boxa m id e (68v): 1H
NMR (DMSO-d6) δ 10.51 (s, 1H), 8.82 (s, 1H), 8.14 (d, 1H),
7.89 (m, 1H), 7.54 (m, 1H), 6.94 (t, 1H), 4.22 (m, 3H), 3.92 (m,
1H), 3.16-2.84 (m, 2H), 2.54 (d, 3H), 2.47 (m, 1H), 1.95 (m,
1H), 1.62-1.17 (m, 15H), 0.91-0.73 (m, 13H); MS (ESI) m/z
550.4 (M + Na)+. Anal. (C25H45N5O7‚0.1H2O) C, H, N.
ter t-Bu tyl (5S)-5-[({(8S,11R,12S)-12-[(Hyd r oxya m in o)-
ca r b on yl]-11-isob u t yl-2,10-d ioxo-1-oxa -3,9-d ia za cyclo-
p en t a d eca n -8-yl}ca r b on yl)a m in o]-6-(m et h yla m in o)-6-
oxoh exylca r ba m a te (68w ): 1H NMR (DMSO-d6) δ 10.52 (s,
1H), 8.82 (s, 1H), 8.17 (d, 1H), 7.87 (m, 1H), 7.47 (m, 1H), 6.95
(t, 1H), 6.76 (t, 1H), 4.19 (m, 3H), 3.93 (m, 1H), 3.16-2.76 (m,
4H), 2.55 (d, 3H), 2.47 (m, 1H), 1.95 (m, 1H), 1.64-1.02 (m,
27H), 0.88 (m, 1H), 0.81 (d, 3H), 0.74 (d, 3H); MS (ESI) m/z
665.5 (M + Na)+. Anal. (C30H54N6O9) C, H, N.
(8S,11R,12S)-N8-{(1S)-2,2-Dim eth yl-1-[(m eth yla m in o)-
ca r bon yl]p r op yl}-N12-h yd r oxy-11-isobu tyl-2,10-d ioxo-1-
oxa-3,9-diazacyclopen tadecan e-8,12-dicar boxam ide (68x)
1H NMR (DMSO-d6) δ 10.52 (s, 1H), 8.82 (s, 1H), 8.31 (d, 1H),
7.97 (m, 1H), 7.26 (m, 1H), 6.95 (t, 1H), 4.23 (m, 2H), 4.15 (d,
1H), 3.93 (m, 1H), 3.14-2.83 (m, 2H), 2.55 (d, 3H), 2.48 (m,
1H), 1.97 (m, 1H), 1.64-1.17 (m, 12H), 0.86-0.82 (m 13H),
0.74 (d, 3H); MS (ESI) m/z 550.4 (M + Na)+. Anal. (C25H45N5O7‚
0.25H2O) C, H, N.
(8S,11R,12S)-N8-[2-(2,6-Dim eth yl-4-m or p h olin yl)-2-ox-
oeth yl]-N12-h yd r oxy-11-isobu tyl-2,10-d ioxo-1-oxa -3,9-d i-
a za cyclop en ta d eca n e-8,12-d ica r boxa m id e (68z): 1H NMR
(DMSO-d6) δ 10.50 (s, 1H), 8.15 (d, 1H), 7.65 (t, 1H), 6.95 (t,
1H), 4.2-3.3 (m, 11H), 3.00 (m, 2H), 2.30 (m, 1H), 2.00 (m,
1H), 1.6-1.2 (m, 12 H), 1.10 (d, 6H), 0.90-0.70 (m, 7H); MS
(ESI) m/z 556.3 (M + H)+. Anal. (C26H45N5O8‚0.5H2O) C, H,
N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-N8-[2-(4-m eth yl-
1-p ip er a zin yl)-2-oxoeth yl]-2,10-d ioxo-1-oxa -3,9-d ia za cy-
clopen tadecan e-8,12-dicar boxam ide (68aa): 1H NMR (DM-
SO-d6) δ 10.50 (s, 1H), 10.00 (br, 1H), 8.15 (d, 1H), 7.72 (t,
1H), 6.95 (t, 1H), 4.2-3.9 (m, 5H), 3.8-3.2 (m, 6H), 3.1-2.9
(m, 4H), 2.80 (s, 3H), 2.46 (m, 1H), 2.00 (m, 1H), 1.6-1.1 (m,
12H), 0.90-0.70 (m, 7H); MS (ESI) m/z 541.3 (M + H)+. Anal.
(C25H44N6O7‚1.4CF3CO2H) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-2,10-d ioxo-N8-[2-
oxo-2-(4-p h en yl-1- p ip er a zin yl)eth yl]-1-oxa -3,9-d ia za cy-
clopen tadecan e-8,12-dicar boxam ide (68bb): 1H NMR (DM-
SO-d6) δ 10.50 (s, 1H), 8.20 (d, 1H), 7.75 (t, 1H), 7.22 (m, 2H),
6.98 (m, 3H), 6.82 (t, 1H), 4.2-3.8 (m, 5H), 3.60 (m, 4H), 3.15
(m, 4H), 3.00 (m, 2H), 2.46 (m, 1H), 2.00 (m, 1H), 1.6-1.2 (m,
h yd r oxy-11-isobu tyl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en -
ta d eca n e-8,12-d ica r boxa m id e (68l): 1H NMR (DMSO-d6)
δ 10.76 (s, 1H), 10.41 (s, 1H), 8.63 (s, 1H), 8.01 (s, 1H), 7.50
(m, 2H), 7.10 (m, 2H), 6.84 (s, 1H), 6.79 (s, 1H), 4.02 (m, 2H),
3.94 (m, 2H), 3.31 (m, 2H), 3.01 (m, 1H), 2.94 (m, 1H), 2.77
(m, 2H), 2.01 (m, 1H), 1.39-1.30 (m, 12H), 0.96 (m, 1H), 0.81
(d, 3H), 0.74 (d, 3H); HRMS (FAB) m/z 562.308110 [(M + H)+
calcd for C28H41FN5O6 562.306285]. Anal. (C28H40FN5O6‚0.6CF3-
CO2H) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-N8-[2-(5-m eth yl-
1H-in d ol-3-yl)eth yl]-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en -
ta d eca n e-8,12-d ica r boxa m id e (68m ): 1H NMR (DMSO-d6)
δ 10.51 (s, 1H), 10.41 (s, 1H), 8.62 (s, 1H), 7.93 (s, 1H), 7.45
(s, 1H), 7.27 (s, 1H), 7.19 (s, 1H), 7.04 (s, 1H), 6.89 (s, 1H),
6.70 (s, 1H), 4.25 (m, 2H), 3.94 (m, 2H), 3.28 (m, 2H), 3.05 (m,
1H), 2.96 (m, 1H), 2.75 (m, 2H), 2.36 (s, 3H), 2.01 (m, 1H),
1.40-1.31 (m, 12H), 0.94 (m, 1H), 0.82 (d, 3H), 0.76 (d, 3H);
HRMS (FAB) m/z 558.329160 [(M + H)+ calcd for (C29H44N5O6)-
558.327853]. Anal. (C29H43N5O6‚0.8CF3CO2H) C, H, N.
(8S,11R,12S)-N12-Hydr oxy-N8-(2-h ydr oxy-2-ph en yleth yl)-
11-isobu tyl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en ta d eca n e-
8,12-d ica r boxa m id e (68n ): 1H NMR (DMSO-d6) δ 10.50 (s,
1H), 8.80 (s, 1H), 8.10 (d, 1H), 7.60 (t, 1H), 7.30 (m, 5H), 6.95
(t, 1H), 5.45 (m, 1H), 4.55 (m, 1H), 4.20 (m, 1H), 4.00 (m, 2H),
3.25 (m, 2H), 3.00 (m, 2H), 2.45 (m, 1H), 2.00 (m, 1H), 1.6-
1.2 (m, 13H), 0.80 (m, 6H); MS (ESI) m/z 543.2 (M + Na)+.
Anal. (C26H40N4O7‚0.7H2O) C, H, N.
Eth yl 4-[({(8S,11R,12S)-12-[(Hyd r oxya m in o)ca r bon yl]-
11-isobu tyl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en ta d eca n -
8-yl}ca r bon yl)a m in o]-1-p ip er id in eca r boxyla te (68o): 1H
NMR (DMSO-d6) δ 10.40 (s, 1H), 8.62 (s, 1H), 7.93 (m, 1H),
7.28 (s, 1H), 6.70 (s, 1H), 4.23 (m, 2H), 4.05 (q, 2H), 3.99 (m,
1H), 3.83 (m, 2H), 3.79 (m, 1H), 3.05 (m, 1H), 2.96 (m, 3H),
2.00 (m, 1H), 1.71 (m, 2H), 1.69(m, 2H), 1.54-1.19 (m, 13H),
1.17 (t, 3H), 0.96 (m, 1H), 0.82 (d, 3H), 0.76 (d, 3H); HRMS
(FAB) m/z. 556.334639 [(M + H)+ calcd for C26H46N5O8
556.334280]. Anal. (C26H45N5O8) C, H, N.
(8S,11R,12S)-N12-H yd r oxy-N8-[(1S,2R)-2-h yd r oxy-2,3-
d ih yd r o-1H-in d en -1-yl]-11-isobu tyl-2,10-d ioxo-1-oxa -3,9-
d ia za cyclop en ta d eca n e-8,12-d ica r boxa m id e (68p ): 1H
NMR (DMSO-d6) δ 10.47 (s, 1H), 8.77 (s, 1H), 8.26 (d, 1H),
7.37 (t, 1H), 7.19-7.05 (m, 3H), 6.98 (d, 1H), 6.93 (t, 1H), 5.13
(m, 1H), 4.97 (d, 1H), 4.40-4.10 (m, 3H), 3.88 (m, 1H), 3.08-
2.82 (m, 2H), 3.00 (dd, 1H), 2.75 (d, 1H), 2.45 (m, 1H), 1.92
(m, 1H), 1.62 (m, 3H), 1.47-1.11 (m, 9H), 0.69 (m 1H), 0.64
(d, 3H), 0.55 (d, 3H); MS (ESI) m/z 555.4 (M + Na)+. Anal.
(C27N40N4O7) C, H, N.
(8S,11R,12S)-N12-H yd r oxy-N8-[(1R ,2S)-2-h yd r oxy-2,3-
d ih yd r o-1H-in d en -1-yl]-11-isobu tyl-2,10-d ioxo-1-oxa -3,9-
d ia za cyclop en ta d eca n e-8,12-d ica r boxa m id e (68q): 1H
NMR (DMSO-d6) δ 10.48 (s, 1H), 8.78 (s, 1H), 8.26 (d, 1H),
7.42 (t, 1H), 7.21-7.01 (m, 4H), 6.92 (t, 1H), 5.07 (m, 1H), 4.98
(d, 1H), 4.34 (m, 2H), 4.18 (m, 1H), 3.89 (m, 1H), 3.11-2.85
(m, 2H), 2.99 (dd, 1H), 2.73 (d, 1H), 2.45 (m, 1H), 1.95 (m, 1H),
1.62 (m, 3H), 1.45-1.11 (m, 9H), 0.83 (m 1H), 0.72 (d, 3H),
0.65 (d, 3H); MS (ESI) m/z 555.4 (M + Na)+. Anal. (C27H40N4O7)
C, H, N.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-2,10-d ioxo-N8-
[(2S)-tetr a h yd r o-2-fu r a n ylm eth yl]-1-oxa -3,9-d ia za cyclo-
p en ta d eca n e-8,12-d ica r boxa m id e (68r ): 1H NMR (DMSO-
d6) δ 10.52 (s, 1H), 8.83 (s, 1H), 8.13 (d, 1H), 7.51 (t, 1H), 6.95
(t, 1H), 4.23 (m, 2H), 3.92 (m, 1H), 3.73 (m, 2H), 3.57-2.82
(m, 6H), 2.41 (m, 1H), 1.94 (m, 1H), 1.79 (m, 3H), 1.58-1.16
(m, 12H), 0.88 (m, 1H), 0.82 (d, 3H), 0.75 (d, 3H); MS (ESI)
m/z 507.4 (M + Na)+. Anal. (C23N40N4O7) C, H, N.

Hydroxamic Acids That Inhibit TNF-R Release
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16 2657
12H), 0.90-0.75 (m, 7H); MS (ESI) m/z 603.4 (M + H)+. Anal.
(C30H46N6O7‚0.5CF3CO2H) C, H, N.
2.45 (m, 3H), 2.00 (m, 1H), 1.6-1.2 (m, 12H), 0.90-0.75 (m,
7H); MS (ESI) m/z 564.3 (M + Na)+. Anal. (C25H43N5O8‚0.5H2O)
C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-2,10-d ioxo-N8-{2-
oxo-2-[4-(2-p yr id in yl)-1-p ip er a zin yl]eth yl}-1-oxa -3,9-d i-
azacyclopen tadecan e-8,12-dicar boxam ide (68cc): 1H NMR
(DMSO-d6) δ 10.50 (s, 1H), 8.20 (d, 1H), 8.15 (d, 1H), 7.75 (t,
1H), 7.55 (t, 1H), 6.95 (t, 1H), 6.82 (d, 1H), 6.65 (t, 1H), 4.3-
3.8 (m, 5H), 3.50 (m, 8H), 3.00 (m, 2H), 2.46 (m, 1H), 2.00 (m,
1H), 1.6-1.2 (m, 12H), 0.90-0.75 (m, 7H); MS (ESI) m/z 604.4
(M + H)+. Anal. (C29H45N7O7‚CF3CO2H‚0.5H2O) C, H, N.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-N8,3-d im et h yl-
2,10-d ioxo-1-oxa -3,9- d ia za cyclop en ta d eca n e-8,12-d ica r -
boxa m id e (69a ): 1H NMR (DMSO-d6) δ 10.55 (s, 1H), 8.30
(d, 1H), 7.40 (m, 1H), 4.30 (m, 1H), 4.00 (m, 1H), 3.70 (m, 1H),
3.30 (m, 1H), 3.00 (m, 1H), 2.76 (s, 3H), 2.55 (d, 3H), 2.45 (m,
1H), 2.05 (m, 1H), 1.7-1.1 (m, 12H), 0.90-0.75 (m, 7H); MS
(ESI) m/z 429.4 (M + H)+. Anal. (C20H36N4O6‚0.2H2O) C, H,
N.
Eth yl 4-{[({(8S,11R,12S)-12-[(Hydr oxyam in o)car bon yl]-
11-isobu tyl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en ta d eca n -
8-yl}ca r b on yl)a m in o]a cet yl}-1-p ip er a zin eca r b oxyla t e
(68d d ): 1H NMR (DMSO-d6) δ 10.50 (s, 1H), 8.18 (d, 1H), 7.75
(t, 1H), 6.95 (t, 1H), 4.4 (m, 1H), 4.20 (m, 2H), 4.02 (q, 2H),
3.95 (m, 2H), 3.40 (m, 8H), 3.00 (m, 2H), 2.45 (m, 1H), 2.00
(m, 1H), 1.6-1.2 (m, 12 H), 1.18 (t, 3H), 0.90-0.70 (m, 7H);
MS (ESI) m/z 599.4 (M + H)+. Anal. (C27H46N6O9‚0.2CF3CO2H)
C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-3-m eth yl-N8-[2-
(4-m or p h olin yl)-2-oxoet h yl]-2,10-d ioxo-1-oxa -3,9-d ia za -
cyclop en ta d eca n e-8,12-d ica r boxa m id e (69b): 1H NMR
(DMSO-d6) δ 10.55 (s, 1H), 8.32 (d, 1H), 7. 75 (m, 1H), 4.40
(m, 1H), 4.00 (m, 2H), 3.70 (m, 2H), 3.60 (m, 4H), 3.40 (m,
5H), 3.00 (m, 1H), 2.80 (s, 3H), 2.45 (m, 1H), 2.02 (m, 1H),
1.7-1.2 (m, 12H), 0.90-0.75 (m, 7H); MS (ESI) m/z 542.3 (M
+ H)+. Anal. (C25H43N5O8‚0.2CF3CO2H) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-N8-[2-(4-h yd r oxy-1-p ip er id i-
n yl)-2-oxoet h yl]-11-isob u t yl-3-m et h yl-2,10-d ioxo-1-oxa -
3,9-d ia za cyclop en ta d eca n e-8,12-d ica r boxa m id e (69c): 1H
NMR (DMSO-d6) δ 10.05 (s, 1H), 8.30 (d, 1H), 7.65 (t, 1H),
4.70 (m, 1H), 4.40 (m, 1H), 4.00 (m, 3H), 3.80 (m, 2H), 3.30
(m, 2H), 3.02 (m, 4H), 2.80 (s, 3H), 2.45 (m, 1H), 2.00 (m, 1H),
1.8-1.2 (m, 16H), 0.90-0.75 (m, 7H); MS (ESI) m/z 556.5 (M
+ H)+. Anal. (C26H45N5O8‚0.5CF3CO2H‚0.8H2O) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-3-m eth yl-N8-[2-
(4-m et h yl-1-p ip er a zin yl)-2-oxoet h yl]-2,10-d ioxo-1-oxa -
3,9-diazacyclopen tadecan e-8,12-dicar boxam ide (69d): 1H
NMR (DMSO-d6) δ 10.50 (s, 1H), 8.30 (d, 1H), 7.95 (t, 1H),
4.40 (m, 2H), 4.00 (m, 4H), 3.70 (m, 1H), 3.40 (m, 4H), 3.00
(m, 4H), 2.81 (s, 3H), 2.78 (s, 3H), 2.45 (m, 1H), 2.00 (m, 1H),
1.6-1.2 (m, 12H), 0.90-0.73 (m, 7H); MS (ESI) m/z 555.5 (M
+ H)+. Anal. (C26H46N6O7‚CF3CO2H‚1.5H2O) C, H, N.
Eth yl 4-{[({(8S,11R,12S)-12-[(Hydr oxyam in o)car bon yl]-
11-isobu tyl-3-m eth yl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en -
t a d eca n -8-yl}ca r b on yl)a m in o]a cet yl}-1-p ip er a zin eca r -
boxyla te (69e): 1H NMR (DMSO-d6) δ 10.55 (s, 1H), 8.20 (d,
1H), 7.60 (d, 1H), 4.35 (m, 1H), 4.00 (m, 3H), 3.8-3.6 (m, 5H),
3.00 (m, 3H), 2.75 (s, 3H), 2.40 (m, 1H), 2.00 (m, 1H), 1.7-1.0
(m, 19H), 0.88-0.70 (m, 7H); MS (ESI) m/z 570.5 (M + H)+.
Anal. (C27H47N5O8‚0.5CF3CO2H‚H2O) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-N8-[2-(4-h yd r oxy-1-p ip er id i-
n yl)-2-oxoeth yl]-11-isobu tyl-2,10-d ioxo-1-oxa -3,9-d ia za cy-
clopen tadecan e-8,12-dicar boxam ide (68ee): 1H NMR (DM-
SO-d6) δ 10.50 (s, 1H), 8.18 (d, 1H), 7.62 (m, 1H), 6.95 (m, 1H),
4.20 (m, 2H), 3.90 (m, 3H), 3.60 (m, 2H), 3.05 (m, 6H), 2.45
(m, 1H), 2.00 (m, 1H), 1.7-1.2 (m, 16H), 0.90-0.75 (m, 7H);
MS (ESI) m/z 542.4 (M + H)+. Anal. (C25H43N5O8‚H2O) C, H,
N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-N8-{2-[4-(1-p ip -
er id in yl)-1-p ip er id in yl]-2-oxoeth yl}-2,10-d ioxo-1-oxa -3,9-
d ia za cyclop en ta d eca n e-8,12-d ica r boxa m id e (68ff): 1H
NMR (DMSO-d6) δ 10.50 (s, 1H), 9.20 (br, 1H), 8.20 (d, 1H),
7.65 (t, 1H), 6.95 (t, 1H), 4.45 (m, 1H), 4.20 (m, 2H), 3.95 (m,
6H), 3.40 (m, 3), 3.00 (m, 4H), 2.55 (m, 1H), 2.00 (m, 1H), 1.8-
1.2 (m, 22H), 0.90-0.75 (m, 7H); MS (ESI) m/z 609.4 (M +
H)+. Anal. (C30H52N6O7‚1.9CF3CO2H) C, H, N.
1-{[({(8S,11R,12S)-12-[(H yd r oxya m in o)ca r b on yl]-11-
isob u t yl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en t a d eca n -8-
yl}ca r bon yl)a m in o]a cetyl}-4-p ip er id in eca r boxylic Acid
(68gg): 1H NMR (DMSO-d6) δ 10.45 (s, 1H), 8.10 (d, 1H), 7.60
(t, 1H), 6.90 (t, 1H), 4.10 (m, 3H), 4.0-3.6 (m, 6H), 3.0-2.8
(m, 3H), 2.0-1.7 (m, 2H), 1.6-1.1 (m, 16H), 0.85-0.68 (m, 7H);
MS (ESI) m/z 592.3 (M + Na)+. Anal. (C26H43N5O9‚0.4CF3-
CO2H) C, H, N.
(8S,11R,12S)-N8-[2-(Dim et h yla m in o)-2-oxoet h yl]-N12
-
h yd r oxy-11-isobu tyl-3-m eth yl-2,10-d ioxo-1-oxa -3,9-d ia za -
cyclop en ta d eca n e-8,12-d ica r boxa m id e (69f): 1H NMR
(DMSO-d6) δ 10.50 (s, 1H), 8.25 (d, 1H), 7.60 (t, 1H), 4.40 (m,
1H), 4.00 (m, 2H), 3.80 (m, 2H), 3.35 (m, 1H), 3.00 (m, 1H),
2.92 (s, 3H), 2.82 (s, 3H), 2.78 (s, 3H), 2.45 (m, 1H), 2.00 (m,
1H), 1.7-1.1 (m, 12H), 0.90-0.75 (m, 7H); MS (ESI) m/z 500.5
(M + H)+. Anal. (C23H41N5O7‚0.2CF3CO2H) C, H, N.
1-{[({(8S,11R,12S)-12-[(H yd r oxya m in o)ca r b on yl]-11-
isob u t yl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en t a d eca n -8-
yl}ca r bon yl)a m in o]a cetyl}-3-p ip er id in eca r boxylic Acid
(68h h ): 1H NMR (DMSO-d6) δ 10.50 (s, 1H), 8.18 (m, 1H),
7.62 (m, 1H), 6.95 (m, 1H), 4.2-3.6 (m, 9H), 3.3-2.7 (m, 3H),
2.40-1.8 (m, 4H), 1.7-1.2 (m, 14H), 0.90-0.75 (m, 7H); MS
(ESI) m/z 592.2 (M + Na)+. Anal. (C26H43N5O9‚0.5CF3CO2H)
C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-N8-[(1S)-1-m eth -
yl-2-(4-m or p h olin yl)-2-oxoeth yl]-2,10-d ioxo-1-oxa -3,9-d i-
azacyclopen tadecan e-8,12-dicar boxam ide (68ii): 1H NMR
(DMSO-d6) δ 10.52 (s, 1H), 8.80 (s, 1H), 8.20 (d, 1H), 7.74 (d,
1H), 6.95 (t, 1H), 4.70 (m, 1H), 4.20 (m, 2H), 3.95 (m, 1H),
3.50 (m, 8H), 3.00 (m, 2H), 2.46 (m, 1H), 2.00 (m, 1H), 1.6-
1.2 (m, 12H), 1.08 (d, 3H), 0.90-0.75 (m, 7H); MS (ESI) m/z
564.3 (M + Na)+. Anal. (C25H43N5O8‚0.5H2O) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-N8-[(1S)-1-(h yd r oxym eth yl)-
2-(4-m or p h olin yl)-2-oxoet h yl]-11-isob u t yl-2,10-d ioxo-1-
oxa-3,9-diazacyclopen tadecan e-8,12-dicar boxam ide (68jj)
1H NMR (DMSO-d6) δ 10.52 (s, 1H), 8.18 (d, 1H), 7.60 (d, 1H),
6.95 (t, 1H), 4.75 (m, 1H), 4.20 (m, 2H), 3.90 (m, 2H), 3.50 (m,
10H), 3.00 (m, 2H), 2.46 (m, 1H), 2.00 (m, 1H), 1.6-1.2 (m,
12H), 0.90-0.75 (m, 7H); MS (ESI) m/z 570.2 (M + Na)+. Anal.
(C25H43N5O9‚0.4CF3CO2H) C, H, N.
(8S,11R,12S)-N12-Hydr oxy-11-isobu tyl-N8-[3-(4-m or ph oli-
n yl)-3-oxop r op yl]-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en ta -
d eca n e-8,12-d ica r boxa m id e (68k k ): 1H NMR (DMSO-d6)
δ 10.52 (s, 1H), 8.10 (d, 1H), 7.56 (t, 1H), 6.95 (t, 1H), 4.20 (m,
1H), 3.9 (m, 2H), 3.6-3.2 (m, 8H), 3.15 (m, 2H), 3.00 (m, 2H),
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-3-m et h yl-2,10-
d ioxo-N8-[2-oxo-2-(1- p yr r olid in yl)eth yl]-1-oxa -3,9-d ia za -
cyclop en ta d eca n e-8,12-d ica r boxa m id e (69g): 1H NMR
(DMSO-d6) δ 10.55 (s, 1H), 8.32 (d, 1H), 7.70 (t, 1H), 4.42 (m,
1H), 4.02 (m, 2H), 3.80 (m, 2H), 3.30 (m, 5H), 3.00 (m, 1H),
2.80 (s, 3H), 2.45 (m, 1H), 2.02 (m, 1H), 1.80 (m, 4H), 1.7-1.2
(m, 12H), 0.90-0.75 (m, 7H); MS (ESI) m/z 526.5 (M + H)+.
Anal. (C25H43N5O7‚H2O) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-3-m eth yl-N8-[2-
(m eth yla m in o)-2-oxoeth yl]-2,10-d ioxo-1-oxa -3,9-d ia za cy-
clop en ta d eca n e-8,12-d ica r boxa m id e (69h ): 1H NMR (DM-
SO-d6) δ 10.55 (s, 1H), 8.35 (d, 1H), 7.90 (t, 1H), 7.75 (q, 1H),
4.40 (m, 1H), 4.05 (m, 1H), 3.75 (m, 1H), 3.60 (d, 2H), 3.30 (m,
1H), 3.00 (m, 1H), 2.80 (s, 3H), 2.60 (d, 3H), 2.45 (m, 1H), 2.00
(m, 1H), 1.60 (m, 2H), 1.40 (m, 10H), 0.90-0.75 (m, 7H); MS
(ESI) m/z 486.5 (M + H)+. Anal. (C22H39N5O7‚0.5CF3CO2H‚
0.4H2O) C, H, N.
(8S,11R,12S)-N8-{(1S)-5-Am in o-1-[(m et h yla m in o)ca r -
bon yl]p en tyl}-N12-h yd r oxy-11-isobu tyl-2,10-d ioxo-1-oxa -
3,9-d ia za cyclop en ta d eca n e-8,12-d ica r boxa m id e (70). To
a solution of 68w (51 mg, 0.079 mmol) in dioxane (1 mL) was
added 4 N HCl in dioxane (2 mL). The solution was stirred at
room temperature for 9 h and concentrated. The residue was

2658 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 16
Xue et al.
triturated with MeOH/ether to provide 70 (38 mg, 82%) as an
HCl salt: 1H NMR (DMSO-d6) δ 10.52 (s, 1H), 8.19 (d, 1H),
7.91 (m, 1H), 7.76 (bs, 3H), 7.58 (d, 1H), 6.95 (t, 1H), 4.16 (m,
2H), 3.82 (m, 2H), 3.09-2.85 (m, 2H), 2.73 (m, 2H), 2.56 (d,
3H), 2.47 (m, 1H), 1.95 (m, 1H), 1.66-1.04 (m, 18H), 0.88 (m,
1H), 0.81 (d, 3H), 0.74 (d, 3H); MS (ESI) m/z 543.4 (M + H)+.
Anal. (C25H46N6O7‚HCl‚0.6MeOH) C, H, N.
ter t-Bu tyl (8S,11R,12S)-11-Isobu tyl-8-({[4-(2-m eth oxy-
2-oxoeth yl)-1,3-th ia zol-2-yl]a m in o}ca r bon yl)-2,10-d ioxo-
1-oxa -3,9-d ia za cyclop en ta d eca n e-12-ca r boxyla te (71). To
a solution of 66a (0.7 g, 1.53 mmol) in THF (5 mL) was added
a solution of 1 N LiOH (5 mL). The solution was stirred at
room temperature for 1 h. EtOAc (50 mL) was added followed
by 1 N HCl (10 mL). The organic phase was separated, washed
with brine 3×, dried (MgSO4), and concentrated. The residue
was taken up in DMF (5 mL). To it were added methyl
2-amino-4-thiazoleacetate hydrochloride salt (0.58 g, 2.8 mmol),
BOP (0.80 g, 1.8 mmol), and DIEA (1.74 mL, 10 mmol). The
solution was stirred at room temperature overnight. EtOAc
(100 mL) was added. The solution was washed with brine 2×,
10% citric acid 2×, brine 2×, NaHCO3 2×, and brine 2×, dried
(MgSO4), and concentrated. The residue was triturated with
ether to give 71 (700 mg, 78%) as a solid: MS (ESI) m/z 619.1
(M + Na)+.
(t, 1H), 7.05 (m, 1H), 4.40 (m, 1H), 3.95 (m, 2H), 3.80 (m, 2H),
3.30 (m, 1H), 3.00 (m, 1H), 2.75 (s, 3H), 2.45 (m, 1H), 2.00 (m,
1H), 1.60 (m, 2H), 1.35 (m, 10H), 0.88-0.72 (m, 7H); MS (ESI)
m/z 549.5 (M + H)+. Anal. (C26H40N6O7‚CF3CO2H‚2.4H2O) C,
H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-3-m eth yl-N8-{2-
[(4-m eth yl-1,3-th iazol-2-yl)am in o]-2-oxoeth yl}-2,10-dioxo-
1-oxa-3,9-diazacyclopentadecane-8,12-dicarboxamide (74b):
1H NMR (DMSO-d6) δ 12.05 (s, 1H), 10.55 (s, 1H), 8.80 (br,
1H), 8.30 (d, 1H), 8.00 (t, 1H), 7.76 (s, 1H), 4.45 (m, 1H), 4.00
(m, 2H), 3.80 (m, 2H), 3.30 (m, 1H), 3.05 (m, 1H), 2.80 (s, 3H),
2.46 (m, 1H), 2.23 (s, 3H), 2.02 (m, 1H), 1.63 (m, 2H), 1.5-1.2
(m, 10H), 0.90-0.75 (m, 7H); MS (ESI) m/z 569.3 (M + H)+.
Anal. (C25H40N6O7S‚0.2CF3CO2H) C, H, N.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-3-m et h yl-2,10-
d ioxo-N8-(2-p yr id in yl)-1-oxa -3,9-d ia za cyclop en ta d eca n e-
8,12-d ica r boxa m id e (74c): 1H NMR (DMSO-d6) δ 10.55 (s,
1H), 10.35 (s, 1H), 8.45 (d, 1H), 8.30 (d, 1H), 8.00 (d, 1H), 7.80
(t, 1H), 7.10 (t, 1H), 4.65 (m, 1H), 4.05 (m, 1H), 3.95 (m, 1H),
3.40 (m, 1H), 3.02 (m, 1H), 2.80 (s, 3H), 2.45 (m, 1H), 2.00 (m,
1H), 1.70 (m, 2H), 1.5-1.2 (m, 10H), 0.90-0.75 (m, 7H); MS
(ESI) m/z 492.5 (M + H)+. Anal. (C24H37N5O6‚CF3CO2H‚
0.5H2O) C, H, N.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-3-m et h yl-2,10-
d ioxo-N8-(3-p yr id in yl)-1-oxa -3,9-d ia za cyclop en ta d eca n e-
8,12-d ica r boxa m id e (74d ): 1H NMR (DMSO-d6) δ 10.58 (m,
2H), 8.95 (s, 1H), 8.50 (d, 1H), 8.42 (d, 1H), 8.20 (d, 1H), 7.65
(t, 1H), 4.40 (m, 1H), 4.05 (m, 1H), 3.80 (m, 1H), 3.40 (m, 1H),
3.05 (m, 1H), 2.80 (s, 3H), 2.45 (m, 1H), 2.05 (m, 1H), 1.70 (m,
2H), 1.5-1.2 (m, 10H), 0.9-0.7 (m, 7H); MS (ESI) m/z 492.4
(M + H)+. Anal. (C24H37N5O6‚CF3CO2H‚0.8H2O) C, H, N.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-3-m et h yl-2,10-
d ioxo-N8-(4-p yr id in yl)-1-oxa -3,9-d ia za cyclop en ta d eca n e-
8,12-d ica r boxa m id e (74e): 1H NMR (DMSO-d6) δ 11.47 (s,
1H), 10.55 (s, 1H), 8.70 (d, 2H), 8.60 (d, 1H), 8.05 (d, 2H), 4.40
(m, 1H), 4.05 (m, 1H), 3.90 (m, 1H), 3.40 (m, 1H), 3.05 (m,
1H), 2.80 (s, 3H), 2.55 (m, 1H), 2.20 (m, 1H), 1.75 (m, 2H),
1.5-1.2 (m, 10H), 0.9-0.7 (m, 7H); MS (ESI) m/z 492.4 (M +
H)+. Anal. (C24H37N5O6‚CF3CO2H‚1.5H2O) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-3-m eth yl-N8-(4-
m eth yl-1,3-th ia zol-2-yl)-2,10-d ioxo-1-oxa -3,9-d ia za cyclo-
p en ta d eca n e-8,12-d ica r boxa m id e (74f): 1H NMR (DMSO-
d6) δ 12.00 (s, 1H), 10.55 (s, 1H), 8.45 (d, 1H), 6.75 (s, 1H),
4.60 (m, 1H), 4.00 (m, 2H), 3.30 (m, 1H), 3.00 (m, 1H), 2.80 (s,
3H), 2.40 (m, 1H), 2.20 (s, 3H), 2.00 (m, 1H), 1.65 (m, 2H),
1.5-1.2 (m, 10H), 0.9-0.7 (m, 7H); MS (ESI) m/z 512.4 (M +
H)+. Anal. (C23H37N5O6S‚0.9CF3CO2H) C, H, N.
TACE a n d MMP Assa ys. The enzymatic activity of porcine
TACE was determined at 25 °C with partially purified porcine
enzyme and a synthetic fluorogenic substrate, Mca-Pro-Leu-
Ala-Gln-Ala-Val-Dpa-Arg-Ser-Ser-Ser-Agr-NH2. (Mca ) [7-meth-
oxycoumarin-4-yl]acetyl, Dpa ) N-3-[2,4-dinitrophenyl]-L-2,3-
diaminopropionyl). Partially purified TACE was obtained from
porcine spleen following a previously described procedure.14
The enzymatic activities of MMPs were determined with
recombinant human version catalytic domains (CHEMICON
International, Inc., 28835 Single Oak Dr., Temecula, CA
92590) and a fluorogenic peptide substrate, Mca-Pro-Leu-Gly-
Leu-Dpa-Ala-Arg-NH2.43 The peptide substrate was diluted to
a final concentration of 10 µM in a buffer containing 50 mM
Tricine (pH 7.5), 100 mM NaCl, 10 mM CaCl2, and 1 mM
ZnCl2. Final enzyme concentrations in the assay were between
0.05 and 10 nM depending on the enzyme and the potency of
the inhibitor. Fluorescence measurements were performed in
a CytoFluor multiwell plate reader, series 4000. Cleavage of
internal quenched substrate liberates emission-active Mca
product, causing an increase in fluorescence emission at 395
nM (the excitation wavelength is 330 nM). The rate of emission
change is proportional to enzyme activity. Initial velocities,
in the presence or absence of inhibitor, were measured as
slopes of the linear portion of the product progress curves.
Apparent Ki values were determined by plotting the inhibitor
Meth yl {2-[({(8S,11R,12S)-12-[(Hyd r oxya m in o)ca r bo-
n yl]-11-isobu tyl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en ta d e-
ca n -8-yl}ca r bon yl)a m in o]-1,3-th ia zol-4-yl}a ceta te (73).
Compound 71 (700 mg, 1.2 mmol) was dissolved in TFA/CH2-
Cl2 (1:1, 30 mL). After the solution was stirred for 2 h at room
temperature, the solvents were removed under reduced pres-
sure. The residue was taken up in DMF (5 mL), and the
solution was cooled in an ice bath. To it were added hydroxy-
lamine hydrochloride (278 mg, 4 mmol), BOP (664 mg, 1.5
mmol), and DIEA (1.04 mL, 6 mmol). The solution was stirred
for 2 h and concentrated under reduced pressure. The residue
was triturated with ether. The resulting solid was purified by
reversed-phase HPLC to provide 73 (240 mg, 33%) as a
powder: 1H NMR (DMSO-d6) δ 12.20 (s, 1H), 10.50 (s, 1H),
8.30 (d, 1H), 7.00 (t, 1H), 6.95 (s, 1H), 4.43 (m, 1H), 4.10 (m,
2H), 3.65 (s, 3H), 3.60 (s, 2H), 3.00 (m, 2H), 2.42 (m, 1H), 2.02
(m, 1H), 1.6-1.1 (m, 12H), 0.88-0.70 (m, 7H); MS (ESI) m/z
578.3 (M + Na)+.
{2-[({(8S,11R,12S)-12-[(H yd r oxya m in o)ca r b on yl]-11-
isob u t yl-2,10-d ioxo-1-oxa -3,9-d ia za cyclop en t a d eca n -8-
yl}ca r bon yl)a m in o]-1,3-th ia zol-4-yl}a cetic Acid (75). To
a solution of 73 (55 mg, 0.1 mmol) in THF (2 mL) was added
1 N LiOH (1 mL). The solution was stirred at room temper-
ature for 2 h and acidified with 1 N HCl. The volatiles were
removed under reduced pressure, and the residue was purified
by reversed-phase HPLC to provide 75 (35 mg) as a powder:
1H NMR (DMSO-d6) δ 12.20 (s, 1H), 10.50 (s, 1H), 8.30 (d, 1H),
7.00 (t, 1H), 6.95 (s, 1H), 4.45 (m, 1H), 4.10 (m, 2H), 3.60 (s,
2H), 3.00 (m, 2H), 2.40 (m, 1H), 2.00 (m, 1H), 1.6-1.2 (m, 12H),
0.88-0.72 (m, 7H); MS (ESI) m/z 542.2 (M+H)+. Anal.
(C23H35N5O8S‚0.3CF3CO2H‚0.8H2O) C, H, N.
(8S,11R,12S)-N12-Hyd r oxy-11-isobu tyl-N8-{4-[2-(4-m or -
p h olin yl)-2-oxoet h yl]-1,3-t h ia zol-2-yl}-2,10-d ioxo-1-oxa -
3,9-d ia za cyclop en ta d eca n e-8,12-d ica r boxa m id e (76). To
a solution of 75 (50 mg, 0.092 mmol), morpholine (40 mg, 0.46
mmol), and DIEA (35 µL, 0.2 mmol) in DMF (2 mL) cooled in
an ice bath was added BOP (44 mg, 0.1 mmol). The mixture
was stirred for 2 h and purified by reversed-phase HPLC to
provide 76 (35 mg) as a powder: 1H NMR (DMSO-d6) δ 12.1
(s, 1H), 10.50 (s, 1H), 8.35 (d, 1H), 7.00 (t, 1H), 6.90 (s, 1H),
4.45 (m, 1H), 4.10 (m, 2H), 3.70 (s, 2H), 3.50 (m, 8H), 3.00 (m,
2H), 2.40 (m, 1H), 2.00 (m, 1H), 1.6-1.2 (m, 12H), 0.90-0.75
(m, 7H); MS (ESI) m/z 611.2 (M + H)+. Anal. (C27H42N6O8S‚
0.4CF3CO2H‚2H2O) C, H, N.
The following compounds were prepared in
a manner
analogous to that described for 73.
(8S,11R,12S)-N12-H yd r oxy-11-isob u t yl-3-m et h yl-2,10-
d ioxo-N8-[2-oxo-2-(2-p yr id in yla m in o)eth yl]-1-oxa -3,9-d i-
a za cyclop en ta d eca n e-8,12-d ica r boxa m id e (74a ): 1H NMR
(DMSO-d6) δ 10.50 (m, 2H), 8.30 (m, 2H), 8.00 (m, 2H), 7.75

Hydroxamic Acids That Inhibit TNF-R Release
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concentration dependence of the fractional velocity for each
enzyme, and fitting the data by a tight binding inhibition
equation.44
TNF -r Hu m a n Wh ole Blood Assa y. Blood is drawn from
normal donors into tubes containing 143 USP units of heparin/
10 mL. The heparinized human blood (225 µL) was plated
directly into 1 mL sterile polypropylene tubes. Compounds
were diluted in DMSO/serum-free medium and added to the
blood samples so the final concentrations of the compounds
were 50, 10, 5, 1, 0.5, 0.1, and 0.01 µM. The final concentration
of DMSO did not exceed 0.5%. Compounds were preincubated
for 15 min before the addition of 100 ng/mL LPS. Plates were
incubated for 5 h in an atmosphere of 5% CO2 in air. At the
end of 5 h, 750 µL of serum-free medium was added to each
tube and the samples were spun at 1200 rpm for 10 min. The
supernatant was collected off the top and assayed for TNF-R
production by a standard sandwich ELISA. The ability of the
compounds to inhibit TNF-R production by 50% compared to
DMSO-treated cultures was given by the IC50 value.
TNF -r Mou se Mod el Stu d ies. Test compounds were
administered to mice po at time zero. Immediately following
compound administration, mice received an ip injection of 20
mg of D-galactosamine plus 10 µg of LPS. One hour later,
animals were anesthetized and bled by cardiac puncture. Blood
plasma was evaluated for TNF-R levels by an ELISA specific
for mouse TNF-R.
P h a r m a cok in etic Stu d ies of SL422 a n d SP 057 in
Bea gle Dogs. Nine male beagle dogs weighing approximately
10 kg were studied. SL422 (0.74 mg/kg) and SP057 (0.50 mg/
kg) each was dosed intravenously to two animals in a 10%
ethanol solution, and blood samples were taken at 6, 15, and
30 min prior to dosing, and at 1, 2, 4, 6, 8, 12, and 24 h
postdosing. Two dogs were administered orally with SL422
(0.863 mg/kg), and three dogs were administered orally with
SP057 (2.2 mg/kg) in a 0.5% methocell suspension. Blood
samples were taken at 15, 30, and 45 min prior to dosing and
at 1, 2, 4, 6, 8, 12, and 24 h postdosing. Plasma concentrations
were determined by LC/MS/MS following solid-phase extrac-
tion. The limit of detection was 10 nM. Pharmacokinetic
parameters were calculated using WINNONLIN, and the data
given are expressed as the mean.
Ack n ow led gm en t. We thank Sherrill A. Nurnberg,
Dianna L. Blessington, Persymphonie B. Miller, Theresa
M. Dimeo, J ohn Giannaras, Robert J . Collins, Tracy L.
Taylor, and Paul J . Strzemienski for performing the in
vitro and in vivo studies, and Thomas A. Emm for the
pharmacokinetic studies.
Su p p or tin g In for m a tion Ava ila ble: Proton NMR spec-
tral data of nontarget compounds. This material is available
free of charge via the Internet at http://pubs.acs.org.
(22) Moreland, L.; Baumgartner, S.; Schiff, M.; Tindall, E.; Fleis-
chmann, R.; Weaver, A.; Ettlinger, R.; Cohen, S.; Koopman, W.;
Mohler, K.; Widmer, M.; Blosch, C. Treatment of rheumatoid
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