Achiral, selective CCK2 receptor antagonists based on a 1,3,5-benzotriazepine-2,4-dione template

Article abstract of DOI:10.1016/j.bmc.2007.12.047

Novel, achiral 1H-1,3,5-benzotriazepine-2,4(3H,5H)-diones have been prepared and structurally characterized. These compounds are potent CCK₂ receptor antagonists that display a high degree of selectivity over CCK₁ receptors.

Full text of DOI:10.1016/j.bmc.2007.12.047

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry 16 (2008) 2974–2983
Achiral, selective CCK2 receptor antagonists based
on a 1,3,5-benzotriazepine-2,4-dione template
John Spencer,a,ꢀ John Gaffen,a Eric Griffin,a Elaine A. Harper,a Ian D. Linney,a
Iain M. McDonald,a,* Sonia P. Roberts,a Mark E. Shaxted,a
Trushar Adatiab and Alan Bashallb
aJames Black Foundation, 68 Half Moon Lane, Dulwich, London SE24 9JE, UK
bDepartment of Health and Human Sciences, London Metropolitan University, North Campus,
166-220 Holloway Road, London N7 8DB, UK
Received 5 September 2007; revised 17 December 2007; accepted 20 December 2007
Available online 25 December 2007
Abstract—Novel, achiral 1H-1,3,5-benzotriazepine-2,4(3H,5H)-diones have been prepared and structurally characterized. These
compounds are potent CCK2 receptor antagonists that display a high degree of selectivity over CCK1 receptors.
ꢀ 2007 Elsevier Ltd. All rights reserved.
1. Introduction
anti-secretory therapy)6 and proliferation of certain GI
tract tumours.7,8
Cholecystokinin (CCK) and gastrin are related polypep-
tides that have the same C-terminal sequence (Gly-Trp-
Met-Asp-Phe-NH2).1 CCK acts on CCK1 receptors in
the periphery where their activation mediates gall blad-
der contraction, bile release, gastric emptying and pan-
creatic enzyme secretion.2 CCK can also stimulate
CCK1 receptors located in the CNS, an action that is
associated with regulation of satiety.3 CCK2 receptors
are present in the CNS also, where their stimulation
by CCK is associated with the mediation of anxiety,
panic and pain.4,5 On the other hand, peripherally lo-
cated CCK2 receptors are primarily activated by gastrin.
In addition to gastrin’s role as a stimulant of gastric acid
secretion, it is a key growth factor for the histamine-
storing enterochromaffin-like cells of the stomach. Thus
peripherally acting CCK2 receptor antagonists have
long attracted interest as one means of treating such gas-
trin-related conditions as gastro oesophageal reflux dis-
order, rebound hypersecretion (following cessation of
Amongst the many non-peptide ligands that have been
devised for CCK receptors, those based on benzodiaze-
pine (BDZ) ring systems are by far the most promi-
nent.9–11 In particular, the prototype non-peptide CCK1
antagonist devazepide is a 1,4-BDZ-based ligand wherein
the compound stereochemistry was pivotal to achieving
high CCK1 affinity and selectivity over CCK2 receptors.12
Thissame ring system formed the core of many potent and
selective CCK2 selective antagonists that followed.
Equally, ligands for both CCK receptor subtypes based
on a 1,5-BDZ ring system have been produced. In the case
of CCK2 receptor antagonists such as 1 (GR 199114X)
their selectivity was influenced by similar structural and
stereochemical influences as for the 1,4-BDZ-based com-
pounds (Fig. 1).13 However, a relatively minor structural
modification to the side chain functionality in an analo-
gous compound, specifically substitution of the methyl
group of the N-1-anilidoacetamide substituent of 2 by iso-
propyl (3), produced a marked change in biological pro-
file, causing switching from antagonist to agonist
behaviour at CCK1 receptors (Table 1).14 More highly
optimized derivatives of this type, such as 4 (GW7854),
inhibited food intake in rats following oral administration
and were explored as anorectic agents.15 Subsequently
1,4-BDZ-based analogues, containing the same side chain
motif, were also shown to display CCK1 receptor-medi-
ated agonism.16
Keywords: CCK; CCK2; Gastrin; Antagonist; Benzotriazepine.
*
Corresponding author. Present address: Argenta Discovery, 8/9 Spire
Green Centre, Flex Meadow, Harlow, Essex CM19 5TR, UK. Tel.:
+44 (0) 1279 645645; fax: +44 (0) 1279 645646; e-mail: iain.
mcdonald@argentadiscovery.com
ꢀ
Present address: School of Science, University of Greenwich at
Medway, Central Avenue, Chatham Maritime, Kent ME4 4TB, UK.
0968-0896/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2007.12.047

J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
2975
compounds were generally less effective than their struc-
turally closest chiral counterparts, because they were
limited to derivatives containing identical substituents
on the N-1 and N-5 positions (8). However, the absence
of a chiral centre in the case of 1,3,5-benzotriazepine-
2,4(3H,5H)-diones enables non-identical substituents
to be incorporated at N-1 and N-5 positions. The SAR
of these novel compounds (9–21) with respect to their
CCK receptor activity and subtype selectivity is
explored.
N
N
O
O
O
N
O
N
H
N
NH
O O
H
N
NH
O O
N
N
N
O
F
1 (GR 199114X)
8
pKi = 8.6 (hCCK2)
pKi = 7.1 (hCCK1)
pIC50 = 7.3 (mCCK2)
pIC50 = 6.1 (mCCK1)
2. Chemistry
Figure 1. Literature activity values and structures of 1,5-BDZ-based
CCK2 antagonists.
There are few reports on the preparation and utility of
1,3,5-benzotriazepine-2,4-diones.22–24 The synthesis of
the 1,3,5-benzotriazepine-2,4-diones as potential CCK
receptor ligands is described in Scheme 1. The differen-
tially alkylated diamines (22–25) were prepared by alkyl-
ation of previously reported or commercially available
N-alkyl-1,2-diamines.13 Treatment of compounds 22–
25 with phenyl isocyanatoformate25 led to the N-1, N-
5-disubstituted 1,3,5-benzotriazepine-2,4-diones 26–29.
Introduction of the N-3 carboxy alkyl substituent was
achieved by base-mediated alkylation with a suitably
protected haloalkyl carboxylic acid followed by depro-
tection to afford 30–34. 1,3,5-Benzotriazepine-2,4-diones
As part of our effort to obtain orally active, non-peptide
CCK2 receptor antagonists, consideration of the influ-
ence of ligand stereochemistry on receptor selectivity
of the BDZ-based antagonists led us to prepare 3-aza-
analogues of 1,4-BDZ-based CCK2 receptor antagonists
such as 6 (YF476) (Fig. 2).17 Significantly, these 1,3,4-
benzotriazepine-based compounds retained selectivity
for CCK2 receptors over CCK1 receptors even though
they were achiral. The most highly optimized derivatives
proved to be potent, orally active inhibitors of gastrin-
mediated gastric acid secretion in vivo.18 One obvious
advantage of an achiral CCK2 receptor antagonist is
that the resolution step, required to obtain all of the
1,4-BDZ-based CCK2 receptor antagonists as single
entities hitherto, is avoided.
O
O
O
O
N
N
NH
O
N
NH
NH
N
N
In this paper we extend the tactic that was used to ob-
tain 7, to 1,5-BDZ-based CCK2 antagonists, thereby
affording 1H-1,3,5-benzotriazepine-2,4(3H,5H)-diones.
This change resulted in the pendant N-3 aryl substituent
being attached to the 1,3,5-benzotriazepine ring by an
acetamide rather than urea linkage, a modification for
which there is precedent in the 1,5-BZD-based CCK li-
gands (e.g., 5).19 Although achiral 1,5-BDZ CCK2
antagonists have been obtained previously,20,21 these
O
NH
CO2H
N
6 (YF476)
7
pKi = 10.1 (hCCK2)
pKi = 7.5 (hCCK1)
pKI = 9.0 (hCCK2)
pKI = 5.0 (hCCK1)
Figure 2. Literature activity values and structures of 1,4-BDZ and
1,3,4-benzotriazepine-based CCK2 antagonists.
Table 1. Literature activity values and structures of 1,5-BDZ-based CCK1 agonists
R
N
O
O
3
1
N
H
N
X
N
Y
5
O O
Compounda
R
X
Y
hCCK2 (pKi)
hCCK1 (pKi)
gpCCK1
%CCKb
Ref.
pEC50
2
Me
i-Pr
i-Pr
i-Pr
NH
NH
NH
CH2
H
Inactive
86
102
6.3c
5.9
7.4
6.4
14
14
15
19
3
4 (GW7854)
H
7.6
8.0
6.1
7.3
6.9
6.8
CO2H
H
5
50
a 3R/3S stereochemistry.
b Percentage contraction of isolated guinea-pig gallbladder produced by test compound (30 lM) relative to that produced by CCK-8 (1 lM).
c pKB from shift of CCK-8 concentration–effect curve in presence of test compound (30 lM).

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J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
R2
R2
N
R2
N
O
O
O
O
O
NH
CO2H
b, then c or d
a
NH
O
N
O
m
NH
R1
N
R1
N
R1
22-25
26-29
30-34
cmpd.
cmpd.
R1
Ph
R2
m
R1
R2
pyrrolidin-1-yl
23, 27 c-C7H13 OBn
30
31
32
33
34
pyrrolidin-1-yl
1
1
1
1
3
22, 26 Ph
c-C7H13 OBn
24, 28
c-C7H13
c-C6H11
c-C6H11
t
-Bu
-Bu
c-C7H13
t-Bu
-Bu
t
t
25, 29 c-C6H11
t-Bu
R2
N
O
O O
X
N
H
N
e, 35f-35j
m
30-34
N
R1
O
36-40
H2N
X
e, 35a-35e
cmpd.
X
H
cmpd.
X
c, d or f
35a
35b
35f
CO2Bn
Me
R2
O
35g
35h
N(Me)Boc
S
N
35c
35d
35e
O O
R3
CH2CO2Bn
N
N
N
N
N
m
H
N
NH 35i
(CH2)2CO2t-Bu
SCH2CO2Bn
N
R1
N
O
N
O
35j
O
9, 10, 12-21
41 (R2 = OBn)
12 (R2 = pyrrolidin-1-yl)
HN
d, e pyrrolidine
Scheme 1. Synthesis of 1,3,5-benzotriazepines. Reagents: (a) PhO2CNCO/DMA; (b) NaH, Br(CH2)mCO2R (m = 1, 3; R = t-Bu, Bn)/DMF; (c)
CF3CO2H; (d) H2, Pd–C/THF–MeOH; (e) EDC, 1-HOBt, DMAP/DMF; (f) NaOH/H2O–CH3OH.
9, 12, 14, 18–21 were obtained directly on reaction of 30,
32–34 with anilines (35a–35e) using standard amide
bond forming conditions. Compounds 10, 15–17 were
obtained by initial reaction of 30, 32 or 33 with anilines
35f–35j, to afford intermediates 36–40, followed by re-
moval of the protecting group on the anilide substituent.
Reaction of 31 with toluidine (35b) afforded intermedi-
ate 41, which yielded compound 12 on manipulation
of the N-1 side chain substituent.
selected examples, of the pentagastrin-stimulated acid
secretion in a perfused, isolated rat stomach assay27
was performed to demonstrate functional CCK2 activity
(Table 2). Compounds 10, 15–17 and 19–21 were tested
as N-methyl-D-glucamine salts. Compound 13 was
tested as the hydrochloride salt.
Initial examples, 9 and 10, displayed micromolar affinity
at hCCK2 receptors with selectivity over hCCK1 recep-
tors. Strict comparison with the structurally closest 1,5-
BZD analogues13 is made difficult as the latter com-
pounds are racemic and were characterised in functional
bioassays. Nonetheless, based on its activity in the rat
stomach assay, where it did not produce acid output on
its own but inhibited pentagastrin-stimulated acid secre-
tion with comparable potency to that determined in the
hCCK2 radioligand binding assay, compound 10 can be
considered to be around 250-fold less potent than its clos-
est 1,5-BZD analogue. Previously published SAR on the
1,5-BDZ CCK2 antagonists had established that en-
hanced CCK2 affinity could be achieved by replacing
the N-5 phenyl ring by alicylic groups such as cyclohexyl
3. Results and discussion
The activity of 1,3,5-benzotriazepines 9–21 as CCK2
receptor antagonists was determined in a radioligand
binding assay by displacement of [125I]-BH-CCK-8S
from human, recombinant CCK2 receptors (hCCK2R)
expressed in NIH3T3 cell membranes.26 To assess their
selectivity for CCK2 receptors over CCK1 receptors,
they were also tested in a similar manner in a human, re-
combinant CCK1 receptor (hCCK1R) radioligand bind-
ing assay.17 Measurement of the inhibition, produced by

J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
Table 2. Biological data for 1,3,5-benzotriazepine-2,4-dione-based CCK2 antagonistsa
2977
R2
N
O
O O
X
N
N
O
m
H
N
R1
b
c
d
Compound
R1
R2
X
m
CCK2
CCK1
CCK2
NTe
9
10
11
12
13
14
15
16
17
18
19
20
21
Ph
Ph
Pyrrolidin-1-yl
Pyrrolidin-1-yl
Pyrrolidin-1-yl
t-Bu
H
1
1
1
1
1
1
1
1
1
1
1
1
3
6.00 0.07
6.42 0.10
7.63 0.09
8.12 0.04
8.02
<5.0
<5.0
6.21
6.35
CO2H
CH3
CH3
6.15 0.21
IAf
c-C7H13
c-C7H13
c-C6H11
c-C6H11
c-C7H13
c-C6H11
c-C7H13
c-C6H11
c-C7H13
c-C6H11
c-C6H11
NTe
t-Bu
t-Bu
NHCH3
2-Methyl-thiazol-4-yl
CH2CO2H
5.63 0.13
5.81
7.71 0.33
7.70 0.35
7.36 0.21
7.58 0.42
8.35 0.33
7.90 0.31
8.49 0.33
7.35 0.28
7.06 0.35
8.02
t-Bu
t-Bu
8.50 0.08
8.20
9.05
5.41 0.05
5.63 0.04
6.55 0.09
5.68 0.06
6.93 0.12
6.50
CH2CH2CO2H
SCH2CO2H
t-Bu
t-Bu
N-Methyl-((2H)-tetrazol-5-yl)-amino
1,2,4-Oxadiazol-3-yl-5(2H)-one
1,2,4-Oxadiazol-3-yl-5(2H)-one
1,2,4-Oxadiazol-3-yl-5(2H)-one
8.37
9.37
t-Bu
t-Bu
8.92 0.41
7.68
t-Bu
7.14 0.02
a Data were generally obtained from at least three separate experiments. Where no SEM was recorded, the data were from two experiments.
b pKI SEM values obtained from competition with 20 pM [125I]-BH-CCK-8S for recombinant, human CCK2 receptors expressed in NIH3T3 cell
membranes.
c pKI SEM values obtained from competition with 20 pM [3H]-L-364,718 for recombinant, human CCK1 receptors expressed in CHO-K1 cells
from at least two separate experiments.
d pA2 SEM values, estimated from single shifts of pentagastrin concentration–effect curves in isolated, lumen-perfused immature rat stomachs.
e Not tested.
f Inactive at concentration tested (10À6 M).
and cycloheptyl.13 In the 1,3,5-benzotriazepine series the
potency increased similarly with the preparation of 11
as judged by comparison with that of 9. Although 11
differed structurally from 9 still further in the nature of
the N-3 substituent, in that it contained a tolyl acetamide,
rather than a phenyl acetamide substituent at the N-3
position, it was at least 40-fold more potent at CCK2
receptors than 9, maintaining a high margin of selectivity
over CCK1 receptors. At a concentration of 10 lM, 11
failed to produce a shift of the pentagastrin concentra-
tion–effect curve in the rat functional bioassay, that might
have been expected based on its affinity in the CCK2
binding assay. This difference in activity between the
CCK2 assays for 11 can be ascribed to differing affinity
for human and rat CCK2 receptors. However, it can also
be attributed to low aqueous solubility of the compound,
as this property is more apparent in the functional tissue
bioassay which is less tolerant of organic co-solvents that
can otherwise be used to aid dissolution of poorly water
soluble compounds in the radioligand binding assay.
ORTEP diagram is presented in Figure 3. Although 26
lacks a substituent on the N-3 position, the seven-mem-
bered ring adopts a pseudo-boat conformation that is
not significantly different to that observed for 1,5-
BDZ-based CCK2 antagonists.13,29
As the presence of a polar substituent on the 3-aryl ring of
10 appeared to confer functional activity (since this was
not observed for compound 11) further derivatives that
incorporated polar groups were prepared. The relation-
ship between functional activity and the presence of polar
substituents was further demonstrated in the case of the
weakly basic compounds 13 and 14, which, although they
contained a cyclohexyl rather than a cycloheptyl group at
the N-5 position, showed only marginally lower potency
in the rat functional bioassay than they produced in the
hCCK2 binding assay. Moreover, the incorporation of
these substituents to obtain 13 and 14 did not significantly
Compound 12 displayed a further increase in affinity rel-
ative to 11 at hCCK2 receptors. This increase is similar
to that produced in the case of 1,4-BDZ containing
CCK2 antagonists, arising from substitution of N-1
acetamide substituents by alkylketomethylene groups
such as pinacoloyl.28 This apparent congruence in
SAR between 1,3,5-benzotriazepine containing com-
pounds and the 1,5-BDZ-based CCK2 antagonists is
consistent with the apparent similarity in core ring con-
formation. Single crystals of intermediate 26 were se-
lected for X-ray diffraction determination and the
Figure 3. ORTEP perspective view of 26 showing 30% probability
displacement ellipsoids.

2978
J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
alter the affinity, relative to 12, for hCCK2 receptors. In
considering incorporation of acid substituents their choice
was influencedby thosethathadnotonlyconferredinvitro
functional activity in the case of 1,3,4-benzotriazepines,
but which had also proved effective in achieving inhibition
of pentagastrin-stimulated acid secretion in in vivo as-
says.18 To this end the acetic acid (15), propionic acid
(16) and thioacetic acid (17) containing compounds were
prepared, each displaying high affinity at hCCK2 recep-
tors, and potency in the functional bioassay within 10-fold
of that determined in the radioligand binding assay. Selec-
tivity over CCK1 receptors of at least 300-fold was main-
tained for these compounds. Acid surrogates had proved
to be particularly effective previously and in line with this
trend the N-methyl amino-tetrazole (18) and oxadiazolone
(19 and 20) derivatives showed a similar profile to the car-
boxylic acid containing compounds across the in vitro bio-
assays, achieving around 1 nM affinity at hCCK2
receptors. In terms of its overall profile, 20 displayed a
greater difference in cross species activity than the equiva-
lent 1,3,4-benzotriazepine-based compound.18
vent systems or on a Biotage Quad purification unit.
1H NMR spectra were recorded on a Bruker DRX-
300 instrument and the chemical shifts (dH) were re-
corded relative to an internal standard Abbreviations:
DCM, dichloromethane; DMA, N,N-dimethylacet-
amide; THF, tetrahydrofuran; DMF, N,N-dimethyl-
formamide; DMAP, 4-(dimethylamino)pyridine; EDC,
1-(3-dimethylaminopropyl)-3-ethylcarbodiimide HCl;
HOBt, 1-hydroxybenzotriazole; TFA, trifluoroacetic
acid. Crystallographic data (excluding structure factors)
for compounds 26 and 29 have been deposited with the
Cambridge Crystallographic Data Centre as supplemen-
tary publication numbers CDC 659555 and CDC
659556, respectively. Copies of these data can be ob-
tained, free of charge, on application to CCDC, 12 Un-
ion Road, Cambridge, CB2 1EZ, UK, (fax: +44 (0)1223
336033). Tables of bond lengths and angles for com-
pounds 26 and 29 can be found in Supplementary data.
Compounds 10, 15–17 and 19–21 were tested as N-
methyl-D-glucamine salts. These salts were prepared by
stirring an aqueous mixture of the compound with one
equivalent of N-methyl-D-glucamine until a solution
was obtained (a minimum amount of 1,4-dioxan was
added if necessary to complete dissolution) and the solu-
tions were freeze-dried. Compound 13 was tested as the
hydrochloride salt which was obtained by suspending
the free-base in saturated methanolic-HCl. The solution
was evaporated to dryness and the resultant hydrochlo-
ride salt freeze-dried from an aqueous solution as above.
Selectivity over CCK1 receptors was maintained through-
out for these 1,3,5-benzotriazepine-based CCK2 antago-
nists. Only by lengthening the N-3 chain, as in 21, was
receptor selectivity significantly diminished. This arose
due tothecombination ofreducedCCK2 activityand high-
er CCK1 affinity being produced by this change. In the ab-
sence of functional assay data it remains unclear whether
21 has intrinsic activity at CCK1 receptors. Further manip-
ulation of 21, through changing the nature of the N-1 sub-
stituent, may ultimately provide CCK1 selective ligands,
particularly since these would bear a strong structural rela-
tionship to 1,5-BZD-based CCK1 agonists such as 5.
5.1. 2-((2-Phenylamino)-phenylamino)-pyrrolidin-1-yl
acetamide (22)
A
mixture of N-phenyl-1,2-phenylenediamine (1 g,
5.4 mmol), 2-bromo-1-pyrrolidin-1-yl ethanone30 (1.05 g,
5.5 mmol) and K2CO3 (0.84 g, 6.1 mmol) in DMF
(20 mL) was heated at 60 ꢁC for 16 h. On cooling, the
mixture was suspended in H2O/EtOAc (1:1/80 mL).
The organic layer was separated, washed successively
with H2O (2· 40 mL), brine (2· 40 mL) and dried
(MgSO4). Filtration and evaporation of the solvent fol-
lowed by chromatography (EtOAc/DCM (1:5)) of the
residue afforded 22 as a red oil (1.01 g, 63%). 1H
NMR (CDCl3) 7.32 (3H, m), 7.07 (1H, m), 6.83–6.64
(5H, m), 5.42 (1H, br s), 5.11 (1H, br s), 3.81 (2H, s),
3.49 (2H, t), 3.38 (2H, t), 1.94 (2H, m), 1.86 (2H, m).
4. Conclusion
In summary, the achiral 1,3,5-benzotriazepine ring sys-
tem has been employed as a template for the synthesis
of a series of potent CCK2 receptor antagonists. Except
for compound 21 these display a high degree of selectiv-
ity over CCK1 receptors. Influenced by some of the
trends in SAR evident in the earlier BDZ-based CCK li-
gands we have prepared derivatives that indicate a high
degree of congruence in CCK2 receptor activity between
the 1,3,5-benzotriazepine-based compounds and the
BDZs. The parallels in SAR, that are consistent with
the structural similarities of the respective ring system,
suggest these 1,3,5-benzotriazepine-containing com-
pounds interact with the CCK2 receptor most likely in
a similar manner. Where precise comparisons can be
made, the 1,3,5-benzotriazepine containing compounds
display a higher degree of selectivity over CCK1 recep-
tors than the BDZ-based compounds, which when cou-
pled with their ease of synthesis, since a resolution step is
avoided, offer a distinct advantage over the prior art.
5.2. (2-Cycloheptylamino-phenylamino)-acetic acid benzyl
ester (23)
A
mixture of N-cycloheptyl-benzene-1,2-diamine13
(1.0 g, 5.0 mmol), K2CO3 (0.69 g, 5.0 mmol) and 2-ben-
zyl bromoacetate (1.15 g, 5.0 mmol) in DMF (20 mL)
was stirred at ambient temperature for 16 h. The mix-
ture was suspended in H2O/EtOAc (1:1/40 mL). The or-
ganic layer was separated, washed successively with H2O
(2· 20 mL), brine (2· 20 mL) and dried (MgSO4). Filtra-
tion and evaporation of the solvent afforded 23 as a red
1
5. Experimental
oil (1.73 g, 98%). H NMR (CDCl3) 7.38 (5H, m), 6.82
(1H, dd), 6.75 (1H, dt), 6.62 (1H, dd), 6.58 (1H, dd),
5.22 (2H, s), 3.94 (3H, m), 3.42 (1H, m), 3.40 (1H, m),
1.97 (2H, m), 1.71–1.56 (10H, m).
Flash column chromatography was performed on
Merck silica gel 60 (40–63 lm) using the reported sol-

J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
2979
1
5.3. 1-(2-Cycloheptylamino-phenylamino)-3,3-dimethyl-
butan-2-one (24)
22 (42%). H NMR (CDCl3) 7.37 (1H, m), 7.27 (2H,
m), 6.99 (1H, m), 6.20 (1H, br s), 4.73 (2H, m), 3.80
(1H, m), 2.20–1.35 (7H, m), 1.34–1.26 (12H, m).
Compound 24 was obtained by the method used in the
preparation of 23 except that 1-bromo-3,3-dimethyl-bu-
tan-2-one was used instead of 2-benzyl bromoacetate
(91%). H NMR (CDCl3) 6.77 (2H, m), 6.61 (2H, m),
4.32 (1H, br s), 4.11 (2H, s), 3.43 (1H, m), 3.27 (1H,
m), 1.72 (1H, m), 1.62-1.41 (11H, m), 1.27 (9H, s).
5.9. (2,4-Dioxo-1-(2-oxo-2-pyrrolidin-1-yl-ethyl)-5-phe-
nyl-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic
acid (30)
1
Step A. Compound 26 (0.14 g, 0.38 mmol) and NaH (60%
suspension in mineral oil/0.02 g, 0.6 mmol) were stirred in
anhydrous DMF (10 mL) at ambient temperature for
0.5 h. tert-Butyl bromoacetate (0.11 g, 0.6 mmol) was
added and stirring was continued for 16 h. The reaction
mixture was suspended in H2O/EtOAc (1:1/25 mL). The
organic layer was separated, washed with H2O (2·
20 mL), brine (2· 20 mL) and dried (MgSO4). Filtration
and evaporation of the solvent followed by chromatogra-
phy (EtOAc/DCM (1:1)) of the oil obtained afforded (2,4-
dioxo-1-(2-oxo-2-pyrrolidin-1-yl-ethyl)-5-phenyl-1,2,4,
5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic acid tert-
butyl ester as a beige solid (0.16 g, 89%). 1H NMR
(CDCl3) 7.54 (2H, d), 7.42 (3H, m), 7.37 (1H, t), 7.17
(1H, dt), 7.07 (1H, dt), 6.84 (1H, dd), 4.67 (1H, d), 4.59
(1H, d), 4.25 (1H, d), 4.13 (1H, d), 3.54 (4H, m), 2.04–
1.85 (4H, m), 1.40 (9H, s).
5.4. 1-(2-Cyclohexylamino-phenylamino)-3,3-dimethyl-
butan-2-one (25)
Compound 25 was obtained by the method used in the
preparation of 23 except that N-cyclohexyl-benzene-1,2-
diamine13 and 1-bromo-3,3-dimethyl-butan-2-one were
used instead of N-cycloheptyl-benzene-1,2-diamine and
1
2-benzyl bromoacetate, respectively (100%). H NMR
(CDCl3) 6.78–6.71 (3H, m), 6.57 (1H, m), 4.20 (1H, br
s), 4.10 (2H, s), 3.23 (2H, m), 2.09 (2H, m), 1.82–1.76
(3H, m), 1.41–1.25 (14H, m).
5.5. 1-(2-Oxo-2-pyrrolidin-1-yl-ethyl)-5-phenyl-1H-1,3,5-
benzotriazepine-2,4(3H,5H)-dione (26)
A
solution of phenyl isocyanatoformate25 (0.6 g,
3.7 mmol) in DMA (3 mL) was added dropwise to a solu-
tion of compound 22 (0.7 g, 2.4 mmol) in DMA (40 mL)
maintained at 90 ꢁC. Heating was continued at 90 ꢁC
for 16 h. After cooling to ambient temperature, the reac-
tion mixture was suspended in H2O/EtOAc (2:1/75 mL).
The organic layer was separated, washed with brine (2·
50 mL) and dried (MgSO4). Filtration and evaporation
of the solvent followed by chromatography (EtOAc/
DCM (1:5) to neat EtOAc) of the oil obtained afforded
Step B. The product of the previous step was suspended
in TFA (4 mL) and stirred at ambient temperature for
1 h. The solution was concentrated in vacuo and re-evap-
orated from DCM (10 mL). The oil obtained was dis-
solved in DCM (20 mL), washed with H2O (20 mL)
and dried (MgSO4). Filtration and evaporation of the
solvent afforded 30 as an orange amorphous solid
1
(0.14 g, 96%). H NMR (CDCl3) 10.29 (1H, br s), 7.47–
7.34 (6H, m), 7.27 (1H, t), 7.10 (1H, t), 6.86 (1H, m),
4.67 (2H, m), 4.25 (2H, m), 3.59 (4H, m), 2.06 (2H, m),
1.94 (2H, m).
1
26 as a white solid (0.43 g, 49%). H NMR (DMSO-d6)
9.08 (1H, s), 7.52-7.23 (6H, m), 7.18 (1H, m), 7.11 (1H,
m), 7.74 (1H, d), 4.72 (2H, m), 3.51 (2H, m), 3.32 (2H,
m), 1.92 (2H, m), 1.77 (2H, m).
5.10. (1-Benzyloxycarbonylmethyl-5-cycloheptyl-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic
acid (31)
5.6. (1-Benzyloxycarbonylmethyl-5-cycloheptyl-2,4-dioxo-
1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic acid (27)
Compound 31 was obtained by steps A and B of the
method used in the preparation of 30 except that 27
was used instead of 26 in step A (41%). 1H NMR
(CDCl3) 10.15 (1H, br s), 7.37–7.24 (9H, m), 5.19 (2H,
m), 4.62 (1H, d), 4.44 (1H, d), 4.17 (2H, s), 3.93 (1H,
m), 2.19–1.44 (12H, m).
Compound 27 was obtained by the method used in the
preparation of 26 except that 23 was used instead of
22 (20%). H NMR (CDCl3) 7.61–7.23 (9H, m), 6.31
(1H, s), 5.20 (2H, s), 4.60 (1H, d), 4.47 (1H, d), 3.93
(1H, m), 2.18–1.21 (12H, m).
1
5.7. 1-Cycloheptyl-5-(3,3-dimethyl-2-oxo-butyl)-1H-
1,3,5-benzotriazepine-2,4(3H,5H)-dione (28)
5.11. (1-Cycloheptyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-dioxo-
1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic acid (32)
Compound 28 was obtained by the method used in the
preparation of 26 except that 24 was used instead of 22
(47%). H NMR (CDCl3) 7.32–7.19 (3H, m), 7.00 (1H,
m), 6.23 (1H, br s), 4.78 (1H, d), 4.67 (1H, d), 3.96 (1H,
m), 2.25 (2H, m), 1.94–1.79 (8H, m), 1.29 (11H, s).
Compound 32 was obtained by steps A and B of the
method used in the preparation of 30 except that 28
was used instead of 26 in step A (82%) 1H NMR
(CDCl3) 7.30–7.17 (3H, m), 7.04 (1H, m), 4.74 (2H,
m), 3.99 (3H, m), 2.28–1.52 (12H, m), 1.27 (9H, s).
1
5.8. 1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-1H-1,3,5-
benzotriazepine-2,4(3H,5H)-dione (29)
5.12. (1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-dioxo-
1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic acid (33)
Compound 29 was obtained by the method used in the
preparation of 26 except that 25 was used instead of
Step A. (1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-ace-

2980
J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
tic acid benzyl ester was obtained by step A of the meth-
od used in the preparation of 30 except that 29 and ben-
zyl 2-bromoacetate were used instead of 26 and tert-
butyl bromoacetate, respectively (88%). 1H NMR
(CDCl3) 7.33–7.18 (8H, m), 6.99 (1H, m), 5.09 (2H,
m), 4.76 (1H, d), 4.68 (1H, d), 4.33 (1H, d), 4.23 (1H,
d), 3.85 (1H, m), 2.18 (1H, m), 1.91–1.34 (6H, m),
1.33–1.25 (11H, m).
Filtration and evaporation of the solvent afforded an
oil from which 9 was obtained as an off-white solid from
DCM/hexanes (0.08 g, 43%). 1H NMR (CDCl3) 8.49
(1H, br s), 7.54 (2H, d), 7.43–7.33 (9H, m), 7.18 (1H,
m), 7.12 (1H, m), 6.88 (1H, d), 4.79 (1H, d), 4.60 (1H,
d), 4.32 (2H, s), 3.53 (4H, m), 1.99–1.84 (4H, m). HRMS
(ES, [M+H]+) calcd for C28H27N5O4 498.2136, found
498.2131.
Step B. The product of the previous step (0.88 g,
1.7 mmol) in THF/MeOH (1:1/30 mL) was stirred with
10% Pd/C (20 mg) under a hydrogen atmosphere at
ambient temperature for 16 h. The reaction mixture
was filtered through a pad of Celite. Concentration of
the filtrate in vacuo to afford an oil from which com-
pound 33 was obtained as a white solid on crystalliza-
tion from DCM/hexanes (0.72 g, 100%). 1H NMR
(CDCl3) 10.40 (1H, br s), 7.36 (1H, m), 7.25 (2H, m),
7.03 (1H, m), 4.75 (2H, m), 4.05 (2H, s), 3.85 (1H, m),
2.27 (1H, m), 1.99–1.65 (6H, m), 1.43–1.20 (12H, s).
5.15. 2-(1-Cycloheptyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-m-
tolyl-acetamide (12)
Compound 12 was obtained by the method used in the
preparation of 9 except that 32 and m-toluidine (35b)
were used instead of 30 and 35a, respectively (16%).
1H NMR (CDCl3) 8.20 (1H, br s), 7.39–7.10 (7H, m),
6.86 (1H, m), 4.77 (2H, m), 4.20 (2H, m), 4.05 (1H,
m), 2.36 (3H, s), 2.30–1.74 (12H, m), 1.25 (9H, s).
HRMS (ES, [M+H]+) calcd for C30H38N4O4 519.2966,
found 519.2966.
5.13. (1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-buty-
ric acid (34)
5.16. 2-(1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-(3-
(2-methyl-thiazol-4-yl)-phenyl)-acetamide (14)
Step A. Compound 29 (0.25 g, 0.7 mmol) and NaH (60%
suspension in mineral oil/0.04 g, 1.0 mmol) were stirred
in anhydrous DMF (10 mL) at ambient temperature
for 0.5 h. Benzyl 4-bromobutanoate31 (0.3 g, 1.2 mmol)
was added and the mixture was heated at 60 ꢁC for
72 h. On cooling, the reaction mixture was suspended
in H2O/EtOAc (1:1/25 mL). The organic layer was sepa-
rated, washed with brine (2· 20 mL) and dried (MgSO4).
Filtration and evaporation of the solvent followed by
chromatography (EtOAc/DCM (1:20–1:5)) of the
residue afforded (1-cyclohexyl-5-(3,3-dimethyl-2-oxo-
butyl)-2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-
3-yl)-butyric acid benzyl ester as a colourless oil. (0.21 g,
Compound 14 was obtained by the method used in the
preparation of 9 except that 33 and 3-(2-methyl-thia-
zol-4-yl)-phenylamine (35c)32 were used instead of 30
and 35a, respectively (70%). 1H NMR (CDCl3) 8.27
(1H, br s), 7.80 (1H, s), 7.46 (1H, d), 7.30 (3H, m),
7.28 (3H, m), 7.08 (1H, m), 4.83 (1H, d), 4.70 (1H, d),
4.21 (2H, m), 3.91 (1H, m), 2.89 (3H, s), 2.31 (1H, m),
1.94–1.85 (5H, m), 1.41 (1H, m), 1.35 (12H, m). HRMS
(ES, [M+H]+) calcd for C32H37N5O4 588.2639, found
588.2633.
5.17. 2-(1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-(3-
(methyl-(2H-tetrazol-5-yl)-amino)-phenyl)-acetamide (18)
1
57%). H NMR (CDCl3) 7.34–7.29 (5H, m), 7.27–7.16
(3H, m), 6.96 (1H, m), 5.01 (2H, s), 4.68 (2H, m), 3.83
(1H, m), 3.51 (2H, m), 2.15–1.75 (8H, m), 1.66–1.18
(11H, m).
Compound 18 was obtained by the method used in the
preparation of 9 except that 33 and N-methyl-N-(2H-tet-
razol-5-yl)-benzene-1,3-diamine (35d)33 were used in-
stead of 30 and 35a, respectively (6%). 1H NMR
(CDCl3) 10.50 (1H, br s), 8.50 (1H, br s), 7.36 (2H,
m), 7.26 (4H, m), 7.02 (1H, m), 7.00 (1H, m), 4.79
(1H, d), 4.67 (1H, d), 3.89 (1H, m), 3.55 (3H, s), 2.20
(1H, m), 1.96–1.63 (6H, m), 1.36–1.23 (12H, m). HRMS
(ES, [M+H]+) calcd for C30H37N9O4 588.3041, found
588.3046.
Step B. Compound 34 was obtained by step B of the
method used in the preparation of 20 except that the
product of the previous step was used instead of (1-
cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-dioxo-1,2,4,5-
tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic acid benzyl
ester (100%). 1H NMR (CDCl3) 7.32–7.18 (3H, m),
6.97 (1H,m), 4.68 (2H, m), 3.76–3.71 (3H, m), 3.50
(2H,m), 2.07–1.80 (8H,m), 1.70–1.23 (11H, m).
5.14. 2-(2,4-Dioxo-1-(2-oxo-2-pyrrolidin-1-yl-ethyl)-5-phe-
nyl-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-phenyl-
acetamide (9)
5.18. 2-(1-Cycloheptyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-(3-
(5-oxo-2,5-dihydro-[1,2,4]oxadiazol-3-yl)-phenyl)-acetam-
ide (19)
A mixture of compound 30 (0.15 g, 0.4 mmol), EDC
(0.1 g, 0.5 mmol), HOBt (0.07 g, 0.5 mmol), NEt3
(0.12 mL, 0.9 mmol), DMAP (10 mg) and aniline (35a)
(0.04 g, 0.4 mmol) in DMF (5 mL) was stirred at ambi-
ent temperature for 18 h. The reaction mixture was di-
luted with EtOAc (20 mL), washed successively with
H2O (2· 20 mL), brine (2· 20 mL) and dried (MgSO4).
Compound 19 was obtained by the method used in the
preparation of 9 except that 32 and 3-(3-amino-phe-
nyl)-2H-[1,2,4]oxadiazol-5-one (35e)34 were used instead
of 30 and 35a, respectively (39%). H NMR (CDCl3)
10.80 (1H, br s), 9.15 (1H, s), 8.02 (1H, s), 7.57–7.07
(8H, m), 4.85 (1H, d), 4.64 (1H, d), 4.31 (2H, s), 4.10
1

J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
2981
(1H, m), 1.72–1.25 (12H, m), 1.20 (9H, s). The com-
pound was further characterized as the N-methyl-D-
glucamine salt. Anal. Found: C, 55.98; H, 6.77; N,
10.91%; C31H36N6O6ÆC7H17NO5Æ1.5H2OÆ0.7dioxan re-
quires: C, 56.16; H, 7.12; N, 11.24%.
salt. Anal. Found C, 54.20; H, 6.03; N, 10.68%;
C29H27N5O6ÆC7H17NO5Æ3.5H2O requires: C, 54.06; H,
6.43; N, 10.51%.
5.23. (3-(2-(1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
acetylamino)-phenyl)-methyl-carbamic acid tert-butyl
ester (37)
5.19. 2-(1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-(3-
(5-oxo-2,5-dihydro-[1,2,4]oxadiazol-3-yl)-phenyl)-acet-
amide (20)
Compound 37 was obtained by the method used in the
preparation of 9 except that 33 and (3-amino-phenyl)-
methyl-carbamic acid tert-butyl ester (35g)17 were used
instead of 30 and 35a, respectively (89%). 1H NMR
(CDCl3) 8.30 (1H, br s), 7.43–7.27 (4H, m), 7.19–7.10
(3H, m), 6.95 (1H, d), 4.78 (1H, d), 4.73 (1H, d), 4.16
(2H, m), 3.90 (1H, m), 3.23 (3H, s), 2.28 (1H, m),
1.89–1.45 (6H, m), 1.45 (9H, s), 1.34–1.25 (12H, m).
Compound 20 was obtained by the method used in the
preparation of 9 except that 33 and 3-(3-amino-phenyl)-
2H-[1,2,4]oxadiazol-5-one (35e)34 were used instead of
1
30 and 35a, respectively (67%). H NMR (CDCl3) 9.15
(1H, s), 7.57–7.07 (9H, m), 4.82 (1H, d), 4.63 (1H, d),
4.31 (2H, m), 3.89 (1H, m), 2.17 (1H,m), 1.84–1.64 (6H,
m), 1.27–1.15 (12H, m). The compound was further char-
acterized as the N-methyl-D-glucamine salt. HRMS (ES,
[M+H]+) calcd for C30H34N6O6ÆC7H17NO5 770.3719,
found 770.3727.
5.24. 2-(1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-(3-
methylamino-phenyl)-acetamide (13)
5.20. 4-(1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-
dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-(3-
(5-oxo-2,5-dihydro-[1,2,4]oxadiazol-3-yl)-phenyl)-butyr-
amide (21)
Compound 13 was obtained by step B of the method
used in the preparation of 35j except that 37 was used
instead
of
benzyl-3-(N-tert-butyloxycarbonylami-
no)phenylthioacetate (32%). 1H NMR (CDCl3) 8.10
(1H, br s), 7.42 (1H, m), 7.28 (2H, m), 7.11–7.01 (3H,
m), 6.45 (1H, m), 6.36 (1H, d), 4.78 (1H, d), 4.67 (1H,
d), 4.19 (2H, m), 3.90 (1H, m), 2.81 (3H, s), 2.28 (1H,
m), 1.83–1.38 (6H, m), 1.35–1.21 (12H, m). The com-
pound was further characterized as the hydrochloride
salt. Anal. Found: C, 62.82; H, 6.76; N, 12.12%;
C39H37N5O4ÆHClÆ0.1dioxan requires: C, 62.51; H, 6.92;
N, 12.40%.
Compound 21 was obtained by the method used in the
preparation of 9 except that 34 and 3-(3-amino-phenyl)-
2H-[1,2,4]oxadiazol-5-one (35e)34 were used instead of
1
30 and 35a, respectively (20%). H NMR (CDCl3) 8.65
(1H, s), 7.89 (1H, s), 7.68 (1H, m), 7.53 (1H, m), 7.37–
7.20 (5H, m), 6.94 (1H, m), 4.90 (1H, d), 4.52 (1H, d),
3.84 (1H, m), 3.51 (2H, m), 2.18–1.47 (9H, m), 1.35–1.26
(16H, m). The compound was further characterized as
the N-methyl-D-glucamine salt. Anal. Found: C, 56.52;
H, 6.75; N, 11.40%. C32H38N6O6ÆC7H17NO5Æ1.5H2OÆ0.1-
dioxan requires: C, 56.76; H, 7.10; N, 11.76%.
5.25. (3-(2-(1-Cycloheptyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
acetylamino)-phenyl)-acetic acid benzyl ester (38)
5.21. 3-(2-(2,4-Dioxo-1-(2-oxo-2-pyrrolidin-1-yl-ethyl)-5-
phenyl-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acet-
ylamino)-benzoic acid benzyl ester (36)
Compound 38 was obtained by the method used in the
preparation of 9 except that 33 and 35h35 were used in-
stead of 30 and 35a, respectively (31%). 1H NMR
(CDCl3) 8.20 (1H, br s), 7.37–7.27 (11H, m), 7.20 (1H,
m), 7.07 (1H, m), 5.12 (2H, s), 4.77 (1H, d), 4.70 (1H,
d), 4.18 (1H, d), 4.13 (1H, d), 4.09 (1H, m), 3.61 (2H,
s), 2.04–1.32 (11H, m), 1.26 (9H, s).
Compound 36 was obtained by the method used in the
preparation of 9, except that 3-amino-benzoic acid ben-
zyl ester (35f) was used instead of 35a. 1H NMR
(CDCl3) 8.67 (1H, br s), 7.85–7.73 (3H, m), 7.53–7.21
(14H, m), 6.90 (1H, m), 5.34 (2H, s), 4.68 (2H, m),
4.30 (2H, m), 3.54 (4H, m), 2.04 (2H, m), 1.91 (2H, m).
5.26. (3-(2-(1-Cycloheptyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
acetylamino)-phenyl)-acetic acid (15)
5.22. 3-(2-(2,4-Dioxo-1-(2-oxo-2-pyrrolidin-1-yl-ethyl)-5-
phenyl-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acet-
ylamino)-benzoic acid (10)
Compound 15 was obtained by step B of the method
used in the preparation of 33 except that 38 was used
instead of (1-cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
Compound 10 was obtained by step B of the method
used in the preparation of 33 except that 36 was used
instead of (1-cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
acetic acid benzyl ester (34%). 1H NMR (DMSO-d6)
10.24 (1H, br s), 8.19 (1H, s), 7.69–7.17 (12H, m), 6.83
(1H, d), 4.91 (1H, d), 4.70 (1H, d), 4.26 (2H, m), 3.44
(4H, m), 1.90 (2H, m), 1.77 (2H, m). The compound
was further characterized as the N-methyl-D-glucamine
1
acetic acid benzyl ester (96%). H NMR (CDCl3) 8.20
(1H, br s), 7.34-7.21 (5H, m), 7.20 (1H, m), 7.19 (1H,
m), 6.97 (1H, m), 4.74 (1H, d), 4.63 (1H, d), 4.17 (2H,
m), 4.00 (1H, m), 3.46 (2H, s), 1.89–1.54 (12H, m),
1.24 (9H, s). The compound was further characterized
as the N-methyl-D-glucamine salt. Anal. Found: C,
57.91; H, 7.59; N, 7.97%; C31H38N4O6ÆC7H17NO5Æ1.5-
H2OÆdioxan requires: C, 57.78; H, 7.62; N, 8.02%.

2982
J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
5.27. 3-(3-(2-(1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
acetylamino)-phenyl)-propionic acid tert-butyl ester (39)
stead of 30 and 35a, respectively (89%). 1H NMR
(CDCl3) 8.33 (1H, br s), 7.40–7.30 (10H, m), 7.27–7.05
(3H, m), 5.14 (2H, s), 4.74 (2H, m), 4.16 (2H, m),
4.05 (1H, m), 4.00 (2H, s), 2.20–1.58 (12H, m), 1.29
(12H, m).
Compound 39 was obtained by the method used in the
preparation of 9 except that 33 and (3-amino-phenyl)-
propionic acid tert-butyl ester (35i)18 were used instead
5.31. (3-(2-(1-Cycloheptyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
acetylamino)-phenylsulfanyl)-acetic acid (17)
1
of 30 and 35a, respectively (100%). H NMR (CDCl3)
8.79 (1H, br s), 7.41 (1H, m), 7.40–7.28 (5H, m), 7.10
(1H, m), 6.94 (1H, m), 4.85 (1H, d), 4.68 (1H, d), 4.15
(2H, s), 3.89 (1H, m), 2.90 (2H, m), 2.62 (2H, m), 2.20
(1H, m), 1.79–1.39 (6H, m), 1.34–1.27 (12H, m), 1.25
(9H, s).
Compound 40 (0.12 g, 0.17 mmol) was dissolved in
MeOH (5 mL) and 1 N NaOH (0.25 mL, 0.25 mmol)
and after stirring at ambient temperature for 16 h
10% KHSO4 solution (2 mL) added. The resulting
white precipitate was collected by filtration, then dis-
solved in DCM (30 mL) and dried (MgSO4). Filtration
and evaporation of the solvent and trituration of the
residue with hexanes afforded 17 as a white solid,
which was isolated by filtration and dried in vacuo
(0.08 g, 77%). 1H NMR (CDCl3) 8.50 (1H, br s),
7.39–7.28 (6H, m), 7.15–7.05 (3H, m), 4.80 (1H, m),
4.70 (1H, m), 4.17 (2H, s), 4.03 (1H, m), 3.62 (2H,
s), 2.29–1.56 (12H, m), 1.26 (12H, m). The compound
was further characterized as the N-methyl-D-glucamine
salt. Anal. Found: C, 53.52; H, 6.97; N, 7.89%;
C31H38N4O6SÆC7H17NO5Æ3.5H2O requires: C, 53.51;
H, 7.33; N, 8.21%.
5.28. 3-(3-(2-(1-Cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
acetylamino)-phenyl)-propionic acid (16)
Compound 16 was obtained by step B of the method used
in the preparation of 30 except that 39 was used instead of
(2,4-dioxo-1-(2-oxo-2-pyrrolidin-1-yl-ethyl)-5-phenyl-1,2,
4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic
acid
1
tert-butyl ester (83%). H NMR (CDCl3) 8.79 (1H, br
s), 7.41 (1H, m), 7.40–7.28 (5H, m), 7.10 (1H, m), 6.94
(1H, m), 4.85 (1H, d), 4.68 (1H, d), 4.15 (2H, s), 3.89
(1H, m), 2.90 (2H, m), 2.62 (2H, m), 2.20 (1H, m), 1.79–
1.39 (6H, m), 1.34–1.27 (12H, m). The compound was
further characterized as the N-methyl-D-glucamine salt.
HRMS (ES, [M+H]+) calcd for C31H38N4O6ÆC7H17NO5
758.3971, found 758.3964.
5.32. (5-Cycloheptyl-2,4-dioxo-3-(m-tolylcarbamoyl-
methyl)-2,3,4,5-tetrahydro-1,3,5-benzotriazepin-1-yl)-
acetic acid benzyl ester (41)
5.29. (3-Amino-phenylsulfanyl)-acetic acid benzyl ester
(35j)
Compound 41 was obtained by the method used in the
preparation of 9, except that 31 and m-toluidine (35b)
were used instead of 30 and 35a, respectively (88%).
1H NMR (CDCl3) 8.21 (1H, br s), 7.61–7.29 (7H, m),
7.22 (2H, m), 6.87 (1H, m), 5.19 (2H, m), 4.64 (1H, d),
4.46 (1H, d), 4.20 (2H, s), 3.98 (1H, m), 2.26 (3H, s),
2.25–1.70 (12H, m).
Step A. A mixture of N-tert-butoxycarbonyl 3-aminothi-
ophenol36 (2.7 g, 12 mmol), benzyl bromoacetate (3.0 g,
13 mmol) and K2CO3 (2.1 g, 15 mmol) in acetone
(40 mL) was stirred at ambient temperature for 72 h.
Diethyl ether was added and the insoluble material re-
moved by filtration. The filtrate was evaporated and chro-
matography (Et2O/pentane (1:3)) of the residue afforded
benzyl-3-(N-tert-butyloxycarbonylamino)phenylthioac-
5.33. 2-(1-Cycloheptyl-2,4-dioxo-5-(2-oxo-2-pyrrolidin-1-
yl-ethyl)-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-N-
m-tolyl-acetamide (11)
1
etate as a white solid (2.67 g, 60%). H NMR (CDCl3)
7.34–7.18 (8H, m), 7.05 (1H, m), 6.34 (1H, br s), 5.16
(2H, s), 3.68 (2H, s), 1.53 (9H, s).
Step A. (5-Cycloheptyl-2,4-dioxo-3-(m-tolylcarbamoyl-
methyl)-2,3,4,5-tetrahydro-1,3,5-benzotriazepin-1-yl)-ace-
tic acid was obtained by step B of the method used in the
preparation of 33 except that 41 was used instead of (1-
cyclohexyl-5-(3,3-dimethyl-2-oxo-butyl)-2,4-dioxo-1,2,4,5-
tetrahydro-1,3,5-benzotriazepin-3-yl)-acetic acid benzyl
Step B. The product of the previous step (2.67 g,
7.2 mmol) in TFA (25 mL) was stirred at ambient tem-
perature for 1 h. After removal of the volatiles in vacuo,
the resulting oil was dissolved in DCM (60 mL), washed
with a saturated NaHCO3 solution (2· 60 mL) and
dried (MgSO4). Filtration and evaporation of the sol-
vent afforded 35j as an oil (1.76 g, 90%). 1H NMR
(CDCl3) 7.34–7.18 (7H, m), 6.72–6.81 (2H, m), 6.54
(1H, br s), 5.18 (2H, s), 3.62 (2H, s).
1
ester (48%). H NMR (CDCl3) 8.31 (1H, br s), 7.38–
7.29 (3H, m), 7.23–7.14 (4H, m), 6.87 (1H, m), 4.52
(2H, m), 4.20 (2H, m), 4.02 (1H, m), 2.25 (3H, m),
2.21–1.40 (12H, m).
Step B. Compound 11 was obtained by the method used in
the preparation of 9, except that the product of the previ-
ous step and pyrrolidine were used instead of 30 and 35a,
5.30. (3-(2-(1-Cycloheptyl-5-(3,3-dimethyl-2-oxo-butyl)-
2,4-dioxo-1,2,4,5-tetrahydro-1,3,5-benzotriazepin-3-yl)-
acetylamino)-phenylsulfanyl)-acetic acid benzyl ester
(40)
1
respectively (91%). H NMR (CDCl3) 8.24 (1H, br s),
7.41–7.14 (7H, m), 6.86 (1H, m), 4.48 (2H, s), 4.21 (2H,
m), 4.04 (1H, m), 3.49 (4H, m), 2.35 (3H, s), 2.30–1.51
(16H, m). HRMS (ES, [M+H]+) calcd for C30H37N5O4
532.2918, found 532.2921.
Compound 40 was obtained by the method used in the
preparation of 9 except that 32 and 35j were used in-

J. Spencer et al. / Bioorg. Med. Chem. 16 (2008) 2974–2983
2983
17. McDonald, I. M.; Austin, C.; Buck, I. M.; Dunstone, D.
J.; Griffin, E.; Harper, E. A.; Hull, R. A.; Kalindjian, S.
B.; Linney, I. D.; Low, C. M.; Pether, M. J.; Spencer, J.;
Wright, P. T.; Adatia, T.; Bashall, A. J. Med. Chem. 2006,
49, 2253–2261.
18. McDonald, I. M.; Black, J. W.; Buck, I. M.; Dunstone,
D. J.; Griffin, E. P.; Harper, E. A.; Hull, R. A.;
Kalindjian, S. B.; Lilley, E. J.; Linney, I. D.; Pether, M.
J.; Roberts, S. P.; Shaxted, M. E.; Spencer, J.; Steel, K.
I.; Sykes, D. A.; Walker, M. K.; Watt, G. F.; Wright,
L.; Wright, P. T.; Xun, W. J. Med. Chem. 2007, 50,
3101–3112.
19. Willson, T. M.; Henke, B. R.; Momtahen, T. M.; Myers,
P. L.; Sugg, E. E.; Unwalla, R. J.; Croom, D. K.;
Dougherty, R. W.; Grizzle, M. K.; Johnson, M. F.;
Queen, K. L.; Rimele, T. J.; Yingling, J. D.; James, M. K.
J. Med. Chem. 1996, 39, 3030–3034.
Acknowledgment
The EPSRC National Mass Spectrometry Service Cen-
tre, University of Wales Swansea, is gratefully acknowl-
edged for providing HRMS measurements.
Supplementary data
Supplementary data associated with this article can be
found, in the online version, at doi:10.1016/
j.bmc.2007.12.047.
References and notes
20. Hagishita, S.; Seno, K.; Kamata, S.; Haga, N.; Ishihara,
Y.; Ishikawa, M.; Shimamura, M. Bioorg. Med. Chem.
1997, 5, 1433–1446.
21. Finizia, G.; Donati, D.; Oliosi, B.; Tranquillini, M. E.;
Ursini, A. Bioorg. Med. Chem. Lett. 1996, 6,
2957–2962.
22. Clifford, D. P.; Jackson, D.; Edwards, R. V.; Jeffrey, P.
Pestic. Sci. 1976, 7, 453–458.
23. Weigert, F. J. J. Org. Chem. 1973, 38, 1316–1319.
24. Bayer, A. G. German Patent Application. DE2036172,
1972.
1. Noble, F.; Wank, S. A.; Crawley, J. N.; Bradwejn, J.;
Seroogy, K. B.; Hamon, M.; Roques, B. P. Pharmacol.
Rev. 1999, 51, 745–781.
2. Crawley, J. N.; Corwin, R. L. Peptides 1994, 15, 731–755.
3. Degen, L.; Matzinger, D.; Drewe, J.; Beglinger, C.
Peptides 2001, 22, 1265–1269.
4. Dauge, V.; Lena, I. Neurosci. Biobehav. Rev. 1998, 22,
815–825.
5. Wiesenfeld-Hallin, Z.; Xu, X. J. Regul. Pept. 1996, 65, 23–
28.
6. Dockray, G. J. Best Pract. Res. Clin. Endocrinol. Metab.
2004, 18, 555–568.
25. Ramirez, F.; Telefus, C. D.; Prasad, V. A. V. Tetrahedron
1975, 31, 2007–2014.
7. Aly, A.; Shulkes, A.; Baldwin, G. S. Biochim. Biophys.
Acta 2004, 1704, 1–10.
26. James Black Foundation Ltd, UK. PCT International
Patent Application. WO 2003041714, 2003.
8. Haigh, C. R.; Attwood, S. E.; Thompson, D. G.;
Jankowski, J. A.; Kirton, C. M.; Pritchard, D. M.; Varro,
A.; Dimaline, R. Gastroenterology 2003, 124, 615–625.
9. Kalindjian, S. B.; McDonald, I. M. Curr. Top. Med.
Chem. 2007, 7, 1195–1204.
10. McDonald, I. M. Exp. Opin. Ther. Patents 2001, 11, 445–
462.
11. Chambers, M. S.; Fletcher, S. R. Prog. Med. Chem. 2000,
37, 45–81.
12. Evans, B. E.; Rittle, K. E.; Bock, M. G.; DiPardo, R. M.;
Freidinger, R. M.; Whitter, W. L.; Lundell, G. F.; Veber,
D. F.; Anderson, P. S.; Chang, R. S.; Lotti, V. J.; Cerino,
D. J.; Chen, T. B.; Kling, P. J.; Kunkel, K. A.; Springer, J.
P.; Hirshfield, J. R. S. J. Med. Chem. 1988, 31, 2235–2246.
13. Bailey, N.; Box, P. C.; Carr, R. A. E.; Cooke, J. W. B.;
Evans, B.; Finch, H.; Head, J. E.; Pass, M.; Shah, P.;
Wheatcroft, J. R. Bioorg. Med. Chem. Lett. 1997, 7, 281–
286.
27. Roberts, S. P.; Harper, E. A.; Watt, G. F.; Gerskowitch,
V. P.; Hull, R. A.; Shankley, N. P.; Black, J. W. Br. J.
Pharmacol. 1996, 118, 1779–1789.
28. Semple, G.; Ryder, H.; Kendrick, D. A.; Szelke, M.; Ohta,
M.; Satoh, M.; Nishida, A.; Akuzawa, S.; Miyata, K.
Bioorg. Med. Chem. 1996, 6, 51–54.
29. Ursini, A.; Capelli, A. M.; Carr, R. A.; Cassara, P.; Corsi,
M.; Curcuruto, O.; Curotto, G.; Dal Cin, M.; Davalli, S.;
Donati, D.; Feriani, A.; Finch, H.; Finizia, G.; Gaviraghi,
G.; Marien, M.; Pentassuglia, G.; Polinelli, S.; Ratti, E.;
Reggiani, A. M.; Tarzia, G.; Tedesco, G.; Tranquillini, M.
E.; Trist, D. G.; Van Amsterdam, F. T. J. Med. Chem.
2000, 43, 3596–3613.
30. Blum, Z.; Ekstrom, M.; Wistrand, L. G. Acta Chem.
Scand. B Org. Chem. Biochem. 1984, 38, 297–302.
31. Baba, A.; Kawamura, N.; Makino, H.; Ohta, Y.;
Taketomi, S.; Sohda, T. J. Med. Chem. 1996, 39, 5176–
5182.
14. Aquino, C. J.; Armour, D. R.; Berman, J. M.; Birkemo, L.
S.; Carr, R. A.; Croom, D. K.; Dezube, M.; Dougherty, R.
W., Jr.; Ervin, G. N.; Grizzle, M. K.; Head, J. E.; Hirst, G.
C.; James, M. K.; Johnson, M. F.; Miller, L. J.; Queen, K.
L.; Rimele, T. J.; Smith, D. N.; Sugg, E. E. J. Med. Chem.
1996, 39, 562–569.
15. Hirst, G. C.; Aquino, C.; Birkemo, L.; Croom, D. K.;
Dezube, M.; Dougherty, R. W., Jr.; Ervin, G. N.; Grizzle,
M. K.; Henke, B.; James, M. K.; Johnson, M. F.;
Momtahen, T.; Queen, K. L.; Sherrill, R. G.; Szewczyk,
J.; Willson, T. M.; Sugg, E. E. J. Med. Chem. 1996, 39,
5236–5245.
16. Sherrill, R. G.; Berman, J. M.; Birkemo, L.; Croom, D.
K.; Dezube, M.; Ervin, G. N.; Grizzle, M. K.; James, M.
K.; Johnson, M. F.; Queen, K. L.; Rimele, T. J.;
Vanmiddlesworth, F.; Sugg, E. E. Bioorg. Med. Chem.
Lett. 2001, 11, 1145–1148.
32. Zhang, M. Q.; Haemers, A.; Vandenberghe, D.; Pattyn, S.
R.; Bollaert, W.; Levshin, I. J. Heterocycl. Chem. 1991, 28,
673–683.
33. Castro, J. L.; Ball, R. G.; Broughton, H. B.; Russell, M.
G.; Rathbone, D.; Watt, A. P.; Baker, R.; Chapman, K.
L.; Fletcher, A. E.; Patel, S.; Smith, A. J.; Marshall, G. R.;
Ryecroft, W.; Matassa, V. G. J. Med. Chem. 1996, 39,
842–849.
34. Merck Sharp and Dohme Ltd. PCT International Patent
Application. WO 9319063, 1993.
35. Eda, M.; Ashimori, A.; Akahoshi, F.; Yoshimura, T.;
Inoue, Y.; Fukaya, C.; Nakajima, M.; Fukuyama, H.;
Imada, T.; Nakamura, N. Bioorg. Med. Chem. Lett. 1998,
8, 919–924.
36. Bartoli, G.; Bosco, M.; Locatelli, M.; Marcantoni, E.;
Massaccesi, M.; Melchiorre, P.; Sambri, L. Synlett 2004,
1794–1798.