Discovery of (R)-7-cyano-2,3,4,5-tetrahydro-1-(1H-imidazol-4-ylmethyl)-3-(phenylmethyl)-4- (2-thienysulfonyl)-1H-1,4-benzodiazepine (BMS-214662), a farnesyltransferase inhibitor with potent preclinical antitumor activity

Article abstract of DOI:10.1021/jm000248z

Continuing structure-activity studies were performed on the 2,3,4,5-tetrahydro-1-(imidazol-4-ylalkyl)-1,4-benzodiazepine farnesyltransferase (FT) inhibitors. These studies demonstrated that a 3(R)-phenylmethyl group, a hydrophilic 7-cyano group, and a 4-s

Full text of DOI:10.1021/jm000248z

© Copyright 2000 by the American Chemical Society
Volume 43, Number 20
October 5, 2000
Expedited Articles
Discover y of (R)-7-Cya n o-2,3,4,5-tetr a h yd r o-1-(1H-im id a zol-4-ylm eth yl)-3-
(p h en ylm eth yl)-4-(2-th ien ylsu lfon yl)-1H-1,4-ben zod ia zep in e (BMS-214662), a
F a r n esyltr a n sfer a se In h ibitor w ith P oten t P r eclin ica l An titu m or Activity
J ohn T. Hunt,*,§ Charles Z. Ding,§ Roberta Batorsky,† Mark Bednarz,‡ Rajeev Bhide,§ Young Cho,§
Saeho Chong,# Sam Chao,‡ J ohnni Gullo-Brown,† Peng Guo,§ Soong Hoon Kim,§ Francis Y. F. Lee,†
Katerina Leftheris,§ Arthur Miller,§ Toomas Mitt,§ Manorama Patel,§ Becky A. Penhallow,† Carol Ricca,†
William C. Rose,† Robert Schmidt,§ William A. Slusarchyk,§ Gregory Vite,§ and Veeraswamy Manne†
Departments of Oncology Chemistry, Chemistry Core Resources, Metabolism and Pharmacokinetics, and Oncology Drug
Discovery, Bristol-Myers Squibb Pharmaceutical Research Institute, P.O. Box 4000, Princeton, New J ersey 08543-4000
Received J une 8, 2000
Continuing structure-activity studies were performed on the 2,3,4,5-tetrahydro-1-(imidazol-
4-ylalkyl)-1,4-benzodiazepine farnesyltransferase (FT) inhibitors. These studies demonstrated
that a 3(R)-phenylmethyl group, a hydrophilic 7-cyano group, and a 4-sulfonyl group bearing
a variety of substituents provide low-nanomolar FT inhibitors with cellular activity at
concentrations below 100 nM. Maximal in vivo activity in the mutated K-Ras bearing HCT-
116 human colon tumor model was achieved with analogues carrying hydrophobic side chains
such as propyl, phenyl, or thienyl attached to the N-4 sulfonyl group. Several such compounds
achieved curative efficacy when given orally in this model. On the basis of its excellent
preclinical antitumor activity and promising pharmacokinetics, compound 20 (BMS-214662,
(R)-7-cyano-2,3,4,5-tetrahydro-1-(1H-imidazol-4-ylmethyl)-3-(phenylmethyl)-4-(2-thienylsulfo-
nyl)-1H-1,4-benzodiazepine) has been advanced into human clinical trials.
In tr od u ction
the C-terminus, followed by C-terminal tripeptide hy-
drolysis, and finally methyl esterification of the new
C-terminal farnesylcysteine. Following the demonstra-
tion that the key step in this sequence is the farnesy-
lation of Ras by the enzyme protein farnesyltransferase
(FT), the great majority of drug discovery effort has
focused on developing FT inhibitors.2,3 Interestingly, the
efficacy of these inhibitors in preclinical antitumor
models has raised the question of the role of Ras-
processing inhibition in the antitumor effects of FT
inhibitors.4
Following an early period in which FT inhibitor
design was focused primarily on thiol-containing pep-
tidomimetics, a wide variety of non-thiol, non-peptidic
inhibitor chemotypes have been discovered.5-7 Our own
efforts resulted in the identification of a potent class of
inhibitors whose distinguishing feature was the pres-
ence of an imidazol-4-ylalkyl group attached to the
Nearly two decades ago, mutated ras genes were first
found to be present in a variety of human tumors.1 Since
that time, substantial efforts have been invested in
defining therapeutic agents which will reverse the
aberrant cell signaling caused by the ras oncogenes. The
signaling functions of both normal and oncogenic Ras
are dependent upon its membrane association, which
is accomplished by posttranslational processing of the
cytosolic Ras protein. This processing involves a se-
quential series of three enzymatic transformations,
namely farnesylation of a cysteine four residues from
* To whom correspondence should be addressed. Tel: (609) 252-
4989. Fax: (609) 252-6601. E-mail: john.hunt@bms.com.
§ Department of Oncology Chemistry.
‡ Department of Chemistry Core Resources.
#
Department of Metabolism and Pharmacokinetics.
† Department of Oncology Drug Discovery.
10.1021/jm000248z CCC: $19.00 © 2000 American Chemical Society
Published on Web 09/13/2000

3588 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 20
Hunt et al.
Sch em e 1a
and 18) or with 7-bromo-2,3,4,5-tetrahydro-3-(phenyl-
methyl)-1H-1,4-benzodiazepine (for analogue 31). 7-Py-
ridyl substituents were introduced by Pd-mediated
couplings of the appropriate stannanes with the bis-
trifluoroacetyl 7-bromo-2,3,4,5-tetrahydro-3-(phenyl-
methyl)-1H-1,4-benzodiazepine.
Biologica l Testin g
Enzyme inhibition assays and cellular assays mea-
suring the phenotypic reversion of H-ras transformed
Rat-1 cells as well as inhibition of anchorage-indepen-
dent growth of H-ras transformed NIH3T3 cells in soft
agar (soft agar growth or SAG assay) were described
previously.8,13,14 Antitumor testing using the Rat-1
model was described previously.8 Antitumor testing
using the HCT-116 model involved subcutaneous (sc)
implantation of tumor fragments followed by intraperi-
toneal (ip) or oral (po) compound administration initi-
ated when tumors had grown to a predetermined size.
For primary screening purposes, this predetermined size
was approximately 100 mg, but for subsequent studies
involving active compounds, the stringency of the test
conditions increased to where tumors were allowed to
grow to between 100 and 300 mg before treatment was
initiated. With the exception of these larger tumor sizes,
the general approach to antitumor testing and efficacy
assessment has been previously described.15
a
Reagents and conditions: (a) D-phenylalanine methyl ester‚HCl,
pyridine, reflux; (b) BH3, THF, reflux; (c) CuCN, NMP, 200 °C;
(d) 2-thienylsulfonyl chloride, CH2Cl2, DIEA; (e) 4-formylimidazole,
Na(OAc)3BH.
1-position of the 2,3,4,5-tetrahydro-1,4-benzodiazepine
nucleus.8 In this report we describe the continuing
exploration of the structure-activity relationships (SARs)
of this class of potent FT inhibitors. These studies have
identified compounds which have marked activity in a
mutated K-Ras-containing human tumor xenograft
model. Results from these studies have led to the
advancement of compound 20 into human clinical trials.
Compound 20 thus joins three other FT inhibitors
(R115777, Sch-66336, L-778123) which have been dis-
closed in the scientific literature to have undergone
testing in cancer patients. 9-12 The oncology community
is eagerly awaiting clinical trial results which will
indicate whether these molecularly targeted therapies
will provide benefit to cancer patients.
Resu lts a n d Discu ssion
Str u ctu r e-Activity Rela tion sh ip s. We recently
described the discovery of a potent class of FT inhibitors
based upon 2,3,4,5-tetrahydro-1,4-benzodiazepines con-
taining an imidazole group attached to N-1.8 The
reported SAR studies focused primarily on imidazol-4-
ylmethyl derivatives in which substituents were varied
on the fused aryl ring as well as on N-4. These studies
demonstrated that a hydrophobic substituent linked to
N-4 via a hydrogen bond-accepting group, as well as a
7- or 8-hydrophobic substituent, was important for
potent enzyme inhibition.
The synthetic scheme for preparing these imidazolyl-
methyltetrahydrobenzodiazepines relied upon initial
construction of the tetrahydrobenzodiazepine ring sys-
tem, followed by modification of the fused aryl ring
substituents as well as the substituents at N-1 and N-4.
Using this straightforward route, exploration of the
effect of C-2 and C-3 substituents was more laborious
in that it involved initial incorporation during ring
system construction. The SARs of these positions were
initially investigated in analogues which contained an
N-1 imidazol-4-ylmethyl group and an N-4 naphthoyl
group, each of which was a constituent of potent
inhibitors in the series in which C-2 and C-3 were
unsubstituted. Limited exploration of C-2 substituents
did not afford analogues of increased inhibitory potency
(data not shown). At C-3 (Table 1), a methyl substituent
produced an analogue of slightly poorer inhibitory
potency (2), while a phenylmethyl substituent afforded
an analogue of slightly increased potency (3). Interest-
ingly, replacement of the critical naphthyl N-4 hydro-
phobe with a methyl group in the 3-phenylmethyl series
led to an analogue (4) only slightly less potent than the
C-3 unsubstituted analogue with a 4-naphthoyl group.
This suggested that the 3-phenylmethyl group of the
Ch em istr y
In general, the 3-substituted-2,3,4,5-tetrahydro-1,4-
benzodiazepine ring systems were prepared as previ-
ously described, by condensation of the appropriately
substituted isatoic anhydride with an amino acid ester
hydrochloride in pyridine at elevated temperature to
form the benzodiazepinedione, followed by reduction
with lithium aluminum hydride or diborane.8 This is
specifically illustrated in Scheme 1 for compound 20.
For selected analogues (14, 15, 19), reaction of the
amino acid with the isatoic anhydride did not afford
cyclization, which was achieved by carbodiimide-medi-
ated amide bond formation. For selected analogues
whose preparation began with commercially available
N-protected amino acids (12, 13, 16, 17), ring formation
was achieved by transformation to the protected amino
acid aldehyde, reductive amination with 2-amino-5-
bromobenzoic acid, and amine deprotection followed by
carbodiimide-mediated cyclization to form the 2,3,4,5-
tetrahydro-1H-1,4-benzodiazepin-5-one. Borane reduc-
tion then afforded the appropriate 2,3,4,5-tetrahydro-
1H-1,4-benzodiazepine. The 7-CN substituent was
introduced by reaction of CuCN with either 7-bromo-
2,3,4,5-tetrahydro-3-(phenylmethyl)-1H-1,4-benzodiaz-
epine (racemic or homochiral 33) or 7-bromo-2,3,4,5-
tetrahydro-4-(methylsulfonyl)-3-(phenylmethyl)-1H-1,4-
benzodiazepine in NMP at elevated temperature. A
phenyl substituent at the 7-position was introduced by
a Pd-mediated coupling of phenylboronic acid at the
stage of the 6-bromoisatoic anhydride (for analogues 10

Discovery of BMS-214662
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 20 3589
Ta ble 1. Interplay of C-3 and N-4 Substituents
compd
R3
R3 isomer
X
R4
R7
FT IC50 (nM)
% reversion (concn)
SAG EC50 (µM)
1a
2c
3c
4c
5b
6c
7c
8c
9c
H
Me
CO
CO
CO
CO
CO
CO
SO2
SO2
SO2
naphth-1-yl
naphth-1-yl
naphth-1-yl
Me
naphth-1-yl
Me
Me
Me
Me
H
H
H
H
Br
Br
Br
Br
Br
456 ( 128
1100 ( 280
276 ( 102
1300 ( 460
228 ( 7
0 (10 µM)
0 (10 µM)
70 (10 µM)
R,S
R,S
R,S
CH2-phenyl
CH2-phenyl
H
CH2-phenyl
CH2-phenyl
CH2-phenyl
CH2-phenyl
5 (5 µM)
30 (1 µM)
80 (1 µM)
70 (1 µM)
R,S
R,S
R
60 ( 24
0.6
0.27
0.29
30 ( 12
10.7 ( 0.4
437 ( 82
S
a
c
Ref 8, compound 8. bRef 8, compound 31. Elemental analysis for C, H, N, halogen.
ethyl (14), as well as a cyclohexylmethyl (12) substitu-
ent, afforded relatively potent inhibitors.
Having established that other 3-position side chains
offered no distinct advantages over the phenylmethyl
group, SAR investigations of the 4-, 7-, and 8-positions
were conducted with the aim of optimizing both enzyme
and cell potency as well as physicochemical properties
(Table 3). Preferred fused aryl substitutions were gener-
ally in accord with those previously reported in the
3-unsubstituted series.8 For example, the 7-(4-pyridyl)
analogue 28 was potent in both enzyme and cellular
assays. While a pyridine substituent allowed us to
moderate the lipophilicity of our inhibitors, it can carry
its own potential liabilities, such as facile in vivo
oxidation of the pyridine nitrogen and the possibility of
cytochrome P450 enzyme inhibition. In our search for
an alternative hydrophilic substituent, it was discovered
that the 7-cyano analogue 25 was a potent FT inhibitor.
In addition, the 7-cyano substituent provided analogues
with substantially improved aqueous solubility. For
example, 25 (81 ng/mL) had nearly 10-fold higher
solubility in 2% DMSO/1X PBS buffer (pH 7.4) com-
pared to the racemic 7-bromo analogue 6 (9.7 ng/mL).
This combination of potent FT inhibitory potency and
increased solubility translated into extremely high
potency in cellular assays. While it is not clear why the
small, linear, and polar nitrile functionality should be
an exceptional replacement for the relatively lipophilic
7-substituents (e.g., bromine, phenyl) which we had
previously identified, we do note that a 4-cyanophenyl-
methyl substituent in several different series of FT
inhibitors has been independently found to provide
potent inhibitors.16-18 Perhaps the nitrile functionality
in these structurally distinct FT inhibitors is playing a
similar role.
F igu r e 1. Pharmacophore model for the binding of imida-
zolylmethyltetrahydrobenzodiazepines to FT.
4-acetyl analogue might be accessing the putative
hydrophobic binding pocket on FT utilized by the
4-naphthoyl group in the 3-unsubstituted series (Figure
1).
Because the 7-bromobenzodiazepine nucleus had pro-
vided somewhat higher FT inhibitory potency in the
3-unsubstituted series, studies of the interplay of the
3- and 4-substituents were primarily carried out using
this fused phenyl substitution pattern. Utilizing the
racemic 3-phenylmethyl nucleus, an N-4 acetyl group
was found to produce a relatively potent inhibitor (6).
The methanesulfonyl analogue 7 produced an even more
potent FT inhibitor and one with a submicromolar IC50
value for morphological reversion of H-ras transformed
Rat-1 cells. Preparation of the enantiomers of this
compound demonstrated that the R-isomer 8 was a
substantially more potent FT inhibitor than the S-
isomer 9. The potent cellular activity of 8 was indistin-
guishable from that of the racemate 7.
Utilizing the N-4 acetyl and methanesulfonyl side
chains, limited additional studies of the C-3 substituent
were performed (Table 2). In general, these analogues
were prepared as racemic mixtures, both for conven-
ience based on availability of starting materials and to
ensure that the optimal isomer would not be missed if
the C-3 configurational preference was dependent on the
nature of the side chain. Relatively poor inhibitory
potency was observed with the 2-naphthyl (19) and
4-pyridyl (18) side chains, suggesting respectively some
steric limitations to the binding pocket as well as some
limitations to the polarity acceptable in the region of
the 4-substituent. However, a variety of other arylalkyl
side chains, including 3-pyridylmethyl (10) and phenyl-
Within the (R)-3-phenylmethyl-7-cyano series, exten-
sive SAR investigations of the 4-substituent demon-
strated a fair amount of tolerance for FT inhibitory
potency, with carboxamides, ureas (e.g., 29), carbamates
(e.g., 30), sulfonamides, and sulfonylureas (e.g., 24) all
providing relatively potent inhibitors (Table 3). How-
ever, the sulfonamides were routinely superior to the
carboxamides both in terms of FT inhibitory potency as
well as cellular activity (e.g., compare 25 and 27). In
fact, sulfonamide substituents as diverse as methyl,

3590 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 20
Ta ble 2. C-3 Substituents
Hunt et al.
compd
X
R3 isomer
R3
R7
FT IC50 (nM)
% reversion (concn)
SAG EC50 (µM)
10a
11
SO2
SO2
SO2
SO2
SO2
CO
SO2
SO2
SO2
CO
R
R,S
R
R,S
R,S
R,S
R,S
R,S
R
pyrid-3-yl
2-Cl-phenyl
c-Hex
3-Cl-phenyl
CH2-phenyl
naphth-1-yl
NHCO-phenyl
4-MeO-phenyl
pyrid-4-yl
phenyl
Br
Br
Br
Br
Br
Br
Br
phenyl
Br
5.95 ( 1.85
18 ( 7
50 (0.1 µM)
70 (1 µM)
90 (10 µM)
70 (10 µM)
70 (1 µM)
5 (1 µM)
0.015
0.46
12a
13a
14a
15a
16
23.5 ( 5.5
24 ( 13
45 ( 9
54.5 ( 14.5
86 ( 39
101 ( 29
383 ( 150
1260 ( 146
0.4
0 (1 µM)
17a
18a
19a
0 (1 µM)
R,S
naphth-2-yl
a
HRMS.
Ta ble 3. N-4 Substituents
compd
R3 isomer
X
R4
R7
FT IC50 (nM)
% reversion (concn)
SAG EC50 (µM)
20a
21e
22b
23b
24e
25c
26e
27e
28e
29c
30e
31d
R
R
R
R
R
R
R,S
R
R
SO2
SO2
SO2
SO2
SO2
SO2
SO2
CO
2-thienyl
CH2CH2NMe2
phenyl
CH2CH2CH3
NMe2
Me
Me
Me
Me
NMe2
OEt
CN
CN
CN
CN
CN
CN
pyrid-3-yl
CN
pyrid-4-yl
CN
1.35 ( 0.05
1.53 ( 0.97
1.77 ( 0.09
1.77 ( 0.57
2.85 ( 0.05
3.0 ( 1.1
7.9 ( 0.1
8.75 ( 2.25
16 ( 4
60 (0.1 µM)
0.025
0.03
0.01
0.01
0.05
0.013
90 (0.1 µM)
80 (0.1 µM)
50 (0.1 µM)
80 (0.1 µM)
10 (1 µM)
0.10
0.04
SO2
CO
CO
R
R
R,S
17 ( 4
40 (1 µM)
50 (1 µM)
70 (1 µM)
CN
phenyl
22 ( 3
54.5 ( 15.5
0.3
0.53
SO2
Me
a
c
e
Anal. for C, H, N, S. bAnal. for C, H, N, S, Cl. Anal. for C, H, N, Cl. dAnal. for C, H, N, F. HRMS.
aryl, and dimethylaminoethyl all afforded low-nanomo-
lar FT inhibitors which produced morphological rever-
sion with IC50 values of less than 100 nM and soft agar
growth inhibition with EC50 values in the 10-50 nM
range.
All of these FT inhibitors which were tested for
inhibition of the related enzyme GGT1 were found to
be highly selective for FT. For example, 20 was over
1000-fold selective for FT, having IC50 values for inhibi-
tion of geranylgeranylation of Ras-CVLL and K-Ras of
1.3 and 2.3 µM, respectively.
ADME a n d In Vivo An titu m or Testin g. The ra-
cemic lead compound 7 was tested for in vivo antitumor
activity in the previously described Rat-1 tumor model,
in which tumor cells were implanted ip in mice and
compound was administered ip once daily for 7 days.8
Compound 7 (30 mg/kg/injection) produced a lifespan
increase compared to untreated controls, with a %T/C
value of 138, where %T/C g 125 was considered an
active result. Limited evaluation of the oral bioavail-
ability of the active enantiomer of 7, homochiral 8, was
conducted in Balb/c mice. Based on the dose-normalized
AUC values, the oral bioavailability was estimated at
about 40%, with an apparent t1/2 of 1.2 h (data not
shown). These results suggested the possibility that 8
might be efficacious when dosed orally in an in vivo
tumor model.
In addition to the H-ras transformed Rat-1 tumor
model, human tumor xenografts were evaluated for
their suitability as in vivo models of FT inhibitor
efficacy. In particular, HCT-116, a human colon tumor
bearing a K-Ras mutation, was evaluated as a sc tumor.
Compound 8 was efficacious against sc HCT-116 when
dosed ip once daily for 10 days (data not shown) at a
dose of 225 mg/kg/injection to yield a log cell kill (LCK)
of 1.7. Similar efficacy was observed when the compound
was dosed once daily po for 10 days (Table 4). Interest-
ingly, a schedule involving intermittent ip dosing of 8
afforded superior efficacy compared to the daily ip
dosing schedule, and in fact, cures were observed (data

Discovery of BMS-214662
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 20 3591
Ta ble 4. In Vivo Antitumor Testing versus sc HCT-116
Tumors
compd
expt no.
optimal dosea
LCK or cures/total
8
10
20
21
22
23
24
25
28
1
2
3
5
2
3
4
3
2
700
600
600
600
600
600
600
600
600
1.5b
0.1b
8/8c
1.0b
7/8c
7/7c
0.2b
0.6b
0.2b
a
All administrations were once daily po for 10 doses, Monday-
Friday. bLCK. Cures.
c
F igu r e 3. Pharmacokinetics of 20 in rats. Levels in plasma
following iv (0) or po (O) administration of a 50 mg/kg dose in
10% ethanol in water.
and an oral bioavailability of 56%. Based upon its high
FT inhibitory potency, excellent antitumor efficacy, good
pharmacokinetics, good physical properties, and straight-
forward synthesis, 20 was selected for advancement into
human clinical trials.
Con clu sion s
Continuing SAR studies were performed on the
2,3,4,5-tetrahydro-1-(imidazol-4-ylalkyl)-1,4-benzodiaz-
epine FT inhibitors. These studies demonstrated that
a 3(R)-phenylmethyl group, a hydrophilic 7-cyano group,
and a 4-sulfonyl group with a variety of substituents
provide potent FT inhibitors with excellent cellular
activity. Maximal in vivo activity was achieved with
hydrophobic side chains such as propyl, phenyl, or
thienyl attached to the N-4 sulfonyl group. Several such
compounds achieved curative efficacy when given orally
in the HCT-116 human colon tumor model. Compound
20 has been advanced into human clinical trials.
F igu r e 2. In vivo antitumor efficacy of orally administered
FT inhibitors in staged HCT-116 human colon tumor xe-
nografts in nude mice. All compound administration was qdx10
Monday-Friday: b, control (no inhibitor); 0, 8, 400 mg/kg;
4, 20, 300 mg/kg (3.5 LCK); 9, 20, 600 mg/kg (8/8 cures).
not shown; ip once every 4 days for 3 injections at a dose
of 450 mg/kg/injection to yield a LCK of >3.5, 4/8 cures).
In vivo antitumor testing in the HCT-116 model using
daily po dosing was performed with selected analogues
from Tables 2 and 3. Many of these analogues were
inactive or weakly active. However, the 2-thienylsulfo-
nyl (20), phenylsulfonyl (22), and propylsulfonyl (23)
analogues were profoundly active in this model, afford-
ing nearly complete cures. Figure 2 shows the graphical
results of a single HCT-116 antitumor test in which 20
was tested head-to-head with 8. As can be seen, 8 was
inactive at the dose shown. Compound 20 showed
curative efficacy at the top dose of 600 mg/kg/day, and
it still showed substantial efficacy at one-half of the
optimal dose (LCK ) 3.5). Compound 23 was also tested
in this same set of experiments, and its activity was
indistinguishable from 20, producing complete cures at
its optimal dose.
Most of the analogues tested for in vivo activity
demonstrated similar inhibitory potency toward FT and
showed cellular effects at similar concentrations. Nev-
ertheless, there was a wide disparity in their in vivo
efficacy in the HCT-116 tumor model. In general, this
could not easily be attributed to differences in the
pharmacokinetic properties of the compounds (data not
shown). The underlying reason for the dramatic differ-
ences in in vivo efficacy is the subject of current
investigation.
Exp er im en ta l Section
P r en yltr a n sfer a se Assa ys. Assays for FT and GGTI
inhibition were performed as previously described except that
>90% purified recombinant human FT (hFT) or purified
recombinant human GGTI (hGGTI) enzymes were used.13 The
amount of enzyme used in the assays was 0.3 nM hFT or 5
nM hGGTI. The assays also contained 10 µM (FT) or 5 µM
(GGTI) ZnCl2 in addition to the components described earlier.
Cell-Ba sed Assa ys. The assays for inhibition of phenotypic
reversion of H-ras transformed Rat-1 cells and anchorage-
independent growth of H-ras transformed NIH3T3 cells in soft
agar were performed as previously described.14 Reversion data
are presented as the concentration of drug required to produce
a given percentage of reversion to the untransformed pheno-
type. Soft agar growth (SAG) data are presented as the
concentration of drug required to inhibit 50% of colony growth.
P h ar m acokin etic Testin g. Limited pharmacokinetic evalu-
ation of 8 was performed in Balb/c mice. Mice were given a
single bolus dose of 8 either intravenously (45 mg/kg) via a
tail vein or orally (90 mg/kg) by gavage in 10% ethanol
solution. Blood samples were collected over 12 h, and the
concentration of 8 was quantitated by an LC/UV assay with a
quantifiable limit of 1 µM. Three mice were sacrificed at each
time point. Limited pharmacokinetic evaluation of 20 was
performed in rats. Male Sprague-Dawley rats (Harlan Spra-
gue-Dawley, Indianapolis, IN) were surgically prepared with
an indwelling jugular vein cannula 1 day prior to drug
administration. Rats were fasted overnight prior to dosing and
fed 8 h after dosing. They were allowed access to water ad
libitum and were conscious and unrestrained during the study.
The dosing vehicle was 10% ethanol in water. Rats were given
single 50 mg/kg bolus doses of 20 either iv (n ) 2) via jugular
The pharmacokinetics of 20 were investigated in rats
at an equal iv and po dose (Figure 3). With iv admin-
istration, the compound was found to have relatively
slow clearance, a moderate volume of distribution, and
a moderate half-life. Upon po administration, the com-
pound was found to give a high Cmax value of 20 µM

3592 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 20
Hunt et al.
vein cannula or orally (n ) 3) by gavage in 10% ethanol
solution. During the 24 h study, aliquots of 500-µL blood
samples were collected at various time points and plasma drug
levels were determined using an LC/MS/MS assay with a
quantifiable limit of 4 nM. Oral bioavailability was estimated
based on the dose-normalized ratio of AUC (area under the
drug concentration vs time curve).
ylm eth yl)-3-(p h en ylm eth yl)-1H-1,4-ben zod ia zep in e, Tr i-
flu or oa ceta te (6). Anal. C, H, N, F. MS (M + H)+ 440; mp
112 °C.
7-Br om o-2,3,4,5-tetr ah ydr o-1-(1H-im idazol-4-ylm eth yl)-
4-(m et h ylsu lfon yl)-3-(p h en ylm et h yl)-1H-1,4-b en zod ia z-
ep in e, Tr iflu or oa ceta te (7). Anal. C, H, N, Br, F. MS (M +
H)+ 476; mp 118-120 °C.
In Vivo An titu m or Testin g. Rat-1 tumor testing was
performed as previously described.8 Therapeutic results are
presented in terms of increases in lifespan reflected by the
relative median survival time (MST) of treated (T) versus
control (C) groups and expressed as a maximum %T/C. With
regard to the HCT-116 tumor model, the basic approach has
been described previously.15 Briefly, mice were implanted sc
with tumor fragments and sorted into treatment and control
groups when their tumors reached a predetermined size (e.g.,
100-300 mg). Compounds were dissolved in 10% ethanol/
sterile water and the solutions were administered ip or po
within 1 h of dissolution. Group sizes were 8 mice. Assessment
of antitumor effectiveness was made by determining the
relative median times for control (C) and treated (T) mice to
grow tumors to a target size. The target size was 500 mg. The
delay in tumor growth (T - C, days) was converted to gross
log cell kill (LCK) values by taking into account the tumor
volume doubling times (TVDT) of control groups in each
individual experiment. A LCK value of 1 or better was
considered an active result. Cures were also used to assess
activity. A mouse was considered cured when no mass larger
than 35 mg was present at the site of tumor implant after a
number of days posttreatment had elapsed equivalent to >10
TVDTs in that experiment. Statistical evaluations of data were
performed using the Gehan’s generalized Wilcoxon test.19
Drug-treated mice dying before the first death in parallel
control mice implanted with the same tumor inoculum, were
considered to have died from drug toxicity. Groups of mice with
more than one death due to drug toxicity were not used in the
evaluation of antitumor efficacy, and the highest dose tested
that did not cause such lethality was termed the maximum
tolerated dose (MTD).
Gen er a l Ch em ica l P r oced u r es. Melting points were
recorded on a Thomas-Hoover capillary apparatus and are
reported uncorrected. IR spectra were recorded on a Mattson
Sirius 100 spectrometer. Proton NMR (1H NMR) and carbon
NMR (13C NMR) spectra were obtained on J EOL FX-270 or
GX-400 spectrometers and are reported relative to tetrameth-
ylsilane (TMS) reference. Analytical and preparative HPLC
were performed on YMC columns (A-302, S-5, 120A ODS, 4.6
× 150 mm; SH-345-15, S-15, 120A ODS, 20 × 500 mm) with
acetonitrile:water gradients containing 0.1% trifluoroacetic
acid. Chromatography was performed under flash conditions
using EM Science silica 0.040-0.063 mm particle size. THF
was distilled from Na/benzophenone. Solutions were dried with
magnesium sulfate unless otherwise noted.
(R)-7-Br om o-2,3,4,5-t et r a h yd r o-1-(1H -im id a zol-4-yl-
m e t h yl)-4-(m e t h ylsu lfon yl)-3-(p h e n ylm e t h yl)-1H -1,4-
ben zod ia zep in e, Hyd r och lor id e (8). Anal. C, H, N, S, Br,
Cl. [R]D ) +58° (c ) 0.4, MeOH); MS (M + H)+ 476; mp 180-
185 °C.
(R)-2,3,4,5-Tet r a h yd r o-1-(1H -im id a zol-4-ylm et h yl)-4-
(m eth ylsu lfon yl)-7-p h en yl-3-(3-p yr id in ylm eth yl)-1H-1,4-
ben zod ia zep in e, Dih yd r och lor id e (10). Anal. C, H, N, S,
F. MS (M + H)+ 474.
(R)-2,3,4,5-Tet r a h yd r o-1-(1H -im id a zol-4-ylm et h yl)-4-
(m eth ylsu lfon yl)-7-p h en yl-3-(4-p yr id in ylm eth yl)-1H-1,4-
ben zod ia zep in e, Dih yd r och lor id e (18). 1H NMR (CD3OD,
400 MHz) δ 8.83 (1H, s), 8.64 (2H, m), 8.38 (1H, m), 7.95 (1H,
m), 7.60-7.15 (8H, m), 6.90 (1H, m), 5.00-4.54 (4H, m), 4.28
(1H, m), 3.68 (2H, m), 3.42 (1H, m), 2.86 (1H, m), 2.36 (3H, s);
MS (M + H)+ 474.
2,3,4,5-T e t r a h y d r o -1-(1H -im id a zo l-4-y lm e t h y l)-4-
(m eth ylsu lfon yl)-7-ph en yl-3-(ph en ylm eth yl)-1H-1,4-ben zo-
d ia zep in e (31). The 7-phenyl group was introduced by a Pd-
catalyzed coupling of phenylboronic acid with 7-bromo-2,3,4,5-
tetrahydro-3-(phenylmethyl)-1H-1,4-benzodiazepine. 1H NMR
(CD3OD, 400 MHz) δ 8.8 (d, 1H), 7.7-7.2 (m, 13H), 6.9 (d, 1H),
4.9-4.2 (m, 6H), 3.6-3.0 (m, 4H), 2.25 (s, 3H); MS (M + H)+
472.
(S )-7-Br om o-2,3,4,5-t e t r a h yd r o-1-(1H -im id a zol-4-yl-
m e t h yl)-4-(m e t h ylsu lfon yl)-3-(p h e n ylm e t h yl)-1H -1,4-
ben zod ia zep in e, Hyd r och lor id e (9). A solution of 7 (100
mg, 0.31 mmol) in 2-propanol (5 mL) was chromatographed
by preparative HPLC using a chiralpak AD column produced
by Chiral Technologies Inc. (50 mm × 500 mm; mobile phase
25:75:0.1 2-propanol:hexane:triethylamine; flow rate 55 mL/
min) to provide isomer A at 36 min and isomer B at 54 min
retention times. Isomer B exhibited the same retention time
and optical rotation (+58°) as 8 (see above). Isomer A ([R]D
)
-67° (c ) 0.1, MeOH)) was assigned the S-configuration (18
mg, 13%). Anal. C, H, N, S, Br, C. MS (M + H)+ 476; mp 180-
185 °C.
4-Acetyl-7-br om o-2,3,4,5-tetr a h yd r o-1-(1H-im id a zol-4-
ylm et h yl)-3-(2-n a p h t h a len ylm et h yl)-1H -1,4-b en zod ia z-
ep in e, Dih yd r och lor id e. (19). A solution of 2-naphthylala-
nine (1.0 g, 4.18 mmol) and bromoisatoic anhydride (1.0 g, 4.15
mmol) in pyridine (50 mL) was refluxed for 4 h. The mixture
was cooled, concentrated and the residue was partitioned
between water (200 mL) and ethyl acetate (200 mL). The
organic layer was washed with water (3 × 100 mL), brine (50
mL), dried and concentrated to yield N-(2-amino-5-bromoben-
zoyl)naphthylalanine as a yellowish glass (450 mg, 38%). A
solution of this material (450 mg, 1.09 mmol), 1-ethyl-3-(3-
dimethylaminopropyl)carbodiimide‚HCl (EDC; 239 mg, 1.2
mmol), N,N-diisopropylethylamine (DIEA; 0.42 mL, 2.4 mmol),
and 1-hydroxybenzotriazole hydrate (162 mg, 1.2 mmol) in
DMF (10 mL) was stirred for 16 h. The mixture was poured
into water (100 mL) and the product was extracted with ethyl
acetate (2 × 50 mL). The combined ethyl acetate layers were
washed with water (3 × 100 mL) and brine (100 mL), dried,
and concentrated to yield 3-(2-naphthalenylmethyl)-7-bromo-
1,4-benzodiazepine-2,5-dione as a brown glass (100 mg, 23%).
MS (M + H)+ 396. This material was carried on to 19 using
previously described procedures.8 1H NMR (CD3OD, 400 MHz)
δ 8.81 (1H, s), 7.84 (4H, m), 7.70 (1H, m), 7.50-7.25 (5H, m),
6.87 (1H, m), 4.73-4.54 (3H, m), 4.43 (1H, m), 3.73 (1H, m),
3.23 (1H, m), 3.05 (1H, m), 2.93 (1H, m), 2.13 (1H, m), 2.05
(3H, s).
The following compounds were prepared using previously
described procedures, using the noted starting materials: 2,
isatoic anhydride and D,L-alanine ethyl ester hydrochloride; 3
and 4, isatoic anhydride and D,L-phenylalanine methyl ester
hydrochloride; 6, 7 and 31, 6-bromoisatoic anhydride and D,L-
phenylalanine methyl ester hydrochloride; 8, 6-bromoisatoic
anhydride and D-phenylalanine methyl ester hydrochloride;
10, 6-phenylisatoic anhydride and D-3-pyridylalanine; 18,
6-phenylisatoic anhydride and D-4-pyridylalanine.8
2,3,4,5-Tetr a h yd r o-1-(1H-im id a zol-4-ylm eth yl)-3-m eth -
yl-4-(1-n aph th alen ylcar bon yl)-1H-1,4-ben zodiazepin e, Hy-
d r och lor id e (2). Anal. C, H, N, Cl. MS (M + H)+ 397; mp
180-185 °C.
2,3,4,5-Te t r a h yd r o-1-(1H -im id a zol-4-ylm e t h yl)-4-(1-
n a p h th a len ylca r bon yl)-3-(p h en ylm eth yl)-1H-1,4-ben zo-
d ia zep in e, Hyd r och lor id e (3). Anal. C, H, N, Cl. MS (M +
H)+ 473; mp 78-80 °C.
2,3,4,5-Tetr ah ydr o-1-(1H-im idazol-4-ylm eth yl)-4-acetyl-
3-(p h en ylm eth yl)-1H-1,4-ben zod ia zep in e, Hyd r och lor id e
(4). Anal. C, H, N, Cl. MS (M + H)+ 428; mp 155-160 °C.
4-Acetyl-7-br om o-2,3,4,5-tetr a h yd r o-1-(1H-im id a zol-4-
4-Acetyl-7-br om o-2,3,4,5-tetr a h yd r o-1-(1H-im id a zol-4-
ylm et h yl)-3-(1-n a p h t h a len ylm et h yl)-1H -1,4-b en zod ia z-
ep in e, Dih yd r och lor id e (15). Prepared as a white solid from
1
1-naphthylalanine as described for 19. H NMR (CD3OD, 400

Discovery of BMS-214662
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 20 3593
MHz) δ 8.53 (1H, s), 7.87 (1H, m), 7.74 (1H, m), 7.55-7.23
(8H, m), 6.74 (1H, m), 4.57-4.43 (2H, m), 4.15 (1H, m), 3.90
(1H, m), 3.83 (1H, m), 3.48 (2H, m), 3.12 (1H, m), 3.00 (1H,
m), 2.06 (2H, m), 2.01 (3H, s); MS (M + H)+ 475.
7-Br om o-2,3,4,5-tetr ah ydr o-1-(1H-im idazol-4-ylm eth yl)-
4-(m et h ylsu lfon yl)-3-(2-p h en ylet h yl)-1H -1,4-b en zod ia z-
ep in e, Dih yd r och lor id e. (14). Prepared as a white solid from
D,L-homophenylalanine as described for 19. 1H NMR (CD3OD,
400 MHz) δ 8.90 (1H, s), 7.59 (1H, s), 7.46-7.26 (7H, m), 6.99
(1H, m), 4.82-4.64 (4H, m), 4.12 (1H, m), 3.55 (1H, m), 3.31
(1H, m), 2.95-2.62 (2H, m), 2.75 (3H, s), 2.09 (1H, m), 1.63
(1H, m); MS (M + H)+ 490.
7-Br om o-3-[(3-ch lor oph en yl)m eth yl]-2,3,4,5-tetr ah ydr o-
1-(1H -im id a zol-4-ylm et h yl)-4-(m et h ylsu lfon yl)-1H -1,4-
ben zod ia zep in e, Dih yd r och lor id e (13). A solution of D,L-
Boc-3-chlorophenylalaninal (800 mg, 2.8 mmol; prepared from
D,L-Boc-3-chlorophenylalanine by reduction of the Weinreb
amide with LAH), 2-amino-5-bromobenzoic acid (660 mg, 3.06
mmol), molecular sieves (3 Å, 7.0 g) and glacial acetic acid (0.2
mL) in MeOH (10 mL) was stirred for 30 min. Sodium
cyanoborohydride (200 mg, 2.99 mmol) was added portionwise
over 30 min. The mixture was stirred for 16 h, cooled to 0 °C
and saturated NaHCO3 (30 mL) was slowly added. The
mixture was stirred for 30 min, concentrated and the residue
was extracted into ethyl acetate (100 mL). The ethyl acetate
layer was washed with water (100 mL), brine (100 mL), dried
and concentrated. Preparative HPLC afforded 2-(3-(3-chlo-
rophenyl)-2-((1,1-dimethylethoxy)carbonylamino)propylamino)-
5-bromobenzoic acid as a clear oil (100 mg, 7%). A solution of
this material (100 mg, 0.21 mmol) in dimethyl sulfide (0.1 mL)
and 4 N HCl in dioxane (10 mL) was stirred for 40 min. The
mixture was concentrated, redissolved in methylene chloride
(20 mL) and concentrated. This latter procedure was repeated
three times to yield 2-(3-(3-chlorophenyl)-2-aminopropylamino)-
5-bromobenzoic acid as a clear glass. This material was carried
on to 13 as described for 19. 1H NMR (CD3OD, 400 MHz) δ
8.53 (1H, s), 7.87 (1H, m), 7.74 (1H, m), 7.55-7.23 (8H, m),
6.74 (1H, m), 4.57-4.43 (2H, m), 4.15 (1H, m), 3.90 (1H, m),
3.83 (1H, m), 3.48 (2H, m), 3.12 (1H, m), 3.00 (1H, m), 2.06
(2H, m), 2.01 (3H, s); MS (M + H)+ 510.
7-bromo-3-[(1,1-dimethylethoxy)methyl]-4-(methylsulfonyl)-
1,2,3,4-tetrahydro-5H-1,4-benzodiazepine. A solution of this
material (1.1 g, 2.8 mmol) in TFA (8 mL) and methylene
chloride (8 mL) was stirred for 3 h and concentrated. Tritu-
ration with ethyl ether afforded a precipitate which was dried
under vacuum to afford 7-bromo-3-(hydroxymethyl)-4-(meth-
ylsulfonyl)-1,2,3,4-tetrahydro-5H-1,4-benzodiazepine (700 mg,
74%) as a white solid. To a solution of this material (50 mg,
0.15 mmol) in methylene chloride (10 mL) was added 2,6-di-
tert-butyl-4-methylpyridine (62 mg, 0.30 mmol). The solution
was cooled to -40 °C under nitrogen. Triflic anhydride (0.85
mL, 0.30 mmol) was added and the solution was stirred for 1
h at -40 °C. Ammonia gas was bubbled through the cold
solution for 10 min and bubbling was continued as the solution
was slowly warmed to room temperature. Ethyl ether (30 mL)
and saturated aqueous sodium bicarbonate solution (30 mL)
were added and the layers were separated. The organic layer
was washed with 1 N aqueous hydrochloric acid. The aqueous
layer was made basic with 5 N aqueous NaOH and the product
was back-extracted using methylene chloride (30 mL). The
organic layer was dried (Na2SO4) and concentrated to 5 mL.
Benzoic acid (26 mg, 0.21 mmol) and EDC (40 mg, 0.21 mmol)
were added and the solution was stirred for 16 h and
concentrated. The residue was chromatographed (silica, 1:5-
1:1; ethyl acetate:hexanes) to afford N-[[7-bromo-2,3,4,5-tet-
rahydro-4-(methylsulfonyl)-1H-1,4-benzodiazepin-3-yl]methyl]-
benzamide (15 mg) as a white solid. Reductive amination was
1
performed routinely to provide 16 as a white solid. H NMR
(CD3OD, 400 MHz) δ 8.80 (1H, s), 7.72 (2H, m), 7.41 (8H, m),
6.81 (1H, m), 4.83 (4H, m), 3.51 (2H, m), 3.21 (2H, m), 2.54
(3H, s); MS (M + H)+ 518.
(R )-7-Cya n o-2,3,4,5-t e t r a h yd r o-1-(1H -im id a zol-4-yl-
m et h yl)-3-(p h en ylm et h yl)-4-(2-t h ien ylsu lfon yl)-1H -1,4-
ben zod ia zep in e, Mon oh yd r och lor id e (20). A stirred solu-
tion of bromoisatoic anhydride (150 g, 0.62 mol), D-phenyla-
lanine methyl ester hydrochloride (127.3 g, 0.59 mol) and
4-(dimethylamino)pyridine (2 g) in pyridine (1500 mL) was
heated at reflux under argon for 3 days. The pyridine was
removed under vacuum and the residue was dissolved in
methylene chloride (3 L). The solution was washed with 10%
HCl solution and brine. The organic solution was dried and
concentrated under vacuum. The solid was collected and dried
to give 152 g (71%) of (R)-7-bromo-2,3,4,5-tetrahydro-3-(phen-
ylmethyl)-1H-1,4-benzodiazepine-2,5-dione (32), mp 220-223
°C; 1H NMR (CDCl3, 500 MHz) δ 2.81-2.89 (m, 1H), 3.09-
3.16 (m, 1H), 3.37 (s, 1H), 3.92-3.99 (m, 1H), 7.03-7.32 (m,
7-Br om o-3-[(2-ch lor oph en yl)m eth yl]-2,3,4,5-tetr ah ydr o-
1-(1H -im id a zol-4-ylm et h yl)-4-(m et h ylsu lfon yl)-1H -1,4-
ben zod ia zep in e, Dih yd r och lor id e (11). Prepared as a
white solid from D,L-Boc-2-chlorophenylalanine as described
1
for 13. H NMR (CD3OD, 400 MHz) δ 8.94 (1H, s), 7.59 (1H,
s), 7.50-7.22 (6H, m), 6.89 (1H, s), 4.72-4.40 (4H, m), 3.65
(1H, m), 3.42-3.25 (2H, m), 3.18 (1H, m), 2.79 (1H, m), 2.33
(3H, s); MS (M + H)+ 510.
5H), 7.64-7.74 (m, 2H), 8.66-8.67 (m, 1H), 10.53 (s, 1H); 13
C
NMR (DMSO-d6) 33.31, 53.69, 115.94, 123.31, 126.43, 128.22,
129.39, 132.63, 134.97, 136.19, 137.76, 166.45, 171.11 ppm;
[R]D ) -182.9° (c ) 1, MeOH). A stirred solution of this
material (30 g, 87 mmol) in anhydrous THF (870 mL) under
argon was treated with a solution of borane-tetrahydrofuran
complex (440 mL of a 1 M solution, 440 mmol) at room
temperature. The solution was slowly heated to reflux and
heated at reflux for 18 h. The mixture was cooled to 0 °C, and
methanol (150 mL) was added to destroy excess borane. The
resultant solution was concentrated under vacuum, the residue
was dissolved in methanol (250 mL), and 7 N HCl solution
(50 mL) was added. This mixture was heated on a steam bath
for 2 h. The solid thus formed was collected, resuspended in
water (400 mL) and the aqueous suspension was made basic
to pH 11 with 5 N NaOH solution and extracted with ethyl
acetate (2 × 300 mL). The organic extracts were combined,
dried, concentrated under vacuum and the residue was
crystallized from methanol and water (9:1) to give 25 g of (R)-
7-bromo-2,3,4,5-tetrahydro-3-(phenylmethyl)-1H-1,4-benzodi-
azepine (33) as a white solid (91%), mp 130-132 °C; 1H NMR
(CDCl3, 500 MHz) δ 1.53 (bs, 1H), 2.62-2.74 (m, 3H), 3.09-
3.15 (m, 1H), 3.27-3.32 (m, 1H), 3.74-3.85 (m, 3H), 6.57(d,
1H, J ) 8.30), 7.12-7.33 (m, 7H); 13C NMR (CDCl3) 40.55,
52.19, 54.22, 61.74, 112.52, 120.25, 126.50, 128.60, 129.30,
130.13, 132.09, 133.96, 138.52, 148.13 ppm; [R]D ) +70.6° (c
) 0.5, MeOH). To a stirred suspension of this material (60 g,
190 mmol) in anhydrous 1-methyl-2-pyrrolidinone (600 mL)
7-Br om o-2,3,4,5-tetr ah ydr o-1-(1H-im idazol-4-ylm eth yl)-
3-[(4-m eth oxyp h en yl)m eth yl]-4-(m eth ylsu lfon yl)-1H-1,4-
ben zod ia zep in e, Dih yd r och lor id e (17). Prepared as a
white solid from D,L-Boc-4-methoxyphenylalanine as described
1
for 13. H NMR (CD3OD, 400 MHz) δ 8.87 (1H, s), 7.45 (1H,
s), 7.25 (2H, m), 6.80 (1H, m), 4.57-4.31 (4H, m), 3.96 (1H,
m), 3.27 (1H, m), 3.08 (1H, m), 2.56 (3H, s), 1.72 (1H, m), 1.65-
1.43 (6H, m), 1.24-1.00 (3H, m), 0.98-0.63 (3H, m); MS (M +
H)+ 506.
(R)-7-Br om o-3-(cycloh exylm eth yl)-2,3,4,5-tetr a h yd r o-
1-(1H -im id a zol-4-ylm et h yl)-4-(m et h ylsu lfon yl)-1H -1,4-
ben zod ia zep in e, Dih yd r och lor id e (12). Prepared as a
white solid from D-Boc-cyclohexylalanine as described for 13.
1H NMR (CD3OD, 400 MHz) δ 8.53 (1H, s), 7.87 (1H, m), 7.74
(1H, m), 7.55-7.23 (8H, m), 6.74 (1H, m), 4.57-4.43 (2H, m),
4.15 (1H, m), 3.90 (1H, m), 3.83 (1H, m), 3.48 (2H, m), 3.12
(1H, m), 3.00 (1H, m), 2.06 (2H, m), 2.01 (3H, s); MS (M + H)+
482.
N-[[7-Br om o-2,3,4,5-t et r a h yd r o-1-(1H -im id a zol-4-yl-
m et h yl)-4-(m et h ylsu lfon yl)-1H-1,4-b en zod ia zep in -3-yl]-
m eth yl]ben za m id e, Dih yd r och lor id e (16). 7-Bromo-3-
[(1,1-dim et h ylet h oxy)m et h yl]-1,2,3,4-t et r a h ydr o-5H -1,4-
benzodiazepine was prepared from Fmoc-O-tert-butylserine as
described for 13, except that Fmoc deprotection was ac-
complished using diethylamine in THF. The methanesulfonyl
group was introduced using standard procedures to afford

3594 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 20
Hunt et al.
under nitrogen was added copper(I) cyanide (51 g, 569 mmol).
The mixture was heated to 200 °C for 3.5 h. The mixture was
slowly added to 15% ethylenediamine solution in water (1.5
L) with vigorous stirring. After 1 h stirring, the slurry was
extracted with EtOAc (3 × 750 mL). The EtOAc extracts were
combined, washed with 10% NH4OH (2 × 750 mL) and brine,
dried over anhydrous Na2SO4 and evaporated to give a black
gum. This was passed through a pad of silica gel (E. Merck
230-400 mesh, 1.2 kg) eluting with EtOAc to give 40 g (80%)
of (R)-2,3,4,5-tetrahydro-3-(phenylmethyl)-1H-1,4-benzodiaz-
chloride. 1H NMR (CD3OD) δ 2.95 (m, 2H), 3.17 (dd, 1H), 3.81
(m, 2H), 4.28 (d, 1H), 4.50 (m, 1H), 4.66 (s, 2H), 6.25 (d, 1H),
7.34 (m, 15H), 8.81 (s, 1H). Anal. C, H, N, S, Cl. MS (M + H)+
484.
(R )-7-Cya n o-1,2,3,5-t e t r a h yd r o-1-(1H -im id a zol-4-yl-
m et h yl)-N,N-d im et h yl-3-(p h en ylm et h yl)-4H -1,4-b en zo-
d ia zep in e-4-su lfon a m id e, Hyd r och lor id e (24). From di-
1
methylsulfamoyl chloride. H NMR (400 MHz, CD3OD): δ 8.9
(s, 1H), 7.5-7.2 (m, 8H), 6.9 (d, 1H, J ) 8 Hz), 4.8-4.4 (m,
4H), 4.25 (m, 1H), 3.75 (m, 1H), 3.38-3.25 (m, 1H), 3.0 (m,
1H), 2.78 (m, 1H), 2.5 (s, 6H); MS (M + H)+ 451.
1
epine-7-carbonitrile (34) as a tan solid, mp 116 °C; H NMR
(CDCl3, 500 MHz) δ 1.72 (bs, 1H), 2.72-2.75 (m, 2H), 2.84-
(R )-7-Cya n o-2,3,4,5-t e t r a h yd r o-1-(1H -im id a zol-4-yl-
m eth yl)-3-(ph en ylm eth yl)-4-m eth ylsu lfon yl-1H-1,4-ben zo-
d ia zep in e, Mon oh yd r och lor id e (25). In this preparation,
the nitrile group was introduced as described above, but
following introduction of the methanesulfonyl group. Anal. C,
H, N, Cl. [R]D ) +218° (c ) 0.23, MeOH); MS (M + H)+ 422;
mp 165 °C.
2.89 (m, 1H), 3.16-3.22 (m, 1H), 3.41-3.46 (m, 1H), 3.88 (qAB
,
2H, J ) 15.07), 4.28 (d, 1H, J ) 4.29), 6.69 (d, 1H, J ) 6.07),
7.32-7.34 (m, 7H); 13C NMR (CD3OD) 40,84, 49.23, 51.62,
51.71, 62.56, 101.42, 119.14, 120.99, 127.56, 129.66, 130.38,
132.71, 134.86, 139.82, 156.29 ppm; [R]D ) -45.6° (HCl salt,
c ) 0.5, MeOH). 2-Thiophenesulfonyl chloride (5 g, 27 mmol)
in CH2Cl2 (5 mL) was added to a solution of this material (6
g, 22.8 mmol) and DIEA (10 mL, 57 mmol) in CH2Cl2 (150 mL)
at 0 °C. The solution was stirred at 0 °C for 10 min and at
room temperature for 10 h. More 2-thiophenesulfonyl chloride
(1 g) was added and stirring was continued for 48 h. The
solution was diluted with CH2Cl2 and washed with 2.5% NaOH
and NaCl. Drying over Na2SO4 and evaporation gave a yellow
solid which was purified by flash chromatography (20% then
40% EtOAc/hexanes) to provide (R)-7-cyano-2,3,4,5-tetrahydro-
3-(phenylmethyl)-4-(2-thienylsulfonyl)-1H-1,4-benzodiaze-
2,3,4,5-T e t r a h y d r o -1-(1H -im id a zo l-4-y lm e t h y l)-4-
(m et h ylsu lfon yl)-3-(p h en ylm et h yl)-7-(3-p yr id in yl)-1H -
1,4-ben zod ia zep in e, Tr ih yd r och lor id e (26). To a solution
of 7-bromo-2,3,4,5-tetrahydro-3-(phenylmethyl)-1H-1,4-benzo-
diazepine (1.61 mmol, 510 mg) and triethylamine (9.66 mmol,
1.35 mL) in CH2Cl2 (10 mL) was added trifluoroacetic anhy-
dride (7.25 mmol, 1.0 mL). The mixture was stirred 30 min,
concentrated and chromatographed on silica to afford the bis-
trifluoroacetate as a white solid (770 mg, 94%). Conversion to
the 7-(pyrid-3-yl) analogue was accomplished by refluxing the
bis-trifluoroacetate with 2 equiv of 3-stannylpyridine and 15
mol % Pd(PPh3)4 in degassed toluene at reflux for 16 h.
Following flash chromatography (10% EtOAc/hexanes) brief
treatment with methanolic KOH afforded 2,3,4,5-tetrahydro-
3-(phenylmethyl)-7-(3-pyridinyl)-1H-1,4-benzodiazepine. This
crude material was carried on to 26 as a yellow solid by the
procedures described for 20. 1H NMR (300 MHz, CD3OD) δ
9.1 (1H, S), 8.91 (1H, s), 8.86 (1H, d, J ) 8.3 Hz), 8.73 (1H, d,
J ) 5.5 Hz), 8.09 (1H, dd, J ) 5.8, 8.1 Hz), 7.67 (2H, br s),
7.53 (1H, S), 7.40-7.18 (m, 5H), 7.04 (1H, d, J ) 8.9), 4.95-
4.4 (1H, m), 3.73 (1H, dd, J ) 5.9, 9.3 Hz), 3.37 (1H, dd, J )
7, 4 Hz), 3.0 (1H, t, J ) 2.5 Hz), 2.75 (1H, dd, J ) 7.5, 4 Hz),
2.57 (1H, s), 2.25 (3H, s); MS (M + H)+ 474.
1
pine (35) as a yellow solid (7.8 g). H NMR (CDCl3, 400 MHz)
δ 2.87-2.95 (m, 1H), 3.06-3.14 (m, 2H), 3.52-3.61 (m, 1H),
4.01-4.08 (m, 1H), 4.34-4.47 (m, 1H), 4.60 (q, 2H, J ) 22.7),
6.14 (d, 1H, J ) 20.0), 6.77 (t, 1H, J ) 10.0), 7.05-7.37 (m,
9H). A solution of this material (7.8 g) and 4-formylimidazole
(3.7 g, 38.5 mmol) in 1:5 AcOH/CH2ClCH2Cl (140 mL) was
stirred for 1 h at 60 °C. NaBH(OAc)3 (7.4 g, 35 mmol) was
added and the stirring was continued for 10 h. Every 8 h, more
4-formylimidazole (1.5 g) and NaBH(OAc)3 (3.0 g) was added
until HPLC analysis showed complete reaction. The solvent
was evaporated. The residue was dissolved in CH2Cl2 (200 mL)
and the solution was stirred with 1 N NaOH (50 mL) for 20
min. The organic layer was separated and washed with 1 N
NaOH (50 mL) twice. The organic phase was dried over Na2-
SO4 and evaporated to give an oil (9 g) which was purified by
preparative HPLC (gradient of aqueous methanol with 0.1%
TFA) and converted to its HCl salt by lyophilizing from 1 N
HCl to give 20 as an off-white solid (4.5 g, 35%). Spectral data
for free base: 13C NMR (100 MHz, CDCl3) 40.0, 47.8, 49.7, 53.3,
59.4, 100.3, 115.0, 115.1, 120.1, 122.3, 127.4, 127.6, 129.1,
129.8, 131.9, 132.0, 132.8, 134.3, 135.9, 136.2, 136.6, 140.7,
152.1 ppm; [R]D ) +250.4° (c ) 1.0, MeOH); IR (KBr) 3150,
2213, 1634, 1512, 1340, 1152, 582 cm-1; MS (M + H)+ 490.
Anal. C, H, N, S. Spectral data for HCl salt: 1H NMR (CD3-
OD, 300MHz) δ 2.90 (m, 2H), 3.15 (m, 2H), 3.90 (m, 1H), 4.3
to 5.1 (m, 4H), 6.40 (d, 7 Hz, 1H), 7.0-7.6 (m, 11H), 8.90 (s,
1H).
(R)-2,3,4,5-Tet r a h yd r o-1-(1H -im id a zol-4-ylm et h yl)-4-
(m et h ylsu lfon yl)-3-(p h en ylm et h yl)-7-(4-p yr id in yl)-1H -
1,4-ben zod ia zep in e, Dih yd r och lor id e (28). A degassed
solution of 32 (55.6 mmol, 19.2 g), 4-stannylpyridine (111
mmol, 40.9 g) and Pd(PPh3)4 (8.24 mmol, 9.6 g) in toluene (2000
mL) was heated at 110 °C for 16 h. The mixture was
concentrated, diluted with 1:1 ether/hexanes and filtered. The
solid was washed with 500 mL of 1:1 ether/hexanes to isolate
16.7 g of (R)-2,3,4,5-tetrahydro-3-(phenylmethyl)-7-(4-pyridi-
nyl)-1H-1,4-benzodiazepine-2,5-dione. The combined filtrate
was concentrated and filtered to yield a second batch (5.8 g,
80% total). This crude material was carried on to 28 as a yellow
1
solid as described for 20. H NMR (400 MHz, CD3OD) δ 8.95
(1H, s), 8.75 (2H, d, J ) 6.5 Hz), 8.33 (2H, d, J ) 6 Hz), 7.9
(2H, br s), 7.55 (1H, s), 7.45-7.25 (5H, m), 7.15 (1H, m), 4.9-
4.6 (3H, m), 4.52 (1H, br s), 3.95 (1H, m), 3.48 (1H, m), 3.36
(1H, s), 2.95 (2H, m), 2.38 (3H, s); MS (M + H)+ 474.
The following compounds were prepared similarly using the
indicated acylating/sulfonylating agent.
(R )-7-Cya n o-2,3,4,5-t e t r a h yd r o-1-(1H -im id a zol-4-yl-
m e t h yl)-3-(p h e n ylm e t h yl)-4-(p r op ylsu lfon yl)-1H -1,4-
ben zod ia zep in e, Mon oh yd r och lor id e (23). From propane-
(R )-7-Cya n o-1,2,3,5-t e t r a h yd r o-1-(1H -im id a zol-4-yl-
m e t h yl)-3-(p h e n ylm e t h yl)-4H -1,4-b e n zod ia ze p in e -4-
ca r boxylic Acid , Eth yl Ester , Hyd r och lor id e (30). A
solution of 34 (1.4 g, 5.3 mmol) and di-tert-butyl dicarbonate
(1.23 g) in THF (20 mL) was stirred for 10 h. The solvent was
evaporated and the residue was chromatographed on silica
(30% AcOEt/hexanes) to provide (R)-2,3,4,5-tetrahydro-3-(phe-
nylmethyl)-4-[(1,1-dimethylethoxy)carbonyl]-1H-1,4-benzodi-
azepine-7-carbonitrile as a white solid (1.2 g). A solution of
this material (1.2 g) and 4-formylimidazole (0.65 g, 2 equiv)
in 1:5 AcOH/CH2Cl2 (30 mL) was stirred at room temperature
for 1 h. NaBH(OAc)3 (1.49 g, 2.1 equiv) was added and stirring
was continued for 14 h. The reaction was quenched with
concentrated NH4OH and diluted with 10% iPrOH in CH2Cl2
(50 mL). The organic phase was separated and washed with 1
N NaOH (2 × 20 mL) and dried over Na2SO4. The solvent was
1
sulfonyl chloride. H NMR (CD3OD, 300MHz) δ 0.82 (t, 3H, J
) 7.27), 1.65-1.70 (m, 2 H), 2.29-2.36 (m, 1H), 2.72-2.79 (m,
1H), 2.85-2.90 (m, 1H), 3.07-3.13 (m, 1H), 3.20-3.27 (m, 1H),
3.55-3.62 (m, 1H), 4.31-4.51 (m, 3H), 6.55 (d, 1H, J ) 8.12),
7.24-7.34 (m, 8H). Anal. C, H, N, S, Cl. MS (M + H)+ 450;
[R]D ) +201° (c ) 1.41, MeOH); mp 140-151 °C.
(R )-7-Cya n o-1,2,3,5-t e t r a h yd r o-1-(1H -im id a zol-4-yl-
m et h yl)-N,N-d im et h yl-3-(p h en ylm et h yl)-4H -1,4-b en zo-
diazepin e-4-car boxam ide, Mon oh ydr och lor ide (29). From
dimethylcarbamoyl chloride. Anal. C, H, N, S, Cl. MS (M +
H)+ 415; [R]D ) +244° (c ) 0.24, MeOH); mp 147-150 °C.
(R )-7-Cya n o-2,3,4,5-t e t r a h yd r o-1-(1H -im id a zol-4-yl-
m eth yl)-3-(ph en ylm eth yl)-4-ph en ylsu lfon yl-1H-1,4-ben zo-
d ia zep in e, Mon oh yd r och lor id e (22). From benzenesulfonyl

Discovery of BMS-214662
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 20 3595
Nomeir, A. A.; Liu, M. Identification of Pharmacokinetically
Stable 3,10-Dibromo-8-chlorobenzocycloheptapyridine Farnesyl
Protein Transferase Inhibitors with Potent Enzyme and Cellular
Activities. J . Med. Chem. 1999, 42, 2651-2661.
evaporated to give (R)-2,3,4,5-tetrahydro-1-(1H-imidazol-4-
ylmethyl)-3-(phenylmethyl)-4-[(1,1-dimethylethoxy)carbonyl]-
1H-1,4-benzodiazepine-7-carbonitrile as a yellow solid (1.2 g).
A solution of this material and HCl-dioxane (4.0M, 5 mL) in
AcOEt (20 mL) was stirred for 2 h and ether (20 mL) was
added. The resulting precipitate was filtered, washed with
ether under nitrogen and dried under vacuum to provide 1.6
g of (R)-2,3,4,5-tetrahydro-1-(1H-imidazol-4-ylmethyl)-3-(phe-
nylmethyl)-1H-1,4-benzodiazepine-7-carbonitrile (35). Routine
(8) Ding, C. Z.; Batorsky, R.; Bhide, R.; Chao, H. J .; Cho, Y.; Chong,
S.; Gullo-Brown, J .; Guo, P.; Kim, S. H.; Patel, M.; Penhallow,
B. A.; Ricca, C.; Rose, W. C.; Schmidt, R.; Slusarchyk, W. A.;
Vite, G.; Yan, N.; Manne, V.; Hunt, J . T. Discovery and
Structure-Activity Relationships of Imidazole-Containing Tet-
rahydrobenzodiazepine Inhibitors of Farnesyltransferase. J .
Med. Chem. 1999, 42, 5241-5253.
1
reaction with ethyl chloroformate then afforded 30. H NMR
(9) Njoroge, F. G.; Taveras, A. G.; Kelly, J .; Remiszewski, S. W.;
Mallams, A. K.; Wolin, R.; Afonso, A.; Cooper, A. B.; Rane, D.;
Liu, Y.-T.; Wong, J .; Vibulbhan, B.; Pinto, P.; Deskus, J .; Alvarez,
C.; Rosario, J . D.; Connolly, M.; Wang, J .; Desai, J . A.; Rossman,
R. R.; Bishop, W. R.; Patton, R.; Wang, L.; Kirschmeier, P.;
Bryant, M. S.; Nomeir, A. A.; Lin, C.-C.; Liu, M.; McPhail, A.
T.; Doll, R. J .; Girijavallabhan, V.; Ganguly, A. K. (+)-4-[2-[4-
(8-Chloro-3,10-dibromo-6,11-dihydro-5H-benzo[5,6]cyclohepta-
[1,2-b]pyridin-11(R)-yl)-1-piperidinyl]-2-oxoethyl]-1-piperidine-
carboxamide (Sch-66336): A Very Potent Farnesyl Protein
Transferase Inhibitor as a Novel Antitumor Agent. J . Med.
Chem. 1998, 42, 2651-2661.
(10) Adjei, A. A.; Erlichman, C.; Davis, J . N.; Cutler, D. L.; Sloan, J .
A.; Marks, R. S.; Hanson, L. J .; Svingen, P. A.; Atherton, P.;
Bishop, W. R.; Kirschmeier, P.; Kaufmann, S. H. A Phase I Trial
of the Farnesyl Transferase Inhibitor SCH66336: Evidence for
Biological and Clinical Activity. Cancer Res. 2000, 60, 1871-
1877.
(11) Zujewski, J .; Horak, I. D.; Bol, C. J .; Woestenborghs, R.; Bowden,
C.; End, D. W.; Piotrovsky, V. K.; Chiao, J .; Belly, R. T.; Todd,
A.; Kopp, W. C.; Kohler, D. R.; Chow, C.; Noone, M.; Hakim, F.
T.; Larkin, G.; Gress, R. E.; Nussenblatt, R. B.; Kremer, A. B.;
Cowan, K. H. Phase I and Pharmcokinetic Study of Farnesyl
Protein Transferase Inhibitor R115777 in Advanced Cancer. J .
Clin. Oncol. 2000, 18, 927-941.
(12) Gibbs, J . B.; Anthony, N. J .; Buser, C. A.; deSolms, S. J .; Graham,
S. L.; Hartman, G. D.; Heimbrook, D. C.; Lobell, R. B.; Koblan,
K. S.; Kohl, N. E.; Williams, T. M. Farnesyltransferase Inhibitors
as Potential Anticancer Agents. Book of Abstracts, 219th
National Meeting of the American Chemical Society, San
Francisco, CA, Mar. 26-30, 2000; Abstr. #288.
(400 MHz, CD3OD) δ 8.9 (1H, J ) 16 Hz), 7.48-7.12 (m, 8H),
6.9 (m, 1H), 5.0-4.4 (m, 5H), 4.8-3.7 (m, 3H), 3.4-3.2 (m, 2H),
2.89-2.7 (m, 2H), 1.03 (m, 3H); MS (M + H)+ 416.
(R)-4-Acetyl-7-cya n o-2,3,4,5-tetr a h yd r o-1-(1H-im ida zol-
4-ylm et h yl)-3-(p h en ylm et h yl)-1H -1,4-b en zod ia zep in e,
Mon oh yd r och lor id e (27). By carbodiimide coupling of 35
with acetic acid. 1H NMR (CD3OD, 300MHz) δ 1.95 (s, 3H),
2.80 (m, 2H), 3.2-3.35 (m, 2H), 3.9 (m, 1H), 4.3 to 5.1 (m, 4H),
6.85 (m, 1H), 7.1 to 7.6 (m, 8H), 8.92 (d, 27 Hz, 1H); MS (M +
H)+ 386.
(R )-7-Cya n o-4-[[2-(d im e t h yla m in o)e t h yl]su lfon yl]-
2,3,4,5-tetr a h yd r o-1-(1H-im id a zol-4-ylm eth yl)-3-(p h en yl-
m eth yl)-4H-1,4-ben zod ia zep in e, Dih yd r och lor id e (21).
2-Chloroethanesulfonyl chloride (1.85 g, 11.4 mmol) was added
to a solution of 35 (1.0 g, 3.8 mmol) and DIEA (2.6 mL, 15.16
mmol) in dichloromethane (16 mL) at 0 °C under argon. After
stirring for 16 h, the mixture was diluted with chloroform (20
mL) and NaHCO3 (5 mL). The layers were separated and the
aqueous layer was extracted with chloroform (2 × 50 mL). The
combined organic extracts were washed with NaHCO3 (2 ×
20 mL) and brine (2 × 50 mL), dried, filtered and concentrated.
Chromatography (silica, 75% then 50% hexanes/EtOAc) af-
forded (R)-7-cyano-4-[(2-chloroethyl)sulfonyl]-2,3,4,5-tetrahy-
dro-1-(1H-imidazol-4-ylmethyl)-3-(phenylmethyl)-4H-1,4-ben-
zodiazepine (0.31 g, 23%). A solution of this material (0.59 g,
1.36 mmol) in a 2 M solution of dimethylamine in THF (15
mL, 30 mmol) was heated in a sealed tube at 60 °C for 16 h.
The mixture was concentrated and the residue was purified
by preparative HPLC (gradient of 30-90% aqueous methanol
with 0.1% TFA). Conversion to the HCl salt afforded 21 (11
(13) Manne, V.; Ricca, C. S.; Brown, J . G.; Tuomari, A. V.; Yan, N.;
Patel, D. V.; Schmidt, R.; Lynch, M. J .; Ciosek, C. P.; Carboni,
J . M.; Robinson, S.; Gordon, E. M.; Barbacid, M.; Seizinger, B.
1
mg, 1.7%). H NMR (400 MHz, CD3OD) δ 8.9 (s,1H), 7.5-7.2
R.; Biller, S. A. Ras Farnesylation as
a Target for Novel
Antitumor Agents: Potent and Selective Farnesyl Diphosphate
Analogue Inhibitors of Farnesyltransferase. Drug Dev. Res. 1995,
34, 121-137.
(m, 8H), 6.9 (m, 1H), 4.8-4.4 (m, 5H), 3.95 (m, 1H), 3.4-3.1
(m, 5H), 3.0-2.7 (m, 8H); MS (M + H)+ 479.
(14) Manne, V.; Yan, N.; Carboni, J . M.; Tuomari, A. V.; Ricca, C. S.;
Brown, J . G.; Andahazy, M. L.; Schmidt, R. J .; Patel, D.; Zahler,
R.; Weinmann, R.; Der, C. J .; Cox, A. D.; Hunt, J . T.; Gordon, E.
M.; Barbacid, M.; Seizinger, B. R. Bisubstrate Inhibitors of
Farnesyltransferase: A Novel Class of Specific Inhibitors of Ras
Transformed Cells. Oncogene 1995, 10, 1763-1779.
Ack n ow led gm en t. Microanalyses, IR spectra, and
mass spectra were kindly provided by the Bristol-Myers
Squibb Department of Analytical Research and Devel-
opment.
(15) Rose, W. C. Taxol-Based Combination Chemotherapy and Other
In Vivo Preclinical Antitumor Studies. J . Natl. Cancer Inst.
Monogr. 1993, 15, 47-53.
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