Second generation 'peptoid' CCK-B receptor antagonists: Identification and development of N-(adamantyloxycarbonyl)-α-methyl(R)-tryptophan derivative (CI-1015) with an improved pharmacokinetic profile

Article abstract of DOI:10.1021/jm970065l

We have previously described the design and development of CI-988, a peptoid analogue of CCK-4 with excellent binding affinity and selectivity for the CCK-B receptor. Due to its anxiolytic profile in animal models of anxiety, this compound was developed as a clinical candidate. However, during its development, it was determined that CI-988 had low bioavailability in both rodent and nonrodent species. In the clinic, it was further established that CI-988 had poor bioavailability. Thus, there was a need to identify an analogue with an improved pharmacokinetic (PK) profile. The poor bioavailability was attributed to poor absorption and efficient hepatic extraction. We envisaged that reducing the molecular weight of the parent compound (5, MW = 614) would lead to better absorption. Thus, we synthesized a series of analogues in which the key α-methyltryptophan and adamantyloxycarbonyl moieties, required for receptor binding, were kept intact and the C-terminus was extensively modified. This SAR study led to the identification of tricyclo[3.3.1.1^3,7]dec-2-yl [1S-[1α(S*)2β]-[2-[(2- hydroxycyclohexyl)amino]-1-(1H-indol-3-ylmethyl)-1-methyl-2- oxoethyl]carbamate (CI-1015, 31) with binding affinities of 3.0 and 2900 nM for the CCK-B and CCK-A receptors, respectively. The compound showed CCK-B antagonist profile in the rat ventromedial hypothalamus assay with a K(e) of 34 nM. It also showed an anxiolytic like profile orally in a standard anxiety paradigm (X-maze) with a minimum effective dose (MED) of 0.1 μg/kg. Although the compound is less water soluble than CI-988, oral bioavailability in rat was improved nearly 10 times relative to CI-988 when dosed in HPβCD. The blood-brain permeability of CI-1015 (31) was also enhanced relative to CI- 988 (5). On the basis of the overall improved pharmacokinetic profile as well as enhanced brain penetration, CI-1015 (31) was chosen as a development candidate.

Full text of DOI:10.1021/jm970065l

38
J . Med. Chem. 1998, 41, 38-45
Secon d Gen er a tion “P ep toid ” CCK-B Recep tor An ta gon ists: Id en tifica tion a n d
Develop m en t of N-(Ad a m a n tyloxyca r bon yl)-r-m eth yl-(R)-tr yp top h a n Der iva tive
(CI-1015) w ith a n Im p r oved P h a r m a cok in etic P r ofile
Bharat K. Trivedi,*,† J anak K. Padia,† Ann Holmes,† Steven Rose,‡ D. Scott Wright,‡ J oanna P. Hinton,‡
Martyn C. Pritchard,§ J on M. Eden,§ Clare Kneen,§ Louise Webdale,§ Nirmala Suman-Chauhan,§ Phil Boden,§
Lakhbir Singh,§ Mark J . Field,§ and David Hill§
Departments of Medicinal Chemistry and Pharmacokinetics and Drug Metabolism, Parke-Davis Pharmaceutical Research,
Division of Warner-Lambert Company, 2800 Plymouth Road, Ann Arbor, Michigan 48105, and Parke-Davis Research Unit,
Cambridge University Fotvie Site, Robinson Way, Cambridge CB2 2QB, U.K.
Received February 3, 1997X
We have previously described the design and development of CI-988, a peptoid analogue of
CCK-4 with excellent binding affinity and selectivity for the CCK-B receptor. Due to its
anxiolytic profile in animal models of anxiety, this compound was developed as a clinical
candidate. However, during its development, it was determined that CI-988 had low
bioavailability in both rodent and nonrodent species. In the clinic, it was further established
that CI-988 had poor bioavailability. Thus, there was a need to identify an analogue with an
improved pharmacokinetic (PK) profile. The poor bioavailability was attributed to poor
absorption and efficient hepatic extraction. We envisaged that reducing the molecular weight
of the parent compound (5, MW ) 614) would lead to better absorption. Thus, we synthesized
a series of analogues in which the key R-methyltryptophan and adamantyloxycarbonyl moieties,
required for receptor binding, were kept intact and the C-terminus was extensively modified.
This SAR study led to the identification of tricyclo[3.3.1.13,7]dec-2-yl [1S-[1R(S*)2â]-[2-[(2-
hydroxycyclohexyl)amino]-1-(1H-indol-3-ylmethyl)-1-methyl-2-oxoethyl]carbamate (CI-1015, 31)
with binding affinities of 3.0 and 2900 nM for the CCK-B and CCK-A receptors, respectively.
The compound showed CCK-B antagonist profile in the rat ventromedial hypothalamus assay
with a Ke of 34 nM. It also showed an anxiolytic like profile orally in a standard anxiety
paradigm (X-maze) with a minimum effective dose (MED) of 0.1 µg/kg. Although the compound
is less water soluble than CI-988, oral bioavailability in rat was improved nearly 10 times
relative to CI-988 when dosed in HPâCD. The blood-brain permeability of CI-1015 (31) was
also enhanced relative to CI-988 (5). On the basis of the overall improved pharmacokinetic
profile as well as enhanced brain penetration, CI-1015 (31) was chosen as a development
candidate.
Cholecystokinin (CCK) has been implicated in the
neurobiology of anxiety and panic disorder. CCK-4, the
tetrapeptide, has been shown to produce panic-like
attacks in humans and produces anxiogenesis in animal
models of anxiety.1 CCK-B receptor antagonists have
been shown to produce anxiolytic-like activity in pre-
clinical studies. Due to continued interest in developing
novel therapeutic agents, which interact selectively with
the cholecystokinin receptors for the treatment of
anxiety, panic disorder, etc., several different classes of
antagonists have been developed.2,3 We previously
reported a series of peptoid analogues with excellent
affinity and selectivity for the CCK-B receptor.4 From
this series of compounds, CI-988 (Figure 1) was devel-
oped as a clinical candidate due to anxiolytic activity
observed in an established in vivo paradigms such as
X-maze in rat.5 However, during the preclinical and
clinical development of this compound, it was deter-
mined that its bioavailability was very low (1-3%) in
rat,6 monkey,7,8 and humans.9 The low bioavailability
F igu r e 1. Chemical structures of CI-988 (5), CI-1015 (31),
and 7.
† Department of Medicinal Chemistry.
‡ Department of Pharmacokinetics and Drug Metabolism.
in preclinical studies was attributed to inefficient
absorption as well as high biliary excretion in part due
§ Parke-Davis Research Unit, U.K.
X Abstract published in Advance ACS Abstracts, December 1, 1997.
S0022-2623(97)00065-4 CCC: $15.00 © 1998 American Chemical Society
Published on Web 01/01/1998

Second-Generation “Peptoid” CCK-B Receptor Antagonists
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 1 39
Sch em e 1. Synthesis of 31 [CI-1015]
(I)
to the high molecular weight (MW ) 614) of the
compound. Thus, there was a need to identify an
analogue with an improved pharmacokinetic profile. It
is of interest to note that similar issues faced the
development of L-365260, a benzodiazepine-based CCK-B
receptor antagonist developed by Merck Laboratories,
which was also shown to have low oral bioavailability
in preclinical studies.10 The lack of oral bioavailability
was attributed in part to the poor aqueous solubility
(<0.002 mg/mL) of the compound. Thus, attempts were
made to improve solubility and absorption of the com-
pound by incorporation of polar ionic functionalities into
the molecule to enhance aqueous solubility.11,12
We approached this problem in the following manner.
Since these analogues were easily accessible from opti-
cally active 2-Adoc-R-methyltrp acid, we chose to pre-
pare analogues modified at the C-terminus. Since the
molecular weight seemed to have contributed to the lack
of oral absorption, we dissected the molecule in four
different parts (Figure 1). We analyzed each part of the
molecule and concluded that fragments A and B,
required for high affinity, contributed minimally toward
the molecular weight. Fragment C, which contributes
30% of the MW, was critical for affinity. However,
fragment D, which represented 38% of the MW, could
be further manipulated since earlier SAR studies had
suggested that binding affinity was maintained while
changing the C-terminus. Additionally, we were also
interested in identifying requisite key structural fea-
tures of compound 3 for high receptor affinity, which
had been identified as one of the most potent ligands
for the CCK-B receptor.13 Thus, we began a systematic
study of the SAR at the C-terminus.
F igu r e 2. X-ray of CI-1015 (31).
were synthesized by first reacting (R)-R-methylbenzy-
lamine with cyclohexane epoxide in the presence of
trimethylaluminum in CH2Cl2.14,15 Separation of the
diastereomers using column chromatography followed
by hydrogenation provided both chiral amino alcohols.
Coupling of these amino alcohols with the acid I
provided corresponding amides 31 and 32. The absolute
stereochemistry for both the amino alcohols was previ-
ously assigned.14,15 On the basis of the comparative
physicochemical data (i.e. melting point, rotation), we
assigned the absolute stereochemistry to the amino
alcohols. This was further corroborated by obtaining
an X-ray crystal structure16 for compound 31 (Figure
2) which confirmed the relative (trans) and absolute
stereochemistry at both the amine and alcohol centers
being S,S. Thus, we assigned the R,R stereochemistry
for the other isomer (32). Similarly, compounds 33 and
34 were also synthesized.
Ch em istr y
Synthesis of these analogues was carried out using
previously described methods.4,13 Thus, coupling of an
amine with N-2-Adoc-R-methyl-(R)-tryptophan (I) using
dicyclohexylcarbodiimde (DCC) and hydroxy benzotria-
zole (HOBt) or pentafluorophenol (PFP) in ethyl acetate
provided targeted analogues. Example 31 (CI-1015) and
32 were synthesized as shown in Scheme 1. The
requisite optically active trans-2(S)-hydroxy-1(S)-ami-
nocyclohexane and the corresponding R,R enantiomer
Resu lts a n d Discu ssion
Earlier studies on this class of compounds had
revealed that replacement of the C-terminus of CI-988
with a phenethylamine moiety provided a compound
with 32 nM affinity for the CCK-B receptor. Interest-
ingly, incorporation of an acetic acid side chain R to the

40 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 1
Trivedi et al.
amine provided compound 3 with 200-fold improvement
in binding affinity with a Ki of 0.15 nM. This is one of
the most potent antagonists from the “dipeptoid” series.
To define which functional group was critical for potency
in 3, we chose to prepare analogues in which the
carboxylate moiety of 3 was first replaced with a phenyl
group (8). Insertion of an additional phenyl provided a
compound with improved affinity and selectivity (8 vs
1); however, it was 100-fold less potent than compound
3. We then prepared a compound in which the phenyl
moiety of 3 was replaced with a carboxylate function
(10). It was anticipated that if the acid functionality
in this part of molecule is rendering potency via H-
bonding interactions, then compound 10 would provide
such an interaction as well. This would provide us the
opportunity to synthesize analogues with significantly
increased hydrophilicity and, thus, improve oral absorp-
tion. However, compound 10, having two carboxylic
acid moieties at the C-terminus, lost all of the binding
affinity whereas the corresponding ester (9) showed
modest binding affinity. These data suggest that having
an additional ionic function in this part of the molecule
is detrimental to binding. Additionally, it confirms that
both a polar carboxylate and a hydrophobic phenyl
moieties are necessary for high affinity for the CCK-B
receptor.
significantly reduced binding affinity. Both compounds
were 4-40-fold less potent than 27.
Further exploration of the SAR revealed that incor-
poration of a hydroxyl moiety on the vicinal carbon (30)
provided a compound with 6.2 nM binding affinity for
the CCK-B receptor. Separation of the individual
diastereomers provided us with compound 31 with
binding affinities of 3.0 and 2900 nM for the CCK-B and
CCK-A receptors, respectively. The other diastereomer
(32) was less potent and selective. Increasing the size
of the cycloalkyl functionality once again provided us
with a set of diastereomers (33, 34) which showed
stereoselective interactions at the receptor. However,
compound 33 was less selective compared to the corre-
sponding cyclohexyl derivative (31). Similar observa-
tions of stereselective interactions for a series of ben-
zodiazepine analogues at the CCK-A and CCK-B
receptors have been reported.17,18
From this in vitro SAR study a few compounds were
further evaluated in the secondary functional assays.
In particular, we compared extensively two compounds
with a polar hydroxyl group (7, 31). On the basis of
our earlier observations that for this class of peptoids
compounds having a free carboxyl group were poorly
absorbed, we chose not to pursue similar compounds
(27) despite their high potency. Thus, compounds 7 and
31 were first evaluated in an in vitro ventromedial
hypothalamus (VMH) assay.19 These analogues showed
Ke values of 1.8 and 34 nM, respectively. These
analogues were also assayed (iv) for their ability to block
pentagastrin-induced acid secretion in the Ghosh and
Schild test in rats. The ED50 values for 7 and 31 were
0.5 and 0.3 mg/kg, respectively. These values compared
well with the ED50 of 0.2 mg/kg for CI-988.
We then prepared a series of compounds in which we
incorporated a variety of small amine side chains at the
C-terminus (Table 1). The binding data suggested that
for this series of compounds, a nonplanar lipophilic
moiety was preferred over a planar hydrophobic func-
tion (12 vs 11). Additionally, N-alkylation of 12 pro-
vided a compound with loss in binding affinity and
selectivity, suggesting that the free NH was required
for potency. On the basis of these preliminary observa-
tions, we focused our efforts toward the synthesis of a
series of analogues having a variety of cycloalkylamines
and hydrazines. A series of hydrazides were prepared
(14-19). Interestingly, the binding affinity improved
with the increase in lipophilicity. Thus, the azabicyclo
octane analogue 17 was 20-fold more potent than 14
with a binding affinity of 6.5 nM for the CCK-B receptor.
Incorporation of a methoxymethyl functionality on to
1-aminopyrrolidine gave the corresponding S (18) and
R (19) (methoxymethyl)pyrrolidine derivatives. These
These analogues were further examined for their
ability to antagonize CCK induced behavioral response
in different animal models in vivo. They were compared
with CI-988 in the elevated rat X-maze test where CI-
988 had previously been shown to have an anxiolytic-
like profile.20 Oral administration of 7 (0.01-100 µg/
kg) and CI-988 (1-10000 µg/kg) 40 min before test
increased time spent by rats on the end sections of the
X-maze with minimum effective doses (MED) of 0.1 and
10 µg/kg, respectively (Figure 3). A reduction in activity
was observed for 7 at the high dose. Similarly, admin-
istration of 31 (0.1-100 µg/kg) 30 min before test also
increased the time spent on the end sections of the
X-maze with a MED of 1.0 µg/kg. The increase in the
percent time and entries onto the end sections of the
X-maze suggests that these compounds possess anxi-
olytic-like activity. None of these analogues altered the
total number of entries, suggesting a lack of effect on
spontaneous locomotor activity. Similarly, compounds
7 and 31 were compared in the light-dark box test in
mice in which standard anxiolytics have been shown to
be active. In this test (Figure 4), these compounds
showed an increase in time spent by mice in the
illuminated side, suggesting anxiolytic-like activity with
MED of 1 and 10 µg/kg, respectively.
analogues
showed
stereospecific interactions at the receptor, and thus, 19
was more potent and selective with binding affinities
of 2.5 and 3430 nM for CCK-B and CCK-A receptors,
respectively. It is also of interest to note that, for the
first time, the C-terminus side chain was lacking a
carboxylic acid moiety which was earlier thought to be
critical for potency.
We then prepared analogues in which we incorporated
additional hydrophilicity by incorporating carboxylic
acid and primary alcohol into the molecules. Geminal
substitution of such functionalities provided analogues
with only modest binding affinity and selectivity (23-
26). However, vicinal substitutions provided analogues
with significantly improved binding affinity. Thus,
compound 27, a mixture of diastereomers, showed an
excellent binding affinity of 0.99 nM for the CCK-B
receptor and showed 700-fold selectivity. Removal of
the acid with a cyano (28) or a methyl group (29)
Encouraged by the in vivo activity observed for these
analogues, we then evaluated compounds 7 and 31 for
their overall pharmacokinetic profile in rat (Table 2).
These compounds were administered orally at 20 mg/
kg and were delivered in a variety of vehicles. A marked

Second-Generation “Peptoid” CCK-B Receptor Antagonists
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 1 41
Ta ble 1. CCK Receptor Binding Affinities
a
b
(R,S)-R-methyltrp acid. IC50 values are average of triplicate determinations for each compound.
improvement in the oral bioavailability (%F) of 31
relative to that for CI-988 (5) was observed with
different formulations studied. The optimal bioavail-
ability (28%) was achieved with 31 dosed as a solution
in (hydroxypropyl)-â-cyclodextrin [50% (w/v) HPâCD in
saline:0.1 N HCl in ethanol (7:3, v/v)]. This formulation
offered nearly 10-fold improvement over CI-988. The
other three formulations had lower yet similar bioavail-
ability of about 10%, which represented about 2-3-fold
improvement relative to CI-988. Oral bioavailability of
7 was also better than that for CI-988 for three of the
four formulations tested. When dosed as a solution in
HPâCD, the highest bioavailability of 16% was achieved
for 7. As a solution in either PEG 400 or dimethyl-
acetamide:water (3:7), bioavailability was comparable
at nearly 10%. However, low bioavailability similar to
that of CI-988 was observed when dosed as an aqueous
suspension in 5% methylcellulose.
Thus, for both the compounds (7 and 31), solution
formulations in general achieved better systemic bio-
availability than did suspension formulations. These
results suggested that the oral absorption of these
neutral compounds was solubility limited. Conse-
quently with the right formulation, overall systemic
exposure of these second-generation “peptoids” could be
markedly improved relative to CI-988. Use of HPâCD

42 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 1
Trivedi et al.
F igu r e 4. Effects of CI-988 (5), 7, and CI-1015 (31) in the
mouse light-dark box. CI 988 and 7 were administered sc in
PEG 400, 40 min before test. CI-1015 (31) was administered
sc 30 min before measuring the time spent in the light side
during the 5 min observation period. Results are shown as the
mean (verticle bars represent ( SEM) of 10 animals per group.
*Significantly different from vehicle treated controls, P < 0.05
(ANOVA followed by Dunnett’s t-test).
F igu r e 3. Effects of CI-988 (5), 7, and CI-1015 (31) in the rat
elevated X-maze. CI 988 and 7 were administered po in PEG
400, 40 min before test. CI-1015 (31) was administered po 30
min before test. The percent time spent (%TOEA) and entries
made (%EOEA) onto the end half sections of the open arms
and the total number of entries (TE) were measured for each
animal during the 5 min test. Results are shown as the mean
(verticle bars represent ( SEM) of 10 animals per group.
*Significantly different from vehicle treated controls, P < 0.05
(ANOVA followed by Dunnett’s t-test).
their ability to penetrate the blood-brain barrier in an
ex vivo experiment. Following the methods described,
brain levels of each drug were determined at 5, 10, and
20 min post iv dose (Table 3). Interestingly, CI-988 (5)
even after administration of 10 mg/kg iv dose showed
only marginal drug levels whereas L-365260, a benzo-
diazepine-based CCK-B antagonist, showed modest
brain levels after 5 min (115 pmol/forebrain) which were
significantly reduced after 20 min (33 pmol/forebrain).
in oral formulations has been found to be safe for
preclinical studies with only about 2% systemic avail-
ability.21,22 Indeed with the decrease in molecular
weight and absence of any ionizable group, the overall
bioavailability was noticeably improved relative to CI-
988.
These new analogues were compared to CI-988 (5) for

Second-Generation “Peptoid” CCK-B Receptor Antagonists
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 1 43
Ta ble 2. Summary of Mean (% RSD) Pharmacokinetic Parameters from Fasted Wistar Rats following a Single Oral 20 mg/kg Dose of
Different CCK-B Antagonistsa
compound
formulation
physical form
Cmax (ng/mL)
tmax (h)
AUC(0-∞) (ng‚h/mL)
t1/2 (h)
%F
5 [CI-988]
saline
solution
solution
suspension
suspension
solution
solution
suspension
solution
solution
solution
32 (19)
5 (51)
2.3 (140)
0.25 (0)
3.0 (38)
3.5 (75)
1.0 (71)
1.8 (89)
2.9 (45)
1.2 (42)
2.8 (54)
0.9 (88)
134 (34)
2.3 (80)
ND
ND
3.2 (44)
<1
HPâCDb
31 [CI-1015]
0.5% MCc
10% PEGd
100% PEGe
HPâCDf
185 (56)
203 (35)
219 (14)
756 (35)
42 (20)
212 (45)
167 (54)
416 (31)
673 (47)
953 (35)
724 (26)
2130 (12)
188 (32)
890 (26)
886 (64)
1440 (26)
1.0 (10)
1.1 (41)
1.3 (22)
0.9 (24)
ND
2.3 (39)
1.1 (18)
1.5 (13)
8.9 (47)
13 (35)
9.6 (25)
28 (12)
2.0 (35)
9.6 (30)
9.6 (66)
16 (30)
7
0.5% MCc
DMA:H2Og
100% PEGd
HPâCDf
a
%RSD ) percent relative standard deviation; Cmax ) maximum plasma concentration; tmax ) time to Cmax; AUC(0-∞) ) area under
b
the plasma concentration-time curve from 0 to infinity; t1/2 ) terminal elimination half-life; %F ) absolute oral bioavailability. HPâCD
) 1:2 mmolar ratio of CI-988 to HPâCD in water. c 0.5% MC ) 0.5% methylcellulose in water. 10% PEG ) 10% PEG400 in water.
d
e 100% PEG ) 100% PEG400. f HPâCD ) 50% (w/v) (hydroxypropyl)-â-cyclodextrin (HPâCD) in saline:ethanol (7:3, v/v). DMA:H2O )
g
dimethylacetamide:water (3:7, v/v).
Ta ble 3. Brain Levels of Selected Antagonists Following iv Administration Determined by Ex Vivo Binding
brain level (pmol/forebrain) after injection
compound dose [1 mg/kg]
5 min
10 min
BLDb
67 ( 14
154 ( 39
68 ( 19
20 min
mice (n)
5 [CI-988]a
L-365260
31 [CI-1015]
7
10 ( 5
115 ( 21
177 ( 46
40 ( 13
8 ( 4
33 ( 3
166 ( 35
43 ( 17
3-6
6
9
11-13
a
b
Dose of 10 mg/kg. BLD ) below level of detection (approximately 1 pmol/forebrain).
rotations were determined with the use of a Perkin-Elmer 241
polarimeter.
Compound 7 produced only modest brain levels over 20
min. However, 31 showed highest levels in the brain
and maintained those levels throughout the experiment.
Thus, it suggests that the higher lipophilicity of 31 (log
P ) 4.3) relative to CI-988 (log P ) 2.0 at pH 7) may in
part contribute to the high degree of brain penetration
observed for this compound.
N-[2-[(2-h yd r oxycycloh exyl)a m in o]-1-(1H -in d ol-3-yl-
m et h yl)-1-m et h yl-2-oxoet h yl]ca r b a m ic Acid Tr icyclo-
[3.3.1.13,7]d ec-2-yl Ester (30) a n d Sep a r a tion of Dia ster -
eom er s 31 a n d 32. A solution of R-methyl-N-[(tricyclo-
[3.3.1.13,7]dec-2-yloxy)carbonyl]-(R)-tryptophan (0.79 g, 2.0
mmol) in ethyl acetate (60 mL) was treated with N,N′-
dicyclohexylcarbodiimide (0.49 g, 2.0 mmol) and 1-hydroxy-
benzotriazole hydrate (0.3 g, 2.0 mmol). After 2 h of stirring
at room temperature, the precipitated solid was filtered and
washed with EtOAc. To the clear filtrate were added trans-
2-aminocyclohexanol hydrochloride (0.3 g, 2.0 mmol) and
triethylamine (0.2 g, 2.0 mmol), and the reaction mixture was
stirred overnight. It was washed with 5% HCl, 5% NaHCO3,
and brine. The organic extract was dried over MgSO4, filtered,
and concentrated. The residue was purified by flash column
chromatography over silica gel using hexanes-ethyl acetate
(1:1) to give 0.65 g of 30.
Con clu sion
In summary, the strategy of lowering molecular
weight of the parent compound CI-988 (5) to address
the issue of low bioavailability has provided 31 with
enhanced bioavailability. Interestingly, this peptoid
molecule, which lacks the carboxylic acid function,
earlier thought to be required for high affinity, main-
tains an excellent binding affinity and selectivity for the
CCK-B receptor. Additionally, it shows antagonist
profile both in vitro as well as in vivo. In two separate
paradigms it shows an anxiolytic profile. Furthermore,
31 not only shows better bioavailability in rodents than
5, but also has enhanced blood-brain penetration.
Thus, on the basis of its overall improved profile, 31 (CI-
1015) has been chosen for further preclinical and clinical
evaluation. It is anticipated that CI-1015 will serve as
a better tool to assess the clinical utility of CCK-B
receptor antagonist as an anxiolytic agent.
Individual diastereomers of 30 were separated by Waters
Model 46K HPLC using silica column (6 µm, Prep Nova Pak).
The mobile phase was ethyl acetate-hexane (6:4), and the flow
rate was 15 mL/min. Compound 31 (mp 125-130 °C) had a
retention time of 11.92 min, and compound 32 (mp 130-137
°C) had a retention time of 13.87 min.
Tr icyclo[3.3.1.13,7]d ec-2-yl [1S-[1r(S*)2â]-[2-[(2-H yd r -
oxycycloh exyl)a m in o]-1-(1H-in d ol-3-ylm eth yl)-1-m eth yl-
2-oxoeth yl]ca r ba m a te (31). (a ) (1S,2S)-tr a n s-2-[(R)-(r-
Meth ylben zyl)am in o]cycloh exan ol (A) an d (1R,2R)-tr a n s-
2-[(R)-(r-Meth ylben zyl)a m in o]cycloh exa n ol (B). To a
well-stirred solution of (R)-R-methylbenzylamine (6.06 g, 5.0
mmol) in CH2Cl2 (60 mL) was added dropwise a solution of
trimethylaluminum (27.5 mL, 2.0 M solution in toluene, 55
mmol) at 0-5 °C. The resulting solution was stirred for
additional 45 min, and a solution of cyclohexane epoxide in
methylene chloride (15 mL) was added dropwise at 0 °C. The
reaction mixture was stirred overnight at room temperature.
The aluminum complex was decomposed by adding NaF (8.82
g, 210 mmol) at 0 °C followed by water (6 mL). After 1 h of
stirring, the reaction mixture was passed through Celite and
concentrated. The residue was purified by flash column
chromatography on silica gel using EtOAc-hexane-Et3N (1:
1:0.1) to provide compound A (4.5 g, 39.%, Rf ) 0.5) and
compound B (4.5 g, 39%, Rf ) 0.32).
Exp er im en ta l Section
Ch em istr y. Materials used were obtained from commercial
suppliers and used without purification, unless otherwise
noted. Melting points were determined on a Thomas-Hoover
melting point apparatus and are uncorrected. NMR spectra
were determined on a Varion Unity 400 or Bruker AM300 or
250 MHz instrument or
a Varion XL-200 spectrometer.
Chemical shifts are expressed as parts per million downfield
from internal tetramethylsilane. Elemental analyses were
determined on a Perkin-Elmer Model 240C elemental analyzer
or were determined by CHN Analysis Limited, Leicester, U.K.,
and are within 0.4% of theory, unless otherwise noted. Optical

44 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 1
Trivedi et al.
(b) (1S,2S)-tr a n s-2-Am in ocycloh exa n ol. A mixture of
(1S,2S)-trans-2-[(R)-(R-methylbenzyl)amino]cyclohexanol (2.2
g, 0.01 mol), 10% Pd/C, and ammonium formate (0.38 g, 0.06
mol) in methanol (75 mL) was stirred overnight at room
temperature. The mixture was passed through Celite and
concentrated. The residue was diluted with ethyl acetate,
washed with water, dried over MgSO4, filtered, and concen-
trated to yield 0.92 g (80%) of (1S,2S)-trans-2-aminocyclohex-
anol.
and comments on the manuscript. We also thank Dr.
Andre Michel of University of Sherbrooke, Canada, for
the determination of X-ray structure for CI-1015.
Su p p or tin g In for m a tion Ava ila ble: Tables of physical
and chemical data of 8-34 and crystallographic data (14
pages). Ordering information is given on any current mast-
head page.
(c) Tr icyclo[3.3.1.13,7]d ec-2-yl [1S-[1r(S*)2â]-[2-[(2-h y-
d r oxycycloh exyl)a m in o]-1-(1H-in d ol-3-ylm eth yl)-1-m eth -
yl-2-oxoeth yl]ca r ba m a te (31). A solution of (R)-2-(adamant-
yloxycarbonyl)-R-methyltryptophan (0.79 g, 2.0 mmol) in ethyl
acetate (60 mL) was treated with dicyclohexylcarbodiimide
(0.495 g, 2.0 mmol) and 1-hydroxybenzotriazole hydrate (0.35
g, 2.2 mmol). After 2 h of stirring at room temperature, the
precipitated urea was filtered. To the clear filtrate was added
(1S,2S)-trans-2-aminocyclohexanol (0.23 g, 2.0 mmol). The
reaction mixture was stirred at room temperature overnight.
The solution was washed with 5% HCl, followed by 5%
NaHCO3 and brine. The organic phase was dried over MgSO4
and concentrated. The residue was purified by flash column
chromatography over silica gel using ethyl acetate-hexane
(1:1) to yield 0.72 g of material which was repurified on Waters
HPLC (Model 46K) using silica column (6 µm, Prep Nova Pak).
The mobile phase was ethyl acetate-hexane (1:1), and the flow
rate was 15 mL/min. Compound 31 was obtained as a solid
(0.4 g, 39.5%, retention time of 9.54 min) having a melting
point of 137-142 °C: NMR (CDCl3) 0.8-2.1 (25H, m), 2.76
(1H, br s), 3.22 (1H, dt, J ) 4.1 and 5.7 Hz), 3.33 (1H, d, J )
14.75 Hz), 3.48 (1H, d, J ) 14.76 Hz), 3.6-3.8 (1H, m), 4.84
(1H, br s), 5.21 (1H, s), 6.0 (1H, d, J ) 8.3 Hz), 7.0-7.23 (3H,
m), 7.38 (1H, d, J ) 8 Hz), 7.61 (1H, d, J ) 7.6 Hz), 8.32 (1H,
s). Anal. C, H, N.
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Biologica l Assa ys. CCK-A and CCK-B receptor binding
assays were performed as previously described.4,13 The phar-
macological activity of these antagonists was evaluated in the
rat elevated X-maze and the mouse light-dark box as de-
scribed by Singh et al.20
The pharmacokinetic studies were performed as follows:
The oral bioavailability and pharmacokinetics of these ana-
logues were evaluated in fasted Wistar rats. Groups of three
to four male wistar rats received either a single oral (po) or iv
dose of a given compound at 20 mg/kg. Due to aqueous
solubility limitations, different vehicles were used for dosing.
Heparinized plasma samples were collected from an implanted
cannula in the jugular vein at serial time out to 24 h. Plasma
samples were frozen until analysis and assayed with validated
HPLC-fluorescence methods for each compound.23 Pharma-
cokinetic parameters were determined by standard noncom-
partmental methods. Absolute oral bioavailability was cal-
culated as the dose normalized ratio of the area under the
curve following po dose to the area under the curve following
iv dose.
Ex vivo binding studies were performed as follows: Mice
were dosed intravenously with 1 mg/kg test compound or
vehicle (saline for 5 and 10% EtOH-60% propylene glycol-
30% saline for 31 and L-365260). Animals were sacrificed at
various times after dosing and brains removed. The forebrain
was dissected from the cerebellum and homogenized in 5 mL
assay buffer (10 mM Hepes, 130 nM NaCl, 4.7 mM KCl, 5 mM
MgCl2, 1 mM EGTA, containing 0.25 mg/mL bacitracin, pH
7.2 at 21 °C). Aliquots (300 µL) of tissue from animal dosed
with either test compound or vehicle were incubated with 50
pM [125I]BH-CCK8s (total assay volume ) 1 mL) for 2 h at 22
°C before filtering under vacuum onto GF\ B filters. The
amount of specifically bound radioligand was determined using
a gamma counter. Calibration curves to each compound were
run in parallel using tissue from vehicle-treated animals and
used to determine the amount of test compound present in
the brain homogenates. The limit of detection using this
methodology was approximately 1 pmol/forebrain.
Ack n ow led gm en t. The authors thank Drs. Bruce
Roth and David Horwell for their support of this work

Second-Generation “Peptoid” CCK-B Receptor Antagonists
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 1 45
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