Design and synthesis of piperidine farnesyltransferase inhibitors with reduced glucuronidation potential

Article abstract of DOI:10.1016/j.bmc.2006.11.007

The design and synthesis of a novel piperidine series of farnesyltransferase (FTase) inhibitors with reduced potential for metabolic glucuronidation are described. The various substitution and exchange of the phenyl group at the C-2 position of the previously described 2-(4-hydroxy)phenyl-3-nitropiperidine 1a (FTase IC₅₀ = 5.4 nM) resulted in metabolically stable compounds with potent FTase inhibition (14a IC₅₀ = 4.3 nM, 20a IC₅₀ = 3.0 nM, and 50a IC₅₀ = 16 nM). Molecular modeling studies of these compounds complexed with FTase and farnesyl pyrophosphate are also described.

Full text of DOI:10.1016/j.bmc.2006.11.007

Bioorganic & Medicinal Chemistry 15 (2007) 1363–1382
Design and synthesis of piperidine farnesyltransferase inhibitors
with reduced glucuronidation potential
Rieko Tanaka,a Almudena Rubio,b Nancy K. Harn,b Douglas Gernert,b
Timothy A. Grese,b Jun Eishima,a Mitsunobu Hara,a Nobuyuki Yoda,a Rui Ohashi,a
Takashi Kuwabara,a Shiro Soga,a Shiro Akinaga,a Shinji Naraa and Yutaka Kandaa,*
aPharmaceutical Research Center, Kyowa Hakko Kogyo Co. Ltd, 1188 Shimotogari, Nagaizumi-cho, Shizuoka 411-8731, Japan
bDiscovery Chemistry Research and Technology, Lilly Research Laboratories, Indianapolis, IN 46285, USA
Received 23 September 2006; revised 3 November 2006; accepted 6 November 2006
Available online 9 November 2006
Abstract—The design and synthesis of a novel piperidine series of farnesyltransferase (FTase) inhibitors with reduced potential for
metabolic glucuronidation are described. The various substitution and exchange of the phenyl group at the C-2 position of the pre-
viously described 2-(4-hydroxy)phenyl-3-nitropiperidine 1a (FTase IC50 = 5.4 nM) resulted in metabolically stable compounds with
potent FTase inhibition (14a IC50 = 4.3 nM, 20a IC50 = 3.0 nM, and 50a IC50 = 16 nM). Molecular modeling studies of these com-
pounds complexed with FTase and farnesyl pyrophosphate are also described.
ꢀ 2006 Elsevier Ltd. All rights reserved.
1. Introduction
The 3-nitro piperidine derivatives 1a and 2a have al-
ready been described as FTase inhibitors (Chart 1).5
Although the compounds have potent FTase inhibitory
activity (1a IC50 = 5.4 nM and 2a IC50 = 3.7 nM), rapid
clearance of these compounds limits their application.
Preliminary pharmacokinetic studies suggested that
glucuronidation of the phenolic groups on the benzene
ring at the C-2 position of the piperidine core could
be one of the responsible factors for the rapid clear-
ance. Here we describe our efforts to identify potent
piperidine inhibitors of FTase with reduced potential
for glucuronidation.
Ras protein plays a key role in cell growth and cell
proliferation in the MAP kinase signal transduction
pathway. Protein farnesyltransferase (FTase), an
enzyme that catalyzes farnesylation of proteins ending
with the CAAX motif (C, cystein; A, aliphatic amino
acid; and X, C-terminal amino acid), is one of the
crucial enzymes involved in the signal transduction
pathway. FTase inhibitors inhibit anchorage-indepen-
dent growth of a variety of transformed cells. A survey
of cancer cell lines has shown that >70% of cells are
sensitive to FTase inhibitors.1 In addition, FTase
inhibitors have been shown to inhibit the growth of
tumors in a number of animal model studies.2,3
Therefore, FTase inhibitors have emerged as promising
anti-cancer drugs.1 Some FTase inhibitors, such as
R115777 (tipifarnib) and SCH66336 (lonafarnib), are
currently being assessed in clinical trials and have dem-
onstrated clinical efficacy for the treatment of human
cancers.4
Br
N
Br
N
O2N
O2N
OH
N
N
HO
HO
Keywords: Farnesyltransferase (FTase) inhibitors; Piperidine deriva-
tives; Reduced glucuronidation; Design and synthesis.
1a
2a
*
Corresponding author. Tel.: +81 55 989 2008; fax: +81 55 986
7430; e-mail: ykanda@kyowa.co.jp
Chart 1.
0968-0896/$ - see front matter ꢀ 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2006.11.007

1364
R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
CHO
Br
Br
Br
N
4
HO
4
5
O2N
O2N
3
5
CO2Me
BH3 SMe2
90%
6
2
3
N
O
N
N
1
1
EtOH, reflux
NH2
73%
CH3NO2/DBU
100%
HO
HO
7
1a
6
N
Br
O2N
conc.HNO3,
CH3COOH
conc.HNO3,
CH3COOH
CO2Me
76%
62%
4
O2N
CHO
Br
O
N
Br
N
HO
O2N
O2N
8
O2N
HO
O2N
EtOH, reflux
NH2
<10%
N
N
HO
10
9
6
N
Pd/C,
H2
BH3 SMe2
63%
44%
Br
O2N
H2N
HO
N
11a
N
Scheme 1. Synthesis of 3-nitropiperidine derivatives at 2 position (1).
2. Chemistry
14a–16a, or sulfamide 20a, respectively. Reductive
alkylation of 11a with acetone and borane dimethyl sul-
fide complex gave isopropyl amino derivative 18a.
LiAlH4 reduction of 12a afforded ethyl amino derivative
17a. Since replacement of the bromo substituent on the
C-4 phenyl ring with ethyl had proven to be acceptable,
4-(2-ethylphenyl)piperidine 11b was also used as starting
material. Reductive alkylation of 11b with aryl or
heteroaryl aldehyde in the presence of sodium triacet-
oxyborohydride afforded 21b and 22a–c by parallel
solution-phase synthesis (Scheme 3).
In Scheme 1, key intermediate 11a was synthesized as
described in two previous papers.5,6 The relative stereo-
chemistry of the piperidine core was determined by
NMR analysis as described in the previous paper.5
Although the 3-component coupling reaction of 4-nit-
robutyrate 4, which was synthesized from the cinnamate
3 by the Michael addition of nitromethane7 in quantita-
tive yield, 3-nitro-4-hydroxybenzaldehyde 8, and amine
6 resulted in a low yield of the desired piperidin-2-one
9, nitration of 7 gave desired 9 in 76% yield. Reduction
of piperidin-2-one 9 with borane–methyl sulfide complex
gave the desired key intermediate 11a in 63% yield.
Alternatively, nitration of piperidine 1a followed by
hydrogenation of the nitro group gave 11a in moderate
yields.
Although the corresponding three-component coupling
reactions using 4-aminomethylimidazole or 4-amino-
methyl-1-trityl-1H-imidazole did not proceed to give
3-nitropiperidine derivative 24, reductive alkylation of
N-unsubstituted 3-nitropiperidine 23 with sodium
triacetoxyborohydride and 4-imidazolecarboxaldehyde
did give 24. Compound 23 was synthesized by the
coupling reaction of 4-nitrobutyrate 4 with benzalde-
hyde and ammonium acetate, followed by the reduction
of the carbonyl group with borane-methyl sulfide
complex. Methylation of 24 with iodomethane and
potassium carbonate afforded a 2:1 mixture of 25 and
Compound 11a was used as a starting material for the
modification of the amino group ortho to the hydroxy
group at the C-2 phenyl (Scheme 2). Treatment of 11a
with acetyl chloride, methyl chloroformate, sulfonyl
chlorides, potassium cyanate, or sulfamoyl chloride8
afforded acetamide 12a, carbamate 13a, sulfonamides

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1365
Br
Br
N
MeO
O2N
R
O2N
CO
HN
SO2
HN
N
N
HO
HO
14a R : Me 16%
15a
16a
Br
N
13a
N
Et
8%
O2N
Ac
HN
p-Tol 30%
MeOCOCl,
CH2Cl2 / DMF
21%
N
RSO2Cl, CH2Cl2 / DMF
HO
AcCl, CH2Cl2
13%
12a
Br
Br
N
KOCN,
AcOH/H2O
O2N
H2N
O2N
LiAlH4, THF
31%
CO
HN
H2N
HO
20%
N
N
HO
11a
19a
Br
N
N
O2N
Acetone,
BH3 SMe2,
46%
NH2SO2Cl,
EtHN
HO
DMA, 41
%
N
17a
Br
N
Br
Me
H2N
O2N
Me
O2N
SO2
HN
HN
HO
N
N
HO
N
18a
20a
Scheme 2. Synthesis of 3-nitropiperidine derivatives at 2 position (2).
Et
Et
N
ArCHO,
NaBH(OAc)3
O2N
O2N
21b
Ar : Ph
61%
H
N
22α
22β
22γ
2-pyridyl 37%
3-pyridyl 78%
4-pyridyl 38%
H2N
HO
Ar
N
N
HO
11b
N
Scheme 3. Synthesis of 3-nitropiperidine derivatives at 2 position (3).
26 (Scheme 4). The regiochemistry of methylation was
determined by the NOE of H NMR.
methyl)pyridine 6 and sodium triacetoxyborohydride
followed by the reduction with sodium cyanoborohy-
dride gave desired 3-allyloxycarbonylpiperidine 36.
The isomerization of the C-3 position of 36 with sodium
allyloxide proceeded to give the 2,3-trans-3,4-trans iso-
mer 38 along with 3-carboxypiperidine 37.
1
Reductive alkylation of 23 with 3-(4-cyanobenzyl)-
3H-imidazol-4-ylcarboxaldehyde afforded imidazole
derivative 31. Synthesis of 3-(4-cyanobenzyl)-3H-
imidazol-4-ylcarboxaldehyde from 4-(hydroxymethyl)
imidazole hydrochloride is shown in Scheme 5.
Acid chloride 39, prepared from 37 with thionyl
chloride, was used for the preparation of many
kinds of novel 3-substituted piperidine derivatives
(Scheme 7). 3-Hydroxymethylpiperidine 41 was pre-
pared by LiAlH4 reduction. Treatment of 39 with
methylamine, methionine methyl ester, and methanol
The syntheses of 2-phenyl-3-carboxypiperidine deriva-
tives are summarized in Scheme 6. Addition of allyl ben-
zoylacetate 32 to 2-ethylcinnamaldehyde 33 afforded
aldehyde 34. Treatment of aldehyde 34 with 3-(amino-

1366
R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1) NH4(OAc), PhCHO
EtOH, reflux, 57%
2) BH3 SMe2, THF, 41%
Br
Br
O2N
O2N
CO2Me
4
N
H
23
HN
NaBH(OAc)3
41%
,
N
OHC
MeI(1.0 eq.),
K2CO3,
Br
Br
Br
N
O2N
O2N
O2N
DMF, r.t.
+
65%
N
N
N
H
N
Me
N
26
25
24
N
N
NMe
2: 1 mixture
Scheme 4. Synthesis of imidazole derivatives.
CN
, AcOH
CN
Br
1) TrCl, TEA
2) Ac2O, Py
NH
NTr
AcO
N
HO
AcO
28
92%
68%
N
N
HBr salt
HCl salt
N
27
29
LiOH, THF, H2O (66%)
CN
1) SO3 Py, DMSO, NEt3
2) 23, NaBH(OAc)3
HO
Br
N
N
O2N
2steps 1.9%
CN
30
N
N
N
31
Scheme 5. Synthesis of 1-(4-cyanobenzyl)imidazolylmethyl derivative 31.
NH
6,
NaBH(OAc)3,
AcOH, CH2Cl2
O
O
Et
CO2Li
CHCl3
O
O
Et
+
O
2 steps 31%
O
O
32
33
34
O
Na(CN)BH3,
THF, EtOH,
HCl, bromocrezole,
green
Et
N
Et
N
Et
N
O
OR
4
O
NaOCH2CH=CH2
O
O
O
3
60%
(37a: 37b = 1:2)
12%
N
N
2
N
37 : R = H
38 : R = CH2CH=CH2
35
36
Scheme 6. Synthesis of 3-carboxypiperidine derivatives.
afforded amides 40, 43, and methyl ester 44, respec-
tively. Methionine amide 45 was prepared from 43
by hydrolysis with LiOH. Treatment of acid chloride
39 with the lithium enolate of di-tert-butylmalonate
followed by decarboxylation afforded 3-acetylpiperi-
dine 42.

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1367
O
Et
N
Et
O
Et
N
MeHN
HO
Me
N
N
N
40
41
42
N
1) CH2(CO2t-Bu)2,
LiHMDS,
NH2Me
LAH
2) AcOH,Ac2O,
camphor sulfonic acid
O
Et
N
O
Et
N
O
Et
SOCl2
MeOH
HO
Cl
MeO
N
N
N
37
39
43
N
1-Met-OMe,
pyridine
SMe
O
SMe
O
Et
O
Et
N
MeO
LiOH
HO
N
N
H
H
O
N
N
45
44
N
Scheme 7. Synthesis of piperidine derivatives at 3 position.
3. Results and discussion
Table 1. Modification of the ortho-position of the hydroxyl group at
the C-2 benzene ring on the 3-nitropiperidine (1)
Although 1a and 2a show potent FTase inhibition, the
metabolic stability of these compounds needs to be
improved. We hypothesized that the glucuronidation
of phenolic hydroxy groups might be the cause of meta-
bolic instability. Our strategy for designing compounds
includes incorporation of substituents at the ortho-posi-
tion of the phenolic hydroxy group and removal of the
groups themselves.
Br
O2N
2
N
HO
X
N
FTase enzyme inhibition was determined by inhibition
of farnesylation of human K-Ras by human FTase, as
previously described.9 In vitro metabolic stability of
selected compounds was evaluated by mouse liver
microsome assay.
Compound
X
FTase IC50 (nM)
1a
2a
H
5.4
3.7
–OH
–OCH3
–NO2
I
46a
10
180
>500
>500
13
47a
11a
Our previous studies5 suggest that the para-hydroxy
group on the C-6 benzene ring may be important for
FTase inhibition, since changing the para-hydroxy
group to a methoxy, amino or amide group resulted
in significantly reduced activity. In order to avoid
glucuronidation while maintaining FTase inhibition,
we therefore evaluated the introduction of substituents
ortho to the hydroxy group of the C-2 benzene ring
(Table 1). Although introduction of nitro or iodo groups
(compounds 10 and 47a) reduced activity, methoxy
–NH2
derivative 46a showed some activity against FTase
(IC50 = 180 nM). Interestingly, ortho-amino derivatives
(11a IC50 = 13 nM and 11b IC50 = 46 nM) retained
FTase inhibition potency.
We hypothesized that introduction of substituents into
the amino group ortho to the phenolic hydroxy group

1368
R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
Table 2. Modification of the ortho-position of the hydroxyl group at
the C-2 benzene ring on the 3-nitropiperidine (2)
Table 3. Modification at 2-position on the 3-nitropiperidine
Br
O2N
Y
O2N
R2
N
X
HO
N
N
N
FTase IC50 (nM)
Compound
R2
FTase IC50 (nM)
5.4
Compound
X
a Y = Br
b Y = Et
1a
HO
1
H
–NH2
5.4
13
50
98
20
46
11
17
18
–NHEt
–NHCH(Me)2
48a
>500
MeO
N
H
N
21
121
23
49a
50a
51a
52a
53a
>500
16
H
N
N
N
H
22a
22b
22c
H
24
N
N
H
N
12
N
36
S
12
13
14
15
16
19
20
–NHCOMe
–NHCO2Me
–NHSO2Me
–NHSO2Et
27
14
25
4.3
5.9
270
15
4.6
N
–NHSO2Tol
–NHCONH2
–NHSO2NH2
>500
6.2
4.5
N
3.0
of 11a may further reduce the rate of glucuronidation
(Table 2). Simple alkylation of the amino group resulted
in decreased potency (17a, 18a, and 21b), however, 2-
pyridylmethylamino derivative 22a (IC50 = 23 nM), 3-
pyridylmethylamino derivative 22b (IC50 = 24 nM),
Other heterocycles such as 3-thiophene 51a (IC50
=
36 nM) or 2-pyridine 52a (IC50 = 25 nM) also showed
FTase inhibition. Interestingly, the 2-pyridine derivative
53a showed no FTase inhibition.
and
4-pyridylmethylamino
derivative
22c
With the identification of non-phenolic replacements for
the R2 group, alternative substituents at N-1 of the
piperidine core (Table 4) and replacement of the C-3
nitro group were also reexamined (Table 5). Although
most of the N-1 modified derivatives lost FTase
inhibition (data not shown), substituted 4-imidazolylm-
ethyl compounds, such as 25 (IC50 = 17 nM) and 31
(IC50 = 44 nM), showed potent FTase inhibition compa-
rable to that of 3-pyridylmethyl compounds. On the
contrary, compound 26, a regio-isomer of 25, and N-un-
substituted imidazole derivative 24 showed no FTase
inhibitory activity (IC50 > 500 nM).
(IC50 = 12 nM) retained FTase inhibition. It is possible
that the nitrogen of the pyridine may have a direct inter-
action with the enzyme, however, molecular modeling
studies of 22c with FTase did not provide support for
this hypothesis. Although the methyl or ethyl sulfon-
amide series showed potent FTase inhibition (14a
IC50 = 4.3 nM, 15a IC50 = 5.9 nM), p-tosyl derivative
16a did not (IC50 = 270 nM). In this case, the bulk of
the substituent may affect FTase inhibition. Urea deriv-
ative 19a (IC50 = 15 nM) and sulfamoylamino derivative
20a (IC50 = 3.0 nM) also showed potent FTase inhibi-
tion. Ethyl analogues such as 14b (IC50 = 4.6 nM), 19b
(IC50 = 6.2 nM), and 20b (IC50 = 4.5 nM) were also
potent inhibitors.
Modification of the nitro group of 3-nitropiperidines,
such as compound 50a, is important to change the phys-
ical and biological character of these series. Table 5 sum-
marizes the structures and FTase inhibition of the
piperidine derivatives at the C-3 position. The com-
pounds with carboxylic acid or hydroxymethyl group
at the C-3 position showed no FTase inhibitory activity
(37 or 41 IC50 > 500 nM). However, 3-acetylpiperidine
42 was found to have potent activity (IC50 = 42 nM)
In an alternative strategy to avoid glucuronidation, we
tried to change the C-2 phenyl group into the other aro-
matic rings without involvement of the phenolic hydro-
xy group (Table 3). Although the methoxy derivative
48a lost FTase inhibition, the simple phenyl derivative
50a showed potent FTase inhibition (IC50 = 16 nM).

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1369
Table 4. Modification at 1-position on the 3-nitropiperidine
hypothesized that the methionine residue may interact
with the FTase like the C-terminal region of Ras pep-
tide, but this hypothesis could not be confirmed by
molecular modeling.
Br
O2N
3.1. Proposed binding model of the piperidine derivatives
for FTase
N
R1
The binding model of the piperidine derivatives in the
Ras protein-binding site of FTase was developed using
the crystal structure of rat FTase, CAAX peptide, and
Farnesyl diphosphate analogue complex.9 Figure 1
shows the proposed binding mode of 1a in the Ras pro-
tein-binding pocket.
Compound
R1
FTase IC50 (nM)
50a
16
N
H
N
In this model, there are three hydrogen bonds. One was
between the O of 3-nitro group and OH of Ser98,
Another was between N of the C-1 pyridine and Zinc
cation of the protein involved water, and last one is be-
tween OH on the C-2 phenyl group and O of Asp359.
This suggests that the direction of the proton in this phe-
nolic hydroxyl group might be important to forming the
hydrogen bond.
24
26
>500
>500
N
N
N
Me
Me
N
25
31
17
44
Binding models of 19b and 22c, with potent FTase
inhibitory activity, are shown in Figure 2. The three
hydrogen bonds, described above, also exist, and impor-
tantly, the direction of the proton in this phenolic
hydroxyl group may be fixed to ASP359 by intermolec-
ular hydrogen bond between 40-OH and 30-NH. As
expected, the substituents on the position are outside
this binding pocket of protein, but the model predicted
that bulky substituents on this position could not be
acceptable.
N
CN
N
N
Table 5. Modification of 3-position on the piperidine
The binding models also suggest that the ortho-position
of the phenolic hydroxyl group may be near to the edge
of the binding pocket and directly outside of the protein.
Some substituents on this position are possible to retain
binding ability because these would be outside this bind-
ing pocket.
X
R3
N
N
3.2. In vitro metabolic stability of C-2 modified piperidine
derivatives
Compound
X
R3
FTase IC50 (nM)
50a
37
41
42
43
38
40
Br
Et
Et
Et
Et
Et
Et
–NO2
–CO2H
16
>500
>500
42
Since 1a was eliminated immediately in mouse in vivo,
in vitro metabolic stability studies of 1a and some
other derivatives using mouse liver microsome were
conducted. Compound 1a was metabolized in mouse
liver microsome mainly as the glucuronide conjugate
and the contribution of P450 to the metabolism was
found to be very small. To assess the contribution of
oxidative metabolism by cytochrome P450, and glucu-
ronidation to the elimination of the compounds,
NADPH and UDPGA, co-enzymes of each metabolic
reaction, were utilized. The residual ratio was calculat-
ed by dividing the concentration of compounds in the
metabolized samples by those in the initial samples.
We checked the metabolic stabilities of several com-
pounds in mouse microsome in the presence of
NADPH and/or UDPGA. Our preliminary results
suggest that the bromo and ethyl group at C-4 phenyl
ring did not greatly affect glucuronidation, therefore we
–CH2OH
–COMe
–CO2Me
95
>500
>500
–CO2CH2CH@CH2
–CONHMe
H
CO2H
CON
SMe
45
Me
19
and 3-methoxycarbonylpiperidine 43 also showed FTase
inhibition (IC50 = 95 nM). Changing the methyl ester to
allyl ester or N-methyl amide resulted in the loss of
FTase inhibitory activity (38 or 40 IC50 > 500 nM).
Interestingly, the L-methionine amide retained potent
inhibition 45 (IC50 = 19 nM). Since the nitropiperidine
inhibitors were shown to be Ras competitive, we

1370
R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
Ser 99 B
OH
Br
O
O
N
N
O
H
N
Water
Zn2+
O
O
Asp 359 B
Figure 1. Proposed binding model of 1a for FTase.
A
Ser 99 B
OH
O
N
O
O
H2N
HN
N
O
H
N
Water
Zn2+
O
O
Asp 359 B
B
Ser 99 B
OH
O
N
O
N
N
HN
O
H
N
Water
O
Zn2+
O
Asp 359 B
Figure 2. (A) Proposed binding model of 19b for FTase. (B) Proposed binding model of 22c for FTase.
used either substituent for the stability tests. As expect-
ed, introduction of a substitution ortho to the phenolic
hydroxy group of C-2 phenyl ring in 1a or 1b resulted
in compounds with reduced propensity for glucuroni-
dation (UDPGA system, uridine 50-diphospho-
glucuronic acid). Although ortho-methoxy compound
48a was still prone to glucuronidation, amino com-
pound 11a (data not shown, ca. 40%), 4-pyridylmeth-
ylamino compound 22c, acetamido compound 12a,
urea compound 19b, and sulfamide compound 20b
were found to be more stable than the simple phenol
compound 1a. In particular, 20b showed the best
stability in assays of the ortho-substituted phenol
compounds (Fig. 3).
Figure 4 shows the metabolic stability of the C-2 phenyl,
thienyl, and pyridyl compounds in the mouse liver
microsome assay. Since there are no phenolic hydroxy

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1371
120
100
80
Y
O2N
N
60
40
HO
X
N
mouse liver microsome : 1 mg/mL
NADPH generating system
UDPGA concentration : 10 mM
Reaction : 30 mim
20
0
Initial concentration : 1 ug/mL
a Each value represents the % remaning.
b NADPH was generated by the mixture of
G-6-P, -NADP+, MgCl2 and G-6-P
dehydrogenase.
1a
48a
2a
Br
22γ
Et
14a
19b
20b
Br
-OMe
180
Y
X
Br
H
5.4
Br
Et
Et
UDPGA
-OH --NHCH2Py -NHSO2Me -NHCONH2 -NHSO2NH2
3.7
12
4.3
6.2
4.5
FTase
UDPGA+NADPHb
IC50 (nM)
Figure 3. Metabolic studies of piperidine derivatives with C-2 substituted phenol parts.
Br
100
O2N
2
80
N
R2
60
N
40
mouse liver microsome : 1 mg/mL
NADPH generating system
UDPGA concentration : 10 mM
20
Reaction : 30 mim
Initial concentration : 1 ug/mL
a Each value represents the % remaning.
b NADPH was generated by the mixture
of G-6-P, -NADP+, MgCl2 and G-6-P
1a
50a
51a
52a
dehydrogenase.
UDPGA
R2
N
UDPGA+NADPHb
S
HO
FTase
5.4
16
36
25
IC50 (nM)
Figure 4. Metabolic studies of C-2 modified piperidine derivatives.
groups in their structures, dramatic improvement in
metabolic stability was observed in each compound.
In order to improve the metabolic stability by steric ef-
fect for glucuronidation, insertion of substituents to the
ortho-position of phenolic hydroxy group was exam-
ined. Some substituents, such as methanesulfonylamino
(14a IC50 = 4.3 nM), urea (19b IC50 = 6.2 nM), and sul-
famoylamino (20a IC50 = 3.0 nM) groups, showed good
metabolic stability in vitro with potent FTase inhibition.
In addition, the simple phenyl compound (50a
Based on the results of in vitro metabolic studies, in vivo
pharmacokinetic and efficacy studies of the selected
compounds such as 20b or 50a are now in progress. De-
tails of these results will be published elsewhere.
IC50 = 16 nM), 3-thiophene compound (51a IC50
=
4. Conclusion
36 nM), and 2-pyridine compound (52a IC50 = 25 nM)
retained the FTase inhibition with good metabolic sta-
bility in vitro. Modifications to the 2-nitro and N-3-pyr-
idylmethyl groups of compound 50a were further
examined in this series. Exchange of the nitro group to
acetyl group (42 IC50 = 42 nM) or to methionine amide
group (45 IC50 = 19 nM) resulted in potent FTase inhib-
itors. In addition, exchange of the N-3-pyridylmethyl
SAR studies around the novel and potent piperidine
inhibitors of FTase, such as 1a and 2a, were carried
out in order to improve their metabolic stability. In vitro
metabolic stability assays suggested that the glucuronide
conjugation of phenolic hydroxy group could be a
primary metabolic pathway.

1372
R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
group to substituted imidazolylmethyl group resulted in
the potent FTase inhibitors, such as 25 (IC50 = 17 nM)
or 31 (IC50 = 44 nM). Further studies of these com-
pounds will be reported in due course.
11.7 Hz, 1H), 3.73 (d, J = 9.4 Hz, 1H), 3.59 (d,
J = 13.9 Hz, 1H), 3.08 (d, J = 13.9 Hz, 1H), 2.90 (m,
1H), 2.39 (m, 1H), 1.89–1.67 (m, 2H). FAB-MS (m/z):
470, 468 (M+H)+.
5.4. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-[3-nitro-4-hy-
droxy]-3-nitro-1-N-(3-pyridylmethyl)-piperidine (10)
5. Experimental
5.1. General methods
To a solution of 1a (300 mg, 0.69 mmol) in acetic acid
(10 mL) was added concentrated nitric acid (0.055 mL,
1.4 mmol) under cooling with ice. After stirred for
1.5 h while elevating the temperature to room tempera-
ture, the reaction solution was poured into water, and
the mixture was neutralized with a dilute aqueous solu-
tion of sodium hydroxide and extracted with a CHCl3/
MeOH, 9:1, mixed solvent. The extract was dried over
sodium sulfate, and the solvent was removed under re-
duced pressure. The resulting residue was chromato-
graphed on silica gel, eluted with CHCl3/MeOH, 98:2,
to afford 10 (220 mg, 62% yield). 1H NMR (CDCl3,
300 MHz) d 10.61 (br s, 1H), 8.51 (dd, J = 4.8, 1.2 Hz,
1H), 8.47 (d, J = 1.7 Hz, 1H), 8.23 (br s, 1H), 7.72 (d,
J = 8.3 Hz, 1H), 7.56–7.53 (m, 2H), 7.37–7.15 (m, 4H),
7.10 (m, 1H), 4.93 (dd, J = 10.8, 9.9 Hz, 1H), 4.08 (m,
1H), 3.90 (d, J = 9.4 Hz, 1H), 3.73 (d, J = 13.6 Hz,
1H), 3.11–3.06 (m, 2H), 2.46 (td, 1H, J = 12.3, 2.0 Hz,
1H), 2.06 (d, J = 11.2 Hz, 1H), 1.71 (m, 1H). FAB-MS
(m/z): 515, 513 (M+H)+.
NMR spectra were recorded on a JEOL Lambda 300
(300 MHz), JEOL GX-270 (270 MHz), JEOL EX-270
(270 MHz) or a JEOL JNM 400 (400 MHz) NMR
spectrometer. Mass spectra were recorded on a JEOL
JMS HX/HX110 mass spectrometer and Micromass
Quattro mass spectrometer.
Elemental analyses were performed on a Perkin-Elmer
Series II Analyzer 2400 elemental analyzer or a Yamato
MT-5 analyzer.
5.2. rac-(4R,5R,6S)-4-(2-Bromophenyl)-6-(4-hydroxy-
phenyl)-5-nitro-1-N-(3-pyridylmethyl)-piperidin-2-one (7)
Methyl 3-(2-bromophenyl)-4-nitrobutylate
4
(61 g,
0.20 mol), 4-hydroxybenzaldehyde 5 (25 g, 0.21 mol),
and 3-(aminomethyl)pyridine 6 (42 mL, 0.41 mol) in
EtOH (1.0 L) were heated under reflux for 20 h. After
removal of EtOH under reduced pressure, the residue
was chromatographed on silica gel, eluted with CHCl3/
MeOH, 19:1, to afford 7 (69 g, 73% yield). Analytically
pure 7 was afforded by recrystallization from hot EtOH.
1H NMR (DMSO-d6, 300 MHz) d 9.62 (s, 1H), 8.38 (d,
J = 4.8 Hz, 1H), 8.11 (s, 1H), 7.78 (d, J = 7.9 Hz, 1H),
7.61 (d, J = 7.9 Hz, 1H), 7.39–7.46 (m, 2H), 7.22–7.27
(m, 2H), 7.14 (d, J = 8.0 Hz, 2H), 6.64 (d, J = 8.0 Hz,
2H), 5.89 (dd, J = 11.0, 9.5 Hz, 1H), 4.94 (d, J =
9.5 Hz, 1H), 4.47 (d, J = 15.8 Hz, 1H), 4.38 (m, 1H),
4.18 (d, J = 15.8 Hz, 1H), 3.05 (dd, J = 16.8, 11.0 Hz,
1H), 2.74 (dd, J = 16.8, 4.7 Hz, 1H). FAB-MS (m/z):
484, 482 (M+H)+. Anal. (C23H20BrN3O4): C, H, N.
(C, 57.27; H, 4.19; N, 8.71%. Found: C, 57.12; H,
4.21; N, 8.67%).
5.5. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(3-amino-4-
hydroxyphenyl)-3-nitro-1-N-(3-pyridylmethyl) piperidine
(11a)
To a solution of the nitro compound 10 (210 mg, 0.41
mmo1) in MeOH (20 mL) was added palladium on car-
bon (21 mg) in a nitrogen atmosphere at room tempera-
ture. After the mixture was refluxed for 5 h, the catalyst
was removed by Celite filtration, and the solvent was
evaporated under reduced pressure. The resulting resi-
due was chromatographed on silica gel, eluted with
CHCl3/MeOH, 19:1, to afford 11a (87 mg, 44% yield).
1H NMR (DMSO-d6, 300 MHz) d 9.14 (br s, 1H), 8.46
(m, 2H), 7.71 (d, J = 6.6 Hz, 1H), 7.65 (d, J = 7.9 Hz,
1H), 7.57 (dd, J = 8.1, 1.1 Hz, 1H), 7.40–7.29 (m, 2H),
7.17 (m, 1H), 6.77 (br s, 1H), 6.63 (d, J = 7.7 Hz, 1H),
6.53 (m, 1H), 5.19 (dd, J = 11.0, 9.6 Hz, 1H), 4.60 (br
s, 2H), 3.86 (m, 1H), 3.67 (d, J = 13.8 Hz, 1H), 3.56
(d, J = 9.1 Hz, 1H), 3.04 (d, J = 13.8 Hz, 1H), 2.87
(dd, J = 11.6, 3.3 Hz, 1H), 2.34 (m, 1H), 1.90 (m, 1H),
1.73 (m, 1H). FAB-MS (m/z): 485, 483 (M+H)+.
5.3. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(4-hydroxyphe-
nyl)-3-nitro-1-N-(3-pyridylmethyl)-piperidine (1a)
To a solution of 7 (40 g, 83 mmol) in THF (2.0 L)
was added borane–methyl sulfide complex (30 mL,
320 mmol) and the mixture was refluxed for 10 h. After
cooling to 4 ꢁC, the reaction was quenched by the addi-
tion of water (50 mL) and the mixture was concentrated.
The resulting residue was heated with aqueous HCl
(3.0 mol/L, 400 mL) and neutralized with aqueous
NaOH (3.5 mol/L, 300 mL) after cooling to room
temperature. The solid formed was recrystallized from
EtOH to give analytically pure 1a (35 g, 90% yield).
mp 229–230 ꢁC. IR (KBr) 2823, 1614, 1560, 1554,
5.6. rac-Aceto-5-[{(3R,4R)-4-(2-bromophenyl)-3-nitro-
1-N-(3-pyridylmethyl)piperidin}-(2S)-2-yl]-2-hydroxy-
anilide (12a)
To a solution of 11a (24 mg, 0.050 mmol) in CH2Cl2
(5.0 mL) was added acetyl chloride (0.0036 mL,
0.050 mol) at room temperature. After stirred for 1 h,
the reaction solution was poured into water and the mix-
ture was extracted with CHCl3/MeOH, 9:1. The extract
was dried over sodium sulfate, and the solvent was evap-
orated under reduced pressure. The resulting residue
was chromatographed on silica gel, eluted with CHCl3,
1473, 1373, 1280, 1250, 1022, 761 cmꢀ1 1H NMR
.
(DMSO-d6, 300 MHz) d 9.53 (s, 1H), 8.44 (m, 1H),
8.16 (br s, 1H), 7.73 (m, 1H), 7.65–7.56 (m, 2H), 7.40–
7.31 (m, 4H), 7.18 (m, 1H), 6.78 (br s, 1H), 6.75 (br s,
1H), 5.34 (dd, J = 9.4, 11.7 Hz, 1H), 3.87 (dt, J = 4.0,

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1373
and re-precipitated in diethyl ether/hexane to afford 12a
(7.2 mg, 13% yield). 1H NMR (CDCl3, 300 MHz) d
8.51–8.49 (m, 2H), 7.97 (m, 1H) 7.69 (d, J = 7.9 Hz,
1H), 7.53 (d, J = 9.0 Hz, 1H), 7.38–7.11 (m, 4H), 7.10
(d, J = 7.3 Hz, 1H), 7.01 (d, J = 7.5 Hz, 1H), 4.97 (dd,
J = 10.8, 8.8 Hz, 1H), 4.05 (m, 1H), 3.80 (d,
J = 13.6 Hz, 1H), 3.76 (d, J = 8.8 Hz, 1H), 3.07-3.05
(m, 2H), 2.42 (m, 1H), 2.27 (s, 3H), 2.03 (d, J =
15.2 Hz, 1H), 1.80 (m, 1H). FAB-MS (m/z): 527, 525
(M+H)+.
(0.0094 mL, 0.10 mmol) and pyridine (0.10 mL) at room
temperature. After stirred at room temperature for
30 min, the reaction solution was poured into water
and the mixture was extracted with CHCl3/MeOH,
9:1. The extract was dried over sodium sulfate, and the
solvent was evaporated under reduced pressure. The
resulting residue was chromatographed on silica gel,
eluted with CHCl3, re-precipitated in diethyl ether/hex-
ane to afford 15a (2.3 mg, 8.0% yield). 1H NMR
(DMSO-d6, 270 MHz) d 9.04 (s, 1H), 8.71 (s, 1H) 8.56
(d, J = 4.6 Hz, 1H), 8.44–8.30 (m, 2H), 7.75 (d,
J = 8.9 Hz, 1H), 7.68 (d, J = 7.6 Hz, 1H), 7.55 (d,
J = 7.9 Hz, 1H), 7.49 (m, 1H), 7.38–7.29 (m, 2H), 7.16
(m, 1H), 6.83 (d, J = 8.3 Hz, 1H), 5.27 (dd, J = 10.6,
9.3 Hz, 1H), 3.86 (m, 1H), 3.70 (d, J = 9.2 Hz, 1H),
3.60 (d, J = 13.5 Hz, 1H), 3.05 (d, J = 13.5 Hz, 1H),
3.00 (q, J = 7.4 Hz, 2H), 2.86 (d, J = 12.2 Hz, 1H),
2.33 (m, 1H), 1.80–1.70 (m, 2H), 1.18 (t, J = 7.4 Hz,
3H). FAB-MS (m/z): 577, 575 (M+H)+.
5.7. rac-N-(5-[{(3R,4R)-4-(2-Bromophenyl)-3-nitro-1-N-
(3-pyridylmethyl)piperidin}-(2S)-2-yl]-2-hydroxyphenyl)-
N0-methyl-carbamate (13a)
To a solution of 11a (48 mg, 0.10 mmol) in CH2Cl2/
DMF (2:1, 3.0 mL) was added methyl chlorocarbonate
(0.0036 mL, 0.050 mmol) at room temperature. After
stirred for 30 min, the reaction solution was poured into
water and the mixture was extracted with a CHCl3/
MeOH, 9:1. The extract was dried over sodium sulfate,
and the solvent was evaporated under reduced pressure.
The resulting residue was chromatographed on silica gel,
eluted with CHCl3, re-precipitated in diethyl ether/hex-
5.10. rac-N-(5-{[(3R,4R)-4-(2-Bromophenyl)-3-nitro-1-N-
(3-pyridylmethyl)piperidin]-(2S)-2-yl}-2- hydroxyphenyl)
p-toluenesulfonamide (16a)
1
ane to afford 13a (12 mg, 21% yield). H NMR (CDCl3,
To a solution of 11a (24 mg, 0.050 mmol) in CH2Cl2
(2.0 mL) was added p-toluenesulfonyl chloride (9.5 mg,
0.050 mmol) at room temperature. After stirred at room
temperature for 2 h, the reaction solution was poured
into water and the mixture was extracted with CHCl3/
MeOH, 9:1. The extract was dried over sodium sulfate,
and the solvent was evaporated under reduced pressure.
The resulting residue was purified by preparative silica
plate with CHCl3/MeOH, 9:1, to afford 16a (9.5 mg,
300 MHz) d 8.51 (br s, 1H), 8.47 (d, J = 3.3 Hz, 1H),
7.67 (d, J = 7.7 Hz, 1H), 7.52 (d, J = 7.7 Hz, 1H), 7.35
(d, J = 7.3 Hz, 1H), 7.27–7.23 (m, 3H), 7.08 (d,
J = 7.2 Hz, 1H), 7.04 (br s, 1H), 6.85 (d, J = 7.3 Hz,
1H), 5.01 (dd, J = 10.8, 8.8 Hz, 1H), 4.04 (m, 1H),
3.94–3.76 (m, 5H), 3.13 (d, J = 13.6 Hz, 1H), 3.02 (d,
J = 12.3 Hz, 1H), 2.47 (m, 1H), 2.03 (d, J = 12.3 Hz,
1H), 1.69 (m, 1H). FAB-MS (m/z): 543, 541 (M+H)+.
1
30% yield). H NMR (DMSO-d6, 270 MHz) d 9.73 (br
5.8. rac-N-([5-{(3R,4R)-4-(2-Bromophenyl)-3-nitro-1-N-
(3-pyridylmethyl)piperidin}-(2S)-2-yl]-2-hydroxyphenyl)
methanesulfonamide (14a)
s, 1H), 9.11 (br s, 1H), 8.46 (br s, 1H), 7.72 (m, 1H),
7.56 (d, J = 4.0 Hz, 1H), 7.37–7.33 (m, 2H), 7.18 (d,
J = 8.2 Hz, 4H), 7.20–7.13 (m, 3H), 6.66 (d,
J = 7.9 Hz, 1H), 5.20 (dd, J = 10.9, 10.2 Hz, 1H), 3.87
(m, 1H), 3.67 (d, J = 9.2 Hz, 1H), 3.50 (d, J = 14.2 Hz,
1H), 3.04 (d, J = 14.2 Hz, 1H), 2.86 (d, J = 10.9 Hz,
1H), 2.41 (m, 1H), 2.24 (s, 3H), 1.82–1.70 (m, 2H).
FAB-MS (m/z): 638, 636 (M+H)+.
To a solution of 11a (48 mg, 0.10 mmol) in CH2Cl2/DMF
(2:1, 3.0 mL) was added methanesulfonyl chloride
(0.0077 mL, 0.10 mmol) at room temperature. After stir-
red for 30 min, the reaction solution was poured into
water and the mixture was extracted with CHCl3/MeOH,
9:1. The extract was dried over sodium sulfate, and the
solvent was evaporated under reduced pressure. The
resulting residue was chromatographed on silica gel, elut-
ed with CHCl3, re-precipitated in diethyl ether/hexane to
afford 14a (8.9 mg, 16% yield). 1H NMR (CDCl3,
300 MHz) d 8.54 (br s, 1H), 8.45 (d, J = 4.0 Hz, 1H),
7.69 (d, J = 7.6 Hz, 1H), 7.69 (m, 1H), 7.53 (d,
J = 8.1 Hz, 1H), 7.34–7.21 (m, 4H), 7.08 (d, J = 7.0 Hz,
1H), 6.92 (d, J = 8.1 Hz, 1H), 4.98 (dd, J = 10.7, 9.8 Hz,
IH), 4.06 (m, 1H), 3.90 (d, J = 14.1 Hz, 1H), 3.82 (d,
J = 9.0 Hz, 1H), 3.18 (d, J = 14.1 Hz, 1H), 3.05 (dd,
J = 13.4,l.6 Hz, 1H), 2.51 (m, 1H), 2.04 (m, 1H), 1.76
(m, 1H), 1.43 (s, 3H). FAB-MS (m/z): 563, 561 (M+H)+.
5.11. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(3-ethylami-
no-4-hydroxyphenyl)-3-nitro-1-N-(3-pyridylmethyl)piper-
idine (17a)
To a suspension of lithium aluminum hydride (20 mg,
0.50 mmol) in THF (100 mL) was added an ice-cold
solution (10 mL) of 12a (52 mg, 0.10 mmol). After stir-
red for 12 h while elevating the temperature to room
temperature, the reaction mixture was poured into dilute
aqueous HCl, followed by stirring for 10 min. The mix-
ture was neutralized with saturated aqueous NaHCO3
and extracted with CHCl3/MeOH, 9:1. The extract was
dried over sodium sulfate, and the solvent was evaporat-
ed under reduced pressure. The resulting residue was
purified by preparative silica plate with CHCl3 and re-
precipitated in diethyl ether/hexane to afford 17a
5.9. rac-N-([5-{(3R,4R)-4-(2-Bromophenyl)-3-nitro-1-N-
(3-pyridylmethyl)piperidin}-(2S)-2-yl]-2-hydroxyphenyl)
ethanesulfonamide (15a)
1
(15 mg, 31% yield). H NMR (DMSO-d6, 270 MHz) d
9.45 (br s, 1H), 8.65 (br s, 1H), 8.48 (dd, J = 4.8,
1.5 Hz, 1H), 7.80 (br s, 1H), 7.59–7.52 (m, 2H),
7.37–7.06 (m, 7H), 4.99 (dd, J = 10.8, 9.9 Hz, 1H),
To a solution of 11a (24 mg, 0.05 mmol) in CH2Cl2
(10 mL)
were
added
ethanesulfonyl
chloride

1374
R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
4.05 (m, 1H), 3.89–3.83 (m, 2H), 3.08–3.03 (m, 2H), 2.45
(m, 1H), 2.04 (m, 1H), 1.66 (m, 1H). FAB-MS (m/z):
513, 511 (M+H)+.
water, and the powder formed was collected by filtra-
tion. The resulting crude product was chromatographed
on silica gel, eluted with chloroform/methanol, 98:2, and
re-precipitated in diethyl ether/hexane to obtain 20a
1
5.12. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(3-isopropyla-
mino-4-hydroxyphenyl)-5-nitro-1-N-(3-pyridylmethyl)pi-
peridine (18a)
(23 mg, 41% yield). H NMR (DMSO-d6, 270 MHz) d
9.87 (br s, 1H), 8.45 (br s, 2H), 7.87 (br s, 1H), 7.75
(d, J = 6.9 Hz, 1H), 7.68 (m, 1H), 7.62–7.59 (m, 2H),
7.39 (m, 1H), 7.31 (m, 1H), 7.21–7.18 (m, 3H), 6.98
(br s, 1H), 6.79 (d, J = 7.9 Hz, 1H), 5.38 (dd, J = 10.9,
9.6 Hz, 1H), 3.87 (m, 1H), 3.69 (d, J = 9.6 Hz, 1H),
3.65 (d, J = 14.2 Hz, 1H), 3.03 (d, J = 14.2 Hz, 1H),
2.88 (m, 1H), 2.37 (m, 1H), 1.88 (m, 1H), 1.65 (m,
1H). FAB-MS (m/z): 564, 562 (M+H)+.
To a solution of 11a (48 mg, 0.1 mmo1) in THF (10 mL)
were added acetone (0.015 mL, 0.20 mmol) and borane–
methyl sulfide complex (0.010 mL, 0.11 mmol) at room
temperature. After stirred for 12 h, the solvent was evap-
orated under reduced pressure. The resulting residue
was dissolved in MeOH (10 mL), and to the solution
was added aqueous HCl (1.0 mol/L, 2.0 mL). After stir-
red at 50 ꢁC for 1 h, the mixture was neutralized with
saturated aqueous NaHCO3 and extracted with CHCl3.
The extract was dried over sodium sulfate, and the sol-
vent was evaporated under reduced pressure. The result-
ing residue was chromatographed on silica gel, eluted
with CHCl3/MeOH, 9:1, to afford 18a (24 mg, 46%
yield). 1H NMR (CDCl3, 270 MHz) d 8.52 (s, 1H),
8.47 (d, J = 6.9 Hz, 1H), 7.75 (d, J = 8.6 Hz, 1H), 7.53
(d, J = 7.6 Hz, 1H), 7.42–7.28 (m, 3H), 7.13 (m, 1H),
6.74–6.64 (m, 3H), 4.99 (dd, J = 10.9, 9.5 Hz, 1H),
4.11 (m, 1H), 3.90 (d, J = 14.2 Hz, 1H), 3.74 (d,
J = 9.5 Hz, 1H), 3.63 (m, 1H), 2.96 (m, 2H), 2.47 (m,
1H), 2.05 (m, 1H), 1.73 (m, 1H), 1.25 (d, J = 6.2 Hz,
3H), 1.17 (d, J = 6.2 Hz, 3H). FAB-MS (m/z): 527, 525
(M+H)+.
5.15. rac-(2R,3S,4S)-4-(2-Ethylphenyl)-2-(3-benzylami-
no-4-hydroxyphenyl)-3-nitro-1-N-(3-pyridylmethyl)piper-
idine (21b)
To a solution of 11b (48 mg, 0.10 mmol) in THF (10 mL)
were added benzaldehyde (0.010 mL, 0.10 mmol), and
sodium triacetoxyborane hydride (210 mg, 1.0 mmol) at
room temperature. After stirred for 12 h, the reaction
mixture was poured into water, and the mixture was neu-
tralized with saturated aqueous NaHCO3 and extracted
with CHCl3. The extract was dried over sodium sulfate,
and the solvent was evaporated under reduced pressure.
The resulting residue was chromatographed on silica gel,
eluted with CHCl3/MeOH, 9:1, to afford 21b (16 mg,
1
61% yield). H NMR (CDCl3, 270 MHz) d 8.55 (br s,
1H), 8.45 (d, J = 4.0 Hz, 1H), 7.50 (d, J = 7.9 Hz, 1H),
7.43–7.11 (m, 10H), 6.87–6.66 (m, 3H), 4.92 (dd,
J = 10.5, 9.9 Hz, 1H), 4.38 (s, 2H), 3.85 (d, J = 13.5 Hz,
1H), 3.67–3.64 (m, 2H), 3.06–2.97 (m, 2H), 2.80–2.52
(m, 2H), 2.37 (m, 1H), 1.90–1.85 (m, 2H), 1.18 (t,
J = 7.6 Hz, 3H). FAB-MS (m/z): 523 (M+H)+.
5.13. rac-N-([5-{(3R,4R)-4-(2-Bromophenyl)-3-nitro-1-N-
(3-pyridylmethyl)piperidin}-(2S)-2-yl]-2-hydroxyphenyl)
urea (19a)
To a solution of 11a (96 mg, 0.20 mmol) in THF
(10 mL) were added acetic acid (1.0 mL) and potassium
cyanate (160 mg, 2.0 mmol) at room temperature. After
stirred for 1 h, the reaction solution was poured into
water and the mixture was extracted with CHCl3/
MeOH, 9:1. The extract was dried over sodium sulfate,
and the solvent was evaporated under reduced pressure.
The resulting residue was chromatographed on silica gel,
eluted with CHCl3, and re-precipitated in diethyl
ether/hexane to afford 19a (21 mg, 20% yield). 1H
NMR (DMSO-d6, 270 MHz) d 10.08 (s, 1H), 8.48 (d,
J = 1.3 Hz, 1H), 8.44 (dd, J = 4.8, 1.4 Hz, 1H), 8.12
(br s, 1H), 8.03 (s, 1H), 7.76–7.71 (m, 2H), 7.57 (dd,
J = 7.9, 1.0 Hz, 1H), 7.39–7.29 (m, 2H), 7.17 (m, 1H),
6.89 (br s, 1H), 6.67 (d, J = 7.9 Hz, 1H), 6.24 (br s,
2H), 5.19 (dd, J = 10.9, 9.9 Hz, 1H), 3.88 (m, 1H),
3.69–3.65 (m, 2H), 3.04 (d, J = 13.5 Hz, 1H), 2.95 (m,
1H), 2.38 (m, 1H), 1.88–1.68 (m, 2H). FAB-MS (m/z):
528, 526 (M+H)+.
5.16. rac-(2R,3S,4S)-4-(2-Ethylphenyl)-2-[3-(2-pyridyl)
methylamino-4-hydroxyphenyl]-3-nitro-1-N-(3-pyridyl-
methyl)piperidine (22a)
According to the procedure for the synthesis of 21b,
11b (48 mg, 0.10 mmol), 2-pyridinecarboxaldehyde
(0.010 mL, 0.10 mmol), and sodium triacetoxyborane
hydride (210 mg, 1.0 mmol) afforded 22a (9.7 mg, 37%
yield). ESI-MS (m/z): 524 (M+H)+.
5.17. rac-(2R,3S,4S)-4-(2-Ethylphenyl)-2-[3-(3-pyridyl)
methylamino-4-hydroxyphenyl]-3-nitro-1-N-(3-pyridyl-
methyl)piperidine (22b)
According to the procedure for the synthesis of 21b, 11b
(48 mg,
0.10 mmol),
4-pyridinecarboxaldehyde
(0.010 mL, 0.10 mmol) and sodium triacetoxyborane
hydride (210 mg, 1.0 mmol) afforded 22b (20 mg, 78%
yield). ESI-MS (m/z): 524 (M+H)+.
5.14. rac-N-([5-{(3R,4R)-4-(2-Bromophenyl)-3-nitro-1-N-
(3-pyridylmethyl)piperidin}-(2S)-2-yl]-2-hydroxyphenyl)
sulfamide (20a)
5.18. rac-(2R,3S,4S)-4-(2-Ethylphenyl)-2-[3-(4-pyridyl)
methylamino-4-hydroxyphenyl]-3-nitro-1-N-(3-pyridyl-
methyl)piperidine (22c)
To a solution of 11a (48 mg, 0.10 mmol) in N,N-dime-
thylacetamide solution (1.0 mL) was added sulfamoyl
chloride (12 mg, 0.10 mmol) at room temperature. After
stirred for 15 min, the reaction solution was poured into
According to the procedure for the synthesis of 21b,
11b (48 mg, 0.10 mmol), 4-pyridinecarboxaldehyde
(0.010 mL, 0.10 mmol), and sodium triacetoxyborane

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1375
hydride (210 mg, 1.0 mmol) afforded 22c (9.4 mg, 36%
yield). ESI-MS (m/z): 524 (M+H)+.
(m, 1H), 6.88 (br s, 1H), 5.03 (dd, J = 10.9, 9.6 Hz,
1H), 4.05 (m, 1H), 3.73 (d, J = 9.6 Hz, 1H), 3.63(s,
3H), 3.55 (d, J = 14.0 Hz, 1H), 3.14–3.02 (m, 2H), 2.27
(m, 1H), 2.05 (m, 1H), 1.61 (m, 1H). ESI-MS (m/z):
5.19. rac-(2R,3S,4S)-4-(2-Bromophenyl)-3-nitro-2-phen-
yl-piperidine (23)
1
458, 456 (M+H)+. Compound 26: H NMR (CDCl3,
270 MHz) d 8.02 (br s, 1H), 7.52 (dd, J = 7.9, 1.0 Hz,
1H), 7.43–7.24 (m, 7H), 7.06 (m, 1H), 6.58 (br s, 1H),
4.97 (dd, J = 11.2, 9.6 Hz, 1H), 3.99 (m, 1H), 3.79 (d,
J = 9.6 Hz, 1H), 3.65 (s, 3H), 3.62 (d, J = 14.0 Hz,
1H), 3.36–3.24 (m, 2H), 2.61 (m, 1H), 2.04 (m, 1H),
1.78 (m, 1H). FAB-MS (m/z): 458, 456 (M+H)+.
Step 1 Methyl 3-(2-bromophenyl)-4-nitrobutyrate (1.8 g,
6.0 mmol), benzaldehyde (0.61 mL, 6.0 mmol), and
ammonium acetate (920 mg, 12 mmol) were heated un-
der reflux in ethanol for 20 h. The solvent was evaporat-
ed under reduced pressure, and the resulting residue was
chromatographed on silica gel, eluted with CHCl3/
MeOH, 98:2, to afford a piperidin-2-one derivative
(1.4 g, 57% yield). ESI-MS (m/z): 440 (M+H)+. Step 2
According to the procedure for the synthesis of 1a, the
resulting piperidin-2-one derivative and borane-methyl
sulfide complex (2.7 mL, 30 mmol) afforded 23
(890 mg, 41% yield). 1H NMR (CDCl3, 270 MHz) d
8.62 (d, J = 2.6 Hz, 1H), 7.56 (d, J = 8.2 Hz, 1H),
7.37–7.21 (m, 5H), 7.22 (d, J = 7.6 Hz, 1H), 7.13–7.08
(m, 2H), 6.27 (d, J = 5.6 Hz, 1H), 5.56 (m, 1H), 4.40
(m, 1H), 4.20 (m, 1H), 3.83 (m, 1H), 2.45 (m, 1H),
1.92 (m, 1H). ESI-MS (m/z): 426 (M+H)+.
5.22. 1-Trityl-1H-imidazol-4-ylmethyl acetate (28)
Step 1 To a solution of 4-(hydroxymethyl)-imidazole
hydrochloride salt (880 mg, 6.5 mmol) in DMF (20 mL)
was added triethylamine (2.3 mL, 16.0 mmol) which
immediately caused a white precipitate to form. After
stirred for 10 min, a solution of chlorotriphenylmethane
(2.0 g, 7.2 mmol) in DMF (15 mL) was added dropwise.
After stirred overnight under a nitrogen atmosphere, the
reaction mixture was poured into ice water and the mix-
ture was filtered. The solid was washed with cold dioxane
and dried under vacuum to afford (1-trityl-1H-imidazol-
4-yl)-methanol as a white powder (2.2 g, 100% yield). 1H
NMR (CD3CO2D, 250 MHz) d 8.56 (d, 1H), 7.57–7.40
(m, 7H), 7.38–7.18 (m, 9H), 4.78 (s, 2H). EI-MS (m/z):
363 (M+Na)+. Step 2 To a suspension of (1-trityl-1H-
imidazol-4-yl)-methanol (2.3 g, 6.5 mmol) in pyridine
(15 mL) was added acetic anhydride (2.0 mL, 20 mmol)
in 5 portions at room temperature over 30 min. After stir-
red under a nitrogen atmosphere overnight, the reaction
mixture had become homogeneous and was diluted with
EtOAc (30 mL) and this mixture was washed with water,
aqueous HCl (5%), and saturated aqueous NaHCO3.
The organic layer was then dried over MgSO4 and the
solvent evaporated to afford 28 (2.3 g, 93% yield) as a
white solid. 1H NMR (CDCl3, 250 MHz) d 8.61 (d,
1H), 7.38–7.26 (m, 9H), 7.18–7.07 (m, 7H), 5.05 (s,
2H), 2.07 (s, 3H). EI-MS (m/z): 405 (M+Na)+.
5.20. rac-(2R,3S,4S)-4-(2-Bromophenyl)-1-(3H-imidazol-
4-ylmethyl)-3-nitro-2-phenyl-piperidine (24)
To a solution of 23 (720 mg, 2.0 mmol) in acetic
acid (5.0 mL) were added 4-imidazolecarboxaldehyde
(190 mg, 2.0 mmol) and sodium triacetoxyborohydride
(2.1 g, 10 mmol) at room temperature. After stirred for
12 h, the reaction solution was poured into water, and
the mixture was neutralized with saturated aqueous NaH-
CO3 and extracted with CHCl3. The extract was dried
over sodium sulfate, and the solvent was evaporated
under reduced pressure. The resulting residue was chro-
matographed on silica gel, eluted with CHCl3/MeOH,
9:1, to afford 24 (360 mg, 41% yield). 1H NMR (CDCl3,
270 MHz) d 7.71 (br s, 1H), 7.52 (d, J = 7.9 Hz, 1H),
7.45–7.26 (m, 7H), 7.07 (m, 1H), 6.82 (br s, 1H), 6.01
(br s, 1H), 4.98 (dd, J = 11.2, 9.9 Hz, 1H), 3.97 (m, 1H),
3.78 (d, J = 9.3 Hz, 1H), 3.65 (d, J = 14.5 Hz, 1H), 3.28
(d, J = 14.5 Hz, 1H), 3.17 (m, 1H), 2.53 (m, 1H), 2.05
(m, 1H), 1.75 (m, 1H). ESI-MS (m/z): 444, 442 (M+H)+.
5.23. 3-(4-Cyano-benzyl)-3H-imidazol-4-ylmethyl acetate
(29)
To a solution of 28 (2.3 g, 6.1 mmol) in EtOAc (20 mL)
was added a-bromo-p-tolunitrile (1.30 g, 6.70 mmol) at
room temperature and heated at 60 ꢁC. After stirred at
60 ꢁC overnight, a white precipitate had formed. The
reaction mixture was filtered and the solid material dis-
solved in MeOH (20 mL) and heated at 60 ꢁC, again.
After stirred at 60 ꢁC for 2 h, the reaction mixture was
cooled and the solvent evaporated. The remaining solid
was triturated with hexane to afford 29 (1.38 g, 68%
yield) as a white powder. 1H NMR (250 MHz, CD3OD)
d 7.80–7.75 (m, 2H), 7.75–7.62 (m, 1H), 7.56–7.47 (m,
2H), 7.30–7.23 (m, 1H), 5.72 (s, 2H), 5.20 (s, 2H), 1.90
(s, 3H). EI-MS (m/z): 256 (M+H)+.
5.21. rac-(2R,3S,4S)-4-(2-Bromophenyl)-1-[(1-N-meth-
ylimidazol)-4-yl-methyl]-3-nitro-2-phenyl-piperidine (25)
and rac-(2R,3S,4S)-4-(2-bromophenyl)-1-[(3-N-meth-
ylimidazol)-4-yl-methyl]-3-nitro-2-phenyl-piperidine (26)
To an ice-cold solution of 24 (160 mg, 0.37 mmol) in
DMF (5.0 mL) were added methyl iodide (0.023 mL,
0.37 mmol) and potassium carbonate (50 mg,
0.36 mmol). After the mixture was stirred for 1 h, the
reaction solution was poured into water and the mixture
was extracted with CHCl3. The extract was dried over
sodium sulfate, and the solvent was evaporated under
reduced pressure. The resulting residue was purified by
preparative silica plate with CHCl3/MeOH, 9:1, to af-
ford 25 (6.7 mg, 4.2% yield), 26 (19 mg, 12% yield),
and a mixture of 25 and 26 (110 mg, 69% yield). Com-
5.24. 4-(5-Hydroxymethyl-imidazol-1-ylmethyl)-benzo-
nitrile (30)
1
pound 25: H NMR (CDCl3, 270 MHz) d 8.00 (br s,
1H), 7.53 (d, J = 8.3 Hz, 1H), 7.43–7.24 (m, 7H), 7.08
To a solution of 29 (1.4 g, 4.1 mmol) in THF/water (3:1,
20 mL) was added lithium hydroxide (520 mg, 12 mmol)

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R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
at room temperature. After stirred for 1 h, the reaction
mixture was diluted with EtOAc (30 mL) and washed
with water, saturated aqueous NaHCO3, and brine.
The organic layer was dried over sodium sulfate, and
the solvent was evaporated under reduced pressure to
afford 30 (530 mg, 60% yield) as a yellow brown solid.
1H NMR (CD3OD, 250 MHz) d 7.68–7.55 (m, 3H),
7.22 (d, 2H), 6.89 (s, 1H), 5.30 (s, 2H), 4.76 (s, 2H),
4.34 (s, 1H). EI-MS (m/z): 213 (M+)+.
5.27. 2-Ethyl-cinnamaldehyde (33)
To a suspension of NaHCO3 (63 g, 0.75 mol) and
tetrabutylammonium chloride hydrate (83 g, 0.30 mol)
in DMF (300 mL), 1-ethyl-2-iodobenzene (43 mL,
0.30 mol) and acrolein (47 mL, 0.60 mol) were added
and the mixture was stirred for 15 min. Palladium ace-
tate (3.4 g, 15 mmol) was added and the mixture was
stirred under nitrogen for 2 days. Additional palladium
acetate (2.0 g, 8.8 mmol) was added at 23 h. An aque-
ous extraction with EtOAc failed to give separable lay-
ers and the organic/aqueous mixture was evaporated.
The resulting residue was loaded directly onto a silica
plug and eluted with hexane/CH2Cl2, 1:1 to 0:1. The
recovered product was dissolved in EtOAc (800 mL)
and the solution was washed with water. The organic
layer was dried over sodium sulfate and evaporated un-
der reduced pressure. The resulting residue was chro-
matographed on silica gel, eluted with hexane/
CH2Cl2, 1:1, to afford 33 (6.1 g, 13% yield). 1H
NMR (CDCl3, 400 MHz) d 9.70 (d, J = 7.3 Hz, 1H),
7.77 (d, J = 16.0 Hz, 1H), 7.56 (d, J = 8.3 Hz, 1H),
7.33 (m, 1H), 7.22 (m, 2H), 6.64 (dd, J = 15.6,
7.8 Hz, 1H), 2.78 (q, J = 7.6, 15.0 Hz, 2H), 1.21 (t,
J = 7.6 Hz, 3H).
5.25. rac-4-(5-{[(2R,3S,4S)-4-(2-Bromophenyl)-3-nitro-2-
phenyl-piperidin]-1-ylmethyl}-imidazol-1-ylmethyl)
benzonitrile (31)
To a solution of 4-(5-hydroxymethyl-imidazol-1-ylmeth-
yl)-benzonitrile 30 (130 mg, 0.61 mmol) in DMSO
(5.0 mL), triethylamine (0.34 mL, 2.4 mmol) was added
followed by the sulfur trioxide pyridine complex
(240 mg, 1.5 mmol) at room temperature under a nitro-
gen atmosphere. After stirred for 40 min, the reaction
mixture was diluted with EtOAc (20 mL) and washed
with water and saturated aqueous NaHCO3. The organ-
ic layer was dried over sodium sulfate, and the solvent
was evaporated under reduced pressure. This residue
was then dissolved in CH2Cl2 (2.0 mL) and cooled to
0 ꢁC under a nitrogen atmosphere. To this solution
was added (220 mg, 0.61 mmol) along with sodium tri-
acetoxyborohydride (190 mg, 0.92 mmol) and the reac-
tion mixture left to warm to room temperature
overnight. After this time the reaction mixture was dilut-
ed in EtOAc (20 mL) and washed with saturated aque-
ous NaHCO3 and brine. The organic layer was dried
over sodium sulfate, and the solvent was evaporated un-
der reduced pressure. This residue was chromato-
graphed on silica gel, to afford 31 (6.4 mg, 1.9% yield).
1H NMR (CD3OD, 250 MHz) d 7.58 (t, 3H); 7.45 (d,
1H), 7.38–7.10 (m, 7H), 7.04 (dt, 1H), 6.95 (d, 2H),
6.81 (s, 1H), 4.78 (s, 4H), 3.91 (td, 1H), 3.56 (d, 1H),
2.92 (dt, 1H), 2.21 (td, 2H), 1.55–1.32 (m, 2H). EI-MS
(m/z): 558, 556 (M+H)+, EI-MS (m/z): 556, 554
(MꢀH)ꢀ.
5.28. rac-3-Allyl [4-(2-ethylphenyl)-2-phenyl-1-N-(3-pyr-
idylmethyl)-2,3-dehydro-piperidin]-3-ylcarboxylate (35)
Step 1 Proline lithium salt (0.26 g, 2.2 mmol) was placed
in a 2-necked flask and evacuated and flushed with
nitrogen 4·. Allyl benzoyl acetate 32 (13.2 g, 65 mmol)
in CHCl3 (10 mL) was transferred to the reaction flask
by cannula. The aldehyde 33 (6.9 g, 43 mmol) in CHCl3
(10 mL) was transferred to the reaction flask by cannula.
Total CHCl3 used was 65 mL. After all reagents and sol-
vent were added to the flask, a subsurface nitrogen
purge was maintained for 15 min. After stirred under
nitrogen for 2 days, the reaction was quenched with
aqueous HCl (2.0 mol/L). The product was extracted
with CH2Cl2. The extracts were washed with brine, dried
over sodium sulfate, and evaporated under reduced
pressure. The resulting residue was chromatographed
on silica gel, eluted with hexane/EtOAc, 9:1, to provide
a mixture of the product isomers 34 with impurities
(9.9 g). Step 2 To an ice-cold mixture of 3-(aminometh-
yl)pyridine (3.0 g, 27 mmol), sodium triacetoxyborohy-
dride (12 g, 54 mmol), and acetic acid (0.80 mL,
14 mmol) in CH2Cl2 (65 mL) was added the aldehyde
34 (9.9 g, 27.0 mmol) in CH2Cl2 (15 mL) dropwise over
1.3 h. After stirred at 0 ꢁC for an additional 30 min, the
ice bath was removed and the reaction mixture was
stirred at room temperature, overnight. The reaction
mixture was washed with saturated aqueous NaHCO3.
The organic layer was dried over sodium sulfate and
evaporated under reduced pressure. The resulting resi-
due was chromatographed on silica gel, eluted with
CH2Cl2/EtOAc, 9:1, to afford 35 (5.9 g, 31% yield over
5.26. 3-Allyl benzoylacetate (32)
Ethyl benzoylacetate (50 mL, 0.29 mol), allyl alcohol
(300 mL, 4.4 mol), and dimethylaminopyridine (11 g,
0.090 mol) were mixed and heated at reflux for 3 days.
The solution was evaporated and NMR indicated
approximately 15% starting material remained. Allyl
alcohol (300 mL, 4.4 mol) was added and the solution
was refluxed for 18 h. The solution was evaporated
and no starting material was observed by NMR.
The resulting residue was dissolved in EtOAc and
the solution washed with saturated ammonium chlo-
ride solution until all dimethylaminopyridine was re-
moved. The organic layer was dried over sodium
sulfate and evaporated under reduced pressure. Com-
pound 32 (32 g, 54% yield) was isolated by vacuum
1
1
distillation (0.02 torr, 130–135 ꢁC). H NMR (CDCl3,
2 steps). H NMR (CDCl3, 250 MHz): d 8.43 (s, 1H),
400 MHz) d 7.90 (d, J = 7.8 Hz, 2H), 7.55 (m, 1H),
7.40 (m, 2H), 5.85 (m, 1H), 5.26 (d, J = 17.0 Hz,
1H), 5.18 (d, J = 10 Hz, 1H), 4.61 (d, J = 5.9 Hz,
2H), 3.99 (s, 2H).
8.34 (s, 1H), 7.44–7.05 (m, 13H), 5.25 (m, 1H), 4.77
(dd, J = 11.0, 1.5 Hz, 1H), 4.70 (dd, J = 17.0, 1.6 Hz,
1H), 4.47 (d, J = 5.5 Hz, 1H), 4.14–3.97 (m, 4 H), 3.09
(dt, J = 13.0, 3.1 Hz, 1H), 2.90 (m, 1H), 2.80 (qn, 1H),

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1377
2.63 (qn, 1H), 1.96 (m, 1H), 1.65 (m, 1H), 1.24
oil. The oil was dissolved in EtOAc (25 mL) and the
products extracted into aqueous HCl (1.0 mol/L). The
combined aqueous fractions were layered with CH2Cl2
and aqueous NaOH (1.0 mol/L) was added to reach
pH 5–6. The products were extracted into CH2Cl2.
The combined organic fractions were washed with aque-
ous NaOH (1.0 mol/L, 2 · 20 mL) to remove the acid
37. The organic layer was dried over sodium sulfate
and evaporated to give the ester 38. The pH of the aque-
ous layer was adjusted to 5–6 with HCl (1.0 mol/L) and
the acid 37 was extracted with CH2Cl2. The organic
layer was dried over sodium sulfate and evaporated to
give the acid 37. Allyloxy-derived impurities were pres-
ent in each sample so both the ester and acid 37 were
separately extracted into acid or base for further purifi-
cation. 37 (0.41 g, 40% yield) and 38 (0.23 g, 25% yield)
were isolated. Compound 37: 1H NMR (CDCl3,
400 MHz): d 8.27 (s, 1H), 8.22 (d, J = 3.9 Hz, 1H),
7.52 (d, J = 7.8 Hz, 1H), 7.39–7.37 (m, 2H), 7.23–7.12
(m, 7 H), 7.07 (m, 1H), 7.02 (d, J = 3.9 Hz, 1H), 3.66
(d, J = 13.7 Hz, 1H), 3.46 (d, J = 9.8 Hz, 1H), 3.27–
3.20 (m, 1H), 3.00–2.85 (m, 3H), 2.69 (m, 1H), 2.50
(m, 1H), 2.22 (m, 1H), 1.77–1.69 (m, 2H), 1.09 (t,
J = 7.5 Hz, 3H). 13C NMR (CDCl3, 63 MHz) d 175.57,
148.66, 146.94, 141.68, 140.27, 137.70, 128.77, 128.72,
128.42, 127.99, 126.63, 126.51, 126.15, 123.62, 70.74,
57.51, 55.93, 52.93, 40.35, 33.35, 25.84, 15.89. FD-MS
(t, J = 7.5 Hz, 3H). ESI-MS (m/z): 439 (M+H)+.
5.29. rac-3-Allyl [(2R,4R)-4-(2-ethylphenyl)-2-phenyl-1-
N-(3-pyridylmethyl)-piperidin]-(3S)-3-ylcarboxylate (36)
Sodium cyanoborohydride (1.0 g, 16 mmol) and bro-
mocresol green (1.0 mg) were added to a solution of
the enamine 35 (5.9 g, 14 mmol) in THF (65 mL) and
absolute EtOH (130 mL). To the resultant blue solution
was added dropwise a solution of concd HCl/EtOH (1:2)
to maintain a yellow color over the course of a day. The
solution was stirred at room temperature, overnight.
Thirty minutes prior to workup, concd HCl/EtOH solu-
tion (10 mL) was added to the reaction mixture. Aque-
ous HCl (5.0 mol/L) was then added at a rate such
that with ice bath cooling, the mixture was kept at
25 ꢁC. A clear yellow solution formed after 80 mL was
added and a total of 100 mL was added. The solution
was basified with aqueous NaOH (5.0 mol/L) cooling
with ice bath to keep the mixture below 26 ꢁC. The
product was extracted with EtOAc. The extract was
dried over sodium sulfate and evaporated under reduced
pressure. The resulting residue, a blue foam, 5.2 g,
was chromatographed on silica gel, eluted with
CH2Cl2/EtOAc, 9:1 to 1:1, to afford 36 as a yellow solid
(3.6 g, 61%) and a borane-complexed product (0.93 g).
Decomposition of borane-complex. To a solution of the
borane-complexed product (0.89 g) in MeOH (20 mL)
was added Amberlyst A-21 resin (9.0 g) and the slurry
was refluxed for 48 h. The mixture was cooled to room
temperature and filtered. After evaporation, the residue
was dissolved in CH2Cl2. The solution was dried over
sodium sulfate and evaporated under reduced pressure
to afford 36 as an oil which crystallized upon standing
(0.74 g, 13% yield). 1H NMR (CDCl3, 400 MHz) d
8.13 (d, J = 7.8 Hz, 2H), 7.44 (d, J = 7.3 Hz, 1H),
6.93–6.67 (m, 8 H), 4.92–4.82 (m, 1H), 4.46 (dd,
J = 11.0, 1.5 Hz, 1H), 4.27 (dd, J = 17.0, 1.7 Hz, 1H),
3.67–3.52 (m, 3H), 3.27 (d, J = 3.9 Hz, 1H), 2.92–2.86
(m, 2H), 2.72–2.62 (m, 3H), 2.44–2.35 (m, 1H), 2.32–
2.23 (m, 1H), 1.87 (dt, J = 12.0, 2.4 Hz, 1H), 1.22–1.18
(m, 1H), 0.89 (t, J = 7.5 Hz, 3H). 13C NMR (CDCl3,
63 MHz) d 170.11, 149.93, 148.20, 141.50, 140.90,
139.23, 136.38, 134.49, 131.91, 128.76, 128.58, 128.42,
127.72, 127.62, 126.70, 125.91, 123.46, 117.03, 69.93,
63.95, 56.53, 54.15, 53.93, 40.59, 25.84, 25.17, 15.79.
ESI-MS (m/z): 441 (M+H)+.
1
(m/z): 401 (M+H)+. Compound 38: H NMR (CDCl3,
400 MHz) d 8.42 (s, 2H), 7.54 (d, J = 7.3 Hz, 1H), 7.40
(s, 2H), 7.31–7.17 (m, 5 H), 7.12–7.00 (m, 3H), 5.07
(m, 1H), 4.74 (m, 1H), 4.54 (m, 1H), 3.90 (m, 1H),
3.80 (m, 1H), 3.73 (d, J = 13.7 Hz, 1H), 3.50 (d,
J = 9.8 Hz, 1H), 3.27 (m, 1H), 3.04–2.88 (m, 3H),
2.75–2.66 (m, 1H), 2.58–2.48 (m, 1H), 2.26 (s, 1H),
1.75 (s, 2H), 1.12 (t, J = 7.5 Hz, 3H). 13C NMR (CDCl3,
63 MHz) d 172.33, 150.18, 148.34, 141.71, 140.82,
140.09, 136.29, 131.61, 128.85, 128.70, 128.27, 127.98,
126.55, 126.46, 126.07, 123.38, 117.30, 70.89, 64.41,
57.92, 56.42, 52.97, 40.45, 33.63, 25.88, 15.83. FD-MS
(m/z): 441 (M+H)+.
5.31. rac-Chloro [(2R,4R)-4-(2-ethylphenyl)-2-phenyl-1-
N-(3-pyridylmethyl)-piperidin]-(3S)-3-ylcarboxylate (39)
The acid 37 (0.37 g, 0.92 mmol) was dissolved in thionyl
chloride (5.0 mL) with a drop of DMF and the solution
was refluxed for 48 h. After cooling to room tempera-
ture, the thionyl chloride was evaporated. Toluene was
added and evaporated. The gummy solid was triturated
with ether to give an orange powder. The ether was
decanted and the trituration repeated twice. The
acid chloride 39 was used directly without further
purification.
5.30. rac-[(2R,4R)-4-(2-Ethylphenyl)-2-phenyl-1-N-(3-
pyridylmethyl)piperidin]-(3S)-3-ylcarbonic acid (37) and
rac-3-allyl [(2R,4R)-4-(2-ethylphenyl)-2-phenyl-1-N-(3-
pyridylmethyl)- piperidin]- (3S)-3-ylcarboxylate (38)
The ester 11a (1.0 g, 2.3 mmol) was treated with aqueous
sodium allyloxide (10%wt) in allyl alcohol (50 mL). The
solution was refluxed for 22 h. After cooling to room
temperature, the solution was diluted with CH2Cl2 and
saturated ammonium chloride solution. The aqueous
solution was neutralized with HCl (1.0 mol/L). The lay-
ers were separated and the aqueous layer was extracted
with CH2Cl2. The combined organic extracts were dried
over sodium sulfate and evaporated to give an orange
5.32. rac-N-Methyl [(2R,4R)-4-(2-ethylphenyl)-2-phenyl-
1-N-(3-pyridylmethyl)-piperidin]-(3S)-3-ylcarboxamide
(40)
The acid chloride 39 (75 mg, 0.18 mmol) and methyl-
amine hydrochloride (0.10 g, 1.5 mmol) were dissolved
in pyridine (1.0 mL) and the mixture was stirred for
48 h. The solvent was evaporated and the residue was
dissolved in CH2Cl2. The solution was treated with

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R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
saturated ammonium chloride solution and the pH
adjusted to 5–6 with aqueous HCl (1.0 mol/mL). The
organic layer was dried over sodium sulfate and evapo-
rated under reduced pressure. The resulting residue, or-
ange oil (72 mg), was chromatographed on silica gel,
eluted with CH2Cl2/EtOAc, 3:1, to provide 40 which
was further purified by aqueous NaOH (1.0 mol/mL)
washes of an EtOAc solution. Final purification was
achieved with a preparative silica plate with CH2Cl2/
EtOAc, 3:1, to afford 40 (20 mg, 27% yield) as a white
solid. 1H NMR (CDCl3, 300 MHz) d 8.41 (s, 2H),
7.55–7.39 (m, 3H), 7.30–7.15 (m, 5 H), 7.07–7.00 (m,
3H), 4.27 (s, 1H), 3.73 (d, 1H), 3.62 (d, 1H), 3.38 (m,
1H), 3.00–2.75 (m, 3H), 2.62–2.45 (m, 2H), 2.28 (m,
1H), 2.03 (d, 3H), 1.75 (m, 2H), 1.13 (t, 3H). FD-MS
(m/z): 414 (M+H)+.
3.40 (m, 1H), 3.25–3.15 (m, 2H), 2.95 (dt, 1H), 2.85
(d, 1H), 2.75–2.60 (m, 1H), 2.55–2.40 (m, 1H), 2.23
(dt, 1H), 1.90–1.65 (m, 2H), 1.10 (t, 3H). 1.00 (s, 3H).
FD-MS (m/z): 399 (M+H)+.
5.35. rac-Methyl [(2R,4R)-4-(2-ethylphenyl)-2-phenyl-1-
N-(3-pyridylmethyl)- piperidin]-(3S)-3-ylcarboxylate (43)
The acid chloride 39 (75 mg, 0.18 mmol) was dissolved
in MeOH (5.0 mL) and the solution was stirred for
48 h. The solvent was evaporated and the residue was
dissolved in CH2Cl2. The solution was washed with sat-
urated aqueous NaHCO3, dried over sodium sulfate,
and evaporated under reduced pressure. The resulting
residue, an orange oil (69 mg), was chromatographed
on silica gel, eluted with hexane/EtOAc, 1:1, to provide
1
43 (38 mg, 51% yield) as an off-white solid. H NMR
5.33. rac-(2R,3S,4R)-4-(2-Ethylphenyl)-3-hydroxymeth-
yl-2-phenyl-1-N-(3-pyridylmethyl)piperidine (41)
(CDCl3, 400 MHz) d 8.41 (s, 2H), 7.54 (d, J = 6.8 Hz,
1H), 7.39 (s, 2H), 7.29–7.14 (m, 5H), 7.09–7.00 (m,
3H), 3.73 (d, J = 13.7 Hz, 1H), 3.50 (d, J = 9.8 Hz,
1H), 3.26 (m, 1H), 2.99–2.87 (m, 6 H), 2.71 (m, 1H),
2.55 (m, 1H), 2.26 (s, 1H), 1.74 (s, 2H), 1.13 (t,
J = 7.5 Hz, 3H). FD-MS (m/z): 415 (M+H)+.
To a solution of 39 (75 mg, 0.18 mmol) in THF
(0.50 mL) was added lithium aluminum hydride
(0.020 g, 0.54 mmol) and the mixture was stirred for
24 h. After the reaction was quenched with wet THF,
the mixture was filtered through Celite with EtOAc.
The filtrate was washed with saturated ammonium chlo-
ride solution. The organic layer was dried over sodium
sulfate and evaporated under reduced pressure. The
resulting residue, orange oil (72 mg), was chromato-
graphed on silica gel, eluted with hexane/EtOAc, 1:1,
5.36. rac-N-(O-Methyl-l-Methionine) [(2R,4R)-4-(2-eth-
ylphenyl)-2-phenyl-1-N-(3-pyridylmethyl)-piperidin]-(3S)-
3-ylcarboxylamide (44)
To a solution of 39 (100 mg, 0.24 mmol) in pyridine
(1.0 mL) was added l-methionine methyl ester hydro-
chloride (0.10 g, 0.48 mmol) at room temperature. After
stirred for 24 h, the solvent was evaporated and the res-
idue was dissolved in EtOAc. The solution was washed
with saturated ammonium chloride solution (2·), aque-
ous NaOH (1.0 mol/L), and saturated ammonium chlo-
ride solution. The organic layer was dried over sodium
sulfate and evaporated under reduced pressure. The
resulting residue was chromatographed on silica gel,
eluted with hexane/acetone/CH2Cl2, 4:2:1, to afford 44
1
to provide 41 (12 mg, 17% yield) as a white solid. H
NMR (CDCl3, 300 MHz) d 8.40 (m, 2 H), 7.53–7.45
(m, 3H), 7.30–7.05 (m, 8 H), 3.65 (d, 1H), 3.40 (d,
1H), 3.15–3.00 (m, 2H), 2.96–2.75 (m, 4 H), 2.60 (m,
1H), 2.15 (dt, 1H), 2.00 (m, 1H), 1.85–1.70 (m, 2H),
1.15 (t, 3H). FD-MS (m/z): 387 (M+H)+.
5.34. rac-(2R,3S,4R)-4-(2-Ethylphenyl)-3-acetyl-2-phen-
yl-1-N-(3-pyridylmethyl)piperidine (42)
1
(49 mg, 37% yield) as a yellow solid. H NMR (CDCl3,
To ice-cold di-tert-butylmalonate (2.7 mL, 12 mmol)
was added lithium bis(trimethylsilyl)amide (12 mL,
1.0 mol/L in THF). After stirred for 30 min, 39 (0.50 g,
1.2 mmol) was added in one portion as a solid. The solu-
tion was stirred for 1 h with the ice bath and then it was
stirred for 48 h. Water was added and the solution was
acidified to pH 5 with aqueous HCl (0.10 mol/L). The
product was extracted with EtOAc. The combined
organic layers were dried over sodium sulfate and evap-
orated to give diester derivative and unreacted di-tert-
butylmalonate. To a solution of this mixture in acetic
anhydride/acetic acid (2:98, 8.0 mL) was added cam-
phorsulfonic acid (42 mg, 0.18 mmol) and refluxed for
3 h. After cooling to room temperature, the solution
was evaporated. Toluene was added and evaporated.
The resulting residue was dissolved in EtOAc and
washed with aqueous NaOH (1.0 mol/L). The organic
layer was dried over sodium sulfate and evaporated un-
der reduced pressure. The resulting residue was purified
by radial chromatography eluted with hexane/EtOAc,
300 MHz): d 8.45 (m, 2H), 7.55–7.40 (m, 3H), 7.30–7.00
(m, 8H), 5.25 (dd, 1H), 4.05 (m, 1H), 3.75–3.55 (m, 2H),
3.45–3.25 (m, 4H), 3.00–2.88 (m, 2H), 2.75 (m, 1H),
2.60–2.45 (m, 2H), 2.28 (m, 1H), 1.85–1.70 (m, 5H),
1.60–1.39 (m, 2H), 1.30–1.05 (m, 5H). FD-MS (m/z):
546 (M+H)+.
5.37. rac-N-(l-methionine) [(2R,4R)-4-(2-ethylphenyl)-2-
phenyl-1-N-(3-pyridylmethyl)-piperidin]-(3S)-3-ylcarbox-
ylamide (45)
To a solution of 44 (33 mg, 0.06 mmol) in THF (1.0 mL)
was added a solution of lithium hydroxide (8.0 mg,
0.19 mmol) in water (1.0 mL). After stirred for 24 h,
the mixture was diluted with saturated ammonium
chloride solution and neutralized with aqueous HCl
(0.10 mol/L). The product was extracted with CH2Cl2.
The combined organic layers were dried over sodium
sulfate and evaporated to give 45 (25 mg, 78% yield)
of acceptable purity. 1H NMR (CDCl3, 300 MHz) d
8.45 (m, 2H), 7.60–7.40 (m, 3H), 7.34–6.85 (m, 8H),
5.60 (m, 1H), 4.00 (m, 1H), 3.85–3.63 (m, 2H), 3.33
(m, 1H), 3.07–2.95 (m, 2H), 2.83–2.68 (m, 2H), 2.53
1
1:1, to afford 42 (60 mg, 13% yield) as a yellow oil. H
NMR (CDCl3, 300 MHz) d 8.45 (br s, 2H), 7.53 (d,
1H), 7.45 (m, 1H), 7.37–7.00 (m, 9 H), 3.70 (d, 1H),

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1379
(m, 1H), 2.35 (m, 1H), 1.90–1.73 (m, 5H), 1.62–1.50 (m,
2H), 1.43–1.35 (m, 2H), 1.18–1.05 (m, 3H). FD-MS (m/
z): 532 (M + H)+.
(m, 2H), 5.13 (dd, J = 11.2, 9.7 Hz, 1H), 3.88–3.71 (m,
7H), 3.31–3.07 (m, 2H), 2.95 (dd, J = 12.1, 3.5 Hz,
1H), 2.02 (td, J = 12.1, 8.9 Hz, 1H). FAB-MS (m/z):
500, 498 (M+H)+.
5.38. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(3,4-dihydr-
oxyphenyl)-3-nitro-1-N-(3-pyridylmethyl) piperidine (2a)
5.40. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(4-hydroxy-3-
iodophenyl)-3-nitro-1-N-(3-pyridylmethyl) piperidine
(47a)
Step 1 According to the procedure for the synthesis
of 7, methyl-3-(2-bromophenyl)-4-nitrobutyrate (81 g,
0.27 mol), 3,4-dihydroxybenzaldehyde (37 g, 0.27 mol),
and 3-(aminomethyl)pyridine (54 mL, 0.53 mol) affor-
ded 4-(2-bromophenyl)-6-(3,4-dihydroxyphenyl)-5-ni-
tro-1-(3-pyridylmethyl) piperidin-2-one (78 g, 60%
yield). 1H NMR (DMSO-d6) d 9.23 (s, 1H), 8.95 (s,
1H), 8.43 (dd, J = 4.6, 1.5 Hz, 1H), 8.18 (d, J = 2.0 Hz,
1H), 7.79 (d, J = 7.2 Hz, 1H), 7.61 (dd, J = 7.8, 1.1 Hz,
1H), 7.41–7.47 (m, 2H), 7.29 (dd, J = 7.8, 4.7 Hz, 1H),
7.22 (dt, J = 7.8, 1.5 Hz, 1H), 6.75 (d, J = 2.0 Hz, 1H),
6.64 (d, J = 8.1 Hz, 1H), 6.56 (dd, J = 8.1, 2.0 Hz, 1H),
5.84 (dd, J = 11.3, 9.8 Hz, 1H), 4.80 (d, J = 9.8 Hz,
1H), 4.65 (d, J = 15.6 Hz, 1H), 4.35 (m, 1 H), 3.99 (d,
J = 15.6 Hz, 1H), 3.02 (dd, J = 16.9, 12.5 Hz, 1H), 2.72
(dd, J = 16.9, 4.6 Hz, 1H). FAB-MS (m/z): 500, 498
(M+H)+. Step 2 To a solution of 4-(2-bromophenyl)-6-
(3,4-dihydroxyphenyl)-5-nitro-1-(3-pyridylmethyl)pip-
eridin-2-one (41 g, 82 mmol) in THF (1.6 L) was added
borane–methyl sulfide complex (30 mL, 320 mmol) and
the mixture was refluxed for 10 h. After cooling to
4 ꢁC, the reaction was quenched by the addition of water
(50 mL) and the mixture was concentrated. The result-
ing residue was heated with aqueous HCl (3.0 mol/L,
400 mL) and neutralized with aqueous NaOH
(3.5 mol/L, 300 mL) after cooling to room temperature.
The solid formed was recrystallized from EtOH to give
analytically pure 2a (24.7 g, 62% yield). 1H NMR
(DMSO-d6, 300 MHz) d 9.05 (s, 1H), 8.97 (s, 1H), 8.44
(m, 2H), 8.42 (m, 1H), 7.75 (m, 1H), 7.66–7.56 (m,
2H), 7.40–7.32 (m, 2H), 7.17 (m, 1H), 6.94 (br s, 1H),
6.72 (m, 1H), 5.25 (dd, J = 9.4, 11.6 Hz, 1H), 3.86 (dt,
J = 4.0, 11.6 Hz, 1H), 3.64 (d, J = 13.8 Hz, 1H), 3.63
(d, J = 9.4 Hz, 1H), 3.05 (d, J = 13.8 Hz, 1H), 2.88 (m,
1H), 2.36 (m, 1H), 1.86–1.71 (m, 2H). FAB-MS (m/z):
486, 484 (M+H)+.
Step 1 According to the procedure for the synthesis
of 7, methyl 3-(2-bromophenyl)-4-nitro-butylate (1.4 g,
4.7 mmol), 4-hydroxy-3-iodobenzaldehyde (1.0 g,
4.7 mmol) and 3-aminomethyl-pyridine (1.0 g,
9.4 mmol) afforded 4-(2-bromophenyl)-6-(4-hydroxy-3-
iodophenyl)-5-nitro-1-(3-pyridylmethyl)-piperidin-2-one
(1.7 g, 60% yield). FAB-MS (m/z): 609, 607 (M+H)+.
Step 2 According to the procedure for the synthesis
of 1a, 4-(2-bromophenyl)-6-(4-hydroxy-3-iodophenyl)-
5-nitro-1-N-(3-pyridylmethyl)-piperidin-2-one (1.3 g,
2.0 mmol) and borane-methyl sulfide complex (1.9 mL,
1
10 mmol) afforded 47a (470 mg, 40% yield). H NMR
(DMSO-d6, 300 MHz) d 10.45 (br s, 1H), 8.44–8.30
(m, 2H), 7.91 (m, 1H), 7.80 (d, J = 7.91 Hz, 1H), 7.57
(d, J = 9.4 Hz, 1H), 7.55 (dd, J = 8.0, 1.1 Hz, 1H),
7.39–7.30 (m, 3H), 7.16 (m, 1H), 6.85 (d, J = 8.0 Hz,
1H), 5.39 (dd, J = 11.0, 9.5 Hz, 1H), 3.85 (m, 1H),
3.73 (d, J = 9.5 Hz, 1H), 3.55 (d, J = 14.0 Hz, 1H),
3.07 (d, J = 14.0 Hz, 1H), 2.88 (m, 1H), 2.48 (m, 1H),
1.87–1.72 (m, 2H). FAB-MS (m/z): 595, 593 (M+H)+.
5.41. rac-(2R,3S,4S)-4-(2-Ethylphenyl)-2-(4-hydroxyphe-
nyl)-3-nitro-1-N-(3-pyridylmethyl)piperidine (1b)
Step 1 According to the procedure for the synthesis
of 7, methyl 3-(2-ethylphenyl)-4-nitrobutylate (1.0 g,
4.0 mmol), 4-hydroxybenzaldehyde (450 mg, 4.0 mmol),
and 3-aminomethylpyridine (0.82 mL, 8.0 mmol)
afforded 4-(2-ethylphenyl)-6-(4-hydroxyphenyl)-5-nitro-
1-N-(3-pyridylmethyl)piperidin-2-one (1.4 g, 82% yield).
Step 2 According to the procedure for the synthesis of
1a, 4-(2-ethylphenyl)-6-(4-hydroxyphenyl)-5-nitro-1-N-
(3-pyridylmethyl)piperidin-2-one (670 mg, 1.6 mmol)
and borane–methyl sulfide complex (0.74 mL, 7.8 mmol)
afforded 1b (420 mg, 64% yield). 1H NMR (CDCl3,
270 MHz) d 8.59 (br s, 1H), 8.48 (dd, J = 5.0, 1.6 Hz,
1H), 8.15 (m, 1H), 7.56 (d, J = 7.9 Hz, 1H), 7.37–7.28
(m, 4H), 7.20–7.13 (m, 2H), 6.87 (d, J = 8.6 Hz, 2H),
4.97 (dd, J = 10.9, 9.6 Hz, 1H), 3.85 (d, J = 9.2 Hz,
1H), 3.70 (d, J = 5.9 Hz, 1H), 7.65 (m, 1H), 3.09–3.04
(m, 2H), 2.79–2.57 (m, 2H), 2.43 (m, 1H), 1.90 (m,
2H), 1.20 (t, J = 7.6 Hz, 3H). FAB-MS (m/z): 434
(M+H)+.
5.39. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(4-hydroxy-3-
methoxyphenyl)-3-nitro-1-N-(3-pyridylmethyl) piperidine
(46a)
Step 1 According to the procedure for the synthesis of
7, methyl 3-(2-bromophenyl)-4-nitrobutylate (300 mg,
1.0 mmol), vanilline (150 mg, 1.0 mmol), and 3-aminom-
ethylpyridine (0.20 mL, 2.0 mmol) afforded 4-(2-ethyl-
phenyl)-6-(4-hydroxy-3-methoxyphenyl)-5-nitro-1-(3-
pyridyl-methyl)piperidin-2-one (320 mg, 63% yield).
FAB-MS (m/z): 514, 512 (M+H)+. Step 2 According
to the procedure for the synthesis of 1a, 4-(2-bromophe-
nyl)-6-(4-hydroxy-3-methoxy-phenyl)-5-nitro-1-N-(3-
pyridylmethyl)piperidin-2-one (135 mg, 0.26 mmol) and
borane–methyl sulfide complex (0.95 mL, 10 mmol)
5.42. rac-(2R,3S,4S)-4-(2-Ethylphenyl)-2-(3-amino-4-
hydroxyphenyl)-3-nitro-1-N-(3-pyridylmethyl)piperidine
(11b)
Step 1 According to the procedure for the synthesis of
10, concentrated nitric acid (1.0 mL, 25 mmol) and 1b
(1.7 g, 3.9 mmol) afforded 4-(2-ethylphenyl)-6-(3-ami-
no-4-hydroxyphenyl)-5-nitro-1-N-(3-pyridylmethyl)pip-
eridin-2-one (1.3 g, 69% yield). 1H NMR (CDCl3,
270 MHz) d 8.51 (d, J = 3.3 Hz, 1H), 8.47 (br s, 1H),
1
afforded 46a (47 mg, 39% yield). H NMR (CD3OD,
300 MHz) d 8.48 (s, 1H), 8.39 (m, 1H), 7.88 (d,
J = 7.9 Hz, 1H), 7.73 (d, J = 6.4 Hz, 1H), 7.53–7.46
(m, 2H), 7.35 (m, 1H), 7.17–7.08 (m, 2H), 6.96–6.86

1380
R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
7.72 (d, J = 7.6 Hz, 1H), 7.57 (d, J = 7.6 Hz, 1H), 7.32–
7.15 (m, 7H), 4.93 (dd, J = 11.2, 9.8 Hz, 1H), 3.86 (d,
J = 9.6 Hz, 1H), 3.76–7.66 (m, 2H), 3.12–3.05 (m, 2H),
2.78–2.57 (m, 2H), 2.43 (m, 1H), 1.95–1.86 (m, 2H),
1.20 (t, J = 7.6 Hz, 3H). FAB-MS (m/z): 463 (M+H)+.
Step 2 According to the procedure for the synthesis of
11a, prepared nitro compound (0.47 g, 1.0 mmol) and
palladium on carbon afforded 11b (0.16 g, 37% yield).
1H NMR (CDCl3, 270 MHz) d 8.52 (br s, 1H), 8.43
(d, J = 4.0 Hz, 1H), 7.58 (d, J = 7.9 Hz, 1H), 7.35–7.11
(m, 5H), 6.80–6.61 (m, 3H), 4.98 (dd, J = 10.9, 9.9 Hz,
1H), 3.86 (d, J = 14.2 Hz, 1H), 3.71–3.63 (m, 2H), 3.
07 (d, J = 14.2 Hz, 1H), 3.01 (d, J = 14.5 Hz, 1H),
2.78–2.55 (m, 2H), 2.40 (m, 1H), 1.90–1.50 (m, 2H),
1.18 (t, J = 7.6 Hz, 3H). FAB-MS (m/z): 433 (M+H)+.
FAB-MS (m/z): 512 (M+H)+. Anal. Calcd For
C26H29Cl2N5O4: C, 65.67; H, 6.15; N, 14.73%. Found:
C, 65.91; H, 6.38; N, 14.45%.
5.46. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(4-methoxy-
phenyl)-3-nitro-1-N-(3-pyridylmethyl)piperidine (48a)
Step 1 According to the procedure for the synthesis
of
7,
methyl
3-(2-bromophenyl)-4-nitrobutylate
(300 mg 1.0 mmol), 4-methoxybenzaldehyde (0.10 mL,
0.90 mmol), and 3-aminomethylpyridine (0.20 mL,
2.0 mmol) afforded 4-(2-bromophenyl)-6-(4-methoxy-
phenyl)-5-nitro-1-N-(3-pyridylmethyl)piperidin-2-one
(220 mg, 45% yield). FAB-MS (m/z): 498, 496 (M+H)+.
Step 2 According to the procedure for the synthesis of
1a, 4-(2-bromophenyl)-6-(4-methoxyphenyl)-5-nitro-1-
N-(3-pyridylmethyl)piperidin-2-one (58 mg, 0.12 mmol)
and borane–methyl sulfide complex (0.047 mL,
5.43. rac-N-(5-{[(3R,4R)-4-(2-Ethylphenyl)-3-nitro-1-N-
(3-pyridylmethyl)piperidin]-(2S)-2-yl}-2-hydroxyphenyl)
methanesulfonamide (14b)
1
0.50 mmol) afforded 48a (18 mg, 30% yield). H NMR
(CDCl3, 300 MHz) d 8.52–8.49 (m, 2 H), 7.61–7.53 (m,
2H), 7.38–7.21 (m, 3H), 7.09 (ddd, J = 8.0, 7.2, 1.9 Hz,
1H), 7.02 (d, J = 8.2 Hz, 2H), 6.84 (d, J = 8.2 Hz, 2H),
4.98 (dd, J = 10.7, 9.7 Hz, 1H), 4.03 (m, 1H), 3.90 (s,
3H), 3.89–3.80 (m, 2H), 3.07–3.03 (m, 2tI), 2.43 (m,
1H), 2.05 (m, 1H), 1.70 (m, 1H). FAB-MS (m/z): 484,
482 (M+H)+.
According to the procedure for the synthesis of 14a,
11b (43 mg, 0.10 mmol) and methanesulfonyl chloride
(0.0077 mL, 0.10 mmol) afforded 14b (23 mg, 45%
1
yield). H NMR (DMSO-d6, 270 MHz) d 10.02 (br s,
1H), 8.76 (br s, 1H), 8.46–8.43 (m, 2H), 7.68 (d,
J = 7.6 Hz, 1H), 7.55 (d, J = 6.3 Hz, 1H), 7.45 (m,
1H), 7.32 (dd, J = 7.6, 4.8 Hz, 1H), 7.20–7.12 (m, 4H),
6.87 (d, J = 8.3 Hz, 1H), 5.17 (dd, J = 10.9, 9.6 Hz,
1H), 3.72 (d, J = 9.6 Hz, 1H), 3.65–3.60 (m, 2H), 3.06
(d, J = 13.5 Hz, 1H), 2.92 (s, 3H), 2.85 (m, 1H), 2.69–
2.51 (m, 2H), 2.40 (m, 1H), 1.81 (br s, 2H), 1.12 (t,
J = 7.6 Hz, 3H). FAB-MS (m/z): 511 (M+H)+.
5.47. rac-(2R,3S,4S)-4-(2-Bromophenyl)-2-(5-benzimi-
dazol)-3-nitro-1-N-(3-pyridylmethyl)piperidine (49a)
Step 1 According to the procedure for the synthesis of
7, methyl 3-(2-bromophenyl)-4-nitrobutylate (120 mg,
0.40 mmol), 1-N-trityl-5-benzimidazolecarboxaldehyde
(190 mg, 2.0 mmol), and 3-aminomethylpyridine
(0.082 mL, 0.80 mmol) afforded crude 4-(2-bromophe-
nyl)-6-(1-N-trityl-5-benzimidazol)-5-nitro-1-N-(3-pyr-
idylmethyl)-piperidin-2-one. Step 2 To a solution of
resulting N-trityl derivatives in MeOH (10 mL) was add-
ed trifluoroacetic acid (0.50 mL), and the mixture was
stirred at room temperature for 3 h. The solvent was
evaporated under reduced pressure, and the residue
was purified by preparative silica plate, diluted with
CHCl3/MeOH, 9:1, to afford 4-(2-bromo-phenyl)-6-
(5-benzimidazol)-5-nitro-1-N-(3-pyridylmethyl) piperi-
din-2-one (32 mg, 2 steps 16% yield). FAB-MS (m/z):
508, 506 (M+H)+. Step 3 According to the procedure
for the synthesis of 1a, 4-(2-bromophenyl)-6-(5-ben-
zimidazol)-5-nitro-1-N-(3-pyridylmethyl)piperidin-2-
one (8.0 mg, 0.016 mmol) and borane–methyl sulfide
complex (0.0047 mL, 0.05 mmol) afforded 49a (0.7 mg,
5.44. rac-N-({5-[(3R,4R)-4-(2-Ethylphenyl)-3-nitro-1-N-
(3-pyridylmethyl)piperidin]-(2S)-2-yl}-2-hydroxy)phe-
nylurea (19b)
According to the procedure for the synthesis of 19a, 11b
(86 mg, 0.20 mmol), acetic acid (1.0 mL), and potassium
cyanate (160 mg, 2.0 mmol) afforded 19b (30 mg 32%
1
yield). H NMR (CDCl3, 270 MHz) d 8.44 (br s, 1H),
8.36 (br s, 1H), 7.92 (br s, 1H), 7.52 (m, 1H), 7.25–
7.09 (m, 4H), 6.94–6.81 (m, 3H), 5.45 (br s, 2H), 4.96
(dd, J = 10.8, 9.6 Hz, 1H), 3.77–3.66 (m, 4H), 2.95–
2.88 (m, 2H), 2.81–2.50 (m, 2H), 2.34 (m, 1H), 1.84–
1.82 (m, 2H), 1.16 (t, J = 7.8 Hz, 3H). FAB-MS (m/z):
476 (M+H)+.
5.45. rac-5-[{(3R,4R)-4-(2-Ethylphenyl)-3-nitro-1-N-(3-
pyridylmethyl)piperidine}-(2S)-yl]-2-hydroxyphenyl
sulfamide (20b)
1
8.9% yield). H NMR (CDCl3, 270 MHz) d 8.49–8.47
(m, 2H), 8.06 (br s, lH), 7.78–7.67 (m, 2H), 7.59–7.48
(m, 3H), 7.35 (m, 1H), 7.28–7.20 (m, 3H), 7.08 (m,
1H), 5.08 (dd, J = 10.8, 9.2 Hz, 1H), 4.09 (m, 1H),
3.99 (d, J = 9.2 Hz, 1H), 3.80 (d, J = 13.9 Hz, 1H),
3.09–3.04 (m, 2H), 2.45 (m, 1H), 2.08 (m, 1H), 1.75
(m, 1H). FAB-MS (m/z): 494, 492 (M+H)+.
According to the procedure for the synthesis of 20a,
11b (200 mg 0.46 mmol), sulfamoyl chloride (79 mg
0.69 mmol), and N,N-dimethylacetamide solution
(5.0 mL) afforded 20b (62 mg 26% yield). 1H NMR
(DMSO-d6, 270 MHz) d 9.84 (br s, 1H), 8.46–8.43 (m,
2H), 7.98 (br s, 1H), 7.73 (d, J = 7.9 Hz, 1H), 7.64–
7.53 (m, 2H), 7.32 (dd, J = 7.9, 5.0 Hz, 1H), 7.19–7.13
(m, 4H), 7.00 (br s, 1H), 6.81 (d, J = 8.3 Hz, 1H), 5.25
(dd, J = 10.2, 10.0 Hz, 1H), 3.84–3.72 (m, 2H), 3.16 (d,
J = 13.5 Hz, 1H), 3.00 (d, J = 11.2 Hz, 1H), 2.81–2.51
(m, 3H), 1.94-1.85 (m, 2H), 1.26 (t, J = 7.6 Hz, 3H).
5.48. rac-(2R,3S,4S)-4-(2-Bromophenyl)-3-nitro-2-phenyl-
1-N-(3-pyridylmethyl)piperidine (50a)
Step 1 According to the procedure for the synthesis of
7, methyl 3-(2-bromophenyl)-4-nitrobutylate (110 mg

R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
1381
0.50 mmol), benzaldehyde (53 mg, 1.0 mmol), and
3-aminomethylpyridine (0.11 mL, 1.0 mmol) afforded
4-(2-bromophenyl)-5-nitro-6-phenyl-1-N-(3-pyridylm-
ethyl)piperidin-2-one (110 mg 47% yield). FAB-MS
(m/z): 468, 466 (M+H)+. Step 2 According to the
procedure for the synthesis of 1a, 4-(2-bromophenyl)-
5-nitro-6-phenyl-1-N-(3-pyridylmethyl)piperidin-2-one
(33 mg 0.072 mmo1) and borane–methyl sulfide com-
plex (0.025 mL, 0.27 mmol) afforded 50a (23 mg 69%
yield). 1H NMR (CDCl3, 300 MHz) d 8.49 (br s,
2H), 7.56–7.47 (m, 4H) 7.41–7.20 (m, 6H), 6.98 (m,
1H), 5.02 (dd, J = 10.7, 10.1 Hz, 1H), 4.07 (m, 1H),
3.86 (d, J = 9.4 Hz, 1H), 3.75 (d, J = 13.7 Hz, 1H),
3.06–2.95 (m, 2H), 2.43 (m, 1H), 2.04 (d,
J = 11.4 Hz, 1H), 1.68 (m, 1H). FAB-MS (m/z): 454,
452 (M+H)+.
for C22H21BrN4O2: C, 58.29; H, 4.67; N, 12.36%.
Found: C, 58.32; H, 4.90; N, 12.37%. FAB-MS (m/
z): 455, 453 (M+H)+.
5.51. rac-(2R,3S,4S)-4-(2-Bromophenyl)-3-nitro-2-(3-pyri-
dyl)-1-N-(3-pyridylmethyl)piperidine (53a)
Step 1 According to the procedure for the synthesis of
7, methyl 3-(2-bromophenyl)-4-nitrobutylate (300 mg,
1.0 mmol), 3-pyridinecarboxaldehyde (100 mg, 1.0
mmol) and 3-aminomethylpyridine (0.20 mL, 2.0 mmol)
afforded 4-(2-bromophenyl)-5-nitro-6-(3-pyridyl)-1-N-
(3-pyridylmethyl)-piperidin-2-one (190 mg, 41% yield).
FAB-MS (m/z): 469, 467 (M+H)+. Step 2 According
to the procedure for the synthesis of 1a, 4-(2-bromophe-
nyl)-5-nitro-6-(3-pyridyl)-1-N-(3-pyridylmethyl)piperi-
din-2-one (190 mg, 0.41 mmol) and borane–methyl
sulfide complex (0.20 mL, 2.1 mmol) afforded 53a
(80 mg, 18% yield). 1H NMR (CDCl3, 270 MHz) d
8.74 (br s, 1H), 8.61 (m, 1H), 8.52–8.48 (m, 2H), 7.83
(d, J = 7.9 Hz, 1H), 7.54 (d, J = 7.9 Hz, 2H), 7.38–7.22
(m, 4H), 7.09 (m, 1H), 5.01 (dd, J = 10.8, 9.8 Hz, 1H),
4.08 (m, 2H), 3.93 (d, J = 9.5 Hz, 1H), 3.70 (d,
J = 13.4 Hz, 1H), 3.11–3.06 (m, 2H), 2.46 (m, 1H),
2.07 (m, 1H), 1.70 (m, 1H). FAB-MS (m/z): 455, 453
(M+H)+.
5.49. rac-(2R,3S,4S)-4-(2-Bromophenyl)-3-nitro-1-N-(3-
pyridylmethyl)-2-(3-thienyl)piperidine (51a)
Step 1 According to the procedure for the synthesis of
7, methyl 3-phenyl-4-nitrobutylate (110 mg, 0.50 mmol),
3-thiophenecarboxaldehyde (0.044 mL 0.50 mmol) and
3-aminomethylpyridine (0.41 mL, 4.0 mmol) afforded
4-(2-bromophenyl)-5-nitro-1-N-(3-pyridylmethyl)-6-(3-
thienyl)piperidin-2-one (75 mg, 32% yield). FAB-MS
(m/z): 474, 472 (M+H)+. Step 2 According to the pro-
cedure for the synthesis of 1a, 4-(2-bromo-phenyl)-5-ni-
tro-1-N-(3-pyridylmethyl)-6-(3-thiophenyl)piperidin-2-
one (30 mg, 0.064 mmol) and borane–methyl sulfide
complex (0.025 mL, 0.27 mmol) afforded 51a (14 mg,
5.52. Enzyme assay of FTase inhibition
FTase activity with full-length K-Ras substrate was
determined by measuring transfer of [3H]farnesyl
from [3H]farnesyl diphosphate to trichloroacetic
acid-precipitable His6-K-Ras4B, respectively. Typical
reaction mixture contained 50 mM Tris–HCl, pH
7.7, MgCl2 (5 mM), ZnCl2 (5 lM), DTT (2 mM),
1
48% yield). H NMR (CDCl3, 300 MHz) d 8.51–8.42
(m, 2H), 7.58–7.52 (m, 2H), 7.40–7.18 (m, 6H), 7.10
(m, 1H), 5.00 (dd, J = l.5, 10.8 Hz, 1H), 4.05–3.96 (m,
2H), 3.81 (d, J = 13.8 Hz, 1H), 3.08 (d, J = 13.8 Hz,
1H), 2.98 (m, 1H), 2.40 (m, 1H), 2.03 (m, 1H), 1.68
(m, 1H). Anal. Calcd For C21H20BrN3O2S: C, 55.03;
H, 4.40; N, 9.17%. Found: C, 54.97; H, 4.52; N,
8.77%. FAB-MS (m/z): 460, 458 (M+H)+.
0.2%
octyl-glucoside,
[3H]farnesyl
diphosphate
(500 nM), 50 ng purified enzyme, indicated concentra-
tion of piperidine compounds or DMSO vehicle con-
trol, and K-Ras4B (1 lM). After 30-min incubation
at room temperature, reactions were stopped with
4% SDS and K-Ras protein was precipitated with
cold 30% trichloroacetic acid (0.5 mL). Precipitated
protein was collected on GF/C filter paper mats
using a Brandel harvester. Filter mats were washed
once with 6% trichloroacetic acid, 2% SDS and
radioactivity was measured in a Wallac 1204 Beta-
plate BS liquid scintillation counter. Percent inhibi-
tion was calculated relatively to the DMSO vehicle
control.
5.50. rac-(2R,3S,4S)-4-(2-Bromophenyl)-3-nitro-2-(2-pyri-
dyl)-1-N-(3-pyridylmethyl)piperidine (52a)
Step 1 According to the procedure for the synthesis of
7, methyl 3-(2-bromophenyl)-4-nitrobutylate (600 mg,
2.0 mmol), 2-pyridinecarboxaldehyde (210 mg, 2.0
mmol)
4.0 mmol) afforded 4-(2-bromophenyl)-6-(2-pyridyl)-5-
and
3-aminomethylpyridine
(0.41 mL,
nitro-1-N-(3-pyridylmethyl)-piperidin-2-one
(96 mg,
10% yield). FAB-MS (m/z): 469, 467 (M+H)+. Step
2 According to the procedure for the synthesis of
5.53. Molecular modeling
1a,
4-(2-bromophenyl)-6-(2-pyridyl)-5-nitro-1-N-(3-
pyridylmethyl)-piperidin-2-one (30 mg, 0.064 mmol)
and borane–methyl sulfide complex (0.074 mL,
0.78 mmol) afforded 52a (14 mg, 48% yield). 1H
NMR (CDCl3, 270 MHz) d 8.72 (d, J = 4.6 Hz, 1H),
8.49 (dd, J = 4.6, 1.8 Hz, 1H), 8.44 (d, J = 1.8 Hz,
1H), 8.11 (td, J = 7.4, 1.8 Hz, 1H), 7.59 (d,
J = 7.9 Hz, 1H), 7.52 (dd, J = 8.2, 1.0 Hz, 1H), 7.40
(m, 1H), 7.29–7.21 (m, 4H), 7.08 (m, 1H), 5.37 (dd,
J = 11.2, 9.9 Hz, 1H), 4.11–4.03 (m, 2H), 3.61 (d,
J = 13.5 Hz, 1H), 3.20 (d, J = 13.5 Hz, 1H), 2.52 (m,
1H), 2.08–2.01 (m, 2H), 1.81 (m, 1H). Anal. Calcd
The crystal structure of rat FTase, CVIM peptide, and
farnesyl diphosphate analogue complex (PDB1QBQ)10
was used for the initial structure of protein. We deleted
the CVIM peptide from the complex and added a water
molecule at the position of the sulfur atom of cysteine of
CVIM peptide.
The initial structures of ligands were built using stan-
dard bond lengths and angles, then conformation search
calculations were performed using MMFF force field11
by SPARTAN.12

1382
R. Tanaka et al. / Bioorg. Med. Chem. 15 (2007) 1363–1382
The initial structure of FTase and 1a complex was
selected from the result of the systematic search includ-
ing the ligand flexibility. Then energy-refinement calcu-
lation was performed using MMFF/AMBER force
field using AMBER.13
References and notes
1. (a) Leonard, D. M.. J. Med. Chem. 1997, 40, 2971–2990;
(b) Sebti, S. M.; Hamilton, A. D. Drug Discov. Today
1998, 3, 26–32; (c) Rowinsky, E. K.; Windle, J. J.; Von
Hoff, D. D. J. Clin. Oncol. 1999, 17, 3631–3652.
2. Sepp-Lorenzino, L.; Ma, Z.; Rands, E.; Kohl, N. E.;
Gibbs, J. B.; Oliff, A.; Rosen, N. Cancer Res. 1995, 55,
5302–5309.
The structures of FTase and 19b, 22c complex were
built using the structure of FTase and 1a complex as
template, then energy-refinement calculations were
performed.
3. (a) Kohl, N. E.; Omer, C. A.; Conner, M. W.; Anthony,
N. J.; Davide, J. P.; Desolmes, S. J.; Giuliani, E. A.;
Gomez, R. P.; Graham, S. L.; Hamilton, K.; Handt, L. K.;
Hartman, G. D.; Koblan, K. S.; Kral, A. M.; Miller, P. J.;
Mosser, S. D.; O’Neill, T. J.; Rands, E.; Schaber, M. D.;
Gibbs, J. B.; Oliff, A. Nat. Med. 1995, 1, 792; (b) Liu, M.;
Bryant, M. S.; Chen, J.; Lee, S.; Yaremko, B.; Lipari, P.;
Malkowski, M.; Ferrari, E.; Nielsen, L.; Prioli, N.; Dell,
J.; Sinha, D.; Syed, J.; Korfmacher, W. A.; Nomeir, A. A.;
Lin, C. C.; Wang, L.; Taveras, A. G.; Doll, R. J.; Njoroge,
F. G.; Mallams, A. K.; Remiszewski, S.; Catino, J. J.;
Girijavallabhan, V. M.; Kirschmeier, P.; Bishop, W. R.
Cancer Res. 1998, 58, 4947–4956.
4. Bell, I. M. Exp. Opin. Ther. Patents 2000, 10, 1813–1831.
5. Nara, S.; Tanaka, R.; Eishima, J.; Hara, M.; Takahashi,
Y.; Otaki, S.; Foglesong, R. J.; Hughes, P. F.; Shelley, T.;
Kanda, Y. J. Med. Chem. 2003, 46, 2467–2473.
6. Bhagwatheeswaran, H.; Gaur, S. P.; Jain, P. C. Synthesis
1976, 615–616.
7. Ono, N.; Kamimura, A.; Kaji, A. Synthesis 1984, 226–227.
8. Okada, M.; Iwashita, S.; Koizumi, N. Tetrahedron Lett.
2000, 41, 7047–7051.
9. Hara, M.; Soga, S.; Shono, K.; Eishima, J.; Mizukami, T.
J. Antibiot. 2001, 54, 182–186.
10. Strickland, L. C.; Windsor, T. W.; Syto, R.; Wang, L.;
Bond, R.; Wu, Z.; Schwartz, J.; Le, V. H.; Beese, L. S.;
Weber, C. P. Biochemistry 1998, 37, 16601–16611.
11. Halgren, T. A. J. Comput. Chem. 1996, 17, 490–519.
12. SPARTAN 5.1.1; Wavefunction, Inc.: Irvine, CA, 1998.
13. AMBER: Case, D. A.; Pearlman, D. A.; Caldwell, J. W.;
Cheatham, T. E., III; Ross, W. S.; Simmerling, C. L.;
Darden, T. A.; Merz, K. M.; Stanton, R. V.; Cheng, A. L.;
Vincent, J. J.; Crowley, M.; Ferguson, D. M.; Radmer, R. J.;
Seibel, G. L.; Singh, U. C.; Weiner, P. K.; Kollman, P. A.
AMBER5; University of California: San Francisco, 1997.
5.54. Metabolic stability
An NADPH-generating system consisted of 20 IU/mL
glucose-6-phosphate dehydrogenase, glucose-6-phos-
phate (2.0 mmol/L), and b-NADP+ (8.0 mmol/L) in
MgCl2 (60 mmol/L) aqueous solution. The compound
(0.0010 mg/mL as final concentration) was pre-incu-
bated with mouse liver microsomes [1.0 mg/mL as fi-
nal concentration in potassium phosphate (100 mmol/
L) buffer, pH 7.4] at 37 ꢁC for 5.0 min. The metabol-
ic reaction was initiated by the addition of the
NADPH-generating system, NADPH-generating sys-
tem containing UDPGA (100 mmol/L) or UDPGA
(100 mmol/L) alone, to the mixture. Initial sample
was obtained by terminating the reaction just after
the start of a reaction by the addition of two vol-
umes of internal standard solution in acetonitrile.
The reaction mixtures were incubated for 30 min at
37 ꢁC and the reactions were terminated by the addi-
tion of two volumes of internal standard solution.
The terminated incubation mixtures, as well as stan-
dard curve samples, composed of the same matrix
materials but without NADPH generating system
and UDPGA, were stirred for 0.5 min and centri-
fuged (20,679g, 5.0 min, 4 ꢁC). The supernatants were
analyzed by HPLC [YMC-Pack ODS C18
(4.6 · 150 mm, YMC), 30 ꢁC, eluted with 50 mmol/L
phosphoric acid (50 mmol/L) buffer solution (pH
3.0)/acetonitrile].