Design and synthesis of orally bioavailable inhibitors of inducible nitric oxide synthase. Identification of 2-azabicyclo[4.1.0]heptan-3-imines

Article abstract of DOI:10.1016/S0968-0896(03)00034-8

Further chemical modification of 2-iminopiperidines fused to cyclopropane rings was performed. Optically active isomers 2 and 13 were synthesized and their biological activity was evaluated. Compound 2 exhibited greater potency and more isoform selectivity than enantiomer 13 in the iNOS inhibition assay. One of the gem-chlorines on the fused cyclopropane moiety of 2 was eliminated to produce 3, which showed reduced potency for iNOS inhibition, as well as 4 with an increased potency. The isoform selectivity of 4 was also much higher than that of 3. This was also true for the corresponding methyl derivatives 6-9. The structure-activity relationship (SAR) study and computer aided docking study of the most optimized structure 4 with human iNOS will also be reported.

Full text of DOI:10.1016/S0968-0896(03)00034-8

Bioorganic & Medicinal Chemistry 11 (2003) 1723–1743
Design and Synthesis of Orally Bioavailable Inhibitors of
Inducible Nitric Oxide Synthase. Identification of
2-Azabicyclo[4.1.0]heptan-3-imines
Yasufumi Kawanaka,a,* Kaoru Kobayashi,b Shinya Kusuda,b Tadashi Tatsumi,b
Masayuki Murota,b Toshihiko Nishiyama,b Katsuya Hisaichi,b Atsuko Fujii,b
Keisuke Hirai,b Masao Naka,b Masaharu Komeno,a Yshihiko Odagaki,b
Hisao Nakaib and Masaaki Todab
aFukui Research Institute, Ono Pharmaceutical Co., Ltd., Technoport, Yamagishi, Mikuni, Sakai, Fukui 913-8538, Japan
bMinase Research Institute, Ono Pharmaceutical Co., Ltd., Shimamoto, Mishima, Osaka 618-8585, Japan
Received 18 December 2002; accepted 24 December 2002
Abstract—Further chemical modification of 2-iminopiperidines fused to cyclopropane rings was performed. Optically active isomers
2 and 13 were synthesized and their biological activity was evaluated. Compound 2 exhibited greater potency and more isoform
selectivity than enantiomer 13 in the iNOS inhibition assay. One of the gem-chlorines on the fused cyclopropane moiety of 2 was
eliminated to produce 3, which showed reduced potency for iNOS inhibition, as well as 4 with an increased potency. The isoform
selectivity of 4 was also much higher than that of 3. This was also true for the correspondingmethyl derivatives 6–9. The structure–
activity relationship (SAR) study and computer aided dockingstudy of the most optimized structure 4 with human iNOS will also
be reported.
# 2003 Elsevier Science Ltd. All rights reserved.
Introduction
cause a marked and sustained increase of blood pres-
sure, indicatingthe importance of NO synthesis by the
vascular endothelium in vasoregulation.8 Due to the
importance of cNOS in normal physiology, selective
inhibition of iNOS would be a favorable characteristic
for a drugtaregtingdiseases mediated by over-
production of NO.
Nitric oxide (NO) is an endogenous chemical mediator
that has a role in various physiological processes, such
as endothelium-dependent vasodilatation, cell-to-cell
communication, and the cytotoxicity of phagocytes.1
NO is produced by at least three isoforms of nitric oxide
synthase (NOS), which include two constitutive iso-
forms (neuronal NOS (nNOS) and endothelial NOS
(eNOS)) and an inducible isoform (iNOS).2ꢀ4 These
three isoforms of NOS catalyze NO production via the
oxidation of l-arginine to l-citrulline. The constitutive
isoforms (cNOS) are regulated by calmodulin and by
the Ca2+ concentration. In contrast, the iNOS binds
strongly to calmodulin, rendering its activity indepen-
dent of the Ca2+ concentration.5
Initial reports about NOS inhibitors focused on struc-
tural analogues of the natural substrate l-arginine. In
recent years, a variety of structures includingnon-
amino acid inhibitors such as aminoguanidines,9 iso-
thioureas,10
2-iminopiperidines11
and
2-amino-
pyridines12 have been reported to be selective inhibitors
of iNOS. We have also reported that dihydropyridin-
2-imines are selective inhibitors of iNOS.13 In an effort
to identify non-amino acid iNOS selective inhibitors
startingfrom the chemical modification of 2-imino-
piperidines,11 we discovered 5-methyl-2-azabicyclo
[4.1.0]heptane-3-imine 1 (Chart 1) as a structurally new
chemical lead. In the process of further optimization of
1, introduction of a gem-dichloro group into the cyclo-
propane ringwas found to be effective for maintaining
Administration of the nonselective NOS inhibitor
NG-methyl-l-arginine (l-NMA)6,7 has been shown to
*Correspondingauthor. Tel.: +81-776-82-6161; fax: +81-776-82-
8420; e-mail: kawanaka@ono.co.jp
0968-0896/03/$ - see front matter # 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0968-0896(03)00034-8

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Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
Chart 1. Molecular design of 2-iminopiperidine fused to a substituted cyclopropane.
potent iNOS inhibition while improvingisoform selec-
tivity, as illustrated in Chart 1. Further chemical modifi-
cation of the cyclopropane moiety in the bicyclic system
was carried out with the hope of identifyinga new selec-
tive iNOS inhibitor. Here we report on the discovery
process for a selective inhibitor of iNOS, compound 4.
with p-methoxybenzylamine gave 22 and 23 at a good yield
(22: 97% yield, 23: 96% yield). Oxidation of 22 and 23
produced the cyclic hemiacetals 24 and 25 at a yield of 57
and 43%, respectively. Acidic dehydration of 24 and 25
produced 26 and 27 at a yield of 60 and 65%, respectively.
As described in Scheme 2, cyclopropanation of 26 with
CHCl3 under alkaline conditions resulted in 28a-anti
(the newly introduced cyclopropane moiety and the
methyl moiety showed anti-stereochemistry: 62% yield)
and 28a-syn (the newly introduced cyclopropane moiety
and the methyl moiety showed syn-stereochemistry:
12% yield), which were separated by column chroma-
tography on silica gel. Cyclopropanation of 26 with
CHBr3 gave 28b-anti (42% yield) and 28b-syn (12%
yield) as a separable mixture, while cyclopropanation of
26 with FCHBr2 gave 28c-anti (36% yield) and 28c-syn
(16% yield) as a separable mixture. Treatment of 28b-
anti with n-BuLi, followed by trappingwith MeI, resul-
ted in 28d-anti (91% yield), stereochemistry of which
was determined by the NOE correlation between the
7-methyl and 5-H as described in Figure 1. Deprotec-
tion of the p-methoxybenzyl moiety of 28a-anti with
Chemistry
Synthesis of all the test compounds presented in Tables 1
and 2 are outlined in Schemes 1–9. Preparation of
enantiomeric isomers 26 and 27 is described in Scheme 1.
The optically active startingmaterial 19, which was
obtained by chemicoenzymatic hydrolysis of the corres-
pondingsymmetrical diester, 14a was converted to 2014b and
d-lactone 2114c by reduction with diborane (94% yield) or
reduction with lithium borohydride followed by acid treat-
ment (90% yield), respectively. Aminolysis of 20 and 21
Table 1. Inhibitory activities of 2-azabicyclo[4.1.0]heptan-3-imines
against hiNOS, heNOS and their isoform selectivityf
.
Structure
Compound
hiNOS
heNOS Selectivity
BF3 OEt2-anisole was followed by removal of the
IC50 (mM) IC50 (mM) heNOS/
hiNOS
chlorines of 29a through reduction with tin hydride,
resultingin a separable mixture consistingof an exo-
isomer 29e (28% yield) and an endo-isomer 29f (53%
2 R1=R2=Cla
3 R1=Cl, R2=Ha
4 R1=H, R2=Cla
5 R1=H, R2=Fa
6 R1=R2=Meb
7 R1=Me, R2=Hb
8 R1=H, R2=Meb
9 R1=H, R2=Mea
10 R1=H, R2=Etb
11 R1=vinyl, R2=Hb
12 R1=H, R2=n-Bub
13 R1=R2=Cla
14 R1=Cl, R2=Ha
15 R1=H, R2=Cla
16 R1=H, R2=Fa
0.020
0.40
0.011
0.031
0.22
0.39
0.093
0.040
0.17
1.8
0.16
0.62
0.12
0.10
0.77
0.68
0.57
0.53
0.59
NTc
NTc
2.20
(>5)d
0.34
8.0
1.6
10.9
3.2
3.5
1.7
6.1
.
yield). Deprotection of 28c-anti with BF3 OEt2-anisole
Table 2. Inhibitory activities of 17 and 18 against hiNOS, heNOS
and their isoform selectivityd
13.3
3.5
Structure
Compound
hiNOS
IC50 (mM) IC50 (mM)
heNOS
Selectivity
heNOS/
hiNOS
NTc
NTc
8.8
3.6
0.25
0.51
0.043
NDe
7.9
17b
0.90
NTc
NTc
NTc
18a
>50
NTc
0.060
0.32
5.3
aChiral.
bRacemic.
aChiral.
cNT: not tested.
bRacemic.
dLess than 50% inhibition at 5 mM.
cNT: not tested.
dAll compounds were prepared as their hydrochlorides.
eND: not determined.
fAll compounds were prepared as their hydrochlorides.

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1725
ꢁ
.
Scheme 1. Synthesis of optically active analogues 26 and 27. Reagents: (a) BH3 SMe2, THF, 0 C (94%); (b) LiOH aq room temperature; (c) LiBH4,
ꢁ
.
THF, 50 C; (d) p-TsOH H2O, benzene, reflux (90% in three steps); (e) PMBNH2, toluene, reflux (22: 97%, 23: 96%); (f) SO3 pyridine, DMSO,
.
Et3N, room temperature (24: 57%, 25: 43%); (g) p-TsOH H2O, toluene, reflux (26: 60%, 27: 65%).
.
.
Scheme 2. Synthesis of 2–5 and 18. Reagents: (a) CHCl3, 50% NaOH aq, aliquat-336 (28a-anti: 62%, 28a-syn: 12%); (b) CHBr3, 50% NaOH aq,
aliquat-336 (28b-anti: 42%, 28b-syn: 12%);ꢁ(c) FCHBr2, 50% NaOH aq, PhCH2NEt3Cl (2ꢁ8c-anti: 36%, 28c-syn: 16%); (d) MeI, n-BuLi, THF,
ꢁ
.
ꢀ78 C (91%); (e) BF3 OEt2, anisole, 100 C (75–88%); (f) Ph3SnH, AIBN, benzene, 80 C (29e: 28%, 29f: 53%; 29g: 41%, 29h: 44%); (g)
Et3O+BFꢀ4 , CH2Cl2; (h) NH3, EtOH; (i) PhCH2NH2, EtOH; (j) 4 M HCl dioxane (28–57% in three steps).
was followed by selective removal of the bromine of 29c
through reduction with tin hydride, resulting in a
separable mixture consistingof an exo-isomer 29g
(41% yield) and an endo-isomer 29h (44% yield).
Compounds 29a, 29e, 29f and 29h were then converted
to their correspondingamidines 2–5 and 18 by the usual
procedure for amidine synthesis.
Preparation of compound 9 is described in Scheme 3.
Treatment of 28d-anti with t-BuOK gave 3015 at a 35%

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Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
ꢁ
.
Scheme 3. Synthesis of 9. Reagents: (a) t-BuOK, THF, room temperature (35%); (b) 10% Pd/C, EtOAc (71%); (c) BF3 OEt2, anisole, 100 C (57%);
(d) Et3O+BFꢀ4 , CH2Cl2; (e) NH3, EtOH; (f) 4 M HCI dioxane (4% in three steps).
Scheme 4. Synthesis of racemic analogues 37 and 38. Reagents: (a) PMBNH2, toluene, room temperature; (b) Ac2O, Et3N, 80 ꢁC (95% in two steps);
ꢁ
ꢁ
.
(c) NaBH4, EtOH. 0 C (95%); (d) MeLi, THF, ꢀ78 C (99%); (e) p-TsOH H2O, toluene, reflux (37: 60%, 38: 99%).
Scheme 5. Synthesis of 6–8 and 12. Reagents: (a) CHBr3, 50% NaOH aq, aliquat-336 (39a-anti: 38%, 39a-syn: 13%); (b) CuSCN, MeLi, Et2O–
HMPA, then MeI (73%); (c) CuSCN, MeLi Et2O–HMPA, then NH4Cl aq (39%); (d) MeI, n-BuLi, THF, ꢀ78 ꢁC (73%); (e) CuI, n-BuLi, THF,
ꢁ
.
then 4 M HCl dioxane (36%); (f) CuSCN, n-BuLi, Et2O–HMPA, then NH4Cl aq (39f-anti: 18%, 39g-anti: 39%); (g) BF3 OEt2, anisole, 100 C (47–
78%); (h) Et3O+BF4ꢀ, CH2CI2; (i) NH3, EtOH; (j) 4 M HCl dioxane (25–74% in three steps).
yield. Hydrogenation of 30 only afforded endo-isomer
31 at a 71% yield. Deprotection of 31 gave 32 (57%
yield), which was converted to 9 accordingto the usual
procedure.
(95% yield), while methylation of 34 with methyl
lithium provided 36 (99% yield). Dehydration of 35 and
36 with p-toluenesulfonic acid produced 37 and 38 (60
and 99% yield, respectively).
Preparation of racemic compounds 37 and 38 is out-
lined in Scheme 4. Aminolysis of 33 with p-methoxy-
benzylamine, followed by formation of an imide with
acetic anhydride, resulted in 34 (95% yield). Partial
reduction of 34 with sodium borohydride afforded 35
Synthesis of racemic compounds 6–8 and 12 is outlined
in Scheme 5. Cyclopropanation of racemic compound
37 with CHBr3 provided 39a-anti (38% yield) and 39a-
syn (13% yield). Dimethylation of 39a-anti with methyl
lithium, followed by trappingwith methyl iodide, afforded

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1727
Scheme 6. Synthesis of 10. Reagents: (a) n-Bu3SnH, toluene, ꢀ10 ꢁC (78%, 41a:41b=1:3); (b) CuI, n-BuLi, THF, then EtI (42a: 20%, 42b: 18%); (c)
BF3 OEt2, anisole, 100 C (39%); (d) Et3O+BF4ꢀ, CH2Cl2; (e) NH3, EtOH; (f) 4 M HCl dioxane (88% in three steps).
ꢁ
.
Scheme 7. Synthesis of 11. Reagents: (a) BF3 OEt2, anisole, 100 C (91%); (b) CuI, CH2=CHMgBr, Et2O–THF (59%); (c) Et3O+BF4ꢀ, CH2Cl2; (d)
ꢁ
.
NH3, EtOH; (e) 4 M HCl dioxane (6% in three steps).
Scheme 8. Synthesis of 13–16. Reagents: (a) CHCl3, 50% NaOH aq, aliguat-336 (46a-anti: 53%, 46a-syn: 14%); (b) ClCHBr2, 50% NaOH aq,
ꢁ
.
PhCH2NEt3Cl (46b-anti: 64%, 46b-syn: 18%); (c) FCHBr2, 50% NaOH aq, PhCH2NEt3Cl (46c-syn: 15%); (d) BF3 OEt2, anisole, 100 C (75–96%);
(e) (TMS)3SiH, BEt3, toluene, room temperature (47c: 44%, 47d: 28%; 48d: 42%, 48e: 54%); (f) Ph3SnH, AIBN, benzene, 80 ꢁC (48g: 22%); (g)
Et3O+BFꢀ4 , CH2Cl2; (h) NH3, EtOH; (i) 4 M HCl dioxane (9–48% in three steps).

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Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
Scheme 9. Synthesis of 17. Reagents: (a) CHꢁBr3, 50% NaOH aq, aliquat-336 (49a-anti: 26%, 49a-syn: 7%); (b) CuSCN, MeLi, Et2O–HMPA, then
NH4Cl aq (65%); (c) BF3 OEt2, anisole, 100 C (90%); (d) Et3O+BF4ꢀ, CH2Cl2; (e) NH3, EtOH; (f) 4 M HCl dioxane (48% in three steps).
.
conditions afforded 44 (91% yield), which was con-
verted to the vinyl intermediate 45 (59% yield). Appli-
cation of the usual amidine synthesis procedure to 45
gave 11.
The preparation of compounds 13–16 is outlined in
Scheme 8. The optically active startingmaterial 27 was
converted to a separable mixture of 46a-c-anti and46a-c-
syn by a cyclopropanation procedure similar to that
described above. Deprotection of 46a-anti and 46b-
Figure 1. The NOE correlation between 7-methyl and 5-H in 28d–anti.
anti gave 47a and 47b, respectively. Reductive debro-
mination of 47b afforded endo-isomer 47c and exo-
isomer 47d at a yield of 44 and 28%, respectively.
Compounds 47a and 47c were then converted to
amidine analogues 13 and 14, respectively, according
to the usual procedure.
39b-anti at a 73% yield. Monomethylation of 39a-anti
was carried out with methyl lithium, followed by trap-
pingof the anion thus formed with aqueous ammonium
chloride, to give exo-isomer 39c-anti at a 39% yield.
Treatment of 39a-anti with n-BuLi, followed by trap-
pingwith methyl iodide, resulted in 39d-anti (73%
yield), the remainingbromine of which was successfully
removed to gave 39e-anti at a 36% yield. Treatment of
39a-anti with n-BuLi in the presence of CuSCN pro-
duced a separable mixture of exo-isomer 39f-anti and
endo-isomer 39g-anti at a yield of 18 and 39%, respec-
tively. Deprotection of the p-methoxybenzyl moiety of
Deprotection of 46b-syn and 46c-syn afforded 48b and
48c, respectively. Reductive debromination of 48b gave
a separable mixture of exo-isomer 48d and endo-isomer
48e at a yield of 42 and 54%, respectively, while reduc-
tive debromination of 48c resulted in a separable mix-
ture of 48f and 48g. Compounds 48e and 48g were then
converted to 15 and 16.
.
39b-anti, 39c-anti, 39e-anti and 39g-anti with BF3 OEt2-
anisole provided 40b, 40c, 40e and 40g, respectively.
Compounds 40b, 40c, 40e and 40g were then converted
to their correspondingamidines 6–8 and 12 by the usual
amidine synthesis procedure.
Preparation of compound 17 is described in Scheme 9.
The racemic startingmaterial 38 was transformed to
49a-anti (26% yield) and 49a-syn (7% yield) by the
usual cyclopropanation procedure, as described above.
Methylation of 49a-anti provided 49b-anti at a 65%
yield, while the deprotection of 49b-anti afforded 50
(90% yield). Compound 50 was then converted to ami-
dine analogue 17 by the usual procedure.
Preparation of compound 10 is described in Scheme 6.
Reductive removal of one of the bromines of 39a-anti
afforded exo-isomer 41a and endo-isomer 41b
(41a:41b=1:3), respectively. Ethylation of 41a and 41b
gave a separable mixture of 42a and 42b at a yield of 20
and 18%, respectively. Deprotection of 42b resulted in
43, which was converted to 10 accordingto the usual
procedure. Compound 11 was prepared as described in
Scheme 7. Deprotection of 39a-anti under the usual
Because of its stereochemical complexity, the structure
of 4 was investigated by X-ray crystallographic analysis
in addition to the usual spectral analyses.16 As shown in
Figure 2, the results of X-ray crystallography support
the structure described in Table 1.

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1729
selectivity because of increased potency in hiNOS inhi-
bition without a change of heNOS inhibition, as illu-
strated in the conversion of 5 to 4 or 16 to 15.
The effect of addingan alkyl substituent to the fused
cyclopropane ringwas also studied. Replacement of the
gem-dichloro group with a gem-dimethyl group gave 6,
which showed a 10-fold reduction in inhibitory activity
against hiNOS and reduced isoform selectivity. Mono-
methyl analogues 7 and 8 showed less inhibitory activity
against both isoforms as well as less isoform selectivity.
Compound 9, the optically active form of the more
potent monomethyl analogue 8, was prepared and eval-
uated. The potency of hiNOS inhibition by 9 was nearly
2-fold greater than by its racemic mixture 8, while
heNOS inhibition remained unchanged. As a result, the
isoform selectivity of 9 was improved when compared
with that of 8. In order to assess the possibility of fur-
ther chemical modifications to the newly introduced
methyl group, compounds 10–12 were prepared and
biologically evaluated. Replacement of the methyl
group in 8 with an ethyl group afforded 10, which
showed a 1.8-fold reduction in the potency of hiNOS
inhibition but no change of heNOS inhibition. As a
result, its isoform selectivity was nearly half that of 8.
Introduction of a higher alkyl group at the same posi-
tion did not improve hiNOS inhibition irrespective of
the stereochemistry, as illustrated in 11 and 12.
Figure 2. X-ray crystallographic analysis of 4.
Results and Discussion
As described in previous papers,13,17 compounds 2–18
were synthesized and biologically evaluated for their
ability to inhibit the two isoforms of human NOS:
hiNOS and heNOS. Isoform selectivity was determined
from the ratio of the IC50 value for these two isoforms
(i.e., heNOS/hiNOS).
Introduction of another methyl group at one of the ring
junctions in 7 provided 17, which showed more than
2-fold reduction in the potency of hiNOS inhibition.
N-Benzylation of the 3-imino group of 4 afforded 18,
which showed a marked reduction of hiNOS inhibition.
The gem-dichloro analogues 2 and 13 were initially
synthesized as optically active forms and evaluated. The
anti-isomer 2 was more potent than 13 in the hiNOS
inhibition assay and showed an 8-fold greater isoform
selectivity.
Amongthe compounds tested, 2 and 417 were selected
for further biological and pharmacodynamic evaluation
because of their excellent biological profile, including
safety data. Additional in vitro activities of 2 and 4 as
well as l-NMMA and aminoguanidine, were evaluated,
Then biological evaluation of their optically active
compounds 3 and 4 was carried out. The biological
profile of optically active compound 4 was improved in
all evaluations (hiNOS inhibition, heNOS inhibition
and isoform selectivity) as compared with that of com-
pound 3. The fluoro analogue 5, which corresponds to
4, was prepared as an optically active form and eval-
uated. Replacement of the chloro group in 4 with a
fluoro group gave 5, which showed a slight reduction in
the potency of hiNOS inhibition and slightly stronger
heNOS inhibition. As a result, the isoform selectivity of
fluoro analogue 5 was nearly 3-fold less than that of 4.
includingthe IC
for mouse iNOS, the Ki values for
50
hiNOS and heNOS and the isoform selectivity deter-
mined from the two Ki values (Table 3).17 The Ki value
of 4 indicated that it was the most potent iNOS inhibi-
tors amongthe compounds tested. Compound 4 also
showed more iNOS selectivity than compound 2. A
non-selective inhibitor, l-NMMA, showed eNOS selec-
tivity. Aminoguanidine showed far less activity than the
other three inhibitors, while it showed quite a good
iNOS selectivity (heNOS/hiNOS=4.8-fold). In order to
evaluate the above-mentioned two compounds 2 and 4
for their ability to inhibit iNOS in vivo, mice were
administered the two test compounds subcutaneously
(sc) at 3 h after lipopolysaccharide (LPS) injection.
Then plasma NOx accumulation from 3 to 6 h after LPS
injection was determined. As shown in Table 3, com-
pounds 2 and 4 inhibited NOx accumulation in plasma
and their ID50 values were 0.023 and 0.010 mg/kg, sc,
respectively. To assess the acute toxicity of each com-
pounds, the maximum tolerated dose (MTD) was
determined. As shown in Table 3, the MTD values of 2
and 4 were 20 and 30 mg/kg, respectively, when a single
Compound 14, the enantiomer of 4, was also prepared
and biologically evaluated, but it showed reduced inhi-
bitory activity against both isoforms when compared
with 4. Compounds 15 and 16 were also evaluated. The
chloro analogue 15 demonstrated slightly stronger
hiNOS inhibition as compared with 16, while its heNOS
inhibitory activity was unchanged. As a result, improved
isoform selectivity was obtained by replacement of the
fluoro group on the cyclopropane ring with a chloro
group, as illustrated in the conversion of 16 to 15.
Replacement of the fluoro group on the fused cyclo-
propane ringwith a chloro group increased the isoform

1730
Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
Table 3. Biological profiles of 2 and 4
IC50
Kt valuesa
iNOS selectivityb
heNOS/hiNOS
Acute toxicity
Mouse NOx
miNOS
hiNOS
heNOS
MTD (mg/kg, iv)
ID50 (mg/kg, sc)
2
4
5.6 (nM)
4.0 (nM)
3.5 (mM)
19.6 (mM)
5.20 (nM)
1.88 (nM)
847 (nM)
39.9 (mM)
43.0 (nM)
18.8 (nM)
250 (nM)
190 (mM)
8.3
10.0
0.3
20
30
3000
NTc
0.023
0.010
26
NTc
l-NMMA
Aminoguanidine
4.8
aEach Ki value was determined from three separate experiments..17
bThe iNOS selectivity is calculated by the formulation; Ki of heNOS/Ki of hiNOS.
cNT: not tested.
Figure 3. Stereo view of a dockingmodel of compound 4 with human iNOS (PDB code 2NSI).
intravenous (iv) dose was given to normal mice. The
MTD values of these two compounds indicated a much
higher potency than that of l-NMMA (3000 mg/kg),
but the MTD/ID50 ratio for NOx accumulation in mice
was 870 for 2 and 3000 for 4. Oral bioavailability of 2
and 4 was excellent in rats (2: 52–100% and 4: 62–83%)
and dogs (2: 57% and 4: 100%).
Computer aided dockingstudy of 4 with the enzyme
was also performed. Accordingly, these two compounds
could be useful tools to help elucidate the role of iNOS
in various disease states and may also have potential as
novel drugs.
Experimental
Figure 3 demonstrates a stereo view of a dockingstudy
of 4 with human iNOS (PDB code 2NSI). This study
was performed usingInsihgt II/Discover molecular
modelingpackaeg (Accelrys, San Dieog, CA) with
CVFF force field on a SGI Octane 2 workstation with
R12000 processors. Clear-cut interaction between the
basic amidine moiety with acidic Glu377 residue and
insertion of the chlorine moiety into a small pocket of
the enzyme were observed.
General directions
Analytical samples were homogeneous as confirmed by
TLC, and afforded spectroscopic results consistent with
the assigned structures. All 1H NMR spectra were
obtained on a Varian Gemini-200, VXR-200s spectro-
meter. Fast atom bombardment mass spectra (FAB-
MS) and electron ionization (EI) were obtained on a
Jeol JMS-DX303HF or PerSeptive Voyager Elite spec-
trometer. Atmospheric pressure chemical ionization
(APCI) was determined on a Hitachi M1200H spectro-
meter. Matrix assisted laser desorption ionization-time
of flight high-resolution mass spectra (MALDI-TOF
HR-MS) were obtained on a PerSeptive Voyager Elite
spectrometer. IR spectra were measured on a Perkin–
Elmer FT-IR 1760X or Jasco FT/IR-430 spectrometer.
Elemental analyses for carbon, hydrogen and nitrogen
were carried out on a Perkin–Elmer PE2400 SeriesII
CHNS/O analyzer. Optical rotations were measured
usinga Jasco DIP-1000 polarimeter. Meltingpoints
Summary
In summary, we explored the SAR for a series of 2-imi-
nopiperidines fused to a substituted cyclopropane ring
and obtained a significant increase of hiNOS inhibition
as well as better isoform selectivity in both in vitro and
in vivo experiments. Amongthe compounds tested,
2
and 4 showed excellent profiles in both the biological and
pharmacodynamic evaluations. The structure of 4 was
finally determined by X-ray crystallographic analysis.

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1731
(mp) were determined by Yanaco micro meltingpoint
apparatus MP-500D and are uncorrected. Column
chromatography was carried out on silica gel (Merck
silica gel 60 (0.063–0.200 mm) or Wako Gel C200 or
Fuji Silysia FL60D). Thin layer chromatography was
performed on silica gel (Merck TLC plate, silica gel 60
F254).
mixture was diluted with EtOAc and washed sequen-
tially with saturated aqueous sodium bicarbonate and
brine. The organic layer was dried over magnesium sul-
fate and evaporated under reduced pressure. The resi-
due was purified by column chromatography on silica
gel (EtOAc/n-hexane, 1/5–1/3) to afford 26 as a pale
yellow oil (1.9 g, 60%). TLC Rf 0.55 (EtOAc/n-hexane,
25
D
1/1); optical rotation ½aŠ ꢀ103.5 (c 1.05, CHCl3); IR
(neat) 3404, 2957, 2836, 1667, 1585, 1514, 1456, 1412,
1386, 1303, 1248, 1212, 1176, 1148, 1105, 1034, 965,
Starting materials
Compounds 19, 20 and 21 were synthesized according
to the literature14aꢀc. Compound 33 is commercially
available.
946, 877, 822, 764, 718, 580, 519 cmꢀ1 1H NMR
;
(200 MHz, CDCl3) d 7.22–7.13 (m, 2H), 6.89–6.80 (m,
2H), 5.96 (dd, J=7.8, 1.4 Hz, 1H), 5.06–4.99 (m, 1H),
4.64 (d, J=14.6 Hz, 1H), 4.57 (d, J=14.6 Hz, 1H), 3.79
(s, 3H), 2.63 (dd, J=17.6, 6.4 Hz, 1H), 2.68–2.49 (m,
1H), 2.29 (dd, J=17.6, 11.4 Hz, 1H), 1.05 (d, J=6.8 Hz,
3H); MS (APCI, Pos.) m/z 232 (M+H)+; HR-MS
(MALDI-TOF, Pos.) calcd for C14H17N1O2+ H+:
232.1338; found: 232.1358.
(3R)-5-Hydroxy-N-(4-methoxybenzyl)-3-methylpentan-
amide (22). To a stirred solution of 20 (5.0 g, 34 mmol)
in toluene (60 mL) was added p-methoxybenzylamine
(4.6 mL, 35.2 mmol) and stirred for 3 h at reflux tem-
perature. After coolingup to room temperature, the
reaction mixture was evaporated under reduced pres-
sure. The resultingresidue was purified by column
chromatography on silica gel (EtOAc to EtOAc/MeOH,
8/1) to afford 22 as a pale yellow solid (8.3 g, 97%).
(3S)-5-Hydroxy-N-(4-methoxybenzyl)-3-methylpentan-
amide (23). Compound 23 was prepared from 21 in
96% yield accordingto the same procedure as described
TLC Rf 0.20 (CHCl3/MeOH, 10/1); optical rotation
½aŠ ꢀ4.1 (c 1.14, CHCl3); IR (KBr) 3277, 2930, 1717,
;
1034, 816, 555, 526, 419 cmꢀ1 1H NMR (200 MHz,
CDCl3) d 7.25–7.16 (m, 2H), 6.91–6.82 (m, 2H), 5.98
(brs, 1H), 4.36 (d, J=5.6 Hz, 2H), 3.79 (s, 3H), 3.66 (t,
J=6.0 Hz, 2H), 2.60 (brs, 1H), 2.29–2.00 (m, 3H), 1.63–
1.30 (m, 2H), 0.98 (d, J=6.4 Hz, 3H); MS (APCI, Pos.)
m/z 252 (M+H)+; HR-MS (MALDI-TOF, Pos.) calcd
for C14H21N1O3+ H+: 252.1560; found: 252.1583.
for the preparation of 22 from 20. Pale yellow solid;
25
D
1637, 1549, 1514, 1459, 1378, 1302, 1250, 1177, 1111,
25
D
TLC Rf 0.28 (EtOAc); optical rotation ½aŠ +4.0 (c
1.05, CHCl3); IR (KBr) 3276, 2930, 2312, 1718, 1637,
1549, 1514, 1460, 1379, 1302, 1250, 1177, 1111, 1052,
1035, 816, 760, 594, 526, 473, 444 cmꢀ1 1H NMR
;
(200 MHz, CDCl3) d 7.21 (d, J=8.8 Hz, 2H), 6.87 (d,
J=8.8 Hz, 2H), 5.84 (brs, 1H), 4.38 (d, J=5.6 Hz, 2H),
3.80 (s, 3H), 3.70–3.60 (m, 2H), 2.30–2.05 (m, 3H),
1.60–1.50 (m, 2H), 0.99 (d, J=6.6 Hz, 3H); MS (APCI,
Pos.) m/z 252 (M+H)+; HR-MS (MALDI-TOF, Pos.)
calcd for C14H21N1O3+ H+: 252.1600; found:
252.1585.
(4R)-6-Hydroxy-1-(4-methoxybenzyl)-4-methylpiperidin-
2-one (24). To a stirred solution of 22 (6.1 g, 24.3
mmol) in DMSO (35 mL) were added Et3N (17 mL, 122
mmol) and sulfur trioxide pyridine complex (19.4 g,
121.9 mmol) at 0 ꢁC under an argon atmosphere. After
stirringfor 4.5 h at room temperature, the reaction was
quenched with water. Then the mixture was treated with
1 M HCl and extracted with EtOAc. The organic layer
was washed with brine, dried over magnesium sulfate
and evaporated under reduced pressure. The residue
was purified by column chromatography on silica gel
(EtOAc/n-hexane, 1/1) to afford 24 as a white crystal
(3.3 g, 57%). TLC Rf 0.50 (EtOAc); mp 88–89 ꢁC; IR
(KBr) 3258, 2951, 2841, 1627, 1514, 1479, 1439, 1413,
1312, 1297, 1281, 1249, 1182, 1170, 1100, 1065, 1026,
974, 931, 903, 838, 818, 768, 742, 701, 619, 582, 528
cmꢀ1; 1H NMR (200 MHz, CDCl3) d 7.26–7.18 (m, 2H),
6.89–6.80 (m, 2H), 5.02–4.90 (m, 2H), 4.32 (d, J=14.6
Hz, 1H), 3.79 (s, 3H), 2.71–2.54 (m, 2H), 2.44–2.20 (m,
1H), 2.09–1.86 (m, 2H), 1.47 (ddd, J=13.6, 12.8, 3.6
Hz, 1H), 1.01 (d, J=6.6 Hz, 3H); MS (APCI, Pos.) m/z
250 (M+H)+; HR-MS (MALDI-TOF, Pos.) calcd for
C14H19N1O3+ H+: 250.1443; found: 250.1432.
(4S)-6-Hydroxy-1-(4-methoxybenzyl)-4-methylpiperidin-
2-one (25). Compound 25 was prepared from 23 in
43% yield accordingto the same procedure as described
for the preparation of 24 from 22. White crystal; TLC
Rf 0.50 (EtOAc); mp 136–137 ꢁC; IR (KBr) 3260, 2951,
1627, 1514, 1479, 1439, 1249, 1182, 1100, 1065, 1026,
1
974, 903, 838, 818, 742, 620, 582, 528 cmꢀ1; H NMR
(200 MHz, CDCl3) d 7.23 (d, J=8.8 Hz, 2H), 6.85 (d,
J=8.8 Hz, 2H), 4.95 (d, J=14.2 Hz, 1H), 5.00–4.88 (m,
1H), 4.34 (d, J=14.2 Hz, 1H), 3.79 (s, 3H), 2.70–2.52
(m, 1H), 2.40–2.18 (brs, 1H), 2.10–1.80 (m, 2H), 1.65–
1.36 (m, 2H), 1.00 (d, J=6.6 Hz, 3H); MS (APCI, Pos.)
m/z 250 (M+H)+; HR-MS (MALDI-TOF, Pos.) calcd
for C14H19N1O3+ H+: 250.1443; found: 250.1469.
(4R)-1-(4-Methoxybenzyl)-4-methyl-3,4-dihydropyridin-
2-one (27). Compound 27 was prepared from 25 in
65% yield accordingto the same procedure as described
for the preparation of 26 from 24. Pale yellow oil; TLC
25
D
Rf 0.55 (EtOAc/n-hexane, 1/1); optical rotation ½aŠ
+104.1 (c 1.06, CHCl3); IR (neat) 2958, 2836, 1668,
1612, 1585, 1513, 1442, 1408, 1384, 1304, 1248, 1212,
1176, 1147, 1109, 1034, 946, 822, 717, 643, 572, 518
cmꢀ1; H NMR (200 MHz, CDCl3) d 7.17 (d, J=8.8
(4S)-1-(4-Methoxybenzyl)-4-methyl-3,4-dihydropyridin-
2-one (26). A solution of 24 (3.4 g, 13.6 mmol) and
p-toluenesulfonic acid monohydrate (150 mg, 0.8 mmol)
in toluene (70 mL) was stirred for 1.5 h at reflux tempera-
ture. After coolingat room temperature, the reaction
1
Hz, 2H), 6.84 (d, J=8.8 Hz, 2H), 5.96 (dd, J=7.7, 1.5
Hz, 1H), 5.03 (dd, J=7.7, 3.4 Hz, 1H), 4.61 (d, J=2.6
Hz, 2H), 3.79 (s, 3H), 2.70–2.50 (m, 2H), 2.40–2.15 (m,

1732
Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1H), 1.05 (d, J=6.8 Hz, 3H); MS (APCI, Pos.) m/z 232
(M+H)+; HR-MS (MALDI-TOF, Pos.) calcd for
C14H17N1O2+ H+: 232.1338; found: 232.1317.
by column chromatography on silica gel (EtOAc/n-hex-
ane, 1/5 to 1/3) to afford 28b-anti as a pale yellow oil
(1.5 g, 42%) and 28b-syn as a pale yellow oil (416 mg,
12%).
General procedure for cyclopropanation
28b-anti: TLC Rf 0.38 (EtOAc/n-hexane, 1/2); optical
rotation ½aŠ +15.1 (c 1.00, CHCl3); IR (neat) 2961,
25
D
2959, 2836, 1667, 1513, 1460, 1415, 1248, 1178, 1032,
(1S,5S,6R)-7,7-Dichloro-2-(4-methoxybenzyl)-5-methyl-
2-azabicyclo[4.1.0]heptan-3-one (28a-anti) and (1R,5S,6S)-
7,7-dichloro-2-(4-methoxybenzyl)-5-methyl-2-azabicyclo
[4.1.0]heptan-3-one (28a-syn). To a stirred solution of
26 (1.2 g, 5.2 mmol) in CHCl3 (12.5 mL, 156 mmol)
were added aliquat-336 (0.1 mL, 0.22 mmol) and 50%
aqueous sodium hydroxide (2.6 g) under an argon
atmosphere. The reaction mixture was stirred for 22 h at
room temperature. After completingthe reaction, the
mixture was treated with saturated aqueous ammonium
chloride and extracted with Et2O. The organic layer was
washed with brine, dried over magnesium sulfate and
evaporated under reduced pressure. The residue was
purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:5–1:3) to afford 28a-anti as a pale
yellow oil (1.0 g, 62%) and 28a-syn as a pale yellow oil
(196 mg, 12%).
1
845, 764 cmꢀ1; H NMR (200 MHz, CDCl3) d 7.30 (d,
J=8.4 Hz, 2H), 6.90 (d, J=8.4 Hz, 2H), 5.47 (d,
J=14.8 Hz, 1H), 3.81 (d, J=14.8 Hz, 1H), 3.81 (s, 3H),
2.98 (d, J=9.8 Hz, 1H), 2.45–2.00 (m, 3H), 1.77 (dd,
J=9.8, 5.4 Hz, 1H), 1.26 (m, 3H); MS (APCI, Pos.) m/z
404 (M+H, 79Br, 81Br)+; HR-MS (MALDI-TOF, Pos.)
calcd for C15H17Br2N1O2+ H+: 401.9704; found:
401.9725.
28b-syn: TLC Rf 0.18 (EtOAc/n-hexane, 1/2); optical
25
D
rotation ½aŠ +31.6 (c 0.63, CHCl3); IR (neat) 2963,
2835, 1651, 1512, 1456, 1415, 1388, 1302, 1248, 1176,
1107, 1033, 849, 773 cmꢀ1; 1H NMR (200 MHz, CDCl3)
d 7.32 (d, J=8.6 Hz, 2H), 6.92 (d, J=8.6 Hz, 2H), 4.93
(d, J=14.2 Hz, 1H), 4.54 (d, J=14.2 Hz, 1H), 3.82 (s,
3H), 3.27 (d, J=10.2 Hz, 1H), 2.50–2.43 (m, 3H), 2.20–
2.10 (m, 1H), 1.28 (d, J=6.2 Hz, 3H); MS (APCI, Pos.)
m/z 404 (M+H, 79Br, 81Br)+; HR-MS (MALDI-TOF,
Pos.) calcd for C15H17Br2N1O2+ H+: 401.9704; found:
401.9708.
28a-anti: TLC Rf 0.36 (EtOAc/n-hexane, 1/2); optical
25
D
2836, 1668, 1612, 1513, 1443, 1415, 1384, 1303, 1248,
rotation ½aŠ +26.3 (c 1.00, CHCl3); IR (neat) 2963,
1176, 1079, 1033, 890, 854, 822, 766, 731, 575, 512 cmꢀ1
;
1H NMR (200 MHz, CDCl3) d 7.32–7.24 (m, 2H), 6.93–
6.85 (m, 2H), 5.44 (d, J=14.4 Hz, 1H), 3.81 (s, 3H),
3.81 (d, J=4.4 Hz, 1H), 2.95 (d, J=9.8 Hz, 1H), 2.39–
2.02 (m, 3H), 1.76 (dd, J=9.8, 5.0 Hz, 1H), 1.25 (d,
J=6.4 Hz, 3H); MS (APCI, Pos.) m/z 316 (M+H, 35Cl,
37Cl)+, 314 (M+H, 35Cl, 35Cl)+; HR-MS (MALDI-
TOF, Pos.) calcd for C15H17Cl2N1O2+ H+: 314.0715;
found: 314.0695.
(1S,5S,6R)-7-Bromo-7-fluoro-2-(4-methoxybenzyl)-5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (28c-anti) and
(1R,5S,6S)-7-bromo-7-fluoro-2-(4-methoxybenzyl)-5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (28c-syn). To a
stirred solution of 26 (200 mg, 0.86 mmol) in CH2Cl2 (1
mL) were added FCHBr2 (0.33 mL, 4.3 mmol), benzyl-
triethylammonium chloride (5.9 mg, 0.03 mmol) and
50% aqueous sodium hydroxide (1 g) at 0 ꢁC under an
argon atmosphere. The reaction mixture was warmed
up to room temperature and stirred for 48 h. After
completingthe reaction, the mixture was treated with
water and extracted with Et2O. The organic layer was
washed with brine, dried over magnesium sulfate and
evaporated under reduced pressure. The residue was
purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:10–1:5) to afford 28c-anti as a pale
yellow oil (107 mg, 36%) and 28c-syn as a pale yellow
oil (47 mg, 16%).
28a-syn: TLC Rf 0.18 (EtOAc/n-hexane, 1/2); optical
25
D
rotation ½aŠ +30.9 (c 0.88, CHCl3); IR (neat) 3447,
2964, 2932, 2837, 1734, 1651, 1513, 1458, 1417, 1392,
1342, 1303, 1250, 1177, 1109, 1035, 929, 891, 845, 821,
712, 630, 584, 522 cmꢀ1; 1H NMR (200 MHz, CDCl3) d
7.35–7.26 (m, 2H), 6.95–6.85 (m, 2H), 4.87 (d, J=14.6
Hz, 1H), 4.57 (d, J=14.6 Hz, 1H), 3.81 (s, 3H), 3.20 (d,
J=10.4 Hz, 1H), 2.62–2.16 (m, 3H), 1.97 (dd, J=10.0,
6.0 Hz, 1H), 1.26 (d, J=6.4 Hz, 3H); MS (APCI, Pos.)
m/z 316 (M+H, 35Cl, 37Cl)+, 314 (M+H, 35Cl, 35Cl)+;
HR-MS
(MALDI-TOF,
Pos.)
calcd
for
1
C15H17Cl2N1O2+ H+: 314.0715; found: 314.0728.
28c-anti: TLC Rf 0.45 (EtOAc/n-hexane, 1:1); H NMR
(200 MHz, CDCl3) d 7.24 (d, J=9.5 Hz, 1H) and 7.23
(d, J=9.5 Hz, 1H), 6.95 (d, J=9.5 Hz, 1H) and 6.94 (d,
J=9.5 Hz, 1H), 5.47 (d, J=14.2 Hz, 0.5H) and 5.19 (d,
J=14.2 Hz, 0.5H), 4.00 (d, J=14.2 Hz, 0.5H) and 3.79
(d, J=14.2 Hz, 0.5H), 3.81 (s, 1.5H) and 3.80 (s, 1.5H),
3.02–2.83 (m, 1H), 2.40–2.00 (m, 3H), 1.82–1.60 (m,
1H), 1.25 (d, J=6.0 Hz, 1.5H) and 1.24 (d, J=6.0 Hz,
1.5H); MS (APCI, Pos.) m/z 342 (M+H)+, 344.
(1S,5S,6R)-7,7-Dibromo-2-(4-methoxybenzyl)-5-methyl-
2-azabicyclo[4.1.0]heptan-3-one (28b-anti) and (1R,5S,6S)-
7,7-dibromo-2-(4-methoxybenzyl)-5-methyl-2-azabicyclo
[4.1.0]heptan-3-one (28b-syn). To a stirred solution of
26 (2 g, 8.6 mmol) in CHBr3 (25 mL, 286 mmol) were
added aliquat-336 (0.1 mL, 0.22 mmol) and 50% aqu-
eous sodium hydroxide (6.4 g) at 0 ꢁC under an argon
atmosphere. The reaction mixture was stirred for 10 h at
0 ꢁC. After completingthe reaction, the mixture was
treated with saturated aqueous ammonium chloride
and extracted with Et2O. The organic layer was washed
with brine, dried over magnesium sulfate and evapo-
rated under reduced pressure. The residue was purified
1
28c-syn: TLC Rf 0.40 (EtOAc/n-hexane, 1/1); H NMR
(200 MHz, CDCl3) d 7.28–7.21 (m, 2H), 6.95–6.82 (m,
2H), 4.99 (d, J=14.3 Hz, 0.5H) and 4.72 (d, J=7.4 Hz,
0.5H), 4.41 (d, J=14.3 Hz, 0.5H) and 3.80 (d, J=7.4
Hz, 0.5H), 3.81 (s, 3H), 3.22–3.02 (m, 1H), 2.85–2.30

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1733
(m, 3H), 1.80–1.60 (m, 1H), 1.24 (d, J=7.2 Hz, 1.5H)
and 1.23 (d, J=6.6 Hz, 1.5H); MS (APCI, Pos.) m/z 342
(M+H)+, 344.
residue was purified by column chromatography on
silica gel (EtOAc/n-hexane, 1:5–2:1) to afford 29e as a
white powder (232 mg, 28%) and 29f as a white powder
(440 mg, 53%).
(1S,5S,6R,7S)-7-Bromo-2-(4-methoxybenzyl)-5,7-dimethyl-
2-azabicyclo[4.1.0]heptan-3-one (28d-anti). To a stirred
solution of 28b-anti (1.4 g, 3.5 mmol), MeI (1.1 mL, 18
mmol) in THF (30 mL) was added n-BuLi (1.6 M in
n-hexane, 2.8 mL, 4.5 mmol) at ꢀ78 ꢁC under an argon
atmosphere and stirred for 30 min. The reaction mixture
was treated with 1 M HCl, warmed up to room tem-
perature and extracted with EtOAc. The organic layer
was washed with brine, dried over magnesium sulfate
and evaporated under reduced pressure. The residue
was purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:5–1:3) to afford 28d-anti as a pale
yellow oil (1.1 g, 91%). TLC Rf 0.45 (EtOAc/n-hexane,
29e: TLC Rf 0.41 (EtOAc/n-hexane, 5/1); mp 117–
25
D
120 ꢁC; optical rotation ½aŠ +7.1 (c 0.35, CHCl3); IR
(KBr) 3197, 2961, 1677, 1456, 1411, 1381, 1353, 1233,
1077, 997, 834, 724, 543, 479 cmꢀ1; 1H NMR (200 MHz,
CDCl3) d 6.40–6.20 (brs, 1H), 2.94 (dt, J=9.2, 2.0 Hz,
1H), 2.85 (dd, J=3.6, 2.0 Hz, 1H), 2.25–1.90 (m, 3H),
1.50–1.38 (m, 1H), 1.26 (d, J=6.6 Hz, 3H); MS (APCI,
Pos.) m/z 160 (M+H, 35Cl)+; HR-MS (MALDI-TOF,
Pos.) calcd for C7H10Cl1N1O1+ H+: 160.0529; found:
160.0538.
29f: TLC Rf 0.27 (EtOAc/n-hexane, 5/1); mp 136–
25
D
137 ꢁC; optical rotation ½aŠ +95.0 (c 1.06, CHCl3); IR
1
2:1); H NMR (200 MHz, CDCl3) d 7.27 (d, J=8.8 Hz,
2H), 6.87 (d, J=8.8 Hz, 2H), 4.94 (d, J=14.2 Hz, 1H),
4.08 (d, J=14.2 Hz, 1H), 3.80 (s, 3H), 2.93 (d, J=9.4
Hz, 1H), 2.31–1.97 (m, 2H), 1.81 (m, 1H), 1.56 (dd,
J=9.4, 6.4 Hz, 1H), 1.42 (s, 3H), 1.18 (d, J=6.6 Hz,
3H); MS (APCI, Pos.) m/z 338 (M+H)+.
(KBr) 3466, 3231, 2968, 2877, 1638, 1474, 1422, 1388,
1358, 1278, 1228, 1199, 1068, 1012, 935, 878, 792, 732,
684, 555 489 cmꢀ1; 1H NMR (200 MHz, CDCl3) d 6.10–
5.80 (brs, 1H), 3.26 (dd, J=7.8, 5.2 Hz, 1H), 2.86 (dd,
J=5.2, 1.2 Hz, 1H), 2.28–2.03 (m, 3H), 1.36–1.25 (m,
1H), 1.23 (d, J=6.4 Hz, 3H); MS (APCI, Pos.) m/z 160
(M+H, 35Cl)+; HR-MS (MALDI-TOF, Pos.) calcd for
C7H10Cl1N1O1+ H+: 160.0529; found: 160.0556.
(1S,5S,6R)-7,7-Dichloro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-one (29a). To a stirred solution of 28a-anti
(993 mg, 3.2 mmol) in anisole (1.7 mL, 15.6 mmol) was
.
added BF3 OEt2 (4.0 mL) and the reaction mixture was
(1S,5S,6R)-7-Bromo-7-fluoro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-one (29c). Compound 29c was prepared from
28c-anti in 88% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
Pale yellow powder; TLC Rf 0.38 (EtOAc/n-hexane,
3:1); 1H NMR (200 MHz, CDCl3) d 6.70–6.40 (brs, 1H),
3.20–3.08 (m, 1H), 2.32–1.90 (m, 3H), 1.85–1.62 (m,
1H), 1.30–1.25 (m, 3H); MS (APCI, Pos.) m/z 224
(M+H)+, 222.
stirred for 35 h at 100 ꢁC under an argon atmosphere.
Then the mixture was quenched with water and then
treated with 5 M NaOH under coolingin an ice bath.
The resultingmixture was extracted with CHCl , and
3
washed with saturated aqueous sodium bicarbonate and
brine. The organic layer was dried over magnesium sul-
fate and evaporated under reduced pressure. The resi-
due was purified by column chromatography on silica
gel (EtOAc/n-hexane, 1:2–2:1) to afford 29a as a white
powder (466 mg, 75%). TLC Rf 0.36 (EtOAc/n-hexane,
(1S,5S,6R,7S) - 7 - Fluoro - 5 - methyl - 2 - azabicyclo[4.1.0]-
heptan-3-one (29g) and (1S,5S,6R,7R)-7-fluoro-5-methyl-
2-azabicyclo[4.1.0]heptan-3-one (29 h). Compounds 29g
and 29h were prepared from 29c in 41 and 44%
yield, respectively accordingto the same procedure as
described for the preparation of 29e and 29f from
29a.
25
D
2/1); mp 98–99 ꢁC; optical rotation ½aŠ +69.6 (c 1.09,
CHCl3); IR (KBr) 3203, 2971, 1661, 1628, 1480, 1352,
1198, 1141, 1077, 1033, 892, 833, 721, 565, 552, 494,
473, 445 cmꢀ1; 1H NMR (200 MHz, CDCl3) d 6.59 (brs,
1H), 3.19 (dd, J=9.8, 1.2 Hz, 1H), 2.25 (dd, J=12.8,
3.2 Hz, 1H), 2.21–2.02 (m, 1H), 2.04 (d, J=12.8 Hz,
1H), 1.79 (ddd, J=9.8, 5.0, 0.6 Hz, 1H), 1.30 (d, J=6.4
Hz, 3H); MS (APCI, Pos.) m/z 198 (M+H, 37Cl,
37Cl)+, 196 (M+H, 37Cl, 35Cl)+, 194 (M+H, 35Cl,
35Cl)+; HR-MS (MALDI-TOF, Pos.) calcd for
C7H9Cl2N1O1+ H+: 194.0139; found: 194.0150.
29g: Pale yellow powder; TLC Rf 0.21 (EtOAc/n-hex-
ane, 4:1); 1H NMR (200 MHz, CDCl3) d 6.45–6.20 (brs,
1H), 4.28 (dd, J=61.6, 2.2 Hz, 1H), 3.10–2.97 (m, 1H),
2.62–2.20 (m, 3H), 2.60–2.40 (m, 1H), 1.25 (d, J=6.6
Hz, 3H); MS (APCI, Pos.) m/z 144 (M+H)+.
(1S,5S,6R,7S)-7-Chloro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-one (29e) and (1S,5S,6R,7R)-7-chloro-5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (29f). To a stir-
red mixture of 29a (1.0 g, 5.2 mmol) in benzene (6.6 mL)
were added Ph3SnH (2.0 g, 5.7 mmol) and azobisisobu-
tylonitrile (42 mg, 0.26 mmol). The reaction mixture
29h: Pale yellow powder; TLC Rf 0.36 (EtOAc/n-hex-
ane, 4:1); 1H NMR (200 MHz, CDCl3) d 5.80–5.60 (brs,
1H), 4.48 (ddd, J=64.8, 6.6, 4.2 Hz, 1H), 2.70–2.61 (m,
1H), 2.29–2.02 (m, 3H), 1.23 (d, J=6.2 Hz, 3H), 1.20–
1.00 (m, 1H); MS (APCI, Pos.) m/z 144 (M+H)+.
ꢁ
was stirred with heatingat 80 C for 4 h under an argon
atmosphere. Under coolingin an ice bath, the reaction
mixture was diluted with EtOAc and then washed with
10% aqueous potassium fluoride. The resultinginso-
luble substance was removed by filtration. The organic
layer was washed with brine and dried over magnesium
sulfate and evaporated under reduced pressure. The
(1S,5S,6S)-2-(4-Methoxybenzyl)-5-methyl-7-methylene-
2-azabicyclo[4.1.0]heptan-3-one (30). To a stirred solu-
tion of 28d-anti (1 g, 3 mmol) in THF (10 mL) was
added t-BuOK (1 g, 9 mmol) at room temperature. The
reaction mixture was stirred for 2 h. After completing
the reaction, the reaction mixture was diluted with

1734
Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
water and extracted with EtOAc. The organic layer was
washed with brine, dried over magnesium sulfate and
evaporated under reduced pressure. The residue was
purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:3–1:1) to afford 30 as a pale yellow
oil (270 mg, 35%). TLC Rf 0.29 (EtOAc/n-hexane, 2:1);
1H NMR (200 MHz, CDCl3) ꢀ 7.23 (d, J=8.8 Hz, 2H),
6.87 (d, J=8.8 Hz, 2H), 5.39 (t, J=1.8 Hz, 1H), 5.24 (t,
J=1.8 Hz, 1H), 4.69 (d, J=14.6 Hz, 1H), 4.59 (d,
J=14.6 Hz, 1H), 3.80 (s, 3H), 3.02 (d, J=6.2 Hz, 1H),
2.30–2.00 (m, 3H), 1.80–1.65 (m, 1H), 1.15 (d, J=6.6
Hz, 3H); MS (APCI, Pos.) m/z 258 (M+H)+.
on silica gel (EtOAc/n-hexane, 10:1–1:1) to afford 34 as
a pale yellow solid (37 g, 95%). TLC Rf 0.82 (EtOAc/
n-hexane, 1:1); mp 50–51 ꢁC; IR (KBr) 3368, 2968, 2871,
1719, 1671, 1611, 1514, 1469, 1428, 1387, 1339, 1291,
1347, 1218, 1175, 1142, 1109, 1062, 1029, 970, 937, 886,
1
832, 806, 776, 646, 628, 595, 562, 518 cmꢀ1; H NMR
(200 MHz, CDCl3) ꢀ 7.32 (d, J=9.0 Hz, 2H), 6.80 (d,
J=9.0 Hz, 2H), 4.88 (s, 2H), 3.77 (s, 3H), 2.80–2.65 (m,
2H), 2.40–2.12 (m, 3H), 1.04 (d, J=6.2 Hz, 3H); MS
(APCI, Pos.) m/z 248 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C14H17N1O3+ H+: 248.1287;
found: 248.1259.
(1R,5S,6S,7R)-2-(4-Methoxybenzyl)-5,7-dimethyl-2-aza-
bicyclo[4.1.0]heptan-3-one (31). Catalytic hydrogenation
of 30 (772 mg, 3 mmol) was carried out in EtOAc (4
mL) in the presence of 10% palladium carbon (80 mg).
The catalyst was removed by filtration. The filtrate was
concentrated under reduced pressure. The residue was
purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:4) to afford 31 as a pale yellow oil
(552 mg, 71%). TLC Rf 0.61 (EtOAc/n-hexane, 1/1); 1H
NMR (200 MHz, CDCl3) d 7.18 (d, J=8.6 Hz, 2H),
6.78 (d, J=8.6 Hz, 2H), 5.21 (d, J=14.2 Hz, 1H), 3.73
(s, 3H), 3.60 (d, J=14.2 Hz, 1H), 2.42 (dd, J=8.4, 6.4
Hz, 1H), 2.21 (dd, J=15.2, 4.6 Hz, 1H), 2.07 (dd,
J=15.2, 12.0 Hz, 1H), 1.74 (m, 1H), 1.05 (d, J=6.6 Hz,
3H), 1.00–0.79 (m, 2H), 0.82 (d, J=5.6 Hz, 3H); MS
(APCI, Pos.) m/z 260 (M+H)+.
6-Hydroxy-1-(4-methoxybenzyl)-4-methylpiperidin-2-one
(35). To a stirred solution of 34 (14 g, 56.6 mmol) in
EtOH (300 mL) was added NaBH4 (4.2 g, 111 mmol) at
0 ꢁC. The reaction mixture was stirred for 2 h at room
temperature. After completingthe reaction, the result-
ingmixture was cooled to 0 ꢁC and treated with 1 M
HCl to adjust the pH value to 7. After removal of the
solvent by evaporation, the resultingmixture was
extracted with EtOAc and washed with brine. The
combined organic layers were dried over anhydrous
sodium sulfate and evaporated under reduced pressure.
The residue was purified by column chromatography on
silica gel (EtOAc/n-hexane, 1:1) to afford 35 as a white
crystal (13.4 g, 95%). TLC Rf 0.24 (EtOAc/n-hexane,
1:1); mp 130–131 ꢁC; IR (KBr) 3259, 2952, 1626, 1514,
1479, 1413, 1299, 1249, 1170, 1101, 1065, 1025, 974,
1
902, 838, 819, 767, 742, 619, 583, 528 cmꢀ1; H NMR
(1R,5S,6S,7R)-5,7-Dimethyl-2-azabicyclo[4.1.0]heptan-
3-one (32). Compound 32 was prepared from 31 in
57% yield accordingto the same procedure as described
for the preparation of 29a from 28a-anti. Pale yellow
powder; TLC Rf 0.38 (EtOAc); 1H NMR (200 MHz,
CDCl3) d 5.86 (brs, 1H), 2.66 (m, 1H), 2.17 (dd,
J=15.2, 5.0 Hz, 1H), 2.05 (dd, J=15.2, 13.0 Hz, 1H),
1.90–1.65 (m, 1H), 1.17 (d, J=6.6 Hz, 3H), 1.07–0.85
(m, 5H); MS (APCI, Pos.) m/z 140 (M+H)+.
(200 MHz, CDCl3) d 7.31 (d, J=9.0 Hz, 2H), 6.80 (d,
J=9.0 Hz, 2H), 5.10–4.70 (m, 2H), 4.40–4.25 (m, 1H),
3.75 (s, 3H), 2.80–2.05 (m, 3H), 1.95–1.60 (m, 2H), 1.47
(ddd, J=13.6, 12.8, 3.6 Hz, 1H), 1.04 (d, J=6.6 Hz,
3H); MS (APCI, Pos.) m/z 250 (M+H)+; HR-MS
(MALDI-TOF, Pos.) calcd for C14H19N1O3+ H+:
250.1443; found: 250.1437.
1-(4-Methoxybenzyl)-4-methyl-3,4-dihydropyridin-2-one
(37). Compound 37 was prepared from 35 in 60% yield
accordingto the same procedure as described for the
preparation of 26 from 24. Pale yellow oil; TLC Rf 0.55
(EtOAc/n-hexane, 1:1); IR (neat) 3421, 2957, 2836,
1668, 1612, 1585, 1513, 1457, 1409, 1385, 1304, 1248,
1212, 1176, 1147, 1105, 1034, 946, 822, 717, 580, 518
1-(4-Methoxybenzyl)-4-methylpiperidine-2,6-dione (34).
To a stirred solution of 3-methylglutaric anhydride (33)
(20 g, 156 mmol) in THF (300 mL) was added p-methoxy-
benzylamine (23 g, 168 mmol) at room temperature and
stirred for 30 min. After completingthe reaction, the
mixture was evaporated and the residue was diluted
with EtOAc. The resultingmixture was treated with
1 M HCl and extracted with EtOAc. The organic layer
was washed with brine, dried over anhydrous magne-
sium sulfate and evaporated to afford a residue, which
was used for the subsequent reaction without further
purification.
1
cmꢀ1; H NMR (200 MHz, CDCl3) d 7.22 (d, J=8.6
Hz, 2H), 6.85 (d, J=8.6 Hz, 2H), 5.96 (dd, J=7.8, 1.5
Hz, 1H), 5.03 (dd, J=7.8, 3.3 Hz, 1H), 4.70–4.50 (m,
2H), 3.79 (s, 3H), 2.80–2.05 (m, 3H), 1.05 (d, J=6.8 Hz,
3H); MS (APCI, Pos.) m/z 232 (M+H)+; HR-MS
(MALDI-TOF, Pos.) calcd for C14H17N1O2+ H+:
232.1337; found: 232.1313.
To a stirred solution of the compound obtained above
in Ac2O (100 mL) was added Et3N (20 mL) at room
temperature. The reaction mixture was heated at 80 ꢁC
for 1 h. After completingthe reaction, the mixture was
cooled to room temperature and evaporated. The resi-
due was diluted with EtOAc and water. The organic
layer was washed with 1 M HCl, saturated aqueous
sodium bicarbonate, brine and dried over anhydrous
magnesium sulfate and evaporated under reduced pres-
sure. The residue was purified by column chromatography
6-Hydroxy-1-(4-methoxybenzyl)-4,6-dimethylpiperidin-2-
one (36). To a stirred solution of 34 (40 g, 162 mmol) in
THF (600 mL) was added MeLi (1 M in Et2O, 174 mL,
174 mmol) at ꢀ78 ꢁC under an argon atmosphere. After
completingthe reaction, the mixture was diluted with
Et2O and saturated aqueous ammonium chloride was
added. The organic layer was washed with brine, dried
over magnesium sulfate and evaporated under reduced
pressure. The residue was purified by column chroma-
tography on silica gel (EtOAc/n-hexane, 1/1) to afford

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1735
36 as a white solid (42 g, 99%). TLC Rf 0.13 (EtOAc/n-
hexane, 1:1); mp 66–68 ꢁC; IR (KBr) 3309, 2963, 1704,
1633, 1548, 1515, 1459, 1370, 1304, 1252, 1173, 1160,
cooled to ꢀ20 ꢁC. A solution of 39a-anti (2.0 g, 5.0
mmol) in Et2O (40 mL) and HMPA (1.4 mL, 8 mmol)
was added to the mixture and stirred for 1.5 h. Then the
reaction mixture was cooled to ꢀ50 ꢁC and treated with
MeI (10 mL, 161 mmol). After stirringfor 30 min, the
reaction was quenched with saturated aqueous ammo-
nium chloride and the resultingprecipitates were
removed by filtration. The filtrate was extracted with
EtOAc. The organic layer was washed with brine, dried
over magnesium sulfate and evaporated under reduced
pressure. The residue was purified by column chroma-
tography on silica gel (EtOAc/n-hexane, 1:5–1:3) to
afford 39b-anti as a pale yellow oil (1.0 g, 73%). TLC Rf
0.55 (EtOAc/n-hexane, 1:1); 1H NMR (200 MHz,
CDCl3) d 7.22 (d, J=8.8 Hz, 2H), 6.85 (d, J=8.8 Hz,
2H), 5.14 (d, J=14.0 Hz, 1H), 3.79 (d, J=14.0 Hz, 1H),
3.79 (s, 3H), 2.29–2.01 (m, 3H), 1.78 (m, 1H), 1.10 (d,
J=6.6 Hz, 3H), 1.00 (s, 3H), 0.86 (s, 3H), 0.69 (dd,
J=8.6, 6.2 Hz, 1H); MS (APCI, Pos.) m/z 274 (M+H)+.
1
1036, 830, 696, 589 cmꢀ1; H NMR (200 MHz, CDCl3)
d 7.20 (d, J=8.8 Hz, 2H), 6.86 (d, J=8.8 Hz, 2H), 5.85
(br, 1H), 4.36 (d, J=5.6 Hz, 2H), 3.80 (s, 3H), 2.63–2.00
(m, 5H), 2.13 (s, 3H), 1.00 (d, J=6.4 Hz, 3H); MS
(APCI, Pos.) m/z 264 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C15H21N1O3+ H+: 264.1600;
found: 264.1585.
1-(4-Methoxybenzyl)-4,6-dimethyl-3,4-dihydropyridin-2-
one (38). Compound 38 was prepared from 36 in 99%
yield accordingto the same procedure as described for
the preparation of 26 from 24. Colorless oil; TLC Rf
0.58 (EtOAc/n-hexane, 1/1); IR (neat) 2957, 2836, 1674,
1614, 1513, 1457, 1388, 1248, 1176, 1110, 1034, 959,
891, 819, 772, 659, 548 cmꢀ1 1H NMR (200 MHz,
;
CDCl3) d 7.11 (d, J=8.8 Hz, 2H), 6.84 (d, J=8.8 Hz,
2H), 4.92 (d, J=15.8 Hz, 1H), 4.89 (s, 1H), 4.70 (d,
J=15.8 Hz, 1H), 3.78 (s, 3H), 2.80–2.20 (m, 3H), 1.85
(s, 3H), 1.04 (d, J=7.0 Hz, 3H); MS (APCI, Pos.) m/z
246 (M+H)+; HR-MS (MALDI-TOF, Pos.) calcd for
C15H19N1O2+ H+: 246.1494; found: 246.1477.
(dl)-(1R,5S,6S,7S)-2-(4-Methoxybenzyl)-5,7-dimethyl-2-
azabicyclo[4.1.0]heptan-3-one (39c-anti). To a stirred
suspension of CuSCN (6.0 g, 50 mmol) in Et2O (35 mL)
was added MeLi (1 M in Et2O, 100 mL, 100 mmol) at
ꢀ78 ꢁC under an argon atmosphere. The reaction mix-
ture was warmed up to ꢀ15 ꢁC in 1.5 h and then cooled
to ꢀ20 ꢁC. A solution of 39a-anti (2.5 g, 6.2 mmol) in
Et2O (45 mL) and HMPA (1.7 mL, 10 mmol) was added
to the mixture. After stirringfor 1.5 h, the reaction
mixture was cooled to ꢀ50 ꢁC, then quenched with
saturated aqueous ammonium chloride and warmed up
to room temperature. After stirringfor 30 min, the
resultingprecipitates were removed by filtration. The
filtrate was extracted with EtOAc. The organic layer
was washed with brine, dried over magnesium sulfate
and evaporated under reduced pressure. The residue
was purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:5–1:3) to afford 39c-anti as a pale
yellow oil (627 mg, 39%). TLC Rf 0.38 (EtOAc/n-hex-
ane, 1:1); 1H NMR (200 MHz, CDCl3) d 7.23 (d, J=8.8
Hz, 2H), 6.85 (d, J=8.8 Hz, 2H), 4.76 (d, J=14.2 Hz,
1H), 4.36 (d, J=14.2 Hz, 1H), 3.80 (s, 3H), 2.28–2.04
(m, 3H), 1.73 (m, 1H), 1.13 (d, J=6.6 Hz, 3H), 0.91 (d,
J=6.2 Hz, 3H), 0.68 (m, 1H), 0.57 (m, 1H); MS (APCI,
Pos.) m/z 260 (M+H)+.
(dl)-(1S,5S,6R) - 7,7 - Dibromo -2 - (4 -methoxybenzyl) - 5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (39a-anti) and
(dl) - (1R,5S,6S) - 7,7 -dibromo -2 -(4 -methoxybenzyl) - 5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (39a-syn). Com-
pounds 39a-anti and 39a-syn were prepared from 37 in
38 and 13% yield, respectively accordingto the same
procedure as described for the preparation of 28b-anti
and 28b-syn from 26.
39a-anti: pale yellow oil; TLC Rf 0.38 (EtOAc/n-hexane,
1:2); IR (neat) 2961, 2836, 1667, 1612, 1513, 1415, 1379,
1248, 1175, 1075, 1033, 845, 764, 721, 581, 503 cmꢀ1; 1H
NMR (200 MHz, CDCl3) d 7.29 (d, J=8.4 Hz, 2H),
6.89 (d, J=8.4 Hz, 2H), 5.47 (d, J=14.8 Hz, 1H), 3.81
(d, J=14.8 Hz, 1H), 3.81 (s, 3H), 2.98 (d, J=9.8 Hz,
1H), 2.40–2.00 (m, 3H), 1.77 (dd, J=9.8, 5.4 Hz, 1H),
1.26 (m, 3H); MS (APCI, Pos.) m/z 404 (M+H, 79Br,
81Br)+; HR-MS (MALDI-TOF, Pos.) calcd for
C15H17Br2N1O2+ H+: 401.9704; found: 401.9717.
39a-syn: pale yellow oil; TLC Rf 0.18 (EtOAc/n-hexane,
1:2); IR (neat) 2962, 2836, 1652, 1513, 1456, 1415, 1389,
1341, 1303, 1249, 1176, 1108, 1033, 885, 849, 773, 582,
545 cmꢀ1; 1H NMR (200 MHz, CDCl3) ꢀ 7.33 (d, J=8.6
Hz, 2H), 6.91 (d, J=8.6 Hz, 2H), 4.93 (d, J=14.2 Hz,
1H), 4.54 (d, J=14.2 Hz, 1H), 3.82 (s, 3H), 3.27 (d,
J=10.2 Hz, 1H), 2.50–2.45 (m, 3H), 2.20–2.10 (m, 1H),
1.28 (d, J=6.2 Hz, 3H); MS (APCI, Pos.) m/z 404
(M+H, 79Br, 81Br)+; HR-MS (MALDI-TOF, Pos.)
calcd for C15H17Br2N1O2+ H+: 401.9704; found:
401.9703.
(dl)-(1S,5S,6R,7S)-7-Bromo-2-(4-methoxybenzyl)-5,7-di-
methyl-2-azabicyclo[4.1.0]heptan-3-one (39d-anti). Com-
pound 39d-anti was prepared from 39a-anti in 73%
yield accordingto the same procedure as described for
the preparation of 28d-anti from 28b-anti. Pale yellow
oil; TLC Rf 0.45 (EtOAc/n-hexane, 2:1); 1H NMR
(200 MHz, CDCl3) d 7.28 (d, J=8.8 Hz, 2H), 6.89 (d,
J=8.8 Hz, 2H), 4.94 (d, J=14.2 Hz, 1H), 4.08 (d,
J=14.2 Hz, 1H), 3.80 (s, 3H), 2.93 (d, J=9.4 Hz, 1H),
2.35–1.95 (m, 2H), 1.81 (m, 1H), 1.56 (dd, J=9.4, 6.4
Hz, 1H), 1.42 (s, 3H), 1.18 (d, J=6.5 Hz, 3H); MS
(APCI, Pos.) m/z 338 (M+H)+.
(dl)-(1S,5S,6S)-2-(4-Methoxybenzyl)-5,7,7-trimethyl-2-
azabicyclo[4.1.0]heptan-3-one (39b-anti). To a stirred
suspension of CuSCN (4.8 g, 40 mmol) in Et2O (30 mL)
was added MeLi (1 M in Et2O, 80 mL, 80 mmol) at
ꢀ78 ꢁC under an argon atmosphere. The reaction mix-
ture was warmed up to ꢀ15 ꢁC over 1.5 h and then
(dl)-(1R,5S,6S,7R)-2-(4-Methoxybenzyl)-5,7-dimethyl-2-
azabicyclo[4.1.0]heptan-3-one (39e-anti). To a stirred
suspension of CuI (2.4 g, 12.8 mmol) in THF (50 mL)
was added n-BuLi (1.6 M in n-hexane, 16 mL, 25.6

1736
Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
mmol) at ꢀ78 ꢁC under an argon atmosphere. The
reaction mixture was warmed up to ꢀ45 ꢁC over 1 h and
then cooled to ꢀ78 ꢁC. A solution of 39d-anti (866 mg,
2.6 mmol) in THF (5 mL) was added to the mixture,
and stirred for 1 h at ꢀ78 ꢁC. After stirringat ꢀ78 ꢁC
for 1 h, the mixture was quenched with 4 M HCl/
EtOAc, and warmed up to room temperature. After
stirred for 30 min, the reaction mixture was treated with
water and extracted with EtOAc. The organic layer was
washed with brine, dried over magnesium sulfate and
evaporated under reduced pressure. The residue was
purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:6–2:1) to afford 39e-anti as a pale
yellow oil (243 mg, 36%). TLC Rf 0.61 (EtOAc/n-hex-
ane, 1:1); 1H NMR (200 MHz, CDCl3) d 7.20 (d, J=8.6
Hz, 2H), 6.80 (d, J=8.6 Hz, 2H), 5.21 (d, J=14.2 Hz,
1H), 3.73 (s, 3H), 3.60 (d, J=14.2 Hz, 1H), 2.42 (dd,
J=8.4, 6.4 Hz, 1H), 2.21 (dd, J=15.2, 4.6 Hz, 1H), 2.07
(dd, J=15.2, 12.0 Hz, 1H), 1.74 (m, 1H), 1.05 (d, J=6.6
Hz, 3H), 1.00–0.75 (m, 2H), 0.82 (d, J=5.6 Hz, 3H);
MS (APCI, Pos.) m/z 260 (M+H)+.
Pale yellow powder; TLC Rf 0.15 (EtOAc/n-hexane,
1
2:1); H NMR (200 MHz, CDCl3) d 5.80 (brs, 1H), 2.36
(d, J=8.4 Hz, 1H), 2.14 (dd, J=14.0, 4.0 Hz, 1H), 1.98
(dd, J=14.0, 2.0 Hz, 1H), 1.73 (m, 1H), 1.15 (d, J=6.6
Hz, 3H), 1.04 (s, 3H), 0.99 (s, 3H), 0.71 (dd, J=8.4, 6.0
Hz, 1H); MS (APCI, Pos.) m/z 154 (M+H)+.
(dl) - (1R,5S,6S,7S) - 5,7 - Dimethyl - 2 - azabicyclo[4.1.0]-
heptan-3-one (40c). Compound 40c was prepared from
39c-anti in 78% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
Pale yellow powder; TLC Rf 0.09 (EtOAc/n-hexane,
1
2:1); H NMR (200 MHz, CDCl3) d 6.12 (brs, 1H), 2.32
(ddd, J=8.4, 3.0, 1.8 Hz, 1H), 2.14 (dd, J=15.0, 5.8 Hz,
1H), 2.01 (d, J=15.0 Hz, 1H), 1.86 (m, 1H), 1.18 (d,
J=6.6 Hz, 3H), 1.02 (d, J=5.8 Hz, 3H), 0.81 (m, 1H),
0.71 (m, 1H); MS (APCI, Pos.) m/z 140 (M+H)+.
(dl) - (1R,5S,6S,7R) - 5,7 - Dimethyl - 2 - azabicyclo[4.1.0]-
heptan-3-one (40e). Compound 40e was prepared from
39e-anti in 68% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
Pale yellow powder; TLC Rf 0.14 (EtOAc/n-hexane,
1:1); 1H NMR (200 MHz, CDCl3) d 5.80–5.60 (brs, 1H),
2.70–2.60 (m, 1H), 2.20–1.90 (m, 2H), 1.85–1.55 (m,
1H), 1.16 (d, J=6.6 Hz, 3H), 1.05–0.97 (m, 5H); MS
(APCI, Pos.) m/z 140 (M+H)+.
(dl) - (1R,5S,6S,7S) - 7 - Butyl - 2 - (4 - methoxybenzyl) - 5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (39f-anti) and
(dl) - (1R,5S,6S,7R) - 7 - butyl - 2 - (4 - methoxybenzyl) - 5 -
methyl-2-azabicyclo[4.1.0]heptan-3-one (39g-anti). To a
stirred suspension of CuSCN (5.1 g, 42 mmol) in Et2O
(70 mL) was added n-BuLi (1.6 M in n-hexane, 51 mL,
82 mmol) at ꢀ78 ꢁC under an argon atmosphere and
warmed up to ꢀ10 ꢁC. After stirringfor 1.5 h, the reac-
tion mixture was cooled to ꢀ78 ꢁC. A solution of 39a-
anti (2.1 g, 5.2 mmol) in Et2O (30 mL) and HMPA (2
mL, 11.5 mmol) was added to the mixture. After stirring
for 1 h, the reaction mixture was poured into saturated
aqueous ammonium chloride under stirring. The reac-
tion mixture was extracted with EtOAc. The organic
layer was washed with brine, dried over magnesium
sulfate and evaporated under reduced pressure. The
residue was purified by column chromatography on
silica gel (EtOAc/n-hexane, 1:5) to afford 39f-anti as a
pale yellow oil (282 mg, 18%) and 39g-anti as a pale
yellow oil (611 mg, 39%).
(dl)-(1R,5S,6S,7R)-7-Butyl-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-one (40g). Compound 40g was prepared from
39g-anti in 47% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
Pale yellow powder; TLC Rf 0.16 (EtOAc/n-hexane,
1:1); 1H NMR (200 MHz, CDCl3) d 5.97 (brs, 1H),
2.80–2.70 (m, 2H), 2.49–2.04 (m, 1H), 1.75 (m, 1H),
1.46–1.20 (m, 6H), 1.18 (d, J=6.6 Hz, 3H), 0.99–0.80
(m, 5H); MS (APCI, Pos.) m/z 182 (M+H)+.
(dl) - (1S,5S,6R,7S) - 7- Bromo - 2 - (4 - methoxybenzyl)- 5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (41a) and (dl)-
(1S,5S,6R,7R)-7-bromo-2-(4-methoxybenzyl)-5-methyl-
2-azabicyclo[4.1.0]heptan-3-one (41b). To a stirred solu-
tion of 39a-anti (8.0 g, 19.8 mmol) in toluene (70 mL)
was added n-Bu3SnH (6.4 mL, 23.8 mmol) at ꢀ10 ꢁC.
After stirringfor 15 h at ꢀ10 ꢁC, the reaction was
quenched with 10% aqueous potassium fluoride and the
mixture was extracted with EtOAc. The organic layer
was washed with brine, dried over magnesium sulfate
and evaporated under reduced pressure. The residue
was purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:5–2:1) to afford 41a and 41b as an
inseparable mixture (5.0 g, 78% yield, 41a:41b=1:3).
1
39f-anti: TLC Rf 0.32 (EtOAc/n-hexane, 1:2); H NMR
(200 MHz, CDCl3) d 7.22 (d, J=8.8 Hz, 2H), 6.85 (d,
J=8.8 Hz, 2H), 4.92 (d, J=14.2 Hz, 1H), 4.20 (d,
J=14.2 Hz, 1H), 3.80 (s, 3H), 2.30–2.05 (m, 3H), 1.75
(m, 1H), 1.27–1.00 (m, 6H), 1.14 (d, J=6.6 Hz, 3H),
0.87 (t, J=7.0 Hz, 3H), 0.73–0.58(m, 2H); MS (APCI,
Pos.) m/z 302 (M+H)+.
1
39g-anti: TLC Rf 0.40 (EtOAc/n-hexane, 1:2); H NMR
(200 MHz, CDCl3) d 7.24 (d, J=8.8 Hz, 2H), 6.85 (d,
J=8.8 Hz, 2H), 5.33 (d, J=14.0 Hz, 1H), 3.79 (s, 3H),
3.63 (d, J=14.0 Hz, 1H), 2.49 (t, J=7.0 Hz, 1H), 2.32–
2.04 (m, 2H), 1.77 (m, 1H), 1.46–1.20 (m, 6H), 1.13 (d,
J=6.4 Hz, 3H), 0.99–0.80 (m, 5H); MS (APCI, Pos.)
m/z 302 (M+H)+.
41a: Pale yellow oil; TLC Rf 0.35 (EtOAc/n-hexane,
1
1:1); H NMR (200 MHz, CDCl3) d 7.27 (d, J=8.8 Hz,
2H), 6.87 (d, J=8.8 Hz, 2H), 4.76 (d, J=14.0 Hz, 1H),
4.42 (d, J=14.0 Hz, 1H), 3.80 (s, 3H), 2.98 (dd, J=10.0,
1.0 Hz, 1H), 2.44 (dd, J=3.0, 1.0 Hz, 1H), 2.30–2.10 (m,
2H), 1.82 (m, 1H), 1.46 (m, 1H), 1.21 (d, J=6.6 Hz,
3H); MS (APCI, Pos.) m/z 324 (M+H)+.
(dl) - (1S,5S,6S) - 5,7,7 - Trimethyl - 2 - azabicyclo[4.1.0]-
heptan-3-one (40b). Compound 40b was prepared from
39b-anti in 49% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
41b: Pale yellow oil; TLC Rf 0.38 (EtOAc/n-hexane,
1
1:1); H NMR (200 MHz, CDCl3) d 7.25 (d, J=8.8 Hz,

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1737
2H), 6.85 (d, J=8.8 Hz, 2H), 5.52 (d, J=14.0 Hz, 1H),
3.79 (s, 3H), 3.64 (d, J=14.0 Hz, 1H), 3.27 (dd, J=7.8,
5.0 Hz, 1H), 2.64 (dd, J=9.4, 5.0 Hz, 1H), 2.48–2.02 (m,
3H), 1.80 (m, 1H), 1.20 (d, J=6.6 Hz, 3H); MS (APCI,
Pos.) m/z 324 (M+H)+.
g, 4.2 mmol) in THF (25 mL) and the reaction mixture
was warmed up to ꢀ15 ꢁC over 1 h. After completing
the reaction, the reaction was quenched with saturated
aqueous ammonium chloride and the mixture was
extracted with EtOAc. The organic layer was washed
with brine, dried over magnesium sulfate and evapo-
rated under reduced pressure. The residue was purified
by column chromatography on silica gel (EtOAc/n-hex-
ane, 1:2–1:1) to afford 45 as a pale yellow powder (375
mg, 59%). TLC Rf 0.23 (EtOAc/n-hexane, 2:1); 1H
NMR (200 MHz, CDCl3) d 5.95 (brs, 1H), 5.46 (ddd,
J=17.0, 10.3, 4.1 Hz, 1H), 5.03 (ddd, J=17.0, 1.5, 0.8
Hz, 1H), 4.94 (dd, J=10.3, 1.5 Hz, 1H), 2.65 (ddd,
J=8.5, 2.7, 1.9 Hz, 1H), 2.27–1.86 (m, 3H), 1.49 (ddd,
J=8.5, 4.8, 2.5 Hz, 1H), 1.22 (d, J=6.4 Hz, 3H), 1.13
(m, 1H); MS (APCI, Pos.) m/z 152 (M+H)+.
(dl) - (1R,5S,6S,7R) - 7 - Ethyl - 2 - (4 - methoxybenzyl) - 5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (42b). To a stir-
red suspension of CuI (7.2 g, 37.8 mmol) in THF (120
mL) was added n-BuLi (1.6 M in n-hexane, 47 mL, 75.6
mmol) at ꢀ78 ꢁC under an argon atmosphere. The
reaction mixture was warmed up to ꢀ45 ꢁC over 1 h. A
solution of 41a and 41b (2.4 g, 7.4 mmol) in THF (45
mL) was added to the mixture and stirred for 1 h at
ꢀ45 ꢁC. Then the reaction mixture was treated with EtI
(15 mL, 188 mmol). After stirringfor 30 min, the reac-
tion was quenched with saturated aqueous ammonium
chloride and the mixture was extracted with EtOAc.
The organic layer was washed with brine, dried over
magnesium sulfate and evaporated under reduced pres-
sure. The residue was purified by column chromato-
graphy on silica gel (EtOAc/n-hexane, 1:6–2:1) to afford
42b as a pale yellow oil (404 mg, 20%). TLC Rf 0.50
(1R,5R,6S)-7,7-Dichloro-2-(4-methoxybenzyl)-5-methyl-
2-azabicyclo[4.1.0]heptan-3-one (46a-anti) and (1S,5R,6R)-
7,7-dichloro-2-(4-methoxybenzyl)-5-methyl-2-azabicyclo
[4.1.0]heptan-3-one (46a-syn). Compounds 46a-anti and
46a-syn were prepared from 27 in 53 and 14% yield,
respectively accordingto the same procedure as descri-
bed for the preparation of 28a-anti and 28a-syn from 26.
1
(EtOAc/n-hexane, 2:3); H NMR (200 MHz, CDCl3) d
7.24 (d, J=8.0 Hz, 2H), 6.84 (d, J=8.0 Hz, 2H), 5.34
(d, J=14.0 Hz, 1H), 3.79 (s, 3H), 3.62 (d, J=14.0 Hz,
1H), 2.49 (dd, J=8.2, 6.6 Hz, 1H), 2.31–2.04 (m, 2H),
1.77 (m, 1H), 1.50–1.16 (m, 2H), 1.13 (d, J=6.6 Hz,
3H), 0.99 (t, J=7.0 Hz, 3H), 0.99–0.85 (m, 2H); MS
(APCI, Pos.) m/z 274 (M+H)+.
46a-anti: pale yellow oil; TLC Rf 0.36 (EtOAc/n-hexane,
25
D
1:2); optical rotation ½aŠ ꢀ27.5 (c 1.03, CHCl3); IR
(neat) 2964, 2932, 2874, 2837, 1669, 1613, 1586, 1515,
1445, 1416, 1384, 1363, 1303, 1280, 1248, 1196, 1176,
1112, 1080, 1033, 970, 890, 855, 823, 766, 731, 703, 626,
1
575, 511 cmꢀ1; H NMR (200 MHz, CDCl3) d 7.28 (d,
(dl)-(1R,5S,6S,7R)-7-Ethyl-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-one (43). Compound 43 was prepared from
42b in 39% yield accordingto the same procedure as
described for the preparation of 29a from 28a-anti.
J=8.6 Hz, 2H), 6.89 (d, J=8.6 Hz, 2H), 5.44 (d,
J=14.0 Hz, 1H), 3.81 (s, 3H), 3.81 (d, J=14.0 Hz, 1H),
2.95 (d, J=10.0 Hz, 1H), 2.37–2.05 (m, 3H), 1.76 (dd,
J=10.2, 5.4 Hz, 1H), 1.25 (d, J=6.2 Hz, 3H); MS
(APCI, Pos.) m/z 316 (M+H, 35Cl, 37Cl)+, 314 (M+H,
35Cl, 35Cl)+; HR-MS (MALDI-TOF, Pos.) calcd for
C15H17Cl2N1O2+ H+: 314.0715; found: 314.0686.
1
White powder; TLC Rf 0.26 (EtOAc/n-hexane, 3:2); H
NMR (200 MHz, CDCl3) d 5.67 (brs, 1H), 2.68 (dd,
J=8.0, 6.0 Hz, 1H), 2.16 (dd, J=15.0, 5.6 Hz, 1H), 2.01
(d, J=15.0 Hz, 1H), 1.75 (m, 1H), 1.32 (m, 2H), 1.17 (d,
J=6.6 Hz, 3H), 1.03–0.84 (m, 2H), 0.96 (t, J=7.0 Hz,
3H); MS (APCI, Pos.) m/z 154 (M+H)+.
46a-syn: pale yellow oil; TLC Rf 0.18 (EtOAc/n-hexane,
25
D
(neat) 2964, 1652, 1513, 1456, 1417, 1392, 1249, 1176,
1:2); optical rotation ½aŠ ꢀ29.7 (c 1.00, CHCl3); IR
(dl)-(1S,5S,6R)-7,7-Dibromo-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-one (44). Compound 44 was prepared from
39a-anti in 91% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
Pale yellow powder; TLC Rf 0.43 (EtOAc/n-hexane,
4:1); mp 106–109 ꢁC; IR (KBr) 3205, 3099, 2960, 1656,
1474, 1452, 1407, 1365, 1352, 1286, 1200, 1121, 1096,
1034, 891, 820, 763, 520 cmꢀ1 1H NMR (200 MHz,
;
CDCl3) ꢀ 7.31 (d, J=8.8 Hz, 2H), 6.89 (d, J=8.8 Hz,
2H), 4.87 (d, J=14.4 Hz, 1H), 4.57 (d, J=14.4 Hz, 1H),
3.81 (s, 3H), 3.20 (d, J=10.2 Hz, 1H), 2.60–2.25 (m,
3H), 2.00–1.90 (m, 1H), 1.27 (d, J=10.0 Hz, 3H); MS
(APCI, Pos.) m/z 314 (M+H, 35Cl, 35Cl)+; HR-MS
(MALDI-TOF, Pos.) calcd for C15H17Cl2N1O2+ H+:
314.0715; found: 314.0708.
1072, 1015, 933, 884, 774, 717, 553 cmꢀ1 1H NMR
;
(200 MHz, CDCl3) d 6.55 (brs, 1H), 3.21 (d, J=9.4 Hz,
1H), 2.30–1.94 (m, 3H), 1.83 (dd, J=9.4, 5.2 Hz, 1H),
1.32 (d, J=6.0 Hz, 3H); MS (APCI, Pos.) m/z 284
(M+H, 79Br, 81Br)+; HR-MS (MALDI-TOF, Pos.)
calcd for C7H9Br2N1O1+ H+: 281.9129; found:
281.9130.
(1R,5R,6S)-7-Bromo-7-chloro-2-(4-methoxybenzyl)-5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (46b-anti) and
(1S,5R,6R)-7-bromo-7-chloro-2-(4-methoxybenzyl)-5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (46b-syn). Com-
pounds 46b-anti and 46b-syn were prepared from 27 in
64 and 18% yield, respectively accordingto the same
procedure as described for the preparation of 28a-anti
(dl)-(1R,5S,6S,7S)-5-Methyl-7-vinyl-2-azabicyclo[4.1.0]-
heptan-3-one (45). To a stirred suspension of CuI (3.1
g, 16 mmol) in Et2O (13 mL) was added vinyl magne-
sium bromide (1 M in THF, 32 mL, 32 mmol) at ꢀ40 ꢁC
under an argon atmosphere and stirring was continued
for 1 h. To this solution was added a solution of 44 (1.2
and 28a-syn from 26 by usingClCHBr
PhCH2NEt3Cl instead of CHCl3 and aliquat-336.
and
2
46b-anti: pale yellow oil; TLC Rf 0.49 (EtOAc/n-hexane,
25
D
1:1); optical rotation ½aŠ ꢀ12.4 (c 1.07, CHCl3); IR

1738
Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
(neat) 2963, 2931, 2836, 1668, 1613, 1585, 1513, 1443,
1415, 1381, 1303, 1280, 1248, 1175, 1112, 1077, 1033,
1400, 1367, 1351, 1280, 1235, 1197, 1136, 1076, 1026,
1
938, 888, 784, 744, 717, 551, 499, 472 cmꢀ1; H NMR
1
964, 885, 848, 817, 793, 765, 727, 576, 508 cmꢀ1; H
(200 MHz, CDCl3) d 6.30–6.15 (brs, 1H), 3.29 (dd,
J=9.6, 1.4 Hz, 0.5H) and 3.11 (dd, J=9.6, 1.4 Hz,
0.5H), 2.30–2.00 (m, 3H), 1.88 (ddd, J=9.7, 5.4, 1.0 Hz,
0.5H) and 1.73 (ddd, J=9.7, 5.4, 1.0 Hz, 0.5H), 1.33–
1.29 (m, 3H); MS (APCI, Pos.) m/z 241, 239 (M+H)+;
NMR (200 MHz, CDCl3) d 7.29 (d, J=8.8 Hz, 2H),
6.90 (d, J=8.8 Hz, 2H), 5.48 (d, J=14.4 Hz, 0.5H) and
5.43 (d, J=14.4 Hz, 0.5H), 3.84 (d, J=9.8 Hz, 0.5H)
and 3.80 (d, J=14.4 Hz, 0.5H), 3.81 (s, 3H), 3.04 (d,
J=9.8 Hz, 0.5H) and 2.88 (d, J=9.8 Hz, 0.5H), 2.20–
2.05 (m, 3H), 1.84 (dd, J=14.6, 5.2 Hz, 0.5H) and 1.69
(dd, J=10.0, 5.2 Hz, 0.5H), 1.30–1.20 (m, 3H); MS
(APCI, Pos.) m/z 360, 358 (M+H)+; HR-MS
(MALDI-TOF, Pos.) calcd for C15H17Br1Cl1N1O2+
H+: 358.0209; found: 358.0205.
HR-MS
(MALDI-TOF,
Pos.)
calcd
for
C7H9Br1Cl1N1O1+ H+: 237.9634; found: 237.9662.
(1R,5R,6S,7S)-7-Chloro-5-methyl-2-azabicyclo[4.1.0]hep-
tan-3-one (47c) and (1R,5R,6S,7R)-7-chloro-5-methyl-2-
azabicyclo[4.1.0]heptan-3-one (47d). To a stirred mix-
ture of 47b (1.0 g, 4.2 mmol) in toluene (5 mL) were
added (TMS)3SiH (2.0 mL, 6.5 mmol) and BEt3 (1 M in
THF, 0.1 mL) and stirringwas continued at room tem-
perature. After completingthe reaction, the reaction
mixture was diluted with EtOAc (10 mL) and washed
with brine (10 mL), dried over magnesium sulfate and
evaporated under reduced pressure. The residue was
purified by column chromatography on silica gel
(EtOAc/n-hexane, 1:5–1:1) to afford 47c as a white
powder (295 mg, 44%) and 47d as a white powder (188
mg, 28%), respectively.
46b-syn: pale yellow oil; TLC Rf 0.40 (EtOAc/n-hexane,
25
D
1:1); optical rotation ½aŠ ꢀ25.1 (c 1.15, CHCl3); IR
(neat) 2963, 1652, 1514, 1456, 1416, 1392, 1342, 1303,
1250, 1177, 1111, 1032, 889, 836, 763, 712, 523 cmꢀ1; 1H
NMR (200 MHz, CDCl3) d 7.32 (d, J=8.8 Hz, 2H),
6.91 (d, J=8.8 Hz, 2H), 4.97 (d, J=14.4 Hz, 0.5H) and
4.84 (d, J=14.4 Hz, 0.5H), 4.63 (d, J=14.4 Hz, 0.5H)
and 4.49 (d, J=14.4 Hz, 0.5H), 3.82 (s, 1.5H) and 3.80
(s, 1.5H), 3.29 (d, J=10.3 Hz, 0.5H) and 3.16 (d,
J=10.3 Hz, 0.5H), 2.65–2.20 (m, 3H), 2.10–1.90 (m,
1H), 1.30–1.25 (m, 3H); MS (APCI, Pos.) m/z 360, 358
(M+H)+; HR-MS (MALDI-TOF, Pos.) calcd for
C15H17Br1Cl1N1O2+ H+: 358.0209; found: 358.0196.
47c: TLC Rf 0.28 (EtOAc); mp 135–136 ꢁC; optical
25
D
rotation ½aŠ ꢀ94.5 (c 1.05, CHCl3); IR (KBr) 3464,
3230, 2974, 2878, 1646, 1475, 1423, 1389, 1358, 1295,
1278, 1229, 1199, 1112, 1069, 1013, 937, 879, 793, 770,
(1S,5R,6R)-7-Bromo-7-fluoro-2-(4-methoxybenzyl)-5-
methyl-2-azabicyclo[4.1.0]heptan-3-one (46c-syn). Com-
pound 46c-syn was prepared from 27 in 15% yield
accordingto the same procedure as described for the pre-
paration of 28a-anti and 28a-syn from 26. Pale yellow oil;
TLC Rf 0.40 (EtOAc/n-hexane, 1:1); 1H NMR (200MHz,
CDCl3) d 7.29 (d, J=8.6 Hz, 2H), 6.88 (d, J=8.6 Hz, 2H),
4.99 (d, J=14.3 Hz, 0.5H) and 4.71 (d, J=14.3 Hz, 0.5H),
4.41 (d, J=7.4 Hz, 0.5H) and 3.78 (d, J=7.4 Hz, 0.5H),
3.81 (s, 3H), 3.23–3.04 (m, 1H), 2.65–2.30 (m, 3H), 2.08–
1.80 (m, 1H), 1.24 (d, J=6.4 Hz, 1.5H) and 1.22 (d, J=6.4
Hz, 1.5H); MS (APCI, Pos.) m/z 342 (M+H)+.
734, 686, 616, 557, 540, 491, 442 cmꢀ1 1H NMR
;
(200 MHz, CDCl3) d 6.00–5.70 (brs, 1H), 3.27 (dd,
J=7.8, 5.2 Hz, 1H), 2.87 (ddd, J=9.4, 5.2, 1.2 Hz, 1H),
2.30–2.00 (m, 3H), 1.36–1.25 (m, 1H), 1.24 (d, J=6.4
Hz, 3H); MS (APCI, Pos.) m/z 160 (M+H)+; HR-MS
(MALDI-TOF, Pos.) calcd for C7H10Cl1N1O1+ H+:
160.0529; found: 160.0540.
47d: TLC Rf 0.42 (EtOAc); mp 118–120 ꢁC; optical
25
D
3049, 2961, 2928, 1677, 1456, 1411, 1382, 1353, 1291,
rotation ½aŠ ꢀ7.5 (c 0.40, CHCl3); IR (KBr) 3195,
1233, 1077, 997, 834, 725, 543, 480 cmꢀ1 1H NMR
;
(1R,5R,6S)-7,7-Dichloro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-one (47a). Compound 47a was prepared from
46a-anti in 79% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
(200 MHz, CDCl3) d 6.50–6.30 (brs, 1H), 2.94 (dt,
J=9.2, 1.9 Hz, 1H), 2.90–2.82 (m, 1H), 2.26–1.90 (m,
3H), 1.55–1.40 (m, 1H), 1.26 (d, J=6.4 Hz, 3H); MS
(APCI, Pos.) m/z 160 (M+H)+.
Pale yellow powder; TLC Rf 0.36 (EtOAc/n-hexane,
25
D
1:2); mp 100–101 ꢁC; optical rotation ½aŠ ꢀ70.5 (c 1.09,
(1S,5R,6R)-7-Bromo-7-chloro-5-methyl-2-azabicyclo[4.1.0]
heptan-3-one (48b). Compound 48b was prepared from
46b-syn in 96% yield accordingto the same procedure as
described for the preparation of 29a from 28a-anti. Pale
CHCl3); IR (KBr) 3436, 2972, 1662, 1630, 1481, 1403,
1373, 1353, 1205, 1078, 1034, 893, 833, 720, 566, 504
cmꢀ1; H NMR (200 MHz, CDCl3) ꢀ 6.37 (brs, 1H),
1
3.19 (dd, J=9.8, 1.4 Hz, 1H), 2.30–1.94 (m. 3H), 1.84–
1.74 (m, 1H), 1.30 (d, J=6.2 Hz, 3H); MS (APCI, Pos.)
m/z 194 (M+H, 35Cl, 35Cl)+; HR-MS (MALDI-TOF,
Pos.) calcd for C7H9Cl2N1O1+ H+: 194.0139; found:
194.0146.
yellow powder; TLC Rf 0.21 (EtOAc/n-hexane, 2/1); mp
25
D
IR (KBr) 3139, 3035, 2349, 1717, 1637, 1402, 1344, 1178,
125–126 ꢁC; optical rotation ½aŠ +20.2 (c 0.11, CHCl3);
886, 813, 712, 524, 473, 464, 438 cmꢀ1 1H NMR
;
(200 MHz, CDCl3) d 8.10–7.80 (brs, 1H), 3.50 (dd, J=10.4,
4.0 Hz, 0.5H) and 3.36 (dd, J=10.4, 4.0 Hz, 0.5H), 3.10–
1.90 (m, 4H), 1.40–1.20 (m, 3H); MS (APCI, Pos.) m/z 241,
239 (M+H)+; HR-MS (MALDI-TOF, Pos.) calcd for
C7H9Br1Cl1N1O1+ H+: 237.9634; found: 237.9668.
(1R,5R,6S)-7-Bromo-7-chloro-5-methyl-2-azabicyclo[4.1.0]
heptan-3-one (47b). Compound 47b was prepared from
46b-anti in 95% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
Pale yellow powder; TLC Rf 0.33 (EtOAc/n-hexane,
2:1); mp 97–99 ꢁC; optical rotation [a]2D5 ꢀ56.9 (c 0.88,
CHCl3); IR (KBr) 3200, 2968, 1660, 1631, 1477, 1455,
(1S,5R,6R,7S)-7-Chloro-5-methyl-2-azabicyclo[4.1.0]hep-
tan-3-one (48d) and (1S,5R,6R,7R)-7-chloro-5-methyl-2-
azabicyclo[4.1.0]heptan-3-one (48e). Compounds 48d

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1739
and 48e were prepared from 48b in 42 and 54% yield,
respectively accordingto the same procedure as descri-
bed for the preparation of 47c and 47d from 47b.
49a-syn: pale yellow oil; TLC Rf 0.31 (EtOAc/n-hexane,
1:2); IR (neat) 2961, 2835, 1652, 1514, 1455, 1404, 1304,
1
1248, 1177, 1108, 1034, 808, 759, 648, 540 cmꢀ1; H
NMR (200 MHz, CDCl3) d 7.28 (d, J=8.8 Hz, 2H),
6.86 (d, J=8.8 Hz, 2H), 5.17 (d, J=15.6 Hz, 1H), 4.29
(d, J=15.6 Hz, 1H), 3.80 (s, 3H), 2.80–2.30 (m, 3H),
1.70–1.50 (m, 1H), 1.66 (s, 3H), 1.31 (d, J=6.4 Hz, 3H);
MS (APCI, Pos.) m/z 420 (M+H, 81Br, 81Br)+, 418
(M+H, 79Br, 81Br)+, 416; HR-MS (MALDI-TOF,
Pos.) calcd for C16H19Br2N1O2+ H+: 415.9861; found:
415.9868.
48d: pale yellow powder; TLC Rf 0.35 (EtOAc); mp 86–
88 ꢁC; optical rotation ½aŠD25-53.9 (c 0.52, CHCl3); IR
(KBr) 3187, 3063, 2962, 1685, 1489, 1396, 1350, 1206,
1050, 997, 886, 786, 519, 501 cmꢀ1; 1H NMR (200 MHz,
CDCl3) d 7.10–6.80 (brs, 1H), 3.04–2.92 (m, 1H), 2.70–
2.24 (m, 2H), 1.80–1.60 (m, 2H), 1.30–0.90 (m, 1H), 1.16
(d, J=6.6 Hz, 3H); MS (APCI, Pos.) m/z 160 (M+H)+.
48e: pale yellow powder; TLC Rf 0.20 (EtOAc); mp 89–
(dl)-(1R,5S,6S,7S)-2-(4-Methoxybenzyl)-1,5,7-trimethyl-
2-azabicyclo[4.1.0]heptan-3-one (49b-anti). Compound
49b-anti was prepared from 49a-anti in 65% yield
accordingto the same procedure as described for the
preparation of 39c-anti from 39a-anti. Pale yellow oil;
TLC Rf 0.24 (EtOAc/n-hexane, 1:3); 1H NMR
(200 MHz, CDCl3) d 7.21 (d, J=8.8 Hz, 2H), 6.82 (d,
J=8.8 Hz, 2H), 4.94 (d, J=14.6 Hz, 1H), 4.21 (d,
J=14.6 Hz, 1H), 3.79 (s, 3H), 2.30–2.02 (m, 2H), 1.70–
1.40 (m, 1H), 1.25 (s, 3H), 1.12 (d, J=6.8 Hz, 3H), 0.89
(d, J=6.2 Hz, 3H), 0.48 (m, 1H), 0.33 (t, J=5.8 Hz,
1H); MS (APCI, Pos.) m/z 274 (M+H)+.
25
91 ꢁC; optical rotation ½aŠ +15.1 (c 0.15, CHCl3); IR
D
(KBr) 3434, 2964, 1638, 1487, 1397, 1353, 1064, 722
cmꢀ1 1H NMR (200 MHz, CDCl3) d 6.70–6.50 (brs,
;
1H), 3.22 (dd, J=8.3, 5.7 Hz, 1H), 3.01 (ddd, J=9.8, 5.7,
4.2 Hz, 1H), 2.65–2.45 (m, 1H), 2.45–2.30 (m, 2H), 1.55–
1.40 (m, 1H), 1.24 (d, J=6.4 Hz, 3H); MS (APCI, Pos.)
m/z 160 (M+H)+; HR-MS (MALDI-TOF, Pos.) calcd
for C7H10Cl1N1O1+ H+: 160.0529; found: 160.0519.
(1S,5R,6R)-7-Bromo-7-fluoro-5-methyl-2-azabicyclo[4.1.0]
heptan-3-one (48c). Compound 48c was prepared from
46c-syn in 75% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
Pale yellow powder; TLC Rf 0.23 (EtOAc/n-hexane,
3:1); 1H NMR (200 MHz, CDCl3) d 6.70–6.55 (brs, 1H),
3.42 (dd, J=10.2, 4.4 Hz, 0.5H) and 3.27 (dd, J=10.4,
4.2 Hz, 0.5H), 2.55–1.00 (m, 4H), 1.32 (d, J=6.2 Hz,
3H); MS (APCI, Pos.) m/z 224 (M+H)+, 222.
(dl)-(1R,5S,6S,7S)-1,5,7-Trimethyl-2-azabicyclo[4.1.0]-
heptan-3-one (50). Compound 50 was prepared from
49b-anti in 90% yield accordingto the same procedure
as described for the preparation of 29a from 28a-anti.
Pale yellow oil; TLC Rf 0.36 (EtOAc); 1H NMR
(200 MHz, CDCl3) d 6.08 (brs, 1H), 2.22–1.86 (m, 3H),
1.32 (s, 3H), 1.15 (d, J=6.6 Hz, 3H), 1.07 (d, J=6.2 Hz,
3H), 0.89 (m, 1H), 0.41 (t, J=4.6 Hz, 1H); MS (APCI,
Pos.) m/z 154 (M+H)+.
(1S,5R,6R,7R)-7-Fluoro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-one (48g). Compound 48g was prepared from
48c in 22% yield accordingto the same procedure as
described for the preparation of 29h from 29c. Pale yel-
low powder; TLC Rf 0.51 (CHCl3/MeOH, 10:1); 1H
NMR (200 MHz, CDCl3) d 6.65–5.30 (brs, 1H), 4.57
(ddd, J=64.0, 6.4, 4.6 Hz, 1H), 2.78 (m, 1H), 2.65–2.33
(m, 2H), 2.16–1.95 (m, 1H), 1.33–1.13 (m, 1H), 1.21 (d,
J=6.2 Hz, 3H); MS (APCI, Pos.) m/z 144 (M+H)+,
124 (M–F)+.
General procedure for preparation of compounds 2–18
(1S,5S,6R)-7,7-Dichloro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (2). To a stirred solu-
tion of 29a (390 mg, 2.0 mmol) in CH2Cl2 (2 mL) was
added triethyloxonium tetrafluoroborate (2 M in
CH2Cl2, 1.5 mL, 3.0 mmol) under an argon atmosphere,
and the reaction mixture was stirred at room tempera-
ture for 20 h. Concentration of the reaction mixture
under reduced pressure gave a residue, which was used
for the subsequent reaction without further purification.
To a stirred solution of the compound obtained above
in EtOH (5 mL) was added saturated ethanolic ammo-
nia (5 mL) under an argon atmosphere at room tem-
perature and stirringwas continued for an additional 40
h. The reaction mixture was diluted with CHCl3 (25
mL) and the resultingprecipitates were removed by fil-
tration. The filtrate was concentrated under reduced
pressure. The reaction mixture was treated with 2 M
NaOH (30 mL) and extracted with CHCl3. The com-
bined organic layers were dried over anhydrous sodium
sulfate and evaporated under reduced pressure. The
residue was converted to its hydrochloride with 4 M
HCl/EtOAc–EtOH at 0 ꢁC and further purified by col-
umn chromatography on silica gel (CHCl3/MeOH, 5:1)
to afford 2 as a beige powder (137 mg, 30%). TLC Rf
(dl)-(1S,5S,6R)-7,7-Dibromo-2-(4-methoxybenzyl)-1,5-di-
methyl-2-azabicyclo[4.1.0]heptan-3-one (49a-anti) and
(dl)-(1R,5S,6S)-7,7-dibromo-2-(4-methoxybenzyl)-1,5-di-
methyl-2-azabicyclo[4.1.0]heptan-3-one (49a-syn). Com-
pounds 49a-anti and 49a-syn were prepared from 38 in
26 and 7% yield, respectively accordingto the same
procedure as described for the preparation of 28b-anti
and 28b-syn from 26.
49a-anti: pale yellow solid; TLC Rf 0.40 (EtOAc/n-hex-
ane, 1:2); mp 118–119 ꢁC; IR (KBr) 3435, 2965, 1655,
1512, 1438, 1305, 1243, 1182, 1033, 856, 756, 588, 512
1
cmꢀ1; H NMR (200 MHz, CDCl3) d 7.24 (d, J=8.8
Hz, 2H), 6.84 (d, J=8.8 Hz, 2H), 5.37 (d, J=15.4 Hz,
1H), 3.83 (d, J=15.4 Hz, 1H), 3.79 (s, 3H), 2.40–2.00
(m, 3H), 1.61–1.40 (m, 1H), 1.50 (s, 3H), 1.31 (d, J=6.2
Hz, 3H); MS (APCI, Pos.) m/z 420 (M+H, 81Br,
81Br)+, 418 (M+H, 79Br, 81Br)+, 416; HR-MS
(MALDI-TOF, Pos.) calcd for C16H19Br2N1O2+ H+:
415.9861; found: 415.9845.
0.35 (CHCl3/MeOH/AcOH, 10/1/1); mp 214–216 ꢁC;
25
D
optical rotation ½aŠ +44.6 (c 0.44, MeOH); IR (KBr)

1740
Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
3231, 3050, 1678, 1521, 1454, 1418, 1385, 1354, 1330,
1286, 1245, 1206, 1174, 1093, 1047, 1021, 986, 933, 894,
that were collected, 4523 were unique. Equivalent
reflections were merged. An empirical absorption cor-
rection was applied.
1
862, 827, 720, 560, 543, 489, 457, 435 cmꢀ1; H NMR
(200 MHz, DMSO-d6) ꢀ 10.41 (brs, 1H), 9.70 (brs, 1H),
7.25 (brs, 1H), 3.55 (d, J=9.8 Hz, 1H), 2.53 (dd,
J=15.6, 5.0 Hz, 1H), 2.36 (dd, J=15.6, 11.8 Hz, 1H),
2.12 (dd, J=9.8, 5.4 Hz, 1H), 2.03–1.80 (m, 1H), 1.27
(d, J=6.6 Hz, 3H); MS and HR-MS analysis showed
only the ion from C7H10Cl2N2 correspondingto the loss
of HCl, MS (FAB, Pos.) m/z 193 (M+H)+, 157 (M–
Cl)+; HR-MS (MALDI-TOF, Pos.) calcd for
C7H10Cl2N2+ H+: 193.0299; found: 193.0323.
The program package teXsan18 was used for analysis
and drawingfiugres. The positions of all the non-H
atoms were determined by the program SHEXLS86,19
and the H atoms of calculated from the coordinates of
non-H atoms and confirmed by the difference Fourier
synthesis.
Crystal data: orthorombic, space group P212121,
a=7.31(3), b=23.23(3), c=5.63(1) A, Z=4, 1910 mea-
sured reflections, 1280 with I>3.00ꢁ(I), 2ꢂ<130.02ꢁ,
R=0.063. Full information on the crystal structure can
be ordered from the CCDC, 12 Union Road, Cam-
bridge CB2 1EZ, UK, upon request quoting the
deposition number CCDC 191009.
(1S,5S,6R,7S)-7-Chloro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (3). Compound 3 was
prepared from 29e in 57% yield accordingto the same
procedure as described for the preparation of 2 from
29a. Brown oil; TLC Rf 0.33 (CHCl3/MeOH/AcOH,
25
D
10:1:1); optical rotation ½aŠ ꢀ5.92 (c 0.39, MeOH); IR
(neat) 3391, 2971, 2360, 2342, 1695, 1682, 1652, 1634,
1520, 1456, 1435, 1385, 1356, 1297, 1243, 1072, 932,
(1S,5S,6R,7R)-7-Fluoro-5-methyl-2-azabicyclo[4.1.0]hep-
tan-3-imine hydrochloride salt (5). Compound 5 was
prepared from 29h in 53% yield accordingto the same
procedure as described for the preparation of 2 from
1
890, 860, 792, 762, 668, 523 cmꢀ1; H NMR (200 MHz,
DMSO-d6) d 10.10–9.85 (brs, 1H), 9.20–9.00 (brs, 1H),
8.80–8.60 (brs, 1H), 3.58 (dd, J=3.9, 2.0 Hz, 1H), 3.19
(dd, J=9.4, 2.0 Hz, 1H), 2.45–2.38 (m, 1H), 2.36–2.05
(m, 2H), 1.75–1.56 (m, 1H), 1.10 (d, J=6.4 Hz, 3H); MS
and HR-MS analysis showed only the ion from
C7H11Cl1N2 correspondingto the loss of HCl, MS
(APCI, Pos.) m/z 159 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C7H11Cl1N2+ H+: 159.0689;
found: 159.0667.
29a. White powder; TLC Rf 0.21 (CHCl3/MeOH/
25
D
AcOH, 10:1:1); mp 157–158 ꢁC; optical rotation ½aŠ
+57.5 (c 0.16, MeOH); IR (KBr) 3089, 2876, 1684,
1627, 1454, 1424, 1365, 1343, 1332, 1233, 1157, 1046,
1032, 1013, 908, 869, 773, 736, 708, 603, 567, 491, 425
;
cmꢀ1 1H NMR (200 MHz, DMSO-d6) ꢀ 10.10–9.90
(brs, 1H), 9.50–9.20 (brs, 1H), 9.15–8.80 (brs, 1H), 4.78
(ddd, J=65.6, 6.2, 4.4 Hz, 1H), 3.05–2.80 (m, 1H),
2.60–2.28 (m, 2H), 2.12–1.82 (m, 1H), 1.45–1.15 (m,
1H), 1.18 (d, J=6.6 Hz, 3H); MS and HR-MS analysis
showed only the ion from C7H11F1N2 correspondingto
the loss of HCl, MS (APCI, Pos.) m/z 143 (M+H)+;
HR-MS (MALDI-TOF, Pos.) calcd for C7H11F1N2+
H+: 143.0985; found: 143.0974.
(1S,5S,6R,7R)-7-Chloro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (4). Compound 4 was
prepared from 29f in 53% yield accordingto the same
procedure as described for the preparation of 2 from
29a. White powder; TLC Rf 0.33 (CHCl3/MeOH/
25
D
AcOH, 10:1:1); mp 234–235 ꢁC; optical rotation ½aŠ
+68.1 (c 1.00, MeOH); IR (KBr) 3130, 2873, 2155,
2086, 2043, 1690, 1456, 1422, 1390, 1361, 1297, 1276,
1234, 1204, 1127, 966, 929, 917, 879, 841, 796, 771, 713,
684, 600, 536, 523, 481, 423 cmꢀ1; H NMR (200 MHz,
(dl)-(1S,5S,6S)-5,7,7-Trimethyl-2-azabicyclo[4.1.0]hep-
tan-3-imine hydrochloride salt (6). Compound 6 was
prepared from 40b in 25% yield accordingto the same
procedure as described for the preparation of 2 from
29a. White powder; TLC Rf 0.46 (CHCl3/MeOH/
AcOH, 15:2:1); mp 125–126 ꢁC; IR (KBr) 2956, 1677,
1
DMSO-d6) d 9.80–9.65 (brs, 1H), 9.30–9.10 (brs, 1H),
8.80–8.60 (brs, 1H), 3.64 (dd, J=7.7, 5.5 Hz, 1H), 3.04
(dd, J=8.9, 5.5 Hz, 1H), 2.41 (d, J=8.2 Hz, 2H), 1.90–
1.70 (m, 1H), 1.55–1.40 (m, 1H), 1.21 (d, J=6.8 Hz,
3H); MS and HR-MS analysis showed only the ion
from C7H11Cl1N2 correspondingto the loss of HCl, MS
(APCI, Pos.) m/z 159 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C7H11Cl1N2+H+: 159.0689;
found: 159.0690; anal. calcd for C7H12Cl2N2; C,
43.10%, H, 6.20%, N, 14.36%, Cl, 36.34%; found; C,
43.08%; H, 6.25%, N, 14.24%.
1456, 1234, 1071, 969, 881, 691 cmꢀ1
;
1H NMR
(200 MHz, CDCl3) d 9.40 (brs, 1H), 8.99 (brs, 1H), 8.71
(brs, 1H), 2.74 (dd, J=15.0, 3.0 Hz, 1H), 2.45 (d, J=8.0
Hz. 1H), 2.10 (m, 1H), 1.74 (m, 1H), 1.22 (d, J=6.6 Hz,
3H), 1.07 (s, 3H), 0.99 (s, 3H), 0.83 (dd, J=8.0, 6.0 Hz,
1H); MS and HR-MS analysis showed only the ion
from C9H16N2 correspondingto the loss of HCl, MS
(APCI, Pos.) m/z 153 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C9H16N2+ H+: 153.1392; found:
153.1373.
X-ray crystallography of 4
(dl)-(1R,5S,6S,7S)-5,7-Dimethyl-2-azabicyclo[4.1.0]hep-
tan-3-imine hydrochloride salt (7). Compound 7 was
prepared from 40c in 74% yield accordingto the same
procedure as described for the preparation of 2 from
29a. Pale yellow oil; TLC Rf 0.40 (CHCl3/MeOH/
AcOH, 15:2:1); IR (neat) 2961, 1677, 1514, 1462, 1384,
Colorless platelet crystals of 4 were obtained from
EtOH/EtOAc (6:1) solution. A suitable crystal of 4
havingthe approximate dimension of 0.10 Â0.20Â0.40
mm was mounted on a glass fiber. All measurements
were made on a Rigaku AFC5R diffractometer with gra-
phite monochromated Cu-Ka radiation and a RU-200
rotatinganode X-ray generator. Of the 9156 reflections
1
1356, 1060, 669 cmꢀ1; H NMR (200 MHz, CDCl3) d
9.80 (brs, 1H), 9.22 (brs, 1H), 8.72 (brs, 1H), 2.66 (dd,

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1741
J=16.0, 4.0 Hz, 1H), 2.43 (dd, J=8.0, 3.0 Hz, 1H), 2.29
(dd, J=16.0, 9.0 Hz, 1H), 1.91 (m, 1H), 1.21 (d, J=6.8
Hz, 3H), 1.05 (d, J=5.2H, 3H), 0.96 (m, 1H), 0.85 (m,
1H); MS and HR-MS analysis showed only the ion from
C8H14N2 correspondingto the loss of HCl, MS (APCI,
Pos.) m/z 139 (M+H)+; HR-MS (MALDI-TOF, Pos.)
calcd for C8H14N2+ H+: 139.1235; found: 139.1225.
procedure as described for the preparation of 2 from
29a. Pale yellow oil; TLC Rf 0.35 (CHCl3/MeOH/
AcOH, 20:4:1); IR (neat) 3122, 1679, 1636, 1525, 1458,
1430, 1382, 1355, 1215, 1185, 1084, 988, 907, 861, 649
cmꢀ1 1H NMR (200 MHz, DMSO-d6) d 10.07 (brs,
;
1H), 9.01 (brs, 1H), 8.61 (brs, 1H), 5.49 (ddd, J=17.2,
10.2, 8.4 Hz, 1H), 5.08 (dd, J=17.2, 1.8 Hz, 1H), 4.95
(dd, J=10.2, 1.8 Hz, 1H), 2.88 (dt, J=8.4, 3.0 Hz, 1H),
2.54 (m, 1H), 2.28 (dd, J=16.0, 7.0 Hz, 1H), 2.11 (m,
1H), 1.82 (m, 1H), 1.29 (m, 1H), 1.09 (d, J=6.6 Hz,
3H); MS and HR-MS analysis showed only the ion
from C9H14N2 correspondingto the loss of HCl, MS
(APCI, Pos.) m/z 151 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C9H14N2+ H+: 151.1235; found:
151.1230.
(dl)-(1R,5S,6S,7R)-5,7-Dimethyl-2-azabicyclo[4.1.0]hep-
tan-3-imine hydrochloride salt (8). Compound 8 was
prepared from 40e in 30% yield accordingto the same
procedure as described for the preparation of 2 from
29a. White powder; TLC Rf 0.15 (CHCl3/MeOH/
AcOH, 20:1:1); mp 183–184 ꢁC; IR (KBr) 3258, 3085,
2964, 1676, 1630, 1458, 1418, 1342, 1204, 1086, 781,
1
747, 706 cmꢀ1; H NMR (200 MHz, DMSO-d6) d 9.75
(brs, 1H), 8.97 (brs, 1H), 8.77 (brs, 1H), 2.66 (dd,
J=8.0, 6.0 Hz, 1H), 2.30 (dd, J=15.5, 4.0 Hz, 1H), 2.19
(dd, J=15.5, 11.5 Hz, 1H), 1.56 (m, 1H), 1.08 (d, J=6.5
Hz, 3H), 1.12–1.02 (m, 1H), 1.00–0.88 (m, 1H), 0.79 (d,
J=6.0 Hz, 3H); MS and HR-MS analysis showed only
the ion from C8H14N2 correspondingto the loss of HCl,
MS (FAB, Pos.) m/z 139 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C8H14N2+ H+: 139.1235; found:
139.1206.
(dl)-(1R,5S,6S,7R)-7-Butyl-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (12). Compound 12
was prepared from 40g in 34% yield accordingto the
same procedure as described for the preparation of 2
from 29a. Beige powder; TLC Rf 0.53 (CHCl3/MeOH/
AcOH, 15:2:1); mp 187–188 ꢁC; IR (KBr) 3015, 2927,
2858, 1678, 1631, 1459, 1419, 1349, 1100, 748, 602 cmꢀ1
;
1H NMR (200 MHz, DMSO-d6) d 9.62 (brs, 1H), 9.15
(brs, 1H), 9.01 (brs, 1H), 2.81–2.71 (m, 2H), 2.17 (dd,
J=16.0, 12.0 Hz, 1H), 1.73 (m, 1H), 1.42–1.25 (m, 6H),
1.23 (d, J=6.6 Hz, 3H), 1.12–1.00 (m, 2H), 0.91 (t,
J=6.2 Hz, 3H); MS and HR-MS analysis showed only
the ion from C11H20N2 correspondingto the loss of
HCl, MS (APCI, Pos.) m/z 181 (M+H)+; HR-MS
(MALDI-TOF, Pos.) calcd for C11H20N2+ H+:
181.1705; found: 181.1710.
(1R,5S,6S,7R)-5,7-Dimethyl-2-azabicyclo[4.1.0]heptan-
3-imine hydrochloride salt (9). Compound 9 was pre-
pared from 32 in 4% yield accordingto the same pro-
cedure as described for the preparation of 2 from 29a.
Beige powder; TLC Rf 0.50 (CHCl3/MeOH/AcOH,
25
D
5:1:1); mp 182–184 ꢁC; optical rotation ½aŠ +17.2 (c
0.10, MeOH); IR (KBr) 3357, 3083, 1680, 1627, 1511,
1459, 1423, 1343, 1282, 1245, 1204, 1180, 1100, 1095,
1070, 978, 712, 528 cmꢀ1; 1H NMR (200 MHz, DMSO-
d6) d 9.70–9.65 (brs, 1H), 9.20–9.00 (brs, 1H), 8.70–8.60
(brs, 1H), 2.73 (t, J=7.5 Hz, 1H), 2.45–2.15 (m, 2H),
1.80–1.50 (m, 1H), 1.15 (d, J=7.0 Hz, 3H), 1.20–0.90
(m, 2H), 0.85 (d, J=6.4 Hz, 3H); MS and HR-MS
analysis showed only the ion from C8H14N2 corre-
spondingto the loss of HCl, MS (APCI, Pos.) m/z 139
(M+H)+; HR-MS (MALDI-TOF, Pos.) calcd for
C8H14N2+ H+: 139.1235; found: 139.1228.
(1R,5R,6S)-7,7-Dichloro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (13). Compound 13
was prepared from 47a in 24% yield accordingto the
same procedure as described for the preparation of 2
from 29a. Beige powder; TLC Rf 0.35 (CHCl3/MeOH/
25
D
AcOH, 10:1:1); mp 213–214 ꢁC; optical rotation ½aŠ
ꢀ45.6 (c 0.45, MeOH); IR (KBr) 3234, 3068, 1678,
1522, 1452, 1418, 1385, 1355, 1330, 1245, 1207, 1093,
1022 cmꢀ1; 1H NMR (200 MHz, DMSO-d6) d 9.66 (brs,
3H), 3.55 (d, J=9.6 Hz, 1H), 2.57–2.28 (m, 2H), 2.11
(dd, J=10.0, 5.6 Hz, 1H), 2.01–1.84 (m, 1H), 1.27 (d,
J=6.6 Hz, 3H); MS and HR-MS analysis showed only
the ion from C7H10Cl2N2 correspondingto the loss of
HCl, MS (FAB, Pos.) m/z 193 (M+H)+; HR-MS
(MALDI-TOF, Pos.) calcd for C7H10Cl2N2+ H+:
193.0299; found: 193.0309.
(dl)-(1R,5S,6S,7R)-7-Ethyl-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (10). Compound 10
was prepared from 43 in 88% yield accordingto the
same procedure as described for the preparation of 2
from 29a. Off-white powder; TLC Rf 0.48 (CHCl3/
MeOH/AcOH, 15:2:1); mp 180–181 ꢁC; IR (KBr) 3089,
1678, 1629, 1418, 1358, 1308, 1205, 889, 801, 761, 701
cmꢀ1; 1H NMR (200 MHz, DMSO-d6) d 9.64 (brs, 1H),
9.35 (brs, 1H), 8.90 (brs, 1H), 2.90–2.70 (m, 2H), 2.15 (dd,
J=16.0, 12.0 Hz, 1H), 1.77 (m, 1H), 1.40–1.20 (m, 2H),
1.24 (d, J=6.6 Hz, 3H), 1.12–0.96 (m, 2H), 1.01 (t, J=6.8
Hz, 3H); MS and HR-MS analysis showed only the ion
from C9H16N2 correspondingto the loss of HCl, MS
(APCI, Pos.) m/z 153 (M+H)+; HR-MS (MALDI-TOF,
Pos.) calcd for C9H16N2+ H+: 153.1392; found: 153.1372.
(1R,5R,6S,7S)-7-Chloro-5-methyl-2-azabicyclo[4.1.0]hep-
tan-3-imine hydrochloride salt (14). Compound 14 was
prepared from 47c in 48% yield accordingto the same
procedure as described for the preparation of 2 from
29a. Beige powder; TLC Rf 0.33 (CHCl3/MeOH/AcOH,
25
D
10:1:1); mp 231–232 ꢁC; optical rotation ½aŠ ꢀ66.9 (c
0.25, MeOH); IR (KBr) 3257, 3031, 2873, 1682, 1628,
1459, 1422, 1390, 1362, 1296, 1276, 1234, 1204, 1071,
1011, 879, 770, 716, 684, 541 cmꢀ1; 1H NMR (200 MHz,
DMSO-d6) d10.05–9.90 (brs, 1H), 9.50–9.30 (brs, 1H),
9.00–8.80 (brs, 1H), 3.64 (dd, J=7.8, 5.6 Hz, 1H), 3.05
(dd, J=8.8, 5.6 Hz, 1H), 2.55–2.30 (m, 2H), 1.96–1.72
(m, 1H), 1.56–1.40 (m, 1H), 1.20 (d, J=6.6 Hz, 3H); MS
(dl)-(1R,5S,6S,7S)-5-Methyl-7-vinyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (11). Compound 11
was prepared from 45 in 6% yield accordingto the same

1742
Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
and HR-MS analysis showed only the ion from
C7H11Cl1N2 correspondingto the loss of HCl, MS
(APCI, Pos.) m/z 159 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C7H11Cl1N2+ H+: 159.0689;
found: 159.0697.
of saturated ethanolic ammonia. Brown oil; TLC Rf
25
D
0.24 (CHCl3/MeOH, 5:1); optical rotation ½aŠ +6.6 (c
0.48, MeOH); IR (neat) 3626, 3585, 3333, 3040, 2970,
2934, 2879, 1661, 1586, 1499, 1486, 1456, 1428, 1382,
1361, 1296, 1280, 1254, 1218, 1072, 870, 752, 697, 522
1
cmꢀ1; H NMR (200 MHz, DMSO-d6) d 10.40–10.20
(1S,5R,6R,7R)-7-Chloro-5-methyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (15). Compound 15
was prepared from 48e in 13% yield accordingto the
same procedure as described for the preparation of 2
from 29a. Beige amorphous; TLC Rf 0.30 (CHCl3/
(brs, 1H), 10.05–9.90 (brs, 1H), 7.50–7.30 (m, 5H), 4.53
(m, 2H), 3.71 (dd, J=7.7, 5.6 Hz, 1H), 3.12 (dd, J=9.2,
5.6 Hz, 1H), 2.70–2.40 (m, 2H), 2.00–1.80 (m, 1H),
1.60–1.45 (m, 1H), 1.23 (d, J=7.0 Hz, 3H); MS and
HR-MS analysis showed only the ion from
C14H17Cl1N2 correspondingto the loss of HCl, MS
(APCI, Pos.) m/z 249 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C14H17Cl1N2+ H+: 249.1159;
found: 249.1131.
MeOH/AcOH, 10:1:1); mp 76–78 ꢁC; Optical rotation
25
D
½aŠ +24.8 (c 0.10, MeOH); IR (KBr) 3289, 3108, 2965,
2876, 1679, 1530, 1511, 1456, 1413, 1376, 1355, 1279,
1249, 1061, 1004, 920, 875, 835, 810, 722, 609, 546, 484,
450, 424 cmꢀ1; 1H NMR (200 MHz, DMSO-d6) d10.50–
10.20 (brs, 1H), 9.20–8.90 (brs, 1H), 8.90–8.60 (brs, 1H),
3.59 (dd, J=8.0, 6.0 Hz, 1H), 3.40–3.10 (m, 1H), 2.70
(dd, J=16.7, 6.2 Hz, 1H), 2.50–2.30 (m, 1H), 2.20 (dd,
J=16.7, 11.0 Hz, 1H), 1.80-1.60 (m, 1H), 1.16 (d, J=6.6
Hz, 3H); MS and HR-MS analysis showed only the ion
from C7H11Cl1N2 correspondingto the loss of HCl, MS
(APCI, Pos.) m/z 159 (M+H)+; HR-MS (MALDI-
TOF, Pos.) calcd for C7H11Cl1N2+ H+: 159.0689;
found: 159.0676.
Biological assays
Preparation of partially purified enzyme and determina-
tion of Ki values. Human eNOS was overexpressed in
Sf-21 cells, by infectingthe cells with baculovirus carry-
ingheNOS cDNA. The hiNOS was overexpressed in
A549 by stimulation with LPS (10 mg/mL) plus cyto-
kines (10 ng/mL TNF-a, 5 ng/mL IL-1b and 100 ng/mL
interferon-g). Human eNOS and iNOS were partially
purified by chromatography on 20,50-ADP-Sepharose
gels. NOS activity was determined by the method for
the conversion of [14C]-l-arginine to l-citrulline with a
minor modification. The conversion rates for various
concentrations of the test compounds and l-arginine
were measured. Dixon and lineweaver-Bulk plots were
constructed to determine the Ki values and the mode of
inhibition. Selectivity was evaluated as the rate of the
IC50 values for heNOS and hiNOS.
(1S,5R,6R,7R)-7-Fluoro-5-methyl-2-azabicyclo[4.1.0]hep-
tan-3-imine hydrochloride salt (16). Compound 16 was
prepared from 48g in 9% yield accordingto the same
procedure as described for the preparation of 2 from
29a. Beige amorphous; TLC Rf 0.28 (CHCl3/MeOH/
25
D
MeOH); IR (KBr) 3430, 2925, 2853, 1734, 1678, 1453,
AcOH, 20:4:1); optical rotation ½aŠ +8.9 (c 0.10,
1
1151, 756, 698 cmꢀ1; H NMR (200 MHz, CDCl3) d
10.8–10.3 (brs, 1H), 8.81 (brs, 2H), 4.67 (m, 1H), 3.12–
2.75 (m, 2H), 2.45–2.10 (m, 2H), 1.50–1.25 (m, 1H), 1.26
(3H, d, J=6.0 Hz, 3H); MS and HR-MS analysis
showed only the ion from C7H11F1N2 correspondingto
the loss of HCl, MS (APCI, Pos.) m/z 143 (M+H)+,
123 (M–F)+; HR-MS (MALDI-TOF, Pos.) calcd for
C7H11F1N2+ H+: 143.0985; found: 143.0987.
Enzyme assay with recombinant mouse iNOS. Recombi-
nant mouse iNOS was purchased from Cayman Che-
mical (Cat. No. 60862) and the inhibitory activities of
the test compounds were evaluated by measuringthe
conversion rate from [14C]-l-arginine to [14C]-l-citrul-
line, and then the IC50 values were determined.
(dl)-(1R,5S,6S,7S)-1,5,7-Trimethyl-2-azabicyclo[4.1.0]-
heptan-3-imine hydrochloride salt (17). Compound 17
was prepared from 50 in 48% yield accordingto the
same procedure as described for the preparation of 2
from 29a. Brown oil; TLC Rf 0.35 (CHCl3/MeOH/
AcOH, 20:1:1); IR (neat) 2962, 1681, 1532, 1456, 1430,
The ID50 value was determined from a log–logit trans-
formation of the dose–response curves (2 and 4; 3, 10,
30 mg/kg, sc, l-NMMA; 10, 30 and 100 mg/kg, sc). The
ID50 value was defined as the concentration of test
compound that produced a 50% inhibition in the NOx
accumulation induced by LPS treatment alone.17 The
MTD was defined as the maximum dose at which no
death was observed within 24 h after an intravenous
injection administration. The doses used were 10, 20, 30,
40, and 50 mg/kg for 2 and 4 and 1000, 2000, 3000,
4000, and 5000 mg/kg for l-NMMA.
1393, 1335, 1277, 1210, 1049, 751 cmꢀ1 1H NMR
;
(200 MHz, DMSO-d6) d 10.17 (brs, 1H), 8.89 (brs, 1H),
8.46 (brs, 1H), 2.42 (dd, J=16.4, 5.4 Hz, 1H), 2.24 (dd,
J=16.4, 7.4 Hz, 1H), 1.98 (m, 1H), 1.31 (s, 3H), 1.20–
0.90 (m, 1H), 1.02 (m, 6H), 0.62 (m, 1H); MS and HR-
MS analysis showed only the ion from C9H16N2 corre-
spondingto the loss of HCl, MS (APCI, Pos.) m/z 153
(M+H)+, 97; HR-MS (MALDI-TOF, Pos.) calcd for
C9H16N2+ H+: 153.1392; found: 153.1363.
Inhibition of NOx accumulation and the maximum toler-
ated dose (MTD) in mice. The test compounds or saline
were administered subcutaneously at 3 h after LPS (10
mg/kg, iv) injection into 7 week old Balb/c mice
(Charles River Japan, Inc.). Blood was collected by
venipuncture from the abdominal aorta under light
anesthesia at 6 h after LPS treatment. Plasma was
obtained by centrifugation and the concentration of
accumulated NOx over 3 h was determined by the
N-[(1S,3E,5S,6R,7R)-7-Chloro-5-methyl-2-azabicyclo
[4.1.0]hept-3-ylidene]-1-phenylmethanamine hydrochloride
salt (18). Compound 18 was prepared from 29f in 28%
yield accordingto the same procedure as described for
the preparation of 2 from 29a usingbenzylamine instead

Y. Kawanaka et al. / Bioorg. Med. Chem. 11 (2003) 1723–1743
1743
method described below. To evaluate the acute toxicity,
the MTD (iv maximum dose where no death was
observed within 24 h after the administration) of the test
compound was determined.
3. Griffith, O. W.; Stuehr, D. J. Annu. Rev. Physiol. 1995, 57,
707.
4. Marletta, M. A. J. Biol. Chem. 1993, 268, 12231.
5. Kerwin, J. F., Jr.; Lancaster, J. R., Jr.; Feldman, P. L.
J. Med. Chem. 1995, 38, 4343.
6. Olken, N. M.; Marletta, M. A. Biochemistry 1993, 32, 9677.
7. Feldman, P. L.; Griffith, O. W.; Hong, H.; Stuehr, D. J.
J. Med. Chem. 1993, 36, 491.
8. Rees, D. D.; Palmer, R. M. J.; Moncada, S. Proc. Natl.
Acad. Sci. U.S.A. 1989, 86, 3375.
9. Misko, T. P.; Moore, W. M.; Kasten, T. P.; Nickols, G. A.;
Corbett, J. A.; Tilton, R. G.; McDaniel, M. L.; Williamson,
J. R.; Currie, M. G. Eur. J. Pharmacol. 1993, 233, 119.
10. Garvey, E. P.; Oplinger, J. A.; Tanoury, G. J.; Sherman,
P. A.; Fowler, M.; Marshall, S.; Harmon, M. F.; Paith, J. E.;
Furfine, E. S. J. Biol. Chem. 1994, 269, 26669.
Measurement of nitrite/nitrate. Nitrite and nitrate, the
oxidized form of nitric oxide that accumulated in the
culture medium and plasma were determined by the use
of nitrite/nitrate colorimetric assay kit (Cayman Che-
mical, Cat. No. 780001). Basically, the nitrate in the
sample was reduced to nitrite with a nitrate reductase
contained in the assay kit; nitrite levels were then deter-
mined spectrophotometrically as the total NOx concen-
tration.
11. Webber, R. K.; Metz, S.; Moore, W. M.; Connor, J. R.;
Currie, M. G.; Fok, K. F.; Hagen, T. J.; Hansen, D. W., Jr.;
Jerome, G. M.; Manning, P. T.; Pitzele, B. S.; Toth, M. V.;
Trivedi, M.; Zupec, M. E.; Tjoeng, F. S. J. Med. Chem. 1998,
41, 96.
12. Hagmann, W. K.; Galdwell, C. G.; Chen, P.; Durette,
P. L.; Esser, C. K.; Lanza, T. J.; Kopka, I. E.; Guthikonda,
R.; Shah, S. K.; MacCoss, M.; Chabin, R. M.; Fletcher, D.;
Grant, S. K.; Green, B. G.; Humes, J. L.; Kelly, T. M.; Luell,
S.; Meurer, R.; Moore, V.; Pacholok, S. G.; Pavia, T.; Wil-
liams, H. R.; Wong, K. K. Bioorg. Med. Chem. Lett. 2000, 10,
1975.
13. Kawanaka, Y.; Kobayashi, K.; Kusuda, S.; Tatsumi, T.;
Murota, M.; Nishiyama, T.; Hisaichi, K.; Fujii, A.; Hirai, K.;
Naka, M.; Komeno, M.; Nakai, H.; Toda, M. Bioorg. Med.
Chem. Lett. 2002, 12, 2291.
14. (a) Leotta, G. J., III.; Overman, L. E.; Welmaker, G. S.
J. Org. Chem. 1994, 59, 1946. (b) Theisen, P. D.; Heathcock,
C. H. J. Org. Chem. 1993, 58, 142. (c) Francis, C. J.; Jones,
J. B. J. Chem. Soc., Chem. Commun. 1984, 579.
Docking study. Dockingstudy was performed using
InsightII/Discover molecular modeling package (Accel-
rys, San Diego, CA) with CVFF force field on a SGI
Octane2 workstation with R12000 processors. Simu-
lated-annealingprocedure in ags phase was used to
search the complex model of the lowest-energy. Initial
protein structure of hiNOS (PDB code 2NSI) was
obtained from the Protein Data Bank. All the hydro-
gens were added and their positions were energetically
optimized. Duringsimulation, all the heavy atoms were
fixed (dielectric constant: "=4r ).
First of all, a complex model of low-energy, consisting
of 4 with hiNOS was constructed, in which 4 was put
near the Glu377 residue of the enzyme to form a salt
bridge with its amidine group. A complex model was
first equilibrated by runningdynamics (dielectric con-
stant: "=4r ) for 6 ps while increasingthe temperature
from 50 to 1200 K by time steps of 1 fs, after which the
resultingconformations were sampled every 5000 steps
over a span of 300 ps at 1200 K to yield 60 snapshots.
Each snapshot was then equilibrated for 4 ps while
decreasingthe temperature from 1200 to 200 K, fol-
lowed by 200 K simulation for 4 ps. Each annealed
model was energetically optimized using the steepest
descent method followed by the conjugate-gradient
method to an energy difference of 0.001 kcal/mol
between successive iterations.
15. Kitatani, K.; Hiyama, T.; Nozaki, H. J. Am. Chem. Soc.
1975, 97, 949.
16. Stereochemistry of syn-isomer and anti-isomer was deter-
mined usingtwo-dimensional NMR (2D NMR) technique
such as nuclear Overhauser effect spectroscopy (NOESY).
NOE observed between spatially close protons as illustrated in
the analysis of 28d–anti (Fig. 1) supported the described ste-
reochemistry. Same spectral analysis was applied to the
correspondingamides prior to the conversion to the amidine
derivatives.
17. Naka, M.; Nanbu, T.; Kobayashi, K.; Kamanaka, Y.;
Komeno, M.; Yanase, R.; Fukutomi, T.; Fujimura, S.; Seo,
H. G.; Fujiwara, N.; Ohuchida, S.; Suzuki, K.; Kondo, K.;
Taniguchi, N. Biochem. Biophys. Res. Commun. 2000, 270,
663.
References and Notes
18. teXsan: Molecular Structure Corporation; 1993.
19. Sheldrick, G. M. In Crystallographic Computing 3; Shel-
1. Moncada, S.; Higgs, E. A. FASEB 1995, 9, 1319.
2. Kerwin, J. F., Jr.; Heller, M. Med. Res. Rev. 1994, 14, 23.
drick, G. M.; Kruger, C.; Goddard, R., Eds.; Oxford Uni-
¨
versity Press, 1985; pp 175–189.