Design and preparation of potent, nonpeptidic, bioavailable renin inhibitors

Article abstract of DOI:10.1021/jm900022f

Starting from known piperidine renin inhibitors, a new series of 3,9-diazabicyclo[3.3.1]nonene derivatives was rationally designed and prepared. Optimization of the positions 3, 6, and 7 of the diazabicyclonene template led to potent renin inhibitors. The substituents attached at the positions 6 and 7 were essential for the binding affinity of these compounds for renin. The introduction of a substituent attached at the position 3 did not modify the binding affinity but allowed the modulation of the ADME properties. Our efforts led to the discovery of compound (+)-26g that inhibits renin with an IC ₅₀ of 0.20 nM in buffer and 19 nM in plasma. The pharmacokinetics properties of this and other similar compounds are discussed. Compound (+)-26g is well absorbed in rats and efficacious at 10 mg/kg in vivo.

Full text of DOI:10.1021/jm900022f

J. Med. Chem. 2009, 52, 3689–3702
3689
Design and Preparation of Potent, Nonpeptidic, Bioavailable Renin Inhibitors†
Olivier Bezenc¸on,* Daniel Bur, Thomas Weller, Sylvia Richard-Bildstein, Lubosˇ Remenˇ, Thierry Sifferlen,
Olivier Corminboeuf, Corinna Grisostomi, Christoph Boss, Lars Prade, Ste´phane Delahaye, Alexander Treiber, Panja Strickner,
Christoph Binkert, Patrick Hess, Beat Steiner, and Walter Fischli
Drug DiscoVery and Preclinical Research, Actelion Pharmaceuticals Ltd., Gewerbestrasse 16, CH-4123 Allschwil, Switzerland
ReceiVed January 9, 2009
Starting from known piperidine renin inhibitors, a new series of 3,9-diazabicyclo[3.3.1]nonene derivatives
was rationally designed and prepared. Optimization of the positions 3, 6, and 7 of the diazabicyclonene
template led to potent renin inhibitors. The substituents attached at the positions 6 and 7 were essential for
the binding affinity of these compounds for renin. The introduction of a substituent attached at the position
3 did not modify the binding affinity but allowed the modulation of the ADME properties. Our efforts led
to the discovery of compound (+)-26g that inhibits renin with an IC50 of 0.20 nM in buffer and 19 nM in
plasma. The pharmacokinetics properties of this and other similar compounds are discussed. Compound
(+)-26g is well absorbed in rats and efficacious at 10 mg/kg in vivo.
Introduction
inhibitors were highly potent peptidomimetics, like compound
1 (zankiren, IC50 value in plasma of 1.1 nM,4 Figure 1), which
suffered from unacceptable low bioavailability. A second
generation of inhibitors appeared in the late 1990s. On one hand
a new compound class arose from the former peptidomimetic
structures and culminated in compound 2 (aliskiren, IC50 value
in buffer of 0.6 nM), which was approved and launched recently,
despite a rather modest oral bioavailability of 4% in humans.5
On the other hand, piperidine derivatives, which were initially
identified from a screening effort, represent a completely new
scaffold for renin inhibitors.6 Extensive efforts in medicinal
chemistry were conducted for new compounds that had no
resemblance with the natural substrate. Compound 3 (Ro-66-
1168; IC50 value in buffer of 0.039 nM) is a prominent member
of this class.6d Unfortunately, these piperidine derivatives were
not suitable for clinical development because of inadequate
pharmacokinetic properties. Recently, ketopiperazine derivatives,
which are a further development of this class, have been found
to be renin inhibitors with IC50 values as low as 4 nM in buffer.7
Very recently, 2,4-diaminopyrimidine derivatives were reported
to inhibit renin quite potently as well.8
Cardiovascular diseases remain a main cause of mortality and
morbidity around the world. In 2005, 17.5 million people died
from cardiovascular diseases, more than from any other illness.1
In particular, hypertension causes 7.1 million premature deaths
per year.2 Moreover, hypertension plays a major etiologic role
in the development of cerebrovascular diseases, ischemic heart
diseases, and cardiac and renal failures.
The renin-angiotensin-aldosterone system (RAASa) is one
of the major and most intensively studied regulating systems
of the arterial blood pressure in humans. It plays a primordial
role not only in cardiovascular diseases but also in renal diseases
and other metabolic diseases as well.3 The RAAS consists of a
two-step cascade. First, the aspartic proteinase renin cleaves its
only known substrate, angiotensinogen, in the rate-limiting step
to the decapeptide angiotensin I. In a second step, angiotensin
I is cleaved either by the metalloproteinase angiotensin-
converting enzyme (ACE) or by the serine proteinase chymase
to the tensoactive octapeptide angiotensin II. The exclusive and
rate-determining function in the RAAS cascade makes renin
an ideal pharmacological drug target.
However, from the point of view of a medicinal chemist, renin
proved to be a very challenging target. First generation renin
Design of a New Class of Renin Inhibitors
On the basis of the temporarily available structural information
for piperidine derived renin inhibitors6c (e.g., compound 3), we
embarked on structure-based design of new compound classes with
improved pharmacokinetic properties. In all three renin structures
(PDB codes 1PR7, 1PR8, 1UHQ, meanwhile retracted from the
data bank) a protonated secondary amine of the piperidine ring
resides between the two catalytically important aspartate residues,
thereby expelling the catalytic water. Two strong hydrogen bonds
between the protonated ring nitrogen and each of the aspartate
carboxylates anchor and orient the piperidine in the active site.
Our initial design efforts started with a piperidine moiety located
as found in the three template structures mentioned above. The
renin inhibitor in 1UHQ occupies part of the S1 and S3 subpockets
with its 3-substituent, while the long and flexible 4-substituent
intrudes in a large newly formed pocket resulting from a reorienta-
tion of the ꢀ-hairpin loop, typically dubbed “flap”, and synchronous
side chain rearrangements of Trp41, Leu73, and Tyr75 (highlighted
in red in Figure 2). The S2 pocket, as well as space typically
occupied by the substrate backbone, remains unoccupied. It was
†
Crystal structures are available at www.rcsb.org. PDB codes are as
follows: 3G62, 3G70, 3G72.
* To whom correspondence should be addressed. Phone: +41 61 565
65 77. Fax: +41 61 565 65 00. E-mail: olivier.bezencon@actelion.com.
a Abbreviations: ACE, angiotensin cleaving enzyme; Ac, acetyl; ADME,
absorption, distribution, metabolism, excretion; Ang, angiotensin; aq,
aqueous; Ar, aryl; arom, aromatic; Boc, tert-butyloxycarbonyl; BSA, bovine
serum albumine; Bu, butyl; n-BuLi, n-butyllithium; conc, concentrated;
DIPEA, diisopropylethylamine; DMAP, 4-N,N-dimethylaminopyridine;
DMF, N,N-dimethylformamide; EDC ·HCl, ethyl-N,N-dimethylaminopro-
pylcarbodiimide hydrochloride; EDTA, ethylenediaminetetraacetic acid;
EIA, enzyme immunoassay; equiv, equivalent(s); ELSD, evaporative light
scattering detector; ESI, electrospray ionization; Et, ethyl; F, bioavailability;
FC, flash chromatography; HOBt, hydroxybenzotriazol; HPLC, high
performance liquid chromatography; LC, liquid chromatography; Me,
methyl; org, organic; MS, mass spectrometry; PBS, phosphate buffered
saline; Ph, phenyl; phe, phenyl; RAAS, renin angiotensin aldosterone
system; Rf, retention factor; sat., saturated; SAR, structure-activity relation-
ship; T1/2, half-life; TBAF, tetrabutylammonium fluoride trihydrate; TBDMS,
tert-butyldimethylsilyl; TBME, tert-butyl methyl ether; THF, tetrahydro-
furan; Tmax, time of maximum concentration; Tf, trifluoromethylsulfonyl;
TLC, thin layer chromatography; tR, retention time.
10.1021/jm900022f CCC: $40.75
2009 American Chemical Society
Published on Web 04/09/2009

3690 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12
Bezenc¸on et al.
Figure 1. Chemical structures of zankiren (1), aliskiren (2), and Ro-66-1168 (3).
Figure 2. Design of the 3,9-diazabicyclononene template from the corresponding piperidine, with their numbering systems, and X-ray structure of
human recombinant renin complexed with (+)-26h.
assumed that the extended hydrophobic interactions in the S1, S3
(filled by the tertiary amide in position 6 of the diazabicyclononene
in Figure 2), and the “flap” pockets (filled by the substituent
attached in position 7 of the diazabicyclononene in Figure 2) are
key contributors to the high affinity of this inhibitor. In order to
extend the number and direction of template exit vectors, positions
2 and 6 of the piperidine ring were bridged with a C-N-C unit
to give a diazabicyclononene moiety (positions 2, 3, and 4 of the
diazabicyclononene in Figure 2). This increased structural versatility
would allow a better fine-tuning of the physicochemical parameters
of our compounds. Introduction of a double bond at the 6,7-position
of this bicyclic system simplified synthetic accessibility and
diminished compound complexity with minor alteration of the
respective exit vectors. The substituent at position 6 had to be
altered, since we wanted to refrain from introducing ether substit-
uents at this position in order to avoid the generation of unstable
vinylic ethers. Fortunately it turned out that the ester group resulting
from the applied synthesis pathway (vide infra) could be easily
converted into an amide which led to very potent compounds after
some further optimization. The substituent in position 4 of the
piperidine ring was transformed to the newly generated position 7
in the bicyclic template but kept constant for initial compounds.
4-bromocrotonate led to tertiary amine 4. Subsequent double
Michael addition with methylamine yielded the corresponding
piperazine in a 2:1 cis/trans mixture. After cleavage of the benzyl
group, the acetate salt of the cis-isomer 5 crystallized selectively.
A subsequent Dieckmann cyclization and protection of the
secondary amine with a Boc group led to the racemic 3,9-
diazabicyclononanone. Resolution by crystallization with (+)-
tartaric acid afforded the pure enantiomer 6 as a key interme-
diate.10
Vinylic triflate 7 (Scheme 2) was prepared from 3,9-
diazabicyclononene 6, using either triflic anhydride or triflimide.
A Negishi coupling with 1-bromo-4-[3-(2-methoxybenzoxy)pro-
poxy]benzene 818 led to 3,9-diazabicyclononene 9 in excellent
yield. Compound 9 still contained a methyl group at the
9-position of the diazabicyclononene system. Since such an
N-methyl group on a tertiary amine is generally cleaved under
rather harsh conditions, such as using an excess of a chloro-
formate at reflux in dichloroethane,11 we decided to replace it
by another group as early as possible in the synthesis. A direct
transprotection with 2,2,2-trichloro-tert-butyl chloroformate
yielded the orthogonally protected compound 10.12 Saponifica-
tion of the ethyl ester proceeded at 80 °C to yield compound
11 together with another regioisomer, in which the double bond
had shifted to its deconjugated 7,8-position (not represented in
Scheme 3). These two compounds were not separated, and
tertiary amide 12 was prepared together with its deconjugated
regioisomer. At this stage both compounds were separated
Chemistry
3,9-Diazabicyclononanone derivative 6 (Scheme 1) was
prepared in three steps using a slightly modified literature
procedure.9 A double N-alkylation of benzylamine with ethyl

Renin Inhibitors
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12 3691
Scheme 1a
a Reagents: (a) Na2CO3, EtOH, 40 °C, 65%; (b) (i) MeNH2, EtOH, H2O, room temp; (ii) H2, Pd/C, EtOH, AcOH, then tert-butyl methyl ether, 47%; (c)
(i) NaH, THF, 45 °C; (ii) Boc2O, Et3N; (iii) (+)-tartaric acid, acetone, 24%.
Scheme 2a
a Reagents: (a) 8, n-BuLi, ZnCl2, Pd(PPh3)4, THF, -78 °C to reflux, 65%; (b) Cl3CC(CH3)2OCOCl, ClCH2CH2Cl, reflux, 46%; (c) NaOH, H2O, EtOH,
80 °C; (d) 2-chlorobenzyl-N-methylamine, EDC ·HCl, HOBt, DMAP, DIPEA, CH2Cl2, room temp, 29% over two steps; (e) HCl, dioxane, CH2Cl2, 0 °C to
room temp, 45%; (f) (i) electrophile, base, CH2Cl2, room temp; (ii) Zn, AcOH, THF, room temp, 41-72% over two steps.
Scheme 3a
a Reagents: (a) (i) HCl, dioxane, CH2Cl2, room temp; (ii) AcCl, DIPEA, CH2Cl2, -78 °C, 68%; (b) NaOH, H2O, EtOH, 80 °C; (c) (i) amine, EDC ·HCl,
HOBt, DMAP, DIPEA, CH2Cl2, room temp; (ii) Zn, AcOH, THF, room temp, 8-46% over three steps.
chromatographically. Selective cleavage of the Boc group
yielded compound 13 as a first scaffold ready for parallel
chemistry. Alkylation or acylation of compound 13 and final
deprotection led to the desired compounds 14a-f.

3692 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12
Bezenc¸on et al.
Scheme 4a
a Reagents: (a) n-BuLi, ZnCl2, Pd(PPh3)4, THF, -78 °C to reflux, 63-85%; (b) Cl3CC(CH3)2OCOCl, CH2ClCH2Cl, reflux, 69-75%; (c) (i) NaOH, H2O,
EtOH, 80 °C, 95%; (ii) TBDMS-Cl, imidazole, DMF, room temp; (iii) K2CO3, THF, MeOH, H2O, room temp; (d) 2,3-dichlorobenzylcyclopropylamine,
EDC ·HCl, HOBt, DMAP, DIPEA, CH2Cl2, room temp, 35-38% over four steps; (e) (i) HCl, dioxane, CH2Cl2; (ii) AcCl, DIPEA, CH2Cl2, -78 °C, 71-84%;
(f) (i) ArOH, 1,1′-(azodicarbonyl)dipiperidine, PBu3, toluene, 80 °C to reflux, 90 min; (ii) Zn, AcOH, THF, room temp, 11-39%.
Table 1. Affinity of the First Diazabicyclononene Derivatives toward
Renin
The Boc group of compound 10 was cleaved as well, and
subsequent acetylation yielded acetamide 15 (Scheme 3). Saponi-
compd
R
IC50 in buffera
IC50 in plasmaa
fication of the ethyl ester led to carboxylic acid 16 that was directly
used for amide couplings. Again, the amide products were obtained
together with their corresponding, deconjugated regioisomers, and
the two compounds were separated chromatographically before
final deprotection. Subsequent removal of the trichloro-tert-
butylcarbamate yielded compounds 17a-l.
14a
14b
14c
14d
14e
14f
4-ClPhCH2CO-
PhCH2SO2-
2-thiophenyl-CH2CO-
CH3CO-
1.0
4.8
1.6
3.2
5.2
6.8
130
170
278
86
86
129
Me-
H-
a IC50 values determined in aqueous buffer or human plasma. Values in
nM.
Finally modifications at the 7-position of the diazabicyclononene
template were undertaken as well. The synthesis started by a
Negishi coupling between vinylic triflate 7 and the corresponding
bromophenyl derivatives 18a and 18b, respectively (Scheme 4,
compounds 19a and 19b). A transprotection with 2,2,2-tert-butyl
chloroformate yielded the corresponding 3,9-diazabicyclonene
derivatives 20a and 20b. Saponification of these ethyl esters and
reintroduction of the hydrolyzed silyl protecting groups afforded
the corresponding carboxylic acids 21a and 21b, again together
with their corresponding regioisomers presenting a shifted double
bond at the 7,8-position. In these cases, subsequent amide couplings
from these regioisomeric mixtures yielded the desired amide
products 22a and 22b, respectively, with the double bond moving
back to the desired position in a ratio ranging from 9:1 to 19:1,
provided 1 equiv of HOBt and a secondary amine bearing an
N-cyclopropyl substituent were engaged.13 Acidic treatment of
amides 22a and 22b cleaved both the Boc and silyl protecting
groups; subsequent acetylations yielded alcohols 23a and 23b.
Construction of libraries via Mitsunobu couplings and final
deprotections yielded compounds 24a-k and 25a-l, respectively.
pounds 14a-f, yielding very promising IC50 values for renin
inhibition (Table 1). The new diazabicyclononene template, with
a tertiary amide substituent replacing the 1-(2-acetylamino-
ethyl)-1,2,3,4-tetrahydroquinolin-7-ylmethoxy substituent present
at the corresponding position in compound 3, was validated.
Compounds 14a-f are highly potent renin inhibitors displaying
single digit nanomolar IC50 values in buffer. Variations at the
N3-position of the 3,9-diazabicyclononene template did not
result in a distinct structure-activity relationship (SAR). In
agreement with our model predicting N3-substituents to reside
at the interface of enzyme and solvent, a phenylacetyl (14a), a
thiophenylacetyl (14c), or the much smaller acetyl derivative
(14d) had similar inhibitory power. Even a sulfonyl group was
tolerated equally well (14b), while very small substituents like
a methyl group (14e) or hydrogen (compound 14f) led to a
marginal loss of affinity in buffer.
IC50 values were also determined in human plasma. Whether
renin should be inhibited in blood, in tissue, or in both is still
a matter of debate. Nevertheless, measuring the inhibitory
affinity of our compounds in human plasma proved to be an
excellent means to take into account the plasma protein binding
and consequently the free fraction available in vivo for renin
Proof of Concept
A small library generated by varying substituents at the N3-
position of the diazabicyclononene template delivered com-

Renin Inhibitors
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12 3693
Table 2. Optimization of the Amide Substituent at Position 6
perpendicular to the C6-C7 double bond. This is due to
repulsions between the R- and Ar-substituents of compounds
17 on one hand, and the hydrogen atom attached at position 5
and the phenyl substituent attached at position 7 of the bicyclic
system on the other hand. (c) The newly introduced R-
substituents in the tertiary series point into a small subpocket
of renin with a cyclopropyl ring (17f vs 14f, Table 1), yielding
best results. Interestingly the linker length between the terminal
phenyl and amide unit did not discriminate IC50 values in the
buffer assay (17c, 17d, and 17e) but the shortest (methylene)
linker led to the lowest IC50 value in plasma. We therefore
concentrated on easily available benzylamides (n ) 1). Opti-
mization of the terminal aryl group was lengthier. After
extensive use of parallel chemistry, we found that small and
hydrophobic ortho- and meta-substituents generally tended to
improve affinity (e.g., 17f, 17g, vs 17h) with optimal results
obtained for the 2,3-dimethyl- (17i) and 2,3-dichloro-substituted
compounds (17j), respectively. Interestingly, compound 17f,
with a 2-chloro substituent, displays a much lower IC50 value
in plasma than compounds 17g and 17h, with a 3- and 4-chloro
substituent, respectively. We do not have any explanation for
this increased potency in plasma. The 2-methyl-3-methoxy
pattern led to a slight loss in activity (17k). Introduction of a
more polar, terminal 4-pyridinyl was well tolerated (17l) and
had a slightly beneficial effect on affinity in plasma.
IC50 in IC50 in
bufferb plasmab
compda
n
R
Ar
17a
17b
17c
17d
17e
17f
17g
17h
17i
1
1
1
2
3
1
1
1
1
1
1
1
H
H
Me
Me
Me
phenyl
2-Cl-phe
phenyl
phenyl
phenyl
458
116
13
8.3
13
0.77
1.5
5.2
0.25
0.20
0.62
1.4
7500
3600
122
167
640
25
242
456
8.8
9.1
29
18
cyclopropyl 2-Cl-phe
cyclopropyl 3-Cl-phe
cyclopropyl 4-Cl-phe
cyclopropyl 2,3-di-Me-phe
cyclopropyl 2,3-di-Cl-phe
17j
17k
17l
cyclopropyl 2-Me-3-MeO-phe
cyclopropyl 2-Me-3-MeO-pyridin-4-yl
a For the preparation of noncommercially available amine, see Supporting
Information. b IC50 values determined in aqueous buffer or human plasma.
Values in nM.
inhibition. Except for compounds (+)-26g and (+)-26h (vide
infra), which were more than 99% protein bound in human,
rat, dog, and monkey plasma, plasma protein binding was not
directly measured. Since the free fraction of all compounds was
presumably very low, a precise, relevant measurement would
have been very delicate. As a matter of fact, we observed an
excellent correlation between IC50 values in plasma and the
pharmacological effects for given inhibitor concentrations in
blood, and we assumed that the ratio of the IC50 values measured
in buffer and in plasma reflected the free fraction of our
compounds in plasma. IC50 values determined in plasma for
compounds 14a-f are shifted upward by about a 100-fold over
the values determined in buffer for compounds with hydrophobic
N3 substituents (compounds 14a and 14c) and by about a 25-
fold for a more polar sulfonamide (compound 14b). Compounds
14e-f with smaller groups at the N3-position displayed shifts
in a range between 15 and 25, making them more potent in
plasma than compounds 14a-c. There seems to exist a certain
correlation between the available free fraction, size and polarity
of the inhibitors: for compounds 14a-c, clogP varies from 5.5
to 6.3, while clogP varies between 4.1 and 4.4 for compounds
14d-f. Similarly, the molecular weight varies from 699 to 729
for compounds 14a-c and from 575 to 617 for compounds
14d-f.
Replacement of the Lengthy Linker and Further
Optimization at the 7-Position
After successfully optimizing position 6 of our template, we
refocused our efforts on position 7. Unfortunately, until this
time all compounds failed to display sufficient metabolic
stability. Compound 17j was rapidly metabolized in human and
rat microsomal suspensions. The most labile part was postulated
to be the terminal methoxybenzyl unit of the substituent in
position 7. We strived to (a) replace this benzylic ether group
and (b) to shorten the lengthy linker between the two aromatic
rings. As suggested by modeling, we focused on two four-atom
tethers, one being a propyl-3-oxy linker (24 series) and the
second a 1,2-ethylenedioxy linker (25 series). The former was
expected to be a bit longer, since the second is known to prefer
a compact, gauche arrangement. A series of differently substi-
tuted terminal phenyl rings was modeled and eventually
introduced.
Optimization of the 6-Position
After validation of the central 3,9-diazabicyclononene as a
useful template, improvement of the inhibitor potency by
optimizing its substituents was undertaken. For the sake of
minimal inhibitor size an acetyl at the N3-position of the bicyclic
system was used in the following libraries. New substituents in
position 6 were introduced by converting acid 16 to a library
of secondary and tertiary amides, respectively (Table 2). While
the secondary amide with an unsubstituted benzyl group (17a)
was only weakly active, the 2-chloro derivative 17b displayed
a slightly improved affinity in buffer. However, converting
compound 17a to the tertiary methylamide 17c improved this
affinity by a factor of more than 30. The superiority of tertiary
amides compared to secondary amides seems to have multiple
reasons. (a) In the group of secondary amides (e.g., 17a) the
amide N-H has to be desolvated in an enthalpically unfavorable
process before binding to renin. Since the inhibitor is unable to
form a new hydrogen bond upon binding, a decreased binding
energy and therefore inhibitory power emerge if compared with
amide 17c. (b) If compared with the secondary amide 17a, the
tertiary amide unit is forced into a different conformation, more
favorable for binding to renin, in which the amide bond is
Since 17j was the most potent compound in our previous
series (17a-l), its 2,3-dichlorobenzylcyclopropylamide moiety
was kept in position 6 for further optimizations. Here again,
the preparation of a large number of compounds led to a rather
complex SAR for the terminal aromatic ring of the 7-substituent,
which was divided into two sections depending on the linker
type between the two aromatic rings (Table 3). The SAR proved
to be a complex mix of steric and electronic effects on the phenyl
ring. The optimal combination of substituents was found after
an extensive use of parallel chemistry only. Both suggested
linkers eventually delivered subnanomolar compounds; however,
they required differently substituted terminal phenyl rings for
optimal results. Indeed, while the propyl-3-oxy linker preferred
to be in an extended conformation, the 1,2-ethylenedioxy unit
is exclusively found in the preferred gauche conformation
(Figure 3a). The conformational preferences of each linker
affected their overall length, and therefore, substitution patterns
at the terminal phenyl rings had to be chosen appropriately. In
both series an unsubstituted terminal phenyl (24a, 25a) turned
out to be only moderately potent. While the introduction of a
chloro substituent in position 2 was beneficial in both series

3694 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12
Bezenc¸on et al.
Table 3. Optimization of the Terminal Phenoxy Moiety at Position 7
7. A smaller substituent in the para-position (25h) only slightly
reduced potency. Shifting the para-substituent to meta-position
and/or changing its size affected affinity only moderately
(25i-k). Interestingly, an additional 3-methyl group added to
our best terminal aromatic was well tolerated (25l). Noteworthy,
compound 25g, which comprises the same 6-substituent as 17j,
displays the same affinity in plasma, clearly demonstrating that
the newly designed, short 1,2-dioxoethylene linker in substituent
7 is at least as good as its longer predecessor in 17j. Compound
25g was so far our most potent renin inhibitor in human plasma.
propyl-3-oxy linker 1,2-ethylenedioxy linker
IC50 in IC50 in
IC50 in IC50 in
Ar
compd buffera plasmaa compd buffera plasmaa
phe
24a
24b
24c 16
24d 38
24e 19
24f
24g
24h
24i
24j
24k
7.7
1.4
699
188
1560
1580
1900
229
60
257
26
50
25a
25b
25c
25d
25e
25f
25g
25h
25i
8.5
4.9
10
7.0
6.6
590
384
1100
694
415
582
7.1
29
54
101
34
40
2-Cl-phe
3-Cl-phe
4-Cl-phe
2-MeO-phe
2-Br-5-F-phe
2,6-diCl-4-Me-phe
2,6-diCl-4-F-phe
2,3,6-triF-phe
3-Cl-2,6-diF-phe
2-Cl-3,6-diF-phe
2,6-diCl-3,4-diMe-pheb
2.4
0.83
1.0
0.48
0.28
0.29
4.6
0.16
0.35
0.79
1.5
0.33
0.30
Further Optimizations
25j
25k
25l
24
While optimizing the 6- and 7-positions of the diazabicy-
clononene, we kept investigating the effect of different substit-
uents at the N3-position. As noticed earlier, the substitution at
this position did not have a profound influence on the affinity
in buffer. The substituted compounds (rac)-26a to (rac)-26c14
displayed very similar values as compounds (+)-26g (ACT-
077825, MK-8141), and (+)- and (-)-26h (Table 4). Interest-
ingly, when both enantiomers of compounds 26h were separated,
we realized that they were almost equally potent toward renin
inhibition. This proved to be generally the case for all diazabi-
cyclononene derivatives that were not substituted at the N3-
position (vide infra for a structural explanation). The upward
shifts of the affinity in plasma tended to be elevated and not
easily predictable for these N3-substituted compounds. Another
common property of the acylated compounds at the N3-position
(compounds (rac)-26a to (rac)-26c) was their poor bioavail-
ability in Wistar rats (at 10 mg/kg), with very variable half-life
times. Removing the polar amide substituent at the N3-position
led to compounds (+)-26g and (+)-26h that were clearly more
bioavailable. The replacement of the central phenyl ring by a
thiazolyl (compound (+)-26d) represents another possibility to
introduce some polarity in this type of lipophilic compounds.
This led to some success, both in vitro and in vivo, but this
compound finally did not display as promising properties as
(+)-26g or (+)-26h regarding its potency in vivo. Furthermore,
compound (+)-26d showed a significant time dependent inhibi-
tion for the CYP 3A4 (shift value of 25 after 30 min of
preincubation). Introduction of polarity in the linker, with tertiary
amines (e.g., compounds (rac)-26e and (rac)-26f), led to a
significant loss of affinity for renin while only gaining little
bioavailability. In general, permeation was most probably the
limiting factor, since T1/2 did not correlate with the bioavailability
for compounds 26a to (+)-26h. All these compounds are
lipophilic, with clogP values varying between 6.6 (26h) and
4.8 (26c). Experimentally, the log D value of compound (+)-
26g was higher than 4.9, and the log D value of compound (+)-
26h was higher than 4.6 (pH 7.4). Enhancing the molecular
weight (MW ) 750 for 26a vs 619 for 26g or 645 for 26h) and
the number of hydrogen bond acceptors (eight acceptors for
26a,b and seven acceptors for 26c,d vs six for 26e-g and five
for 26h) proved to be extremely detrimental for the bioavailability.
a IC50 values determined in aqueous buffer or human plasma. Values in
nM. b For preparation of the corresponding phenol, see Supporting Informa-
tion.
Figure 3. X-ray structure analysis: (a) linker and terminal aromatic
ring of 7-substituents in (+)-26h (green) and (+)-26g (yellow); (b)
view of 7-substituent of (+)-26g (blue) with 1,2-oxoethylene linker in
preferred gauche conformation and exit vector of linker forced out of
plane by two bulky chlorines in position 2,6 of terminal phenyl ring.
(24b, 25b), shifting this halogen to the meta-position (24c) or
especially the para-position (24d) turned out to be quite
unfavorable in the series with the extended propyl-3-oxy linker.
This negative trend was less pronounced in the second series
(compare 24b-d vs 25b-d). With a very long and flexible
linker as found in 3, a terminal 2-methoxybenzyl substituent
yielded a very potent inhibitor.6 However, in our propyl-3-oxy
linker series, a terminal 2-methoxyphenyl (24e) was even less
active than the unsubstituted phenyl ring. All compounds
comprising a terminal 2,6-disubstituted phenyl turned out to be
superior to compounds lacking this substitution pattern (compare
compounds a-f vs g-l in 24 and 25 series). Two reasons might
be responsible for this required substitution pattern. First, the
optimal hydrophobic 2-chloro substituent fits snuggly into a
hydrophobic pocket while the 6-substituent fills a gap between
the terminal aromatic rings of template substituents 6 and 7.
Second, the two phenyl substituents force the O-C vector of
the linker out of the plane defined by the phenyl ring. This
conformationally strained arrangement is enforced by this 2,6-
substitution pattern and turned out to be favorable for binding
into the active site (Figure 3b). Inhibitors with an additional
4-phenyl substituent were slightly less potent in the longer
propyl-3-oxy linker series (24 series) probably because of steric
repulsions (compare 24g and 24h with 25g, and 25h). The SAR
for compounds comprising the shorter 1,2-ethylenedioxy linker
was slightly different. Inhibitors comprising an unsubstituted
phenyl (25a) or a monosubstituted phenyl (25b-e) displayed
moderate affinities only. However, a substituent at the 4-position
(25d) was well accepted here. An extensive study with polysub-
stituted phenoxy derivatives revealed the 2,6-dichloro-4-meth-
ylphenyl (25g) to be the optimal terminal aromatic in position
Fine optimization of the IC50 values in plasma by modulating
the substituents at positions 6 and 7 with the finding described
above led to the identification of two optimized compounds,
one with a 1,2-ethylenedioxy linker at position 7 of the 3,9-
diazabicyclononene (compound (+)-26g), and compound (+)-
26h including the extended propyl-3-oxy linker at its 7-sub-
stituent. The substituents in position 6 were marginally different
in compounds 26g and 26h, respectively. Interestingly, (+)-
26g demonstrated decreased metabolic activation potential and

Renin Inhibitors
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12 3695
Table 4. Further Optimization at the N3-Position and Further Variations
b
c
compd
Ar
2-Cl-phe
2-Cl-phe
2-Cl-phe
Ph
R
IC50 in buffera
IC50 in plasmaa
T1/2
Tmax
F d
(rac)-26a
(rac)-26b
(rac)-26c
(+)-26d
(rac)-26e
(rac)-26f
(+)-26g
(+)-26h
(-)-26h
2-Br-5-F-phe
2,3,6-triF-phe
2,3,6-triF-phe
CO(CH2)3CONH2
CO(CH2)2CONH2
COCH2OH
0.55
0.55
1.8
0.67
2.9
56
56
356
23
41
120
19
11
4.5
4
3
4
15
12
17
24
33
2.3
6.0
0.3
0.5
3
0.5
5.2
3.6
7.5
6.3
5.5
2-Me-3-MeO-phe
2-Me-3-MeO-phe
2,3-diMe-phe
2,3-diCl-phe
2-Cl-3,6-diF-phe
2,3,6-triCl-phe
2,6-diCl-4-Me-phe
2-Cl-3,6-diF-phe
2-Cl-3,6-diF-phe
2.3
4
10
10
H
H
H
0.20
0.40
0.80
36
88
2,3-diCl-phe
a IC50 values determined in aqueous buffer or human plasma. Values in nM. b In h. c In h. d In %.
Figure 5. (a) Superposition of X-ray structures of human recombinant
renin complexed with (+)-26h (yellow) and (-)-26h (magenta)
respectively. (b) Side view of the bicyclic templates of (+)-26h (yellow,
protein in blue) with (-)-26h superimposed (magenta, protein green).
Figure 4. Mean arterial blood pressure (MAP) recordings before
(control) and after a single oral administration of 10 mg/kg compound
(+)-26g to double transgenic rats. Data are expressed as mean ( SEM.
on blood pressure was observed at about 11 h and Tmax in Wistar
rats was 10 h.
Structural Studies by X-ray Crystallography
less in vitro covalent binding to rat and human liver microsomal
During our medicinal chemistry campaign we prepared
compound (+)-26h and its enantiomer (-)-26h. Much to our
surprise, these two enantiomers were almost equally potent as
renin inhibitors in buffer and plasma, respectively, a performance
that is rarely seen for compounds of this size and structural
complexity. X-ray structures of both compounds ((+)- and (-)-
26h) complexed to human recombinant renin provided a
stupefying simple explanation for this quite unexpected result.
A superposition of these two structures revealed the two large
substituents in positions 6 and 7 of our bicyclic template
superimposed almost perfectly (Figure 5a). However, the central
bicyclic template that is responsible for hydrogen bond contact
with the two catalytically important aspartates was oriented quite
differently. In compound (+)-26h nitrogen N9 that is located
in the short bridge of the template is perfectly situated between
the two aspartates, thereby expelling the catalytically important
water and forming two strong H-bonds, one with each of the
neighboring aspartates (Figure 5b). In (-)-26h the whole
template is rotated 180° and shifted away from the aspartates
proteins than (+)-26h.15
Pharmacodynamics
Oral activity was assessed in female double transgenic rats
implanted with a telemetry blood pressure transmitter as
previously described.16 These animals are expressing both
human renin and angiotensinogen genes. If left untreated, they
develop hypertension and severe albuminuria with serious
vascular lesions in the kidney, resulting in ∼50% mortality at
7-9 weeks of age.17 The blood pressure recordings before and
after oral administration of compound (+)-26g are shown in
Figure 4. Compound (+)-26g at 10 mg/kg reduced the blood
pressure by about 30 mmHg, and the duration of action was
close to 36 h. This compound displayed a good pharmacody-
namic-pharmacokinetic relationship, since the maximal effect

3696 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12
Bezenc¸on et al.
mol) was added again, and the mixture was stirred at 40 °C for
4 h. The conversion was not complete yet, and ethyl bromocrotonate
(150 mL, 0.82 mol) was added again. The mixture was stirred for
16 h at 40 °C. The conversion was complete. The solvents were
removed to one-half under reduced pressure, and toluene (58 L)
was added. The mixture was washed with water (2 × 28 L). The
org layer was extracted with aq 1 M HCl (3 × 36 L). The combined
aq extracts were covered with EtOAc (60 L), and the pH was
adjusted to 12 by addition of aq 30% NaOH (40 L). The layers
were separated, and the aq phase was extracted back with EtOAc
(31 L). The combined org extracts were washed with water and
evaporated under reduced pressure. EtOH (36 L) was added, and
the solvents were removed under reduced pressure. The residue
was dried under high vacuum to yield a crude mixture containing
the title compound (5.9 kg, 65%). This mixture was use without
further without purification. 1H NMR (CDCl3) 7.31-7.26 (m, 5H),
6.97 (ddd, J1 ) 15.7 Hz, J2 ) J3 ) 5.8 Hz, 2H), 6.05 (d, J ) 15.7
Hz, 2H), 4.20 (q, J ) 7.2 Hz, 4H), 3.63 (s, 2H), 3.24 (dd, J1 ) 5.8
such that N3 of the three atom bridge is now perfectly located
between the two aspartates also establishing hydrogen bonds
with each of the aspartates. Obviously, the hydrophobic interac-
tions dominate the positioning of these inhibitors with the
pseudosymmetry of the central template being able to establish
the necessary hydrogen bonds in either orientation. Substitution
of either of the two nitrogen atoms abolishes this pseudosym-
metry and completely abrogates this observed effect.
Conclusions
A series of renin inhibitors was discovered by de novo design
and lead optimization. The most preferred compound of this
series, compound (+)-26g, proved to be highly potent in renin
inhibition, well absorbed in rats, and efficacious at 10 mg/kg
in a double transgenic rat model.
Experimental Section
Hz, J2 ) 1.3 Hz, 4H), 1.31 (t, J ) 7.2 Hz, 6H). LC-MS: tR
)
0.79 min. MS (ESI): m/z 332.38 (M + 1)+.
All solvents and reagents were purchased from commercial
sources (Fluka, Aldrich, Lancaster, Acros, Riedel-de-Haen). Pal-
ladium catalysts were purchased from Strem Chemicals. Solvents
were used undistilled. Dry solvents were purchased on molecular
sieves and used directly. All reactions were run under a nitrogen
atmosphere. TLC was run on glass plates from Merck coated with
silica gel 60F254. Flash chromatography (FC) was run either in a
glass column filled with silica gel 60 from Fluka under a slight
positive pressure or using a Flash Master Personal from Argonaut,
equipped with IsoluteSPE columns Flash Si II and with a Gilson
FC204 fraction collector. Analytical LC-MS was run on one of
the following columns: (a) Waters XBridge C18 column, 5 µm,
4.6 mm × 50 mm; (b) Zorbax Extend SB-AQ column, 5 µm, 4.6
mm × 50 mm. Gradient was from CH3CN/H2O (5:95) to 100%
CH3CN over 1.5 min. Flow was 4.5 mL/min. For the determination
of the enantiomeric ratio, a ChiraCel OD column, 10 µm, 4.6 mm
× 50 mm, was used under isocratic conditions (2% EtOH with
0.1% Et3N and 98% hexane, flow rate of 0.8 mL/min, over 12 min).
The LC-MS machines consisted of a Gilson Liquid Handler Micro
215, of Hewlett-Packard Agilent 1100 series pumps, of a Dionex
P580 makeup pump, of a Dionex MS detector, of a Dionex UVD
340U UV detector, and of a Sedex 85 LT-ELSD detector. Chemical
shifts are in ppm, using Me4Si (0 ppm) or CHCl3 (7.27 ppm) as
cis-(6-Ethoxycarbonylmethyl-1-methylpiperazin-2-yl)acetic Acid
Ethyl Ester (5). A solution of compound 4 (11.3 kg, 34.1 mol) in
EtOH (70 L) was charged to a 100 L reactor. MeNH2 (30% in
EtOH, 5.2 L, 37.5 mol) was added at room temperature, and the
mixture was stirred at room temperature overnight. Since the
conversion was not complete, MeNH2 (30% in EtOH, 250 mL, 1.84
mol) was added again, and the mixture was stirred overnight at
room temperature. Since the conversion was not complete, MeNH2
(30% in EtOH, 260 mL, 1.91 mol) was added again, and the mixture
was stirred overnight at room temperature. AcOH (1.1 L) was
added, and the pH was adjusted to 5-6. Pd/C (10% on charcoal,
628 g) was added, and the mixture was hydrogenated at 1230 mbar
of H2 at room temperature for 3 days. The mixture was filtered
through Celite, and the filter cake was washed with EtOH. The
filtrate was charged to a clean reactor and concentrated under
reduced pressure. Toluene (34 L) was added, and the solvents were
partially (11 L) removed under reduced pressure. The resulting
solution was heated to 40 °C, and TBME (52 L) was added. The
mixture was stirred for 20 min at 40 °C, was cooled within 3 h to
1 °C, and was stirred at this temperature overnight. The resulting
suspension was filtered off, washed with cold TBME (2 × 15 L),
and dried under a stream of N2 for 4 days to yield the acetate salt
of the title compound as a colorless solid (4.37 kg). The mother
liquor (93 L) was charged to the reactor and was concentrated under
reduced pressure. After removal of 70 L of solvents, the resulting
solution was heated to 40 °C. TBME (40 L) was added, and the
mixture was stirred at 40 °C for 20 min. The mixture was cooled
to 1 °C over 2 h and stirred at this temperature overnight. The
suspension was filtered, the filter cake was washed with TBME,
and the colorless residue was dried overnight under a stream of N2
to yield the acetate salt of the title compound as a colorless solid
(730 g). The mother liquor (91 L) was charged to the reactor and
was concentrated under reduced pressure. After removal of 66 L
of solvents, the resulting solution was heated to 40 °C. The mixture
was cooled to 1 °C over 2 h and stirred at this temperature for 3
days. The mixture was cooled to -10 °C for 2 h. The suspension
was filtered, the filter cake was washed with TBME, and the
colorless residue was dried for 4 h under a stream of N2 to yield
the acetate salt of the title compound as a colorless solid (251 g).
The three crystallization crops were pulled together to yield the
title compound as its acetate salt (5.35 kg, 47%). 1H NMR (CD3OD)
4.16 (q, J ) 7.1 Hz, 4H), 3.16 (d, J ) 10.9 Hz,2H), 2.97 (m, 2H),
2.90 (d, J ) 11.4 Hz, 2H), 2.65 (dd, J1 ) 15.8 Hz, J2 ) 4.2 Hz,
2H), 2.56 (dd, J1 ) 15.8 Hz, J2 ) 6.2 Hz, 2H), 2.31 (s, 3H), 1.95
(s, 3H), 1.28 (t, J ) 7.1 Hz, 6H). LC-MS: tR ) 0.57 min. MS
(ESI): m/z 273.41 (M + 1)+.
1
reference. The attribution of the signals of the H and 13C NMR
spectra was based on COSY, HSQC, and HMBC experiments; as
much as is possible, an assignment of the NMR signals is described
(see Supporting Information; aromatic signals are not attributed and
are characterized with the subscript arom, and Cq stands for a
nonattributed quaternally carbon atom). The NMR spectra of
compounds 8-26 are complex; they consist of at least two rotamers
and sometimes display broad signals. The minor rotamer is
characterized with a superscripted R. Because of the complexity
of the spectra, the integral could not always be determined
unambiguously. Unless specified otherwise, the NMR spectra were
recorded on a Bruker 400 MHz machine. Spectra measured at 300
MHz were recorded on a Varian machine. IR spectra were recorded
on a Perkin-Elmer Spectrum One apparatus, with samples dissolved
in CHCl3 and prepared as films. Elemental analysis and Karl Fischer
titrations were run at Solvias AG. Optical rotations were measured
on a Jasco P-1030 apparatus. Purity of the final compounds was
assessed by combustion analysis and/or LC-MS. When assessed
by LC-MS, the purity was determined according to the UV trace
at 230 and 254 nm.
4-[Benzyl(3-ethoxycarbonylallyl)amino]but-2-enoic Acid Ethyl
Ester (4). A 100 L enamelled reactor was charged with EtOH (55
L). Ethyl 4-bromocrotonate (85%, 12.5 kg, 54.8 mol) was added,
and the feeding tank was rinsed with EtOH (8 L). DIPEA (15 L,
86 mol) was added at room temperature, and the mixture was heated
to 40 °C. Benzylamine (2.95 kg, 27.4 mol) was added over 25 min.
The inner temperature increased from 38 to 55 °C. After complete
addition, the mixture was stirred at 40 °C overnight. Since the
conversion was incomplete, ethyl 4-bromocrotonate (420 mL, 2.29
(1R,5S)-9-Methyl-7-oxo-3,9-diazabicyclo[3.3.1]nonane-3,6-dicar-
boxylic Acid 3-tert-Butyl Ester 6-Ethyl Ester (6). Toluene (77 L)
was charged to a 100 L reactor, and t-BuOK (6.48 kg, 57.7 mol)
was added at room temperature. The suspension was heated to 40
°C, and the acetate salt of compound 5 (5.35 kg, 16.1 mol) was

Renin Inhibitors
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12 3697
added portionwise. The mixture was stirred at 40 °C for 16 h and
was cooled to 0-5 °C. AcOH (8.0 L) was added, keeping the inner
temperature below 8 °C. The resulting orange-brown suspension
was concentrated under reduced pressure until a thick suspension
was obtained (60 L of toluene removed). EtOAc (26 L) was added,
and the mixture was cooled to 0-5 °C. Et3N (14 L, 100 mol) was
added until a pH of 8-9 was reached. A solution of Boc2O (3.81
kg, 17.4 mol) in EtOAc (4.2 L) was added over 25 min. The reaction
mixture was stirred for 2 h at 0-5 °C and allowed to warm to
room temperature over 2 h. The mixture was stirred at room
temperature overnight. The mixture was washed with aq 10%
K2CO3 (16 L). Since the layer separation was sluggish, water (5
L) and EtOAc (10 L) were added. The layers separated overnight.
The org layer was washed with water (2 × 16 L). The combined
aq layers were extracted back with EtOAc (3 × 16 L). The
combined org layers were concentrated under reduced pressure (83
L of org solvents removed). The remaining org mixture was diluted
with toluene (40 L), and the resulting mixture was extracted with
aq 2.5 M NaOH (3 × 20 L and 4 × 10 L). The combined aq extracts
were acidified with aq 6 M HCl (18 L) to pH 6-7 and were
extracted back with methylcyclohexane (4 × 40 L). The combined
org extracts were filtered, and the solvents were removed under
reduced pressure. The residue was dried under high vacuum to yield
the crude racemic title compound (rac)-6 (4.13 kg, 80%) as a
slightly yellow oil.
Anal. Calcd for C16H26N2O5: C, 58.88; H, 8.03; N, 8.58. Found: C,
58.59; H, 7.87; N, 8.57.
(1R,5S)-9-Methyl-7-trifluoromethanesulfonyloxy-3,9-
diazabicyclo[3.3.1]non-6-ene-3,6-dicarboxylic Acid 3-tert-Butyl Es-
ter 6-Ethyl Ester (7). A solution of compound 6 (50 g, 153 mmol)
in dry THF (900 mL) under N2 was cooled to 0 °C. NaH (55% in
oil, 8.35 g, 191 mmol) was added slowly (gas evolution was seen)
while keeping the temperature under 10 °C. The mixture was stirred
for 1 h, and Tf2NPh (65.7 g, 183 mmol) was added. The reaction
mixture was stirred overnight while warming to room temperature.
The reaction mixture was poured onto ice/distilled water mixture
(1.5 L), and the solvents were partially removed under reduced
pressure. The aq residue was extracted with EtOAc (3 × 350 mL).
The combined org layers were washed with water (375 mL) and
brine (350 mL). The org layer was dried over MgSO4, filtered, and
the solvents were removed under reduced pressure to yield a yellow
oil. Purification of the crude by FC (EtOAc/heptane 5:95 to 3:7)
yielded the title compound (61.3 g, 87%) as a yellow oil. Rf )
0.20 (EtOAc/heptane 1:4). 1H NMR (CDCl3) 4.33 (q, J ) 7.0 Hz,
1H), 4.16 (d, J ) 12.6 Hz, 1H), 4.09 (d, J ) 12.3 Hz, 1HR), 4.00
(d, J ) 13.2 Hz, 1H), 3.83 (s, 1HR), 3.77 (s, 1H), 3.19 (d, J ) 12.7
Hz, 1H), 3.04 (broad, d, J ≈ 9.9 Hz, 2H), 2.75 (dd, J ) 18.8, 6.4
Hz, 1H), 2.436 (s, 3H), 2.24 (d, J ) 19.3 Hz, 1H), 1.40 (broad, s,
9H), 1.37 (t, J ) 7.1 Hz, 3H). IR (film) ν 2977, 2934, 1694 cm-1
.
Anal. Calcd for C17H25F3N2O7S: C, 44.54; H, 5.50; F, 12.43; N,
6.11; S, 6.99. Found: C, 45.02; H, 5.42; F, 12.42; N, 6.04; S, 7.07.
[R]2D5 +23.0 (1.0, MeOH). MS (ESI): tR ) 0.87, m/z 561.69 (M +
1)+.
Compound (rac)-6 (100 g, 306 mmol) was dissolved in acetone
(500 mL). Separately, (+)-tartaric acid (18.4 g, 122 mmol) was
dissolved in acetone (1.0 L). The tartaric acid solution was added
dropwise to the bicyclononane solution under stirring. A few crystals
of (1S,5R)-9-methyl-7-oxo-3,9-diazabicyclo[3.3.1]nonane-3,6-di-
carboxylic acid 3-tert-butyl ester 6-ethyl ester (+)-tartaric acid salt
were added, and the suspension was continuously stirred for 18 h
at room temperature. The suspension was filtered, and the precipitate
was washed with acetone (100 mL). The precipitate was dried under
high vacuum to yield (1S,5R)-9-methyl-7-oxo-3,9-diazabicyclo[3.3.1]-
nonane-3,6-dicarboxylic acid 3-tert-butyl ester 6-ethyl ester (+)-
tartaric acid salt (40 g). The mother liquor was concentrated under
reduced pressure, and the oily residue was diluted with methyl-
tert-butyl ether (300 mL) and water (300 mL). NaHCO3 (11 g)
was slowly added under stirring until a pH value of 8 was reached.
The layers were separated, and the aq layer was extracted with
methyl tert-butyl ether (100 mL). The combined org extracts were
washed with water (200 mL), and the solvents were removed under
reduced pressure. The residue was dissolved in acetone (500 mL),
and a solution of (-)-tartaric acid (21.9 g, 145 mmol) in acetone
(500 mL) was added dropwise under stirring. A few crystals of
(1R,5S)-9-methyl-7-oxo-3,9-diazabicyclo[3.3.1]nonane-3,6-dicar-
boxylic acid 3-tert-butyl ester 6-ethyl ester (-)-tartaric acid salt
were added, and the suspension was continuously stirred for 18 h
at room temperature. The suspension was filtered, and the precipitate
was washed with acetone (100 mL). The precipitate was dried under
high vacuum to yield (1R,5S)-9-methyl-7-oxo-3,9-diazabicyclo[3.3.1]-
nonane-3,6-dicarboxylic acid 3-tert-butyl ester 6-ethyl ester (-)-
tartaric acid salt (55 g). This salt was recrystallized from isopropanol
(145 mL). The obtained crystals were dissolved in a stirred mixture
of water (275 mL) and CH2Cl2 (275 mL). NaHCO3 (27 g) was
added in portions until a pH of 8 was reached. The layers were
separated, and the aq layer was extracted with CH2Cl2 (150 mL).
The combined org extracts were washed with water (275 mL), dried
over Na2SO4, and filtered. The solvents were removed under
reduced pressure to yield the title compound (30 g, 30% for the
resolution step) that crystallized spontaneously. Mp ) 54.8-55.3
°C. Chiral HPLC: tR ) 8.43 min, er ) 99.9:0.01. 1H NMR (CDCl3,
400 MHz) 12.1 and 12.0 (s, broad, 1H), 4.23 (m, 2H), 4.13 and
3.92 (d and m, J ) 12.9 Hz, 1H, rot), 3.92 and 3.80 (m and d, J )
12.7 Hz, 1H, rot), 3.57 and 3.53 (s, 1H, rot), 3.13 and 3.02 (m and
d, J ) 10.9 Hz, 1H, rot), 3.06 and 2.94 (d, and m, J ) 12.8 Hz,
1H, rot), 2.94 (s, 1H), 2.63 (m, 1H), 2.34 (s, 3H), 2.13 and 2.01 (d
and d, J ) 19.2 and 18.8 Hz, 1H, rot), 1.41 and 1.34 (s, 9H, rot),
(1R,5S)-7-{4-[3-(2-Methoxybenzyloxy)propoxy]phenyl}-9-meth-
yl-3,9-diazabicyclo[3.3.1]non-6-ene-3,6-dicarboxylic Acid 3-tert-
Butyl Ester 6-Ethyl Ester (9). A solution of bromide 818 (53.5 g,
152 mmol) in THF (1050 mL) was treated at -78 °C with n-BuLi
(1.6 M in hexane, 130 mL, 208 mmol) under N2. After 30 min of
stirring at -78 °C, a solution of ZnCl2 (1 M in THF, 249 mL, 249
mmol) was added and the mixture was allowed to warm to room
temperature. Triflate 7 (63.4 g, 138 mmol) in THF (40 mL) was
added, followed by Pd(PPh3)4 (4.00 g, 3.00 mmol). The mixture
was heated to 65 °C for 1 h and after cooling to room temperature
was quenched with aq sat. NH4Cl (400 mL). The mixture was
partitioned between EtOAc and water. The org layer was concen-
trated, and the residue was purified by FC (5-50% gradient EtOAc
in heptane) to provide the title compound (52.4 g, 65%). 1H NMR
(CDCl3): 7.37 (d, J ) 7.3 Hz, 1H), 7.28 (d, J ) 6.5 Hz, 1HR), 7.26
(d, J ) 7.8 Hz, 1H), 7.07 (d, J ) 7.8 Hz, 2H), 6.95 (t, J ) 7.4 Hz,
1H), 6.88 (d, J ) 8.2 Hz, 1H), 6.83 (d, J ) 8.0 Hz, 2H), 4.59 (s,
2H), 4.24 (d, J ) 13.0 Hz, 1H), 4.11 (t, J ) 6.0 Hz, 2H), 4.06-3.87
(m, 2H and 2HR), 3.94 (broad, d, J ≈ 13.6 Hz, 1H), 3.83 (s, 3H),
3.77 (s, 1H), 3.72 (t, J ) 5.9 Hz, 2H), 3.19 (d, J ) 13.2 Hz, 1H),
3.12 (d, J ) 12.7 Hz, 1H), 2.98 (broad, d, 1H), 2.57 (dd, J ) 19.6,
6.3 Hz, 1H), 2.47 (s, 3HR), 2.43 (s, 3H), 2.36 (d, J ) 19.8 Hz,
1H), 2.12 (broad, t, J ≈ 6.0 Hz, 2H and 1HR), 1.97 (d, J ) 18.8
Hz, 1H), 1.42 (broad, s, 9HR), 1.38 (broad, s, 9H), 0.94 (t, J ) 6.8
Hz, 2H). IR (film) ν 2930, 1689 cm-1. Anal. Calcd for C33H44N2O7:
C, 68.25; H, 7.64; N, 4.82. Found: C, 66.35; H, 7.51; N, 4.73. [R]D25
+7.0 (1.00, MeOH). LC-MS: tR ) 0.92 min. MS (ESI): m/z 581.51
(M + 1)+.
(1R,5S)-7-{4-[3-(2-Methoxybenzyloxy)propoxy]phenyl}-3,9-
diazabicyclo[3.3.1]non-6-ene-3,6,9-tricarboxylic Acid 3-tert-Butyl
Ester 6-Ethyl Ester 9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester
(10). A suspension of starting material 9 (20.6 g, 35 mmol),
NaHCO3 (29.8 g, 355 mmol), and ꢀ,ꢀ,ꢀ-trichloro-tert-butylchlo-
roformate (17 g, 71 mmol) in 1,2-dichloroethane (360 mL) was
stirred under N2 at 80 °C overnight. After cooling to room
temperature, the mixture was filtered, concentrated, and the residue
was purified by FC (2-50% gradient EtOAc in heptane) to provide
1
the title compound (12.6 g, 46%). H NMR (CDCl3) 7.38 (d, J )
7.3 Hz, 1H), 7.26 (d, J ) 7.8 Hz, 1H), 7.04 (d, J ) 6.5 Hz, 2H),
6.95 (t, J ) 7.4 Hz, 1H), 6.88 (d, J ) 8.2 Hz, 1H), 6.84 (d, J ) 7.9
Hz, 2H), 5.27 (s, 1H), 5.23 (s, 1HR), 5.12 (s, 1HR), 4.58 (s, 2H),
4.42 (broad, s, 1H and 1HR), 4.32 (d, J ) 13.4 Hz, 1H), 4.13-3.92
(m, 3H and 5HR), 4.10 (t, J ) 6.1 Hz, 2H), 3.83 (s, 3H), 3.71 (t,
1.32 (m, 3H). IR (film): ν 2973, 2930, 2146, 1681, 1635 cm-1
.

3698 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12
Bezenc¸on et al.
J ) 5.8 Hz, 2H), 3.14 (d, J ) 13.3 Hz, 1H and 1HR), 3.06 (d, J )
11.5 Hz, 1H and 1HR), 2.81 (dd, J ) 19.6, 6.1 Hz, 1H), 2.53 (d, J
) 19.8 Hz, 1H), 2.11 (t, J ) 5.9 Hz, 2H), 1.99 (s, 3H), 1.95 (s,
3H), 1.41 (broad, s, 9HR), 1.38 (broad, s, 9H), 1.03 (t, J ) 6.7 Hz,
product was used in the next step without purification. A suspension
of the obtained crude, acylated/sulfonated compound (∼0.4 mmol)
and Zn powder (4 mmol) in THF (10 mL) was stirred in the
presence AcOH (0.25 mL) at room temperature for 3 h. The mixture
was filtered through Celite, and the solvents were evaporated under
reduced pressure. The residue was dissolved in CH2Cl2 (10 mL),
and aq 0.1 M EDTA (5 mL) was added. The mixture was stirred
at room temperature for 6 h. The org layer was concentrated under
reduced pressure, and the residue was purified by FC (1-5% MeOH
in CH2Cl2) to provide compounds 14a-d.
2H). IR (film)
ν . Anal. Calcd for
2978, 1695 cm-1
C37H47N2O9Cl3 ·0.5H2O: C, 57.04; H, 6.16; N, 3.59. Found: C,
57.34; H, 6.09; N, 3.68. [R]2D5 -0.3 (0.99;, MeOH). LC-MS: tR )
1.22 min. MS (ESI): m/z 771.41 (M + 1)+.
(1R,5S)-7-{4-[3-(2-Methoxybenzyloxy)propoxy]phenyl}-3,9-
diazabicyclo[3.3.1]non-6-ene-3,6,9-tricarboxylic Acid 3-tert-Butyl
Ester 9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester (11). A mixture
of ethyl ester 10 (5.20 g, 6.70 mmol) in aq 1 M NaOH (74 mL)
and EtOH (176 mL) was stirred at 90 °C for 20 h. The EtOH was
evaporated, and the mixture was diluted with EtOAc (400 mL)
before aq 1 M HCl (30 mL) was added. The org layer was
concentrated to provide the title compound together with its
deconjugated regioisomer (5.10 g, quant), which was used in the
next step without purification. LC-MS: tR ) 1.15 min. MS (ESI):
m/z 743.47 (M + 1)+.
(1R,5S)-6-[(2-Chlorobenzyl)methylcarbamoyl]-7-{4-[3-(2-meth-
oxybenzyloxy)propoxy]phenyl}-3,9-diazabicyclo[3.3.1]non-6-ene-
3,9-dicarboxylic Acid 3-tert-Butyl Ester 9-(2,2,2-Trichloro-1,1-
dimethylethyl) Ester (12). A solution of acid 11 (4.9 g, 6.6 mmol),
HOBt (1.1 g, 8.0 mmol), EDC·HCl (3.2 g, 117 mmol), DIPEA
(3.4 g, 26 mmol), and DMAP (0.2 g, 1.6 mmol) in CH2Cl2 (140
mL) was stirred at room temperature for 30 min. 2-Chloro-N-
methylbenzylamine19 (3.10 g, 19.9 mmol) was added, and the
reaction mixture was stirred at room temperature for 7 days.
Aqueous 1 M HCl (40 mL) was added. The org layer was separated,
concentrated, and the residue was purified by FC (1-4% gradient
MeOH in CH2Cl2) to provide the title compound (1.70 g, 29% over
two steps). 1H NMR (CDCl3) 7.39 (d, J ) 7.2 Hz, 1H), 7.32-6.99
(m, 7H), 6.96 (t, J ) 7.5 Hz, 1H), 6.88 (d, J ) 8.1 Hz, 1H),
6.85-6.72 (m, 2H), 4.78 (d, J ) 15.9 Hz, 1H), 4.75-4.37 (m, 6H
and 5HR), 4.10 (broad, d, 2H), 3.83 (s, 3H), 3.74 (t, J ) 5.6 Hz,
2H), 3.50-2.98 (m, 3H and 3HR), 2.74, 2.70, 2.54, and 2.43 (3H
and 3HR), 2.37-2.21 (m, 1H and 1HR), 2.14 (t, J ) 5.9 Hz, 2H),
1.99 (s, 3H), 1.94 (s, 3H), 1.33 (broad, s, 9HR), 1.29 (broad, s,
9H). IR (film) ν 2925, 1697, 1607 cm-1. Anal. Calcd for
C43H51N3O8Cl4: C, 58.71; H, 5.84; N, 4.78. Found: C, 58.60; H,
5.83; N, 4.98. [R]2D5 +47.6 (1.02, MeOH). LC-MS: tR ) 1.26 min.
MS (ESI): m/z 880.65 (M + 1)+.
(1R,5S)-3-Acetyl-7-{4-[3-(2-methoxybenzyloxy)propoxy]phenyl}-
3,9-diazabicyclo[3.3.1]non-6-ene-6,9-dicarboxylic Acid 6-Ethyl Es-
ter 9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester (15). A solution of
starting material 10 (37.1 g, 48.2 mmol) in CH2Cl2 (360 mL) was
cooled to 0 °C. HCl (4 M in dioxane, 163 mL) was added, and the
mixture was stirred at 0 °C for 20 min and at room temperature
for 3.5 h. The mixture was diluted with CH2Cl2 (150 mL) before
aq 10% Na2CO3 (200 mL) was carefully added. The org layer was
concentrated under reduced pressure. The residue was dissolved
under N2 in THF (750 mL) and cooled to -78 °C. DIPEA (33
mL, 193 mmol), DMAP (0.59 g, 4.8 mmol), and AcCl (3.8 mL,
53 mmol) were added, and the mixture was stirred at -78 °C for
30 min. MeOH (250 mL) was added, the mixture was allowed to
warm up to room temperature, and the solvents were evaporated
under reduced pressure. The mixture was diluted with EtOAc (200
mL) before aq 1 M HCl (200 mL) was added. The org layer was
concentrated under reduced pressure, and the residue was purified
by FC (10-55% EtOAc in heptane) to provide the title compound
1
(23.4 g, 68%). H NMR (CDCl3) 7.37 (d, J ) 7.4 Hz, 1H), 7.26
(d, J ) 7.4 Hz, 1H), 7.02-6.92 (m, 3H), 6.90-6.81 (m, 3H), 5.21
(broad, s, 1HR), 5.16 (s, 1H), 4.86 (d, J ) 13.4 Hz, 1HR), 4.82 (d,
J ) 13.9 Hz, 1HR), 4.76 (d, J ) 13.4 Hz, 2H), 4.58 (s, 2H), 4.51
(d, J ) 7.2 Hz, 1HR), 4.46 (d, J ) 6.5 Hz, 1H), 4.10 (t, J ) 6.3
Hz, 2H), 4.06-3.91 (m, 3H), 3.83 (s, 3H), 3.78 (dd, J ) 13.4, 2.2
Hz, 1HR), 3.71 (t, J ) 6.1 Hz, 2H), 3.49 (dd, J ) 13.4, 2.2 Hz, 1H
and 1HR), 2.98-2.81 (m, 2H and 2HR), 2.42 (dd, J ) 19.4, 2.2
Hz), 2.11 (t, J ) 6.2 Hz, 2H), 2.00, 1.99, 1.96, 1.95 (4s, 4 × 3H),
0.98 (t, J ) 7.1 Hz, 2H). IR (film) ν 2927, 1706, 1651 cm-1. Anal.
Calcd for C34H41N2O8Cl3: C, 57.35; H, 5.80; N, 3.93. Found: C,
57.54; H, 5.87; N, 3.85. [R]2D5 +55.3 (1.01, MeOH). LC-MS: tR
) 1.17 min. MS (ESI): m/z 711.41 (M + 1)+.
(1R,5S)-3-Acetyl-7-{4-[3-(2-methoxybenzyloxy)propoxy]phenyl}-
3,9-diazabicyclo[3.3.1]non-6-ene-6,9-dicarboxylic Acid 9-(2,2,2-
Trichloro-1,1-dimethylethyl) Ester (16). A mixture of ethyl ester
15 (20.1 g, 28.2 mmol) in aq 1 M NaOH (430 mL) and EtOH (600
mL) was stirred at 90 °C for 2 h. The EtOH was evaporated under
reduced pressure, and the mixture was diluted with EtOAc (400
mL) before aq 1 M HCl (200 mL) was added. The org layer was
concentrated to provide the title compound (17.8 g, 93%), which
was used in the next step without purification. LC-MS: tR ) 1.07
min. MS (ESI): m/z 683.15 (M + 1)+.
(1R,5S)-6-[(2-Chlorobenzyl)methylcarbamoyl]-7-{4-[3-(2-meth-
oxybenzyloxy)propoxy]phenyl}-3,9-diazabicyclo[3.3.1]non-6-ene-
9-carboxylic Acid 2,2,2-Trichloro-1,1-dimethylethyl Ester (13). A
solution of starting material 12 (6.6 g, 7.5 mmol) in CH2Cl2 (85
mL) was cooled to 0 °C. HCl (4 M in dioxane, 85 mL) was added,
and the mixture was stirred at 0 °C for 30 min and at room
temperature for 1 h. Aqueous 2.5 M NaOH (100 mL) was carefully
added. The org layer was separated, concentrated, and the residue
was purified by FC (1-4% gradient MeOH in CH2Cl2) to provide
1
the title compound (2.6 g, 45%). H NMR (CDCl3) 7.39 (d, J )
7.2 Hz, 1H), 7.33-6.99 (m, 7H), 6.96 (t, J ) 7.4 Hz, 1H),
6.91-6.75 (m, 3H), 6.63 (d, J ) 7.5 Hz, 1H), 6.55 (d, J ) 7.3 Hz,
1H), 4.76 (d, J ) 15.7 Hz, 1H), 4.68-4.54 (m, 4H and 1HR),
4.43-4.26 (m, 1H and 2HR), 4.42 (d, J ) 15.8 Hz, 1H and 1HR),
4.12 (broad, d, 2H), 3.83 (s, 3H), 3.75 (t, J ) 5.6 Hz, 2H), 3.45,
3.41 (d and d, J ) 12.1 Hz, J ) 11.8 Hz, 2H), 3.28-2.87 (m, 2H
and 3HR), 3.20 (d, J ) 15.9 Hz, 1H), 2.75, 2.71, and 2.59 (3H and
2 × 3HR), 2.25-2.03 (m, 2H and 1HR), 2.10 (t, J ) 5.7 Hz, 2H),
2.00 (s, 3H), 1.94 (s, 3H), 1.93 (s, 3H), 1.86 (s, 3H). IR (film) ν
2908, 1698, 1606 cm-1. Anal. Calcd for C38H43N3O6Cl4: C, 58.55;
H, 5.56; N, 5.39. Found: C, 58.40; H, 5.51; N, 5.40. [R]2D5 +18.6
(1.0, MeOH). LC-MS: tR ) 1.08 min. MS (ESI): m/z 780.11 (M
+ 1).
General Procedure for the Preparation of Compounds 14a-d.
To a solution of compound 13 (0.4 g, 0.51 mmol) and acyl chloride
or sulfonyl chloride (0.56 mmol) in CH2Cl2 (5 mL) was added
DIPEA (0.67 mmol) at 0 °C. The mixture was stirred for 10 min
at 0 °C. The mixture was diluted with CH2Cl2 (5 mL), and aq 1 M
HCl (10 mL) was added. The org layer was concentrated, and the
General Procedure for the Preparation of Compounds 17a-l.
A solution of acid 16 (1.6 g, 2.3 mmol), HOBt (0.45 g, 3.2 mmol),
EDC·HCl (1.2 g, 6.2 mmol), DIPEA (1.3 g, 10 mmol), and DMAP
(80 mg, 0.6 mmol) in CH2Cl2 (25 mL) was stirred at room
temperature for 30 min. The desired amine (7.0 mmol) was added,
and the reaction mixture was stirred at room temperature for 1-5
days. The mixture was diluted with CH2Cl2 (25 mL) before aq 1
M HCl (25 mL) was added. The org layer was concentrated under
reduced pressure, and the residue was purified by FC (35-50%
gradient EtOAc in heptane) to provide the corresponding amide
products. A suspension of these amide products (∼1 mmol) and
Zn powder (14 mmol) in THF (20 mL) was efficiently stirred in
the presence AcOH (0.5 mL) at room temperature for 3 h. The
mixture was filtered through Celite, and the solvents were evapo-
rated under reduced pressure. The residue was dissolved in CH2Cl2
(10 mL), and aq 0.1 M EDTA (5 mL) was added. The mixture
was stirred at room temperature for 6 h. The org layer was
concentrated under reduced pressure, and the residue was purified
by FC (1-5% MeOH in CH2Cl2) to provide the products 17a-l.

Renin Inhibitors
Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12 3699
(1R,5S)-7-{4-[3-(tert-Butyldimethylsilanyloxy)propyl]phenyl}-9-
methyl-3,9-diazabicyclo[3.3.1]non-6-ene-3,6-dicarboxylic Acid 3-tert-
Butyl Ester 6-Ethyl Ester (19a). A solution of bromide 18a (55.6
g, 168.8 mmol) in THF (1000 mL) was treated at -78 °C with
n-BuLi (1.6 M in hexane, 144 mL, 230 mmol) under N2. After 30
min of stirring at -78 °C a solution of ZnCl2 (1 M in THF, 276
mL, 276 mmol) was added, and the mixture was allowed to warm
to room temperature. Triflate 7 (70.35 g, 153 mmol) in THF (200
mL) was added, followed by Pd(PPh3)4 (4.6 g, 3.8 mmol). The
mixture was heated at 65 °C for 1 h and, after cooling to room
temperature, was quenched with aq sat. NH4Cl (500 mL). The
mixture was partitioned between EtOAc and water. The org layer
was concentrated, and the residue was purified by FC (CH2Cl2/
(1R,5S)-7-{4-[2-(tert-Butyldimethylsilanyloxy)ethoxy]phenyl}-3,9-
diazabicyclo[3.3.1]non-6-ene-3,6,9-tricarboxylic Acid 3-tert-Butyl
Ester 6-Ethyl Ester 9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester
(20b). A suspension of starting material 19b (20 g, 35.7 mmol),
NaHCO3 (30 g, 356.6 mmol), and ꢀ,ꢀ,ꢀ-trichloro-tert-butylchlo-
roformate (17.1 g, 71.3 mmol) in 1,2-dichloroethane (250 mL) was
stirred under N2 at 80 °C overnight. After cooling to room
temperature, the mixture was filtered and concentrated under
reduced pressure, and the residue was purified by FC (2-15%
gradient EtOAc in heptane) to provide the title compound (20.1 g,
75%). 1H NMR (CDCl3): 7.04 (broad, d, J ≈ 7.5 Hz, 2H), 6.84 (d,
J ) 8.1 Hz, 2H), 5.26 (broad, s, 1HR), 5.23 (broad, s, 1H), 5.12
(broad, s, 1HR), 4.41 (broad, s, 1H and 1HR), 4.31 (d, J ) 13.5 Hz,
1H), 4.15-3.89 (m, 7H), 3.22-2.93 (m, 2H and 2HR), 2.84 (dd, J
≈ 18.2, 7.6 Hz, 1H), 2.78 (dd, J ≈ 18.1, 7.0 Hz, 1HR), 2.53 (broad,
d, J ≈ 19.7 Hz, 1H and 1HR), 1.99 (s, 3H), 1.94 (s, 3H), 1.41 (broad,
s, 9HR), 1.37 (broad, s, 9H), 1.02 (t, J ) 6.9 Hz, 3H), 0.92 (s, 9H),
0.11 (s, 6H). IR (film) ν 2929, 2857, 1697, 1604 cm-1. Anal. Calcd
for C34H51N2O8Cl3Si: C, 54.43; H, 6.85; N, 3.73. Found: C, 54.48;
H, 6.77; N, 3.74. [R]2D5 +1.5 (1.02, MeOH). LC-MS: tR ) 1.28
min. MS (ESI): m/z 751.52 (M + 1)+.
(1R,5S)-7-{4-[3-(tert-Butyldimethylsilanyloxy)propyl]phenyl}-3,9-
diazabicyclo[3.3.1]non-6-ene-3,6,9-tricarboxylic Acid 3-tert-Butyl
Ester 9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester (21a). A mixture
of ethyl ester 20a (25.0 g, 33.5 mmol) in aq 1 M NaOH (370 mL)
and EtOH (870 mL) was stirred at 80 °C for 12 h. The EtOH was
evaporated under reduced pressure, and the mixture was diluted
with EtOAc (400 mL), before aq 1 M HCl (240 mL) was added.
The org layer was concentrated under reduced pressure to provide
a mixture of the desilylated acid together with its deconjugated
regioisomer (24.5 g), which was used in the next step without
purification.
A solution of this crude acid mixture (24.5 g), imidazole (9.1 g,
134 mmol), and TBDMS-Cl (13.3 g, 83.7 mmol) in DMF (500
mL) was stirred under N2 overnight. The mixture was partitioned
between heptane (450 mL) and aq sat. NH4Cl (450 mL). The org
layer was concentrated, and the residue was diluted in THF (420
mL). MeOH (140 mL), water (140 mL), and K2CO3 (4.5 g, 32.6
mmol) were added, and the mixture was stirred at room temperature
for 1.5 h. The MeOH was evaporated under reduced pressure, and
the mixture was diluted with Et2O (300 mL) before aq sat. NH4Cl
(300 mL) was added. The org layer was concentrated to provide
the crude title compound mixed with its deconjugated regioisomer
(24.5 g), which was used in the next step without purification.
LC-MS: tR ) 1.23 min. MS (ESI): m/z 721.41 (M + 1)+.
(1R,5S)-7-{4-[2-(tert-Butyldimethylsilanyloxy)ethoxy]phenyl}-3,9-
diazabicyclo[3.3.1]non-6-ene-3,6,9-tricarboxylic Acid 3-tert-Butyl
Ester 9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester (21b). A mixture
of ethyl ester 20b (49.3 g, 65.7 mmol) in aq 1 M NaOH (788 mL)
and EtOH (1100 mL) was stirred at 90 °C for 5 h. The EtOH was
evaporated under reduced pressure, and the mixture was diluted
with EtOAc (400 mL) before aq 1 M HCl (240 mL) was added.
The org layer was concentrated under reduced pressure to provide
the crude desilylated carboxylic acid together with its deconjugated
regioisomer (40 g). MS (ESI): m/z 607.34 (M + 1)+. A solution of
this crude product (40 g), imidazole (13.4 g, 197.2 mmol), and
TBDMS-Cl (29.7 g, 197 mmol) in DMF (350 mL) was stirred under
N2 overnight. The mixture was partitioned between heptane (450
mL) and aq sat. NH4Cl (450 mL). The org layer was concentrated
under reduced pressure, and the residue was diluted in THF (450
mL). MeOH (250 mL), water (120 mL), and K2CO3 (22.8 g, 164
mmol) were added, and the mixture was stirred at room temperature
for 1.5 h. The solvents were partially evaporated under reduced
pressure, and the mixture was diluted with Et2O (450 mL) before
aq sat. NH4Cl (450 mL) was added. The org layer was concentrated
under reduced pressure to provide the title compound mixed with
its deconjugated regioisomer (45.4 g), which was used in the next
step without purification. LC-MS: tR ) 1.20 min. MS (ESI): m/z
723.39 (M + 1)+.
1
MeOH ) 98/2) to provide the title compound (54.4 g, 63%). H
NMR (CDCl3) 7.13 (d, J ) 7.4 Hz, 2H), 7.06 (d, J ) 7.0 Hz, 2H),
4.24 (d, J ) 12.9 Hz, 1H), 4.03-3.84 (m, 3H), 3.77 (s, 1H), 3.70
(s, 1HR), 3.65 (t,, J ) 6.0 Hz, 2H), 3.18 (d, J ) 13.2 Hz, 1H), 3.12
(d, J ) 12.7 Hz, 1H), 2.98 (broad, s, 1H), 2.68 (t, J ) 7.6 Hz, 2H),
2.59 (dd, J ) 19.7, 6.6 Hz, 1H), 2.47 (s, 3HR), 2.43 (s, 3H), 2.36
(d, J ) 19.8 Hz, 1H), 1.84 (q, 2H), 1.43 (broad, s, 9HR), 1.39 (broad,
s, 9H), 0.93 (s, 9H), 0.86 (t, J ) 7.2 Hz, 2H), 0.12 (s, 6H). IR
(film) ν 2929, 2857, 1693 cm-1. Anal. Calcd for C31H50N2O5Si: C,
66.63; H, 9.02; N, 5.01. Found: C, 66.25; H, 8.92; N, 4.98. [R]D25
+1.66 (1.28, MeOH). LC-MS: tR ) 0.98 min. MS (ESI): m/z
559.61 (M + 1)+.
(1R,5S)-7-{4-[2-(tert-Butyldimethylsilanyloxy)ethoxy]phenyl}-9-
methyl-3,9-diazabicyclo[3.3.1]non-6-ene-3,6-dicarboxylic Acid 3-tert-
Butyl Ester 6-Ethyl Ester (19b). A solution of bromide 18b20 (14.7
g, 44.4 mmol) in THF (300 mL) was treated at -78 °C with n-BuLi
(1.6 M in hexane, 37.8 mL, 60.5 mmol) under N2. After the mixture
was stirred for 30 min at -78 °C, ZnCl2 (1 M in THF, 73 mL, 73
mmol) was added, and the mixture was allowed to warm to room
temperature. Triflate 7 (18.5 g, 40.3 mmol) in THF (10 mL) was
added, followed by Pd(PPh3)4 (1.4 g, 1.2 mmol). The mixture was
heated at 65 °C for 1 h and, after cooling to room temperature,
was quenched with aq sat. NH4Cl (300 mL). The mixture was
partitioned between EtOAc and water. The org layer was concen-
trated, and the residue was purified by FC (CH2Cl2/MeOH ) 98
/2) to provide the title compound (19.2 g, 85%). 1H NMR (CDCl3):
7.07 (d, J ) 8.4 Hz, 2H), 6.84 (d, J ) 8.4 Hz, 2H), 4.24 (d, J )
12.9 Hz, 1H), 4.08-3.87 (m, 7H), 3.77 (s, 1H), 3.18 (d, J ) 14.5
Hz, 1H), 3.12 (d, J ) 12.0 Hz, 1H), 2.98 (d, J ) 5.8 Hz, 1H), 2.57
(dd, J ) 20.1, 6.5 Hz, 1H), 2.42 (s, 3H, NCH3), 2.35 (d, J ) 19.7
Hz, 1H), 1.40 (broad, s, 9H), 0.93 (s, 12H), 0.12 (s, 6H). IR (film)
ν 2928, 2856, 1683, 1607 cm-1. Anal. Calcd for C30H48N2O6Si: C,
64.25; H, 8.63; N, 5.00. Found: C, 64.08; H, 8.44; N, 5.04. [R]D25
+4.5 (1.0, MeOH). LC-MS: tR ) 0.98 min. MS (ESI): m/z 561.69
(M + 1)+.
(1R,5S)-7-{4-[3-(tert-Butyldimethylsilanyloxy)propyl]phenyl}-3,9-
diazabicyclo[3.3.1]non-6-ene-3,6,9-tricarboxylic Acid 3-tert-Butyl
Ester 6-Ethyl Ester 9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester
(20a). A suspension of starting material 19a (33.8 g, 60.4 mmol),
NaHCO3 (50 g, 604 mmol), and ꢀ,ꢀ,ꢀ-trichloro-tert-butylchloro-
formate (29.9 g, 121 mmol) in 1,2-dichloroethane (700 mL) was
stirred under N2 at 80 °C overnight. After cooling to room
temperature, the mixture was filtered, concentrated under reduced
pressure, and the residue was purified by FC (5-15% gradient
EtOAc in heptane) to provide the title compound (30.7 g, 69%).
1H NMR (CDCl3) 7.13 (d, J ) 7.5 Hz, 2H), 7.03 (d, J ) 6. Hz,
2H), 5.67 (s, 1H), 5.24 (s, 1HR), 5.14 (s, 1HR), 4.41 (broad, s, 1H
and 1HR), 4.32 (d, J ) 13.2 Hz, 1H), 4.13 (d, J ) 11.8 Hz, 1HR),
4.09 (d, J ) 11.4 Hz, 1H), 4.05-3.87 (m, 2H), 3.65 (t, J ) 6.1
Hz, 2H), 3.14 (d, J ) 13.3 Hz, 1H and 1HR), 3.07 (d, J ) 13.0 Hz,
1H), 2.83 (broad, dd, J ≈ 20.0, 7.0 Hz, 1H), 2.68 (t, J ) 7.4 Hz,
2H), 2.52 (d, J ) 19.5 Hz, 1H), 2.36 (d, J ) 19.8 Hz, 1H), 1.99 (s,
3H), 1.94 (s, 3H), 1.89-1.78 (m, 2H), 1.43 (broad, s, 9HR), 1.39
(broad, s, 9H), 1.02-0.84 (m, 12H), 0.86 (t, J ) 7.2 Hz, 2H), 0.07
(s, 6H). IR (film) ν 2977, 2932, 1694 cm-1. [R]D25 +1.44(1.45,
MeOH). LC-MS: tR ) 1.37 min. MS (ESI): m/z 748.86 (M +
1)+.

3700 Journal of Medicinal Chemistry, 2009, Vol. 52, No. 12
Bezenc¸on et al.
(1R,5S)-7-{4-[3-(tert-Butyldimethylsilanyloxy)propyl]phenyl}-6-
[cyclopropyl-(2,3-dichlorobenzyl)carbamoyl]-3,9-diaza-
bicyclo[3.3.1]non-6-ene-3,9-dicarboxylic Acid 3-tert-butyl Ester
9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester (22a). A solution of acid
21a (23.9 g, 33.2 mmol), HOBt (5.50 g, 39.9 mmol), EDC ·HCl
(16.2 g, 83.0 mmol), DIPEA (17.5 g, 132.8 mmol), and DMAP
(1.0 g, 8.3 mmol) in CH2Cl2 (530 mL) was stirred at room
temperature for 30 min. N-Cyclopropyl-N-(2,3-dichlorobenzy-
l)amine (21.5 g, 99.6 mmol) was added, and the reaction mixture
was stirred at room temperature for 7 days. The mixture was diluted
with CH2Cl2 (300 mL) before aq 1 M HCl (400 mL) was added.
The org layer was concentrated under reduced pressure, and the
residue was purified by FC (3-7% gradient acetone in heptane) to
provide the title compound (18.7 g, 35% over two steps). 1H NMR
(CDCl3) 7.36 (d, J ) 7.6 Hz, 1H), 7.17-6.99 (m, 5H), 6.96 (d, J
) 7.6 Hz, 1H), 6.85 (d, J ) 7.6 Hz, 1HR), 4.78 (broad, d, J ≈ 15.2
Hz, 2H), 4.73-4.44 (m, 2H and 4HR), 4.35 (d, J ) 15.4 Hz, 1H),
4.23 (d, J ) 13.0 Hz, 1H), 4.09 (d, J ) 13.0 Hz, 1HR), 3.70-3.57
(m, 2H), 3.31-2.98 (m, 1H and 3HR), 3.16 (broad, d, J ≈ 12.8
Hz, 1H), 3.04 (d, J ) 12.5 Hz, 1H), 2.77-2.58 (m, 2H), 2.34 (d,
J ≈ 18.2 Hz, 1H), 2.27 (d, J ≈ 18.0 Hz, 1HR), 2.01 (s, 3H), 1.97
(s, 3H), 1.92-1.57 (m, 3H and 1HR), 1.54 (broad, s, 9HR), 1.48
(broad, s, 9H), 0.94 (s, 9H), 0.96-0.74 and 0.68-0.17 (m, 1H and
1HR), 0.07 (s, 6H). IR (film) ν 2928, 2856, 1698 cm-1. Anal. Calcd
for C43H58N3O6Cl5Si: C, 56.24; H, 6.37; N, 4.58. Found: C, 56.79;
H, 6.59; N, 4.65. [R]2D5 +139.9 (0.98, MeOH). LC-MS: tR ) 1.45
min. MS (ESI): m/z 917.87 (M + 1)+.
) 7.7 Hz, 1H), 4.74 (s, 1H and 1HR), 4.72 (broad, d, J ≈ 14.9 Hz,
1H), 4.68 (broad, d, J ≈ 16.3 Hz, 1H), 4.65-4.00 (m, 1H and 2HR,
H-1), 4.48 (broad, d, J ) 12.8 Hz, 1H), 4.41 (d, J ) 15.9 Hz,
1HR), 4.38 (d, J ) 15.7 Hz, 1H), 3.73-3.61 (m, 2H), 3.46 (d, J )
14.7 Hz, 1H), 3.22 (dd, J ) 18.3, 7.5 Hz, 1H), 3.13 (d, J ) 18.1,
7.4 Hz, 1HR), 2.88 (broad, d, J ) 12.8 Hz, 1H), 2.70 (q, J ) 7.4
Hz, 2H), 2.38-2.21 (m, 1H and 1HR), 2.29 (s, 3H), 2.27 (s, 3HR),
2.03 (s, 3H), 2.01 (s, 3H), 1.98 (s, 3HR), 1.93-1.60 (m, 3H and
1HR), 0.95-0.77, 0.73-0.51, 0.45-0.31 and 0.29-0.19 (m, 4H
and 4HR). 13C NMR (CDCl3) 173.07, 172.87, 171.24, 171.21 (CO
and COR), 151.59 (Cq), 150.61 (CqR), 142.59 (Cq), 142.44 (CqR),
138.49 (Cq), 138.22 (CqR), 137.14 (Cq), 137.07 (CqR), 133.06 (Cq),
133.00 (CqR), 131.30 (CqR), 131.00 (Cq), 129.04 (CHarom), 129.02
(CHarom), 128.79 (CHarom), 128.70 (CHarom), 128.60 (CHRarom),
128.53 (CHRarom), 127.42 (CHRarom), 127.26 (CHarom), 127.15
(CHarom), 127.07 (CHarom), 126.73 (CHRarom), 126.56 (CHarom),
126.47 (CHarom), 126.36 (CHarom), 126.18 (CHRarom), 125.42 (Cq),
106.65 (CCl3), 106.53 (CRCl3), 89.32 (Me2C-O), 88.95 (Cq,
Me2CR-O), 62.02 (C12), 51.00, 49.54 (C5 and C5R), 50.52 (C4),
49.97 (C4R), 49.33, 49.02 (C14 and C14R), 47.55, 45.82 (C1 and
C1R), 46.92, 46.77 (C2 and C2R), 34.23 (C11R), 33.97 (C11), 33.17,
32.83 (C8 and C8R), 31.73 (C10), 31.21, 31.12 (C15 and C15R),
21.77, 21.69, 21.61, 21.32, and 21.29 (CH3CO, CRH3CO,
(CH3)2C-O, (CRH3)2C-O), 10.72, 10.54, 6.58, 6.00 (C16 and
C16R). IR (film) ν 2925, 2159, 2025, 1703, 1620 cm-1. Anal. Calcd
for C34H38N3O5Cl5: C, 54.75; H, 5.13; N, 5.63. Found: C, 54.52;
H, 5.15; N, 5.34. [R]2D5 +177.0 (1.02, MeOH). LC-MS: tR ) 1.15
min. MS (ESI): m/z 746.43 (M + 1)+.
(1R,5S)-7-{4-[2-(tert-Butyldimethylsilanyloxy)ethoxy]phenyl}-6-
[cyclopropyl-(2,3-dichlorobenzyl)carbamoyl]-3,9-
diazabicyclo[3.3.1]non-6-ene-3,9-dicarboxylic Acid 3-tert-Butyl Es-
ter 9-(2,2,2-Trichloro-1,1-dimethylethyl) Ester (22b). A solution of
acid 21b (40 g, 55.4 mmol), HOBt (9.9 g, 72.0 mmol), EDC ·HCl
(28.4 g, 145.2 mmol), DIPEA (29.2 g, 221.5 mmol), and DMAP
(1.7 g, 13.8 mmol) in CH2Cl2 (650 mL) was stirred at room
temperature for 30 min. N-Cyclopropyl-N-(2,3-dichlorobenzy-
l)amine (34.0 g, 157.3 mmol) was added, and the reaction mixture
was stirred at room temperature for 7 days. The mixture was diluted
with CH2Cl2 (650 mL) before aq 1 M HCl (450 mL) was added.
The org layer was concentrated under reduced pressure, and the
residue was purified by FC (3-20% gradient acetone in heptane)
(1R,5S)-3-Acetyl-6-[cyclopropyl(2,3-dichlorobenzyl)carbamoyl]-
7-[4-(2-hydroxyethoxy)phenyl]-3,9-diazabicyclo[3.3.1]non-6-ene-9-
carboxylic Acid 2,2,2-Trichloro-1,1-dimethylethyl Ester (23b). A
solution of starting material 22b (20.4 g, 22.2 mmol) in CH2Cl2
(250 mL) was cooled to 0 °C. HCl (4 M in dioxane, 75 mL) was
added, and the mixture was stirred 0 °C for 20 min and at room
temperature for 3 h. The mixture was diluted with CH2Cl2 (150
mL) before aq 1 M NaOH (100 mL) was carefully added. The org
layer was concentrated under reduced pressure, the residue was
dissolved under N2 in THF (250 mL), and the solution was cooled
to -78 °C. DIPEA (15.2 mL, 88.7 mmol), DMAP (0.27 g, 2.2
mmol), and AcCl (1.6 mL, 22.2 mmol) were added, and the mixture
was stirred at -78 °C for 15 min. MeOH (250 mL) was added,
and the mixture was allowed to warm to room temperature. The
mixture was diluted with EtOAc (200 mL) before aq 1 M HCl
(200 mL) was added. The org layer was concentrated under reduced
pressure, and the residue was purified by FC (EtOAc) to provide
1
to provide the title compound (21.0 g, 41% over two steps). H
NMR (CDCl3) 7.33 (broad, s, 1H), 7.17-7.02 (m, 3H), 6.85-6.69
(m 3H), 4.81-4.32 (m, 5H and 5HR), 4.23 (d, J ) 13.5 Hz, 1H),
4.13-3.93 (m, 4H and 1HR), 3.28-3.0 (m, 2H and 3HR), 3.04
(broad, d, J ≈ 12.6 Hz, 1H), 2.32 (d, J ≈ 18.0 Hz, 1H), 2.26 (d, J
≈ 18.1 Hz, 1HR), 2.17 (broad, d, J ≈ 21.7 Hz, 1HR), 2.01 (s, 3H),
1.97 (s, 3H), 1.82-1.59 (m, 1H and 1HR), 1.53 (broad, s, 9HR),
1.48 (broad, s, 9H), 0.94 (s, 9H), 0.94-0.73 and 0.68-0.17 (m,
1H and 1HR), 0.13 (s, 6H). IR (film) ν 2928, 2856, 1698, 1608
cm-1. Anal. Calcd for C42H56N3O7Cl5: C, 54.82; H, 6.13; N, 4.57.
Found: C, 55.02; H, 6.09; N, 4.56. [R]2D5 +126.0 (1.0, MeOH).
LC-MS: tR ) 1.34 min. MS (ESI): m/z 920.65 (M + 1)+.
1
the title compound (11.8 g, 71%). H NMR (CDCl3) 7.36 (t, J )
6.7 Hz, 1H), 7.18-7.02 (m, 3H), 6.83 (d, J ) 8.4 Hz, 1H), 6.77
(d, J ) 8.4 Hz, 1H), 6.69 (t, J ) 9.7 Hz, 1H), 4.74 (s, 1H and
1HR), 4.72 (broad, d, J ≈ 14.4 Hz, 1H), 4.62 (broad, d, J ≈ 16.0
Hz, 1H and 2HR), 4.58-4.47 (m, 1H and 2HR), 4.46 (broad, d, J
) 10.4 Hz, 1H and 1HR), 4.40 (d, J ) 12.4 Hz, 1HR), 4.36 (d, J )
16.2 Hz, 1H), 4.11-4.02 (m, 2H), 4.01-3.92 (m, 2H), 3.48 (d, J
) 14.1 Hz, 1HR), 3.44 (d, J ) 14.0 Hz, 1H), 3.20 (dd, J ) 18.1,
7.4 Hz, 1H), 3.10 (d, J ) 17.9, 7.4 Hz, 1HR), 2.88 (broad, d, J ≈
11.1 Hz, 1H), 2.38-2.20 (m, 1H and 1HR), 2.29 (s, 3H), 2.27 (s,
3HR), 2.03 (s, 3H), 2.01 (s, 3H), 1.99 (s, 3HR), 1.97 (s, 3H),
1.82-1.66 (m, 1H and 1HR), 0.92-0.77, 0.72-0.50, 0.47-0.30,
and 0.29-0.19 (m, 4H and 4HR). IR (film) ν 2925, 1703, 1607
cm-1. Anal. Calcd for C33H36N3O6Cl5: C, 52.99; H, 4.85; N, 5.62.
Found: C, 54.21; H, 5.30; N, 5.25. [R]2D5 +117.3 (1.0, MeOH).
LC-MS: tR ) 1.12 min. MS (ESI): m/z 748.36 (M + 1)+.
(1R,5S)-3-Acetyl-6-[cyclopropyl(2,3-dichlorobenzyl)carbamoyl]-
7-[4-(3-hydroxypropyl)phenyl]-3,9-diazabicyclo[3.3.1]non-6-ene-9-
carboxylic Acid Trichloromethyl Ester (23a). A solution of starting
material 22a (18.5 g, 20.1 mmol) in CH2Cl2 (180 mL) was cooled
to 0 °C. HCl (4 M in dioxane, 68 mL) was added, and the mixture
was stirred 0 °C for 20 min and at room temperature for 3 h. The
mixture was diluted with CH2Cl2 (100 mL) before aq 1 M NaOH
(100 mL) was carefully added. The org layer was concentrated
under reduced pressure, the residue was dissolved under N2 in THF
(380 mL), and the solution was cooled to -78 °C. DIPEA (13.8
mL, 80.5 mmol), DMAP (0.246 g, 2.0 mmol), and CH3COCl (1.36
mL, 19.1 mmol) were added, and the mixture was stirred at -78
°C for 15 min. MeOH (100 mL) was added, and the mixture was
allowed to warm to room temperature. The mixture was diluted
with EtOAc (150 mL) before aq 1 M HCl (150 mL) was added.
The org layer was concentrated under reduced pressure, and the
residue was purified by FC (CH2Cl2/MeOH 98.2) to provide the
General Conditions for the Preparation of Compounds 24a-k
and 25a-l. To a solution of alcohol 23a (1.0 g, 1.3 mmol) or 23b
(1.66 g, 2.2 mmol), 1,1′-(azodicarbonyl)dipiperidine (2.7 mmol/
4.5 mmol), and the desired phenol (2.7 mmol/4.4 mmol) in toluene
(30 mL) was added PBu3 (85%, 4.0 mmol/6.7 mmol) at room
temperature, and the reaction mixture was stirred at 80 °C for 12 h.
Heptane (75 mL) was added, and the mixture was filtered. The
solvents were removed under reduced pressure, and the residue was
dissolved in CH2Cl2 (40 mL). At 0 °C aq H2O2 (25% in water, 8
mL) was added and the mixture was stirred at 0 °C for 5 min and
1
title compound (12.7 g, 84%). H NMR (CDCl3) 7.36 (t, J ) 6.6
Hz, 1H), 7.18-7.00 (m, 4H), 6.77 (d, J ) 7.7 Hz, 1H), 6.72 (d, J

Renin Inhibitors
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at room temperature for 1.5 h. Aqueous 1 M NaOH (20 mL) and
brine (20 mL) were added. The org layer was concentrated under
reduced pressure, and the residue was purified by FC (20-35%
EtOAc in heptane) to provide the corresponding crude phenyl ether.
A suspension of this crude product and of Zn powder (6.6 mmol)
in THF (20 mL) was stirred in the presence of AcOH (0.4 mL) at
room temperature for 3 h. The mixture was filtered through Celite,
the solvents were evaporated under reduced pressure, and the
residue was dissolved in CH2Cl2 (3 mL). Aqueous 0.1 M EDTA
(5 mL) was added. The mixture was stirred at room temperature
for 6 h. The org layer was concentrated, and the residue was purified
by FC (3-5% MeOH in CH2Cl2) to provide the corresponding title
product.
Acknowledgment. We thank Remy Dureau, Patrick Eckert,
Martin Faes, Sven Glutz, Luke Harris, Bela Humer, Daniel
Marchal, Sophie Moujon, Emmanuelle Andre´, Pascal Rebmann,
Daniel Trachsel, Gabriela Vorburger, Alain Chambovey, An-
toinette Amrein, Geoffroy Bourquin, Fabienne Drouet, Aude
Weigel, Markus Rey, and Daniel Wanner for their help and
engagement on this program. We thank Synphabase AG as well
for the preparation of compound 6 in kilogram scale. We thank
Dr. Clemens Schulze-Briese, Takashi Tomizaki, and their group
for their support in setting up the beamline.
Supporting Information Available: Experimental details of
some precursors, full spectroscopic data of all compounds with
assignments of the 1H and 13C NMR signals, X-ray structure analysis
results, and details of the inhibition assay, pharmacokinetic experi-
ments on the rat, and pharmacodynamic experiments on double
transgenic rats. This material is available free of charge via the
Internet at http://pubs.acs.org.
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