A novel series of 4-piperidinopyridine and 4-piperidinopyrimidine inhibitors of 2,3-oxidosqualene cyclase-lanosterol synthase

Article abstract of DOI:10.1021/jm000139k

A novel series of 4-piperidinopyridines and 4-piperidinopyrimidines showed potent and selective inhibition of rat 2,3-oxidosqualene cyclase-lanosterol synthase (OSC) (e.g. 26 IC₅₀ rat = 398 ± 25 nM, human = 112 ± 25 nM) and gave selective oral inhibition of rat cholesterol biosynthesis (26 ED₈₀ = 1.2 ± 0.3 mg/kg, n = 5; HMGCoA reductase inhibitor simvastatin ED₈₀ = 1.2 ± 0.3 mg/kg, n = 5). The piperidinopyrimidine OSC inhibitors have a significantly lower pK_a than the corresponding pyridine or the previously reported quinuclidine OSC inhibitor series. This indicates that other novel OSC inhibitors may be found in analogues of this series across a broader pK_a range (6.0-9.0). These series may yield novel hypocholesterolemic agents for the treatment of cardiovascular disease.

Full text of DOI:10.1021/jm000139k

4964
J . Med. Chem. 2000, 43, 4964-4972
A Novel Ser ies of 4-P ip er id in op yr id in e a n d 4-P ip er id in op yr im id in e In h ibitor s
of 2,3-Oxid osqu a len e Cycla se-La n oster ol Syn th a se
George R. Brown,* David M. Hollinshead, Elaine S. E. Stokes, David Waterson, David S. Clarke,
Alan J . Foubister, Steven C. Glossop, Fergus McTaggart, Donald J . Mirrlees, Graham J . Smith, and Robin Wood
AstraZeneca, Alderley Park, Macclesfield, Cheshire SK10 4TG, U.K.
Received March 24, 2000
A novel series of 4-piperidinopyridines and 4-piperidinopyrimidines showed potent and selective
inhibition of rat 2,3-oxidosqualene cyclase-lanosterol synthase (OSC) (e.g. 26 IC50 rat ) 398
( 25 nM, human ) 112 ( 25 nM) and gave selective oral inhibition of rat cholesterol
biosynthesis (26 ED80 ) 1.2 ( 0.3 mg/kg, n ) 5; HMGCoA reductase inhibitor simvastatin
ED80 ) 1.2 ( 0.3 mg/kg, n ) 5). The piperidinopyrimidine OSC inhibitors have a significantly
lower pKa than the corresponding pyridine or the previously reported quinuclidine OSC inhibitor
series. This indicates that other novel OSC inhibitors may be found in analogues of this series
across a broader pKa range (6.0-9.0). These series may yield novel hypocholesterolemic agents
for the treatment of cardiovascular disease.
Sch em e 1. Oxidosqualene Cyclization to Lanosterol
In tr od u ction
The HMGCoA reductase inhibitory drugs (statins) are
now well-established as agents for lowering plasma
cholesterol levels and have widespread therapeutic use
in the treatment of cardiovascular disease. Their use is
strongly supported1,2 by long-term clinical trials, which
showed significant reductions in the mortality rates of
patients with cardiovascular disease. This successful
therapy with the statin class of hypocholesterolemic
agents has led to a search for other novel inhibitors of
the cholesterol biosynthesis pathway3 that should also
afford a lowering of plasma cholesterol levels. The area
of greatest interest to date has been inhibition at the
squalene synthase (SQS) step.4 In addition to work on
novel SQS inhibitors,5,6 we have considered7 the inhibi-
tion of other enzyme steps of the cholesterol biosynthesis
pathway as potential targets for medicinal chemistry.
We sought new inhibitors at later biosynthetic steps
where the pathway is dedicated3 to the biosynthesis of
cholesterol, as others have previously reasoned4,8 in the
design of both SQS and 2,3-oxidosqualene cyclase (OSC;
EC 5.4.99.7) inhibitors. The enzyme targets considered
were thus beyond the formation of farnesyl pyrophos-
phate and before the formation of lanosterol. In this
approach inhibition of steps before SQS is avoided
because they are involved in protein prenylation and
the synthesis of key biosynthetic precursors such as
ubiquinone. Inhibition of steps after the formation of
lanosterol is also avoided because it previously afforded3
toxic inhibitors by accumulation of higher sterols such
as desmosterol. This analysis leads to squalene oxida-
tion and the subsequent cyclization (Scheme 1, 1-3) of
2,3-oxidosqualene (OS, 1) by OSC, as optimal steps for
enzyme inhibition. Herein we describe a novel series of
4-piperidinopyridine and 4-piperidinopyrimidine inhibi-
tors of OSC together with their in vivo effects on
cholesterol biosynthesis as a whole.
reported,8 and many of these compounds closely re-
semble the enzyme substrate 1 or the cyclization
product lanosterol 3. These mimics tend to have high
lipophilicity (CLOGP9 > 7) and molecular weight (>400).
The synthesis of further analogues of these mostly weak
inhibitors was rejected on the grounds that they rep-
resented a poor medicinal chemistry starting point for
the discovery of an orally active inhibitor of cholesterol
biosynthesis. Instead a compound was sought which
would potentially interfere with the cyclization of 1 to
the putative protolanosteryl cation 2 (Scheme 1). This
cation is widely accepted10 as the first tetracyclic
intermediate in the cyclization (1-3) of oxidosqualene
to lanosterol. In addition we sought, as part of our
rationale, to mimic the cation formed in the acid-
catalyzed opening of the 2,3-oxirane ring of 1 with a
physiologically protonated amine. The structures of the
similarly derived potent and orally active OSC inhibi-
tors (Chart 1): 4a (BIBB 515),11 5 (BIBX 79),12 6a (Ro
48-8071),13 and 6b (Ro 44-2103)14 were also considered
in this analysis. The more rigid inhibitors 4a , 5, and
6b were used as templates for directed 2D-searching of
the >300000 compounds in the AstraZeneca Alderley
Park compound collection. A requirement was included
for the presence of a terminal nitrogen lone pair at the
position potentially corresponding to the oxygen lone
A considerable number of OSC inhibitors have been
* To whom correspondence should be addressed. Phone: 01625-
515918. Fax: 01625-516667. E-mail: George.R.Brown@astrazeneca.com.
10.1021/jm000139k CCC: $19.00 © 2000 American Chemical Society
Published on Web 11/18/2000

Piperidinopyridine and -pyrimidine Inhibitors of OSC
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 26 4965
Ch a r t 1
in the Experimental Section. This method involved the
reaction of 4-carboxy-N-(4-pyridyl)piperidine15 (27) with
SOCl2 and subsequent acylation of N-Boc-piperazine in
the presence of Et3N. The Boc protective group was
removed with TFA to afford 28. The substituted pip-
eridine 28 was reacted with an appropriately substi-
tuted benzenesulfonyl chloride in the presence of Et3N
to give 8-17. Compounds 18 and 19 (Table 3) were
similarly obtained by reaction of 28 with 4-bromobenzyl
bromide and 4-bromobenzoyl chloride, respectively, but
for 18 the base used was NaH.
A second synthetic route B (Scheme 3) was used to
prepare compounds 7, 20-22, 25, and 26 (Table 4). 7
was synthesized by reaction of N-4-iodophenylsulfo-
nylpiperazine with the acid chloride of 27 in the pres-
ence of Et3N. In a second variation N-4-bromophenyl-
sulfonylpiperazine was allowed to react with 29 in
CH2Cl2 to afford 30 and the Boc group removed with
TFA to give 31. The final synthetic step for compounds
22, 25, and 26 was reaction of 31 with the appropriate
chloro heterocycle in the presence of Et3N in boiling
EtOH or THF. For 20 and 21 the 2-chloroethyl isocy-
anate and 2-chloroethyl isothiocyanate, respectively,
were allowed to react with 31 in THF before subsequent
ring closure in boiling H2O.
Synthetic route C (Scheme 4) was used for the
synthesis of 23 and 24 (Table 4). Isonipecotic acid was
reacted with CbzCl in the presence of Na2CO3 to give
32, which was condensed with N-Boc-piperazine in the
presence of HOBt and EDAC (1-[3-(dimethylamino)-
propyl]-3-ethylcarbodiimide hydrochloride) to afford 33.
The Cbz group was removed by hydrogenation over 10%
Pd/C at 25 °C to afford 34. 34 was allowed to react with
2-chloropyrimidine in boiling EtOH containing Et3N to
give 35a . The Boc protective group was removed from
35a with TFA to give 35b, which gave 23 after reaction
with 4-bromophenylsulfonyl chloride in Et3N/THF. For
the pyridazine 24, regioselective16 alkylation with 3,4,5-
trichloropyridazine gave the 5-substituted 35c as the
main product. The two additional 3- and 4-chlorine
atoms were removed by catalytic reduction with hydro-
gen over 10% Pd/C to give 35d . The Boc protecting group
was removed from 35d with TFA to afford 35e, and
reaction of 35e with 4-bromophenylsulfonyl chloride in
Et3N/THF yielded 24.
Sch em e 2. Synthetic Route Aa
Biologica l Resu lts a n d Discu ssion
a
(a) SOCl2, Et3N, TFA, CH2Cl2; (b) Et3N, pyridine, 0 °C.
The strategy employed for evaluation of compounds
as OSC inhibitors has been fully described in relation
to our study of quinuclidine inhibitors of OSC.7 Com-
pounds were assessed first in oral cholesterol biosyn-
thesis tests in rats before in vitro confirmation that the
in vivo activity found was derived from inhibition of the
target OSC enzyme. In summary, compounds were
assessed for inhibition of cholesterol biosynthesis in rats
2 h after an oral dose of test compound. First the
cholesterol biosynthetic precursor pattern found in the
HPLC analysis of extracts of liver samples from orally
dosed rats was compared with the same profile from
control animals. These HPLC chromatograms showed
whether accumulated substrates for cholesterol biosyn-
thesis enzymes were present and thereby gave an
indication of which steps of cholesterol biosynthesis had
been inhibited: i.e. it enabled determination of whether
pair of the oxirane ring of 1 at the enzyme active site.
This search led to recognition (Chart 1) that the
substituted pyridine 7 resembled the known11 inhibitor
4a and was a suitable medicinal chemistry starting
point for optimization of OSC inhibition. Novel ana-
logues 8-26 (Tables 1, 3, and 4) of 7 were synthesized
and examined for oral inhibition of rat cholesterol
biosynthesis and as inhibitors of rat microsomal OSC.
Inhibition of cholesterol biosynthesis was accepted as a
surrogate for plasma cholesterol-lowering activity, as
seen with the clinically used statin drugs.
Ch em istr y
Compounds 8-17 were prepared in multiparallel
fashion by synthetic route A (Scheme 2) with the
synthesis of 14 being described as a typical compound

4966 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 26
Sch em e 3. Synthetic Route Ba
Brown et al.
a
(a) CH2Cl2; (b) TFA, Na2CO3; (c) chloro heterocycle, Et3N, EtOH; (d) THF; (e) H2O, 100 °C.
Sch em e 4. Synthetic Route Ca
a
(a) CbzCl, Na2CO3, 2 M HCl; (b) EDAC, HOBt, DMF, N-Boc-piperazine; (c) H2, Pd/C, EtOH; (d) chloro heterocycle, Et3N, THF, TFA,
CH2Cl2, 25 °C; (e) Et3N, THF.
the enzyme inhibition was selective for the OSC step.
This analysis of the in vivo mode of action was seen as
an important process for rejecting compounds with
inhibitory actions at other biosynthetic steps outside of
our interest. In addition it alleviated the need for much
in vitro selectivity testing at the other cholesterol
biosynthetic steps. Second, the extent of the inhibition
of cholesterol biosynthesis was also measured from the
same HPLC chromatograms and initially expressed as
percent inhibition after a 2 mg/kg (Table 1) oral dose of
test compound (n ) 2). Compounds were initially tested
in rats at this relatively low dose for two reasons: (a) 7
and 8 gave >70% inhibition of cholesterol biosynthesis
when tested at 5 mg/kg (n ) 2), making this dose level
potentially unsuitable for differentiating between test
compounds in this series, and (b) the aim of the study
was to discover inhibitors of the cholesterol biosynthesis
pathway with at least similar inhibitory potency to that
found for the widely prescribed HMGCoA reductase
inhibitors, (e.g. simvastatin ED80 ) 1.2 ( 0.3 mg/kg in
this test). Key compounds were retested to obtain an
IC50 for OSC inhibition and a more detailed confirma-
tory oral ED50 or ED80 for inhibition of rat cholesterol
biosynthesis. The trifluoromethyl-substituted oxazoli-
dine 4b17 was used as a standard OSC inhibitor, and
the HMGCoA reductase inhibitor simvastatin was the
standard compound used for cholesterol synthesis in-
hibition.
Compound 7 was examined in this manner and it
afforded an IC50 of 82 ( 14 nM for the inhibition of rat
OSC with a 52% inhibition of cholesterol biosynthesis
at 2 mg/kg (Tables 1 and 2). The subsequent ap-
proximate ED80 found for 7 (shallow dose response) was
<20 mg/kg and the ED50 2.0 ( 0.3 mg/kg. Thus overall
7 was a potent inhibitor of cholesterol biosynthesis by
inhibition at the OSC step. This level of inhibition of

Piperidinopyridine and -pyrimidine Inhibitors of OSC
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 26 4967
Ta ble 1. Inhibition of Rat Cholesterol Biosynthesis after a 2
Ta ble 3. Comparison of Percent Inhibition for
mg/kg Oral Dose
Phenylpiperazine Link Groups in the Rat
compd
R
formula
mp, °C
% inhiba
% inhib
7
8
9
4-I
H
4-Br
3-Br
4-F
C21H25IN4O3S
211-213
143-146
188-189
gum
130-133
144-146
133-135
168-169
64-66
52
53
69
43
46
52
10
12
38
7
cholesterol ED50 cholesterol
C
C
C
C
C
C
C
C
21H26N4O3S
OSC at biosynth at biosynth inhib,
21H25BrN4O3S
21H25BrN4O3S
21H25FN4O3S
21H25FN4O3S
21H24F2N4O3S
22H25N5O3S
compd
formula
1 µMa
2 mg/kgb
mg/kgc
10
11
12
13
14
15
16
17
4b
9
18
19
C21H25BrN4O3S
21H27BrN4O
22H27BrN4O2
100
100
96
69
48
28
2.0
1.8
C
C
3-F
2,4-di-F
4-CN
4-OMe
3-CF3
4-Me
n ) 2, (9%. n ) 2, (9.0%, oral dose. c n ) 5, (0.3%, oral
a
b
22H28N4O4S
dose.
C21H25F3N4O3S
160-164
201-203
151-152
would have a different conformational orientation in
relation to the piperazino ring, compared to the benzoyl
group of 19. These in vitro findings contrast with the
drug design paradigm used for the similarly shaped
benzophenone class of OSC inhibitors: e.g. Ro 44-2103
(6b).13 In this inhibitor series an electrophilic carbonyl
group was incorporated at a structural position remote
from the terminal basic group (in a similar position to
group X in 9, 18, and 19, Table 3). The intention here
was to interact with OSC at both the intermediate
protosteryl cation site and the 2,3-oxirane ring of 1. The
good inhibitory activity found for the benzyl-linked 18
suggests that the piperidinopyridine series of inhibitors
does not have the bifunctional interaction with the OSC
enzyme postulated for Ro 44-2103 (6b).
C
C
22H28N4O3S
23H21F3N2O2
66
93
a
(9.0%, n ) 2; inhibition of cholesterol biosynthesis after an
oral dose.
Ta ble 2. Inhibition of Rat and Human OSC by
4-N-Piperazinopyridines
IC50, nMa
compd
rat
human
7
8
9
4b
82b
326c
228c
90b
143 ( 33
161 ( 39
n ) 5. Error ) (14 nM. c Error ) (25 nM.
a
b
cholesterol biosynthesis, however, compared unfavor-
ably with the superior ED80 of 1.2 ( 0.3 mg/kg found
for the clinically used simvastatin, indicating that an
improved in vivo biosynthesis inhibition was desirable.
The 4-iodo atom in the phenyl ring of 7 was considered
to be potentially unstable in vivo; it contributed signifi-
cantly to the molecular weight as well as added over 1
unit to the CLOGP value of 7 compared to hydrogen
substitution at the same position. Hence using a parallel
synthesis procedure, synthetic analogues of 7 were
prepared (Table 1, 8-17). Preliminary structure-activ-
ity information for phenyl ring substitution was ob-
tained from the determination of the inhibition of rat
cholesterol biosynthesis following oral dosing. The in-
hibitory results obtained after a 2 mg/kg oral dose
indicated that inhibition of cholesterol biosynthesis
comparable to that found with 4b and 7 was only
achievable at this dose with a very limited group of
lipophilic substituents. Confirmatory IC50 data for
inhibition of rat and human OSC (human IC50 data were
obtained in the same manner as described in the
Experimental Section for rat IC50 data) were determined
for compounds 7-9 (Table 2); the inhibitory potency was
greater at the human OSC enzyme. On the basis of the
preliminary structure-activity information derived from
in vitro and in vivo testing, 9 was selected as a lead
structure for further structural variation.
Structural variation of the lead structure 9 involving
the synthesis of compounds containing other hetero-
cycles in place of the pyridine ring led to several potent
inhibitors of rat cholesterol biosynthesis (Table 4). For
example the oxazoline 20 and thiazoline 21 were very
potent inhibitors of rat cholesterol biosynthesis, whereas
the thiadiazole 22 gave no inhibition of rat cholesterol
biosynthesis at a 2 mg/kg oral dose. Examination of the
HPLC chromatograms derived from the extraction of
liver samples of rats dosed with 20 and 21 showed,
however, that both compounds were nonselective inhibi-
tors blocking a number of cholesterol biosynthetic steps.
Particularly the conversion of desmosterol to cholesterol
was inhibited. Thus, despite their inhibitory potency 20
and 21 were not examined further because of this
potential to accumulate higher sterols, for which our
design hypothesis predicted toxic effects. The 2-substi-
tuted pyrimidine 23, which did not have the terminal
nitrogen atom present in the 4-pyridyl and 4-pyrimidyl
compounds (Table 4), did not inhibit rat OSC at a 1 µM
concentration or rat cholesterol biosynthesis at an oral
dose of 2 mg/kg. By contrast the pyridazine 24 did
inhibit cholesterol biosynthesis, but much more potent
cholesterol biosynthesis inhibition was found for the
4-substituted pyrimidine 25 and its 2-methyl analogue
26. The inhibitory activity of both of these compounds
was explored in detail (Table 5) against rat and human
OSC (human IC50 data were obtained in the same
manner as described in the Experimental Section for
rat IC50 data) and cholesterol biosynthesis. The rat oral
ED80 values for 25 and 26 were similar to the result for
simvastatin (25, 1.4 ( 0.3 mg/kg; 26, 1.2 (0.3 mg/kg;
simvastatin, 1.2 ( 0.3 mg/kg).
Changing the sulfonyl link of 9 to methylene (18) and
carbonyl (19) afforded OSC inhibitors (Table 3), but 9
and 18 were slightly more potent inhibitors of choles-
terol biosynthesis in vivo than 19. Interestingly 9, 18,
and 19 were all inhibitors of OSC even though the
linked phenylsulfonyl and benzyl groups of 9 and 18

4968 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 26
Brown et al.
Ta ble 4. Inhibition of Rat OSC and Rat Cholesterol Biosynthesis for Pyridine Ring Analogues
a
b
% Inhib ( 15%, n ) 2. % Inhib ( 9%, n ) 2, after an oral dose.
Ta ble 5. Comparison of Inhibitory Activities of Pyridino- and
Pyrimidinopiperidines
vivo than the standard quinuclidine OSC inhibitor 36
(Table 5) but were similar in potency (both in vitro and
in vivo) to the reported11-13 lipophilic amine OSC
inhibitors 4a , 4b, 5, 6a , and 6b, e.g. 4b in Table 5.
IC50, nMa
rat inhib cholesterol biosynthb
compd
rat
human
ED50, mg/kg
ED80, mg/kg
Our previous work7 in the quinuclidine series of OSC
inhibitors suggested that an inhibitor with a pKa close
to 9 might be necessary for inhibition of OSC (e.g. 36,
pKa ) 9.1), which is similar to that found (pKa values
were measured spectrophotometrically using a Beckman
DU-7 spectrophotometer) for the pyridine-substituted
inhibitors 7 and 9 (Table 6, pKa ) 9.2). This contrasts
with the much lower pKa of 4.7 found for the weakly
basic phenyloxazoline inhibitor 4b used as a standard
OSC inhibitor and the wide pKa range of the other OSC
inhibitors cited11-13 above. In addition some recently
described polyprenylated mimics (Chart 2, 37-39)18-20
of the putative carbocation intermediates of OS cycliza-
tion with a wide pKa range have also shown good in vitro
9
25
26
4b
36
228c
116c
398c
90d
161 ( 39
37 ( 9
2.0 ( 0.3
0.5 ( 0.2
0.3 ( 0.2
0. 4 ( 0.2
2.0 ( 0.3
>2.0
1.4 ( 0.3
1.2 ( 0.3
1.0 ( 0.3
112 ( 25
58d
35 ( 9
a
b
c
n ) 5. n ) 5, inhibition after an oral dose. Error ) (25 nM.
dError ) (14 nM.
Comparison of the inhibitory properties of the pyri-
dino- and pyrimidino-substituted piperazines (Table 5)
indicated that substitution with pyrimidine afforded
compounds with significant in vitro OSC inhibitory
potency and with superior in vivo inhibition of rat
cholesterol biosynthesis compared with the correspond-
ing pyridine 9. These pyrimidines were more potent in

Piperidinopyridine and -pyrimidine Inhibitors of OSC
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 26 4969
Ta ble 6. Comparison of Physical Properties in the OSC
Inhibitory Series
(with SiMe4 as an internal standard) and mass spectra were
measured on a MS902 Kratos (AEI) instrument. Elemental
analyses were determined on a Perkin-Elmer series II-2400
analyzer. Reactions were carried out under an atmosphere of
argon and column chromatography was on E. Merck silica gel
(Kieselgel 60, 230-400 mesh) or neutral alumina N (ICN
Biomedicals). Bond Elut columns were silica Mega Bond Elut
(Varian). Solvents were dried over MgSO4 before evaporation.
Exa m p le of Syn th etic Rou te A. 1-(1-(4-P yr id yl)p ip er i-
d in -4-ylca r bon yl)p ip er a zin e (28). 2715 (4.12 g, 20 mmol)
was suspended in dry CH2Cl2 (30 mL) and treated with SOCl2
(3 mL, 41 mmol) dropwise with cooling at 0 °C. The mixture
was stirred for 1 h followed by the removal by evaporation of
the solvent and the excess SOCl2. The resulting gum was
dissolved in CH2Cl2 (80 mL) and added slowly, with cooling to
a solution of N-Boc-piperazine (3.72 g, 20 mmol) in CH2Cl2 (100
mL) and Et3N (15 mL, 107 mmol). The reaction mixture was
stirred for 2 h and the solvent removed by evaporation. The
residue was crystallized from EtOAc to give, as a waxy yellow
solid, 4-(tert-butoxycarbonyl)-1-(4-pyridylpiperidin-4-ylcarbo-
nyl)piperazine (3.74 g, 50%): 1H NMR (DMSO-d6) δ 1.38(s,
9H), 1.58(m, 4H), 2.90(m, 3H), 3.36(m, 6H), 3.50(bs, 2H), 3.91-
(d, J ) 12.5 Hz, 2H), 6.79(d, J ) 6.4 Hz, 2H), 8.09(d, J ) 6.4
Hz, 2H); EI-MS m/z 374 (M + H).
compd
pKa
CLOGP9
7
9
25
26
36
4b
9.2
9.2
6.1
7.2
9.1
4.7
3.7
3.4
2.6
3.2
4.7
5.3
Ch a r t 2
This product was dissolved in dry CH2Cl2 (50 mL), treated
with TFA (5.3 mL, 68 mmol) and stirred under an argon
atmosphere at room temperature for 3 h. The CH2Cl2 solvent
was removed by evaporation to afford a brown oil which slowly
solidified after treatment with an excess of saturated NaHCO3
solution. This solid was taken up in CH2Cl2, and the solution
washed with H2O, brine and dried. The resultant solution was
evaporated to dryness to yield an oil, which slowly crystallized
to give, as a yellow solid, 28 (2.45 g, 90%): mp 131-132 °C;
1H NMR (DMSO-d6) δ 1.53(m, 2H), 1.78(m, 2H), 3.03(m, 5H),
3.241(m, 2H), 3.74(m, 4H), 4.20(d, J ) 12.5 Hz, 2H), 7.17(d, J
) 6.4 Hz, 2H), 8.18(d, J ) 6.4 Hz, 2H), exchangeable proton
not observed; EI-MS m/z 274 (M + H).
1-(4-Cya n op h en ylsu lfon yl)-4-(1-(4-p yr id yl)p ip er id in -4-
ylca r bon yl)p ip er a zin e (14). 28 (0.411 g, 1.5 mmol) was
dissolved in dry CH2Cl2 (20 mL) and stirred at 0 °C. Et3N (0.56
mL, 4.0 mmol) was added to the resulting solution followed
by the dropwise addition of a solution of 4-cyanobenzenesulfo-
nyl chloride (0.33 g, 1.6 mmol) in dry pyridine (20 mL). The
reaction mixture was stirred at 0 °C for 10 min and allowed
to warm to room temperature and stirred for a further 1 h.
The CH2Cl2 and pyridine were evaporated, the residue dis-
solved in H2O (60 mL) and the solution extracted with EtOAc
(3 × 50 mL). The organic extracts were washed with H2O,
brine, dried and evaporated to dryness. The product was
recrystallized from isohexane/EtOAc to give, as a colorless,
hygroscopic solid, 14 (512 mg, 78%): mp 168-169 °C; 1H NMR
(CDCl3) δ 1.83(m, 4H), 2.64(m, 1H), 2.88(m, 2H), 3.10(s, 4H),
3.68(bs, 4H), 3.88(m, 2H), 6.64(d, J ) 7.4 Hz, 2H), 7.86(d, J )
8.2 Hz, 2H), 8.03(d, J ) 7.4 Hz, 2H), 8.14(d, J ) 8.2 Hz, 2H).
Anal. (C22H25N5O3S‚0.5H2O) C, H, N.
OSC inhibitory activity (IC50 values in the nanomolar
and low-micromolar ranges). Our replacement of the
pyridine ring by pyrimidine (e.g. in 25 and 26) reduced
the inhibitor pKa to 6 and 7, respectively: i.e. in this
series compounds in a lower pKa range were also able
to give potent inhibition at the OSC enzyme site. Use
of information from the recently described21 photoaf-
finity labeling of OSC (aimed at locating binding sites
for 6a , Ro 48-8071) should allow a better definition of
binding sites in this series. Hence the relevance of
inhibitor pKa to OSC inhibitor binding should be clari-
fied.
Con clu sion s
A novel series of 4-piperidinopyridines and 4-piperi-
dinopyrimidines was synthesized, which afforded potent
inhibitors of both rat and human 2,3-oxidosqualene
cyclase-lanosterol synthase, and was shown in rats to
give (after oral dosing) selective inhibition of cholesterol
biosynthesis at the OSC step. This oral inhibition of rat
cholesterol biosynthesis for compounds 25 and 26 was
comparable with that found for the clinically used
HMGCoA reductase inhibitor simvastatin and the cited
standard orally active lipophilic amine inhibitors of
OSC. In line with the wide pKa range of known OSC
inhibitors, the piperidinopyrimidine inhibitors had a
significantly lower pKa than the corresponding pyridine
series or the previously reported quinuclidine OSC
inhibitor series. This indicated that further novel OSC
inhibitors may be found in either series by exploring a
broader range of inhibitor pKa (5.0-9.0). A more de-
tailed pharmacological and toxicological investigation
of the pyrimidines 25 and 26 may afford novel hypoc-
holesterolemic agents for the treatment of cardiovas-
cular disease.
4-(4-Br om op h en ylm eth yl)-1-(1-(4-p yr id yl)p ip er id in -4-
ylca r bon yl)p ip er a zin e (18). 28 (0.722 g, 2.64 mmol) was
dissolved in dry DMF (22 mL) and treated with NaH (0.19 g,
50% dispersion in mineral oil, 4.0 mmol) under an argon
atmosphere. The resultant mixture was allowed to stir for 30
min before the addition of 4-bromobenzyl bromide (0.66 g, 2.64
mmol). The reaction was stirred at room temperature for 2 h
and quenched by pouring into H2O, basifying with saturated
NaHCO3 solution and extracting with Et2O. The combined
organic extracts were washed with H2O, brine, dried and
evaporated to afford a cream solid which was recrystallized
twice from EtOAc/isohexane to give, as a colorless solid, 18
1
(945 mg, 83%): mp 148-149 °C; H NMR (DMSO-d6) δ 1.58-
(m, 4H), 2.28(m, 4H), 2.92(t, J ) 12.0 Hz, 3H), 3.48(m, 4H),
3.89(d, J ) 13.0 Hz, 2H), 6.75(d, J ) 6.4 Hz, 2H), 7.26(d, J )
8.2 Hz, 2H); 7.46(d, J ) 8,2 Hz, 2H), 8.13(d, J ) 6.4 Hz, 2H);
EI-MS m/z 430 (M + H). Anal. (C21H27BrN4O) C, H, N.
Exp er im en ta l Section
A. Ch em istr y. Melting points were determined with a
Buchi apparatus and are uncorrected. The 1H NMR spectra
were determined with a Bruker AM (200 MHZ) spectrometer
4-(4-Br om oben zoyl)-1-(1-(4-p yr id yl)p ip er id in -4-ylca r -
bon yl)p ip er a zin e (19). Compound 19 was prepared in a

4970 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 26
Brown et al.
manner similar to 14 but using 4-bromobenzoyl chloride as
starting material to give, as a colorless solid, 19 (74%): mp
214-215 °C; H NMR (DMSO-d6) δ 1.60(m, 4H), 2.86(t, J )
12.5 Hz, 3H), 3.50(m, 8H), 3.92(d, J ) 12.5 Hz, 2H), 6.80(d, J
) 7.4 Hz, 2H), 7.37(d, J ) 8.3 Hz, 2H), 7.61(d, J ) 8.3 Hz,
2H), 8.13(d, J ) 7.4 Hz, 2H); EI-MS m/z 457 (M + H). Anal.
(C22H25BrN4O2) C, H, N.
5-chloro-3-methyl-1,2,4-thiadiazole, to give, as a colorless solid
after crystallization from EtOH, 22 (64%): mp 173-174 °C;
1H NMR (CDCl3) δ 1.73(m, 2H), 1.85(m, 2H), 2.39(s, 3H), 3.02-
(m, 4H), 3.12(t, J ) 11.7 Hz, 2H), 3.27(m, 1H), 3.65(m, 4H),
3.92(d, J ) 11.7 Hz, 2H), 7.58(d, J ) 7.9 Hz, 2H), 7.68(d, J )
7.9 Hz, 2H); EI-MS m/z 514 (M + H). Anal. (C19H24BrN5O3S2)
C, H, N.
1
Exa m p les of Syn th etic Rou te B. 1-(4-Iod op h en ylsu l-
fon yl)-4-(1-(4-p yr id yl)p ip e r id in -4-ylca r b on yl)p ip e r a -
zin e (7). Thionyl chloride (0.21 mL, 2.86 mmol) was added to
a suspension of 27 (0.29 g, 1.42 mmol) in CH2Cl2 (5 mL) and
the mixture stirred for 0.5 h. The solvent was evaporated to a
give a solid, which was suspended in CH2Cl2 (5 mL) and added
to a suspension of N-4-iodophenylsulfonylpiperazine (0.5 g,
1.42 mmol) in CH2Cl2 (5 mL) and Et3N (1.4 mL, 10 mmol).
The mixture was stirred for 18 h and the organic phase washed
with H2O, dried and evaporated to give a solid. The solid was
purified by chromatography on silica gel in 5% MeOH/CH2Cl2
to give, as a colorless solid, 7 (0.235 g, 31%): mp 211-213 °C;
1H NMR (DMSO-d6) δ 1.52(m, 4H), 2.82(m, 1H), 2.95(m, 6H),
3.55(m, 4H), 3.92(d, J ) 13.1 Hz, 2H), 6.75(d, J ) 5.7 Hz, 2H),
7.48(d, J ) 8.3 Hz, 2H), 8.04(d, J ) 8.3 Hz, 2H), 8.11(d, J )
3.8 Hz, 2H); EI-MS m/z 541 (M + H). Anal. (C21H25IN4O3S) C,
H, N.
1-(4-P yr im id in yl)-4-(1-(4-br om op h en ylsu lfon yl)p ip er -
a zin -4-ylca r bon yl)p ip er id in e (25). 31 (170 mg, 0.4 mmol)
and 4-chloropyrimidine‚2HCl (78 mg. 0.41 mmol) in absolute
EtOH (10 mL) and Et3N (0.5 mL, 3.5 mmol) were heated under
reflux for 2 h. The solution was evaporated under vacuum and
H2O (50 mL) added. The aqueous mixture was extracted with
EtOAc (2 × 50 mL), washed with water, brine and dried. The
EtOAc was evaporated under vacuum to give an oil which was
dissolved in EtOAc and purified by flash column chromatog-
raphy on neutral alumina (ICN Alumina N 32-63) eluting with
an increasing concentration of MeOH/EtOAc (0-10%). The
resulting solid was recrystallized from a mixture of EtOAc/
THF/isohexane and then from MeCN to give, as a solid, 25
1
(155 mg, 75%): mp 197-198 °C; H NMR (DMSO-d6) δ 1.80-
(m, 4H), 2.70(m, 1H), 3.05(m, 6H), 3.65(bs, 4H), 4.40(m, 2H),
6.50(d, J ) 6.4 Hz, 1H), 7.65(d, J ) 8.0 Hz, 2H), 7.76(d, J )
8.0 Hz, 2H), 8.20(d, J ) 6.4 Hz, 1H), 8.60(s, 1H); EI-MS m/z
494 (M + H). Anal. (C20H24BrN5O3S) C, H, N.
4-(1-(4-Br om op h en ylsu lfon yl)p ip er a zin -4-ylca r bon yl)-
p ip er id in e (31). 2918 (1.30 g, 4 mmol) was added to 4-(4-
bromobenzenesulfonyl)piperazine (1.23 g, 4 mmol) in CH2Cl2
(50 mL) and the mixture stirred for 40 h. The CH2Cl2 solution
was washed with H2O, dried and evaporated to a foam which
crystallized on trituration with EtOAc. The solid was purified
by chromatography on a silica Bond Elut column, eluting with
2% MeOH/ CH2Cl2 to give, as a colorless solid, 30 (1.54 g,
1-(2-Meth yl-4-p yr im id in yl)-4-(1-(4-br om op h en ylsu lfo-
n yl)p ip er a zin -4-ylca r bon yl)p ip er id in e (26). 4-Chloro-2-
methylpyrimidine (135 mg, 1.07 mmol) was added to a solution
of 31 (415 mg, 1.0 mmol) in THF (15 mL) containing Et3N (0.2
mL, 1.5 mmol). The mixture was heated at reflux for 16 h,
allowed to cool and the THF evaporated. The residue was
treated with H2O (20 mL) and the aqueous phase extracted
with EtOAc (3 × 20 mL). The combined organic phases were
washed with saturated brine (1 × 20 mL) dried and evaporated
to give an oil which was purified by column chromatography
on silica gel. Elution with CH2Cl2/MeOH/0.88 NH4OH (96:3:
1) gave an oil on evaporation. Trituration with Et2O (10 mL)
gave, as a colorless solid, 26 (152 mg, 30%): mp 200-202 °C;
1H NMR (CDCl3) δ 1.70(m, 4H), 2.50(s, 3H), 2.67(m, 1H), 2.94-
(m, 2H), 3.05(m, 4H), 3.68(m, 4H), 4.41(d, J ) 12.4 Hz, 2H),
6.30(d, J ) 6.1 Hz, 1H), 7.62(d, J ) 8.3 Hz, 2H), 7.70(d, J )
8.3 Hz, 2H), 8.10(d, J ) 6.1 Hz, 1H); EI-MS m/z 508 (M + H).
Anal. (C21H26BrN5O3S) C, H, N.
Exa m p les of Syn th etic Rou te C. 4-(N-Boc-p ip er a zin -
4-ylca r bon yl)p ip er id in e (34). Na2CO3 (4.52 g, 42.6 mmol)
was stirred with isonipecotic acid (5.0 g, 38.7 mmol) in H2O
(80 mL) and benzyl chloroformate (6.82 g, 5.71 mL, 40 mmol)
added. The mixture was stirred for 18 h and acidified with 2
M HCl, before extraction with EtOAc. The organic phase was
washed with H2O, dried and evaporated to an oil. The oil was
purified by column chromatography on silica gel in CH2Cl2,
eluting with 5% MeOH/CH2Cl2 to give after evaporation, as a
colorless oil 32 (4.45 g, 44%): 1H NMR (CDCl3) δ 1.67(m, 2H),
1.92(m, 2H), 2.51(m, 1H), 2.96(m, 2H), 4.09(m, 2H), 5.13(s, 2H),
7.33(m, 5H),), exchangeable protons not observed; EI-MS m/z
264 (M + H).
1
75%): mp 209-210 °C; H NMR (CDCl3) δ 1.34(s, 9H), 1.65-
(m, 4H), 2.51(m, 1H), 2.72(m, 2H), 3.03(t, 4H), 3.63(m, 4H),
4.10(d, 2H), 7.58(d, 2H), 7.68(d, 2H); EI-MS m/z 516 (M + H).
30 (1.53 g, 2.96 mmol) was stirred in TFA (10 mL, 130 mmol)
for 1 h, and the TFA evaporated. H2O (25 mL) was added and
the mixture made basic with excess of Na2CO3 to pH 11. The
aqueous was extracted with EtOAc and the organic phase
washed with brine, dried and evaporated to give, as a colorless
1
solid, 31 (693 mg, 57%): mp 158-162 °C; H NMR (DMSO-
d6) δ 1.43(m, 4H), 2.47(m, 2H), 2.62(m, 1H), 2.90(m, 6H), 3.53-
(m, 4H), 7.65(d, 2H), 7.87(d, 2H), exchangeable protons not
observed; EI-MS m/z 416 (M + H). Anal. (C16H22BrN3O3S‚
0.75H2O) C, H, N.
1-(2-Oxazolin yl)-4-(1-(4-br om oph en ylsu lfon yl)piper azin -
4-ylca r bon yl)p ip er id in e (20). 31 (1.01 g, 2.4 mmol) was
dissolved in dry THF (50 mL) and 2-chloroethyl isocyanate
(0.225 mL, 2.6 mmol) was added. The reaction mixture was
stirred for 2 h. The solution was diluted with EtOAc, filtered
through a plug of neutral alumina and evaporated to dryness
to afford a colorless, waxy solid. This solid was dissolved in
H2O (100 mL) and heated to 100 °C for 0.5 h. The clear solution
was basified with excess of NH4OH (d ) 0.880) and extracted
with CH2Cl2 (3 × 30 mL). The extracts were washed with H2O,
brine and dried. The organic phase was evaporated to dryness
and the residue recrystallized from EtOAc/isohexane to give,
as a colorless solid, 20 (448 mg, 48%): mp 210-212 °C; 1H
NMR (DMSO-d6) δ 1.41(m, 4H), 2.70(m, 3H), 2.90(m, 4H), 3.55-
(m, 6H), 3.72(d, J ) 12.4 Hz, 2H), 4.18(t, J ) 7.9 Hz, 2H),
7.62(d, J ) 8.2 Hz, 2H), 7.83(d, J ) 8.2 Hz, 2H); EI-MS m/z
485 (M + H). Anal. (C19H25BrN4O4S) C, H, N.
1-Hydroxybenzotriazole (4.67 g, 34.5 mmol) and 1-(3-di-
methylaminopropyl)-3-ethylcarbodiimide hydrochloride (4.79
g, 25.3 mmol) were added to a solution of 32 (6.0 g, 23 mmol)
and Boc-piperazine (4.24 g, 23 mmol) in DMF (75 mL). The
mixture was stirred for 18 h and evaporated. Et2O was added
to the residue, and the organic layer washed with 2 M NaOH,
saturated citric acid and H2O. The organic phase was dried
and evaporated to give, as a colorless waxy solid, 33 (7.4 g,
76%): 1H NMR (CDCl3) δ 1.48(s, 9H), 1.73(m, 4H), 2.64(m,
1H), 2.87(m, 2H), 3.48(m, 8H), 4.23(d, 2H), 5.12(s, 2H), 7.33-
(m, 5H); EI-MS m/z 432 (M + H).
1-(2-Th ia zolin yl)-4-(1-(4-b r om op h en ylsu lfon yl)p ip er -
a zin -4-ylca r bon yl)p ip er id in e (21). Compound 21 was pre-
pared in a manner similar to 20 by using 2-chloroethyl
isothiocyanate as the starting material to give 21, as a colorless
1
solid, (71%): mp 208-210 °C; H NMR (DMSO-d6) δ 1.40(m,
2H), 1.49(m, 2H), 2.72(m, 1H), 2.95(m, 6H), 3.22(t, J ) 7.9 Hz,
2H), 3.52(m, 4H), 3.72(d, J ) 12.3 Hz, 2H), 3.82(t, J ) 7.9 Hz,
2H), 7.62(d, J ) 8.2 Hz, 2H), 7.83(d, J ) 8.2 Hz, 2H); EI-MS
m/z 501 (M + H). Anal. (C19H25BrN4O3S2) C, H, N.
1-(3-Meth yl-1,2,4-th ia d ia zin -5-yl)-4-(1-(4-br om op h en yl-
su lfon yl)p ip er a zin -4-ylca r b on yl)p ip er id in e (22). Com-
pound 22 was synthesized as for 25 below, but starting with
33 (3.71 g, 8.6 mmol) was hydrogenated over 5% Pd/C (120
mg) in EtOH (100 mL) at atmospheric pressure for 18 h. The
catalyst was filtered and the EtOH evaporated. Trituration
of the residue with Et2O gave, as a colorless solid, 34 (1.81 g,
71%): 1H NMR (CDCl3) δ 1.47(s, 9H), 1.67(m, 4H), 2.64(m,
3H), 3.14(m, 2H), 3.50(m, 8H); EI-MS m/z 298 (M + H). Anal.
(C15H27N3O3) C, H, N.

Piperidinopyridine and -pyrimidine Inhibitors of OSC
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 26 4971
1-(2-P yr im id in yl)-4-(1-(4-br om op h en ylsu lfon yl)p ip er -
a zin -4-ylca r bon yl)p ip er id in e (23). 2-Chloropyrimidine (230
mg, 2.0 mmol) and Et3N (0.4 mL, 2.9 mmol) were added to a
solution of 34 (594 mg, 2.0 mmol) in EtOH (20 mL), and the
mixture heated under reflux for 2 h. The EtOH was evaporated
and the residue shaken with EtOAc (80 mL) and H2O (40 mL).
The organic phase was separated, washed with brine, dried
and evaporated. The residue was chromatographed on a silica
Bond Elut column in 10% EtOAc/isohexane eluting with 50%
EtOAc/isohexane to give, as a colorless solid, 35a (390 mg,
52%): mp 149-150 °C; 1H NMR (DMSO-d6) δ 1.35(m. 4H),
1.37(s, 9H), 2.94(m, 3H), 3.32(m, 4H), 3.50(m, 4H), 4.62(d, J
) 12.5 Hz, 2H), 5.46(t, J ) 5.5 Hz, 1H), 8.33(d, J ) 4.6 Hz,
2H); EI-MS m/z 376 (M + H). Anal. (C19H29N5O3) C, H, N.
A solution of TFA (0.7 mL 9.0 mmol) and 35a (355 mg, 0.9
mmol) in CH2Cl2 (20 mL) was stirred at 25 °C for 16 h. The
mixture was evaporated and triturated in EtOAc (25 mL) and
saturated NaHCO3 solution (15 mL). The EtOAc phase was
dried and evaporated to an oil, which solidified to give 35b
dosqualene in ethanol (1 mL, 0.65 mg/mL) was added to an
ethanolic solution (1 mL) of radiolabeled oxidosqualene (stock
solution 18 µL, 1 mCi/mL), and compounds were dissolved in
DMSO as follows: molecular weight/1000 × 7.875 ) mg/mL
required to give a 10-4 M stock solution. Dilutions were made
from the stock to give 10-5 M, 10-6 M, etc., concentrations of
test compound. Phosphate-buffered Tween (281 µL) was placed
in 5-mL disposable vials and compound solution (4 µL) added
and mixed well. Microsomes (14.6 µL) were added and the vials
preincubated for 10 min at 37 °C. Oxidosqualene mix (15 µL)
was added and the mixture incubated for a further 1 h. The
reaction was terminated by the addition of 16% KOH in 20%
aqueous ethanol (315 µL). The samples were heated at 80 °C
for 2 h, hexane (2 × 5 mL) was added, and the samples were
whirl-mixed for 10 s. The hexane phases were separated, blown
down with N2 and the residue was dissolved in CH3CN:i-PrOH
(300 µL, 4:1). The samples were chromatographed using a
Hichrom 3ODS1 column with an isocratic elution using CH3-
CN/i-PrOH (95:5, flow rate 1 mL/min). The output from the
UV detector was connected to a radiochemical detector to
visualize radiolabeled sterol intermediates. The reaction rate
was measured as the conversion of oxidosqualene to lanosterol,
and the effects of compounds were expressed as an inhibition
of this process. IC50 values were obtained using the Origin
curve-fitting program supplied by MicroCal Software Inc., with
a tight binding background.
In vivo a ssa y for in h ibition of ch olester ol biosyn th esis
in r a ts: Female rats (130 ( 20 g) were acclimatized to reverse
lighting and compounds orally dosed (n ) 2) in DMSO/HPMC
(hydroxypropylmethylcellulose). 1 h later tritiated mevalonate
(2.5 µCi) was administered, and after a further 1 h the rats
were sacrificed. A weighed piece of liver (ca 0.5 g) was
saponified in KOH/EtOH (2 mL, 1:9 w/v) at 80 °C for 2 h, and
the mixture diluted (2 mL) before extraction with isohexane
(5 mL). The solvent was evaporated under N2 at 40 °C and
the residue dissolved in i-PrOH/CH3CN (300 µL, 1:4); aliquots
of this solution (100 µL) were examined by HPLC (Spherisorb
column S3ODS1-1590; 10 cm × 4.6 mm; flow rate 1 mL/min;
eluants CH3CN/H2O 95:5, CH3CN/i-PrOH 95:5, CH3CN/i-
PrOH 80:20). The column eluates were monitored by an on-
line radiochemical detector. In this way an HPLC lipid profile
was obtained for each compound relating radioactivity counts
to elution times. Chromatographic peaks obtained at different
retention times were identified by comparison to the retention
time of standard intermediates in the cholesterol biosynthesis
pathway (e.g. oxidosqualene, 4.8 min; squalene, 7.0 min;
farnesyl pyrophosphate, 2.0 min; cholesterol, 13.1 min), so that
a selectivity profile could be obtained in addition to the extent
of the inhibition of the biosynthesis of cholesterol from peak
area.
1
(175 mg, 64%): mp 109-111 °C; H NMR (DMSO-d6) δ 1.45-
(m, 2H), 1.63(m, 2H), 2.63(m, 4H), 2.91(m, 3H), 3.37(m, 4H),
4.62(d, J ) 12.0 Hz, 2H), 6.55(t, J ) 4.4 Hz, 1H), 8.32(d, J )
4.4 Hz, 2H); EI-MS m/z 276 (M + H).
4-Bromophenylsulfonyl chloride (130 mg, 0.5 mmol) was
added to a solution of 35b (137.5 mg, 0.5 mmol) and Et3N (0.9
mL, 6.5 mmol) in THF (10 mL), and the mixture stirred for
18 h. The mixture was evaporated and EtOAc (15 mL) and
H2O (25 mL) added. The insoluble solid was collected and
combined with the dried and evaporated EtOAc phase to give,
as a colorless solid, 23 (206 mg, 83%): mp 211-213 °C; 1H
NMR (DMSO-d6) δ 1.38(m, 2H), 1.62(m, 2H), 2.90(m, 7H), 3.61-
(m, 4H), 4.61(d, J ) 12.0 Hz, 2H), 6.57(t, J ) 4.4 Hz, 1H),
7.66(d, J ) 8.4 Hz, 2H), 7.87(d, J ) 8.4 Hz, 2H), 8.31(d, J )
4.4 Hz, 2H); EI-MS m/z 494 (M + H). Anal. (C20H24BrN5O3S)
C, H, N.
1-(5-P yr id a zin yl)-4-(1-(4-br om op h en ylsu lfon yl)p ip er -
a zin -4-ylca r bon yl)p ip er id in e (24). Compound 24 was pre-
pared by the same method as 23, but 3,4,5-trichloropyridazine
was used in the reaction with 34 (Scheme 4) to give, as a
colorless solid, 35c (42%): mp 180-182 °C; 1H NMR (DMSO-
d6) δ 1.46(s, 9H), 1.81(m, 2H), 2.08(m, 2H), 2.76(m, 1H), 3.05-
(m, 2H), 3.51(m, 8H), 3.81(m, 2H), 8.68(s, 1H); EI-MS m/z 444
(M + H). Anal. (C19H27Cl2N5O3) C, H, N.
35c (360 mg, 0.75 mmol) and Et3N (0.2 mL, 2.2 mmol) were
hydrogenated in EtOH (40 mL) over 10% Pd/C (50 mg) at 1
atm for 16 h. The catalyst was filtered and the mixture
evaporated. The residue was partitioned between H2O and
CH2Cl2 and the organic phase dried and evaporated to give,
as a foam, 35d (264 mg, 93%): 1H NMR (DMSO-d6) δ 1.48(s,
9H), 1.92(m, 4H), 2.78(m, 1H), 3.05(m, 2H), 3.51(m, 8H), 3.93-
(m, 2H), 6.67(dd, J ) 6.3 Hz and J ) 3.3 Hz, 1H), 8.67(d, J )
6.3 Hz, 1H), 8.82(d, J ) 3.3 Hz, 1H); EI-MS m/z 376 (M + H).
TFA (1 mL, 12 mmol) was added to 35d (250 mg, 0.6 mmol)
in CH2Cl2 (5 mL), and after 2 h the mixture was evaporated
to give an oil. The oil was dissolved in saturated brine and
10M NaOH added. The mixture was extracted with CH2Cl2
and the solvent dried and evaporated to afford a gum 35e (93
mg, 52%): 1H NMR (CDCl3) δ 1.90(m, 4H), 2.77(m, 1H), 2.90-
(s, 4H), 3.02(m, 2H), 3.55(m, 4H), 3.95(m, 2H), 6.63(dd, J )
6.3 Hz and J ) 3.3 Hz, 1H), 8.67(d, J ) 6.3 Hz, 1H), 8.82(d, J
) 3.3 Hz, 1H); EI-MS m/z 276 (M + H).
4-Bromophenylsulfonyl chloride (78 mg, 0.3 mmol) was
added to a solution of 35e (84 mg, 0.3 mmol) and Et3N (0.063
mL, 0.45 mmol) in THF (4 mL) at ambient temperature, and
after 1 h THF was evaporated. The residue was partitioned
in H2O (10 mL) and EtOAc (10 mL), and the insoluble product
collected. The product was crystallized from MeOH to give,
as a cream solid, 24 (32 mg, 22%): mp 83-86 °C; EI-MS m/z
494 (M + H). Anal. (C20H24BrN5O3S) C, H, N.
Ack n ow led gm en t. We thank our colleagues A. W.
Faull and R. P. Walker for the original synthesis of
compounds 7-10.
Su p p or tin g In for m a tion Ava ila ble: Spectral data and
elemental analyses for compounds in Table 1. This material
is available free of charge via the Internet at http://pubs.ac-
s.org.
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mM phosphate buffer (100 mL). A solution of cold oxi-

4972 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 26
Brown et al.
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