Design and synthesis of hepatoselective, pyrrole-based HMG-CoA reductase inhibitors

Article abstract of DOI:10.1016/j.bmcl.2007.05.096

This manuscript describes the design and synthesis of a series of pyrrole-based inhibitors of HMG-CoA reductase for the treatment of hypercholesterolemia. Analogs were optimized using structure-based design and physical property considerations resulting i

Full text of DOI:10.1016/j.bmcl.2007.05.096

Bioorganic & Medicinal Chemistry Letters 17 (2007) 4538–4544
Design and synthesis of hepatoselective, pyrrole-based
HMG-CoA reductase inhibitors
Jeffrey A. Pfefferkorn,* Yuntao Song, Kuai-Lin Sun, Steven R. Miller,
Bharat K. Trivedi, Chulho Choi, Roderick J. Sorenson, Larry D. Bratton,
Paul C. Unangst, Scott D. Larsen, Toni-Jo Poel, Xue-Min Cheng,
Chitase Lee, Noe Erasga, Bruce Auerbach, Valerie Askew, Lisa Dillon,
Jeffrey C. Hanselman, Zhiwu Lin, Gina Lu, Andrew Robertson, Karl Olsen,
Thomas Mertz, Catherine Sekerke, Alexander Pavlovsky, Melissa S. Harris,
Graeme Bainbridge, Nicole Caspers, Huifen Chen and Matthias Eberstadt
Pfizer Global Research & Development, Michigan Laboratories, 2800 Plymouth Road, Ann Arbor, MI 48105, USA
Received 25 April 2007; revised 24 May 2007; accepted 30 May 2007
Available online 6 June 2007
Abstract—This manuscript describes the design and synthesis of a series of pyrrole-based inhibitors of HMG-CoA reductase for the
treatment of hypercholesterolemia. Analogs were optimized using structure-based design and physical property considerations
resulting in the identification of 44, a hepatoselective HMG-CoA reductase inhibitor with excellent acute and chronic efficacy in
a pre-clinical animal models.
Ó 2007 Elsevier Ltd. All rights reserved.
Coronary heart disease (CHD) is the leading cause of
death in the United States at an annual cost of more
than $150 billion.1 Given that hypercholesterolemia, or
elevated serum cholesterol, is a key risk factor for
CHD, substantial efforts have been undertaken to miti-
gate this condition.2 Currently, the standard of care for
treating hypercholesterolemia is the use of HMG-CoA
reductase inhibitors, also known as statins, which block
the rate-limiting step of cholesterol biosynthesis.3 As a
class, statins have proven remarkably safe and effective
for both primary prevention of coronary heart disease
and secondary prevention of coronary events.4
Education Program (NCEP-ATP-III) has modified its
treatment guidelines recommending that highest risk
CHD patients reach a treatment goal of LDL-C
<70 mg/dl, down from the previous treatment goal of
100 mg/dl for this patient group.6 Moreover, other
researchers have suggested that optimal LDL-C levels
to prevent atherosclerosis and CHD might be even low-
er, in the range of 50–70 mg/dl.7
Achieving such aggressive LDL-C reductions in patients
typically requires the use of high dose statins alone or in
combination with complimentary agents such as the
cholesterol absorption inhibitor ezetimibe.8 A potential
limitation of high dose statin therapy is statin-induced
myalgia, the muscle pain or weakness that sometimes
accompanies statin therapy.9 While the overall incidence
of myalgia is low (2–7% of patients in trials), the likeli-
hood of occurrence increases with drug dose, and it can
be a key factor in preventing patient compliance with a
treatment regimen.9
In recent years, results from several clinical trials
(TNT, MIRACAL, PROVE-IT, and others) have dem-
onstrated that increasingly aggressive LDL-C-lowering
therapy may offer additional protection against CHD
relative to earlier, less aggressive, treatment standards.5
In light of this evidence, the National Cholesterol
The mechanism of statin-induced myalgia is complex
but thought to involve, in part, inhibition of HMG-
CoA reductase in non-hepatic tissues (particularly
Keywords: HMG-CoA reductase inhibitor; Statin; Hypercholesterolemia.
*
Corresponding author. Tel.: +1 860 686 3421; fax: +1 860 715
4608; e-mail: jeffrey.a.pfefferkorn@pfizer.com
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.05.096

J. A. Pfefferkorn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4538–4544
Br O2N
4539
muscle) resulting in the disruption of the biosynthesis of
isoprenoid-derived biomolecules.9 Encouragingly, the
potential for statin-induced myalgia can be reduced by
targeting HMG-CoA reductase inhibitors to hepatic
tissues and limiting peripheral exposure.10 Moreover, it
has been demonstrated that the hepatoselectivity of a
given HMG-CoA reductase inhibitor is related to its
degree of lipophilicity.10,11 In general, lipophilic statins
tend to achieve higher levels of exposure in non-hepatic
peripheral tissues, whereas more hydrophilic statins tend
to be more hepatoselective. These differences in hepa-
toselectivity can be rationalized by the fact that
lipophilic statins passively and non-selectively diffuse
into both hepatocyte and non-hepatocyte cells, while
hydrophilic statins rely largely on active transport into
hepatocyte cells to exert their effects.10,12 In particular
the organic anion transporting polypeptide (OATP)
family of membrane transporters has been reported to
be important for the hepatic uptake of hydrophilic
statins such as rosuvastatin and pravastatin.10d,12
a
F
F
F
F
c
NO2
4
5
N
CHO
Bu
b
10
F
F
6
8
d
H
N
N
H
CO2Et
H
CO2Et
e
12
11
F
F
F
F
Given this precedent, we recently undertook a discovery
effort to identify novel, potent, and hepatoselective
HMG-CoA reductase inhibitors which might be useful in
helping patients reach increasingly aggressive LDL-C
reduction goals. We sought to accomplish this through
modification of the atorvastatin (1) template with a
focus on altering the central pyrrole heterocycle. These
efforts included replacement of the pyrrole with alterna-
tive 5-membered heterocycles as reported elsewhere13 as
well as the evaluation of an isomeric pyrrole core repre-
sented by prototype inhibitor 2 (Fig. 1). Comparison of
atorvastatin (1) and 2 illustrates the transposition of the
3,5-dihydroxyhexanoic acid side chain from the 1-posi-
tion (atorvastatin) to the 2-position of the central
pyrrole of 2. Herein, we describe the structure–activity
studies undertaken on inhibitor 2 aimed at optimizing
hepatoselectivity and efficacy.14
f
N
N
OHC
CO2Et
OHC
CO2H
g
F
F
F
F
13
14
Scheme 1. Synthesis of functionalized pyrrole 14. Reagents and
conditions: (a) AgNO2, Et2O, 0 °C, 12 h, 36%; (b) n-BuNH2, C6H6,
80 °C, 2 h, 100%; (c) AcOH, 25 °C, 12 h, 82%; (d) CNCH2CO2Et,
DBU, THF, 25 °C, 12 h, 38%; (e) KOH (powdered), i-PrI, DMSO,
25 °C, 1.5 h, 62%; (f) POCl3, DMF, dichloroethane, 80 °C, 3 h, 45%;
(g) NaOH, MeOH, 60 °C, 2 h, 100%.
Schemes 1 and 2 illustrate the synthesis of prototype
inhibitor 2 as a representative example of this series of
inhibitors. As shown, a Barton–Zard synthesis was uti-
lized for construction of the central pyrrole heterocy-
cle.15 Initially, 4-fluorobenzyl bromide (4) was treated
with silver nitrite to afford 1-fluoro-4-nitromethyl-benz-
ene (5). In parallel, 4-fluorobenzaldehyde (6) was
converted to imine 8 by treatment with n-butylamine.
Subsequent reaction of 5 and 8 in acetic acid afforded
nitro-olefin 10. To complete the pyrrole synthesis, inter-
mediate 10 was reacted with ethyl isocyanatoacetate in
the presence of DBU to generate pyrrole 11 in modest
yield. N-Alkylation of 11 was then accomplished using
i-propyl iodide and powdered KOH in DMSO to pro-
vide pyrrole 12, which was subjected to Vilsmeir–Haack
formylation to afford pyrrole carboxaldehyde 13. Final-
ly, the ester of 13 was saponified with aqueous NaOH to
generate carboxylic acid 14 as a key intermediate for
subsequent analog synthesis.
O
OH OH
CO2H
PhHN
N
1: atorvastatin
F
O
OH OH
N
CO2H
PhHN
R1
As outlined in Scheme 2, the amide functionality of this
class of inhibitors was installed by initial conversion of
carboxylic acid 14 to the corresponding acid chloride
15 via treatment with SOCl2. Subsequent reaction of
15 with aniline then afforded anilide 16. Installation of
the 3,5-dihydroxyhexanoic acid side chain was
2: R1 = F
3: R1 = H
A
B
F
Figure 1. Structures of atorvastatin (1) and isomeric HMG-CoA
reductase inhibitors 2 and 3.

4540
J. A. Pfefferkorn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4538–4544
of the terminal ester to provide compound 2 as its car-
boxylate sodium salt.
O
O
N
N
b
CHO
CHO
PhHN
R
Separately, the route outlined in Schemes 1 and 2 was
utilized, with appropriate substitution of alternative
building blocks, to prepare all other analogs described
in these studies.
F
a
F
F
F
16
14: R = OH
15: R = Cl
Lastly, during the course of this work, a second genera-
tion process chemistry synthesis of this class of inhibi-
tors was also developed to facilitate the preparative
scale synthesis of selected compounds for pre-clinical
efficacy and safety studies. These efforts have been
recently reported.18
O
OTBS
c
Ph3P
CO2Me
17
O
O
OR
N
CO2Me
PhHN
Structural biology. The design of analogs in this series
was enabled through structural biology studies begin-
ning with determination of the X-ray structure of inhib-
itor 2 bound in the active site of HMG-CoA reductase
(see Fig. 2).19 Not surprisingly, inhibitor 2 binds to
18: R = TBS
19: R = H
d
F
F
20
HMG-CoA in a mode similar to that of other statins
e
with its 3,5-dihydroxyhexanoic acid side chain engaged
in an extensive hydrogen bonding network with residues
including: Ser-684, Asp-690, Lys-691, Lys 692, Glu 559
and Asp-767. Like both atorvastatin and rosuvastatin,
the carbonyl of the amide motif of inhibitor 2 also
accepts a hydrogen bond from Ser-556, and the 4-flour-
ophenyl A-Ring of 2 engages in a p-stacking interaction
with Arg-590.
O
OH OH
N
CO2Me
PhHN
20
F
F
Considering the design of novel analogs, examination of
Figure 2 revealed that the phenyl ring of the anilide was
in a relatively accommodating, and partially solvent-ex-
posed, binding pocket. This suggested that modification
at this position might be tolerated and possibly provide
an opportunity to modulate inhibitor lipophilicity while
still maintaining potency in order to achieve the desired
balance between hepatoselectivity and efficacy. This
strategy guided our structure–activity optimization stud-
ies as described below.
f,g
O
OH OH
N
CO2Na
PhHN
2
F
F
Scheme 2. Synthesis of inhibitor 2. Reagents and conditions: (a)
SOCl2, 80 °C; (b) PhNH2, Et3N, THF, 25 °C, 12 h, 57% (two steps); (c)
toluene, 110 °C, 48 h, 66%; (d) HF (48% in H2O), MeCN, 0 °C, 2 h,
99%; (e) Et2B(OMe), NaBH4, AcOH (cat), THF, À78 °C, 2 h, 58%; (f)
10% Pd/C, H2, EtOH/THF (1:1), 25 °C, 3 h, 100%; (g) NaOH, EtOH,
1 h, 100%.
accomplished via a Wittig olefination reaction between
pyrrole carboxaldehyde 16 and stabilized phosphonium
yield 17 (prepared stereoselectively in eight steps from
diethyl 3-hydroxyglutarate according to the method of
Konoike and Araki16). In that event, 16 and 17 were
reacted at elevated temperature to afford 18 in 66% yield
as exclusively the E-isomer. The TBS-protected side-
chain hydroxyl of 18 was then liberated by treatment
with aqueous HF to provide b-hydroxy ketone 19 which
was subjected to a stereoselective syn-reduction with Et2-
BOMe and NaBH4 to produce syn-diol 20.17 Finally, the
synthesis was completed by hydrogenation of the side-
chain olefin over 10% Pd/C, followed by saponification
˚
Figure 2. X-ray crystal structure (2.1 A resolution) of inhibitor 2
bound to the active site of HMG-CoA reductase.

J. A. Pfefferkorn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4538–4544
4541
On a separate note, additional inspection of the X-ray
structure of Figure 2 revealed, that in its bound confor-
mation, the amide NH-hydrogen and the ortho-hydro-
gen of the phenyl A-ring of inhibitor 2 are in close
amine-primed hamsters. In this study, animals were or-
ally dosed with drug (10 mg/kg) once daily for 7 days.
Upon completion of treatment, the percent reduction
of serum LDL-C was determined by comparison to an
untreated control group. Analogs demonstrating prom-
ising single dose efficacy in this model were then evalu-
ated in a full dose–response study to enable an ED50
determination.
˚
proximity (1.9 A) to one another suggesting the possibil-
ity of creating an intramolecular connection to create
conformationally restricted analogs. This observation
is explored in greater detail in the following
manuscript.14b
As shown in Table 1, initial analog efforts focused on the
addition of substituents to the anilide ring system as
highlighted in Table 1. While the unsubstituted phenyl
analogs 2 or 3 had good acute in vivo efficacy, they lacked
sufficient selectivity against myocytes with IC50 = 145 nM
and IC50 = 229 nM, respectively. To improve this selec-
tivity, increasingly hydrophilic substituents were added
in an effort to reduce inhibitor lipophilicity. As shown,
analogs (22–28) bearing these polar substituents were less
lipophilic (clogD < 0) and maintained good activity
against HMG-CoA reductase. As expected these more
hydrophilic analogs also exhibited good hepatoselectivity
with myocyte IC50 > 500 nM. Unfortunately, achieving
this improved selectivity through the addition of polar
groups resulted in a concomitant decrease in acute
in vivo efficacy as shown. This reduced efficacy was pre-
sumably a result of reduced oral absorption due to low
membrane permeability. This observation highlighted a
key challenge: the balance of selectivity and efficacy.
Structure–activity studies. Structure–activity studies
(Tables 1–4) on this template focused on modification
of the amide moiety, since this region was the most syn-
thetically amenable and was anticipated to be tolerant to
modification based upon our structural biology studies.
All new analogs were evaluated in an HMG-CoA reduc-
tase enzyme inhibition assay.21 Additionally, the ability
of analogs to block cholesterol synthesis in both rat
hepatocyte and myocyte cell lines was evaluated, and
comparison of these two values was utilized as a mea-
surement of hepatoselectivity. As a guideline, inhibitors
with potent hepatocyte activity (IC50 < 10 nM) and
good hepatoselectivity [(myocyte IC50 > 500 nM) and
(myocyte/hepatocyte ratio preferably >1000)] were
selected for further evaluation in an acute animal effi-
cacy model. In this model, mice were initially dosed with
drug (1 mg/kg) and then, after 0.5 h, given an intraperi-
toneal injection of 14C sodium acetate. After 4.5 h, a
whole blood sample was obtained and analyzed for
14C cholesterol levels which were compared to untreated
control animals to determine percent inhibition of acute
cholesterol synthesis. Analogs exhibiting acute in vivo
activity comparable to or better than the benchmark
rosuvastatin (CrestorÒ) were subsequently evaluated in
a chronic LDL-lowering study performed in cholestyr-
The evaluation of a series of benzyl amides is high-
lighted in Table 2. Paralleling the anilide series, the
unsubstituted benzyl amide 29 had moderate acute effi-
cacy, but lacked sufficient hepatoselectivity (myocyte
IC50 = 44 nM). The addition of increasingly polar sub-
stituents again led to an improvement in selectivity while
at the same time generally reducing in vivo efficacy.
Table 1. Structure and biological activity of anilide analogs 2, 3, and 21–2821
OH OH
O
O
R2
R1
N
O-Na+
N
H
X
F
Compound
R1
R2
X
HMG-CoA
IC50 (nM)
Inhibition of cellular
cholesterol synthesis
Mouse acute inhibition
cholesterol synthesis
(1 mg/kg) [%]
clogD
(pH 7.4)
Hepatocyte
IC50 (nM)
Myocyte
IC50 (nM)
Simvastatin
Rosuvastatin
49
1.3
0.27
0.3
0.4
1.0
1.2
0.5
0.2
0.1
0.7
0.3
0.1
150
250
50
À45
À82
À54
À75
À57
À22
À48
À20
À45
À10
À32
NA
4.41
À2.63
0.40
3.1
6.6
12.4
1.8
0.7
0.4
0.3
0.3
1.4
2.9
0.3
21
2
OMe
H
H
F
H
H
F
145
229
524
626
549
684
1110
1790
12,600
0.31
0.24
3
H
OH
H
22
23
24
25
26
27
28
H
OH
F
À0.11
À0.43
À1.19
À0.73
À1.04
À0.88
À1.29
H
C(O)NMe2
H
F
H
H
H
H
H
C(O)NH2
C(O)NMe2
SO2NH2
CO2H
H
H
H
H
NA, not active.

4542
J. A. Pfefferkorn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4538–4544
Table 2. Structure and biological activity of benzyl amide analogs 29–3621
OH OH
O
O
N
O-Na+
N
R1
H
R2
F
Compound
R1
R2
HMG-CoA
IC50 (nM)
Inhibition of cellular
cholesterol synthesis
Mouse acute
inhibition cholesterol
synthesis (1 mg/kg) [%]
clogD (pH 7.4)
Hepatocyte
IC50 (nM)
Myocyte
IC50 (nM)
Simvastatin
Rosuvastatin
49
1.3
0.27
0.3
0.4
1.5
0.1
0.7
0.1
4.9
0.1
150
250
44
À45
À82
À46
À52
—
4.41
À2.63
1.77
3.1
0.8
0.8
0.7
1.8
0.2
1.2
1.2
2.2
29
30
31
32
33
34
35
36
H
H
H
OMe
CH2OMe
H
310
211
130
650
3100
6520
7920
1.68
1.51
H
OMe
—
1.68
1.21
H
C(O)NH2
CN
H
À47
À22
À38
À7
0.29
0.01
H
H
C(O)NMe2
CO2H
À0.27
Table 3. Structure and biological activity of heterocyclic amide analogs 37–4121
OH OH
O
O
N
R1
O-Na+
N
H
F
Compound
R1
HMG-CoA
IC50 (nM)
Inhibition of cellular
cholesterol synthesis
Mouse acute
inhibition cholesterol
synthesis (1 mg/kg) [%]
clogD (pH 7.4)
Hepatocyte
IC50 (nM)
Myocyte
IC50 (nM)
Simvastatin
Rosuvastatin
49
1.3
0.3
150
250
À45
À82
4.41
À2.63
3.1
29
37
38
39
40
41
0.8
1.5
0.8
0.6
2.6
3.4
0.3
1.5
0.1
0.9
0.5
0.5
44
1010
À46
À33
À39
À19
N/A
À11
1.77
1.16
N
N
S
N
472
0.28
N
3230
0.18
N
HN
10,400
10,400
0.33
H
N
À0.03
N

J. A. Pfefferkorn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4538–4544
Table 4. Structure and biological activity of alkyl amide analogs 42–4721
4543
O
OH OH
O
N
R1
O-Na+
N
R2
F
Compound
R1
R2
HMG-CoA
IC50 (nM)
Inhibition of cellular
cholesterol synthesis
Mouse acute inhibition
cholesterol synthesis
(1 mg/kg) [%]
Hamster 7-day
LDL-lowering
(10 mg/kg) [%]
clogD (pH 7.4)
Hepatocyte
IC50 (nM)
Myocyte
IC50 (nM)
Simvastatin
Rosuvastatin
49
1.3
0.27
0.6
1.2
0.7
0.9
1.6
2.0
150
250
À45
À82
À85
À83
À76
À85
À73
À70
—
4.41
À2.63
À0.48
À0.32
À0.21
À1.87
0.56
3.1
3.0
8.8
12
À25
À11
À12
À24
—
42
43
44
45
46
47
H
H
2450
5300
2270
1580
1930
2780
H
Me
Et
H
Me
H
Me
i-Pr
c-Pr
20
15
—
À24
H
3.8
0.05
In an effort to influence the selectivity/efficacy balance
without the use of polar substituents, we next evaluated
a series of heterocyclic amides as outlined in Table 3.
Encouragingly, nearly all of these analogs (37–41) exhib-
ited good hepatoselectivity; unfortunately, all com-
pounds had inferior acute efficacy relative to the
benchmark compounds.
evaluated these analogs in a 7-day LDL-lowering assay
conducted in cholestyramine-primed hamsters. Initially,
analogs were evaluated at a single oral dose of 10 mg/kg
and compared against the benchmark rosuvastatin at
the same dose. As highlighted in Table 4, compounds 44
and 47 exhibited comparable efficacy to rosuvastatin at
this dose. Metabolic stability studies (data not shown)
suggested that 44 was more metabolically robust than
47, and as such 44 was selected for a full dose–response
study in this hamster model. Doses of 0.1, 0.5, 1, 5, 10,
and 50 mg/kg of 44 were evaluated in separate animal
groups (n = 10 each) resulting in an ED50 = 3 mg/kg for
this inhibitor. For comparison purposes, rosuvastatin
was evaluated at a dose of 10 mg/kg in an additional ani-
mal group during this study. As illustrated, 44 and rosu-
vastatin exhibited similar efficacies at this dose (Fig. 3).
As shown in Table 4, we also evaluated whether or not
non-aromatic amides would afford the desired balance
of selectivity and efficacy. Interestingly, primary (42),
secondary alkyl (43, 44, 46, 47) and tertiary alkyl amides
(45) all demonstrated excellent hepatoselectivity with
myocyte IC50 > 1000 nM. Furthermore, all of the ana-
logs in this series exhibited significant acute efficacy rel-
ative to the benchmark standards.
In order to further characterize the therapeutic potential
of the primary and alkyl amides of Table 4 (42–47) we
In conclusion, we have described the design, synthesis,
and biological evaluation of a series of pyrrole-based
HMG-CoA reductase inhibitors based upon an isomeric
variation of the atorvastatin template. Through struc-
ture–activity studies, we identified a subset of alkyl
amide containing analogs that exhibited excellent hepa-
toselectivity as well as good acute and chronic efficacy in
animal models of dyslipidemia. Of particular interest
was inhibitor 44 which was 3200-fold selective for inhib-
iting cholesterol synthesis in hepatocytes as opposed to
myocytes, yet it maintained pre-clinical LDL-lowering
efficacy comparable to that of rosuvastatin. It is antici-
pated that additional pre-clinical characterization of this
compound will represent an important step in the iden-
tification of a next generation HMG-CoA reductase
inhibitor useful for helping patients to achieve increas-
ingly aggressive LDL-lowering goals.
20
10
0
-10
-20
-30
-40
44
ED50 = 3.0 mg/kg
Rosuvastatin
-50
0.01
0.1
1
10
100
Dose (mg/kg)
Figure 3. Effect of 44 on plasma LDL in hamsters dosed orally for 7
days. All groups were sacrificed 2 h post-dose on the final day of
dosing. Data represent means SEM of the percent change from the
control group (n = 10/group). Rosuvastatin was tested in a similar
manner at a dose of 10 mg/kg.
Acknowledgments
We acknowledge the technical contributions made by
Fang Sun, Loola Al-Kassim, Susan Holly, Cindy

4544
J. A. Pfefferkorn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 4538–4544
E.; Newton, R. S.; O’Brien, P.; Picard, J. A.; Roark, W.
H.; Sekerke, C. S.; Sliskovic, D. R.; Wilson, M. W.
J. Med. Chem. 1991, 34, 466; (b) Koga, T.; Shimada, Y.;
Kuroda, M.; Tsujita, Y.; Hasegawa, K.; Yamazaki, M.
Biochim. Biophys. Acta 1990, 1045, 115; (c) Balasubrama-
nian, N.; Brown, P. J.; Catt, J. D.; Han, W. T.; Parker, R.
A.; Sit, S. Y.; Wright, J. J. J. Med. Chem. 1989, 32, 2038;
(d) Joshi, H. N.; Fakes, M. G.; Serajuddin, A. M. Pharm.
Pharmacol. Commun. 1999, 5, 267.
Spessard, and Siradanahalli Guru. We also thank Barry
Finzel for his assistance with the X-ray structure.
References and notes
1. American Heart Association Statistics Committee and
Stroke Statistics Subcommittee, 2007 Update. Dallas, TX:
American Heart Association, 2007.
12. Hsiang, B.; Zhu, Y.; Wang, Z.; Wu, Y.; Sasseville, V.;
Yang, W.-P.; Kirchgessner, T. G. J. Biol. Chem. 1999, 274,
37161.
13. The use of other heterocyclic cores will be described in
separate, forthcoming reports.
14. (a) For additional studies on this pyrrole template, see:
Bratton, L. D.; Auerbach, B.; Choi, C.; Dillon, L.;
Hanselman, J. C.; Larsen, S. D.; Lu, G.; Olsen, K.;
Pfefferkorn, J. A.; Robertson, A.; Sekerke, C.; Trivedi, B.
K.; Unangst, P. C. Bioorg. Med. Chem. 2007, in press.
doi:10.1016/j.bmc.2007.05.031; (b) Pfefferkorn, J. A.;
Choi, C.; Song, Y.; Trivedi, B. K.; Larsen, S. D.; Askew,
V.; Dillon, L.; Hanselman, J. C.; Lin, Z.; Lu, G.;
Robertson, A.; Sekerke, C.; Auerbach, B.; Pavlovsky,
A.; Harris, M. S.; Bainbridge, G.; Caspers, N. Bioorg.
Med. Chem. Lett. 2007, in press. doi:10.1016/
j.bmcl.2007.05.097.
2. (a) LaRosa, J. C.; Grundy, S. M.; Walters, D. D.; Shear,
C.; Barter, P.; Fruchart, J.-C.; Gotto, A. M.; Greten, H.;
Kastelein, J. J. P.; Shepard, J.; Wenger, N. K. N. Engl. J.
Med. 2005, 352, 1425; (b) Schwartz, G.; Olsson, A. G.;
Ezekowitz, M. D.; Ganz, P.; Oliver, M. F.; Waters, D.;
Zeiher, A.; Chaitman, B. R.; Leslie, S.; Stern, T. JAMA
2001, 285, 1711; (c) Cannon, C. P.; Braunwald, E.;
McCabe, C. H.; Rader, D. J.; Rouleau, J. L.; Belder, R.;
Joyal, S. V.; Hill, K.; Pfeffer, M. A.; Skene, A. M. N. Engl.
J. Med. 2004, 350, 1495.
3. For a review, see: (a) McKenney, J. M. Clin. Cardiol. 2003,
26(Suppl. III), 32.
4. (a) Scandinavian Simvastatin Survival Study Investiga-
tors. Lancet, 1994, 344, 1383; (b) The long-term interven-
tion with pravastatin in ischemic disease (LIPID) study
group. N. Engl. J. Med. 1998, 339, 1349; (c) Shepard, J.;
Cobbe, S. M.; Ford, I.; Isles, C. G.; Lorimer, A. R.;
MacFarlane, P. W.; McKillop, J. H.; Pachard, C. J. N.
Engl. J. Med. 1995, 333, 1301; (d) LaRosa, J. C.; He, J.;
Vupputuri, S. JAMA 1999, 282, 2340.
15. Barton, D. H. R.; Zard, S. C. Chem. Commun. 1985, 1098.
16. Konoike, T.; Araki, Y. J. Org. Chem. 1994, 59, 7849.
17. Chen, K.-M.; Hardtmann, G. E.; Prasad, K.; Repic, O.;
Shapiro, M. J. Tetrahedron Lett. 1987, 28, 155.
5. (a) LaRosa, J. C.; Grundy, S. M.; Walters, D. D.; Shear,
C.; Barter, P.; Fruchart, J.-C.; Gotto, A. M.; Greten, H.;
Kastelein, J. J. P.; Shepard, J.; Wenger, N. K. N. Engl. J.
Med. 2005, 352, 1425; (b) Schwartz, G.; Olsson, A. G.;
Ezekowitz, M. D.; Ganz, P.; Oliver, M. F.; Waters, D.;
Zeiher, A.; Chaitman, B. R.; Leslie, S.; Stern, T. JAMA
2001, 285, 1711; (c) Cannon, C. P.; Braunwald, E.;
McCabe, C. H.; Rader, D. J.; Rouleau, J. L.; Belder, R.;
Joyal, S. V.; Hill, K.; Pfeffer, M. A.; Skene, A. M. N. Engl.
J. Med. 2004, 350, 1495.
6. Grundy, S. M.; Cleeman, J. I.; Merz, N. B.; Brewer, B.;
Clark, L. T.; Hunninghake, D. B.; Pasternak, R. C.;
Smith, S. C.; Stone, N. J. Circulation 2004, 110, 227.
7. O’Keefe, J. H.; Cordain, L.; Harris, W. H.; Moe, R. M.;
Vogel, R. J. Am. Coll. Cardiol. 2004, 43, 2142.
18. Pfefferkorn, J. A.; Bowles, D. M.; Kissel, W.; Bolyles, D.
C.; Choi, C.; Larsen, S. D.; Song, Y.; Sun, K.-L.; Miller,
S.; Trivedi, B. K. Tetrahedron, in press.
19. The structure of HMG-CoA reductase (HMGR) in
complex with statins has previously been reported (see
Ref. 20). For the purpose of this study a modified version
of the published clone was used. Briefly HMGR was
expressed as an N-terminal His 6 protein truncated from
amino acid 441–875 with the mutation M485I in the E. coli
strain BL21 STAR. Protein was purified via Ni-affinity
and gel filtration chromatography. HMGR was concen-
trated to 20 mg/ml then complexed with statins at a
concentration of 0.5mM and incubated at 4 °C for 1 h.
Crystals were prepared in hanging drops from a crystal-
lization solution of 28% PEG 4000, 0.1 mM Tris, pH 8.5,
0.2 M LiSO4, and 50 mM DTT at 20 °C. Diffraction
quality crystals (P21) were obtained after micro-seeding,
harvested after 1 week, and cryo protected in 25% ethylene
8. For a review, see Patel, T. N.; Shishehbor, M. H.; Bhatt,
D. L. Eur. Heart J. 2007, 28, 664.
9. (a) Brukert, E.; Hayem, G.; Dejager, S.; Yau, C.; Begaud,
B. Cardiovasc. Drugs Ther. 2005, 19, 403; (b) Baer, A. N.;
Wortmann, R. L. Curr. Opin. Rheumatol. 2007, 19, 67; (c)
Tiwari, A.; Bansal, V.; Chugh, A.; Mookhtiar, K. Expert
Opin. Drug Saf. 2006, 5, 651; (d) Athar, H.; Shah, A. R.;
Thompson, P. D. Future Lipidol. 2006, 1, 143; (e) Franc,
S.; Dejager, S.; Bruckert, E.; Chauvenet, M.; Giral, P.;
Turpin, G. Cardiovasc. Drugs Ther. 2003, 17, 459.
10. (a) Hamelin, B. A.; Turgeon, J. Trends Pharm Sci. 1998,
19, 26; (b) Rosenson, R. S.; Tangney, C. C. JAMA 1998,
279, 1643; See also (c) Schachter, M. Fundam. Clin.
Pharmacol. 2004, 19, 117; (d) Ho, R. H.; Tirona, R. G.;
Leake, B. F.; Glaeser, H.; Lee, W.; Lemke, C. J.; Wang,
Y.; Kim, R. B. Gastroenterology 2006, 130, 1793.
˚
glycol. Data were collected at 2 A resolution at the
Industrial Macromolecular Crystallography Association
Collaborative Access Team (IMCA-CAT) beam-line ID-
17 at the Advanced Photon Source (Argonne National
Laboratory). All data were processed and scaled using
HKL2000. Data for Figure 2 have been deposited at the
PDB under filename 2Q1L.
20. (a) Istvan, E. S.; Deisenhofer, J. Science 2001, 292, 1160;
(b) Istvan, E. Atherosclerosis Suppl. 2003, 4, 3.
21. In vitro IC50 values are reported as arithmetic means for
n P 2 independent measurements unless otherwise
noted. In vivo measurements are represented as the
percent change from untreated control group (n = 10
animals/group). For detailed assay protocols, see Ref.
14a.
11. (a) Roth, B. D.; Bocan, T. M.; Blankey, C. J.; Chucho-
lowski, A. W.; Creger, P. L.; Creswell, M. W.; Ferguson,