α-substituted malonester amides: Tools to define the relationship between ACAT inhibition and adrenal toxicity

Article abstract of DOI:10.1021/jm970560h

We prepared a series of α-substituted malonester amides that were evaluated for their ability to inhibit acyl-CoA:cholesterol O-acyl transferase activity in vitro and to lower plasma total cholesterol levels in a variety of cholesterol-fed animal models.

Full text of DOI:10.1021/jm970560h

682
J . Med. Chem. 1998, 41, 682-690
r-Su bstitu ted Ma lon ester Am id es: Tools To Defin e th e Rela tion sh ip betw een
ACAT In h ibition a n d Ad r en a l Toxicity
Drago R. Sliskovic,*,† J oseph A. Picard,† Patrick M. O’Brien,† Peggy Liao,† W. Howard Roark,† Bruce D. Roth,†
Maureen A. Anderson,‡ Sandra Bak Mueller,‡ Thomas M. A. Bocan,‡ Richard F. Bousley,‡
Katherine L. Hamelehle,‡ Reynold Homan,‡ J ames F. Reindel,§ Richard L. Stanfield,‡ Daniel Turluck,| and
Brian R. Krause‡
Parke-Davis Pharmaceutical Research Division, Warner-Lambert Company, 2800 Plymouth Road, Ann Arbor, Michigan 48105
Received August 20, 1997
We prepared a series of R-substituted malonester amides that were evaluated for their ability
to inhibit acyl-CoA:cholesterol O-acyl transferase activity in vitro and to lower plasma total
cholesterol levels in a variety of cholesterol-fed animal models. Compounds of this series were
also useful in examining the relationship between adrenal toxicity and ACAT inhibition. One
compound from this series, 9f, was a potent inhibitor of ACAT in both the microsomal and
cellular assays. It was also bioavailable as determined by both a bioassay and a HPLC-UV
assay. This compound was evaluated in both guinea pig and dog models of adrenal toxicity
and compared to tetrazole amide 15. In the most sensitive species, the dog, both of these
compounds achieved good plasma levels; however, compound 9f caused adrenal necrosis,
whereas compound 15 had no effect on the adrenal gland. This adds to the growing body of
evidence that the adrenal toxicity observed with ACAT inhibitors may not be mechanism related.
In tr od u ction
(2) with ethyl malonyl chloride (3), followed by hydroly-
sis and re-esterification with a desired alcohol.4 How-
ever, attempts to repeat this procedure for the synthesis
of certain R-substituted malonester amides failed. Thus,
attempted monohydrolysis of diethyl phenylmalonate
(4a ) with alcoholic base followed by acidification led to
decarboxylation to yield the acid (5a ).5 However, this
route was successful for compounds 7b,c and 14a ,b
(Scheme 1).
An alternative route (Scheme 2) to the majority of
target compounds avoided this problem in most cases.
Deprotonation of ethyl phenylacetate (6) with LDA at
-78 °C followed by quenching with an aryl isocyanate
gave the malonester amides (7). Hydrolysis, followed
by acidification with 1 M HCl gave the acids (8) which
were then esterified with a suitable alcohol, using DCC
in dichloromethane at 0 °C, to yield the target com-
pounds (9a -p ). Interestingly, hydrolyses of esters of
type 7 proved capricious, since decarboxylations oc-
curred on random occasions when Ar ) 2,4,6-trimeth-
oxyphenyl or the R-substituent was 2-pyridyl. To syn-
thesize these analogues, the formation of the desired
ester was the first step, thus avoiding the troublesome
alkaline hydrolysis, acidification, and re-esterification
(Scheme 3). Thus, the acids (10) were converted to the
esters (11) by esterification with a suitable alcohol using
DCC in dichloromethane at 0 °C. Deprotonation fol-
lowed by quenching with an aryl isocyanate gave the
desired compounds (12a -e).
Inhibition of ACAT activity in monocyte-macrophages
of the arterial wall is an area of intense interest in the
field of experimental atherosclerosis prevention, since
direct inhibition of arterial ACAT may reduce the
formation of macrophage-enriched fatty streaks and
contribute to plaque stabilization by direct lipid deple-
tion.1 Our continuing research has primarily focused
on ACAT inhibitors that possess the necessary struc-
tural features to achieve sufficient plasma levels of
active drug to inhibit the arterial enzyme.2 However,
issues surrounding the toxic effect of ACAT inhibitors
on the adrenal gland continue to be pervasive in this
field.3 There is still no conclusive data as to whether
the toxic effects on the adrenal gland are due to the
mechanism of action of these drugs. In addition, if these
toxicities are associated with ACAT inhibition, then the
goal of selectively inhibiting ACAT in the artery wall
(a desirable peripheral target) without inhibiting ACAT
in the adrenal gland (an undesirable peripheral target)
may be difficult one to achieve in a therapeutic agent.
In this paper, we describe the synthesis and structure-
activity relationships of a series of R-substituted mal-
onester amides and describe our efforts to address the
adrenal toxicity issues surrounding the identification of
the best candidates for long-term antiatherosclerotic
studies.
Ch em istr y
The unsubstituted malonester amides (1) were previ-
ously synthesized by the acylation of suitable anilines
Resu lts a n d Discu ssion
Biologica l Meth od s. Compounds prepared were
initially tested for ACAT inhibition in hepatic mi-
crosomes isolated from cholesterol-fed rats (LAI screen).6
Using murine IC-21 macrophages (obtained from the
American Type Culture Collection, Rockville, MD) as a
model system for a cell type found in arterial lesions,
* To whom correspondence and reprint requests should be ad-
dressed.
† Department of Medicinal Chemistry.
‡ Department of Vascular and Cardiac Diseases.
§ Department of Pathology and Experimental Toxicology.
| Department of Pharmacokinetics and Drug Metabolism.
S0022-2623(97)00560-8 CCC: $15.00 © 1998 American Chemical Society
Published on Web 02/05/1998

R-Substituted Malonester Amides
Sch em e 1. Method Aa
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 5 683
a
Reagents: (a) KOH, EtOH, then 1 M HCl; (b) C6H4-1,2-(COCl)2; (c) 2,6-diisopropylphenyl aniline, Et3N, CH2Cl2; (d) NaOH, MeOH
then 1 M HCl; (e) DCC, CH2Cl2, R3OH.
Sch em e 2. Method Ba
Sch em e 3. Method Ca
a
Reagents: (a) DCC, CH2Cl2, R3OH; (b) LDA, ArNCO.
diet and consisting of peanut oil (PO) (5.5%), cholesterol
(C) (1.5%), cholic acid (CA) (0.5%)] was assessed. The
TC concentration was then measured and reported as
a percent change from controls.7 This same diet was
also used in a chronic screen (PCC), with the rats
receiving both drug and diet simultaneously for 7 days.8
In a second chronic model (CPCC), hypercholesterolemia
was first established (1 week, PCC diet) and then the
compounds were coadministered with diet for 1 week.
Efficacy in this model is defined as (1) the ability to
reduce non-HDL-C (high-density lipoprotein-cholester-
ol), defined as the amount of cholesterol in apo B
containing lipoproteins, and (2) also the ability to
elevate the diet-induced low levels of HDL-C.7
a
Reagents: (a) LDA, ArNCO; (b) NaOH, MeOH/THF then 1 M
HCl; (c) DCC, CH2Cl2, R3OH.
selected compounds were also evaluated in a cellular
assay that measured the ability of the compounds to
inhibit ACAT in a cell type found in atherosclerotic
lesions.7 Acute in vivo activity (APCC) was measured
in rats (male Sprague-Dawley strain, 200-225 g,
Charles River, Portage, MI) dosed with the test com-
pound orally as suspensions in water containing car-
boxymethylcellulose (1.5%) and Tween-20 (0.2%). After
dosing, the ability of the compound to prevent the rise
in plasma total cholesterol (TC) after the consumption
of a high-fat, high-cholesterol meal [known as the PCC
Efficacy in cholesterol-fed dogs (CDOG) [female
Beagles, 6-9 kg (Marshall Labs, Northrose, NY)] was
also assessed. Oral dosing (capsule) of the test sub-
stance was begun when the plasma TC concentration

684 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 5
Sliskovic et al.
Ta ble 1. Physical and Biological Properties of the R-Substituted Malonester Amides
acute in vivo (%∆TC)d
compd
no.
in vitroc
mp (°C) IC50 (µM)
30
mg/kg
3
Ar
R1
Ph
R2
R3
method
formulab
mg/kg
7a
7b
7c
9a
9b
9c
9d
9e
9f
9g
9h
9i
9j
9k
9l
9m
9n
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
H
H
CH2CH3
CH2CH3
B
A
A
B
B
B
B
B
B
B
B
B
B
B
B
B
B
B
B
C
C
C
C
C
A
A
C23H29NO3
C17H24FNO3 116-119
175-177
>5
>1
>1
>1
-23
F
1
CH3
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
CH3 CH2CH3
(CH2)2CH3
C19H29NO3
C24H31NO3
C28H39NO3
C29H41NO3
C30H43NO3
C31H45NO3
C33H49NO3
C35H53NO3
C37H57NO3
C39H61NO3
C28H39NO3
C33H49NO3
C35H53NO3
C32H47NO6
C28H39NO3
C36H55NO3
C37H57NO3
C30H43NO6
C36H55NO3
126-128
148-150
108-110
120-122
124-126
104-105
102-104
99-101
89-91
-24**
-20*
e
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
(CH2)6CH3
(CH2)7CH3
(CH2)8CH3
(CH2)9CH3
(CH2)11CH3
(CH2)13CH3
(CH2)15CH3
(CH2)17CH3
CH(CH3)(CH2)4CH3
CH(CH3)(CH2)9CH3
CH(CH3)(CH2)11CH3
CH(CH3)(CH2)11CH3
C(CH3)2(CH2)3CH3
C(CH3)2(CH2)11CH3
C(CH3)2(CH2)13CH3
(CH2)11CH3
(CH2)11CH3
(CH2)11CH3
(CH2)11CH3
C(CH3)2(CH2)11CH3
(CH2)11CH3
0.050
-71***
-56***
-67***
-58***
-63***
-49***
-26
-47***
-46**
-53***
-27***
-29**
0.013
0.012
0.013
0.010
0.016
0.037
0.200
0.040
0.013
0.022
0.030
0.011
0.012
0.072
0.013
0.056
0.008
0.180
0.020
0.032
0.021
78-81
-25
104-105
102-104
94-97
-67***
-37***
-61***
-59***
-72***
-26*
-61***
-46***
-48***
-65***
f
2,4,6-(OMe)3Ph Ph
84-86
g
h
i
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
Ph
Ph
Ph
79-82
9o
9p
89-91
98-101
93-95
-37*
12a
12b
12c
12d
12e
14a
14b
2,4,6-(OMe)3Ph Ph
-51***
-21**
-43**
-9
-51***
2,4,6-i-Pr)3Ph
2,6-(i-Pr)2Ph
2,4,6-(i-Pr)3Ph
2,4,6-(OMe)3Ph Ph
2,6-(i-Pr)2Ph
2,6-(i-Pr)2Ph
Ph
124-126
2-Pya
C32H48N2O3 44-47
j
2-Pya
C35H54N2O3 97-99
C33H49NO6
52-54
NT
-36**
NT
F
CH3
C27H44FNO3 oil
C31H53NO3
k
CH3 CH(CH3)(CH2)11CH3
oil
a
b
2-Pyridyl. C, H, N analysis within 0.4% of theoretical values unless otherwise noted. c In vitro ACAT inhibition determined in rat
d
liver microsomes. Denotes percent change in total cholesterol (TC) in cholic acid (0.5%)-cholesterol (1.5%)-peanut oil (5.5%)-fed rats
(APCC screen). See ref 7 for complete protocol. Significantly different from control using the unpaired Student’s two-tailed t-test: *p <
0.05, **p < 0.01, ***p < 0.001. NT ) not tested. e C: calcd, 75.56; found, 76.88. N: calcd, 3.67; found, 4.14. f N: calcd, 2.59; found, 3.41.
g
h
i
j
k
N: calcd, 3.20; found, 4.52. N: calcd, 2.55; found, 2.99. N: calcd, 2.48; found, 3.32. C: calcd, 75.09; found, 74.63. C: calcd, 76.33;
found, 75.64.
plateaued at 300 mg/dL after initiating a PCC diet
(same composition as described above for the rats). This
is about 2-fold higher than the levels found in chow-fed
dogs. Efficacy was expressed as the percent change in
plasma TC before and after treatment.6,7
Resu lts a n d Discu ssion
Str u ctu r e-Activity Rela tion sh ip s. In previous
studies with a series of fatty acid anilides8 or the
unsubstituted malonester amides,4 the retention of
significant ACAT inhibitory activity depended upon the
incorporation of the 2,6-diisopropylphenyl or 2,4,6-
trimethoxyphenyl substitution pattern into the anilide
moiety. Thus, both of these substitution patterns (and
the 2,4,6-triisopropylphenyl pattern) were used during
the course of this study and it was shown that all three
anilides gave compounds that were potent inhibitors of
ACAT activity in vitro (Table 1). For example, com-
pounds 9f, 9l, 9m , and 12a showed comparable potency
in vitro and equivalent efficacy in the acute in vivo
model. However, compound 12b, although of compa-
rable potency, was much less efficacious in vivo, prob-
ably due to its very high lipophilicity.
Like the unsubstituted malonester amides,4 varying
the chain length of the alkyl esters, in general, led to
increased in vitro potency and in vivo efficacy. In the
unsubstituted malonester amides, the optimum chain
length was C12 (dodecyl); however, in the R-substituted
series the optimal length was any length between C8
(octyl, 9c) and C14 (tetradecyl, 9g). All of these com-
pounds (9c-g) showed both equivalent potency at
inhibiting ACAT activity in vitro and equivalent efficacy
in vivo at 30 mg/kg. Potency differences were noted in
vivo, since at the lower dose of 3 mg/kg, the more
Oral bioavailability was assessed using two methods.
Rabbits (Male New Zealand Whites, 1.2-1.5 kg, Kuipers
Farms, Gary, IN) were dosed (in the daily meal contain-
ing 3% peanut oil and 3% coconut oil) with the test
compound at 25 and 50 mg/kg. Blood samples were
obtained predose and at intervals up to 24 h. The
corresponding plasma samples were assayed, following
solvent extraction, by an HPLC-UV assay. A similar
assay was developed for chow-fed dogs. The limit of
quantitation for both assays was 20 ng/mL. A rabbit
bioassay (ABIO) assessed oral bioactivity by incubating
extracts of plasma, from animals dosed with the inhibi-
tors, with rat liver microsomes. The ability of the
extract to inhibit the ACAT reaction was assessed, and
the data are expressed as relative percent inhibition.9
Adrenal toxicity was evaluated in both chow-fed guinea
pigs (male Hartley strain, 500-600 g, Charles River,
Portage, MI) and dogs. For guinea pigs, the drugs were
dissolved or suspended in oleic acid to facilitate absorp-
tion and dosed at 100 mg/kg for 2 weeks. Toxicity was
defined by the incidence, severity, and complexity of
adrenal histopathologic alterations in the zona fascicu-
lata.10 For dogs, the drug was administered as bulk
drug in capsule at a dose of 100 mg/kg for 2 weeks.11

R-Substituted Malonester Amides
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 5 685
Ta ble 3. Biological Activity Summary for Compound 9f
Ta ble 2. Efficacy Data for Selected Compounds in the Chronic
Cholesterol-Fed Rat Modela
test
9f
compd no.
dose (mg/kg)
%∆ non HDL-C
%∆ HDL-C
LAI (IC50, µM)
0.01
0.02
9b
9e
9f
9g
9j
9k
9l
9m
12a
10
10
30
30
10
30
30
30
30
-21
21
2
-5
148***
28
MAI2 (IC50, µM)
APCC (%∆ TC)a,c
-6330, -293
-80***
-57**
-18
-64**
-75***
-85***
-71*
PCC-OA (%∆ non HDL-C)a,c
PCC-CMC (%∆ non HDL-C)a,c
CPCC (%∆ non HDL-C)a,c
CDOG (%∆ TC)a,c
-541 (+29% ∆ HDL)
-541 (+23% ∆ HDL)
-8030 (+148% ∆ HDL)
-4310
-1
97***
215***
272***
127**
plasma levels (ng/mL)a,b
ABIO (% inhibition)a,b
69-13125, 75-14050
9925, 10050
a
b
Dose is superscript (mg/kg). Determined in rabbits. c Unless
a
For experimental details, see ref 6. Significantly different from
control using the unpaired two-tailed t-test: *p < 0.05, **p < 0.01,
***p < 0.001.
otherwise noted, the changes were significantly different from
control, p < 0.05 using analysis of variance followed by Fisher’s
multiple range test.
lipophilic compounds 9f and 9g were less effective.
Although not as potent as compounds 9c-g, the heptyl
ester (9b) was the most effective compound in the acute
in vivo screen. Compounds outside of this range (C7-
C14) were less active both in vitro and especially in vivo
[e.g. C3 (9a ), C16 (9h ), and C18 (9j)]. Branching the alkyl
chain gave equivocal results. By comparing compounds
with the same number of carbon atoms in the alkyl ester
moiety, it was possible to show both an advantage for
branching (compare 9g to 9l) and a disadvantage for
branching (compare 9f to 9k ). Addition of another
methyl group, as in 9o, at the branchpoint of 9l had a
detrimental effect on in vivo activity. Interestingly, the
2-methyl-2-hexyl (9n ) and 2-heptyl (9j) esters, like the
1-heptyl ester (9b), were among the most effective
agents both in vitro and in vivo.
As shown in Table 3, compound 9f is a potent ACAT
inhibitor in both the microsomal LAI assay (IC50 ) 0.01
µM) and in the cellular MAI2 assay (IC50 ) 0.02 µM).
This latter observation is important in that it shows that
the compound can cross cell membranes and inhibit
ACAT activity in a cell type found in atherosclerotic
lesions. In addition to the in vivo activities described
previously, compound 9f was evaluated in a number of
other cholesterol-fed animal models. In the chronic
version (PCC) of the APCC screen, it lowers plasma non-
HDL-C to the same extent irrespective of whether an
oleic acid vehicle (PCC-OA) or an aqueous vehicle is
employed (PCC-CMC). This is a very important point
since, typically, such lipophilic inhibitors only show
efficacy when dosed in an oil vehicle.12 In addition, the
compound also produced cholesterol lowering (43% at
10 mg/kg) in the cholesterol-fed dog model. This model
reflects the situation faced in the clinic, where bulk drug
is dosed in soft gelatin capsules. No emesis was noted
in this study.
Replacing the R-phenyl in 9f by the R-2-pyridyl (as
in 12c) or the R-fluoro (as in 14a ) led to significant
decreases in in vivo activity, despite potent in vitro
activity.
It was also determined if plasma drug levels capable
of inhibiting arterial ACAT can be achieved on oral
dosing. This was measured using two methods. The
first was a direct measurement, by an HPLC-UV assay,
of plasma levels achieved in the rabbit after oral dosing.
At doses of 25 and 50 mg/kg, plasma levels of 69-131
and 75-140 ng/mL were achieved, respectively. On
the basis of macrophage ACAT inhibition (MAI2) re-
sults (IC50 ) 20 nM at 10 ng/mL), there is more
than sufficient drug to inhibit arterial macrophage
ACAT. In addition, in the bioassay (ABIO) there was
almost complete inhibition of ACAT in this assay at the
doses tested (25 and 50 mg/kg), thus suggesting the
presence of active drug (or active metabolite) in the
plasma.
Ad r en otoxic Com p a r ison w ith th e r-Su bstitu ted
Tetr a zole Am id es. Compound 9f is a potent inhibitor
of ACAT and a potent hypocholesterolemic agent in two
rodent models of hypercholesterolemia, both acutely and
chronically. We also showed that this compound inhib-
ited cholesterol esterification in murine IC-21 macro-
phages and was bioactive in rabbits. The major hurdle
facing bioavailable ACAT inhibitors is adrenal toxicity.
We have shown that compound 9f is bioavailable and
capable of inhibiting ACAT in the tissue of interest
(macrophage in the artery wall); we must now deter-
mine that it is possible to have a beneficial effect in one
peripheral tissue (artery wall) without having a non-
beneficial effect in another peripheral tissue (adrenal
gland).
Several of the more active compounds in Table 1 were
then evaluated in the clinically relevant, chronic cho-
lesterol-fed rat model of preestablished hypercholester-
olemia (Table 2). In this model, the compounds with
the shorter alkyl ester chains (9b and 9e) were not as
effective as initially observed in the acute in vivo screen.
This observation was also noted for compound 9j, a
compound previously shown to possess a desirable pro-
file both in vitro and in in vivo. The lack of correlation
between the acute and chronic in vivo screens cannot
be explained at this time, although it may relate to the
ability of the compounds to reach the liver and prevent
the production of cholesteryl ester-rich lipoproteins. The
best activity in the chronic model was observed with
alkyl side chains longer than 10 carbon atoms. Com-
pound 9f was very effective, giving a 80% decrease in
plasma non-HDL-C and a 148% increase in HDL-C. The
2,4,6-trimethoxyphenyl analogue (12a ) was comparable
to 9f. Compound 9g, with the longer tetradecyl side
chain, had little effect on increasing plasma HDL levels.
In contrast to compound 9g, compounds with 14 carbon
atoms that were branched, showed good efficacy. Thus,
both compounds 9l and 9m lowered plasma non-HDL-C
by 75% and 85%, respectively, and elevated HDL-C by
215% and 272%, respectively. These compounds suf-
fered from the significant drawback of existing as a pair
of inseparable diastereomers.
After a consideration of all the biological and chemical
data, compound 9f was chosen as a model compound
for further pharmacological evaluation.

686 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 5
Sliskovic et al.
The key issue requiring resolution is whether the
adrenal toxicity observed with a number of ACAT
inhibitors is due to inhibition of cholesterol esterification
in that tissue (i.e., mechanism-based) or is unrelated
to enzyme inhibition. Some evidence suggests that the
adrenal toxicity observed is unrelated to ACAT inhibi-
tion. The first report of toxicity, in beagle dogs, with
an ACAT inhibitor (PD 132301-2) revealed the now
characteristic adrenal lesion consisting of adrenocortical
degeneration with necrosis in the zona fasciculata and
reticularis.13 Further mechanistic studies in guinea pig
adrenocortical cells in culture, however, suggested that
the observed cytotoxicity was not a result of ACAT
inhibition but of ATP depletion resulting from direct
inhibition of mitochondrial respiration.14 Shortly there-
after, reports of adrenal toxicity with other potent
bioavailable ACAT inhibitors appeared in the literature.
Thus, RP 73163 given to guinea pigs at doses of 100,
300, and 1000 mg/kg for 3 weeks produced dose-related
histopathologic changes in the adrenal cortex consisting
of diffuse coarse vacuolation of zona fasciculata cells
similar to that observed in dogs with PD 132301-2.
These authors also concluded that the adrenal toxicity
was not related to ACAT inhibition, since the pharma-
cologically inactive enantiomer of RP 73163 also caused
similar toxic effects.15 Further mechanistic studies
suggested that both RP 73163 and its enantiomer were
metabolized to a common product which was an inhibi-
tor of adrenal cholesterol 17R-reductase activity. The
authors speculated that this inhibition of cholesterol
17R-reductase activity may be the source of the coarse
vacuolation produced by RP 73163, although ACAT
inhibition could not be ruled out.16 It was recently
disclosed that FR 145237, a potent ACAT inhibitor
possessing direct antiatherosclerotic activity at the
artery wall,17 also produced adrenal toxicity in normal
rabbits after a single iv dose.18 Surprisingly, FR 145237
did not induce adrenal toxicity in low-density lipoprotein
receptor deficient WHHL rabbits, despite producing
adrenal drug concentrations equivalent to those found
in the normal rabbits. These authors speculated that,
because of the structural similarity between FR 145237
and PD 132301-2, the toxicity induced by FR 145237
may not be due to ACAT inhibition but to inhibition of
mitochondrial respiration. Another study in rabbits
demonstrated severe cellular damage with XP767 at
doses between 10 and 100 mg/kg; however, a structur-
ally related analogue, XR920, produced no adrenal
changes, despite having similar in vitro potency to
XP767.19 Long-term incubations of potent ACAT in-
hibitors (SaH 58-035 and CP-113,818) with cholesterol-
enriched mouse peritoneal macrophages led the authors
to conclude that ACAT inhibition in these cells increased
cell toxicity due to the build-up of intracellular free
cholesterol concentrations. This was supported by the
observations that cell toxicity paralleled the increase in
intracellular free cholesterol concentrations and that
removal of free cholesterol by the addition of extracel-
lular cholesterol “acceptors” or by blocking intracellular
sterol transport relieved the ACAT inhibitor-induced
toxicity. Thus, it was proposed that ACAT inhibition
in vivo could induce cell death by destabilization of the
plasma membrane upon cholesterol enrichment, unless
sufficient cholesterol acceptors were present.20
Despite these findings of adrenal toxicity with sys-
temically available ACAT inhibitors, the recent results
in ACAT-deficient transgenic mice would tend to sup-
port the position that the observed toxicity is not related
to ACAT inhibition, since these animals developed
normally and had no evidence of adrenal dysfunction
although their adrenal cholesteryl esters were markedly
reduced.21
The adrenal glands of guinea pigs given compound
9f at doses of 10-100 mg/kg had minimal to mild in-
creases in coarse vacuolation and reduced fine vacuola-
tion of zona fasciculata cortical cells. There was no evi-
dence of adrenal cortical necrosis at any dose. The
beagle dog has been reported to be the most sensitive
species for evaluating the adrenal effects of ACAT inhi-
bitors.22 Thus, compound 9f was evaluated in this
model and compared with the tetrazole amide 15, a
potent bioavailable ACAT inhibitor previously shown to
be nontoxic to the adrenal gland in guinea pigs.9
Adrenal toxicity was observed in dogs given compound
9f at 100 mg/kg for 3 weeks. This toxicity was charac-
terized by zonal degeneration and necrosis of zona
fasciculata cortical cells. There was also less wide-
spread degeneration and necrosis of zona reticularis
cortical cells. These changes often resulted in substan-
tial thinning and/or near complete collapse of these
zones. The inner portion of the zona reticularis at the
corticomedullary junction persisted. Collapsed stroma
and minor infiltrates of small mononuclear inflamma-
tory cells remained in regions of substantial cortical loss.
These changes were not observed in dogs administered
compound 15 at the same dose for the same duration of
treatment (Figure 1). Plasma drug blood levels for
compounds 9f and 15 were determined by HPLC. Both
compounds achieved blood levels far in excess of the IC50
necessary to inhibit macrophage ACAT. These levels
were highly variable and entirely consistent with the
very lipophilic nature of the compounds. For compounds
9f and 15, blood levels of 200-690 and 240-310 ng/mL
were achieved, respectively. Thus, even though both
compounds achieved blood levels adequate to inhibit
ACAT in the periphery, only one compound (9f) was
shown to be adrenotoxic.
Thus, we have demonstrated in the most sensitive
species (dog) that it is possible to identify potent ACAT
inhibitors, such as 15, that are bioavailable and not
adrenotoxic.
Con clu sion s
We have identified a series of R-substituted malon-
ester amides that were shown to be potent ACAT
inhibitors in vitro and hypocholesterolemic agents in
vivo. Compound 9f was shown to be a potent inhibitor
of both microsomal and cellular ACAT and was particu-
larly effective in a rodent model of preestablished
hypercholesterolemia. Despite its lipophilicity, com-

F igu r e 1. Adrenotoxic comparison between compounds 9f and 15: Adrenal cortex from an untreated dog (A) and dogs given compounds 9f (B) and 15 (C) at 100 mg/kg for 2 weeks.
Notice the collapse of the zona fasciculata and reticularis (between hash marks) in the 9f-treated dog (B). Collapsed stroma and islands of cortical cells (asterisks) remain in this
region. The adrenal cortex of the compound 15-treated dog is (C) comparable to the untreated dog. Zona glomerulosa (ZG) and adrenal medulla (M) are indicated. Normal and
collapsed zona fasciculata/reticularis is shown as an area between side hash marks. Hematoxylin and Eosin stain. Magnification of A, B, and C ) 95×.

688 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 5
Sliskovic et al.
yield 8 (Ar ) 2,6-diisopropylphenyl) (32.12 g, 82%) as a white
pound 9f, was readily absorbed and plasma levels,
capable of inhibiting ACAT in the periphery, were easily
attained. This compound was then used as a pharma-
cological tool to define the relationship between adrenal
toxicity and ACAT inhibition. Consequently, it was
shown to be toxic to the adrenal gland of beagle dogs.
In the same study, compound 15, a structurally unre-
lated potent ACAT inhibitor, achieved similar plasma
drug levels and was shown to be nontoxic to the adrenal
glands. This finding would tend to support the argu-
ment that adrenal toxicity is unrelated to inhibition of
ACAT activity.
1
solid: mp 201-203 °C; H NMR (CDCl3) δ 8.7 (s, 1H), 7.0-
7.5 (m, 8H), 4.60 (s, 1H), 2.75 (br m, 2H), 1.09 (s, 6H), 1.06 (s,
6H) ppm. Anal. (C21H25NO3‚0.5 H2O) C, H, N.
(c) (()-Ben zen ea cetic Acid , r-[[[2,6-Bis(1-m eth yleth yl)-
p h en yl]a m in o]ca r bon yl]-, Dod ecyl Ester (9f). To a dichlo-
romethane solution (1.3 L) of 8 (40 g, 0.1178 mol) was added
1-dodecanol (21.96 g, 0.1178 mol) at 0 °C under nitrogen with
stirring. To this was added dicyclohexylcarbodiimide (25.53
g, 0.124 mol). An immediate precipitate resulted and the
resulting suspension was allowed to warm to room-tempera-
ture overnight. This was then filtered and the solution was
concentrated in vacuo. The residue was flash chromato-
graphed (eluting with a gradient of 10-25% ethyl acetate/
hexane) to give 44.97 g (75%) of 9f as a white solid: mp 102-
104 °C; 1H NMR (CDCl3) δ 8.0 (s, 1H), 7.5 (d, 2H), 7.4 (m, 3H),
7.3 (tr, 1H), 7.1 (d, 2H), 4.70 (s, 1H), 4.2 (m, 2H), 2.90 (br m,
2H), 1.7 (m, 2H), 1.3 (s, 18H), 1.05 (s, 12H), 0.95 (tr, 3H) ppm.
Anal. (C33H49NO3) C, H, N.
Exp er im en ta l Section
Unless otherwise noted, materials were obtained from
commercial suppliers and used without further purification.
All organic extracts were dried over MgSO4 except when
otherwise noted. Column chromatography was performed on
Merck silica gel 60 (230-400) mesh). Melting points were
determined on a Thomas-Hoover melting point apparatus and
are uncorrected. Nuclear magnetic resonance spectra were
determined on either a Bruker 250 MHz, Varian XL 300 MHz,
or Varian Unity 400 MHz instrument. Chemical shifts (δ) are
expressed as parts per million (ppm) downfield from internal
tetramethylsilane. Elemental analyses for carbon, hydrogen,
and nitrogen were performed on a Perkin-Elmer 240C elemen-
tal analyzer and are within 0.4% of theory unless noted
otherwise. HPLC analyses of heparinized dog plasma samples
were analyzed with a Waters 600E HPLC equipped with a 717
plus autosampler and 490E programmable multiwavelength
Compounds 9a -e and 9g-p were synthesized according to
the procedures described in method B above and exemplified
for compound 9f; their physical and biological properties are
listed in Table 1.
Alter n a tive P r oced u r e for P r ep a r in g selected r-Su b-
st it u t ed Ma lon est er Am id es. Met h od C: Syn t h esis of
(()-Ben zen ea cetic Acid , r-[[(2,4,6-Tr im eth oxyp h en yl)-
a m in o]ca r bon yl]-1,1-d im eth yltr id ecyl Ester (12e). Ben -
zen ea cetic Acid , 1,1-Dim eth yltr id ecyl Ester (11, R1 ) P h ,
R3 ) C(CH3)2(CH2)11CH3). To a dichloromethane (100 mL)
solution of phenylacetic acid (3.5 g, 0.0213 mol) at 0 °C under
an inert nitrogen atmosphere with stirring was added 2-meth-
yl-2-tetradecanol (4.87 g, 0.0213 mol). After 10 min, dicyclo-
hexylcarbodiimide (4.84 g, 0.0234 mol) was added. An imme-
diate precipitate resulted and the resulting suspension was
allowed to warm to room-temperature overnight. This was
then filtered and the solution was concentrated in vacuo. The
residue was flash chromatographed (eluting with 5% ethyl
acetate/hexane) to give 2.0 g (27%) of the title compound as a
low melting solid: 1H NMR (CDCl3) δ 7.2-7.3 (m, 5H), 3.51
(s, 2H), 1.7 (m, 2H), 1.4 (s, 6H), 1.2 (m, 20H), 0.9 (tr, 3H) ppm.
Anal. (C23H38O2) C, H, N.
detector. The column used was a reverse phase Altima C18
5
µM column. The mobile phase used was 100% acetonitrile at
a flow rate of 1.0 mL/min with a detection wavelength of 214
nM.
Gen er a l P r oced u r e for P r ep a r in g r-Su bstitu ted Ma -
lon ester a m id es. Meth od B: Syn th esis of (()-Ben zen e-
a cetic Acid , r-[[[2,6-Bis(1-m eth yleth yl)p h en yl]a m in o]-
ca r bon yl]-, Dod ecyl Ester (9f). (a ) (()-Ben zen ea cetic
Acid, r-[[[2,6-Bis(1-m eth yleth yl)ph en yl]am in o]car bon yl]-
, Eth yl Ester (7a ). To a THF solution (40 mL) of diisopro-
pylamine (22.53 mL, 0.16 mol) at -40 °C under nitrogen
with stirring was added n-BuLi (100.5 mL, 1.6 M in hexanes,
0.1607 mol). After 10 min, the resulting LDA solution was
cooled to -78 °C and a solution of ethyl phenyl acetate (6,
24 g, 0.146 mol) in THF (150 mL) was added. A yellow
suspension resulted that was stirred at -78 °C for an ad-
ditional 30 min. To this was then added a THF (80 mL)
solution of 2,6-diisopropylphenyl isocyanate (31.24 g, 0.146
mol) dropwise. The resulting mixture was stirred at -78 °C
for 3 h before quenching with 1 N HCl (120 mL). Ethyl acetate
(300 mL) was added and the organic layer separated. This
was washed with water (2 × 200 mL), brine (200 mL), and
dried. Filtration, concentration in vacuo, and recrystallization
from ethyl acetate/hexane gave 7a (42.25 g, 78%) as a white
(()-Ben zen ea cetic Acid , r-[[(2,4,6-Tr im eth oxyp h en yl)-
a m in o]ca r bon yl]-1,1-d im eth yltr id ecyl Ester (12e). To a
THF solution (20 mL) of diisopropylamine (1.01 mL, 0.0072
mol) at -40 °C under an inert nitrogen atmosphere with
stirring was added n-BuLi (3.6 mL, 2.01 M in hexanes, 0.0072
mol). After 15 min, a THF solution (10 mL) of benzeneacetic
acid, 1,1-dimethyltridecyl ester (1.5 g, 0.00656 mol), was added
dropwise. This mixture was cooled to -78 °C for 30 min and
a THF solution (10 mL) of 2,4,6-trimethoxyphenyl isocyanate23
(1.37 g, 0.00656 mol) was added dropwise. After stirring for
3 h at -78 °C, the reaction was quenched by the addition of 1
N HCl (10 mL). The solution was warmed to room tempera-
ture and partitioned between ethyl acetate and water. The
organic layer was separated, washed with water (1 × 100 mL)
and brine (1 × 100 mL), and dried. After filtration and
concentration, flash chromatography (eluting with a gradient
of 25-50% ethyl acetate/hexane) gave 0.87 g (24%) of 12e as
a white solid: mp 52-54 °C; 1H NMR (CDCl3) δ 8.0 (br s, 1H),
7.2-7.7 (m, 5H), 6.1 (s, 6H), 4.6 (s, 1H), 3.8 (s, 9H), 1.7 (m,
1
solid: mp 175-177 °C, H NMR (CDCl3) δ 8.0 (s, 1H), 7.05-
2H), 1.4 (s, 6H), 1.2 (m, 20H), 0.9 (tr, 3H) ppm. Anal. (C33H49
NO6) C, H, N.
-
7.5 (m, 8H), 4.75 (s, 1H), 4.2-4.4 (m, 2H), 2.85 (br m, 2H), 1.3
(tr, 3H), 1.09 (s, 6H), 1.06 (s, 6H) ppm. Anal. (C23H29NO3) C,
H, N.
Compounds 12a -d were synthesized according to the
procedures described in method C above and exemplified for
compound 12e; their physical and biological properties are
listed in Table 1.
Alter n a tive P r oced u r e for P r ep a r in g selected r-Su b-
st it u t ed Ma lon est er Am id es. Met h od A: Syn t h esis of
(()-P r op a n oic Acid , 3-[[2,6-Bis(1-m eth yleth yl)p h en yl]-
a m in o]-2-flu or o-3-oxo-, Dod ecyl Ester (14a ). (a ) (()-
P r opan oic Acid, 3-[[2,6-Bis(1-m eth yleth yl)ph en yl]am in o]-
2-flu or o-3-oxo, Eth yl Ester (7b). Monoethyl fluoromalonate24
(10.37 g, 0.068 mol) and phthaloyl dichloride (13.84 mL, 0.096
mol) were heated together at 100 °C for 2.5 h. After cooling,
(b) (()-Ben zen ea cetic Acid , r-[[[2,6-Bis(1-m eth yleth -
yl)p h en yl]a m in o]ca r bon yl] (8, Ar ) 2,6-d iisop r op ylp h en -
yl). To a methanol (750 mL)/THF (100 mL) solution of 7a
(33.55 g, 0.0913 mol) was added 1 N aqueous NaOH (91.3 mL,
0.0913 mol). The resulting solution was stirred overnight at
room temperature. It was then concentrated in vacuo and the
resulting residue was redissolved in water (500 mL). This was
then washed with ethyl ether (2 × 200 mL) and the aqueous
layer was cooled in ice water and acidified with 1 N HCl. A
white solid precipitated, which was filtered and air-dried to

R-Substituted Malonester Amides
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 5 689
(4) Sliskovic, D. R.; Picard, J . A.; Roark, W. H.; Essenburg, A. D.;
Krause, B. R.; Minton, L. L.; Reindel, J . F.; Stanfield, R. L.
Inhibitors of Acyl-CoA:cholesterol O-acyl transferase (ACAT) as
hypocholesterolemic agents. The synthesis and biological activity
of a series of malonester amides. Bioorg. Med. Chem. Lett. 1996,
6, 713-718.
the resulting solid was distilled (70-80 °C/15 mmHg) to yield
the corresponding acid chloride (4.43 g, 44%), which was used
in the next step without any further purification.
To a dichloromethane solution (120 mL) of 2,6-diisopropyl-
aniline (2.97 mL, 0.0206 mol) and triethylamine (3.6 mL,
0.0257 mol) at 0 °C was added the acid chloride (3.93 g, 0.0257
mol), obtained in the previous step, under an inert nitrogen
atmosphere with stirring. The resulting red solution was
stirred at 0 °C for 1 h and then quenched by the addition of 1
N HCl (20 mL). The organic layer was separated, washed with
water (1 × 100 mL) and brine (1 × 100 mL), and dried. After
filtration and concentration, the residue was flash chromato-
graphed (eluting with 10% ethyl acetate/hexane) to yield 3.46
g (55%) of 7b as a white solid: mp 116-119 °C; 1H NMR
(5) Vliet, E. B.; Marvel, C. S.; Hsueh, C. M. 3-Methylpentanoic acid.
Org. Synth. 1943, 2, 416-417.
(6) Krause, B. R.; Black, A.; Bousley, R.; Essenburg, A. D.; Cornicelli,
J . A.; Holmes, A.; Homan, R.; Kieft, K. A.; Sekerke, C.; Shaw-
Hes, M. K.; Stanfield, R. L.; Trivedi, B. K.; Woolf, T. Divergent
Pharmacological Activities of PD 132301-2 and CL 277,082, Urea
Inhibitors of Acyl CoA:Cholesterol Acyl-Transferase. J . Phar-
macol. Exp. Ther. 1993, 267, 734-743.
(7) White, A. D.; Purchase II, C. F.; Picard, J . A.; Anderson, M. K.;
Bak Mueller, S.; Bocan, T. M. A.; Bousley, R. F.; Hamelehle, K.
L.; Krause, B. R.; Lee, P.; Stanfield, R. L.; Reindel, J . F.
Heterocyclic Amides: Inhibitors of Acyl-CoA:Cholesterol O-
Acyltransferase with hypocholesterolemic activity in several
species and antiatherosclerotic activity in the rabbit. J . Med.
Chem. 1996, 39, 3908-3919.
(8) Roth, B. D.; Blankley, C. J ., Hoefle, M. L.; Holmes, A.; Roark,
W. H.; Trivedi, B. K.; Essenburg, A. D.; Kieft, K. A.; Krause, B.
R.; Stanfield, R. L. Inhibitors of Acyl-CoA:Cholesterol Acyltrans-
ferase. 1. Identification and Structure-Activity Relationships
of a Novel Series of Fatty Acid Anilide Hypocholesterolemic
Agents. J . Med. Chem. 1992, 35, 1609-1617.
(9) O’Brien, P. M.; Sliskovic, D. R.; Picard, J . A.; Lee, H. T.; Purchase
II, C. F.; Roth, B. D.; White, A. D.; Anderson, M. K.; Bak Mueller,
S.; Bocan, T. M. A.; Bousley, R. F.; Hamelehle, K. L.; Homan,
R.; Lee, P.; Krause, B. R.; Lee, P.; Reindel, J . F.; Stanfield, R.
L.; Turluck, D. Inhibitors of Acyl-CoA:Cholesterol O-Acyltrans-
ferase. Synthesis and pharmacological activity of (()-2-do-
decyl-R-phenyl-N-(2,4,6-trimethoxyphenyl)-2H-tetrazole-5-aceta-
mide and structurally related tetrazole derivatives. J . Med.
Chem. 1996, 39, 2354-2366.
(CDCl3) δ 7.58 (br s, 1H), 7.1-7.4 (m, 3H), 5.2 (d, 1H, J H-F
)
49 Hz), 4.4 (q, 2H), 3.0 (hept, 2H), 1.4 (tr, 3H), 1.2 (m, 12H)
ppm. Anal. (C17H24FNO3) C, H, N.
(b ) (()-P r op a n oic Acid , 3-[[2,6-Bis(1-m et h ylet h yl)-
p h en yl]a m in o]-2-flu or o-3-oxo-, Dod ecyl Ester (14a ). To
a methanol (70 mL) solution of 7b (3.0 g, 0.0097 mol) was
added 1 N aqueous NaOH (9.7 mL, 0.0097 mol). The resulting
solution was stirred at room temperature for 2 h. It was then
concentrated in vacuo and the resulting residue was redis-
solved in water (50 mL). This was washed with ethyl acetate
(2 × 20 mL) and the aqueous layer was cooled in ice water
and acidified with 1 N HCl. This was extracted with ethyl
acetate and the resulting organic extract was dried. Filtration
and concentration gave the corresponding acid (2.5 g, 92%):
1
mp 139-141 °C; H NMR (CDCl3) δ 7.90 (br s, 1H), 7.2-7.4
(m, 3H), 5.5 (d, 1H, J H-F ) 47 Hz), 2.9 (hept, 2H), 1.2 (m, 12H)
ppm. Anal. (C15H20FNO3) C, H, N.
(10) Dominick, M. A.; Bobrowski, W. A.; MacDonald, J . R.; Gough,
A. W. Morphogenesis of a zone-specific adrenocortical cytotoxicity
in guinea pigs administered PD 132301-2, an inhibitor of acyl-
CoA:cholesterol O-acyltransferase. Toxicol. Pathol. 1993, 21, 54-
62.
(11) Wolfgang, G. H. I.; Robertson, D.G.; Welty, D. F.; Metz, A. L.
Hepatic and adrenal toxicity of a novel lipid regulator in beagle
dogs. Fundam. Appl. Toxicol. 1995, 26, 272-281.
(12) Trivedi, B. K.; Purchase, T. S.; Holmes, A.; Augelli-Szafran, C.
E.; Essenburg, A. D.; Hamelehle, K. L.; Stanfield, R. L.; Bousley,
R. F.; Krause, B. R. Inhibitors of Acyl-CoA:Cholesterol Acyl-
transferase (ACAT). 7. Development of a series of substituted
N-phenyl-N′-[1-phenylcyclopentyl)methyl]ureas with enhanced
hypocholesterolemic activity. J . Med. Chem. 1994, 37, 1652-
1659.
(13) Dominick, M. A.; McGuire, E. J .; Reindel, J . F.; Bobrowski, W.
A.; Bocan, T. M. A.; Gough, A. W. Subacute toxicity of a novel
inhibitor of acyl-CoA:cholesterol acyltransferase in beagle dogs.
Fundam. Appl. Toxicol. 1993, 20, 217-224.
To a dichloromethane solution (100 mL) of the acid obtained
above (0.7 g, 0.0025 mol) was added 1-dodecanol (0.56 mL,
0.0025 mol) at 0 °C under nitrogen with stirring. To this
was added dicyclohexylcarbodiimide (0.56 g, 0.0027 mol).
The resulting suspension was allowed to warm to room
temperature overnight. This was then filtered and the fil-
trate was partitioned between dichloromethane and water.
The organic layer was separated, washed with water (1 × 50
mL) and brine (1 × 50 mL), and dried. After filtration and
concentration, the residue was flash chromatographed (eluting
with a gradient of 10-25% ethyl acetate/hexane) to give
0.44 g (39%) of 14a as a viscous oil: 1H NMR (CDCl3) δ 7.6
(br s, 1H), 7.2-7.4 (m, 3H), 5.4 (d, 1H, J H-F ) 49 Hz), 4.2
(tr, 2H), 3.0 (hept, 2H), 1.7 (m, 2H), 1.3 (m, 30H), 0.9 (tr, 3H)
ppm. Anal. (C27H44FNO3) C, H, N. Compounds 7c and
14b were synthesized according to the procedures described
in method A above and exemplified for compounds 7b and
14a ; their physical and biological properties are listed in
Table 1.
(14) Vernetti, L. A.; MacDonald, J . R.; Wolfgang, G. H. I.; Dominick,
M. A.; Pegg, D. G. ATP Depletion is associated with cytotoxicity
of a novel lipid regulator in guinea pig adrenocortical cells.
Toxicol. Appl. Pharmacol. 1993, 118, 30-38.
(15) Riddell, D.; Bright, C. P.; Burton, B. J .; Bush, R. C.; Harris, N.
V.; Hele, D.; Moore, U. M.; Naik, K.; Parrott, D. P.; Smith, C.;
Williams, R. J . Hypolipidemic properties of a potent and bio-
available alkylsulphinyl-diphenylimidazole ACAT inhibitor (RP
73163) in animals fed diets low in cholesterol. Biochem. Phar-
macol. 1996, 52, 1177-1186.
Ack n ow led gm en t. We thank Dr. G. A. McCluskey
and staff for spectral and analytical determinations. We
also extend our gratitude to Walter Bobrowski for the
preparation of the photomicrographs.
(16) Albaladejo, V.; Catinot, R.; Moronvalle-Halley, V.; Kriszt, W.;
Schorsch, F.; Picaut, P.; Baron, H.; Ballet, F. RP 73163, A novel
systemic ACAT-inhibitor induces coarse vacuolation and impairs
steroid hormone metabolism in guinea pig adrenocortical cells.
Book of Abstracts-EUROTOX ‘94, p 3.
(17) Matsuo, M.; Ito, F.; Konto, A.; Aketa, M.; Tomoi, M.; Shimomura,
K. Effect of FR 145237, a novel ACAT inhibitor, on atherogenesis
in cholesterol-fed and WHHL rabbits. Evidence for a direct effect
on the arterial wall. Biochim. Biophys. Acta 1995, 1259, 254-
260.
(18) Matsuo, M.; Hashimoto, M.; Suzuki, J .; Iwanami, K.; Tomoi, M.;
Shimomura, K. Difference between normal and WHHL rabbits
in susceptibility to the adrenal toxicity of an acyl-CoA:cholesterol
acyltransferase inhibitor, FR 145237. Toxicol. Appl. Pharmacol.
1996, 140, 387-392.
(19) Wilde, R. G.; Billheimer, J . T.; Germain, S. J .; Hausner, E. A.;
Meunier, P. C.; Munzer, D. A.; Stoletenborg, J . K.; Gillies, P. J .;
Burcham, D. L.; Huang, S.-M.; Klaczkiewicz, J . D.; Ko, S. S.;
Wexler, R. R. ACAT inhibitors derived from hetero-Diels-Alder
cycloadducts of thioaldehydes. Bioorg. Med. Chem. 1996, 4,
1493-1513.
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