Discovery of chiral N,N-disubstituted trifluoro-3-amino-2-propanols as potent inhibitors of cholesteryl ester transfer protein [1]

Full text of DOI:10.1021/jm000337b

© Copyright 2000 by the American Chemical Society
Volume 43, Number 24
November 30, 2000
Communications to the Editor
for CETP polymorphism in the Framingham Offspring
Discover y of Ch ir a l N,N-Disu bstitu ted
Tr iflu or o-3-a m in o-2-p r op a n ols a s P oten t
In h ibitor s of Ch olester yl Ester Tr a n sfer
P r otein
Study identified a B2B2 allele in men associated with
significantly reduced CETP activity, elevated HDL-C,
and a reduced risk for CHD.10 Administration of TP-2,
a CETP-neutralizing monoclonal antibody, to hamsters
produced a dramatic reduction in CETP activity with a
concomitant elevation of HDL-C.11 Recently, a CETP
inhibitor, which binds to CETP via a disufide bond, has
been shown to raise HDL-C, lower LDL-C, and inhibit
the progression of atherosclerosis in rabbits.12 On the
basis of these observations, a CETP inhibitor is expected
to block the balanced exchange of CE and TG between
HDL and VLDL or LDL such that the LDL-C/HDL-C
ratio and risk of CHD are significantly reduced. Prefer-
ably, a CETP inhibitor would be specific for CETP and
not disrupt the integrity of other lipoproteins.
Richard C. Durley,* Margaret L. Grapperhaus,
Mark A. Massa, Deborah A. Mischke,
Barry L. Parnas, Yvette M. Fobian, Nigam P. Rath,†
Dorothy D. Honda, Ming Zeng, Daniel T. Connolly,
Deborah M. Heuvelman, Bryan J . Witherbee,
Kevin C. Glenn, Elaine S. Krul, Mark E. Smith, and
J ames A. Sikorski
Pharmacia Discovery Research,
700 Chesterfield Parkway North, St. Louis, Missouri 63198,
and Department of Chemistry, University of Missouri-St.
Louis, St. Louis, Missouri 63121
Small-molecule CETP inhibitors from a variety of
structural classes including sterols, polycyclic natural
products, and heterocycles are known.7 Each class of
known inhibitors appears to require a central core ring
for activity. Representative members of these classes
typically exhibit modest micromolar IC50 values for
inhibiting CETP-mediated transfer of [3H]CE from HDL
to LDL under buffered assay conditions in vitro.7 To
date, no CETP inhibitor class has been described that
exhibits submicromolar activity when this CETP-medi-
ated transfer process is assayed in the presence of
human plasma.7 However, recently a series of tetrahy-
dronaphthalene derivatives have been reported with
low-nanomolar potency in vitro13 and a series of tet-
rahydoquinolines derivatives have been reported (activ-
ity not given).14
Received August 4, 2000
In tr od u ction . Epidemiological studies have demon-
strated an inverse relationship between serum high-
density cholesterol (HDL-C) levels and the incidence of
coronary heart disease (CHD).1 Low levels of HDL-C
represent a significant independent risk factor in CHD
whether these patients have elevated plasma low-
density cholesterol (LDL-C).2 Several clinical studies
have shown reduced CHD events with treatments that
raised HDL-C.3,4 Multiple approaches have been identi-
fied with the potential to elevate HDL-C levels.5 Here
we describe our efforts to identify a novel simple class
of potent cholesteryl ester transfer protein (CETP)
inhibitors.
CETP is a plasma glycoprotein that mediates the
transfer of cholesteryl ester (CE) from HDL to very low-
density lipoprotein (VLDL) and LDL with a balanced
exchange of triglyceride (TG).6-8 CETP lowers HDL-C
by moving CE from HDL that is known to protect
against CHD into proatherogenic VLDL and LDL. In
contrast, CETP deficiency in humans results in hyper-
alphalipoproteinemia, i.e., elevated HDL-C.9 Analysis
In this Communication we describe the identification
and initial modification of N,N-disubstituted trifluoro-
3-amino-2-propanols as an unusually simple class of
CETP inhibitor. Subsequent optimization produced the
chiral (R)-(+)-propanol derivative 1a having low- na-
nomolar potency in vitro under buffered conditions. The
activity of 1a was also maintained at submicromolar
levels in the presence of human serum. This novel class
thus represents the first alicyclic series exhibiting
significant inhibitory activity for the CETP system.
* To whom correspondence should be addressed. Tel: (636)737-6792.
Fax: (636)737-7425. E-mail: richard.c.durley@monsanto.com.
† University of Missouri-St. Louis.
10.1021/jm000337b CCC: $19.00 © 2000 American Chemical Society
Published on Web 11/10/2000

4576 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24
Communications to the Editor
Sch em e 1a
a
Reagents: (a) 1,1,1-trifluoroepoxypropane, Yb(CF3SO3)3,
CH3CN, 50 °C, 2 h.
Sch em e 2a
F igu r e 1. ORTEP drawing of 9 from X-ray analysis.
of 4 and 5 with sodium triacetoxyborohydride also gave
3. In cases where the benzaldehyde 5 was difficult to
access, radical bromination of a suitable toluene 7 gave
the bromomethylbenzene 8 which could be used to
prepare 3 by reaction with an excess of aniline 4.
Surprisingly, the potent 3-amino-2-propanol 2k was
shown by analytical chiral chromatography to consist
of a 7:1 mixture of enantiomers. Independent chiral GC
analysis confirmed that the commercial 1,1,1-trifluoro-
2,3-oxirane reagent also contained a 7:1 mixture of the
individual (S)-(-)- and (R)-(+)-enantiomers. The indi-
vidual enantiomers of 2k were conveniently isolated by
standard preparative chiral chromatography. The activ-
ity16 was shown to reside in the minor (+)-isomer, 1a ,
which was prepared independently from the reaction of
N-(3-phenoxyphenyl)[[3-(1,1,2,2-tetrafluoroethoxy)phe-
nyl]methyl]amine with the known chiral epoxide, (R)-
(+)-1,1,1-trifluoro-2,3-epoxypropane.17 To confirm the
(R)-configuration at the chiral carbon in (+)-1a , a
crystalline analogue 9 was prepared from the reaction
of N-(3,4,5-trimethoxyphenyl)[[3-(trifluoromethylthio)-
phenyl]methyl]amine with (R)-(+)-1,1,1-trifluoro-2,3-
epoxypropane, and its structure was determined by
X-ray crystallography (Figure 1).
a
Reagents: (a) cyclohexane, heat, -H2O; (b) NaBH4, CH3OH;
(c) NaBH(OAc)3, AcOH, DCE, rt; (d) NBS, AIBN, CCl4, heat; (e)
cyclohexane, heat.
These results have been presented at the 220th ACS
National Meeting, Washington, DC, in which 1a was
identified as SC-795.15
Ch em istr y. N,N-Disubstituted trifluoro-3-amino-2-
propanols 2a -k were readily prepared from the ring-
opening reaction of commercially available 1,1,1-trifluoro-
2,3-oxirane of unspecified enantiomeric composition
with the appropriate N-benzylaniline 3 (Scheme 1). This
reaction proceeded smoothly in the presence of ytter-
bium(III) triflate in warm acetonitrile. The ytterbium
triflate helps the reaction proceed at lower temperature
and minimizes the need for large excess quantities of
this volatile epoxide due to thermal loss. This epoxide
ring-opening reaction proceeded with complete regiose-
lectivity, since none of the isomeric 2-amino-3-propanol
product was detected. The required N-benzylanilines 3
were conveniently prepared by standard reductive ami-
nation or alkylation sequences (Scheme 2). The aniline
4 was treated with an appropriate benzaldehyde 5 and
dehydrated to generate imine 6 which was then reduced
with sodium borohydride to give the N-benzylaniline 3.
Reduction of imine 6 to 3 with [3H]sodium borohydride
provided a convenient entry to radiolabeled analogues.17
Alternatively, direct reductive amination of a mixture
Resu lts a n d Discu ssion . An initial chemical lead
2a , identified through screening a combinatorial library,
had promising activity (IC50 40 µM) as a CETP inhibitor
in a simple buffer assay.16,18,19 In a similar assay but in
the presence of human serum which provided the source
of the LDL, VLDL, and human CETP, the inhibitory
activity (IC50 > 200 µM) was markedly reduced for 2a .
The IC50 value in human serum is indicative of the
inhibitory activity in the target tissue, human blood,
when other plasma lipoproteins are present. We suspect
that this value is lower than that in buffer due to
nonspecific binding of the inhibitors to nontarget blood

Communications to the Editor
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 24 4577
Ta ble 1. Structure and CETP Inhibitory Properties of
N,N-Disubstituted Trifluoro-3-amino-2-propanols
significant potency in the presence of human serum
(IC50 6 µM).
When 2k was analyzed by chiral chromatography, an
unexpected 7:1 ratio of enantiomers was observed. The
individual enantiomers were obtained by preparative
chiral chromatography and analyzed for CETP inhibi-
tory activity. The minor (+)-enantiomer 1a had an IC50
0.02 µM, while the major (-)-enantiomer 1b exhibited
much less activity (IC50 0.8 µM). The minor (+)-
enantiomer 1a also displayed submicromolar activity
(IC50 0.6 µM) in the presence of human serum. More-
over, 1a could be prepared independently from the
known chiral epoxide, (R)-(+)-1,1,1-trifluoro-2,3-ep-
oxypropane,17 suggesting that the (+)-1a enantiomer
contained the (R)-configuration. Since no unequivocal
structural data has been presented on such compounds,
we prepared a related crystalline analogue 9 from the
same sample of chiral (R)-(+)-1,1,1-trifluoro-2,3-ep-
oxypropane,17 determined its structure by X-ray analy-
sis (Figure 1), and confirmed that the alcohol configu-
ration in 9 was indeed (R). From these data we conclude
that the structure of (+)-1a also has the (R)-configura-
tion.
More detailed biochemical binding studies comparing
the relative affinities and binding properties of 2k , 1a ,
and 1b have been completed and are reported sepa-
rately.16 The results of these efforts demonstrate that
these inhibitors reversibly block both CETP-mediated
TG and CE transfer. They associate with HDL and LDL
but do not disrupt the structure of these lipoproteins.
Their CETP inhibitory activity is highly specific, since
they do not inhibit phospholipid transfer protein or
lecithin cholesterol acyl transferase. Competition ex-
periments showed that the potent reversible inhibitor
1a prevented CE binding to CETP and that 1a bound
approximately 5000-fold more efficiently to CETP than
CE.
IC50 (µM)
IC50 (µM) in
inhibitor
R1
R2
3-CF3
3-CF3
H
3-CH3
2-CF3
3-OCF3
4-OCF3
in buffera human seruma,b
2a
2b
2c
2d
2e
2f
2g
2h
2i
3-F
H
40
15
>100
>100
>100
12
>200
>200
ND
ND
ND
>200
ND
40
3-F
3-F
3-F
3-F
3-F
15
1
25
4
3-phenoxy 3-OCF3
4-phenoxy 3-OCF3
3-phenoxy 4-OCF3
3-phenoxy 3-OCF2CF2H
ND
120
6
2j
2k
0.2
a
b
ref 18. ND, not determined.
Ta ble 2. Structure and CETP Inhibitory Properties of Chiral
N,N-Disubstituted Trifluoro-3-amino-2-propanols
IC50 (µM)
IC50 (µM) in
inhibitor
in buffera
human seruma,b
1a
1b
9
0.02
0.8
5
0.6
20
ND
a
b
Ref 18. ND, not determined.
proteins. The simplicity of the chemistry needed to
prepare 2a suggested that a wide variety of structural
modifications could be readily incorporated to explore
the structure-activity relationships (SAR) with this
interesting class (Table 1).
As summarized below, several key structural changes
provided insights into the SAR and suggested a direc-
tion for further modifications. Removing the aniline 3-F
group from 2a to give 2b (IC50 15 µM) retained activity,
while removing the benzylic 3-CF3 group as in 2c or
replacing the benzylic 3-CF3 substituent in 2a with a
3-CH3 moiety as in 2d produced a significant reduction
in potency. These results indicated that the benzylic
3-CF3 substituent was an important contributor to the
activity while the aniline ring was open to broader
modification. Similarly, moving the benzylic 3-CF3 group
to an ortho position as in 2e also dramatically reduced
activity (IC50 > 100 µM). Ortho substituents would likely
affect the respective orientation of the two phenyl rings,
and this in turn may be critical for potency. Alterna-
tively, changing the benzylic 3-CF3 group in 2a to a
3-OCF3 group as in 2f (IC50 12 µM) improved the
potency by over 3-fold. Activity similar to 2f was also
observed with the 4-OCF3 substituent as in 2g.
In conclusion, we have discovered chiral N,N-disub-
stituted trifluoro-3-amino-2-propanols as a new simple
class of potent CETP inhibitors. The most potent inhibi-
tor 1a also exhibited submicromolar activity in the
presence of human serum. We are currently optimizing
this series to identify even more potent analogues.
Ack n ow led gm en t. We thank Shengtian Yang and
J ohn Likos for NMR assistance and J ames Doom and
Gary Mappes for mass spectral analysis. We also thank
Anne Daiss for elemental analysis. We are grateful to
Brad McKinnis for chiral chromatography analysis.
Su p p or tin g In for m a tion Ava ila ble: Experimental de-
tails and analytical data for the preparation of compounds 1a ,
1b, 2a -k , and 9 as well as details of the X-ray structure for
9. This material is available free of charge via the Internet at
http://pubs.acs.org.
Modifying the 3-F substituent in 2f with a 3-phenoxy
group as in 2h (IC50 1 µM) identified the first low-
micromolar inhibitor, and more importantly, 2h now
exhibited some inhibitory activity in the presence of
human serum (IC50 40 µM). Moving the 3-phenoxy
aniline substituent in 2h to the 4-position as in 2i (IC50
25 µM) significantly reduced activity. Similarly, moving
the benzylic 3-OCF3 group in 2h to the 4-position as in
2j (IC50 4 µM) also reduced activity. Subsequent exten-
sion of the benzylic 3-OCF3 group in 2h using a
3-(1,1,2,2-tetrafluoroethoxy) moiety identified 2k (IC50
0.2 µM) with nanomolar potency, and 2k also had
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