Synthesis and Structure-Activity Relationships of 1-Arylmethyl-5-aryl-6-methyluracils as Potent Gonadotropin-Releasing Hormone Receptor Antagonists

Article abstract of DOI:10.1021/jm030472z

Based on the SAR from bicyclic gonadotropin-releasing hormone (GnRH) antagonists such as 6-aminomethyl-7-aryl-pyrrolo [1,2-a] pyrimid-4-ones (5) and 2-aryl-3-aminomethyl-imidazolo-[1,2-a]pyrimid-5-ones (6a,b), a series of novel uracil compounds (8) were d

Full text of DOI:10.1021/jm030472z

J . Med. Chem. 2004, 47, 1259-1271
1259
Syn th esis a n d Str u ctu r e-Activity Rela tion sh ip s of
1-Ar ylm eth yl-5-a r yl-6-m eth ylu r a cils a s P oten t Gon a d otr op in -Relea sin g
Hor m on e Recep tor An ta gon ists
Zhiqiang Guo,*,† Yun-Fei Zhu,† Timothy D. Gross,† Fabio C. Tucci,† Yinghong Gao,† Manisha Moorjani,†
Patrick J . Connors, J r.,† Martin W. Rowbottom,† Yongsheng Chen,† R. Scott Struthers,‡ Qiu Xie,‡
J ohn Saunders,† Greg Reinhart,‡ Ta Kung Chen,§ Anne L. Killam Bonneville,§ and Chen Chen*,†
Department of Medicinal Chemistry, Department of Exploratory Discovery, and Department of Preclinic Development,
Neurocrine Biosciences, Inc., 10555 Science Center Drive, San Diego, California 92121
Received September 19, 2003
Based on the SAR from bicyclic gonadotropin-releasing hormone (GnRH) antagonists such as
6-aminomethyl-7-aryl-pyrrolo[1,2-a]pyrimid-4-ones (5) and 2-aryl-3-aminomethyl-imidazolo-
[1,2-a]pyrimid-5-ones (6a ,b), a series of novel uracil compounds (8) were derived as GnRH
antagonists. The synthesis and SAR studies of 6-methyluracils as human GnRH receptor
antagonists are discussed herein. Introduction of a small methyl substituent at the â-position
of the N3 side-chain improved the GnRH binding potency by 5-10-fold. Introduction of a methyl
group of (R)-configuration at the R-carbon of the N-3 side-chain gave a modest improvement
in binding affinity over the unsubstituted ethylene analogues. This modification enabled us to
make uracil compounds without the labile 2-pyridylethyl motif on the basic nitrogen while
still maintained excellent potency against the hGnRH receptor.
In tr od u ction
“flare effect”, which is exhibited by the peptide agonists
due to their initial overstimulation of the receptor.4
Nevertheless, both peptide agonists and antagonists
require parenteral administration, typically in depot
form due to their poor oral bioavailability. By contrast,
small molecule GnRH antagonists offer the potential for
oral administration and therefore could gain wider
acceptance from patients. In response to that need,
intensive efforts have been initiated for the development
of orally active small molecule GnRH antagonists by
many laboratories.5-10
Gonadotropin-releasing hormone (GnRH), also known
as luteinizing hormone-releasing hormone (LHRH), is
the decapeptide (pGlu-His-Trp-Ser-Tyr-Gly-Leu-Arg-
Pro-Gly-NH2), hormone produced in and secreted by the
hypothalamus in a pulsatile manner.1,2 It acts on the
pituitary gland to stimulate the secretion of both lutein-
izing hormone (LH) and follicle-stimulating hormone
(FSH). These gonadotropins, in turn, act on the repro-
ductive organs, where they participate in the regulation
of gonadal steroid production, spermatogenesis in males,
and follicular development in females. Several repro-
ductive disease conditions such as endometriosis, uter-
ine fibroids, and prostate cancer are caused by over-
stimulation of the reproductive organs by the gonadal
steroids and thus can be treated by suppression of the
pituitary-gonadal axis. Continuous administration of
high dose of GnRH or its analogues (agonists) desensi-
tizes the receptor with consequent decline in gonadal
gametogenesis and steroid production. This phenom-
enon has successfully led to an extensive application of
GnRH peptide agonists for treatment of a variety of
reproductive diseases. Currently, depot forms of peptidic
GnRH agonists, represented by leuprorelin are used to
treat such conditions through a receptor down-regula-
tion mechanism to suppress gonadal steroid production.3
On the another hand, recent clinical evidence has shown
that peptidic GnRH antagonists can act immediately at
the receptor through competition with endogenous
GnRH to inhibit the reproductive system and therefore
alleviate disease symptoms without the concomitant
Compound 1 (T-98475, Figure 1) and its analogues
are the first small molecules reported5 to have high
affinity on the human GnRH receptor (IC50: 0.2 nM)
albeit with reduced binding affinity for the rat receptor
(IC50: 60 nM).5 In addition, a series of papers on
quinolones and tryptamines as potent GnRH antago-
nists were recently published. For example, quinolone
2 possesses high binding affinity (IC50: 0.4 nM) for the
human GnRH receptor as well as the rat GnRH receptor
(IC50: 4 nM).6 Indole derivatives such as 3 were reported
to be potent and orally bioavailable GnRH antagonists.7
Most recently, TAK-013 (IC50: 0.2 nM) and its ana-
logues were reported as potent and orally active hGnRH
antagonists.8 Most of the above compounds have high
binding affinity to the hGnRH receptor, but they all
have high molecular weight (1 and 4 above 650; 2 and
3 above 500).
In the previous papers,9,10 we disclosed the SAR study
of 6-aminomethyl-7-aryl-pyrrolo[1,2-a]pyrimid-4-ones (5,
Figure 2) and 2-aryl-3-aminomethyl-imidazolo[1,2-a]-
pyrimid-5-ones (6a ,b) as potent hGnRH receptor an-
tagonists. The SAR of 5 revealed that the binding
affinity was enhanced by a 2-pyridyl group on the side
chain of the 6-aminomethyl group and a para substitu-
ent (R3) on the 7-phenyl group (e.g. 5, R1 ) CH3, R2 )
* Correponding authors. For Z. Guo, Tel: 1-858-658-7657. Fax:
1-858-658-7601. E-mail: zguo@neurocrine.com. For C. Chen, Tel:
1-858-658-7634. Fax: 1-858-658-7619. E-mail cchen@neurocrine.com.
† Department of Medicinal Chemistry.
‡ Department of Exploratory Discovery.
§ Department of Preclinic Development.
10.1021/jm030472z CCC: $27.50 © 2004 American Chemical Society
Published on Web 02/04/2004

1260 J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5
Guo et al.
Sch em e 1a
F igu r e 1. Small molecule GnRH antagonists.
a
Reagents and conditions: (a) 2,6-difluorobenzyl bromide, TBAF,
THF/DMF, 59%; (b) NaIO4, OsO4 (cat.), THF/H2O, quant.; (c) Br2,
HOAc, 58%; (d) 2-(N-methyaminoethyl)-pyridine, NaBH(OAc)3,
DCM, 73%; (e) boronic acid or boronic acid pinacol ester, Pd(PPh3)4,
K2CO3, toluene/H2O, reflux, 30-80%.
vation led us to successfully design the 2-alkyl ana-
logues 7, which possess lower molecular weights but
maintain the potent GnRH antagonistic activity of 6b.11
On the basis of these SAR results, we further postulated
that the whole left side of 7 including the alkyl group
and the five-membered ring might not be necessary for
binding, which would lead to much smaller monocyclic
compounds. To test this hypothesis, the uracil was
chosen as one of several possible monocyclic core
structures due to its synthetic accessibility. The initial
SAR studies of this novel monocyclic series (8) as human
GnRH receptor antagonists were discussed recently.12
The best compound (13b) from the initial SAR study
has a Ki value of 34 nM, which is less potent than some
of the bicyclic analogues. We rationalized that reducing
the flexibility of the alkylamino side chain at the
3-position should be beneficial for binding energy, as
long as one of the low-energy conformers fits into the
binding pocket. A small alkyl group such as a methyl
substituent at the ethylene linker could serve such a
purpose. Here we report the synthesis and SAR studies
of the 1-arylmethyl-3-(2-aminoethyl)-5-aryl-6-methyl-
uracil series.
F igu r e 2. From bicyclic to monocyclic GnRH antagonists.
Ch em istr y
The initial approach to synthesize the fully substi-
tuted uracils is outlined in Scheme 1. 3-Allyl-6-methyl-
uracil (9) was obtained by condensation of N-allylurea
with ethyl acetoacetate using a known procedure.13 It
was then treated with 2,6-difluorobenzyl bromide in the
presence of n-tetrabutylammonium fluoride in THF to
afford 10 in 59% yield, which underwent oxidative
cleavage of the olefin with NaIO4, followed by bromi-
nation at the 5-position to yield 11 in 58% yield (two
steps). Reductive amination of the aldehyde 11 with
2-(N-methyl-2-aminoethyl)pyridine, one of the favored
side-chains from the bicyclic series, in the presence of
NaBH(OAc)3, gave 12 in 73% yield. Finally, 12 was
subjected to Suzuki coupling reactions with a variety
of arylboronic acids or boronic acid pinacol esters to
afford the desired products 13(a -v) in 30-80% yields.
While the above synthesis of the 6-methyluracil core
offers quick access to the desired products 13, its
F igu r e 3. X-ray structure for 16.
2-PyCH2CH2; R3 ) n-PrCONH). Furthermore, SAR
studies of 6b indicated that an aryl group, preferably
the 3-methoxyphenyl group, could replace the 6-car-
boxylate of 6a . Introduction of the 6-aryl group led to a
different SAR profile in the two series represented by
6a and 6b. Thus, while the para substituent (R3) on the
phenyl ring of the molecule 6a was crucial for binding,
it had minimal effect on the activity of 6b. This obser-

Gonadotropin-Releasing Hormone Receptor Antagonists
J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5 1261
Sch em e 2a
ethane, followed by reaction with NaBH(OAc)3 to afford
the desired chiral amines 25 and 26.
To examine the substitution effect at the R-position
of the N3 side chain, we also prepared compounds 34
and 35, and the synthesis is shown in Scheme 4.
Commercial (R)- or (S)-1-amino-2-propanol (R-27 or
S-27, Aldrich, 97% ee) was protected as its tert-butyl
carbamate derivate and then used in the Mitsunobu
alkylation of uracil 19 (Scheme 4). The Mitsunobu
reaction on the secondary alcohol R-28, or S-28, occurred
with complete inversion of the chiral center, leading to
the corresponding alkylated uracil S-29, or R-29, with
(S)-, or (R)-configurations, in ∼70% yield. The stereo-
chemical integrity of S-29 and R-29 was confirmed by
chiral HPLC analysis,15 and the absolute (R)-stereo-
chemistry of R-29 was determined by X-ray crystallog-
raphy,12c which was possible due to the anomalous
contributions of the heavy atom (bromine).16 The 5-(3-
methoxyphenyl)- or 5-(2-fluoro-3-methoxyphenyl) group
was introduced via a Suzuki coupling reaction of the
appropriate boronic acids with bromides S-29 or R-29.
For example, compound 31 was obtained in 53% yield,
which was treated with trifluoroacetic acid in CH2Cl2
to furnish the corresponding amine 33 in 95% yield. 32
and 33 were then reacted with a variety of aldehydes
and ketones, under reductive conditions, to give the
corresponding secondary amines 34 and 35.
a
Reagents and conditions: (a) urea, water, HCl, reflux, 79%;
(b) diketene, HOAc, reflux, 40 min, 69%; (c) diketene, pyridine,
rt, 24 h, 60%; (d) HOAc, reflux, 1 h, 95%.
application is limited in several respects. First, the
condensation of allylurea with ethyl acetoacetate gave
low yield (∼7%) and a long reaction time (12 days).
Second, the N3 substituent was introduced into the
molecule at the beginning of synthesis, thus limiting the
SAR study at this site. Accordingly, a new and improved
synthetic route was then developed, as shown in Scheme
2 and 3. Thus, 2,6-difluorobenzylamine and urea were
refluxed in HCl/water solution to form the benzyl urea
15 (Scheme 2), which was crystallized from EtOAc in
79% yield. Urea 15 and diketene were then refluxed in
acetic acid for 40 min to yield a mixture of N1- and N3-
substituted 6-methyluracils (16 and 17) in 3:1 ratio.
1-(2,6-Difluorobenzyl)-6-methyluracil (16) was separated
from its N3-isomer (17) by recrystallization in acetic
acid. This two-step sequence afforded 16 in about 25%
yield from 14.12b A more efficient procedure was sub-
sequently developed to afford the N1-substituted 6-me-
thyluracil 16 exclusively, wherein compound 15 was
first treated with diketene in pyridine at ambient
temperature for 24 h, and the intermediate 18 was
separated in 60% yield. 18 was then refluxed in acetic
acid for 1 h and the desired product 16 crystallized out
from the reaction mixture in 95% yield (Scheme 2). The
structure assignments of 16 and 17 were confirmed by
NOE NMR experiments. (In compound 16, for instance,
there was a clear NOE of the 6-methyl signal upon
irradiation of the benzylic protons at N1; this effect was
absent in 17.)14
Resu lts a n d Discu ssion s
In Vitr o Recep tor Bin d in g Stu d ies. All synthe-
sized compounds were evaluated for their ability to
inhibit des-Gly10[125I-Tyr5,DLeu6,NMeLeu7,Pro9-NEt]-
GnRH radioligand binding to the cloned hGnRH recep-
tor stably expressed in HEK293 cells using a 96-well
filtration assay format.17
Our initial SAR studies of the uracil series indicated
that at N3 position, [N-methyl-N-(2-pyridylethyl)]-ami-
noethyl group was the key feature for obtaining highly
potent hGnRH antagonists.12a Thus, this preferred side-
chain was fixed at N3 position, while the impact of the
substitution on the 5-phenyl group to binding affinities
was examined and the data are summarized in Table
1.18 The unsubstituted phenyl compound 13a was more
than 10-fold less potent than the corresponding 3-meth-
oxyphenyl analogue 13b, indicating that the 3-methoxy
group may function as a possible hydrogen-bond accep-
tor. Replacement of the methoxyl group by a 3,4-
methylenedioxy (13c) or 3,4-ethylenedioxy (13d ) gen-
erated equally potent molecules. However, replacement
with the difluoro-3,4-methylenedioxy group (13e, 470
nM) reduced the binding affinity significantly, which
might be caused by reduced hydrogen bonding ability
of the substituent. Shifting the methoxy group from the
3- to the 4-position also resulted in a substantial
reduction in potency (13f, 230 nM), while extending the
4-methoxy to 4-ethoxy led to further decrease in potency
(13g, 680 nM). Combining 3- and 4-methoxy groups into
the phenyl ring resulted in compound (13h ) with very
poor binding, and incorporation of the 3,4,5-trimethoxy-
phenyl group yielded an inactive compound (13i). The
importance of hydrogen bonding ability of the substit-
uents to the binding affinity was further demonstrated
by 13m -o, in which simple alkyl substituents without
any oxygen atom led to lower potency. Although a strong
Uracil 16 was brominated in acetic acid to yield 19
in 88% yield. Compound 19 was treated with either N-t-
Boc-R-alaninol or its corresponding S-isomer under
Mitsunobu conditions in THF using di-tert-butyl-azodi-
carboxylate as the coupling reagent to give R-20 or S-20
in high yield (94% and 96%, respectively) with complete
retention of their chirality (Scheme 3). Coupling of 20
with 3-methoxyphenylboronic acid or 2-fluoro-3-meth-
oxyphenylboronic acid under the Suzuki coupling condi-
tions catalyzed by Pd(0) yielded 21 or 22 in about 45%
yield. Deprotection of the amino-Boc group of 21 and
22 with TFA gave the amino compounds 23 and 24 in
quantitative yields. These amines (23 and 24) were
treated, under reductive alkylation conditions, with a
variety of aldehydes and ketones (R1R2CO) in dichloro-

1262 J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5
Guo et al.
Sch em e 3a
a
Reagents and conditions: (a) Br2, HOAc, 88%; (b) N-t-Boc-R-alaninol or N-t-Boc-S-alaninol, PPh3, di-tert-butyl-azodicarboxylate, THF,
94% or 96%; (c) 3-methoxyphenyl boronic acid or 2-fluoro-3-methoxyphenyl boronic acid, Pd(Ph3P)4, Na2CO3, toluene/H2O, reflux, ∼45%;
(d) TFA/CH2Cl2 (1:1), quant.; (e) aldehyde or ketone, NaBH(OAc)3, dichloroethane.
Sch em e 4a
a
Reagents and conditions: (a) (Boc)2O, Et3N, CH2Cl2, 0 °C to rt, 2 h, quant.; (b) R-28 or S-28, DEAD, Ph3P, THF, rt, 17 h, ∼70%; (c)
3-methoxyphenyl boronic acid or 2-fluoro-3-methoxyphenyl boronic acid, Pd(Ph3P)4, K2CO3, toluene/EtOH/H2O, 110 °C, 18 h, 90% or
53%; (d) TFA/CH2Cl2 1:1 v/v, rt, 1 h, 95%; (e) aldehyde or ketone, MeOH, NaBH4, rt, 1 h.
electron-withdrawing fluorine atom at 3-position re-
sulted in weakly active analogue (13q, 1200 nM),
switching the fluorine to the 2-position gave a 5-fold
increase in binding affinity in comparison to unsubsti-
tuted phenyl analogue (13a ). When the 2-fluorine and
the 3-methoxy substituents were combined into the
5-phenyl group of the uracil, a dramatic increase in
binding affinity was achieved and 13s became the most
potent compound in this series (3 nM). This phenom-
enon may be explained by a combination of effects
caused by the stereo-effect of the 2-fluoro and hydrogen
bond acceptor by the 3-methoxy group. We also specu-
late that the 2-fluoro group, being slightly larger than
a proton, could impede the free rotation of the 5-phenyl
ring,19 thus force the 5-phenyl ring into a perpendicular
conformation with respect to the uracil core, which in
turn, may be preferred for a π-π interaction with the
GnRH receptor. Extending the 5-phenyl ring to a larger
aromatic ring system (13u and 13v) did not give
noticeable change in binding affinity.

Gonadotropin-Releasing Hormone Receptor Antagonists
J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5 1263
Ta ble 1. Binding Affinities of Compounds 13a -v to the Human GnRH Receptor18
With this success, we also rationalized that restriction
of the flexible ethylene side chain on the N3 of uracil
may also be beneficial for the binding, and introducing
a small alkyl group such as a methyl at the R- or
â-position could serve such purpose. In addition, the
readily available chiral 2-amino-1-propanol (alaninol)
and 1-amino-2-propanol enabled us to examine the
stereo-effect. The results from compounds 25 and 26,
prepared from R- and S-2-amino-1-propanols, are sum-
marized in Table 2. The previous SAR studies from both
bicyclic 5-7 and uracil 8 demonstrated the 2-(2-pyridyl)-
ethylamino side-chain was the most preferred, and this
preference continued in compound 25 and 26 with a
â-methylated ethylene side-chain, albeit not signifi-
cantly. For example, R-25c, which has the 2-(2-pyridyl)-
ethylamino group, was merely 2-fold better in binding
affinity than the corresponding benzyl analogue R-25a ,
while the 2-pyridylmethyl derivative R-25b was 5-fold
more active than R-25c. Interestingly, for N-benzyl
derivatives, R- and S-isomers were almost equally
active. As expected, introducing the 2-fluoro-3-methoxy-
phenyl group at 5-position greatly improved the binding
affinity. S-26a was 6 times more potent than S-25a ,
while R-26b was more than 10 times better than R-25b.
Compound R-26d with a thiazolylmethyl side-chain also
showed good activity. For (S)-derivatives, both benzyl
26a and benzylic-like side-chains, such as thiophene-
methyl and furan-methyl 26e, 26f demonstrated good
activities as well as small alkyl groups such as 26g, 26h ,
and 26i (Ki ) 4.1, 4.5, and 5.0 nM, respectively) also
generated highly potent compounds.
The SAR of compounds 34 and 35 with a methyl
group on the R-position of the N-3 side-chain was also
examined and the results are shown in Table 3. Unlike
the compounds 25 and 26, the (R)-enantiomeric prefer-
ence for the receptor was consistently confirmed by
alkylation of amines S-32 and R-32 with various alde-
hydes and ketones.12c For instance, the N-benzylated
compound S-34a (Ki ) 370 nM) was about 5 times less
potent than its counterpart R-34a (Ki ) 75 nM). It is
clear that benzylic side-chains were preferred over
aliphatic ones for this subseries. The substitution pat-
tern of the benzylic side-chain was also investigated.
Introduction of an o-methyl group R-34b caused little
change in binding affinity compared to R-34a whereas
o-methoxy compound (R-34c) was about 4-fold more
potent. Interestingly, there was no obvious difference
in the binding affinity between these compounds in the

1264 J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5
Guo et al.
Ta ble 2. Binding Affinities of Compounds 25 and 26 to the
Ta ble 3. Binding Affinities of Compounds 34 and 35 to the
Human GnRH Receptor18
Human GnRH Receptor18
Ta ble 4. Functional Assay Data to the Human GnRH
Receptor, and Binding Affinities to the Rats and Monkey GnRH
Receptors for Selected Compounds18
compd human Ki (nM) IC50 (nM) rats Ki (nM) monkey Ki (nM)
13b
34
25
16
38
0.5
6.1
4.0
27
14
5.5
10
5.7
2200
25000
6400
10000
3200
4900
7600
7500
22000
7900
610
550
43
820
87
260
200
50
S-26a
R-26b
R-26d
S-26e
S-26f
S-26g
S-26h
S-26i
R-35c
1.1
13
4.0
15
4.1
4.5
5.0
3.9
S-isomers, 34a -c all had Ki values around 350 nM. As
expected, introducing a 5-(2-fluoro-3-methoxy)-phenyl
group generated highly potent compounds. R-35b (Ki )
7 nM) was about 10 times more potent than its coun-
terpart R-34b, while R-35c (Ki ) 3.9 nM) was about 5
times better than R-34c.
87
30
affinity on the rat GnRH receptor. For example, com-
pound S-26h exhibited 10 times weaker binding affinity
on the monkey GnRH receptor (Ki ) 50 nM) and 1000
times lower affinity on the rat GnRH receptor (Ki ) 7.5
µM) than the high affinity on the human GnRH receptor
(Ki ) 4.5 nM).
Selected compounds were also studied for their phar-
macokinetic properties in mice, and the results are
summarized in Table 5. Compound 13b from the initial
series with a 2-pyridylethylamino side-chain was meta-
bolically labile.12a The high clearance rate data in mice
(CL ) 120 mL/min‚kg) suggested other degradation
routes in addition to hepatic removal (hepatic blood flood
in mice is about 90 mL/min‚kg). Sample analysis from
hepatic portal vein indicated that this compound was
also poorly absorbed (4.7% by comparing oral hepatic
concentration versus iv plasma concentration). Com-
pounds from the alaninol series had improved pharma-
F u r th er in Vitr o a n d in Vivo Stu d ies. To ensure
that these high binding affinity molecules were func-
tional antagonists, selected compounds were tested for
their ability to inhibit Ca2+ influx induced by GnRH in
RBL cells transfected with the GnRH receptor. The
results shown in Table 4 indicate that all these com-
pounds are potent functional antagonists with compa-
rable IC50 values versus their Ki values. For example,
compound S-26h exhibited a Ki value of 4.5 nM on
human GnRH receptor binding, and it’s also a functional
antagonist with IC50 value of 5.5 nM in the Ca2+ influx
assay. Similar to our previous results,9a these com-
pounds displayed species differences in their binding to
the GnRH receptors.20 They exhibited reduced binding
affinity on the monkey GnRH receptor and much lower

Gonadotropin-Releasing Hormone Receptor Antagonists
J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5 1265
(25 µg protein in 130 µL binding buffer) to 50 µL of [125I]-
labeled GnRH peptide (∼100 000 cpm), and 20 µL of competitor
at increasing concentrations. The reaction was terminated
after 90 min by filtration and washing (2×) with phosphate-
buffered saline. Bound radioactivity was measured by remov-
ing the filters from the plate, and the filters were counted
directly on a gamma counter. Ki values were calculated from
competition binding data using nonlinear least squares regres-
sion analysis by the Prism software package (GraphPad
Software) with the Cheng-Prusoff equation.
Ta ble 5. In Vivo Pharmacokinetics of Selected Compounds in
Mice
clearance
AUC (oral)
compd
F (%)
t1/2 (h)
(mL/min‚kg)
(ng/mL‚h)
13bb
1.6
6.0
8.2
2.2
0.4
1.3
0.9
0.6
120
97.3
129
70.7
23
104
110
57
R-26d a
S-26ga
S-26h c
a
b
Dosed at 10 mg/kg iv and 10 mg/kg po. Dosed at 5 mg/kg iv
and 10 mg/kg po. c Dosed at 2.5 mg/kg iv and 10 mg/kg po.
Ca 2+ In flu x Assa y. In h ibition of Gn RH-Stim u la ted
Ca 2+ F lu x. Functional activity of the compounds for the
human GnRH receptor was determined by inhibition of GnRH-
stimulated Ca2+ flux. RBL cells stably expressing the full-
length human GnRH receptor were seeded into 96-well, black-
wall clear bottom plates (Corning) at a density of 50 000 cells/
well and the cells allowed to attach overnight. The attached
cells were then loaded with the Ca2+ sensitive dye, Fluo-4
(Molecular Probes), by incubation in loading medium [(DMEM
with 20 mM Hepes, 10% FBS, 2 µM Fluo-4, 0.02% pluronic
acid (Molecular Probes), and 2.5 mM probenecid (Sigma)] for
1 h at 37 °C. The cells were then washed three times with
assay buffer (Hanks balanced salt, 20 mM Hepes, 2.5 mM
probenecid). Compounds at increasing concentrations in assay
buffer were preincubated with the cells for 1 min prior to
stimulation with GnRH (5 nM). Measurement of fluorescence
due to GnRH-stimulated Ca2+ flux was performed according
to manufacturer’s instructions on a FLIPR system (Molecular
Devices, FLIPR384 system). IC50 values for the inhibition of
GnRH-stimulated Ca2+ flux were calculated using the Prism
software package (GraphPad Software) with a “sigmoidal
dose-response (variable slope)” option for curve fitting.
cokinetic profiles. For example, R-26d and S-26g had
reasonable oral exposure in mice with absolute oral
bioavailability (F) of 6.0 and 8.2%, respectively, despite
high clearance in this species. Compound S-26h also
had high clearance in mice, which may account for its
low oral bioavailability, but this compound exhibited
reasonable absorption (42%) by comparing the blood
sample concentration from hepatic portal vein of oral
dose versus iv plasma concentration. This compound
had moderate volume distribution (4 L/kg) and short
half-life in mice (0.6 h). However, in an in vitro human
liver microsome assay, this compound was projected to
have a systemic clearance of 8.1 mL/min‚kg, a much
lower value than 13b, which had a clearance value of
18.1 mL/min‚kg. On the basis of in vitro human liver
microsomes stability and in vivo mouse absorption data,
one can predict this compound should have oral bio-
availability of 27% in humans.
Ch em istr y. Elemental analysis results are indicated by
atom symbols and are within 0.4% of theoretical values except
where indicated. H NMR spectra were recorded on a Varian
Con clu sion
1
In conclusion, we have successfully designed and
synthesized a series of novel and potent monocyclic
uracil GnRH antagonists. A novel, convergent cycliza-
tion procedure for assembly of the 6-methyl uracil core
was developed. Introduction of a small methyl substitu-
ent at the â-position of the N3 side-chain improved the
GnRH binding potency by 5-10-fold. We have also
demonstrated that introduction of a methyl group of (R)-
configuration at the R-carbon of the N-3 side-chain gave
a modest improvement in binding affinity over the
unsubstituted ethylene analogues. This modification
enabled us to make uracil compounds without the labile
2-pyridylethyl motif on the basic nitrogen while still
maintaining excellent potency against the hGnRH
receptor. More importantly, these new uracil compounds
have lower molecular weights (∼500), which should be
beneficial in the discovery of bioavailabile compounds.
Further modification in terms of metabolic stability and
oral bioavailability will be published in due course.
Spectrometer (Mercury 300 Hz) using TMS as the internal
standard and CDCl3 as solvent except where indicated. Final
products were purified by Gilson preparatory HPLC system,
which was connected to a mass spectrometer and fraction
collector. The fraction collector was triggered by the desired
mass. All final compounds after purification were reanalyzed
by reverse phase HPLC-MS system (HP-4500 series with APCI
mode for mass detection) and determined to be at least 98%
pure based on two UV absorbance wavelengths (220 nM, 254
nM) and total ion current (TIC) monitoring from the mass
spectrometer.
3-Allyl-6-m eth ylu r a cil (9). To the allylic urea (25 g, 0.25
mol) in anhydrous ethanol (10 mL) were added ethyl acetoace-
tate (31.86 mL, 0.25 mol) and 10 drops of concentrated HCl
solution. The mixture was stored at rt for 12 days in a
desiccator and then concentrated and redissolved in MeOH
(200 mL). KOH (22.5 g, 0.40 mol) was added and the solution
refluxed for 1 h, acidified slowly by 6 N HCl. The resulting
precipitates were collected by filtration. The solid was washed
with water and dried under vacuum at 50 °C for 1 day to give
2.7 g of white solid (9, 6.5%). 1H NMR (CDCl3): δ 2.16 (s, 3H),
4.52 (d, J ) 5.3 Hz, 2H), 5.18 (d, J ) 10.2 Hz, 1H), 5.23 (d, J
) 18.2 Hz, 1H), 5.60 (s, 1H), 5.82-5.93 (m, 1H), 10.30 (s, 1H).
Anal. (C8H10N2O2) C, H, N.
Exp er im en ta l Section
Com p etitive Ra d ioliga n d Bin d in g Assa y. The affinity
of the compounds for the human GnRH receptor was deter-
mined by competitive displacement of the GnRH receptor
radioligand, [125I-Tyr5,DLeu6,NMeLeu7,Pro9-NEt]GnRH. HEK293
cells stably expressing the full-length human GnRH receptor
were harvested, resuspended in 5% sucrose, and homogenized
using a polytron homogenizer (2 × 15 s). Nuclei debri were
removed by centrifugation (3000g for 5 min), and the super-
natant centrifuged (20 000g for 30 min, 4 °C) to collect the
membrane fraction. The final membrane preparation was
resuspended in binding buffer (10 mM HEPES (pH 7.5), 150
mM NaCl, and 0.1% BSA) and stored at -70 °C. Binding
reactions were performed in a Millipore MultiScreen 96-well
filtration plate assembly with polyethylenimine-coated GF/C
membranes. The reaction was initiated by adding membranes
1-(2,6-Diflu or oben zyl)-3-a llyl-6-m eth ylu r a cil (10). To 9
(2.6 g, 15.7 mmol) in DMF (20 mL) was added tetrabutylam-
monium fluoride (1M in THF, 25 mmol, 25 mL), followed by
addition of 2,6-difluorobenzyl bromide (4.14 g, 20 mmol). The
mixture was stirred at r.t for 2 days and then concentrated.
The crude product was partitioned between ethyl acetate (200
mL) and water (100 mL). The organic layer was separated,
dried over Na2SO4, and purified by column chromatography
with silica gel (ethyl acetate/hexane) to recover the starting
1
material (0.39 g) and the desired product (10, 2.7 g, 59%). H
NMR (CDCl3): δ 2.26 (s, 3H), 4.52-4.55 (m, 2H), 5.18(s, 2H),
5.13-5.23 (m, 2H), 5.63 (s, 1H), 5.81-5.92 (m, 1H), 6.86-6.95
(m, 2H), 7.23-7.33 (m, 1H). MS (CI) m/z 293.1 (MH+). Anal.
(C15H14N2O2) C, H, N.

1266 J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5
Guo et al.
1-(2,6-Diflu or oben zyl)-3-(2-oxoeth yl)-5-br om o-6-m eth -
ylu r a cil (11). To 10 (1.46 g, 5 mmol) in THF/H2O (20 mL/10
mL) were added OsO4 (200 mg, 0.79 mmol) and NaIO4 (3.2 g,
15 mmol). After the solution was stirred at rt for 2 h, additional
NaIO4 (1 g, 4.6 mmol) was added and stirred for another 1 h,
followed by the addition of ethyl acetate (200 mL) and water
(100 mL). The organic layer was separated and dried to yield
the crude aldehyde (1.5 g). MS: 372.9/374.9 (MH+).
To the crude aldehyde (1.5 g, 5.1 mmol) in acetic acid (20
mL) was added bromine (0.28 mL, 5.5 mmol) in acetic acid (5
mL) dropwise over 10 min. The mixture was stirred at rt for
1 h and then concentrated and resuspended in ethyl acetate
(200 mL). The organic layer was washed with water (2 × 100
mL), dried, concentrated, and purified by column chromatog-
raphy with silica gel to give the compound 11 as a white solid
(1.1 g, 58%). 1H NMR (CDCl3): δ 2.55 (s, 3H), 4.87 (s, 2H),
5.33 (s, 2H), 6.90-6.96 (m, 2H), 7.26-7.33 (m, 1H), 9.59 (s,
1H). MS (CI) m/z 373.0/375.0 (MH+).
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-eth yl)-a m in o]-eth yl}-5-br om o-6-m eth ylu r a cil (12). To
a mixture of the aldehyde 11 (6.71 g, 18.0 mmol) and N-methyl-
2-(2-pyridyl)ethylamine (2.5 g, 18.3 mmol) in 1,2-dichloroeth-
ane (100 mL) was added NaBH(OAc)3 (7.6 g, 36 mmol) in 4
portions within 10 min. The mixture was stirred at rt for 2 h
and then diluted with water (100 mL) and chloroform (200
mL) with stirring for 5 min. The organic layer was separated,
dried over Na2SO4, and concentrated to yield a brown oil, which
was purified by silica gel chromatography using the CHCl3/
MeOH system to yield the desired product 12 (6.5 g, 73%). 1H
NMR (CDCl3): δ 2.39 (s, 3H), 2.50 (s, 3H), 2.70 (t, J ) 7.5 Hz,
2H), 2.78-2.95 (m, 4H), 4.14 (t, J ) 7.5 Hz, 2H), 5.32 (s, 2H),
6.69-6.87 (m, 2H), 7.06-7.13 (m, 1H), 7.20 (d, J ) 7.5 Hz,
1H), 7.25-7.31 (m, 1H), 7.58 (dt, J ) 7.5, 1.8 Hz, 1H), 8.50 (d,
J ) 4.5 Hz, 1H). MS (CI) m/z 493.1/495.1 (MH+).
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-p h en yl-6-m et h yl-u r a cil (13a ).
P r oced u r e A: To a sealable tube containing 12 (49.3 mg, 0.1
mmol) were added phenylboronic acid (18 mg, 0.15 mmol) and
K2CO3 (47.4 mg, 0.3 mmol), followed by addition of toluene (5
mL) and water (2 mL). The slurry was degassed by N2 for 10
min, and then Pd(Ph3P)4 (12 mg, 0.01 mmol) was added. The
tube was sealed and heated at 90 °C overnight. After being
cooled to rt, the organic layer was separated and concentrated.
The crude product was purified by prep TLC plate (CHCl3/
MeOH/NH4OH ) 850/150/2) to give 13a (24 mg, 49%). 1H NMR
(CDCl3): δ 2.14 (s, 3H), 2.41 (s, 3H), 2.72-2.78 (m, 2H), 2.84-
2.93 (m, 4H), 4.15 (t, J ) 7.2 Hz, 2H), 5.29 (s, 2H), 6.90 (t, J
) 8.1 Hz, 2H), 7.06-7.41 (m, 8H), 7.56 (dt, J ) 1.8 and 7.9
Hz, 1H), 8.48 (d, J ) 4.4 Hz, 1H). MS (CI) m/z 491.1 (MH+).
Anal. (C28H28F2N4O2‚0.4MeOH) C, H, N.
1H). HRMS (FAB) m/z calcd for C29H28F2N4O4 (MH+) 535.2157,
found 535.2134.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-(3,4-et h ylen ed ioxy-p h en yl)-6-
m eth yl-u r a cil (13d ). P r oced u r e B: To a sealable tube
containing 12 (49.3 mg, 0.1 mmol) was added 3,4-ethylene-
dioxyphenylboronic acid pinacol ester (45 mg, 0.16 mmol),
followed by addition of toluene (2 mL), EtOH (0.8 mL), K2CO3
(0.12 mL of a 2 M solution in water), and Ba(OH)2 (40 µL of a
saturated aqueous solution). The slurry was bubbled with N2
for 10 min, and then Pd(Ph3P)4 (12 mg, 0.01 mmol) was added.
The tube was sealed and heated at 90 °C for 12 h. After being
cooled to ambient temperature, the organic layer was sepa-
rated and concentrated. The crude product was purified by
prep TLC plate (CHCl3/MeOH/NH4OH ) 850/150/2) to give
13d (38 mg, 69%). 1H NMR (CDCl3): δ 2.14 (s, 3H), 2.39 (s,
3H), 2.70-2.75 (m, 2H), 2.78-2.88 (m, 2H), 2.90-2.98 (m, 2H),
4.12 (m, 2H), 4.25 (s, 4H), 5.27 (s, 2H), 6.65-6.70 (m, 2H),
6.85-6.92 (m, 3H), 7.06-7.10 (m, 1H), 7.18-7.29 (m, 2H), 7.56
(dt, J ) 2.0 and 7.6 Hz, 1H), 8.48 (d, J ) 4.4 Hz, 1H). MS (CI)
m/z 549.2 (MH+). Anal. (C30H30F2N4O4) C, H; N: calcd, 10.21;
found, 9.29.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-[3,4-(2,2-d iflu or o-m et h ylen e-
d ioxy)-p h en yl]-6-m eth yl-u r a cil (13e). Prepared according
1
to Procedure B, 26 mg, 46%. H NMR (CDCl3): δ 2.15 (s, 3H),
2.39 (s, 3H), 2.70-2.75 (m, 2H), 2.82-2.91 (m, 4H), 4.13 (t, J
) 7.0 Hz, 2H), 5.28 (s, 2H), 6.87-6.93 (m, 3H), 7.05-7.30 (m,
5H), 7.57 (dt, J ) 1.8 and 7.9 Hz, 1H), 8.48-8.50 (m, 1H). MS
(CI) m/z 571.0 (MH+). Anal. (C29H26F4N4O4) C, H, N.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-eth yl)-a m in o]-eth yl}-5-(4-m eth oxy-p h en yl)-6-m eth yl-
u r a cil (13f). Prepared according to Procedure A, 14 mg, 27%.
1H NMR (CDCl3): δ 2.15 (s, 3H), 2.40 (s, 3H), 2.72-2.77 (m,
2H), 2.83-2.93 (m, 4H), 3.81 (s, 3H), 4.13 (dd, J ) 7.0 and
14.1 Hz, 2H), 5.28 (s, 2H), 6.88-6.93 (m, 4H), 7.10-7.26 (m,
5H), 7.56 (dt, J ) 1.8 and 7.9 Hz, 1H), 8.48 (d, J ) 4.1 Hz,
1H). HRMS (FAB) m/z calcd for C29H30F2N4O3 (MH+) 521.2364,
found 521.2339.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-(4-et h oxy-p h en yl)- 6-m et h yl-
u r a cil (13g). Prepared according to Procedure A, 22 mg, 42%.
1H NMR (CDCl3): δ 1.25 (t, J ) 7.0 Hz, 3H), 2.14 (s, 3H), 2.40
(s, 3H), 2.71-2.77 (m, 2H), 2.83-2.92 (m, 4H), 4.03 (q, J )
7.0 Hz, 2H), 4.13 (t, J ) 7.0 Hz, 2H), 5.28 (s, 2H), 6.88-6.93
(m, 4H), 7.08-7.29 (m, 5H), 7.56 (dt, J ) 1.8 and 7.9 Hz, 1H),
8.48 (d, J ) 4.5 Hz, 1H). HRMS (FAB) m/z calcd for
C
30H32F2N4O3 (MH+) 535.2521, found 535.2494.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-(3,4-d im et h oxy-p h en yl)-6-m e-
th ylu r a cil (13h ). Prepared according to Procedure A, 19 mg,
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-(3-m et h oxyp h en yl)-6-m et h yl-
u r a cil d i-tr iflu or oa ceta te (13b). To 12 (49.3 mg, 0.1 mmol)
in toluene/H2O (3 mL/2 mL) in a sealable tube were added
3-methoxyphenylboronic acid (30.2 mg, 0.2 mmol) and K2CO3
(47.4 mg, 0.3 mmol). The mixture was degassed with N2 for
10 min, and then Pd (PPh3)4 (11 mg, 0.01 mmol) was added.
The mixture was sealed and stirred vigorously at 90 °C for 14
h. After being cooled to rt, the aqueous layer was removed and
the organic layer was concentrated and purified by Prep LC-
MS to obtain the desired product 13b (45 mg, 60%) as TFA
1
34%. H NMR (CDCl3): δ 2.16 (s, 3H), 2.50 (s, 3H), 2.77-2.83
(m, 2H), 2.93-3.02 (m, 4H), 3.84 (s, 3H), 3.85 (s, 3H), 4.18 (t,
J ) 7.0 Hz, 2H), 5.28 (s, 2H), 6.71-6.93 (m, 3H), 7.07-7.36
(m, 5H), 7.56 (dt, J ) 1.8 and 7.5 Hz, 1H), 8.48 (d, J ) 4.0 Hz,
1H). HRMS (FAB) m/z calcd for C30H32F2N4O4 (MH+) 551.2469,
found 551.2459.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-eth yl)-a m in o]-eth yl}-5-(3,4,5-tr im eth oxy-ph en yl)-6-m e-
th ylu r a cil (13i). Prepared according to Procedure A, 45 mg,
1
77%. H NMR (CDCl3): δ 2.15 (s, 3H), 2.40 (s, 3H), 2.72-2.77
1
salt. H NMR (CDCl3): δ 2.10 (s, 3H), 3.00 (s, 3H), 3.45-3.78
(m, 2H), 2.83-2.93 (m, 4H), 3.83 (s, 6H), 3.85 (s, 3H), 4.15 (t,
J ) 7.0 Hz, 2H), 5.28 (s, 2H), 6.39 (s, 2H), 6.91 (t, J ) 8.3 Hz,
2H), 7.07-7.30 (m, 3H), 7.57 (dt, J ) 1.8 and 7.9 Hz, 1H), 8.48
(d, J ) 4.1 Hz, 1H). Anal. (C31H34F2N4O5) C, H, N.
(m, 6H), 3.80 (s, 3H), 4.30 (br s, 2H), 5.25 (s, 2H), 6.81-6.83
(m, 2H), 6.88-6.95 (m, 3H), 7.24-7.35 (m, 2H), 7.60 (m, 1H),
7.89 (d, J ) 7.0 Hz, 1H), 8.13 (t, J ) 8.0 Hz, 1H), 8.62 (s, 1H).
HRMS (FAB) m/z calcd for C29H30F2N4O3 (MH+) 521.2364,
found 521.2369. Anal. (C29H30F2N4O3‚2TFA‚H2O) C, H, N.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-eth yl)-a m in o]-eth yl}-5-(3-m eth oxym eth yl-p h en yl)- 6-
m eth ylu r a cil (13j). Prepared according to Procedure B, 24
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-(3,4-m et h ylen ed ioxy-p h en yl)-
6-m eth yl-u r a cil (13c). Prepared according to Procedure A,
37 mg, 70%. 1H NMR (CDCl3): δ 2.15 (s, 3H), 2.40 (s, 3H),
2.71-2.77 (m, 2H), 2.84-2.92 (m, 4H), 4.13 (t, J ) 7.0 Hz, 2H),
5.28 (s, 2H), 5.96 (s, 2H), 6.63 (t, J ) 7.9 Hz, 2H), 6.81-7.30
(m, 6H), 7.58 (dt, J ) 1.8 and 7.9 Hz, 1H), 8.48 (d, J ) 4.2 Hz,
1
mg, 45%. H NMR (CDCl3): δ 2.13 (s, 3H), 2.43 (s, 3H), 2.75-
2.82 (m, 2H), 2.88-3.01 (m, 4H), 3.39 (s, 3H), 4.14-4.18 (m,
2H), 4.42 (d, J ) 13.6 Hz, 2H), 5.28 (s, 2H), 6.73-6.93 (m,
3H), 7.08-7.47 (m, 5H), 7.56 (dt, J ) 1.8 and 7.9 Hz, 1H),
7.76-7.81 (m, 1H), 8.48 (d, J ) 4.1 Hz, 1H). HRMS (FAB) m/z
calcd for C30H32F2N4O3 (MH+) 535.2520, found 535.2499.

Gonadotropin-Releasing Hormone Receptor Antagonists
J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5 1267
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-(4-m et h oxym et h yl-p h en yl)-6-
m eth ylu r a cil (13k ). Prepared according to Procedure B, 13
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-eth yl)-a m in o]-eth yl}-5-(2-flu or o-3-m eth oxy-p h en yl)-6-
m eth ylu r a cil (13s). Prepared according to Procedure A, 33
1
1
mg, 62%. H NMR (CDCl3): δ 2.15 (s, 3H), 2.46 (s, 3H), 2.61-
mg, 25%. H NMR (CDCl3): δ 2.14 (s, 3H), 2.40 (s, 3H), 2.72-
2.69 (m, 2H), 3.10-3.17 (m, 4H), 3.89 (s, 3H), 4.23-4.28 (m,
2H), 5.29 (s, 2H), 6.84-7.32 (m, 8H), 7.59 (td, J ) 6.0 and 1.8
Hz, 1H), 8.48 (d, J ) 4.8 Hz, 1H). HRMS (FAB) m/z calcd for
2.77 (m, 2H), 2.82-2.93 (m, 4H), 3.40 (s, 3H), 4.13 (t, J ) 7.0
Hz, 2H), 4.46 (s, 2H), 5.28 (s, 2H), 6.88-6.93 (m, 2H), 7.06-
7.37 (m, 7H), 7.56 (dt, J ) 1.8 and 7.9 Hz, 1H), 8.48 (d, J )
4.3 Hz, 1H). HRMS (FAB) m/z calcd for C30H32F2N4O3 (MH+)
535.2520, found 535.2510.
C
29H29F3N4O3 (MH+) 539.2272, found 539.2268.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-e t h yl)-a m in o]-e t h yl}-5-(4-vin yl-p h e n yl)-6-m e t h yl-
u r a cil (13t). Prepared according to Procedure A, 24 mg, 46%.
1H NMR (CDCl3): δ 2.15 (s, 3H), 2.40 (s, 3H), 2.71-2.77 (m,
2H), 2.83-2.93 (m, 4H), 4.14 (t, J ) 7.2 Hz, 2H), 5.23 (d, J )
11.0 Hz, 1H), 5.29 (s, 2H), 5.76 (d, J ) 17.5 Hz, 1H), 6.70 (dd,
J ) 11.0 and 17.5 Hz, 1H), 6.91 (t, J ) 8.3 Hz, 2H), 7.06-7.43
(m, 7H), 7.57 (dt, J ) 1.8 and 7.5 Hz, 1H), 8.49 (d, J ) 4.7 Hz,
1H). HRMS (FAB) m/z calcd for C30H30F2N4O2 (MH+) 517.2415,
found 517.2391.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-e t h yl)-a m in o]-e t h yl}-5-(n a p h t h a le n -2-yl)-6-m e t h yl-
u r a cil (13u ). Prepared according to Procedure A, 19 mg, 35%.
1H NMR (CDCl3): δ 2.18 (s, 3H), 2.41 (s, 3H), 2.72-2.78 (m,
2H), 2.84-2.95 (m, 4H), 4.17 (t, J ) 7.5 Hz, 2H), 5.32 (s, 2H),
6.93 (t, J ) 8.1 Hz, 2H), 7.06-7.88 (m, 11H), 8.49 (d, J ) 3.9
Hz, 1H). MS (CI) m/z 541.1 (MH+). Anal. (C32H30F2N4O2‚
0.4MeOH) C, H, N.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-(d iben zofu r a n -3-yl)-6-m et h yl-
u r a cil (13v). Prepared according to Procedure A, 30 mg, 52%.
1H NMR (CDCl3): δ 2.14 (s, 3H), 2.40 (s, 3H), 2.76-2.94 (m,
6H), 4.18 (t, J ) 7.0 Hz, 2H), 5.25 (d, J ) 16.2 Hz, 1H), 5.46
(d, J ) 16.2 Hz, 1H), 6.93 (t, J ) 8.3 Hz, 2H), 7.05-7.56 (m,
9H), 7.93 (dd, J ) 2.2 and 6.6 Hz, 2H), 8.48 (d, J ) 4.1 Hz,
1H). MS (CI) m/z 581.1 (MH+). Anal. (C34H30F2N4O3‚0.5MeOH)
C, H, N.
1-(2,6-Diflu or o-ben zyl)-3-(3-oxo-bu tyr yl)-u r ea (18). Dike-
tene (12.9 mL, 166.7 mmol, reagent stabilized with CuSO4)
was added dropwise over 10 min to a stirred solution of 2,6-
difluorobenzyl urea (15, 28.18 g, 151.5 mmol) in pyridine (300
mL), cooled at 0 °C (ice/water bath) under N2. The resulting
gold-yellow solution was allowed to reach room temperature
and stirred for 24 h. The resulting suspension was filtered,
the solids were rinsed with diethyl ether and dried under
vacuum to give 18 (24.38 g, 60%). 1H NMR (CDCl3): δ 2.12 (s,
3H), 3.55 (s, 2H), 4.45 (d, J ) 5.7 Hz, 2H), 7.06-7.15 (m, 2H),
7.35-7.45 (m, 1H), 8.60 (br, 1H), 10.42 (br, 1H). MS (CI) m/z
271.1 (MH+).
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-et h yl)-a m in o]-et h yl}-5-(4-m et h ylsu lfa n yl-p h en yl)-6-
m eth ylu r a cil (13l). Prepared according to Procedure A, 37
1
mg, 69%. H NMR (CDCl3): δ 2.14 (s, 3H), 2.40 (s, 3H), 2.48
(s, 3H), 2.72-2.77 (m, 2H), 2.83-2.93 (m, 4H), 4.13 (t, J ) 7.0
Hz, 2H), 5.26 (s, 2H), 6.75-6.93 (m, 3H), 7.07-7.47 (m, 5H),
7.58 (dt, J ) 1.8 and 7.8 Hz, 1H), 7.78 (dd, J ) 3.5 and 8.3 Hz,
1H), 8.49 (d, J ) 4.2 Hz, 1H). HRMS (FAB) m/z calcd for
C29H30F2N4O2S (MH+) 537.2136, found 537.2137.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-eth yl)-a m in o]-eth yl}-5-(p-tolyl)-6-m eth ylu r a cil (13m ).
Prepared according to Procedure A, 24 mg, 47%. 1H NMR
(CDCl3): δ 2.14 (s, 3H), 2.35 (s, 3H), 2.40 (s, 3H), 2.71-2.77
(m, 2H), 2.83-2.93 (m, 4H), 4.14 (t, J ) 7.0 Hz, 2H), 5.28 (s,
2H), 6.90 (t, J ) 8.3 Hz, 2H), 7.06-7.30 (m, 7H), 7.56 (dt, J )
1.8 and 7.8 Hz, 1H), 8.48 (d, J ) 4.2 Hz, 1H). MS (CI) m/z
505.0 (MH+). Anal. (C29H30F2N4O2‚0.4MeOH) C, H, N.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-eth yl)-a m in o]-eth yl}-5-(4-isop r op yl-p h en yl)-6-m eth yl-
u r a cil (13n ). Prepared according to Procedure A, 28 mg, 52%.
1H NMR (CDCl3): δ 1.24 (d, J ) 7.0 Hz, 6H), 2.15 (s, 3H), 2.39
(s, 3H), 2.71-2.76 (m, 2H), 2.83-2.93 (m, 5H), 4.13 (t, J ) 7.3
Hz, 2H), 5.29 (s, 2H), 6.90 (t, J ) 8.3 Hz, 2H), 7.06-7.30 (m,
7H), 7.55 (dt, J ) 1.8 and 7.5 Hz, 1H), 8.49 (d, J ) 4.8 Hz,
1H). MS (CI) m/z 533.1 (MH+). Anal. (C31H34F2N4O2‚H2O) C,
H, N.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-eth yl)-a m in o]-eth yl}-5-(3-isop r op yl-p h en yl)-6-m eth yl-
u r a cil (13o). Prepared according to Procedure A, 22 mg, 41%.
1H NMR (CDCl3): δ 1.25 (d, J ) 6.6 Hz, 6H), 2.14 (s, 3H), 2.40
(s, 3H), 2.72-2.78 (m, 2H), 2.84-2.93 (m, 5H), 4.13 (t, J ) 7.5
Hz, 2H), 5.29 (s, 2H), 6.91 (t, J ) 8.3 Hz, 2H), 6.99-7.33 (m,
7H), 7.56 (dt, J ) 1.8 and 7.5 Hz, 1H), 8.49 (d, J ) 4.2 Hz,
1H). HRMS (FAB) m/z calcd for C31H34F2N4O2 (MH+) 533.2728,
found 533.2726.
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-e t h yl)-a m in o]-e t h yl}-5-(3-ch lor o-p h e n yl)-6-m e t h yl-
u r a cil (13p ). Prepared according to Procedure A, 28 mg, 53%.
1H NMR (CDCl3): δ 2.14 (s, 3H), 2.39 (s, 3H), 2.70-2.75 (m,
2H), 2.83-2.92 (m, 4H), 4.12 (t, J ) 7.0 Hz, 2H), 5.29 (s, 2H),
6.91 (t, J ) 7.9 Hz, 2H), 7.06-7.33 (m, 7H), 7.56 (dt, J ) 1.8
and 7.9 Hz, 1H), 8.49 (d, J ) 4.6 Hz, 1H). MS (CI) m/z 524.8
(MH+). Anal. (C28H27ClF2N4O2) C, H, N.
1-(2,6-Diflu or o-ben zyl)-6-m eth yl-u r a cil (16). The acyl
urea obtained above (18, 24.3 g, 90.0 mmol) was suspended
in glacial acetic acid (180 mL) and the mixture refluxed for 1
h. The resulting solution was then cooled and poured onto ice/
water (∼1.5 L). The white solids were collected by vacuum
filtration and dried in a vacuum oven at 50 °C for 24 h to give
1
16 (21.3 g, 94%). H NMR (DMSO-d6): δ 2.22 (s, 3H), 5.07 (s,
2H), 5.53 (s, 1H), 7.06-7.13 (m, 2H), 7.36-7.42 (m, 1H), 11.24
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-e t h yl)-a m in o]-e t h yl}-5-(3-flu or o-p h e n yl)-6-m e t h yl-
u r a cil (13q). Prepared according to Procedure A, 15 mg, 29%.
1H NMR (CDCl3): δ 2.14 (s, 3H), 2.40 (s, 3H), 2.71-2.76 (m,
2H), 2.84-2.93 (m, 4H), 4.13 (t, J ) 7.3 Hz, 2H), 5.29 (s, 2H),
6.88-7.39 (m, 9H), 7.58 (dt, J ) 1.8 and 7.5 Hz, 1H), 8.48 (d,
J ) 4.2 Hz, 1H). HRMS (FAB) m/z calcd for C28H27F3N4O2
(MH+) 509.2164, found 509.2187.
(s, 1H). MS (CI) m/z 253.0 (MH+).
5-Br om o-1-(2,6-d iflu or o-b en zyl)-6-m et h yl-u r a cil (19).
To a solution of 16 (1.26 g, 5.0 mmol) in acetic acid (20 mL)
was added bromine (0.28 mL, 5.5 mmol) in acetic acid (5 mL)
dropwise over 10 min. The mixture was stirred at rt for 1 h
and then purged with N2 to remove most of the bromine. The
resulting solid was collected by filtration and washed with
ether (three times) to give compound 19 as a white solid (1.46
g, 88%). 1H NMR (DMSO-d6): δ 2.46 (s, 3H), 5.20 (s, 2H), 7.06-
7.12 (m, 2H), 7.36-7.46 (m, 1H), 11.86 (s, 1H). MS (CI) m/z
330.0/332.9 (MH+).
5-Br om o-1-(2,6-d iflu or oben zyl)-3-[2-(S)-N-Boc-a m in o-
p r op yl]-6-m eth ylu r a cil (S-20). A solution of N-(tert-butyl-
oxycarbonyl)-L-alaninol (1.75 g, 10 mmol) in anhydrous THF
(15 mL) was treated with 5-bromo-1-(2,6-difluorobenzyl)-6-
methyluracil (19, 3.31 g, 10 mmol) and triphenylphosphine
(3.15 g, 12 mmol) at ambient temperature, and then di-tert-
butylazo dicarboxylate (2.76 g, 12 mmol) was introduced. The
reaction mixture was stirred at ambient temperature for 16
1-(2,6-Diflu or ob en zyl)-3-{2-[N-m et h yl-N-(2-p yr id in -2-
yl-e t h yl)-a m in o]-e t h yl}-5-(2-flu or o-p h e n yl)-6-m e t h yl-
u r a cil Tr iflu or oa ceta te (13r ). Prepared according to Pro-
cedure A and then purified on Prep LC-MS to give 13r (35
1
mg, 56%) as TFA salt. H NMR (CDCl3): δ 2.17 (s, 3H), 3.03
(s, 3H), 3.47-3.55 (m, 2H), 3.65-3.76 (m, 4H), 4.27-4.46 (m,
2H), 5.15 (d, J ) 15.9 Hz, 1H), 5.37 (d, J ) 15.9 Hz, 1H), 6.92
(t, J ) 8.3 Hz, 2H), 7.08-7.36 (m, 5H), 7.69 (t, J ) 6.6 Hz,
1H), 7.98 (d, J ) 7.9 Hz, 1H), 8.23 (t, J ) 7.7 Hz, 1H), 8.65 (d,
J ) 5.2 Hz, 1H). HRMS (FAB) m/z calcd for C28H27F3N4O2
(MH+) 509.2164, found 509.2169.

1268 J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5
Guo et al.
1
h, and volatiles were evaporated. The residue was partitioned
between saturated NaHCO3/H2O and EtOAc. The organic layer
was dried (sodium sulfate), evaporated, and purified by flash
chromatography (silica, 1:2 EtOAc/hexanes) to give compound
TFA salt, 203 mg, 56%. H NMR (CDCl3): δ 1.25 and 1.27 (d,
J ) 6.4 Hz, 3H), 2.13 and 2.14 (s, 3H), 3.39 and 3.48 (m, 1H),
3.87 and 3.88 (s, 3H), 3.96 (d, J ) 9.6 Hz, 1H), 4.02 (m, 1H),
4.15 and 4.16 (d, J ) 14.0 Hz, 1H), 4.31 (m, 1H), 5.10 and
5.22 (d, J ) 16.0 Hz, 1H), 5.25 and 5.28 (d, J ) 16.0 Hz, 1H),
6.78 (m, 1H), 6.88 (t, J ) 8.4 Hz, 2H), 6.96 (m, 1H), 7.09 (m,
1H), 7.22-7.36 (m, 6H). MS (CI) m/z 524.1 (MH+). Anal.
(C29H28F3N3O3‚TFA) C, H, N.
1
S-20 (4.69 g, 96.1%). H NMR (CDCl3): δ 1.15 (d, 3H, J ) 6.3
Hz), 1.36 (s, 9H), 2.51 (s, 3H), 3.86 (d, J ) 9.3 Hz, 1H), 4.09-
4.15 (m, 2H), 4.75-4.80 (m, 1H), 5.15 (d, J ) 15.6 Hz, 1H),
5.46 (d, J ) 15.6 Hz, 1H), 6.90 (t, J ) 8.2 Hz, 2H), 7.22-7.32
(m, 1H). MS (CI) m/z 388.0, 390.0 (MH+ - Boc).
1-(2,6-Diflu or oben zyl)-3-[2-(R)-2-p yr id ylm eth yla m in o-
p r op yl]-5-(2-flu or o-3-m eth oxyp h en yl)-6-m eth ylu r a cil Di-
h yd r och lor id e (R-26b). P r oced u r e E: A solution of R-24
(216 mg, 0.5 mmol) in MeOH (5 mL) was added 2-pyridine-
carboxyaldehyde (80 mg, 0.75 mmol), and the reaction mixture
was stirred at ambient temperature for 10 h. NaBH4 (56 mg,
1.5 mmol) was added, and the reaction mixture was kept at
ambient temperature for 10 min. Volatiles were evaporated,
and the residue was partitioned between saturated NaHCO3/
water and dichloromethane. The organic layer was dried
(sodium sulfate), evaporated, and purified by flash chroma-
tography to give R-26b, which was converted to the HCl salt
5-Br om o-1-(2,6-d iflu or oben zyl)-3-[2-(R)-N-Boc-a m in o-
p r op yl]-6-m eth ylu r a cil (R-20). The procedure used for
preparation of S-20 was repeated here with N-(tert-butyloxy-
carbonyl)-D-alaninol (1.75 g, 10 mmol) and 5-bromo-1-(2,6-
difluorobenzyl)-6-methyluracil (19, 3.31 g, 10 mmol) as the
1
starting materials to give compound R-20 (4.61 g, 94.2%). H
NMR (CDCl3): δ 1.15 (d, 3H, J ) 6.0 Hz), 1.46 (s, 9H), 2.51 (s,
3H), 3.83 (d, J ) 9.3 Hz, 1H), 4.09-4.15 (m, 2H), 4.76-4.81
(m, 1H), 5.15 (d, J ) 15.8 Hz, 1H), 5.46 (d, J ) 15.8 Hz, 1H),
6.90 (t, J ) 8.1 Hz, 2H), 7.22-7.32 (m, 1H). MS (CI) m/z 388.0,
390.0 (MH+-Boc).
1
3-[2-(R)-Am in o-p r op yl]-1-(2,6-d iflu or o-ben zyl)-5-(2-flu -
or o-3-m eth oxy-p h en yl)-6-m eth ylu r a cil Tr iflu or oa ceta te
(R-24). To a solution of compound R-20 (1.0 g, 2.05 mmol) in
bezene/EtOH/ethylene glycol dimethyl ether (20/2/22 mL) were
added 2-fluoro-3-methoxyphenylboronic acid (435 mg, 2.56
mmol) and saturated Ba(OH)2/water (∼0.5 M, 15 mL). The
reaction mixture was deoxygenated with N2 for 10 min,
tetrakis(triphenylphosphine) palladium (0) (242 mg, 0.21
mmol) was added, and the reaction mixture was heated at 80
°C overnight under the protection of N2. The reaction mixture
was partitioned between brine and EtOAc. The organic layer
was dried (sodium sulfate), evaporated, and purified by flash
chromatography (silica, 40% EtOAc/hexanes) to give compound
R-22 (450 mg, 41.2%): MS (CI) m/z 434.2 (MH+ - Boc).
A solution of R-22 (267 mg, 0.5 mmol) in dichloromethane
(2 mL) was added TFA (2 mL), and the reaction mixture was
stirred at ambient temperature for 1 h. Volatiles were evapo-
rated, and the residue was partitioned between saturated
NaHCO3/water and EtOAc. The organic layer was dried
(sodium sulfate) and evaporated to give compound R-24 (241
mg, 97%); a sample of the product was purified by reverse
phase HPLC (C-18 column, 15-75% ACN/water) to give a TFA
salt: 1H NMR (CDCl3): δ 1.07 and 1.17 (d, J ) 6.6 Hz, 3H),
2.13 and 2.14 (s, 3H), 3.86 (s, 3H), 3.81-3.98 (m, 2H), 4.23
and 4.35 (dd, J ) 14.0, 9.6 Hz, 1H), 5.12-5.36 (m, 2H), 6.73-
7.33 (m, 6H), 7.66 (br s, 2H). MS (CI) m/z 434.10 (MH+).
3-(2-(S)-Am in o-p r op yl)-1-(2,6-d iflu or o-ben zyl)-5-(2-flu -
or o-3-m eth oxy-p h en yl)-6-m eth yl-u r a cil Tr iflu or oa ceta te
(S)-24. The procedure used for the preparation of R-24 was
repeated here with S-20 (978 mg, 2 mmol) as the starting
materials to give compound S-24 in two steps (390 mg, 45%);
a sample was purified on HPLC as the TFA salt: 1H NMR
(CDCl3): δ 1.09 and 1.18 (d, J ) 6.6 Hz, 3H), 2.13 and 2.14 (s,
3H), 3.86 (s, 3H), 3.80-3.97 (m, 2H), 4.23 and 4.35 (dd, J )
14.0, 9.6 Hz, 1H), 5.11-5.38 (m, 2H), 6.75-7.31 (m, 6H), 7.73
(br s, 2H). MS (CI) m/z 434.10 (MH+).
Gen er a l P r oced u r e C for th e P r ep a r a tion of Com -
p ou n d s 26. To compound 24 (216 mg, 0.5 mmol) in dichloro-
ethane (10 mL) was added a carbonyl compound (0.6 mmol),
and the mixture was stirred at room temperature for 10 min.
Then NaBH(OAc)3 (255 mg, 1.2 mmol) was added, and the
mixture was further stirred for 10 h and then diluted with
DCM, washed with water, dried (sodium sulfate), and evapo-
rated. The crude product was purified by flash chromatography
(silica, 20 to 50% EtOAc/hexanes). Alternately, the crude
product was purified by Prep LC-MS to give the desired
compound as a TFA salt.
P r oced u r e D. The free base was converted to the hydro-
chloride salt by dissolution in dichloromethane and addition
of 1 N HCl solution in ether, followed by removal of the solvent
under vacuum to provide the HCl salt as amorphous solid.
1-(2,6-Diflu or oben zyl)-3-[2-(S)-ben zyla m in op r op yl]-5-
(2-flu or o-3-m et h oxyp h en yl)-6-m et h ylu r a cil Tr iflu or o-
a ceta te (S-26a ). Prepared according to Procedure C as the
as described in procedure D (160 mg, 46%). H NMR (DMSO-
d6): δ 1.25 and 1.26 (d, J ) 6.8 Hz, 3H), 2.19 (s, 3H), 3.58 (m,
1H), 3.95 (s, 3H), 4.07 (m, 1H), 4.23 (dd, J ) 6.4, 14.0 Hz, 1H),
4.35 (d, J ) 14.0 Hz, 1H), 4.42 (d, J ) 14.0 Hz, 1H), 5.25 (s,
2H), 6.76 (m, 1H), 7.12 (t, J ) 8.4 Hz, 2H), 7.17 (m, 1H), 7.19
(m, 1H), 7.42 (m, 1H), 7.44 (dd, J ) 4.4, 8.2 Hz, 1H), 7.48 (d,
J ) 7.6 Hz, 1H), 7.89 (dt, J ) 2.0, 8.0 Hz, 1H), 8.60 (d, J ) 4.4
Hz, 1H), 9.17 (br s, 2H), 9.98 (br s, 1H). MS (CI) m/z 525.2
(MH+). Anal. (C28H27F3N4O3‚2HCl) C, H, N.
1-(2,6-Diflu or ob en zyl)-3-{2-(R)-[(2-t h ia zolylm et h yl)-
a m in o]p r op yl}-5-(2-flu or o-3-m et h oxyp h en yl)-6-m et h yl-
u r a cil Hyd r och lor id e (R-26d ). Prepared according to Pro-
cedure E and D to give R-26d as the HCl salt (201 mg, 71%).
1H NMR (DMSO-d6): δ 1.25 and 1.27 (d, J ) 5.1 Hz, 3H), 2.19
(s, 3H), 3.60-3.65 (m, 1H), 3.86 (s, 3H), 4.02-4.09 (m, 1H),
4.23 (dd, J ) 10.5, 5.1 Hz, 1H), 4.62-4.67 (m, 2H), 5.25 (s,
2H), 6.74-6.78 (m, 1H), 7.11 (t, J ) 6.3 Hz, 2H), 7.18-7.42
(m, 3H), 7.87 (dd, J ) 0.6, 2.4 Hz, 1H), 7.91 (d, J ) 2.4 Hz,
1H), 9.39 (br s, 1H), 9.54 (br s, 1H). MS (CI) m/z 531.2 (MH+).
Anal. (C26H25F3N4O3S‚HCl‚H2O) C, H, N.
1-(2,6-Diflu or ob en zyl)-3-{2-(S)-[(3-m et h yl-2-t h iop h e-
n ylm eth yl)a m in o]p r op yl}-5-(2-flu or o-3-m eth oxyp h en yl)-
6-m eth ylu r a cil Hyd r och lor id e (S-26e). Prepared according
to Procedure E and D to give S-26e as the HCl salt (151 mg,
52%). 1H NMR (DMSO-d6): δ 1.23-1.28 (m, 3H), 2.20 (s, 3H),
2.23 (s, 3H), 3.60-3.65 (m, 1H), 3.86 (s, 3H), 4.01-4.07 (m,
1H), 4.15-4.21 (m, 1H), 4.33-4.38 (m, 2H), 5.26 (s, 2H), 6.77-
6.81 (m, 1H), 6.93 (d, J ) 3.6 Hz, 1H), 7.11 (t, J ) 6.3 Hz,
2H), 7.19-7.54 (m, 4H), 8.98 (brs, 2H). MS (CI) m/z 502.2
(MH+). Anal. (C28H28F3N3O3S‚HCl) C, H, N.
1-(2,6-Diflu or ob en zyl)-3-{2-(S)-[(5-et h yl-2-fu r a n ylm e-
th yl)a m in o]p r op yl}-5-(2-flu or o-3-m eth oxyp h en yl)-6-m e-
th ylu r a cil Hyd r och lor id e (S-26f). Prepared according to
Procedure E and D to give S-26f as the HCl salt (237 mg, 82%).
1H NMR (DMSO-d6): δ 1.17 (t, J ) 6.7 Hz, 3H), 1.20 and 1.21
(d, J ) 6.7 Hz, 3H), 2.19 (s, 3H), 2.61 (q, J ) 6.7 Hz, 2H),
3.50-3.55 (m, 1H), 3.86 (s, 3H), 4.03-4.08 (m, 1H), 4.14-4.23
(m, 3H), 5.25 (s, 2H), 6.12 (d, J ) 2.4 Hz, 1H), 6.45 (d, J ) 2.4
Hz, 1H), 6.73-6.79 (m, 1H), 7.11 (t, J ) 6.3 Hz, 2H), 7.18-
7.42 (m, 3H), 9.18 (br s, 2H). MS (CI) m/z 542.3 (MH+). Anal.
(C29H30F3N3O4‚HCl) C, H, N.
1-(2,6-Diflu or oben zyl)-3-[2-(S)-(1-eth ylp r op yl)a m in o-
pr opyl]-5-(2-flu or o-3-m eth oxyph en yl)-6-m eth ylu r acil Hy-
d r och lor id e (S-26g). Prepared according to Procedure C and
D to give S-26g as the HCl salt (214 mg, 79%). 1H NMR
(DMSO-d6): δ 0.89 (t, J ) 7.6 Hz, 6H), 1.15 and 1.16 (t, J )
6.0 Hz, 3H), 1.24-1.28 (m, 1H), 1.35-1.39 (m, 1H), 1.43-1.46
(m, 1H), 1.62-1.66 (m, 1H), 2.17 and 2.18 (s, 3H), 3.33 (m,
1H), 3.53-3.58 (m, 1H), 3.86 (s, 3H), 4.03-4.08 (m, 1H), 4.16
(dd, J ) 9.9, 6.0 Hz, 1H), 5.28 (s, 2H), 6.74-6.79 (m, 1H), 7.12
(t, J ) 8.0 Hz, 2H), 7.18-7.42 (m, 3H), 8.60 (br s, 1H), 8.83
(br s, 1H). MS (CI) m/z 504.2 (MH+). Anal. (C27H32F3N3O3‚HCl‚
0.5Et2O) C, H, N.

Gonadotropin-Releasing Hormone Receptor Antagonists
J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5 1269
1-(2,6-Diflu or oben zyl)-3-[2-(S)-cyclopen tylam in opr opyl]-
5-(2-flu or o-3-m eth oxyp h en yl)-6-m eth ylu r a cil Hyd r och lo-
r id e (S-26h ). Prepared according to Procedure C and D to give
(s, 3H), 2.97 (dd, J ) 12.9, 6.0 Hz, 1H), 3.36 (dt, J ) 13.5, 5.4
Hz, 1H), 3.88 (s, 3H), 5.00-5.08 (m, 1H), 5.23-5.39 (m, 2H),
6.78-7.30 (m, 6H). MS (CI) m/z 434.10 (MH+).
1
S-26h as the HCl salt (226 mg, 84%). H NMR (DMSO-d6): δ
1-(2,6-Diflu or oben zyl)-3-[1-(R)-m eth yl-2-(2-m eth yl-ben z-
yla m in o)-eth yl]-5-(2-flu or o-3-m eth oxyp h en yl)-6-m eth yl-
u r a cil Hyd r och lor id e (R-35b). To a solution of R-33 (650
mg, 1.5 mmol) in MeOH (15 mL) was added o-tolualdehyde
(1.80 g, 15 mmol), and the mixture stirred for 1 h at rt. NaBH4
(283 mg, 7.5 mmol) was added and reaction mixture stirred
at ambient temperature for 15 min, concentrated, and purified
by flash chromatography (20% EtOAc/hexanes). The crude
material was treated according to procedure D to yield a white
1.17 (d, J ) 6.8 Hz, 3H), 1.51 (brs, 2H), 1.62 (brs, 2H), 1.71
(br s, 2H), 1.92 (br s, 2H), 2.17 (s, 3H), 3.40-3.45 (m, 1H),
3.57-3.64 (m, 1H), 3.85 (s, 3H), 4.11 (br s, 1H), 4.13 (br s, 1H),
5.27 (s, 2H), 6.74-6.79 (m, 1H), 7.12 (t, J ) 8.4 Hz, 2H), 7.17
(d, J ) 5.2 Hz, 1H), 7.19-7.42 (m, 2H), 9.01 (br s, 1H), 9.10
(br s, 1H). MS (CI) m/z 502.2 (MH+). Anal. (C27H30F3N3O3‚HCl)
C, H, N.
1-(2,6-Diflu or oben zyl)-3-[2-(S)-cyclobu tylam in opr opyl]-
5-(2-flu or o-3-m eth oxyp h en yl)-6-m eth ylu r a cil Hyd r och lo-
r id e (S-26i). Prepared according to Procedure C and D to give
1
solid R-35b as the HCl salt (232 mg, 27%). H NMR (CDCl3):
δ 1.41 (d, J ) 6.2 Hz, 3H), 2.07 (s, 3H), 2.28 and 2.29 (s, 3H),
2.94 (t, J ) 10.4 Hz, 1H), 3.84 (s, 3H), 3.93 (m, 1H), 4.06 (d, J
) 13.0 Hz, 1H), 4.23 (d, J ) 13.0 Hz, 1H), 5.14 and 5.15 (d, J
) 16.4 Hz, 1H), 5.25 and 5.32 (d, J ) 16.4 Hz, 1H), 5.45 (m,
1H), 6.73 (m, 1H), 6.87 (m, 2H), 6.92-7.30 (m, 7H). MS (CI)
m/z 538.2 (MH+). Anal. (C30H30F3N3O3‚HCl) C, H, N.
1
S-26i as the HCl salt (202 mg, 77%). H NMR (DMSO-d6): δ
1.12 (d, J ) 5.1 Hz, 3H), 1.72-1.79 (m, 2H), 2.18 (s, 3H), 2.20-
2.26 (m, 4H), 3.37-3.42 (m, 1H), 3.77-3.83 (m, 1H), 3.86 (s,
3H), 4.03-4.08 (m, 2H), 5.27 (s, 2H), 6.75-6.80 (m, 1H), 7.12
(t, J ) 6.3 Hz, 2H), 7.18-7.42 (m, 3H), 9.13 (br s, 2H). MS
(CI) m/z 488.3 (MH+). Anal. (C26H28F3N3O3‚HCl‚0.5H2O) C, H,
N.
1-(2,6-Diflu or o-ben zyl)-5-(2-flu or o-3-m eth oxy-p h en yl)-
3-[1-(R)-m eth yl-2-(2-m eth oxy-ben zyla m in o)-eth yl]-6-m e-
th ylu r a cil Hyd r och lor id e (R-35c). The procedure used for
the preparation of R-35b was repeated here with R-33 (650
mg, 1.5 mmol) and o-anisaldehyde (2.04 g, 15 mmol) to give
R-35c as a white solid, which was treated under procedure D
1-N-Boc-a m in o-p r op a n -2-ol (S-28). To a solution of (S)-
(+)-1-amino-2-propanol (5.18 g, 67 mmol) and TEA (4.14 g,
70 mmol) in DCM (80 mL) was added di-tert-butyl dicarbonate
(15.5 g, 69 mmol), and the reaction mixture was stirred at 0
°C under N2 for 1 h. The reaction mixture was partitioned
between saturated NaHCO3/H2O and DCM and washed with
brine. The organic layer was dried (sodium sulfate) and
evaporated to give R-28 (11.6 g, 99%). 1H NMR (CDCl3): δ
1.18 (d, 3H, J ) 6.3 Hz), 1.45 (s, 9H), 2.95-3.05 (m, 1H), 3.24-
3.29 (m, 1H), 3.87-3.93 (m, 1H), 5.30 (s, 1H).
3-[2-N-Boc-a m in o-1-(R)-m et h yl-et h yl]-5-b r om o-1-(2,6-
d iflu or o-ben zyl)-6-m eth ylu r a cil (R-29). A solution of 1-N-
Boc-amino-propan-2-ol (S-28, 1.92 g, 11 mmol) in anhydrous
THF (30 mL) was treated with 5-bromo-1-(2,6-difluorobenzyl)-
6-methyluracil (19, 3.31 g, 10.5 mmol) and triphenylphosphine
(4.11 g, 15.7 mmol) at ambient temperature, and then diethyl
azodicarboxylate (2.73 g, 15.7 mmol) was added dropwise. The
reaction mixture was stirred at ambient temperature for 12
h, and volatiles were evaporated. The residue was partitioned
between saturated NaHCO3/H2O and EtOAc. The organic layer
was dried (sodium sulfate), evaporated, and purified by flash
chromatography (silica, 1:9 EtOAc/hexanes) to give compound
R-29 (3.58 g, 70%). 1H NMR (DMSO-d6): δ 1.21 (d, 3H, J )
6.9 Hz), 1.29 (s, 9H), 2.46 (s, 3H), 3.14-3.19 (m, 1H), 3.36-
3.41 (m, 1H), 4.87-4.93 (m, 1H), 5.14 (d, J ) 16.5 Hz, 1H),
5.28 (d, J ) 16.5 Hz, 1H), 7.04-7.09 (m, 2H), 7.33-7.43 (m,
1H), 8.97 (br s, 1H). MS (CI) m/z 388.0, 390.0 (MH+-Boc).
1
to yield the product as the HCl salt (274 mg, 31%). H NMR
(CDCl3): δ 1.60 and 1.61 (d, J ) 6.4 Hz, 3H), 2.08 (s, 3H),
2.84 and 2.94 (m, 1H), 3.58 and 3.67 (s, 3H), 3.62 (m, 1H),
3.87 and 3.90 (s, 3H), 3.87 (m, 1H), 4.41 and 4.43 (m, 1H),
5.16 and 5.17 (d, J ) 16.0 Hz, 1H), 5.26 and 5.27 (d, J ) 16.0
Hz, 1H), 5.43-5.47 (m, 1H), 6.65-7.37 (m, 10H), 8.62 and 8.76
(br s, 1H), 10.7 and 10.8 (br s, 1H). MS (CI) m/z 554.2 (MH+).
Anal. (C30H30F3N3O4‚HCl) C, H, N.
Hu m a n Liver Micr osom es Sta bility Stu d ies. Microso-
mal incubations were performed in the presence of an NADPH-
generating system composed of 10 mM MgCl2, 1 mM NADP+,
46 mM glucose-6-phosphate, and 3 units/mL glucose-6-
phosphate dehydrogenase in 70 mM potassium phosphate
buffer (pH 7.4); all concentrations are relative to the final
incubation volume. The GnRH compounds were added in
DMSO to a final concentration of 50 µM (1% final DMSO
concentration). Incubations were conducted at 37 °C in dupli-
cate and were terminated by the addition of 0.1 mL of acetic
acid in acetonitrile (6:94; v/v) to each 0.25 mL of incubation
volume. Precipitated proteins were removed by centrifugation,
and supernatants were analyzed by HPLC with UV detection.
Separation of parent compound from putative metabolites was
obtained on a Zorbax C8 HPLC column (5 µm, 150 × 4.6 mm)
and a mobile phase gradient from 100% trifluoroacetic acid in
water (1:1000; v/v) to 100% trifluoroacetic acid in acetonitrile
(1:1000; v/v) in 15 min; injection volumes were 50 µL. The
GnRH compounds were detected at UV absorbance wave-
lengths of 240 and 255 nm. Testosterone (Sigma Chemical Co.,
St Louis, MO) was used as a positive control for viability of
the microsomal preparation; testosterone assays were per-
formed as described above, and with UV detection at 240 nm.
Or a l Absor p tion in Mice. Oral bioavailability studies were
conducted in CD-1 mice, and test articles were administered
to the mice by oral gavage and intravenous injection (10 mg/
kg; N ) 3/time point) in water solution. Terminal blood
samples were taken at predetermined time points for compos-
ite sampling. For in vivo first-pass effect studies, the mice were
anesthetized with isoflurane by using either a calibrated
vaporizer and induction chamber, anesthetic chamber, or open
drop method at the time of sampling. A retro-orbital eye bleed
was performed on each mouse so that 300-600 µL of whole
blood was drawn. The mouse was then checked to ensure a
deep anesthesia. The mouse was laid on its back and the
midline of the stomach opened up. The hepatic portal vein was
occluded as near as possible to the liver. A catheter was placed
into the hepatic portal vein such that the catheter was pointed
toward the intestines. A 300-600 µL sample was taken from
the hepatic portal vein using a 600 µL tube that contained
1.2-2 mg EDTA/mL of blood. Blood samples from each dosing
3-[2-N-Boc-a m in o-1-(R)-m et h yl-et h yl]-1-(2,6-d iflu or o-
ben zyl)-5-(2-flu or o-3-m eth oxy-ph en yl)-6-m eth ylu r acil (R-
31). To a solution of compound R-29 (2.83 g, 5.8 mmol) in
toluene/water (20/10 mL) were added 2-fluoro-3-methoxyphe-
nylboronic acid (1.87 g, 11 mmol) and K2CO3 (1.60 g, 11.6
mmol). The reaction mixture was deoxygenated with N2 for
10 min, tetrakis(triphenylphosine)palladium(0) (693 mg, 0.6
mmol) was added, and the reaction mixture was heated at 90
°C for 12 h in a sealed flask under the protection of N2. The
reaction mixture was partitioned between brine and EtOAc.
The organic layer was dried (sodium sulfate), evaporated, and
purified by flash chromatography (silica, 10 to 40% EtOAc/
hexanes) to give compound R-31 (1.64 g, 53%). 1H NMR
(CDCl3): δ 1.41 (s, 9H), 1.59 (s, 3H), 2.11 (s, 3H), 3.28-3.37
(m, 1H), 3.88 (s, 3H), 4.96-5.01 (m, 1H), 5.20-5.31 (m, 3H),
6.79-7.28 (m, 6H), 7.68 (br s, 1H). MS (CI) m/z 434.1 (MH+
Boc).
-
3-[2-Am in o-1-(R)-m eth yl-eth yl]-1-(2,6-d iflu or o-ben zyl)-
5-(2-flu or o-3-m eth oxy-p h en yl)-6-m eth ylu r a cil (R-33). To
a solution of R-31 (1.12 g, 2.1 mmol) in dichloromethane (20
mL) was added TFA (20 mL), and the reaction mixture was
stirred at ambient temperature for 2 h. Volatiles were evapo-
rated, and the residue was partitioned between saturated
NaHCO3/water and EtOAc. The organic layer was dried
(sodium sulfate) and evaporated, to give compound R-33 (863
1
mg, 95%). H NMR (CDCl3): δ 1.43 (d, J ) 6.9 Hz, 3H), 2.11

1270 J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5
Guo et al.
Bioorg. Med. Chem. Lett. 2001, 11, 1727-1731. (d) Chu, L.;
Hutchins, J . E.; Weber, A. E.; Lo, J . L.; Yang, Y. T.; Cheng, K.;
Smith, R. G.; Fisher, M. H.; Wyvratt, M. J .; Goulet, M. T. Initial
structure-activity relationship of a novel class of nonpeptidyl
GnRH receptor antagonists: 2-arylindoles. Bioorg. Med. Chem.
Lett. 2001, 11, 509-513. (e) Chu, L.; Lo, J . L.; Yang, Y. T.; Cheng,
K.; Smith, R. G.; Fisher, M. H.; Wyvratt, M. J .; Goulet, M. T.
SAR studies of novel 5-substituted 2-arylindoles as nonpeptidyl
GnRH receptor antagonists. Bioorg. Med. Chem. Lett. 2001, 11,
515-517. (f) Lin, P.; Marino, D.; Lo, J .-L.; Yang, Y. T.; Cheng,
K.; Smith, R. G.; Fisher, M. H.; Wyvratt, M. J .; Goulet, M. T.
2-(3,5-Dimethylphenyl)tryptamine Derivatives that Bind to the
GnRH Receptor. Bioorg. Med. Chem. Lett. 2001, 11, 1073-76.
(g) Lin, P.; Parikh, M.; Lo, J .-L.; Yang, Y. T.; Cheng, K.; Smith,
R. G.; Fisher, M. H.; Wyvratt, M. J .; Goulet, M. T. Heterocyclic
Derivatives of 2-(3,5-Dimethylphenyl)tryptamine as GnRH Re-
ceptor Antagonists. Bioorg. Med. Chem. Lett. 2001, 11, 1077-
80.
route were taken at predetermined time points for pharma-
cokinetic analysis. All plasma samples were flash frozen in
liquid nitrogen within 10 min of sampling and stored in -76
°C or below until analysis. The bioanalytical method applied
for the measurement of test articles in plasma along with
added internal standard consisted of precipitation with 200
µL of acetonitrile from 50 µL of plasma, centrifugation, and
recovery the supernatant, which was evaporated in a vacuum
and then reconstituted in acetonitrile-water solution before
introduction into an LC-MS/MS system for analysis. The lower
limit of quantification (LLOQ) for the analytical methods was
5 ng/mL of test article in plasma. All pharmacokinetic param-
eters were calculated from a noncompartmental model using
WinNonlin program version 3.2.
Refer en ces
(8) Sasaki, S.; Cho, N.; Nara, Y.; Harada, M.; Endo, S.; Suzuki, N.;
Furuya, S.; Fujino, M. Discovery of a Thieno[2,3-d]pyrimidine-
(1) Huirne, J . A. F.; Lambalk, C. B. Gonadotropin Releasing
Hormone Receptor Antagonists. Lancet 2001, 358, 1793-803.
(2) (a) Matsuo, H.; Baba, Y.; Nair, R. M. G.; Arimura, A.; Schally,
A. V. Structure of the Porcine LH- and FSH-releasing Hormone.
I. The Proposed Amino Acid Sequence. Biochem. Biophys. Res.
Commun. 1971, 43, 1334-1339. (b) Amoss, M.; Burgus, R.;
Blackwell, R. Butcher, Vale. W.; Fellows, R.; Guillemin, R.
Purification, Amino Acid Composition and N-Terminus of the
Hypothalamic Luteinizing Hormone Releasing Factor (LRF) of
Ovine Origin. Biochem. Biophys. Res. Commun. 1971, 44, 205-
210.
(3) Fujino, M.; Fukuda, T.; Shinagawa, S.; Kobayashi, S.; Yamazaki,
I.; Nakayama, R.; Seely, J . H.; White, W. F.; Rippel, R. H.
Synthetic Analogues of Luteinizing Hormone Releasing Hormone
(LHRH) Substituted in Position 6 to 10. Biochem. Biophys. Res.
Commun. 1974, 60, 406-13.
(4) Goulet, M. T. Gonadotropin Releasing Hormone Antagonists. In
Annual Reports in Medicinal Chemistry; Bristol, J . A., Ed.;
Academic Press: New York, 1995; Vol. 30, p 169.
2,4-dione Bearing
a p-Methoxyureidophenyl Moiety at the
6-Position: A Highly Potent and Orally Bioavailable Non-
Peptide Antagonist for the Human Luteinizing Hormone-Releas-
ing Hormone Receptor. J . Med. Chem. 2003, 46, 113-24.
(9) (a) Zhu, Y.-F.; Struthers, R. S.; Connors, P. J ., J r., Gao, Y.; Gross,
T. D.; Saunders: J .; Wilcoxen, K.; Reinhart, G. J .; Ling, N.; Chen,
C. Initial Structure-Activity Relationship Studies of a Novel
Series of Pyrrolo[1, 2-a]pyrimid-7-ones as GnRH Receptor
Antagonists. Bioorg. Med. Chem. Lett. 2002, 12, 339-402. (b)
Zhu, Y.-F.; Wilcoxen, K.; Saunders: J .; Guo, Z.; Gao, Y.; Connors,
P. J ., J r., Gross, T. D.; Tucci, F. C.; Struthers, R. S.; Reinhart,
G. J .; Xie, Q.; Chen, C. A Novel Synthesis of 2-Arylpyrrolo[1,2-
a]pyrimid-7-ones and Their Structure-Activity Relationships as
Potent GnRH Receptor Antagonists. Bioorg. Med. Chem. Lett.
2002, 12, 403-406.
(10) (a) Wilcoxen, K.; Zhu, Y.-F.; Connors, P. J ., J r.; Saunders, J .;
Gross, T. D.; Gao, Y.; Reinhart, G. J .; Struthers, R. S.; Chen, C.
Synthesis and Initial Structure-Activity Relationships of a
Novel Series of Imidazolo[1,2-a]pyrimid-4-ones as Potent GnRH
receptor antagonists. Bioorg. Med. Chem. Lett. 2002, 12, 2179-
2184. (b) Gross, T. D.; Zhu, Y.-F.; Saunders: J .; Gao, Y.; Connors,
P. J ., J r.; Guo, Z.; Struthers, R. S.; Reinhart, G. J .; Chen, C.
Synthesis and Structure-Activity Relationships of a Novel
Series of Imidazolo[1,2-a]pyrimid-4-ones as Potent GnRH recep-
tor antagonists. Bioorg. Med. Chem. Lett. 2002, 12, 2185-2187.
(11) Zhu, Y.-F.; Guo, Z.; Gross, T. D.; Gao, Y.; Connors, P. J ., J r.;
Struthers, R. S.; Xie, Q.; Tucci, F. C.; Reinhart, G. J .; Wu, D.;
Saunders: J .; Chen C. Design and Structure-Activity Relation-
ships of 2-Alkyl-3-Aminomethyl-6-(3-Methoxyphenyl)-7-Methyl-
8-(2-Fluorobenzyl)imidazolo[1,2-a]pyrimid-5-ones as Potent GnRH
Receptor Antagonists. J . Med. Chem. 2003, 46, 1769-72.
(12) (a) Zhu, Y.-F.; Gross, T. D.; Guo, Z.; Connors, P. J ., J r.; Gao, Y.;
Tucci, F. C.; Struthers, R. S.; Reinhart, G. J .; Saunders, J .; Chen,
T. K.; Bonneville, A. L. K.; Chen, C. Identification of 1-Aryl-
methyl-3-(2-aminoethyl)-5-aryluracil as Novel Gonadotropin-
Releasing Hormone Receptor Antagonists. J . Med. Chem. 2003,
46, 2023-26. (b) Guo, Z.; Zhu, Y.-F.; Tucci, F. C.; Gao, Y.;
Struthers, R. S.; Saunders, J .; Gross, T. D.; Xie, Q.; Reinhart,
G. J .; Chen, C. Synthesis and Structure-Activity Relationships
of 1-Arylmethyl-3-(2-Aminopropyl)-5-Aryl-6-Methyluracils as
Potent GnRH Receptor Antagonists. Bioorg. Med. Chem. Lett.
2003, 13, 3311-3315. (c). Tucci, F. C.; Zhu, Y.-F.; Guo, Z.; Gross,
T. D.; Connors, P. J ., J r.; Struthers, R. S.; Reinhart, G. J .;
Saunders, J .; Chen, C. Synthesis and Structure-Activity Rela-
tionships of 1-Arylmethyl-3-(1-Methyl-2-Amino)Ethyl-5-Aryl-6-
Methyluracils as Antagonists of the Human GnRH Receptor.
Bioorg. Med. Chem. Lett. 2003, 13, 3317-3322.
(5) Cho, N.; Harada, M.; Toshihiro, I.; Imada, T.; Matsumoto, H.;
Hayase, Y.; Sasaki, S.; Furuya, S.; Suzuki, N.; Okubo, S.; Ogi,
K.; Endo, S.; Onda, H.; Fujino, M. Discovery of a Novel, Potent,
and Orally active Nonpeptide Antagonist of Human Luteinizing
Hormone -Releasing Hormone (LHRH) Receptor. J . Med. Chem.
1998, 41, 4190-4195.
(6) (a) DeVita, R. J .; Holling, D. D.; Goulet, M. T.; Wyvratt, M. J .;
Fisher, M. H.; Lo, J .-L.; Yang, Y. T.; Cheng, K.; Smith, R. G.
Identification and initial structure-activity relationships of a
novel non-peptide quinolone GnRH receptor antagonist. Bioorg.
Med. Chem. Lett. 1999, 9, 2615-20. (b) DeVita, R. J .; Goulet,
M. T.; Wyvratt, M. J .; Fisher, M. H.; Lo, J .-L.; Yang, Y. T.; Cheng,
K.; Smith, R. G. Investigation of the 4-O-alkylamine substituent
of non-peptide quinolone GnRH receptor antagonists. Bioorg.
Med. Chem. Lett. 1999, 9, 2621-24. (c) Walsh, T. F.; Toupence,
R. B.; Young, J .; Huang, S. X.; Ujjainwalla, F.; DeVita, R. J .;
Goulet, M. T.; Wyvratt, M. J .; Fisher, M. H.; Lo, J .-L.; Ren, N.;
Yudkovitz, J . B.; Yang, Y. T.; Cheng, K.; Smith, R. G. Potent
antagonists of gonadotropin releasing hormone receptors derived
from quinolone-6-carboxamides. Bioorg. Med. Chem. Lett. 2000,
10, 443-47. (d) Young, J . R.; Huang, S. X.; Chen, I.; Walsh, T.
F.; DeVita, R. J .; Wyvratt, M. J .; Goulet, M. T.; Ren, N.; Lo, J .-
L.; Yang, Y. T.; Yudkovitz, J . B.; Cheng, K.; Smith, R. G.
Quinolones as gonadotropin releasing hormone (GnRH) antago-
nists: simultaneous optimization of the C(3)-aryl and C(6)-
substituents. Bioorg. Med. Chem. Lett. 2000, 10, 1723-27. (e)
DeVita, R. J .; Walsh, T. F.; Young, J . R.; J iang J . L.; Ujjainwalla,
F.; Toupence, R. B.; Parikh, M.; Huang, S. X.; Fair, J . A.; Goulet,
M. T.; Wyvratt, M. J .; Lo, J .-L.; Ren, N.; Yudkovitz, J . B.; Yang,
Y. T.; Cheng, K.; Cui, J .; Mount, G.; Rohrer, S.; Schaeffer, J .
M.; Rhodes, L.; Drisko, J . E.; McGowan, E.; MacIntyre, D. E.;
Vincent, S.; Carlin, J . R.; Cameron, J .; Smith, R. G. A Potent,
Nonpeptidyl 1H-Quinolone Antagonist for the Gonadotropin-
Releasing Hormone Receptor. J . Med. Chem. 2001, 44, 917-22.
(7) (a) Ashton, W. T.; Sisco, R. M.; Kieczykowski, G. R.; Yang, Y.
T.; Yudkovitz, J . B.; Cui, J .; Mount, G. R.; Ren, R. N.; Wu, T. J .;
Shen, X.; Lyons, K. A.; Mao, A. H.; Carlin, J . R.; Karanam, B.
V.; Vincent, S. H.; Cheng, K.; Goulet, M. T. Orally Bioavailable,
Indole-based Nonpeptide GnRH Receptor Antagonists With High
Potency and Functional Activity. Bioorg. Med. Chem. Lett. 2001,
11, 2597-2602. (b) Ashton, W. T.; Sisco, R. M.; Yang, Y. T.; Lo,
J . L.; Yudkovitz, J . B.; Cheng, K.; Goulet, M. T. Substituted
indole-5-carboxamides and -acetamides as potent nonpeptide
GnRH receptor antagonists. Bioorg. Med. Chem. Lett. 2001, 11,
1723-1726. (c) Ashton, W. T.; Sisco, R. M.; Yang, Y. T.; Lo, J .
L.; Yudkovitz, J . B.; Gibbons, P. H.; Mount, G. R.; Ren, R. N.;
Butler, B. S.; Cheng, K.; Goulet, M. T. Potent nonpeptide GnRH
receptor antagonists derived from substituted indole-5-carbox-
amides and -acetamides bearing a pyridine side-chain terminus.
(13) Grahner, B.; Winiwarter, S.; Lanzner, W.; Mueller. C. E.
Synthesis and Structure-Activity Relationships of Deazaxan-
thines: Analogues of Potent A1- and A2-Adenosine Receptor
Antagonists. J . Med. Chem. 1994, 37, 1526-1534.
(14) The crystal structure of 16 was also determined (Figure 3).
(15) The enantiomeric excesses (% ee) of compounds S-29 and R-29
were determined by HPLC, using a long Pirkle Covalent (R,R)
WHELK column (25 cm × 4.6 mm), manufactured by REGIS.
The samples were eluted with an isocratic 80:20 (v/v) mixture
of hexanes and ethanol, respectively. For S-29, tR ) 11.10 min,
ee ) >99%. For R-29, tR ) 9.93 min, ee ) >99%.
(16) R-29 was crystallized from slow diffusion of diethyl ether into a
dichloromethane solution. Crystallographic data (excluding
structure factors) for the structure in this paper have been
deposited with the Cambridge Crystallographic Data Centre as
supplementary publication numbers CCDC 204509. Copies of
the data can be obtained, free of charge, on application to CCDC,

Gonadotropin-Releasing Hormone Receptor Antagonists
J ournal of Medicinal Chemistry, 2004, Vol. 47, No. 5 1271
12 Union Road, Cambridge, CB2 1EZ, UK (FAX: +44 (0)1223
336033 or e-mail: deposit@ccdc.cam.ac.uk).
tions. Key compounds were assayed in three to eight indepen-
dent experiments.
(17) Perrin, M. H.; Haas Y.; Rivier, J . E.; Vale, W. W. Gonadotropin-
releasing hormone binding to rat anterior pituitary membrane
homogenates. Comparison of antagonists and agonists using
radiolabeled antagonist and agonist. Mol Pharmacol. 1983, 23,
44-51.
(18) On each assay plate a standard antagonist of comparable affinity
to those being tested was included as a control for plate-to-plate
variability. Overall, Ki values were highly reproducible with an
average standard deviation of 45% for replicate Ki determina-
(19) Atropical diastereoisomers were observed by NMR and HPLC
for compounds with 2-fluoro or 2-fluoro-3-methoxy analogues.
(20) Two compounds (R-26d and S-26g) were selected and tested for
their binding activity against over 10 different G-protein-coupled
receptors in-house. Neither of them showed more than 30%
inhibition against any of these GPCR receptors at 10 µM
concentration.
J M030472Z