Potent nonpeptide GnRH receptor antagonists derived from substituted indole-5-carboxamides and -acetamides bearing a pyridine side-chain terminus

Article abstract of DOI:10.1016/S0960-894X(01)00275-X

A pyridine side-chain terminus has been incorporated into the indole-5-carboxamide and indole-5-acetamide series of GnRH antagonists. Potent activity was observed in binding and functional assays. Certain branched or cyclic tertiary amides were identified

Full text of DOI:10.1016/S0960-894X(01)00275-X

Bioorganic & Medicinal Chemistry Letters 11 (2001) 1727–1731
Potent Nonpeptide GnRH Receptor Antagonists Derived from
Substituted Indole-5-carboxamides and -acetamides Bearing a
Pyridine Side-Chain Terminus
Wallace T. Ashton,a,* Rosemary M. Sisco,a Yi Tien Yang,b Jane-Ling Lo,b
Joel B. Yudkovitz,b Patrice H. Gibbons,b George R. Mount,b Rena Ning Ren,b
a
Bridget S. Butler,b Kang Chengb and MarkT. Goulet
aDepartment of Medicinal Chemistry, Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065-0900, USA
bDepartment of Biochemistry and Physiology, Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065-0900, USA
Received 16 March 2001; accepted 19 April 2001
Abstract—A pyridine side-chain terminus has been incorporated into the indole-5-carboxamide and indole-5-acetamide series of
GnRH antagonists. Potent activity was observed in binding and functional assays. Certain branched or cyclic tertiary amides were
identified as preferred in each series. Alkylation of the side-chain secondary amine had generally unfavorable effects. Variations of
the gem-dialkyl substituents in the indole-5-acetamide series were also investigated. # 2001 Elsevier Science Ltd. All rights
reserved.
As described in our previous paper,1 we have sought
orally active, nonpeptide receptor antagonists of gona-
dotropin-releasing hormone (GnRH; also known as
luteinizing hormone-releasing hormone, or LHRH).
Such compounds could be desirable drugs for ther-
apeutic applications requiring suppression of sex hor-
mone levels.2 Potential uses include treatment of certain
hormone-dependent cancers, uterine fibroids, and
endometriosis, as well as assisted reproduction proto-
cols.2,3 Following previous workfrom these labora-
tories,4 we reported potent GnRH antagonists derived
from indole-5-carboxamides and -acetamides, exempli-
fied by structures 1 and 2.1 Our original compounds
contained phenol or aryl methanesulfonamide groups
terminating the side chain at the 3-position of the
indole. Because of unfavorable pharmacokinetics, alter-
native end groups had been investigated. It was repor-
ted5 that 3-pyridyl and 4-pyridyl termini, as in 3 and 4,
were compatible with good potency at the GnRH
receptor. We now describe the incorporation of the
pyridyl terminus into the indole-5-carboxamide and
-acetamide series of GnRH antagonists. A series of
indole-5-carboxamides incorporating pyridyl end
groups was prepared according to Scheme 1. The 3- and
4-pyridylbutanols 56 were oxidized under Swern condi-
tions5 to the aldehydes 6.7 Reductive amination with the
*Corresponding author. Fax: +1-732-594-5350; e-mail: wally_ashton@merck.com
0960-894X/01/$ - see front matter # 2001 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(01)00275-X

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W. T. Ashton et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1727–1731
tryptamine derivative 71 furnished 8. After saponification
of the ester, the secondary amine was protected to give Cbz
derivative 9. Next, carboxylic acid 9 was coupled with
amines in the presence of PyBOP reagent. Finally, depro-
tection by catalytic hydrogenolysis afforded the amide
products 10–13 and 15–35. The N-methylated analogue 14
was obtained from 13 by reductive amination using
paraformaldehyde and sodium cyanoborohydride.
further characterize many of the compounds: inhibition
of GnRH-stimulated LH release from rat primary
pituitary cells9 and inhibition of GnRH-stimulated
phosphatidylinositol (PI) hydrolysis in cloned Chinese
hamster ovary (CHO) cells stably expressing the human
GnRH receptor.9
It is apparent from the data in Table 1 that addition of a
carboxamide to the 5-position of the indole dramati-
cally increases binding affinity to the rat GnRH receptor
compared to the parent analogues 3 and 4. Although
potency at the rat receptor did not vary greatly among
the amides, all of which had subnanomolar IC50 values,
greater differences were seen for the human receptor.
Several bulky, hydrophobic, branched or cyclic tertiary
amides (24–28 and 31) were most favorable for binding
to the human GnRH receptor. Some of these same
compounds, notably 26–28, were also among the most
effective in the LH release and PI turnover assays.
Nevertheless, the results from the functional assays did
not always trackwith those of the binding studies.
Thus, the diethyl amide 13 proved to be considerably
more active (IC50=12 nM) in the PI inhibition experi-
ment than would have been predicted from its perfor-
mance in the other tests. In all of the assays, the
attachment point of the terminal heterocycle (i.e.,
3-pyridyl or 4-pyridyl) had little effect.
A series of analogues containing gem-dialkylacetamide
substituents, as in 2, was prepared as shown in Scheme
2. By known methods,8 ethyl (4-nitrophenyl)acetate (36)
was dialkylated and then hydrogenated to give 37.
These intermediates were converted to the tryptamines
38 by our previously described procedures.1 Further
elaboration to target compounds 39–61 was carried out
as described for Scheme 1.
In the binding assay, compounds were initially tested
for the ability to compete with the GnRH receptor
agonist [125I]-buserelin for binding to the rat GnRH
receptor9 in the presence of 0.1% BSA. During the
course of this study, the rat receptor binding assay was
replaced by a similar human GnRH receptor binding
assay,9 which was more sensitive to structure–activity
relationships and was considered more relevant to the
drug design process. Two functional assays were used to
Scheme 1. Conditions: (i) (a) oxalyl chloride, DMSO, CH2Cl2, À60 ꢀC; (b) Et3N, À65 ꢀC; (ii) (a) MgSO4, CDCl3, À5 to À10 ꢀC; (b) NaBH4, MeOH,
À5 ꢀC; (iii) Cbz-Cl, i-Pr2NEt, CH2Cl2–THF, À78 ꢀC; (iv) (a) KOH, MeOH–H2O, Á; (b) H+; (v) R1R2NH, PyBOP, Et3N, CH2Cl2; (vi) H2,
Pd(OH)2/C, 2-methoxyethanol; (vii) paraformaldehyde, NaBH3CN (4 equiv), AcOH (10 equiv), powdered 4 A molecular sieves, MeOH–THF.
Scheme 2. Conditions: (i) (a) NaH, DMF; (b) R3I, R4I; (ii) H2, Pd/C, EtOH; (iii) procedures described in ref 1.

W. T. Ashton et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1727–1731
1729
Except for the primary amide 39, most of the indole-5-
acetamides (Table 2) were potent antagonists at the
human GnRH receptor, several having IC50 values of 1–
3 nM. Low nanomolar IC50 values at this receptor were
achieved with smaller amide N-substituents than in the
indole-5-carboxamide series. Again, 3- and 4-pyridyl
analogues were similar in potency. Among the most
effective compounds in the binding and functional
Table 1. Inhibition of GnRH receptor binding, LH release, and PI turnover by indole-5-carboxamides with pyridine end groups
5
Compd
X
Pyridine linkR
GnRH binding
IC50 (nM)a
LH release
IC50 (nM)b
PI turnover
IC50 (nM)c
Rat
Human
3d
4d
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
[H]e
[H]e
Me2N
Me2N
Et2N
Et2N
Et2N
3
4
3
4
3
4
4
3
4
3
4
3
3
4
3
4
3
4
3
4
4
H
H
H
H
H
H
Me
H
H
H
H
H
H
H
H
H
H
H
H
H
H
40
16
0.7
0.8
0.6
0.5
2600
1460
400
470
430
1200
1000
1300
3600
>1300
830
400
1000
240
150
380
23
18
36
190
12
58
HOCH2CH2N(Et)
HOCH2CH2N(Et)
(MeOCH2CH2)2N
(CF3CH2)2N
i-PrNH
0.6
0.6
36
45
36
55
11
260
610
91
1200
110
i-PrN(Et)
i-Pr2N
n-Bu2N
n-Bu2N
i-Bu2N
i-Bu2N
0.3
0.4
0.4
0.7
0.6
0.2
0.4
56
120
80
16
22
5.0
7.1
2.3
2.8
1.9
Cyclohexyl-N(Et)
Cyclohexyl-N(Et)
Cyclooctyl-N(Et)
140
130
81f
15
29
3
H
0.6
0.5
5100
30
4
H
>1300
31
3
H
3.6
360
62
32
33
34
35
4
3
4
4
H
H
H
H
0.3
0.4
0.3
340
1600
1400
32
260
aInhibition of binding of [125I]buserelin to rat or human pituitary GnRH receptor.
bInhibition of GnRH-stimulated LH release from rat pituitary cells.
cInhibition of GnRH-stimulated [3H]inositol phosphate hydrolysis.
dRef 5.
eSubstituent in place of XCO- at indole 5-position.
fModified assay conditions using 0.2 nM instead of 2 nM GnRH.

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W. T. Ashton et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1727–1731
assays were the 7-azanorbornane amides 53 and 54
despite the relatively poor performance of the corre-
sponding analogue 35 in the indole-5-carboxamide ser-
ies. Other particularly active analogues in the PI
turnover assay included the diisobutyl amide 45 and the
2,5-dimethylpyrrolidine amide 50. N-Methylation of the
secondary amine (55; cf. 54) had no significant effect on
potency in the receptor binding and LH release assays,
as was also the case for 14 in Table 1. However, a 6-fold
loss in activity was observed for 55 versus the parent 54
in the PI turnover assay. Increasing the size of the
N-alkyl group (56–58) tended to reduce activity in the
assays. In our earlier series,1 gem-dimethylation of the
acetamide side chain was found to enhance activity, and
this substitution pattern may also be favorable for
reducing metabolism. Here, replacing one or both of the
Table 2. Inhibition of GnRH receptor binding, LH release, and PI turnover by indole-5-acetamides with pyridine end groups
5
Compd
R3
R4
X
Pyridine linkR
hGnRH
IC50 (nM)a
LH release
IC50 (nM)b
PI turning
IC50 (nM)c
39
40
41
42
43
44
45
46
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
H2N
Me2N
Me2N
Et2N
Et2N
i-Bu2N
i-Bu2N
c-HxN(Et)
3
3
4
3
4
3
4
4
H
H
H
H
H
H
H
H
32
350
[0.5]d
[0.3]d
3.3
680
930
240
770
400
140
960
350e
190
94
58
22
8.3
54
3.2
1.7
2.8
2.3
47
48
49
50
51
52
53
54
55
56
57
58
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
3
3
4
4
4
4
3
4
4
4
4
4
H
1.0
5.0
4.8
2.8
1.8
11
62
H
290
340
160
1200
540
73e
61
H
H
15
38
H
H
59
H
1.4
1.4
1.5
2.2
3.1
8.2
14
H
71e
18
Me
Et
i-Pr
i-Bu
71e
110
90
730e
200
300
(continued on next page)

W. T. Ashton et al. / Bioorg. Med. Chem. Lett. 11 (2001) 1727–1731
1731
Table 2 (continued)
5
Compd
R3
R4
X
Pyridine linkR
hGnRH
IC50 (nM)a
LH release
IC50 (nM)b
PI turning
IC50 (nM)c
59
60
61
Me
Et
Et
Et
4
4
4
H
H
H
1.0
1.7
1.2
240e
260e
38
40
-(CH2)3—
170
aInhibition of binding of [125I]-buserelin to human pituitary GnRH receptor.
bInhibition of GnRH-stimulated LH release from rat pituitary cells.
cInhibition of GnRH-stimulated [3H]inositol phosphate hydrolysis.
dData for inhibition of rat GnRH receptor.
eModified assay conditions using 0.2 nM instead of 2 nM GnRH.
gem-dimethyl substituents by ethyl (59 and 60) or con-
necting them in a four-membered ring (61) was reasonably
well tolerated but offered no advantages.
Simeone, Mamta Parikh, and Dr. Peter Lin for pre-
paration of certain intermediates.
In summary, the combination of a 3- or 4-pyridyl ter-
minus on the indole-3-side chain and carboxamide or
acetamide substituents at the indole 5-position afforded
potent antagonists at rat and human GnRH receptors.
These compounds were also effective in blocking
GnRH-stimulated release of LH from rat pituitary cells
and GnRH-stimulated PI turnover in CHO cells
expressing cloned human GnRH receptors. Nonpolar
tertiary amines were most potent in each series. Among
the indole-5-carboxamides, a preferred analogue was
the bulky N-cyclooctyl-N-ethyl amide 28, with IC50
values of 1.9, 81, and 15 nM in the receptor binding, LH
release, and PI turnover assays, respectively. A some-
what different structure–activity relationship was
observed for amides in the indole-5-acetamide series.
Here the most favorable amides were those derived
from diisobutylamine (45), 2,5-dimethylpyrrolidine (50),
and 7-azanorbornane (53 and 54). For example, 53 had
IC50 values of 1.4, 73, and 14 nM in the binding, LH
release, and PI turnover assays, respectively. In both
series, other structural modifications, involving varia-
tion of the gem-dialkyl substituents or alkylation on the
secondary amine of the side chain, had neutral or nega-
tive effects. Further structural variations, leading to
GnRH antagonists with improved pharmacokinetics
and in vivo activity, will be described in a subsequent
report.
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Acknowledgements
The authors are grateful to Amy Bernickand Dr.
Lawrence Colwell for providing mass spectral data and
to MarkLevorse, Robert Frankshun, Glenn Reynolds,
Judith Pisano, Joseph Leone, Dr. Ranjit Desai, Joseph
9. Walsh, T. F.; Toupence, R. B.; Young, J. R.; Huang, S. X.;
Ujjainwalla, F.; DeVita, R. J.; Goulet, M. T.; Wyvratt, M. J., Jr.;
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