Tetrahydroisoquinoline 1-carboxamides as growth hormone secretagogues

Article abstract of DOI:10.1016/j.bmcl.2005.02.040

Several novel series of tetrahydroisoquinoline 1-carboxamides were prepared and shown to be potent growth hormone (GH) secretagogues. Among them, carbamate 12a-E2 displays excellent in vivo activity by increasing plasma GH 10-fold in an anesthetized IV rat model.

Full text of DOI:10.1016/j.bmcl.2005.02.040

Bioorganic & Medicinal Chemistry Letters 15 (2005) 1799–1802
Tetrahydroisoquinoline 1-carboxamides as growth
hormone secretagogues
James J. Li,a,* Haixia Wang,a Fucheng Qu,a Christa Musial,a Joseph A. Tino,a,*
Jeffrey A. Robl,a Dorothy Slusarchyk,b Rajasree Golla,b Ramakrishna Seethala,b
Kenneth Dickinson,b Leah Giupponi,b Gary Grover,b Paul Sleph,b Neil Flynn,b
Brian J. Murphy,b David Gordon,b Melissa Kungc and Robert Stoffelc
aDiscovery Chemistry, Bristol-Myers Squibb Pharmaceutical Research Institute, PO Box 5400, Princeton, NJ 08543, USA
bMetabolic Disease Research, Bristol-Myers Squibb Pharmaceutical Research Institute, PO Box 5400, Princeton, NJ 08543, USA
cLead Discovery, Bristol-Myers Squibb Pharmaceutical Research Institute, 5 Research Parkway, Wallingford, CT 06492, USA
Received 4 January 2005; revised 10 February 2005; accepted 14 February 2005
Abstract—Several novel series of tetrahydroisoquinoline 1-carboxamides were prepared and shown to be potent growth hormone
(GH) secretagogues. Among them, carbamate 12a–E2 displays excellent in vivo activity by increasing plasma GH 10-fold in an anes-
thetized IV rat model.
ꢀ 2005 Elsevier Ltd. All rights reserved.
The availability of recombinant human growth hormone
(GH) in mid 1980s has generated numerous potential
uses for GH replacement therapy such as the treatment
of aged related functional muscular decline in the el-
derly.1 However, significant drawbacks, including cost,
parental administration, and side effects, limit the clini-
cal use of GH.2 An alternative approach of promoting
endogenous GH release with growth hormone releasing
peptides (GHRPs) was later discovered by Bowers et al.3
GHRPs and peptidomimetic analogs, collectively called
GH secretagogues, can increase pulsatile GH release via
activation of the novel G-protein coupled receptor
GHS-R1a, mimicking physiological conditions in hu-
mans.4 The discovery of new GHSs has received consid-
erable attention since GHS therapy may offer potential
advantages over existing GH replacement therapy.5 Sev-
eral GH secretagogues have advanced to clinic studies,
including MK-677 (Merck),6 CP-424391 (Pfizer),7 and
LY-444711 (Lilly).8 One common structural feature
among these compounds is the presence of a dipeptide
scaffold containing an aminoisobutyric acid (Aib) and
a phenylpropylglycine or O-benzylserine residue (Fig.
1). In an attempt to identify novel GH secretagogues,
we initiated a high throughput screen of the BMS
H
H
N
N
O
NH2
O
NH2
O
O
O
O
N
N
O
N
N
N
Me
SO2Me
MK-677
CP-424391
H
N
NH2
O
O
N
BnO
HN
O
N
N(iPr)2
N
H
N
Me
N
Boc
O
1 (HTS Lead)
MeO
Keyword: Growth hormone secretagogues.
*
Corresponding authors. Tel.: +1 609 818 7124; fax: +1 609 818 6810
(J.L.); tel.: +1 609 818 5049 (J.T.); e-mail addresses: james.li@bms.
com; joseph.tino@bms.com
LY-444711
Figure 1. Representative GH secretagogues and HTS lead 1.
0960-894X/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.02.040

1800
HO
J. J. Li et al. / Bioorg. Med. Chem. Lett. 15 (2005) 1799–1802
HO
The general synthetic sequence for analogs related to 1 is
O
O
c
a, b
outlined in Scheme 1. The key step for tetrahydroiso-
quinoline formation was accomplished by condensation
of glyoxylic acid with 3-hydroxy-phenethylamine (2) in
the presence of dilute hydrochloric acid. The methyl es-
ter 3 was generated from the intermediate acid to aid in
isolation and purification. From methyl ester 3, several
standard synthetic manipulations provided the late-
stage amine intermediate 6. With 6 in hand, an auto-
mated parallel solution phase synthesis was used for
the rapid preparation and isolation of analogs 7–10.14
Alternately, conventional amide coupling of acid 5 with
various amines gave analogs 11, allowing for the evalu-
ation of the basic amino side chain. A copper promoted
diaryl ether synthesis was adopted to provide analogs 12
in good yield.15 During the initial round of optimiza-
tion, all analogs were screened in vitro as racemic com-
pounds.16 Potent analogs were then evaluated in an
anesthetized IV rat model to determine their effect on
plasma GH levels.17,18
O
+
H
OMe
OH
NH
NH2
2
3
HO
BnO
O
O
O
d, e
OMe
O
OH
O
N
N
O
4
5
i
BnO
O
NHR
e, f, j
f, g
N
O
O
11
ArO
BnO
O
O
We hypothesized that there were three major pharmaco-
phore components enabling lead 1 to function as a GHS
agonist: a hydrophobic region, a basic amine, and a car-
bonyl hydrogen bond acceptor (Fig. 2).
N(iPr)2
N(iPr)2
NH
NH
h
N
O
NH
6
O
12
Structure–activity relationships were first directed to-
ward exploration of the Boc group in 1. Several series
of amides 7, ureas 8, sulfonamides 9, and carbamates
10 were prepared and their in vitro data are summarized
in Table 1. No clear SAR trend could be seen from
amide analogs 7, with most resulting in a decrease in
potency except for compounds 7j,p, and q. Several ureas
8a–c and sulfonamides 9a–e were also tested, but found
to be less active in the functional activity. Carbamate
analogs 10e–g in which a methyl group of the Boc moi-
ety is replaced with a methyl ester, methyl ketone or
dimethylamide respectively, showed a 5- to 10-fold
improvement of in vitro potency relative to 1. As com-
pared with the endogenous ligand Ghrelin, several ana-
logs never reach maximum response even at 10 lM, as
noted in Table 1.
BnO
O
N(iPr)2
NH
R
N
O
7
8
9
(amides)
(ureas)
(sulfonamides)
10 (carbamates)
Scheme 1. Reagents: (a) 5% HCl; (b) MeOH, TMSiCl; (c) Boc2–O,
Et3N; (d) NaH, BnBr; (e) NaOH; (f) 2-diisopropylaminoethylamine,
EDAC, HOAT; (g) 4 N HCl in dioxane, MeOH; (h) see Ref. 14; (i)
˚
RNH2, EDAC, HOAT; (j) ArB(OH)2, Cu(OAc)2, pyridine, Et3N, 4 A
molecular sieves.
compound collection. Through this effort, tetrahydro-
isoquinoline 1 was identified as a full GHS agonist with
an EC50 of 11.7 nM. Herein we describe the SAR
surrounding this novel class of GH secretagogues.
A set of primary, secondary, and tertiary amines
through analogs 11 (Table 2) were prepared to assess
the importance of terminal diisopropylamino group in
1. We also included compound 11e which contained a
terminal a,a-dimethyl amine in an attempt to model
the aminoisobutyrate subunit found in the literature ser-
ies (Fig. 2). In addition, different chain lengths between
the amino group and tetrahydroisoquinoline core were
One commonly used method for in vitro evaluation of
GHS activity has been the measurement of GH release
from isolated rat pituitary cells.9 However, the results
from this assay tend to be highly variable and the proce-
dure is not amendable for high throughput screen.10
Since the GHS receptor has been shown to signal
through a calcium phospholipase C mediated pathway,
it is expected that agonists can be identified by their abil-
ity to stimulate intracellular calcium mobilization using
a calcium sensitive fluorescent probe in an automated
instrument such as the fluorescence imaging plate reader
(FLIPR).11 A cell based functional FLIPR assay was
subsequently developed from H4 glioma cells in which
expression of the endogenous human GHS receptor
was enhanced by RAGE-activation.12,13
basic amine
O
O
N(iPr)2
NH
N
O
hydrophobic region
O
hydrogen bond acceptor
Figure 2. Proposed pharmacophore model for HTS lead 1.

J. J. Li et al. / Bioorg. Med. Chem. Lett. 15 (2005) 1799–1802
Table 1. In vitro potency of Boc replacement analogs 7–10
1801
of 0.49 nM vs 11.7 nM) improvement in functional
potency (Table 3). Further modifications with substi-
tuted phenyl ethers led to the identification of several
subnanomolar GHS agonists such as 12f,h. The two
enantiomers of 12a were separated to probe the
stereochemical effect of tetrahydroisoquinoline 1-carb-
oxamide side chain. The second eluting enantiomer
(12a–E2) was 15-fold more potent than the first one
(12a–E1) and both are full agonists.19
Compda
R
EC50 (nM)b
1
7a
t-BuO
t-BuCH2
t-Buc
Me
11.7
20.9
7b
7c
50% @ 10 lM
257
7d
7e
Ph
2-MeCOPh
2-FPh
7.6
7.3
7f
7g
17.8
30% @ 10 lM
9.0
2-EtOPhc
PhCH2
7h
7i
To assess the in vivo efficacy of this novel series of GH
secretagogues, select analogs were evaluated in an anes-
thetized IV rat model (Table 4).17,18 Two carbamates
10e and f (EC50 of 1.5–2.0 nM) were found to be effica-
cious; however, a slightly more potent carbamate 12h
(EC50 = 0.49 nM) only showed a marginal effect in this
model. Treatment of rats with the single enantiomer
12a–E2 (EC50 = 0.46 nM) generated over a 10-fold in-
crease in plasma GH, resulting in the most potent ana-
log identified in the IV rat model for the current series.
2-FPhCH2
2-MeOPhCH2
3-MeOPhCH2
4-MeOPhCH2
PhCH2CH2
PhCH2CH2CH2
PhCH(OH)
PhCH(CH2OH)
PhCH(Me)
PhCH(COMe)
PhCH(NHCOMe)
t-BuNH
43
2.1
7j
7k
7l
6.4
45
7m
7n
26.7
13.1
7o
7p
39
5.6
7q
7r
3.4
9.2
7s
8a
19.7
14.4
In summary, we have identified a novel series of tetrahy-
droisoquinolines as potent GH secretagogues. System-
atic SAR studies have resulted in compounds 12a,g,
and h with excellent in vitro potency (EC50 ꢀ 0.5 nM)
in the GHS functional assay. Carbamate 12a–E2
showed an 10-fold increase in plasma GH levels in an
8b
8c
PhNHc
43% @ 10 lM
190
Me2N
MeSO2
c
9a
9b
47% @ 10 lM
50% @ 10 lM
21
c
BuSO2
PhSO2
BnSO2
9c
9d
7.3
21
9e
Me2NSO2
n-PrO
c-HexO
10a
10b
10c
10d
10e
10f
10g
10h
Ghrelin
3.4
4.9
Table 3. In vitro potency of phenyl analogs 12
PhCH2O
PhO
27
<5% @ 10 lM
Compda
Ar
EC50 (nM)b
MeO2CCMe2O
MeOCCMe2O
Me2NCOCMe2O
NCCMe2O
1.5
2.0
0.85
9.8
1.4
1
PhCH2
Ph
Ph
11.7
0.49
7.6
12a
12a–E1
12a–E2
12b
12c
Ph
3-FPh
0.46
4.3
a Compounds 7o–s are diastereoisomers, all others are racemates.
b See Ref. 12 for detailed description.
c These compounds have a maximum intrinsic activity less than 50% at
10 lM as compared to endogenous ligand Ghrelin.
4-ClPh
3-CF3Ph
4-CF3Ph
3-NO2Ph
3-MeOPh
4-MeOPh
4-MeSPh
4-CHOPh
3-AcNHPh
0.73
1.5
12d
12e
2.3
12f
0.66
0.46
0.49
2.4
12g
12h
12i
Table 2. In vitro potency of analogs 11 with modification of 2-
diisopropylaminoethyl amide side chain
12j
12k
1.4
12
Compd
R
EC50 (nM)a
a Compound 12a–E1 & 12a–E2 are single enantiomers separated from
12a; all others are racemates.
1
CH2CH2N(i-Pr)2
CH2CH2NH(i-Pr)
CH2CH2NMe(i-Pr)
CH2CH2NEt2
11.7
219
b See Ref. 12 for detailed description.
11a
11b
11c
11d
11e
11f
11g
41
19.6
CH2CH2NMe2
CH2CMe2NH2
46% @ 10 lM
47% @ 10 lM
<5% @ 10 lM
257
Table 4. In vivo activity in the IV rat model
CH2CH2CH2NH(i-Pr)
CH2CMe2CH2NH2
Compd
EC50 (nM)
GH SEM(%)a
Responderc
a See Ref. 12 for detailed description.
10e
10f
1.5
2.0
746 227
666 326
172 49
5
5
4
5
5
12h
12a–E2
0.49
0.46
probed. As can be seen in Table 2, the diisopropylamino
moiety in 1 appears to be optimal for maintaining po-
tent GHS functional activity since all modifications re-
sulted in a substantial loss in potency for the series.
1283 367
367 154b
a Percent (%) of plasma GH increase at an IV dose of 5.23 lmol/kg.
b Percent (%) of plasma GH increase at an IV dose of 1.74 lmol/kg.
c Number of rats with greater than 100% GH increase over the control
group, there are five rats in both control and compound treated
groups.
Simple replacement of the benzyl moiety in 1 with a phe-
nyl group provided compound 12a with a 20-fold (EC50

1802
J. J. Li et al. / Bioorg. Med. Chem. Lett. 15 (2005) 1799–1802
10. Pong, S. S.; Chuang, L.-Y. P.; Dean, D. C.; Nargund, R.
P.; Patchett, A. A.; Smith, R. G. Mol. Endocrinol. 1996,
10, 57.
anesthetized IV rat model. Selected analogs are being
further evaluated.
11. (a) Sullivan, E.; Tucker, E. M.; Dale, I. L. Methods Mol.
Biol. 1999, 114, 125–133; (b) Schroeder, K. S.; Neagle, B.
D. J. Biomol. Screen. 1996, 1, 75.
Acknowledgments
The authors would like to thank Drs. William R. Ewing
and Robert Zahler for the helpful discussion during the
manuscript preparation.
12. The EC50 was measured by determining intracellular
calcium concentration in a FLIPR assay with Ghrelin as
having 100% intrinsic functional activity. All EC50 values
are the averages of multiple measurements.
13. For details about random activation of gene expression
(RAGE)TM technology (Athersys, Inc.), see: Harrington,
J. J.; Sherf, B.; Rundlett, S.; Jackson, P. D.; Perry, R.;
Cain, S.; Leventhal, C.; Thornton, M.; Ramachandran,
R.; Whittington, J.; Lerner, L.; Costanzo, D.; McElligott,
K.; Boozer, S.; Mays, R.; Smith, E.; Veloso, N.; Klika, A.;
Hess, J.; Cothren, K.; Lo, K.; Offenbacher, J.; Danzig, J.;
Ducar, M. Nat. Biotechnol. 2001, 19, 440.
14. For details, see (a) Li, J. J.; Tino, J. A. US 6,469,024, 2002;
(b) Lawrence, R. M.; Biller, S. A.; Fryszman, O. M.; Poss,
M. A. Synthesis 1997, 553.
15. Evans, D. A.; Katz, J. L.; West, T. R. Tetrahedron Lett.
1998, 39, 2937.
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16. The enantiomers were later separated for several ana-
logs (unpublished results). It was typically observed
that one enantiomer is about 5- to 15-fold more potent
than the other, and both enantiomers are full GHS
agonists.
17. Fasted male Wistar rats (200–250 g) were anesthetized via
intraperitoneal injection with ketamine (30 mg) and xyl-
azine (10 mg) per kilogram body weight. This treatment not
only serves to anesthetize the animals, but also stabilizes
the baseline plasma GH concentrations at approximately
60 ng/mL.18 The drug or vehicle (10% ethanol, 0.09%
saline V/V) was administered intravenously at a volume of
10 mL/kg to a group of five rats respectively. After 15 min,
1.5 mL of blood sample was drawn from the abdominal
aorta. Plasma samples were then assayed for rat growth
hormone by radioimmunoassay using a modification of
the kit supplied by the National Pituitary Hormone Center
(Dr. A. Parlow, Harbor-UCLA Medical Center, Los
Angeles, CA).
18. Chihara, K.; Arimura, A.; Schally, A. V. Endocrinology
1979, 104, 1434.
19. Chiral separations were performed on a preparative
ChiralPakAD Column (5 · 50 cm) with 20% isopropa-
nol/hexanes. The first eluting isomer is assigned as 12a–E1,
and second one is assigned as 12a–E2, but absolute
configurations have not been assigned.
9. Howard, A. D.; Feighner, S. D., et al. Science 1996, 273,
974.