5892 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 24
Letters
In summary, we report a potent, highly water soluble (>100
mg/mL at pH 4), orally active GH secretagogue containing a
novel tetrazole linker. The safety profile and acceptable phar-
macokinetic properties of 2 were suitable for long-term efficacy
evaluation studies for the prevention of frailty and treatment
for cancer cachexia as well as wasting syndrome, and as a result,
the compound was advanced for clinical evaluation.
Figure 2. Pharmacodynamic response to a single ascending oral dose
of 2 in conscious beagle dogs.
Acknowledgment. We thank Dr. Robert Zahler for his
advice and suggestions for the work and the manuscript.
suggested that a secondary or cyclic amine might be suitable
as a replacement for the primary amine. Indeed, the pyrrolidine-
based secondary alcohol 12 (EC50 ) 1.0 nM) was identified as
one of the most potent compounds in this series. However, the
pharmacokinetic profile of compound 12 was inferior to that
of compound 2. Efforts to address the potential liability of the
secondary alcohol in 12 by incorporation of the tertiary alcohol
13 (EC50)10.1 nM) resulted in a 10-fold reduction in potency.
In contrast to that of the acyclic acetamide 14, the pyrrolidine
acetamide 15 exhibited good in vitro functional potency (EC50
) 3.3 nM) as a mixture of diastereomers. Unfortunately, the
PK profile of 15 was unacceptable due to its low bioavailability
in a rat model (F < 5%). The carbamate moiety was found to
be a metabolically stable linker for the chemotype; the overall
hydroxybutyl carbamic acid gave excellent potency and PK
properties. Other attempts to replace the carbamate in 2 with
an amide, urea, or reversed carbamate resulted in similar or less
potent analogues; the results are to be reported elsewhere.
The in vivo efficacy of compounds was evaluated in an acute
anesthetized intravenously dosed rat model, monitoring plasma
GH increases.16 In this model, 2 showed excellent activity,
producing a rapid and robust GH plasma elevation when
administered iv to rats (ED50 ) 0.08 µmol/kg). Figure 2 shows
the growth hormone pharmacodynamic response following
single oral doses of 2 in conscious beagle dogs. The vehicle
(water) treated control animals showed no GH increase over
the 4 h time course of the experiment (data not shown). At a
dose of 0.52 µmol/kg, only a minimal increase in plasma GH
was observed. Significant increases in plasma GH were achieved
at the doses of 1.74 and 5.23 µmol/kg, and a further increase in
plasma GH was observed at the dose of 17.4 µmol/kg. The
response plateaued at 52.3 µmol/kg. The maximum GH
concentration occurred between 15 and 60 min after dosing and
remained elevated for up to 120 min.
The pharmacokinetic characteristics of compound 2 were
evaluated in fasted rats, cynomolgus monkeys, and beagle dogs
at single oral doses of 15, 10, and 10 µmol/kg, respectively.
The iv dose was 5 µmol/kg for all species. The oral bioavail-
ability of 2 in rats was 9%, with a half-life of 0.7 h. In monkeys,
the oral bioavailability was 12%, with a half-life of 1.1 h.
Compound 2 demonstrated superior exposure in dogs relative
to the other species, with an oral bioavailability of 40% and a
2.3 h half-life. The low oral bioavailability of 2 in rats and
monkeys was probably attributable to its high plasma
clearance (213 and 37 mL/min/kg in rats and monkeys,
respectively). For comparison, the clearance rate of 2 in dogs
was 21 mL/min/kg.
Compound 2 was found to have no significant activity (>50%
inhibition at 10 µM) in the PanLabs SpectrumScreen in vitro
assay panel17 and was not hepatotoxic up to 250 µg/mL in a
human hepatocyte assay. The IC50 values for compound 2 were
all greater than 100 µM against human CYP enzymes 3A4, 2C9,
2C19, 2D6, and 1A2. Ames testing also showed that compound
2 was not mutagenic to the tester strains TA 98 and TA 100.
Supporting Information Available: Experimental procedure
for the preparation of 2 and 1H NMR, HPLC, and HRMS data for
compounds 2-18. This material is available free of charge via the
References
(1) (a) Strobl, J. S.; Thomas, M. J. Human growth hormone. Pharmacol.
ReV. 1993, 46, 1-34. (b) Rosen, T.; Johannsson, G.; Johansson, J.-
O.; Bengtsson, B.-A. Consequences of growth hormone deficiency
in adults and the benefits and risks of recombinant human growth
hormone treatment. Horm. Res. 1995, 43, 93-99.
(2) Momany, F. A.; Bowers, C. Y.; Reynolds, G. A.; Chang, D.; Hong,
A.; Newlander, K. Design, synthesis, and biological activity of
peptides which release growth hormone in vitro. Endocrinology 1981,
108, 31-39.
(3) (a) Sartor, O.; Bowers, C. Y.; Chang, D. Parallel studies of His-D-
Trp-Ala-Trp-D-Phe-Lys-NH2 and human pancreatic growth
hormone-releasing factor-44-NH2 in rat primary pituitary cell mono-
layer culture. Endocrinology 1984, 116, 952-957. (b) Bowers, C.
Y.; Momany, F. A.; Reynolds, G. A.; Hong, A.; Newlander, K. On
the in vitro and in vivo activity of a new synthetic hexapeptide that
acts on the pituitary to specifically release growth hormone.
Endocrinology 1984, 114, 1537-1545.
(4) (a) Akman, A. S.; Girard, M.; O’Brien, L. F.; Ho, A. K.; Chik, C. L.
Mechanisms of actions of a 2nd generation growth hormone-releasing
peptide (Ala-His-D-beta-Nal-Ala-Trp-D-Phe-Lys-NH2) in rat
anterior-pituitary cells. Endocrinology 1993, 132, 1286-1291. (b)
For a review on the clinical actions of GHRPs, see: Ghigo, E.; Arvat,
E.; Muccioli, G.; Camanni, F. Growth hormone releasing peptides.
Eur. J. Endocrinol. 1997, 136, 445-460, and references therein.
(5) (a) Smith, R. G.; Cheng, K.; Pong, S. S.; Hickey, H.; Jacks, T.; Butler,
B.; Chan, W.-S.; Chaung, L. Y. P.; Judith, F.; Taylor, J. A.; Wyvratt,
M. J.; Fisher, M. H. A novel non-peptidyl growth hormone
secretagogue. Science 1993, 260, 1640-1643. (b) Pong, S.-S.;
Chaung, L.-Y. P.; Dean, D. C.; Nargund, R. P.; Patchett, A. A.; Smith,
R. G. Identification of a new G-protein-linked receptor for growth
hormone secretagogues. Mol. Endocrinol. 1996, 10, 57-61.
(6) (a) Schoen, W. R.; Pisano, J. M.; Prendergast, K.; Wyvratt, M. J.,
Jr.; Fisher, M. H.; Cheng, K.; Chan, W. W. S.; Butler, B.; Smith, R.
G.; Ball, R. G. A novel 3-substituted benzazepinone growth hormone
secretagogue (L-692,429). J. Med. Chem. 1994, 37, 897-906. (b)
Patchett, A. A.; Nargund, R. P.; Tata, J. R.; Chen, M.-H.; Barkat, K.
J.; Johnston, D. B. R.; Cheng, K.; Chan, W. W.-S.; Butler, B.; Hickey,
G.; Jacks, T.; Schleim, K.; Pong, S.-S.; Chaung, L.-Y. P.; Chen, H.
Y.; Frazier, E.; Leung, K. H.; Chiu, S.-H. L.; Smith, R. G. Design
and biological activities of L-163,191 (MK-0677): A potent orally
active growth hormone secretagogue. Proc. Natl. Acad. Sci. U.S.A.
1995, 92, 7001-7005.
(7) (a) Yang, L.; Morriello, G.; Patchett, A. A.; Leung, K.; Jacks, T.;
Cheng, K.; Schleim, K. D. Feeney, W.; Chan, W. W.-S.; Chiu, S.
L.; and Smith, R. G. 1-[2(R)-(2-Amino-2-methylpropionylamino)-
3-(1H-indol-3-yl)propionyl]-3-benzylpiperidine-3(S)-carboxylic Acid
Ethyl Ester (L-163,540): A potent, orally bioavailable, and short-
duration growth hormone secretagogue. J. Med. Chem. 1998, 41, 2439
-2441. (b) Hansen, T. K.; Ankersen, M.; Hansen, B. S.; Raun, K.;
Nielsen, K. K.; Lau, J.; Peschke, B.; Lundt, B. F.; Thgersen, H.;
Johansen, N. L.; Madsen, K.; Andersen, P. H. Novel orally active
growth hormone secretagogues. J. Med. Chem. 1998, 41, 3705-3714.
(c) Carpino, P. A.; Lefker, B. A.; Toler, S. M.; Pan, L. C.; Hadcock,
J. R.; Murray, M. C.; Cook, E. R.; DiBrino, J. N.; DeNinno, S. L;.
Chidsey-Frink, K. L;. Hada, W. A.; Inthavongsay, J.; Lewis, S. K.;
Mangano, F. M.; Mullins, M. A.; Nickerson, D. F., Ng, O.; Pirie, C.
M.; Ragan, J. A.; Rose, C. R.; Tess, D. A.; Wright, A. S.; Yu, L.;
Zawistoski, M. P.; Pettersen, J. C.; DaSilva-Jardine, P. A.; Wilson,
T. C.; Thompson, D. D. Discovery and biological characterization
of capromorelin analogues with extended half-lives. Bioorg. Med.