1064
M. Sawa et al. / Bioorg. Med. Chem. Lett. 15(2005) 1061–1064
Table 3. Binding affinity and cell permeability of tryptamine-based
b3-agonists
monolayer permeability. The cinnamylamine analog
16i exhibited good permeability and showed improved
oral bioavailability in rats. Further studies are being
conducted to optimize the in vitro profile of this series
and will be reported in due course.
Compd
Binding Ki, nMa
Permeabilityb
b3
3.3
b1
b2
Papp · 10À6 cm/s
3a
44
150
48
23
18
25
nd (low)
7.3 (high)
2.1 (high)
16b
16i
33
15
Acknowledgements
a Binding potency is reported as Ki, the binding inhibition constant,
determined by inhibition of 125I-iodocyanopindolol binding.
b See footnote c in Table 1.
net) for their assistance in editing this manuscript.
b3-agonistic activity (16d and 16e). Interestingly, 16e
showed strong activity against b2-AR (EC50 = 1.5 nM),
while the potency against b1-AR was extremely weak.
Acetylene substituents tended to exert little influence
on the potency for b3-AR (16f and 16g vs 3g).
References and notes
1. Harada, H.; Hirokawa, Y.; Suzuki, K.; Hiyama, Y.; Oue,
M.; Kawashima, H.; Yoshida, N.; Furutani, Y.; Kato, S.
Bioorg. Med. Chem. Lett. 2003, 13, 1301–1305.
2. Mizuno, K.; Sawa, M.; Tateishi, H.; Harada, H.; Oue, M.;
Tsujiuchi, H.; Furutani, Y.; Kato, S. Bioorg. Med. Chem.
Lett. 2004, 14, 5959–5962.
3. Sawa, M.; Tateishi, H.; Mizuno, K.; Harada, H.; Oue, M.;
Tsujiuchi, H.; Furutani, Y.; Kato, S. Bioorg. Med. Chem.
Lett. 2004, 14, 5963–5966.
4. de Souza, C. J.; Burkey, B. F. Curr. Pharm. Des. 2001, 7,
1433–1449.
5. Yamaguchi, O. Urology 2002, 59(Suppl 5A), 25–29.
6. Ghera, E.; Plemenitas, A.; Ben-David, Y. Synthesis 1984,
504–506.
A very interesting observation was made upon the
expansion of the benzene ring to the naphthalene ring
(Table 2). The b3-agonistic potency of 16h increased
4-fold compared to that of 3g. As with the olefin com-
pounds, geminal substitution of the double bond was ex-
pected to decrease the potency for b3-AR, we therefore,
designed cinnamylamine analog 16i. As expected, cinn-
amylamine analog 16i showed strong activity for b3-
AR (EC50 = 1.0 nM, IA = 97%). This compound also
showed excellent selectivity against b1-AR and modest
activity against b2-AR.
7. Vallin, K. S. A.; Larhed, M.; Hallberg, A. J. Org. Chem.
2001, 66, 4340–4343.
8. In a previous report,1 we used CHO cells expressing a high
level of b3-AR to measure compound activity, that is the
receptor densities were 150,000 receptors/cell (b3-AR),
12,000 receptors/cell (b1-AR), and 30,000 receptors/cell
(b2-AR). To better evaluate the subtype selectivity, we
used CHO cells expressing low densities of b3-AR (13,000
receptors/cell), and high densities of b1- and b2-ARs in this
report (320,000 and 600,000 receptors/cell, respectively).
The CHO cells expressing either human b1-, b2-, or b3-AR
were prepared as described in Kato, S.; Harada, H.;
Taoka, I.; Kawashima, H. PCT Patent Application, WO
2000044721, 2000; Kato, S.; Harada, H.; Hirokawa, Y.;
Yoshida, N.; Kawashima, H. PCT Patent Application,
WO 9616938, 1996.
9. The apparent permeability coefficients (Papp) were deter-
mined in the apical-to-basolateral (A-to-B) direction
across the cellÕs monolayers cultured on polycarbonate
membrane filters.
10. Compound 16i was dosed orally at 5 mg/kg in 1%
hydroxypropyl cellulose aqueous solution and intrave-
nously at 0.5 mg/kg in a vehicle consisting of 10% DMSO,
10% EtOH, 20% polyethylene glycol and saline. After
dosing, blood samples were collected at 5 and 15 min (iv
only), and 0.5, 1, 2, 4, 6, 8, 24 h (ivand po) to determine
plasma drug concentrations by LC/MS/MS.
To evaluate the possibility of this series for drug candi-
dates, the selected compounds were subjected to recep-
tor binding assays. As shown in Table 3, the binding
potency of crotyl analog 16b decreased by one order
of magnitude compared to that of 3a (Ki = 33 and
3.3 nM, respectively). Furthermore, 16b exhibited rever-
sal selectivity in binding regarding b3 versus b2-AR
compared with the agonistic activity. Cinnamylamine
analog 16i also showed a decreased binding constant
(Ki = 15 nM), but possessed moderate selectivity against
b1 and 2-AR. Since the cinnamylamine analog 16i
showed high Caco-2 cellular permeability (Papp = 2.1
Papp · 10À6 cm/s), the pharmacokinetic property was
determined in rats (5 mg/kg po, 0.5 mg/kg iv).10 As ex-
pected from the Caco-2 experiments, a large improve-
ment was observed with the cinnamylamine analog
16i; its oral bioavailability in the rat was 21%. In addi-
tion, the half-life of 16i was 4.2 h. These data indicate
that the sulfonamide moiety is detrimental to oral
absorption.
In conclusion, we have discovered aniline analogs as po-
tent and selective b3-agonists with excellent Caco-2 cell