Design and synthesis of KNT-127, a δ-opioid receptor agonist effective by systemic administration

Article abstract of DOI:10.1016/j.bmcl.2010.08.083

We have reported previously the novel δ-opioid agonist, SN-28, which was more potent in in vitro assays than the prototype δ-agonists, TAN-67 and SNC-80. However, when administered by subcutaneous injection, this compound showed no analgesic effect at dosages greater than 30 mg/kg in the acetic acid writhing test. We speculated that SN-28 was not effective in the test because the presence of the charged ammonium groups prevented its penetration through the blood-brain barrier. On the basis of our proposal, we designed the novel δ-agonist, KNT-127, which was effective with systemic administration.

Full text of DOI:10.1016/j.bmcl.2010.08.083

Bioorganic & Medicinal Chemistry Letters 20 (2010) 6302–6305
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design and synthesis of KNT-127, a d-opioid receptor agonist effective
by systemic administration
Hiroshi Nagase a, , Toru Nemoto a, Ayaka Matsubara a, Manabu Saito a, Naoshi Yamamoto a, Yumiko Osa a,
⇑
Shigeto Hirayama a, Mayumi Nakajima b, Kaoru Nakao b, Hidenori Mochizuki b, Hideaki Fujii a
a School of Pharmacy, Kitasato University, 5-9-1, Shirokane, Minato-ku, Tokyo 108-8641, Japan
b Pharmaceutical Research Laboratories, Toray Industries, Inc., 6-10-1, Tebiro, Kamakura, Kanagawa 248-8555, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
We have reported previously the novel d-opioid agonist, SN-28, which was more potent in in vitro assays
than the prototype d-agonists, TAN-67 and SNC-80. However, when administered by subcutaneous injec-
tion, this compound showed no analgesic effect at dosages greater than 30 mg/kg in the acetic acid writh-
ing test. We speculated that SN-28 was not effective in the test because the presence of the charged
ammonium groups prevented its penetration through the blood–brain barrier. On the basis of our pro-
posal, we designed the novel d-agonist, KNT-127, which was effective with systemic administration.
Ó 2010 Elsevier Ltd. All rights reserved.
Received 3 August 2010
Revised 17 August 2010
Accepted 18 August 2010
Available online 21 August 2010
Keywords:
Opioid
Naltrexone
Analgesics
d-Receptor
O
Three types of opioid receptors (l, d, j) are now well estab-
O
Et2N
lished not only by pharmacological studies, but also through
Et2N
H
N
N
molecular biological studies.1 For the past three decades, consider-
H
N
N
OCH3
OH
able effort has been expended on obtaining an opioid
j-selective
CH3
CH3
agonist to eliminate undesirable morphine-like side effects like
addiction. In 1982, U-50,488H was discovered to be a highly
H3C
H3C
selective
j-agonist by researchers at Upjohn (now merged with
Pfizer).2–4 Since then, numerous research groups modified its
structure and succeeded in preparing more selective and potent
SNC80
BW373U86
j
-agonists. These compounds had a potent antinociceptive effect
12
5
OH
N
1
11 10
H
2
H3C
H3C
9
8
in animal models and also succeeded eliminating the morphine-
like side effects. However, all these compounds have structures
N
3
4
N
N
6
7
O
H
OH
OH
TAN-67
OH
Cl
Cl
O
OMI
O
N
N
N
Figure 2. The structures of BW373U86, SNC-80, and TAN-67.
CH3
O
O ·HCl
CH3
N
OH
TRK-820
U-50,488H
Upjohn
8
14
9
H
4'
5'
8'
H3C 17
7
6'
7'
N
13
5
16
10
15
6
N
Figure 1. The structures of U-50,488H and TRK-820.
12
4
11
1
3
OH
SN-28
2
Abbreviations: BBB, blood–brain barrier; s.c., subcutaneous; i.t., intrathecal;
CBM, cyclobutylmethyl.
⇑
Corresponding author. Tel.: +81 3 5791 6372; fax: +81 3 3442 5707.
E-mail address: nagaseh@pharm.kitasato-u.ac.jp (H. Nagase).
Figure 3. The structure of SN-28.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.08.083

H. Nagase et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6302–6305
6303
(Nalfurafine hydrochloride) which has a structure quite different
from U-50,488H. Unlike U-50488H, TRK-820 bears a tyrosine–gly-
cine dipeptide unit which is a structural motif commonly found in
endogenous opioid peptides (Fig. 1).5,6
hydrogen
bond
H
O
N
The new compound showed neither addiction nor aversion and
was launched in Japan as an antipruritic agent for kidney dialysis
patients last year. This is the first opioid drug that does not cause
O
O
OH
addiction. Now that we obtained a selective j-agonist without mor-
naltrexone
phine-like side effects and aversion, our next target was a highly
selective and potent d-agonist. Nonpetide d-opioid agonists
BW373U86,7 SNC-80,8,9 and OMI10,11 have been described and we
have also reported the highly selective d-agonist, TAN-67(Fig. 2).12,13
However, these prototypical compounds were not potent
enough to examine the intrinsic pharmacological effects of the
d-receptor and even now we do not know if d-receptor contributes
to addiction, as occurs with morphine.
Figure 4. The hydrogen bond formed the lone electron pair on the 17-nitrogen and
the 14-hydroxy group in naltrexone.
OH
OAc
Troc
N
N
O
a) b)
O
O
O
Quite recently, we have reported a novel d-agonist, SN-28,
which showed about 15 times and 334 times more potent agonist
activities than TAN-67 and SNC-80, respectively and almost equiv-
alent selectivity to TAN-67 in in vitro assays (Fig. 3).14 However, at
dosages over 30 mg/kg administered by subcutaneous (s.c.) injec-
tion, SN-28 showed no analgesic effect in the acetic acid writhing
test.6 To date, no d-agonist exists that is therapeutically effective
by systemic administration and this problem illustrates the diffi-
culties of developing peptidic compounds as drugs. In an effort to
understand why SN-28 (by the s.c. route) was not effective in the
test, we proposed that SN-28 does not penetrate through the
blood–brain barrier (BBB). To confirm this hypothesis, we exam-
ined the analgesic effect of SN-28, administered by intrathecal
injection (i.t. injection), in the acetic acid writhing test. As we
OCH3
OH
OCH3
1
2
R
N
OH
H3C
O
N
e)
c)
f)
O
O
OCH3
OCH3
R = H: 3
R = CH3: 4
d)
5
OH
H3C
N
N
had expected, SN-28 showed
a
strong analgesic effect
OR
(ED50 = 0.095 nM) in this test, which supported the idea that the
compound does not cross the BBB. On the basis of the above pos-
tulate, we designed a novel d-agonist, KNT-127, which is effective
in systemic administration. Here we report the design and synthe-
sis of KNT-127.
TAN-67 and SN-28 have two basic nitrogens: a quinoline moiety
and a 17-nitrogen. Both would contribute to high polarity of the
compounds. The quinoline group is considered to be an essential
part of the address for d-receptor agonist activity.12,13 Therefore,
when we designed and synthesized TAN-67 by use of the mes-
sage-address concept15 and the accessory site hypothesis,16 we
chose the quinoline group as the address moiety. On the other
R = CH3: 6
R = H: KNT-127
g)
Scheme 1. Reagents and conditions: (a) Ac2O, reflux; (b) TrocCl, K2CO3,
CHCl2CHCl2, reflux, 85% (two steps from 1); (c) 12 N KOH, DMSO, reflux; (d) HCHO,
CH3COONa, CH3COOH, H2, 10% Pd–C, rt, 72% (two steps from 2); (e) 1 N HCl, 80 °C,
90%; (f) 2-aminobenzaldehyde; CH3SO3H, EtOH, reflux, 75%; (g) BBr3, CH2Cl2,
0 °C ? rt, 85%.
similar to that of U-50,488H, with the [N–C–C–N(sp2)] pharmaco-
phore sequence (Fig. 1), and showed severe aversion like psychoto-
mimetic effect. We recently reported a novel
j-agonist, TRK-820
OH
R
OH
N
f)
RN
a) or b) or c, d)
or e, d)
O
3
O
O
OCH3
OCH3
R = benzyl: 7 (95%); a)
R = phenethyl: 8 (79%); b)
R = CBM: 9 (86%); c), d)
11
R = benzyl:
(92%)
12
R = phenethyl:
(92%)
13
R = CBM:
(85%)
R = c-Pen-methyl: 10 (55%); e), d)
R = c-Pen-methyl: 14 (95%)
OH
OH
RN
g)
RN
h)
N
N
OCH3
OH
R = benzyl: 15 (93%)
R = benzyl: SYK-3 (77%)
R = phenethyl: 16 (84%)
R = CBM: 17 (77%)
R = phenethyl: SYK-4 (93%)
R = CBM: SYK-14 (88%)
R = c-Pen-methyl: 18 (87%)
R = c-Pen-methyl: SYK-69 (70%)
Scheme 2. Reagents and conditions: (a) BnBr, K2CO3, DMF, rt, 95%; (b) PhCH2CHO, NaBH(OAc)3, CH2Cl2, rt, 79%; (c) c-BuCOCl, NEt3, CH2Cl2, rt; (d) LiAlH4, THF, 0 °C ? rt, 9: 86%
(two steps from 3), 10: 55% (three steps from 2); (e) c-PenCOCl, NEt3, CH2Cl2, rt; (f) 1 N HCl, 80 °C; (g) 2-aminobenzaldehyde, CH3SO3H, EtOH, reflux; (h) BBr3, CH2Cl2,
0 °C ? rt.

6304
H. Nagase et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6302–6305
Thus obtained KNT-127 showed higher selectivity for the d-
receptor [Ki (nM) = 21.3 ( ), 153 ( ), 0.16 (d) and Ki ratio: 133.5
/d), 960.5 ( /d)] than did SN-28 [Ki (nM) = 10.8( ), 49.4 ( ),
0.14 (d) and Ki ratio: 77.1 ( d), 352.9 ( d)].
Next, using i.t. injection, we compared the analgesic activity of
the newly obtained compound with that of SN-28 in the acetic acid
writhing method in mice. This compound showed almost the same
potent agonistic activity (ED50 = 0.149 nM) as did SN-28
(ED50 = 0.095 nM) in this test. Finally, we examined the analgesic
effect of KNT-127 given by s.c. injection in the acetic acid writhing
test in mice. As we expected, a very strong dose dependent analge-
sia was observed (ED50 = 1.2 mg/kg), which was about 30 times
more potent than that of TAN-67 (ED50 = 31.4 mg/kg).12
OH
OH
N
l
j
a)
N
O
(l
j
l
j
O
O
l
j
OCH3
N
OCH3
19
1
OH
b)
N
OR
20
R = CH3:
R = H: SYK-33
c)
Thus, we confirmed our above postulation that SN-28 could not
readily cross the BBB and our design to improve the defect of SN-28
led to the potent analgesic agent (KNT-127), with effective sys-
temic administration. Having obtained a lead compound for the
d-agonist, we next attempted to vary the substituent at 17-nitro-
gen in an effort to address the structure–activity relationship. 14-
Hydroxymorphinan derivatives with various 17-substituents were
synthesized in a similar manner to the synthesis of KNT-127
(Schemes 2–4). 17-Isobutyl derivative 19 was obtained by the
reductive cleavage of the 17-cyclopropylmethyl substituent in
compound 1 (Scheme 3).25,26 The results of binding assays of the
thus obtained compounds SYK-3, SYK-4, SYK-14, SYK-33, SYK-58,
and SYK-69 along with those of reference compounds SN-28,
KNT-127, and TAN-67 appear in Table 1.
Scheme 3. Reagents and conditions: (a) 48%HBr, MeOH, PtO2, H2, rt, 56%; (b) 2-
aminobenzaldehyde, CH3SO3H, EtOH, reflux, 70%; (c) BBr3, CH2Cl2, 0 °C ? rt, 72%.
OH
OH
N
N
a)
O
N
OCH3
OR
22
R = H: SYK-58
21
R = CH3:
b)
Scheme 4. Reagents and conditions: (a) 2-aminobenzaldehyde, CH3SO3H, EtOH,
reflux, 71%; (b) BBr3, CH2Cl2, 0 °C ? rt, 61%.
KNT-127 and SYK-4 showed the highest
the highest /d ratio was obtained in SYK-33 and SYK-69. Although
SYK-58 had the strongest affinity for the d-receptor, its selectivities
for the d-receptor over the - and -receptors were low. Among
these newly synthesized 14-hydroxymorphinan derivatives, KNT-
127 had sufficient affinity and the most balanced and highest
selectivity for the d-receptor. Moreover, its affinity and selectivity
for the d-receptor was superior to those of the prototypical d-ago-
nist, TAN-67. These results suggested that the 17-substituent
would be a very important structural determinant for the study
of 14-hydroxymorphinan derivatives with the quinoline moiety
that serve as d-selective opioid ligands.
In conclusion, we designed and synthesized 14-hydroxymorph-
inan derivatives to improve the penetration of SN-28 through the
BBB by use of the hydrogen bond formed between the 14-hydroxy
group and the lone electron pair on the 17-nitrogen. One of these
compounds, KNT-127, showed the most balanced and highest
j/d selectivity, whereas
hand, the basic 17-nitrogen could form ammonium ions under
physiological conditions, and the resulting ionized compound
may be less likely to penetrate through the BBB. We postulated
that the lone electron pair on the 17-nitrogen in SN-28 requires
protection from forming of an ammonium ion. We reported al-
ready that the 14-hydroxy group in naltrexone could form a hydro-
gen bond with the lone electron pair on the 17-nitrogen to produce
a less polar compound (Fig. 4).17 Based on the above discussion, we
designed and synthesized 14-hydroxy-SN-28, that is, KNT-127. The
morphinan 1 was synthesized from naltrexone by the reported
method.18–20 The 17-cyclopropylmethyl group of compound 1
was converted to a 17-methyl group by a previously reported
method.21,22 The treatment of compound 5 with 2-aminobenzalde-
hyde in the presence of CH3SO3H provided compound 6.12 The
resulting compounds 6 was demethylated with BBr3 to afford
KNT-127 (Scheme 1).23,24
l
l
j
Table 1
The binding affinity and selectivity of SN-28, KNT-127, TAN-67, SYK-3, 4, 14, 33, 58, and 69 for the opioid
l
-, d-, and
j
-receptorsa
R1
R2
N
N
OH
Compounds
R1
R2
Affinity (Ki, nM)
Selectivity
l
d
j
l/d
j/d
SN-28
H
OH
Methyl
Methyl
10.8
21.3
284.14
924.3
53.2
10.6
153.9
3.55
0.14
0.16
1.44
6.38
0.57
0.21
0.51
0.14
0.78
49.4
153
732.45
122.8
558
5.72
48.8
1.95
33.6
77.1
133.5
197.5
144.8
93.7
51.3
301.7
25.4
352.9
960.5
508.7
19.2
983.4
27.7
95.7
13.9
43.2
KNT-127
TAN-67b
SYK-3
OH
OH
OH
OH
OH
OH
Benzyl
Phenethyl
Cyclobutylmethyl
Isobutyl
Cyclopropylmethyl
Cyclopentylmethyl
SYK-4
SYK-14
SYK-33
SYK-58
SYK-69
277.9
357.7
a
The binding affinity of each compound was evaluated by displacement of [3H]DAMGO (
brain ( and d) or the guinea pig cerebellum ( ).
Ref. 14.
l j) binding to membranes of mouse whole
), [3H]DPDPE (d), and [3H]U-69,593 (
l
j
b

H. Nagase et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6302–6305
6305
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the acetic acid writhing test even when it was given by s.c. admin-
istration. Its systemically active antinociceptive effect was about
30 times more potent than that of a prototypical d-agonist, TAN-
67. The pharmacological character of the d-receptor has not yet
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known to contribute to the analgesic effects and to severe side ef-
fects like addiction, while the -agonist has antinociceptive and
antipruritic effects without addiction. As KNT-127 induced a
marked effect at low dose through systemic administration, it is
expected to be a useful tool to clarify the real pharmacological
effects via the d-receptor including analgesia and addiction.
l-receptor is
j
Acknowledgments
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We acknowledge the financial supports from Shorai Foundation
for Science and the Uehara Memorial Foundation. We also
acknowledge the Institute of Instrumental Analysis of Kitasato Uni-
versity, School of Pharmacy for its facilities.
22. Nemoto, T.; Fujii, H.; Narita, M.; Miyoshi, K.; Nakamura, A.; Suzuki, T.; Nagase,
H. Bioorg. Med. Chem. Lett. 2008, 16, 4304.
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;
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