Communications to the Editor
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 25 4913
Ta ble 1. Opioid Antagonist Potencies in Smooth Muscle Preparations
Ke (nM)a
selectivity ratio
compd
compd
EK (κ)b
M (µ)b
DADLE (δ)c
κ/µ
κ/δ
1
2
3
4
nor-BNI
GNTI
NTI
0.56
0.14
45
13.7
30
29.4
16.13
10.6
115
0.13
9.08
25
208
(δ/µ ) 226)
5
19
799
(δ/κ ) 346)
3
CNGNTI
3.13
a
Ke ) [antagonist]/(IC50 ratio - 1), where the IC50 ratio is the IC50 of the agonist in the presence of the antagonist (100 nM) divided
by the IC50 of the agonist alone in the same preparation. Ke values were calculated from an average of at least three IC50 ratio values.
Determined using the guinea-pig ileum preparation. c MVD determined using mouse vas-deferens preparation. The agonists employed
b
were ethylketazocine (EKC), morphine (M), and [D-Ala2,D-Leu5]enkephalin (DADLE).
Ta ble 2. Effect on Binding Affinity of Mutational Exchange of Glu297 in the κ Receptor and Lys303 in the µ Receptor
Ki ( SEM (n), nMa
wt κ
Fana
κ[E297K]
Fmut
wt µ
µ[K303E]
b
c
c
c
compd
Fmut
Fmut
1
0.12 ( 0.04 (8)
1.0
12.5 ( 0.92 (5)
123
101.9 ( 10.2 (3)
800
0.77 ( 0.13 (5)
9.3
2
4
0.09 ( 0.01 (3)
0.75
0.27 ( 0.01 (3)
2.25
12.9 ( 0.48 (2)
143
2.48 ( 0.82 (3)
9.2
9.23 ( 1.39 (3)
0.06 ( 0.02 (3)
0.67
0.29 ( 0.05 (3)
1.1
103
4.04 ( 1.23 (3)
15
a
The Ki values were determined in competition binding using [3H]diprenorphine in transiently expressed rat COS-7 cells and analyzed
b
by whole cell binding. The number of individual determinations is indicated in parentheses (n). Fana ) Ki(analogue)/Ki(nor-BNI on the
wt κ receptor). Fmut ) mutational factor, Ki(mutant receptor)/Ki(wt κ receptor).
c
Ack n ow led gm en t. We thank Susanne Hummel-
gaard, Michael Powers, Veronika Philips, and Reynold
Francis for their capable technical assistance in obtain-
ing the in vitro pharmacological data and M. Germana
Paterlini for the colored graphics. This work was sup-
ported by the National Institute on Drug Abuse.
Refer en ces
F igu r e 2. Extracellular loop regions (ELII and ELIII) of the
κ opioid receptor (acidic residues are shaded).
(1) Dhawan, B. N.; Cesselin, F.; Raghubir, R.; Reisine, T.; Bradley,
P. B.; Portoghese, P. S.; Hamon, M. International Union of
Pharmacology. XII. Classification of Opioid Receptors. Pharma-
col. Rev. 1996, 48, 567.
(2) Portoghese, P. S.; Lin, C.-E.; Farouz-Grant, F.; Takemori, A. E.
SAR of N17′ Substituted Norbinaltorphimine Congeners. Role
of the Basic N17′ Group in the Interaction with a Putative
Address Subsite on the κ-Opioid Receptor. J . Med. Chem. 1994,
37, 1495-1500.
(3) Schwyzer, R. ACTH: A Short Introductory Review. Ann. N. Y.
Acad. Sci. 1977, 247, 3-26.
(4) Hjorth, S. A.; Thirstrup, K.; Grandy, D. K.; Schwartz, T. W.
Analysis of Selective Binding Epitopes for the κ-Opioid Receptor
Antagonist nor-Binaltorphimine. Mol. Pharmacol. 1995, 47,
1089.
(5) Metzger, T. G.; Paterlini, M. G.; Portoghese, P. S.; Ferguson, D.
M. Applications of the Message-Address Concept to the Docking
of Naltrexone and Selective Naltrexone-Derived Opioid Antago-
nists into Opioid Receptor Models. Neurochem. Res. 1996, 21,
1287-1294.
(6) Portoghese, P. S.; Sultana, M.; Takemori, A. E. Design of
Peptidomimetic δ-Opioid Receptor Antagonists using the Mes-
sage-Address Concept. J . Med. Chem. 1990, 33, 1714-1720.
(7) Olmsted, S. L.; Takemori, A. E.; Portoghese, P. S. A Remarkable
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[(N2-Alkylamidino)methyl]naltrindole Derivatives as a Novel
Class of Kappa Opioid Receptor Antagonists. J . Med. Chem.
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Antagonists. U.S. Patent 5,457,208, Oct 10, 1995.
(8) The predicted pKa values were calculated using ACD/ILab Web
predicted for 25 °C and zero ionic strength in aqueous solutions.
(9) Kim, K. Y.; Qian, L. Improved Method for the Preparation of
Guanidines. Tetrahedron Lett. 1988, 29, 3183. Poss, M. A.;
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5933-5936.
role of an acidic residue in this position for conferring κ
selectivity. In this regard it is noteworthy that GNTI
has extremely high affinity for both wt κ and µ[K303E]
receptors. Interestingly, the finding that 4 has one-
third to one-fifth the affinity of 2 for the wt κ and mutant
µ receptors suggests that a H-bonding component also
is involved in the interaction of the glutamate residue
with the guanidinyl group.
Significantly, the κ “address” recognition locus for nor-
BNI (1) and GNTI (2) differs from that of the endog-
enous peptide agonist dynorphin-A, which requires the
highly acidic extracellular loop II for high potency and
selectivity (Figure 2).15-17 Thus, it appears that the
binding of peptidic κ agonists and nonpeptide κ antago-
nists may involve a common “message”3 recognition
cavity but different “address” recognition loci.18
In conclusion, this study suggests that the κ “address”
recognition locus for κ antagonists is determined pri-
marily by the glutamate residue. Placement of this
acidic wt κ residue into an equivalent position in the µ
receptor facilitates recognition, as reflected by the
binding of the indolomorphinan GNTI (2) and the
bivalent ligand nor-BNI (1) to wt κ and mutant µ-
[K303E] receptors. These data provide persuasive
evidence that the recognition loci for nor-BNI and GNTI
are very similar in the κ receptor and that the µ[K303E]
receptor also contains an analogous recognition cavity.