Analgesic 14-Alkoxymorphinans
J ournal of Medicinal Chemistry, 2003, Vol. 46, No. 19 4185
4,5r-E p oxy-3-h yd r oxy-5â,17-d im et h yl-14â-[(3-p h en yl-
p r op yl)oxy]m or p h in a n -6-on e Hyd r obr om id e (7‚HBr ). A
mixture of 6 (2.23 g, 4.98 mmol) and 48% HBr (13 mL) was
refluxed for 20 min and then allowed to cool to room temper-
ature. The mixture was treated with MeOH (20 mL) and
evaporated almost to dryness. This procedure was repeated
twice whereas the evaporation the last time was carried out
until dryness. The residue (2.48 g beige crystals) was recrys-
tallized from MeOH (15 mL) to give 2.26 g (88%) of pure
7‚HBr: white crystals; mp > 270 °C (dec); IR (KBr) 3473 (OH),
1725 (CdO) cm-1; 1H NMR (DMSO-d6) δ 9.42 (s, br, NH+), 8.58
(s, br, OH), 7.36-7.16 (m, 5 arom H), 6.69 (d, J ) 8.2, 1 arom
H), 6.64 (d, J ) 8.2, 1 arom H), 2.97 (d, J ) 4.6, CH3N+), 1.52
(s, CH3-C(5)); 13C NMR (DMSO-d6) δ 209.7, 142.9, 141.7, 139.9,
128.8, 128.3 (2C), 128.2 (2C), 125.8, 120.5 119.9, 118.0, 94.4,
75.4, 60.8, 59.4, 48.9, 46.7, 41.3, 33.1, 31.7, 30.7, 24.2, 23.5,
17.0; CI-MS m/z 434.3 (M+ + 1). Anal. (C27H31NO4‚HBr) C, H,
N.
the phasetransfer catalyst) and brine (200 mL), dried
(Na2SO4), and evaporated to give 1.25 g of a yellow oil.
Purification of this crude product by flash chromatography
(silica gel, elution with CH2Cl2/MeOH/concentrated NH4OH
solution, 250:2:0.5) afforded 1.00 g of a colorless oil which was
converted into the hydrochloride salt with Et2O/HCl and
recrystallized from Et2O/i-PrOH to yield 0.86 g (50%) of pure
10‚HCl: slightly yellow powder; mp 135-138 °C (dec); IR
(KBr) 1700 (CdO) cm-1 1H NMR (DMSO-d6) δ 9.36 (s, br,
;
NH+), 7.31-7.18 (m, 5 arom H), 7.05 (d, J ) 8.4, 1 arom H),
6.95 (d, J ) 8.4, 1 arom H), 3.80 (CH3O), 3.79 (CH3O), 2.81 (d,
J ) 4.8, CH3N+), 1.53 (d, J ) 7.0, CH3-C(5)); 13C NMR (DMSO-
d6) δ 209.6, 151.7, 147.2, 142.2, 134.9, 128.2 (2C), 128.1 (2C),
125.8, 125.6, 122.9, 112.4, 74.5, 60.5, 59.8, 56.7, 55.8, 48.2, 47.1,
44.7, 40.7, 32.6, 31.6, 30.4, 26.5, 25.3, 24.0, 12.6; CI-MS m/z
464.3 (M+ + 1). Anal. (C29H37NO4‚HCl‚1.0H2O) C, H, N.
Op ioid Recep tor Bin d in g Assa ys. Membranes were
prepared from whole rat or guinea-pig brains as previously
described.24 Briefly, tissues were homogenized in 5 volumes
of ice-cold 50 mM Tris-HCl buffer (pH 7.4). After centrifugation
at 40 000g for 20 min at 4 °C, the membrane pellets were
resuspended in 30 volumes of 50 mM Tris-HCl buffer and
incubated at 37 °C for 30 min. The centrifugation step
described above was repeated, the final pellet was resuspended
in 50 mM Tris-HCl buffer and stored at -70 °C until use.
Binding experiments were performed in 50 mM Tris-HCl
buffer in a final volume of 1 mL containing 0.3-0.5 mg protein
as described.24 Rat brain homogenates were incubated either
with the µ-selective radioligand [3H]DAMGO (0.5 nM, 45 min,
35 °C)25 or with the δ-selective radioligand [3H][Ile5,6]deltorphin
II (0.5 nM, 45 min, 35 °C).26 Guinea pig brain homogenates
were incubated with the κ-selective radioligand [3H]U69,593
(1 nM, 30 min, 30 °C)27 in the presence of increasing concen-
trations of the test compound. Reactions were terminated by
rapid filtration through Whatman GF/B pretreated with 0.1%
polyethyleneimine ([3H]U69,593) or GF/C ([3H]DAMGO and
[3H][Ile5,6]deltorphin II) glass fiber filters using a Brandel Cell
Harvester, followed by three washings with 5 mL of ice-cold
50 mM Tris-HCl buffer. The bound radioactivity was measured
in Ultima Gold scintillation cocktail, using a Beckman LS1701
liquid scintillation counter. Nonspecific binding was defined
in the presence of 10 µM unlabeled naloxone. Protein concen-
tration was determined using bovine serum albumine as a
standard.28 All experiments were carried out in duplicate. The
values presented are the mean ( SEM of 3-4 independent
experiments. Competition inhibition constant (Ki) values were
calculated with the GraphPad Prism (version 3.0, San Diego,
CA) program.
In Vivo Assa ys. All animals received care according to the
Guide for the Care and Use of Laboratory Animals, U.S.
Department of Health and Human Services, 1985. The facili-
ties are certified by the American Association for the Ac-
creditation for Laboratory Animal Care. These studies were
approved by the Institutional Animal Care and Use Committee
at Virginia Commonwealth University.
Gen er a l Meth od s. ICR male mice (Harlan-Sprague-
Dawley, Inc., Indianapolis, IN) weighing 20-30 g were used.
Each animal was tested once only. All drugs were given by
the subcutaneous (sc) route. At least three doses were tested,
and 6-10 animals/dose were used. The drugs were dissolved
in 5% hydroxypropyl-â-cyclodextrin in water or in dilute HCl
(vehicles).
4,5r-Ep oxy-5â,17-d im eth yl-14â-[(3-p h en ylp r op yl)oxy]-
3-[(p r op -2-yn yl)oxy]m or p h in a n -6-on e H yd r och lor id e
(8‚HCl). A mixture of 7 (0.38 g, 0.88 mmol, the free base was
obtained from the hydrobromide), K2CO3 (0.37 g, 2.68 mmol),
propargyl bromide (0.32 g, 2.66 mmol), and acetone (25 mL)
was refluxed for 7 h. The inorganic material was filtered off,
and the filtrate was evaporated to give 0.46 g of an orange oil.
Purification of this crude product by flash chromatography
(silica gel, elution with CH2Cl2/MeOH/concentrated NH4OH
solution, 250:2:0.5) afforded 0.35 g of a yellow oil which was
precipitated as hydrochloride from Et2O to give 0.22 g (49%)
of pure 8‚HCl: white powder; mp 130-133 °C; IR (KBr) 1727
1
(CdO) cm-1; H NMR (DMSO-d6) δ 9.38 (s, br, NH+), 7.34-
7.19 (m, 5 arom H), 6.92 (d, J ) 8.4, 1 arom H), 6.78 (d, J )
8.4, 1 arom H), 4.81 (d, J ) 1.0, HCCCH2), 3.59 (t, J ) 1.0,
HCCCH2), 2.94 (d, J ) 3.8, CH3N+), 1.53 (s, CH3-C(5)); 13C
NMR (DMSO-d6) δ 209.4, 144.4, 141.8, 140.1, 129.3, 128.3 (2C),
128.2 (2C), 125.7, 124.0, 120.0, 117.2, 95.3, 79.1, 78.4, 75.2,
60.8, 59.1, 56.8, 48.9, 46.4, 41.3, 33.0, 31.8, 30.6, 24.3, 23.8,
23.5, 16.9; CI-MS m/z 472.3 (M+ + 1). Anal. (C30H33NO4‚HCl‚
1.0H2O) C, H, N.
4-H yd r oxy-3-m et h oxy-5â,17-d im et h yl-14â-[(3-p h en yl-
p r op yl)oxy]m or p h in a n -6-on e Hyd r och lor id e (9‚HCl). To
a refluxing mixture of 6 (3.70 g, 8.27 mmol), NH4Cl (4.50 g,
84 mmol), and EtOH (50 mL) was added 5.50 g activated zinc
powder (84 mmol) in five portions. After refluxing for 47 h,
the inorganic material was filtered off, and the filtrate was
evaporated to give 6.35 g of a white foam. This was dissolved
in H2O (200 mL) and alkalinized with concentrated NH4OH
solution, and the mixture was extracted with CH2Cl2 (1 × 100
mL, 4 × 50 mL). The combined organic layers were washed
with H2O (4 × 200 mL) and brine (200 mL), dried (Na2SO4),
and evaporated to give 3.93 g of a yellow oil. Purification of
this crude product by flash chromatography (silica gel, elution
with CH2Cl2/MeOH/concentrated NH4OH solution, 250:2:0.5)
afforded 2.99 g (80%) of a yellow oil which was not crystallized
for the next step. An analytical sample was precipitated as
hydrochloride from Et2O and recrystallized from Et2O/
i-PrOH: slightly yellow powder; mp > 210 °C (dec); IR (KBr)
1
3415 (OH), 1700 (CdO) cm-1; H NMR (DMSO-d6) δ 9.64 (s,
br, NH+), 8.88 (s, OH), 7.33-7.17 (m, 5 arom H), 6.91 (d,
J ) 8.2, 1 arom H), 6.70 (d, J ) 8.2, 1 arom H), 3.78 (CH3O),
2.80 (d, J ) 3.4, CH3N+), 1.28 (d, J ) 6.6, CH3-C(5)); 13C
NMR (DMSO-d6) δ 210.5, 146.7, 144.3, 142.2, 128.2 (2C),
128.1 (2C), 127.6, 125.9, 125.6, 118.2, 110.5, 74.6, 61.9, 60.4,
56.9, 56.0, 47.2, 46.8, 44.3, 32.7, 31.7, 30.6, 25.4, 25.2, 24.1,
13.5; CI-MS m/z 450.3 (M+ + 1). Anal. (C28H35NO4‚HCl‚
1.1H2O) C, H, N.
Hot P la te Test (HP ). A modified method previously
described was used.29 A 1000 mL Pyrex beaker (bottom
removed) was placed on the hot plate maintained at 56 °C.
The test was initiated by placing a mouse in the specially
designed beaker. This arrangement served to confine a mouse
to a specific area of the hot plate. Each mouse was exposed to
the hot plate for two trials spaced 5 min apart. Only mice that
gave a control response latency in the range of 6-10 s on both
trials served as subjects. Each subject received a dose of test
drug and 30 min later was again tested on the hot plate.
Activity was scored as positive if the mouse jumped, licked,
or shook its paws at least 5 s beyond its average control
3,4-Dim eth oxy-5â,17-d im eth yl-14â-[(3-p h en ylp r op yl)-
oxy]m or p h in a n -6-on e H yd r och lor id e (10‚H Cl). A mix-
ture of 9 (1.55 g, 3.44 mmol), 40% tetrabutylammonium
hydroxide solution (20 g), dimethyl sulfate (0.48 g, 3.80 mmol),
and CH2Cl2 (15 mL) was vigorously stirred under N2 at room
temperature. After 12 h, H2O (300 mL) was added, and the
mixture was extracted with CH2Cl2 (3 × 50 mL). The combined
organic layers were washed with H2O (6 × 200 mL, to remove