Derivatives of tramadol for increased duration of effect

Article abstract of DOI:10.1016/j.bmcl.2005.10.024

Tramadol is a centrally acting opioid analgesic structurally related to codeine and morphine. Analogs of tramadol with deuterium-for-hydrogen replacement at metabolically active sites were prepared and evaluated in vitro and in vivo.

Full text of DOI:10.1016/j.bmcl.2005.10.024

Bioorganic & Medicinal Chemistry Letters 16 (2006± 691–694
Derivatives of tramadol for increased duration of effect
*
Liming Shao, Craig Abolin, Michael C. Hewitt, Patrick Koch and Mark Varney
Sepracor Inc., 84 Waterford Drive, Marlborough, MA 01752, USA
Received 23 August 2005; revised 6 October 2005; accepted 7 October 2005
Available online 27 October 2005
Abstract—Tramadol is a centrally acting opioid analgesic structurally related to codeine and morphine. Analogs of tramadol with
deuterium-for-hydrogen replacement at metabolically active sites were prepared and evaluated in vitro and in vivo.
Ó 2005 Elsevier Ltd. All rights reserved.
Tramadol (± ±-cis is used for the treatment of moderate
1
develop an improved tramadol analog with a longer
half-life.
to moderately severe pain. Tramadol has a relatively
short duration of analgesic effect due to extensive first
pass metabolism and as such is dosed as frequently as
2
Our approach to tramadol analogs took advantage of
4
the primary kinetic isotope effect to slow CYP450-med-
1
00 mg every 4–6 h. The analgesic activity of tramadol
is thought to be the result of a dual mechanism—the
parent (1± acts as an inhibitor of norepinephrine and
serotonin reuptake, and the major O-desmethyl metabo-
iated metabolism (Fig. 2±. Replacing hydrogen with deu-
terium at metabolically active sites can result in a slower
metabolism due to the reduced rate of cleavage of a C–D
bond relative to a C–H bond. This approach has been
shown to be effective for a number of pharmacological
lite (M1± is a potent agonist at l-opioid receptors
3
(
Fig. 1±.
5
6
agents including amphetamine, butethal, and mor-
7
phine. For tramadol, formation of metabolite M1 is
The opioid antagonist naloxone only partially inhibits
the analgesic activity of tramadol in animal tests. Not
unlike other opioids, tramadol causes a number of side
effects including constipation, nausea, dizziness, and
somnolence. A tramadol analog that had a longer
half-life (and therefore required less-frequent dosing±
and was devoid of opioid side effects would have thera-
peutic benefit. Herein, we wish to disclose our efforts to
both the primary route of metabolism and is responsible
for its opioid-like effects. By slowing down metabolism
of tramadol, it was anticipated that the formation of
M1 would be reduced, providing a longer-acting drug
with potentially fewer opioid-related side effects. To-
ward that end, the hydrogen atoms on the O-methyl
and N-methyl groups were replaced by deuterium as
shown in structures D3, D6, D9, and D6 M1.
OCH3
OH
OR1
OH
OH
CH3
CH3
OH
N
N
R2
N
R2
CH3
CH3
1
M1
1
2
D R = CD , R = CH
3
3
3
Figure 1. (± ±-Tramadol 1 and metabolite (± ±-M1.
1
2
D R = CH , R = CD
6
3
3
1
2
D M R = H, R = CD
3
6
1
1
2
D R = CD , R = CD
3
9
3
Keywords: Tramadol; Metabolic stability; Deuterium isotope effect;
Opioids.
Dramadol analogs
*
Corresponding author. Tel.: +1 5083577467; fax: +1 5083577467;
e-mail: liming.shao@sepracor.com
Figure 2. D-tramadol (dramadol± analogs.
0
960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.10.024

692
L. Shao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 691–694
O
OR2
OH
R1 1. Mg, THF, 4 or 5
1. n-BuLi, PHPh , THF
2
N
R1
2
. HCl, Et O
2. HCl, Et
2
O
OH
2
OH
R1
R1
1
2
3
R = CH
N
R1
N
3
OR2
R1
1
HCl
R = CD3
HCl
1
2
1
6
7
(D ) R = CH , R = CD
8 (D
6
M1) R = CD
3
3
3
3
Br
1
2
(D ) R = CD , R = CH
6
3
3
2
4
5
R = CH
3
2
R = CD3
Scheme 1. Synthesis of D3, D6, and D6 M1 metabolites.
mation seen by LCMS and HPLC analysis. This led
to an alternate strategy to synthesize D9 that relied
on a lithium–halogen exchange reaction that had been
outlined previously in the literature, and was suc-
cessfully employed using Mannich base 3 and aromat-
ic bromide 5 (Scheme 2±.
OCD3
CD3
1. t-BuLi, Et O, 5
2
N
HCl
10
CD3
2. HCl, Et O
2
OH
CD
N
3
OCD3
CD3
Crude 9 (D9± was also converted to its HCl salt and
recrystallized several times in isopropanol to provide a
Br
9 (D )
9
1
1
9
3% pure cis isomer.
Scheme 2. Synthesis of 9 (D9±.
The receptor binding affinity and functional monoamine
uptake inhibition by tramadol and the deuterated tram-
adol analogs are shown in Table 1.
The syntheses of the deuterated tramadol analogs
were relatively straightforward and began with Man-
nich base 2 or 3 (Scheme 1±. Specifically, addition of
the Grignard reagent 4 or 5 to ketones 2 or 3 provid-
ed crude 6 (D3) and 7 (D6± as mixtures of diastereo-
mers (76% cis for D6±. The abundance of the cis
isomer could be improved to 93% after formation of
the HCl salt and multiple recrystallizations from iso-
As expected, the deuterated analog of M1 retained the
l-opioid receptor binding affinity present in metabolite
M1. In general, the deuterated analogs retained in vitro
activity comparable to their non-deuterated parent mol-
ecules, with a slightly diminished reuptake inhibition of
norepinephrine compared to tramadol.
8
propanol. Demethylation of 7 (D6± occurred without
9
incident to provide phenol 8 (D6 M1±. Purification
The in vitro metabolic stability results for the deuterated
1
2,13
was effected via formation of the HCl salt and recrys-
tallization from isopropanol/ethyl acetate to give the
phenol salt as an analytically pure white powder.
The reaction to form D9 was attempted several times
with starting materials 3 and 5, with little product for-
tramadol compounds are detailed in Table 2.
Based
on disappearance of parent compound in incubation
media, the most stable compound of the six tested in hu-
man liver microsomes and hepatocytes was clearly the
D9-tramadol derivative 9. Tramadol and D6-tramadol
Table 1. l-Opioid binding, and 5-HT and NE reuptake inhibition of (± ±-cis tramadol and deuterated derivatives
OR2
OR3
R1
N
R1
1
2
R
3
R
Compound
R
IC50 (nM±
l
5-HT
NE
1
Tramadol
Metabolite M1
CH
CH
CH
3
3
3
CH
H
3
H
H
H
H
H
H
7600
47
4300
4600
1900
3100
9900
1100
790
>10,000
3600
6
7
8
9
(D3±
(D6±
CD
CH
H
3
>10,000
>10,000
43
CD
3
3
3200
(D6 M1±
(D9±
CD
CD
3
3
6700
2000
CD
3
5300
Note: IC50 is the concentration required to inhibit 50% binding to l-opioid receptors or reuptake of 5-hydroxytryptamine (5-HT± and norepinephrine
NE±.
(

L. Shao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 691–694
693
Table 2. Summary of in vitro metabolic stability of tramadol and
deuterated derivatives
hepatocytes and microsomes in Figures 3 and 4, respec-
tively. Consistent with a deuterium isotope effect on the
metabolically labile O-methyl group, 6 (D3± and 9 (D9±
slowed the formation of the primary metabolite M1 by
approximately 5-fold. The formation rate of metabolite
M1 from 7 (D6± was nearly indistinguishable from that
of tramadol in human hepatocytes.
a
Half-life (min±/rank
b
Compound
c
HLM
Hepatocytes
9
8
(D9±, (+/À±-cis
7210/1
747/5
2820/2
744/4
741/3
547/6
3610/1
1650/2
1560/3
1330/4
968/5
(M1 D6±, (+/À±-cis
Metabolite M1, (+/˱-cis
6
1
7
(D3±, (+/À±-cis
The in vivo activities of tramadol (1±, 7 (D6±, and 9 (D9±
were tested in the rat tail-flick model. Tramadol (1±, 7
(D6±, and 9 (D9± were administered at a dose of
Tramadol, (+/˱-cis
(D6±, (+/À±-cis
852/6
a
Calculated from samples taken at 0, 180, and 360 min for hepatocytes
and 0, 90, and 180 min for microsomes.
50 mg/kg intraperitoneally. Intraperitoneal morphine
b
c
(5 mg/kg± and saline were used as controls. Analgesic
activity (tail-flick latency± was evaluated at 1, 2, 4, and
Ranked from 1, the most stable to 6, the least stable.
HLM = human liver microsomes.
8 h after the dose. The results of efficacy (tail-flick laten-
cy± are shown in Figure 5.
7
were the least stable compounds in human liver micro-
somes and hepatocytes.
Tramadol and two of its deuterated derivatives (7 (D6±
and 9 (D9±± significantly lengthened tail-flick latency
when tested at 1 and 2 h after intraperitoneal adminis-
tration of 50 mg/kg. Tramadol and 9 (D9± were also
effective when tested 4 h after dosing. By 8 h the effects
of the test compounds were no longer statistically signif-
icant. As expected, morphine increased tail-flick latency
at the 1 h time point, but at later times the effect was not
significantly greater than that of vehicle. The results
indicate that the test compounds effectively reduced
acute nociception for 2–4 h following a single acute
intraperitoneal dose.
Metabolic stability was also evaluated based on the for-
mation rate of metabolite M1 from tramadol (1± and
three tramadol deuterated derivatives capable of pro-
ducing metabolite M1. The results are summarized for
100
D6 Tramadol
D3 Tramadol
D9 Tramadol
8
0
0
0
0
In conclusion, the substitution of deuterium for hydro-
gen in the methyl groups of tramadol did not adversely
affect the in vitro binding affinity. In vitro testing in hu-
man microsomes confirmed that replacing hydrogen
with deuterium in the metabolically labile O-methyl po-
sition of 1 slowed the formation of the primary metabo-
lite M1 by approximately 5-fold. The deuterated
derivatives 7 (D6± and 9 (D9± were active analgesics in
the rat tail-flick model, but were not superior to trama-
dol in terms of potency or duration of effect. Deuterium
6
4
2
0
180
360
Incubation Time (min)
Figure 3. Stability of tramadol analogs in human hepatocytes.
25
**
**
**
**
**
1
00
20
15
10
5
0
**
D6 Tramadol
D3 Tramadol
D9 Tramadol
**
**
*
80
60
40
20
60
120
240
480
Minutes
Saline Control
Morphine Control
Tramadol
D6
D9
90
180
Incubation Time (min)
Dunnett's test: *, **/ p<0.05, p<0.01 (n = 9 - 10 per group)
Figure 4. Stability of tramadol analogs in human liver microsomes.
Figure 5. Effect of test compounds on tail-flick latency.

694
L. Shao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 691–694
1
for hydrogen replacement at metabolically active sites
had no obvious deleterious effects in vivo but did not
result in a longer duration of effect. In this case,
deuteration at metabolically active sites produced a
pharmacological agent equipotent in vivo with
tramadol.
9. NMR data for 18 (D6 M1±: H NMR (400 MHz, CD
3
OD±
.14–7.11 (m, 1H±, 6.90–6.88 (m, 2H±, 6.62–6.60 (m, 1H±,
.91–2.85 (m, 1H±, 2.61 (dd, J = 2.57, 13.2 Hz, 1H±, 2.13–
7
2
2
1
3
.07 (m, 1H±, 1.94–1.43 (m, 8H±; C NMR (100 MHz,
OD± 158.8, 150.5, 130.6, 117.1, 114.7, 113.2, 75.8,
1.8, 43.1, 41.5, 27.1, 26.2, 22.4; mp: 221–226 °C; TLC
20% MeOH in CH Cl ± R = 0.23.
CD
3
6
(
2
2
f
1
0. Draper, R. W.; Hou, D.; Iyer, R.; Lee, G. M.; Liang, J. T.;
Mas, J. L.; Vater, E. J. Org. Process Res. Dev. 1998, 2, 186.
1
11. NMR data for 19 (D9±: H NMR (400 MHz, CD OD±
References and notes
3
7.27–7.24 (m, 1H±, 7.09–7.04 (m, 2H±, 6.80 (d, J = 8.07 Hz,
1H±, 2.94 (dd, J = 9.16, 13.2 Hz, 1H±, 2.21–2.16 (m, 1H±,
1
. (a± Raffa, R. B.; Friderichs, E.; Reimann, W.; Shank, R.
P.; Good, E. E.; Vaught, J. L. J. Pharmacol. Exp. Ther.
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1.95–1.50 (m, 8H±; C NMR (100 MHz, CD OD± 161.0,
3
3
1
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150.1, 130.5, 117.9, 112.7, 112.0, 75.6, 61.5, 42.8, 41.3,
27.0, 25.9, 22.1; mp: 176–182 °C; TLC (10% MeOH in
CH Cl ± R = 0.45.
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2
f
12. General procedure for microsomal stability assay in
Human liver microsomes: The test compounds at 23 lM
were incubated separately with human liver microsomes
(4 mg protein/mL± at 37 ± 1 °C in 3-mL incubation
mixtures containing potassium phosphate buffer
(50 mM, pH 7.4±, MgCl2 (3 mM±, and EDTA (1 mM±.
Reactions were started by the addition of the NADPH-
generating system. At designated times (0, 90, and
180 min±, a 500-lL aliquot was removed from the
incubation and added to 500 lL acetonitrile to terminate
the reaction. The amount of unchanged test compound
was quantified by LC/MS/MS. Where appropriate, the
amount of O-desmethyl metabolite formed was also
assessed by LC/MS/MS.
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for
Ò
13. General procedure for in vitro stability assay in human
hepatocytes. Test compounds (23 lM± were incubated
separately with a pool (n = 2± of cryopreserved human
hepatocytes (2 million cells/mL± in WaymouthÕs+ (Way-
mouthÕs medium [without phenol red] supplemented
with FBS (4.5%±, insulin (5.6 lg/mL±, glutamine
(3.6 mM±, sodium pyruvate (4.5 mM±, and dexametha-
sone (0.9 lM±± at the final concentrations indicated.
Each test article was added to incubations in 2.5 lL
(1%± of methanol. Reactions were started when placed
in the incubator. At designated times (0, 180, and
360 min±, a 250-lL aliquot was removed from the
incubation and added to 250 lL acetonitrile to terminate
the reaction. Precipitated protein was removed by
centrifugation (920g for 10 min at 10 °C±. Amount of
unchanged parent compound was quantified as follows;
an aliquot (20 lL± of the supernatant fraction was
transferred to 1 mL of internal standard (50 ng/mL
dextrorphan in 1:1 methanol/water, 51-fold dilution± and
analyzed by LC/MS/MS. Additional incubations were
performed with hepatocytes in which the test article was
replaced with 7-ethoxycoumarin (500 lM, marker sub-
strate± to determine if the hepatocytes were metaboli-
cally competent.
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. NMR datafor 16 (D3±: H NMR (400 MHz, CD OD± 7.36–
1
3
7
2
1
1
2
.31 (m, 1H±, 7.16–7.11 (m, 2H±, 6.85–6.83 (m, 1H±, 3.04–
.97 (m, 1H±, 2.74–2.64 (m, 7H±, 2.34–2.26 (m, 1H±, 1.97–
.60 (m, 8H±; C NMR (100 MHz, CD OD± 161.3, 150.5,
3
1
3
30.6, 118.2, 113.0, 112.2, 75.8, 61.8, 46.2, 42.8, 42.6, 41.6,
7.3, 26.0, 22.4; mp: 180–184 °C; TLC (10% MeOH in
1
= 0.48. NMR data for 17 (D6±: H NMR
CH
(
2
Cl
2
± R
f
400 MHz, CD
3
OD± 7.30–7.27 (m, 1H±, 7.10–7.05 (m, 2H±,
.83–6.80 (m, 1H±, 3.79 (s, 3H±, 2.98–2.93 (m, 1H±, 2.66–
6
2
1
3
.62 (m, 1H±, 2.25–2.20 (m, 1H±, 1.94–1.51 (m, 8H±;
OD± 162.3, 151.4, 131.6, 119.1, 113.8,
13.2, 76.7, 62.6, 56.5, 43.8, 42.4, 27.9, 27.0, 23.3; mp: 175–
80 °C; TLC (10% MeOH in CH Cl ± R = 0.43.
C
NMR (100 MHz, CD
1
1
3
2
2
f