Unique spirocyclopiperazinium salt. Part 4: Modification of dispirocyclopiperazinium (DSPZ) salts as analgesics

Article abstract of DOI:10.1016/j.bmcl.2007.07.010

In order to improve the analgesic activity of lead compound 7a, two series of dispirocyclopiperazinium (DSPZ) salts 9a-h, 10a-e and compounds 14, 15 were synthesized and evaluated for their in vivo analgesic activity both by acetic acid induced writhing t

Full text of DOI:10.1016/j.bmcl.2007.07.010

Bioorganic & Medicinal Chemistry Letters 17 (2007) 5078–5081
Unique spirocyclopiperazinium salt. Part 4: Modification
of dispirocyclopiperazinium (DSPZ) salts as analgesics
Ang Li,a Xin Wang,a Cai-Qin Yue,b Jia Ye,b Chang-Ling Lib and Run-Tao Lia,c,
*
aDepartment of Chemical Biology, School of Pharmaceutical Sciences, Peking University, Beijing 100083, PR China
bDepartment of Molecular and Cellular Pharmacology, School of Pharmaceutical Sciences,
Peking University, Beijing 100083, PR China
cState Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University,
Beijing 100083, PR China
Received 28 April 2007; revised 5 July 2007; accepted 6 July 2007
Available online 13 July 2007
Abstract—In order to improve the analgesic activity of lead compound 7a, two series of dispirocyclopiperazinium (DSPZ) salts 9a–
h, 10a–e and compounds 14, 15 were synthesized and evaluated for their in vivo analgesic activity both by acetic acid induced writh-
ing test and hot plate test. Compounds 9h, 14, and 15 exhibited better analgesic activities than 7a. Several important structure–activ-
ity relationships were revealed from this study: (1) the introduction of aryl group would obviously improve the activity; (2) it was
favorable to enhance the analgesic activity and reduce the toxicity to incorporate alkyl group with suitable length in the molecule; (3)
carbamate analogues displayed lower toxicity than carboxylic ester analogues; (4) hydroxylation and chlorination of lead compound
could increase the analgesic activity in hot plate test.
ꢀ 2007 Elsevier Ltd. All rights reserved.
Pain is a pervasive public health problem. The chief
treatments of pain are agents that bind at opioid recep-
tors and inhibitors of cyclooxygenase.1 However, both
general classes of agents have undesirable side effects
associated with their use,2 and this has prompted a
search for mechanistically different analgesic agents.
Over the past few years, considerable efforts have been
directed toward the identification of ligands selective
for subtypes of nAChR and several high affinity com-
pounds have been reported.3
sented mainly by the a4 b2 subtype). The structural sim-
ilarity of the two analgesic compounds encouraged us to
further study piperazinium salts as analgesics. Hence, we
synthesized several more classes of piperazinium salts,7–12
including nonspirocyclopiperazinium salts (NSPZ) (3 and
4), monospirocyclopiperazinium (MSPZ) (5 and 6),
dispirocyclopiperazinium (DSPZ) salts (7 and 8), and sev-
eral compounds were found to own good analgesic activ-
ity (Fig. 1).11,12
However, due to the poor lipophilicity of piperazinium
salts, those compounds could not cross the blood–brain
barrier easily, the pharmacological experiments of com-
pound 3 (R = methyl, R0 = phenethyl, n = 6) proved
that its analgesic effective dose of subcutaneous injection
was 4000 times higher than that of intracerebroventric-
ular injection,13 which hinted that improving CNS per-
meability of piperazinium salts may be a valid method
to increase the analgesic activity of quaternary ammo-
During our study on the synthesis and biological activity
of quaternary piperazinium salts,4,5 we found a signifi-
cant analgesic piperazinium salt 1, whose structure is
similar to a well-known nicotinic agonist N1,N1di-
methyl-N4-phenylpiperazinium iodide (DMPP, 2).
DMPP is considered to represent a unique ligand among
hundreds of nicotinic agonists,6 because it does not fit
any proposed pharmacophore for nicotinic binding, yet
it presents a Ki = 250 nM as a nicotinic receptor of the
rat brain labeled by [3H]cytisine (thought to be repre-
nium salts. Therefore, we selected
a promising
compound 7a (80.97%, 0.04 mmol/kg, sc, in writhing
test) as the lead compound to improve its lipophilicity.
Ester formation is the most commonly employed
approach for increasing lipophilicity of highly polar
parent compounds.14 Besides, carbamates were usually
used in cholinocepter agonists and acetylcholinesterase
Keywords: Dispirocyclopiperazinium salts; Analgesic; Structure–activ-
ity relationship; Synthesis.
*
Corresponding author. Tel.: +86 10 82801504; fax: +86 10
82716956; e-mail: lirt@mail.bjmu.edu.cn
0960-894X/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.07.010

A. Li et al. / Bioorg. Med. Chem. Lett. 17 (2007) 5078–5081
5079
O
C
N+
N
I-
Br- HCl
N+
N
Cl
b
a
C6H5
N
NH
HN
NH
OH
2 (DMPP)
1
11
R
R
2Br- 2HCl
N R'
N+
N+
R=Alkyl, Benzyl
R'=H, Alkyl
n=2,4,6,8,10
HN
HN
N
C6H5CO N
C6H5CO N
N
N
R'
N
c
d
e
(CH2)n
OH
CHOH
N
3
12
13
H3C
R
CH3
R=Methyl, Benzyl
n=2,4,6,8
N+
N CO(CH2)nCO N
N+
2Br-
R
4
N+
N+
N
O
N+
N+
N
N
f
OCR 2Br-
OH 2Br-
O
N
X-
N+
NR
N+
N
R Br-
HCl
9
14
R=Aryl
R=Alkyl
g
9a-g: R=(CH2)nCH3
n=0,2,4,7,10,14,16
9h: R=Ph
6
5
h
7a: n=3 7b: n=6
7c: n=8 7d: n=10
2Br-
N+
N
(CH2)n
N
N+
N+
N+
N
N
N+
N+
N
O
7
OCNHR 2Br-
Cl 2Br-
O
O
N
N+
N C (CH2)n C N
N+
2Br-
15
n=1-6
10
8
10a-d: R=(CH2)nCH3
n=2,4,7,10
10e: R=Ph
Figure 1. Known quaternary ammoniums with analgesics.
Scheme 1. Synthesis of compounds 9, 10, and 15. Reagents and
conditions: (a) C6H5COCl, AcOH, 56%; (b) ClCH2CH(OH)CH2Cl
(0.5 equiv), K2CO3, EtOH, reflux, 61%; (c) 10% HCl; (d) NaOH, 85%
overall; (e) Br(CH2)4Br (2 equiv), NaHCO3, EtOH, reflux, 76%; (f)
(RCO)2O, DMAP, DMF, 70–100 ꢁC, 50–70%; (g) RNCO, TEA,
DMF, 70–100 ꢁC, 40–60%; (h) SOCl2, DMF, rt, 29%.
inhibitors.15,16 Hence, two series of DSPZ salt conju-
gates carboxylic esters 9 and carbamates 10 were de-
signed (Fig. 2).
The designed compounds (9 and 10) were synthesized as
outlined in Scheme 1. The reaction of two equivalents of
1-benzoyl piperazine 11 with one equivalent of 1,3-
dichloro-2-propanol in the presence of potassium
carbonate provided 12. Deprotection of 12 in 10%
hydrochloric acid, followed by neutralization with aque-
ous sodium hydroxide, gave the 1,3-di(1-piperazyl)-2-
propanol 13. One equivalent of 13 was reacted with
two equivalents of 1,4-dibromobutane to yield the key
intermediate dispirocyclopiperazinium bromide 14.
Treatment of compound 14 with SOCl2 afforded the
compound 15.
trying several different methods, we found that 14 was
treated with 20 equivalents of corresponding acid anhy-
drides in DMF with 4-dimethylaminopyridine (DMAP)
as catalyst for 3h at 70–100 ꢁC affording the expected
products 9 in 50–70% yields. The compounds 9 were
purified by column chromatography on neutral alumi-
num oxide using methanol/ethyl acetate as the eluents.
Compounds 10 were obtained from the reaction of 14
with corresponding isocyanates which were prepared
according to the literature.17 The reaction was carried
out in DMF with triethylamine (TEA) as base for 7 h
at 70–100 ꢁC giving the products in 40–60% yields.
The purification of products 10 was similar to that of
series 9.
Compounds 9 were synthesized from 14. Because of its
poor solubility and steric blocks, quaternary ammonium
salts 9 were not obtained from conventional acylation
reagents such as acid chloride and carboxylic acid. After
All newly synthesized compounds were tested for their
in vivo analgesic activity by acetic acid induced writhing
test and hot plate test using reported method.13 Gener-
ally, we chose 0.04 mmol/kg as the dose of subcutaneous
injection. If causing death of mice, the dose would be re-
duced. The results are summarized in Table 1.
N+
N+
N
N
N+
N+
N
N
O
O
2Br-
2Br-
OCNHR
OCR
10
9
9a-g: R=(CH2)nCH3
n=0,2,4,7,10,14,16
9h: R=Ph
10a-d: R=(CH2)nCH3 n=2,4,7,10
10e: R=Ph
As shown in Table 1, the analgesic activity data have an
approximate normal distribution for the compounds 9a–g.
For instance, when n = 4, 7, and 10, compounds in series
Figure 2. Structures of compounds 9 and 10.

5080
A. Li et al. / Bioorg. Med. Chem. Lett. 17 (2007) 5078–5081
Table 1. Analgesic activity of compounds 7a, 9, 10, 14, 15 by acetic acid induced writhing test and hot plate test in mice
N+
N+
N
N
N+
N+
N
N
N+
N+
N
N
O
O
2Br-
X
OCNHR 2Br-
OCR 2Br-
7a: X=H
14: X=OH
15: X=Cl
9
10
Compound
R(X)
Me
Writhing test
Hot plate
Dose (mmol/kg)
Analgesic activitya (%)
Dose (mmol/kg)
Analgesic activityb (%)
7a
9a
0.04
81.0
32.1
36.4
69.5
57.8
55.5
25.3
0
0.04
0.01c
0.01c
0.01c
0.04
0.04
0.04
0.04
0.04
0.04
0.04
0.04
0.04
0.04
0.04
0.02c
64.2
35.7
0.005c
0.005c
0.005c
0.02c
0.04
9b
9c
Me(CH2)2–
Me(CH2)4–
Me(CH2)7–
36.4
ꢀ13.4
62.9
79.6
9d
9e
Me(CH2)10
Me(CH2)14
Me(CH2)16
Ph–
–
9f
9g
–
0.04
ꢀ10.3
ꢀ10.3
117.0
26.4
–
0.04
9h
0.02c
0.04
44.5
70.9
75.2
36.4
39.0
79.7
88.9
51.9
10a
10b
10c
10d
10e
14
Me(CH2)2–
Me(CH2)4–
Me(CH2)7–
0.04
0.04
ꢀ17.9
4.6
Me(CH2)10
–
0.04
ꢀ5.2
42.8
Ph–
OH
Cl
0.02c
0.04
91.8
98.4
15
0.02c
a % Inhibition = 100 ꢀ (A/B · 100), where A = incidence of writhing in the treated group and B = incidence of writhing in the control group,
occurring from the 5th to 10th min after administration of the noxious agents.
b % Elevation of pain threshold = 100 · (A ꢀ B)/B, where A = latency time of treated group and B = latency time of control group. The time interval
from placing animals on the surface of the hot plate to a licking of the hind paws or jumping was defined as hot plate latency.
c Toxic at higher dose.
9 showed higher analgesic activity (9c: n = 4, 69.5%,
dose 0.005 mmol/kg in writhing test; 9d: n = 7, 57.8%,
dose 0.02 mmol/kg in writhing test; 62.9%, dose
0.04 mmol/kg in hot plate; 9e: n = 10, 55.5% in writhing
test and 79.6% in hot plate, dose 0.04 mmol/kg). These
results demonstrate that substituents with suitable
length will maintain definite analgesic effects. Similarly,
when n = 4, compound in series 10 exhibited the definite
analgesic activity (10b: n = 4, 75.2%, writhing test).
analgesic activity as well. Surprisingly, compound 14
showed excellent analgesic activity in both tests (88.9%
in writhing test, 91.8% in hot plate, dose 0.04 mmol/
kg), which encouraged us to further synthesize the chlo-
ride derivative 15. Differing from compound 14, the
analgesic activity of compound 15 was notably different
in both tests. Its hot plate test gave significant analgesic
activity (98.4%, dose 0.02 mmol/kg in hot plate test),
whereas writhing test only showed moderate analgesic
activity (51.9%, dose 0.04 mmol/kg). Therefore, intro-
duction of hydroxyl or chloride on the link between
two spirocyclopiperiziniums may benefit the analgesic
activity.
It is also found from Table 1 that the carbamate ana-
logues 10 displayed lower toxicity than carboxylic ester
analogues 9. Meantime, carbamates 10 only showed
remarkable analgesic activity in writhing test. These re-
sults suggest that introduction of carbamoyl groups may
be favorable to decrease the toxicity and enhance the
selectivity.
To explore whether these two series compounds were
hydrolyzed by esterase and released the parent com-
pound 14 after subcutaneous injection, we selected com-
pounds 9a, 9e, 9g, and 10d to study their hydrolytic
ability in buffer, 10% mice blood, and 10% mice brain
homogenate using reported method.18 TLC was used
to monitor the result. It was found that all tested com-
pounds were stable within 3 h whether in buffer or in
biological matrices. Therefore, these compounds may
act as their original forms. This result can explain the
reason that only derivatives with medium length of side
chain had higher activity, and coincides with our previ-
ous conclusion too.12
Compounds 9h (R = Ph) and 10e (R = Ph) were de-
signed to investigate the influence of aromatic group
on the pharmacological activity. The results showed that
the compounds 9h (117.0%, dose 0.04 mmol/kg in hot
plate) and 10e (R = Ph, 79.7%, dose 0.02 mmol/kg in
writhing test) exhibited the highest analgesic activity in
corresponding analogues. These results give us an inspi-
ration that the introduction of aryl group may be an
available route for the modification of this kind of
compounds.
In summary, two series of dispirocyclopiperazinium
salts were synthesized and evaluated for their in vivo
analgesic activity in both acetic acid writhing test and
Compound 14 is the key intermediate for the synthesis
of compounds 9 and 10. In comparison, we tested its

A. Li et al. / Bioorg. Med. Chem. Lett. 17 (2007) 5078–5081
5081
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played lower toxicity than carboxylic ester analogues;
(4) both hydroxylation and chlorination of lead com-
pound could increase the analgesic activity. These re-
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Acknowledgment
This research was supported by the fund of National
Science Foundation of China (NSFC 20372006).
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