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T. Assaad et al. / Bioorg. Med. Chem. Lett. 16 (2006) 2654–2657
Table 1. IC50 values (nM) at the human VAChT compared to the literature data
b
b
Compound
R
Series 6a
Series 60
Series 7a
Series 70
a
b
c
d
e
f
H
CH2C6H5
ꢁ100
5000 ( 700)
83 ( 7)
>100
>100
2900 ( 400)
30 ( 7)
22 ( 6)
21 ( 8)
25 ( 10)
—
57 ( 26)
11 ( 4)
66 ( 24)
>100
CH2–o-Br–C6H4
CH2–m-Br–C6H4
CH2–p-Br–C6H4
CH2CH3
55 ( 8)
—
310 ( 130)
47 ( 18)
46 ( 20)
>100
>1000
>100
>100
—
—
—
—
—
—
>1000
>100
>100
g
h
i
CH2CH@CH2
CH2C(CH3)@CH2
(E)-CH2–CH@CHCH3
CH2C„CH
—
—
ꢁ100
>100
>100
>100
—
—
j
>1000
>100
k
(E)-CH2–CH@CHI
—
a Values are means of three experiments, standard deviation is given in parentheses. In the same conditions, (ꢀ)-vesamicol showed an IC50
=
20 ( 2.3) nM.
b Literature data obtained on VAChT isolated from the electric organ of Torpedo californica,17 ( )-vesamicol showed a Kd = 34 ( 6) nM.
natant rich in synaptic-like microvesicles containing
human VAChT. Competition was allowed to proceed
for 24 h to ensure equilibration of low concentrations
of target compounds. It could not be carried out at
37 °C due to the length of incubation. However, binding
of vesamicol is not strongly temperature dependent.
seems not to influence the binding affinity. These results
contrast with those obtained previously for trozamicol
and prezamicol congeners.17,18 For these compounds,
the 1,4 orientation of the nitrogen relative to the hydroxyl
group (trozamicol series) was found to be preferred.
Regarding the nature of the R group, substitution of
compounds 6a and 7a with alkyl, alkenyl, and alkynyl
groups was based on results obtained for hydroxylated
decahydroquinoline vesamicol derivatives.18 In this
study, a (E)-iodoprop-2-enyl group was successfully
introduced to obtain a compound with nanomolar
VAChT affinity. Contrary to these results, the N-alkyl-
ation of compounds 6a or 7a failed to obtain com-
pounds with VAChT potency. Only some of the
benzyl derivatives displayed IC50 less than 100 nM
(6b–d and 7c–d). Moreover, the position of the bromide
atom on the benzyl group induces variability in the
VAChT affinity as the para-substituted derivatives
(6e and 7e) showed low VAChT affinities. Here also,
these results contrast with those obtained for benzyl-
substituted prezamicol or trozamicol derivatives as the
biological activity was not influenced by the position
of the substituent on the ring.17
Because IBVM and MIBT are compounds with high
affinities for the VAChT; IC50 values have only been
determined for potent compounds with IC50 lower than
100 nM. Affinities of synthesized compounds are pre-
sented in Table 1.
The only structural difference between prezamicol ana-
logs (60a–e),17 and compounds 6a–e, trozamicol analogs
(70a–e)17 and compounds 7a–e is the replacement of the
4-phenylpiperidine group for a N-phenylpiperazine
group. Comparison of their in vitro affinities would pro-
vide information on the impact of an additional nitro-
gen at this part of the molecule. Even compounds 6a
(IC50 ꢁ 100 nM) and 7a (IC50 > 100 nM) displayed
higher affinities for the VAChT compared to prezamicol
(60a;
IC50 = 5000 nM)
and
trozamicol
(70a;
IC50 = 2900 nM),17 compounds 6b,6c, and 6e
(IC50 = 57, 11, and >100 nM, respectively) exhibited
similar affinities for the VAChT compared to their pre-
zamicol analogs 60b,60c, and 60e (IC50 = 83, 55, and
310 nM, respectively).17 Moreover, even compounds 7c
and 7d (IC50 = 47 and 46 nM, respectively) showed
VAChT affinities in the same range of magnitude as
their trozamicol analogs 70c and 70d (IC50 = 22 and
21 nM, respectively), while compounds 7b and 7e
(IC50 = >100 nM) are less potent compared to com-
pounds 70b and 70e (IC50 = 30 and 25 nM, respectively).
Because the substitution of the 4-phenylpiperidine group
for N-phenylpiperazine generated compounds with or
without improved affinities for the VAChT, an addition-
al nitrogen at this part of the molecule cannot be consid-
ered as essential for the binding to the VAChT.
Although it is well known that interaction at the
VAChT binding site is stereoselective,9 only racemic
mixtures have been screened. We can expect that enan-
tiomeric separation of compounds 6b–d and 7c–d would
provide compounds with improved VAChT affinity. In
parallel, as compound 6c is the most potent of this ser-
ies, its radiolabeling with bromide-76 but also with
iodine-125/123 is underway to further evaluate its in vivo
pharmacological profile.
Acknowledgments
This work was supported by Grant (82/2001 and 582/
2003) In Aid scientific research from the AECS (Atomic
Energy Commission of Syria). This work was partially
supported by DEST and the French Embassy in Austra-
lia under the FAST program FR040051. This study was
funded in part by the EC-FP6-project DiMI, LSHB-CT-
2005-512146. We thank Jeffrey D. Erickson of the
Louisiana State University Health Sciences Center,
Except for the benzyl group for which the VAChT affinity
could be considered significantly different for compounds
6b and 7b, the VAChT affinity in series 6 and 7 is in the
same range for each R substituent. In these series of com-
pounds, the orientation of the nitrogen in the piperidine
ring relative to the hydroxyl group (in position 3 or 4)