New PDE4 inhibitors based on pharmacophoric similarity between papaverine and tofisopam

Article abstract of DOI:10.1016/j.bmcl.2011.08.036

Pharmacophoric comparison between papaverine and tofisopam led to identify three new series of microto sub-micromolar inhibitors of phosphodiesterase-4, including 7,8-dialkoxy-2,3-benzodiazepin-4-one derivatives, 7,8-dialkoxy-1,4- benzodiazepin-2-one derivatives, and dialkoxybenzophenone derivatives.

Full text of DOI:10.1016/j.bmcl.2011.08.036

Bioorganic & Medicinal Chemistry Letters 21 (2011) 6567–6572
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
New PDE4 inhibitors based on pharmacophoric similarity
between papaverine and tofisopam
Frédéric J.J. Bihel a, , Hélène Justiniano b, Martine Schmitt a, Malik Hellal a, Mohamed A. Ibrahim a,
⇑
Claire Lugnier b, Jean-Jacques Bourguignon a
a Laboratoire d’Innovation Thérapeutique, UMR 7200, CNRS, Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, 67400 Illkirch Graffenstaden, France
b Biophotonique et Pharmacologie, UMR 7213, CNRS, Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, 67400 Illkirch Graffenstaden, France
a r t i c l e i n f o
a b s t r a c t
Article history:
Pharmacophoric comparison between papaverine and tofisopam led to identify three new series of micro-
to sub-micromolar inhibitors of phosphodiesterase-4, including 7,8-dialkoxy-2,3-benzodiazepin-4-one
derivatives, 7,8-dialkoxy-1,4-benzodiazepin-2-one derivatives, and dialkoxybenzophenone derivatives.
Ó 2011 Elsevier Ltd. All rights reserved.
Received 26 May 2011
Revised 4 August 2011
Accepted 6 August 2011
Available online 12 August 2011
Keywords:
Benzodiazepine
Benzophenone
Phosphodiesterase
Cyclic nucleotide phosphodiesterases (PDEs) represent an
important class of enzymes for the cellular regulation, by hydrolyz-
ing the intracellular cyclic AMP (cAMP) and cyclic GMP (cGMP).
Among all the PDE families,1 increasing interest has been focused
on cAMP-specific PDE4 family, which appears as a potential target
for the development of new anti-asthmatic and anti-inflammatory
drugs.2,3
PDE inhibitors are generally hydrophobic compounds with rel-
atively large chemical diversity. Among them, the alkaloid papav-
erine 1a is presenting micromolar IC50 towards PDE4,4,5 while its
diethoxy-derivative ethaverine 1b is exhibiting a sub-micromolar
activity (Table 1).6 In terms of pharmacophoric fragments, papav-
erine 1a presents a H-bond acceptor system (imine nitrogen)
flanked by two dimethoxy phenyl rings. A drug repositioning strat-
egy allowed recently identifying tofisopam 2, clinically used as
anxiolytic drug, as a sub-micromolar PDE4 inhibitor.7 While
belonging to the class of 2,3-benzodiazepines, tofisopam exhibits
common pharmacophoric pattern with papaverine.
both tofisopam-related analogs and 2,3-benzodiazepin-4-one
derivatives I (Scheme 1).8 The commercially available 3,4-dialk-
oxy-phenylacetic acids were esterified in presence of methanol
upon acidic catalysis, before to be regioselectively acylated at the
6-position of the phenyl ring, in presence of tin(II) chloride. The
cyclization was performed with hydrazine hydrate upon acidic
catalysis, leading to 2,3-homophthalazin-4-ones 4. A treatment of
4 with sodium hydride in presence of methyl iodide led to deriva-
tives 5. However, the amide function of 4 could also be activated
into imino-chloride, which could further be methylated to yield
tofisopam-related analogs 6a–c.
Next, we investigated the hypothesis to remove the alkoxy
groups at the 7- and 8-positions of the 2,3-benzodiazepin-4-one
scaffold I, while keeping them at the 3- and 4-positions of the
free-rotating ring. In the absence of alkoxy groups at the 7- and
8-positions of benzodiazepinone, another synthetic pathway has
been developed (Scheme 2). Commercially available 2-iodobenzoic
acid was activated into the corresponding aroyl chloride, before to
be involved in a Friedel–Crafts reaction in presence of dialkoxyben-
zene. Then, a copper-catalyzed reaction was performed on the
resulting 2-iodobenzophenone 8, leading to the substitution of
the iodine atom by a diethylmalonate moiety.9
Based on the hypothesis of a common pharmacophoric pattern
between papaverine and tofisopam, this work describes the SAR
analysis of three novel series, 7,8-dialkoxy-2,3-benzodiazepinones
(series I), 7,8-dialkoxy-1,4-benzodiazepinones (series II) and 4,5-
dialkoxybenzophenones (series III) as phosphodiesterase-4 inhibi-
tors (Fig. 1).
After alkaline hydrolysis and in situ decarboxylation, the 2,3-
benzodiazepin-4-ones 9 were obtained by condensation with
hydrazine hydrate. N-methylation of the amide function was even-
tually performed with methyliodide to give the compounds 10.
We next considered 7,8-dialkoxy-1,4-benzodiazepin-2-ones II
We first initiated the synthesis of 7,8-dialkoxy-2,3-benzodiaze-
pine compounds, using a convenient synthetic pathway leading to
as valuable carba-azaisosteres of 2,3-benzodiazepin-4-ones
(Fig. 1). From a synthetic point of view, we first synthesized the
I
⇑
Corresponding author.
E-mail address: fbihel@unistra.fr (F.J.J. Bihel).
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.08.036

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F. J. J. Bihel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6567–6572
1,4-benzodiazepin-2-one, using either pallado-catalyzed cross-
Table 1
PDE4 inhibition of reference compounds
coupling reactions (Suzuki–Miyaura reaction (14a–f), Sonogashira
reaction (14h–i)), or amination reaction (14g). Eventually, we syn-
thesized a series of benzophenones 15a–j (series III), as seco deriv-
atives of benzodiazepinones following classical Friedel–Crafts
reaction (Scheme 4). Amination step was performed on 4-bromo-
benzophenone derivatives using Buchwald cross coupling reac-
tion.12 All the synthesized compounds were tested as PDE4
inhibitors following standard procedure.13 IC50 values are summa-
rized in Tables 2–5.
RO
MeO
MeO
N
N
RO
N
The tofisopam-analog girisopam 6a,14 another well-known anx-
iolytic agent, appeared to be five-fold less active on PDE4 than its
parent drug 2. tofisopam and girisopam exhibit two main struc-
tural differences: the first one is the presence of two H-bond accep-
tor methoxy groups in 3- and 4-position of the exocycle for
tofisopam, in comparison with a withdrawing chlorine atom at po-
sition 3 for girisopam; the second one is the presence of a small
hydrophobic ethyl group at 5-position of the tofisopam seven-
member ring. Based on our working hypothesis dealing with phar-
macophoric pattern analogy between papaverine 1a and tofisopam
2, the ethaverine-diethoxy groups were introduced into girisopam
analogs 6b and 6c. In spite of the presence of a chlorine atom on
the exocycle, affinities of both compounds for PDE4 exhibited a
gain of one order of magnitude in comparison with girisopam, with
OR
1a - R = Me
1b - R = Et
2
OMe
OR
MeO
Compound
Name
IC50 (lM)
1a
1b
2
Papaverine
Ethaverine
Tofisopam
1.1
0.4
0.68
1,3-benzoxazine-2,4-diones 11 following standard procedures
(Scheme 3).10 The 1,4-benzodiazepin-2-ones 12 were obtained by
condensation with methyl glycinate, then the benzamide function
was activated into imino-chloride 13 using POCl3 in presence of
dimethylaniline.11 Starting from compounds 13, we were able
to introduce a diversity of decorations at the 5-position of the
IC50 values of 0.4 and 0.2 lM, respectively. This result validates our
working hypothesis dealing with pharmacophoric pattern analogy
between papaverine and tofisopam.
2
R
O
O
N
RO
RO
RO
2
N
R
O
N
RO
RO
N
RO
Ar
Ar
Ar
I
II
III
Figure 1. Novel classes of PDE4 inhibitor.
2
2
R
R
2
R
O
RO
RO
RO
RO
RO
CO Me
2
CO H
2
a, b
c
NH
O
N
RO
3
4
1
1
R
R
e, f
d
2
R
2
R
O
RO
RO
RO
N
N
N
N
RO
1
6
R
5
1
R
Scheme 1. Reagents and conditions: (a) H2SO4(cat), MeOH, reflux; (b) R1-COCl, SnCl2, rt; (c) NH2–NH2ÁH2O, EtOH, 150 °C, 3 h, then AcOH, EtOH, reflux, 30 min; (d) MeI, NaH,
THF; (e) POCl3, dimethylaniline, CHCl3, 120 °C; (f) MeLi, THF, À78 °C.

F. J. J. Bihel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6567–6572
6569
CO2Et
I
I
CO2Et
O
a, b
c
O
CO2H
1
R
7
8
1
R
d, e
O
O
f
N
NH
N
N
1
R1
R
10
9
Scheme 2. Reagents and conditions: (a) SOCl2, DMF, 60 °C; (b) AlCl3, DCE, (EtO)2benzene; (c) CH(CO2Et)2, CuI, picolinic acid, Cs2CO3, dioxane, 90 °C; (d) LiOH, MeOH, H2O,
reflux; (e) NH2–NH2ÁH2O, EtOH, 150 °C, 3 h, then AcOH, EtOH, reflux, 30 min; (f) MeI, NaH, THF.
We next investigated the replacement of the girisopam methyl-
imine moiety by an amide group, leading to compound 5c, and an
O
IC50 of 7.7
l
M (Table 3). As observed with 6c, a chlorine atom at 4-
M.
N
R
R
N
O
O
R
R
position (5d) appeared to be more active with an IC50 of 1.5
l
a
The substitution of the phenyl group by a 2-benzothienyl ring
led to 5g with a micromolar affinity. Then the N-methylation of
the amide function led to 5e with a 10-fold increase in affinity
O
NH
(IC50 = 0.3 lM). The same series of compound was synthesized in
O
11
12
the diethoxy-series leading to compounds 5i–m. All of them ap-
peared to be more potent towards PDE4 than their corresponding
dimethoxy analogs 5a–h. Especially, 5m was found the most po-
b
tent inhibitor of PDE4 with an IC50 of 0.09
better than tofisopam.
lM, about seven-fold
O
O
N
1
N
R
R
R
R
So far, we keep as a constant the presence of alkoxy groups at 7-
and 8-positions of the bicycle. However, tofisopam and papaverine
exhibit the same alkoxy moieties at the 3- and 4-positions of the
free-rotating ring. So, we investigated the hypothesis to remove
the alkoxy groups at the 7- and 8-positions of the 2,3-benzodiaze-
pin-4-one scaffold, while keeping them at the 3- and 4-positions of
the exocycle. Diethoxy analog 9 appeared to be 4-fold more potent
than 5i, which carries the diethoxy moieties at 7- and 8-positions.
As previously observed, N-methylation of the amide led to a better
c
N
N
R
Cl
14
13
Scheme 3. Reagents and conditions: (a) Methyl glycinate hydrochloride, pyridine,
reflux, 6 h, then add AcOH, 130 °C, 12 h; (b) POCl3, dimethylaniline, CHCl3, 120 °C;
(c) 14a–f: R1-B(OH)2, K3PO4, Pd(PPh3)4, DMF, 100 °C, 12 h; 14g: piperidine, EtOH,
110°, 48 h; 14h: phenylacetylene, PdCl2(PPh3)2, CuI, Et3N, PPh3, 55 °C, 3 h; 14i: 14h,
Pd/C, MeOH, DCM, H2 (70 psi), 48 h.
affinity (10, IC50 = 0.14 lM). This last result seems to show that
among the two pairs of dialkoxy groups exhibited by papaverine
O
O
1
R O
R1O
Cl
a
R2
R2
1
R1O
R O
16a-e
O
O
R1O
1
R O
b
3
R
R1O
N
1
R O
Br
16f-j
2
R
Scheme 4. Reagents and conditions: (a) AlCl3, nitrobenzene, reflux, 12 h; (b) Pd(OAc)2—5%, BINAP—6%, Cs2CO3, dioxane, 90 °C, 7 h.

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F. J. J. Bihel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6567–6572
Table 4
Table 2
PDE4 inhibition of 1,4-benzodiazepin-2-ones
PDE4 inhibition of tofisopam-related derivatives
2
R2
R
O
N
R
R
R
N
N
R
N
1
R
Compound
R
R1
R2
IC50 (lM)
13a
14a
14b
14c
14b
14d
14e
14h
14f
OMe
OMe
OMe
OMe
OEt
Ph
Ph
3-Cl-Ph
4-Cl-Ph
Ph
H
>20
18
R1
Me
Me
Me
H
Me
Me
Me
Me
Me
Me
>20
>20
0.95
0.86
2.3
1.6
8.4
>20
>20
Compound
R
R1
R2
IC50 (lM)
Tofisopam
6a (Girisopam)
6b
6c
OMe
OMe
OEt
3,4-MeO
3-Cl
Et
H
Et
Et
0.68
3.2
0.39
0.21
OEt
Ph
OMe
OMe
OMe
OMe
OMe
2-Benzothienyl
Ph-C„C-
Ph-CH@CH-
Ph-(CH2)2-
3-Cl
4-Cl
OEt
14i
14g
1-piperidinyl
Table 3
PDE4 inhibition of 2,3-benzodiazepin-4-ones
O
Table 5
R
PDE4 inhibition of benzophenone derivatives
Et O
N
R
2
N
R
O
Et O
R
1
Compound
R
R1
R2
IC50 (lM)
1
R
15
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
9
10
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OEt
OEt
OEt
OEt
OEt
H
Ph
Ph
3-Cl-Ph
4-Cl-Ph
3,4-Cl2-Ph
2-Naphtyl
2-Benzothienyl
2-Benzothienyl
Ph
3-Cl-Ph
4-Cl-Ph
2-Benzothienyl
2-Benzothienyl
3,4-EtO2-Ph
3,4-EtO2-Ph
H
Me
H
H
H
H
H
Me
H
H
H
H
Me
H
Me
7.70
5.1
7.7
1.5
Compound
R
R1
IC50 (lM)
1.64
1.50
1.20
0.30
0.80
1.4
0.8
0.26
0.087
0.19
0.14
15
OEt
OEt
OEt
OEt
OEt
OEt
OMe
OEt
OEt
OEt
OEt
—
2.9
1.4
>20
7.9
4.3
5.0
9.6
3.6
17.0
2.3
2.2
15a
15b
15c
15d
15e
15f
15g
15h
15i
15j
2-Cl-Ph
4-Cl-Ph
3,4-Cl2-Ph
1-Naphtyl
2-Naphtyl
1-Piperidino-
1-Piperidino-
4-Ph-piperazin-1-yl-
4-Bn-piperazin-1-yl-
N–Me–N–Bn-4-amino
H
and tofisopam, the one carried by the free-rotating ring may be
more important for affinity.
of seco benzodiazepinones derivatives (Table 5). Most of the benz-
ophenones exhibited a range of inhibition between 1 and 10 M,
slightly less active than the reference compound papaverine. The
poor inhibition observed with compound 15b appeared to be very
interesting in the way where it is the seco-derivative of girisopam-
l
We next considered 7,8-dialkoxy-1,4-benzodiazepin-2-ones II as
valuable carba-azaisosteres of 2,3-benzodiazepin-4-ones I (Fig. 1).
Structurally-related diazepinoindoles CI-1018 have already been re-
ported by Burnouf et al. as potent PDE4 inhibitors, but this com-
pound does not exhibit any of the alkoxy groups, characteristic of
thepapaverineortofisopamderivatives.15 Globally, 1,4-benzodiaze-
pin-2-one derivatives appeared to be less potent on PDE4 than their
corresponding 2,3-benzodiazepin-4-one isosteres (Table 4). For
related 6c (IC50 = 0.21 lM). One hypothesis to explain this drastic
loss of inhibition may rely on the position of the diethoxy substit-
uents. We previously observed that affinity is increased with the
diethoxy substituents on the free rotating aromatic ring (9). Fol-
lowing this hypothesis applied to benzophenone would localize
the chlorine atom of 15b at the 7-position of girisopam-related
6c, resulting in a drastic loss of affinity. This hypothesis could then
explain the good affinity of 15a, in which the chlorine atom at 2-
position may induce some specific features (geometric, lipophilic)
mimicking the seven-member ring closure of tofisopam. This last
result allowed us to definitively link the different series I, II and
III through a common pharmacophoric pattern represented in
Figure 2. This pharmacophoric pattern can also be observed into
example, the 2-benzothienyl derivative 14e (IC50 = 2.3
found 10-fold less potent than its isostere 5h (IC50 = 0.30
l
M) was
l
M). How-
ever, the diethoxy substituents led again to a better affinity than
their corresponding dimethoxy substituted derivatives.
The results obtained in both series of benzodiazepinones I and
II, along with tofisopam and papaverine, highlight the hypothesis
of a common pharmacophoric pattern resumed into the dialkoxy-
benzophenonimine scaffold. In order to validate this pharmaco-
phoric hypothesis, we considered benzophenones III as a series

F. J. J. Bihel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6567–6572
6571
O
EtO
EtO
EtO
NH
N
N
EtO
NH
O
5i
9
IC50 = 0.8 µM
I
C50 = 0.19 µM
O
H
N
EtO
EtO
EtO
O
N
EtO
Cl
15a
14b
IC50 = 1.4 µM
IC50 = 0.95 µM
Figure 2. Common pharmacological pattern between series I, II and III.
the 4-(2,3-dimethoxyphenyl)-phthalazin-1-one derivatives, de-
scribed by Van der Mey et al. as submicromolar PDE4 inhibitors.16
One of the main negative properties of benzophenone scaffold is
its lack of aqueous solubility, so we tried to circumvent this prob-
lem by introducing some water-solubilizing groups, based on a
substituted nitrogen atom at para-position. The electronic doublet
of nitrogen has the capability to be a hydrogen bond acceptor, as
well as an alkoxy group. Indeed, compounds 15i–j were found as
Inhibitory activities of these new PDE4 inhibitors were
screened on several phosphodiesterases (Table 6). While tofiso-
pam 2 exhibits submicromolar to micromolar inhibition towards
PDE2, PDE3 and PDE4, 2,3-benzodiazepin-4-ones 5i and 9, but
also benzophenone 15a appeared to be more selective towards
PDE4 in comparison with other PDE (IC50 >10
lM). Moreover,
kinetic experiments performed with compounds
9
and 15a
confirmed a mechanism by competitive inhibition for both kinds
of inhibitors (Fig. 3)
micromolar inhibitors of PDE4, with IC50 about 2 lM.
Table 6
Inhibitory activities towards several phosphodiesterases
Compound
IC50 (lM)
PDE1
PDE2
PDE3
PDE4
PDE5
a
Tofisopam 2
Girisopam 6a
6b
6c
5i
nd
nd
nd
nd
37%
0.90
4.00
44%
5.90
22%
0.68
3.20
0.39
0.21
0.80
0.19
1.40
36%
12.5
nd
39%
30%
a
a
nd
4%
a
a
a
38%
a
a
a
a
a
5%
30%
12%
24%
a
a
a
a
9
15a
3%
21%
27%
46%
17%
a
a
a
4%
a
Percentage of inhibition at 10 lM; nd: not determined.
Figure 3. Lineweaver–Burk plots of compounds 10 and 15a inhibitory effects on purified PDE4 activity (using 0.25–4
lM cAMP as substrate). Apparent Km value for
PDE4 = 0.71 M. (a) 10 Ki = 0.066
l
l
M (d: 0 M; j: 3.10À8 M; N: 1.10À7 M); (b) 15a Ki = 0.42 M (d: 0 M; j: 1.10À6 M;N: 3.10À6 M).
l

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F. J. J. Bihel et al. / Bioorg. Med. Chem. Lett. 21 (2011) 6567–6572
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tern common to both PDE4 inhibitors papaverine and tofisopam.
This result led us to design three novel series of potent and selec-
tive PDE4 inhibitors exhibiting micromolar to sub-micromolar IC50
values.
Acknowledgments
JChem for Excel was used for structure database management,
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