PDE5 inhibitors: An original access to novel potent arylated analogues of tadalafil

Article abstract of DOI:10.1016/j.bmcl.2006.10.069

A method to access totally new analogues of tadalafil was explored. The Buchwald reaction was adapted and used to replace the methyl group of tadalafil by various aryl groups. Inhibition potencies on PDE5 of these analogues were determined and proved to be comparable to the one of tadalafil. Using the same route, compounds with the same level of activity but improved water solubility were produced by introducing a pyridine or a pyrimidine ring. This original route also opens access to new unpatented compounds.

Full text of DOI:10.1016/j.bmcl.2006.10.069

Bioorganic & Medicinal Chemistry Letters 17 (2007) 789–792
PDE5 inhibitors: An original access to novel potent
arylated analogues of tadalafil
Terence Beghyn,* Candide Hounsou and Benoit P. Deprez
Inserm U761 ‘Biostructures and Drug Discovery’ Lille, F-59006, France
Universite´ de Lille2, Faculte´ de Pharmacie, Institut Pasteur de Lille, France
Received 14 September 2006; revised 23 October 2006; accepted 24 October 2006
Available online 27 October 2006
Abstract—A method to access totally new analogues of tadalafil was explored. The Buchwald reaction was adapted and used to
replace the methyl group of tadalafil by various aryl groups. Inhibition potencies on PDE5 of these analogues were determined
and proved to be comparable to the one of tadalafil. Using the same route, compounds with the same level of activity but improved
water solubility were produced by introducing a pyridine or a pyrimidine ring. This original route also opens access to new
unpatented compounds.
Ó 2006 Elsevier Ltd. All rights reserved.
Phosphodiesterases (PDEs) affect the cellular levels of
the cyclic nucleotides cAMP and cGMP, which are in-
volved in many physiological processes such as immuni-
ty, cardiac- and smooth-muscle contraction, apoptosis,
ion-channel conductance and growth control. Inhibiting
these enzymes is therefore an attractive strategy in the
development of smooth-muscle relaxants and drugs to
treat inflammatory diseases, asthma, depression and
many other diseases.1 The cGMP-specific PDE5 is the
major PDE isozyme in the corpus cavernosum, and con-
trols penile erection. PDE5 inhibitors used in the clinic
amplify the NO-cCMP pathway and enhance the nor-
mal process leading to penile erection. The launch of
ViagraTM (sildenafil, 1) (Figure 1)2 as the first oral treat-
ment for male erectile dysfunction revolutionized the
treatment of this condition. Today, LevitraTM (vardena-
fil, 2), a closely related structural analogue of sildenafil,
and CialisTM (tadalafil, 3), a tetrahydro-b-carboline
derivative,3,4 are also available (Fig. 1).
O
N
O
N
O
H
N
N
O
HN
O
HN
2
N
N
12
N
12a N
6
O
N
H
O
S
N
O
O
S
N
O
H
O
O
N
N
Vardenafil, 2
Figure 1. Structure of sildenafil, vardenafil and tadalafil.
Sildenafil, 1
Tadalafil, 3
launched, multiple analogues were synthesized in order
to optimize the selectivity profile against these PDEs.5
The N-methyl group of tadalafil is directed toward a
large unoccupied pocket of the active site of PDE5.
Our objective was to enlarge the structural diversity of
this tadalafil-derived family through the replacement
of the N2-methyl group by novel aryl substituants,
and to evaluate the compatibility of these modifications
with PDE5 inhibitory activity.
These three PDE5 inhibitors exhibit different PDE selec-
tivity profiles, in particular in their relative affinities for
PDE5, PDE6 (found in photoreceptor cells) and PDE11
(of yet unclear biological function). Since tadalafil was
Although the N2-phenyl derivative 6a (Scheme 1) dis-
plays a significant inhibitory activity against PDE5,6
no other arylated analogues of tadalafil were reported.
This important gap in the structural space surrounding
tadalafil can be explained by synthetic difficulties. In
the only synthetic route to tadalafil analogs described
Keywords: PDE5; Tadalafil; Tetrahydrobetacarboline; N-arylation;
Cuprous iodide; Epimerisation.
*
Corresponding author. Address: 3, rue du Professeur Laguesse 59000
Lille, France. Tel.: +33 3 20 96 40 36; fax: +33 3 20 96 47 09; e-mail:
terence.beghyn@univ-lille2.fr
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.10.069

790
T. Beghyn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 789–792
O
H
O
O
H
O
O
H
N
Cl
2
N
H
N
O
N
O
H2N
N
O
O
O
O
+
N
H
N
H
N
H
H
H
H
O
O
O
4
Scheme 1. Reagents and conditions: (i) MeOH reflux, NEt3, 10 days, <5% yield.
6a
5
so far, the terminal step is the formation of the diketo-
piperazine ring from an N-substituted glycyltetrahydro-
carboline generated in situ (Scheme 1). This cyclisation
is very efficient when N2 is not substituted or alkylated,
whereas N-phenylglycyltetrahydrocarbolin 5 is only very
slowly converted to the desired compound 6a (only 5%
yield after 10 days of reaction).
K2CO3 or Cs2CO3. The nature of the base is important
and the best results are described with K3PO4. Reac-
tions are best performed in toluene, DMF, THF or
dioxane, at temperatures ranging from 25 to 110 °C.
In such conditions, lactams are arylated in good
yields.7
These conditions were explored using iodobenzene as a
model arylating reagent. It is known that only the cis
isomers significantly inhibit PDE54 in this family. There-
fore, a particular attention has been paid to the config-
uration of the tetrahydrocarboline system. Indeed,
upon heating in basic conditions, one can expect the for-
mation of the thermodynamically favored trans isomer
by epimerization of one of the chiral centers of Nortad-
alafil 7a, leading to compound 6b or 6c represented in
Scheme 3. The conditions described by Buchwald gave
in quantitative yield a product arylated exclusively at
the targeted N2 position, leaving untouched the indolic
nitrogen, described as potentially reactive.8 However,
NMR analyses revealed that the product isolated was
in the trans configuration. Measurement of optical rota-
tion showed that the product formed was 6c and not 6b
(Scheme 3).
To fill the diversity gap in this family, we propose to
introduce the N2 substituent at the last step of the syn-
thesis, using the mild conditions described by Buchwald
et al. for the arylation of amides and lactams.7 We
would thus close the diketopiperazine ring to form Nor-
tadalafil 7a in high yield, and introduce the aromatic
substituent afterward, tapping in a large array of com-
mercially available aryl or heteroaryl halides. Our start-
ing material Nortadalafil 7a (in the 6(R), 12a(R) cis
configuration) was easily prepared in two steps from
D-tryptophane methyl ester, piperonal, chloroacetyl-
chloride and ammonia (Scheme 2).
The procedures described by Buchwald et al. for the
arylation of amides, carbamates and lactams typically
use 0.2–10 mol% of cuprous iodide, 5–20 mol% of a
diamine ligand and a mineral base such as K3PO4,
In order to avoid the deleterious isomerisation, we low-
ered the temperature and the amount of base while using
a higher stoichiometry of catalyst. The reaction mixtures
were analyzed by NMR after simple workup in ammo-
O
H
O
H
O
O
O
O
NH+3 Cl
NH+2
Cl
O
i
+
N
N
H
H
H
O
O
H
H
O
H
N
N
O
12a N
Arylation site
12a
I
N
O
6
O
6
O
H
O
H
O
H
N
H
H
N
H
O
N
H
i
2
Cl
N
O
N
O
O
ii
iii
O
O
O
O
N
H
N
H
H
Nortadalafil 7a Cis 6(R), 12a(R)
6a Cis 6(R), 12a(R) :
6b Trans 6(S), 12a(R) :
0%
0%
H
O
O
6c Trans 6(R), 12a(S) : 100%
4
7a Cis 6(R), 12a(R)
Scheme 3. Arylation of Nortadalafil. Reagent and conditions: Nor-
tadalafil (1 equiv, 133 mM), CuI 10 mol%, ( )-trans-1,2-diaminocy-
clohexane 20 mol%, iodobenzene (1 equiv), K3PO4 (3 equiv), dioxane,
at 110 °C.
Scheme 2. Reagents and conditions: (i) isopropanol reflux, 92%; (ii)
chloroacetylchloride, chloroform, triethylamine, ꢀ10 °C, 85%; (iii)
NH3, methanol, 45 °C, 70%.

T. Beghyn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 789–792
791
nia, removing the base and the copper complexes.
cis:trans ratios displayed in Table 1 were evaluated
directly from NMR spectra.
Decreasing temperature allowed a reduction of epimer-
ization but still half of the final product was epimerized
leading to a 50:50 mixture of cis:trans products. Even if
long time reaction were required, a further decrease of
the temperature to 14 °C (solidification of dioxane)
proved to be a good compromise to prepare arylated
analogs of tadalafil in the cis configuration.
Table 1. Reaction conditions
Com- Temp. Catalyst
pound (°C)
equiva
% Reaction 6a (cis)/
Conversion time 6c(trans)
1
2
3
110
28
0.1
1.6
2.0
100
90
3 h
5 days
15 days
0/100
50/50
95/5
Conditions (3)9 from Table 1 were used to prepare a set
of 11 analogues of tadalafil. Arylated compounds were
isolated in moderate to high yields (35–80%), after thick
layer chromatography (Table 2).
14
80
a Ratio ligand/CuI = 2.
Depending on aryl group introduced, various cis:-
trans ratio were obtained. Interestingly, epimerization
was reduced with electron-donating rings and in-
creased significantly with electron-withdrawing rings
such as 4-nitrophenyl 18, 3,5-pyrimidyl 17, 4-trifluo-
romethyl 15, and in lesser extend 3-pyridyl 14 (cis:-
trans ratio, respectively, 0.2; 0.9; 1.5 and 1.9). This
observation is consistent with an epimerization of
carbon 12a (and not 6) which acidity is increased
in function of the electron-withdrawing property of
the aryl ring.
Table 2. Yields of cis and trans arylated Nortadalafil derivatives
R
R
O
H
O
N
N
H
12a
12a
N
O
N
O
6
6
O
O
H
H
N
N
H
H
trans configuration
6(R), 12a(S)
cis configuration
6(R), 12a(R)
6a, 8a-18a
O
O
6c, 8c-18c
Compound
R
Isolated
yield (%)
cis:trans
ratio
All these new unpatented tadalafil analogues with cis
configuration were evaluated for PDE5 inhibition in a
competitive immunoassay using bovine PDE510
(Table 3).
cis
80
trans
6
0
—
*
*
Our results show that aryl substituents are globally well
tolerated (IC50: 18–377 nM). Both electron-donating
and -withdrawing groups are tolerated at meta or para
positions. Hydrophobic substituents seem to be unfa-
vorable (para-trifluoromethyl 15a, para-chloro 13a,
para-butyl 12a). Interestingly, heterocycles can advanta-
geously replace the benzene ring. Indeed a 5-fold in-
crease in activity is observed when benzene is replaced
by thiophene in compound 11a. Compounds bearing a
pyridine or pyrimidine (respectively 14a and 17a) are
also more active that the parent compound 6a.
Although the best activity is obtained with the para-
methoxyphenyl substituent (9a, 16 nM), the series of
more polar heteroaryl-substituted compounds appears
more interesting, in particular 14a and 17a, both more
8
52
47
65
60
48
28
43
30
25
17
11
0
5
—
O
9
9.4
4.0
3.5
3.2
1.9
1.9
1.5
1.4
0.9
0,2
*
*
10
11
12
13
14
15
16
17
18
16
17
15
15
23
20
18
18
49
Cl
S
*
*
Cl
*
*
Table 3. Inhibitory activity for compounds 6a and 8a–18a
Compound
IC50 PDE5 nM standard
error
N
Tadalafil 3
6a
8a
3.3
91.0
0.6
CF3
10
18
4
68.0
16.0
*
*
9a
S
10a
11a
12a
13a
14a
15a
16a
17a
18a
141.0
18.0
30
4
152.0
134.0
28.0
52
43
5
N
N
377.0
57.0
53.0
55
9
NO2
7
55
122.0
*

792
T. Beghyn et al. / Bioorg. Med. Chem. Lett. 17 (2007) 789–792
Table 4. Kinetic aqueous solubility of some heteroarylated com-
pounds compared to that of tadalafil and N-phenylnortadalafil
References and notes
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2. Terrett, N. K.; Bell, A. S.; Brown, D.; Ellis, P. Bioorg.
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3. Daugan, A.; Grondin, P.; Ruault, C.; Le Monnier de
Gouville, A.-C.; Costes, H.; Kirilovsky, J.; Hyafil, F.;
Compound
Aqueous solubility
PBS buffer pH 7.4 (lM)
Tadalafil
6a
14a
9.7
2.4
33
`
Labaudiniere, R. . J. Med. Chem. 2003, 46, 4525.
17a
25
4. Daugan, A.; Grondin, P.; Ruault, C.; Le Monnier de
Gouville, A.-C.; Costes, H.; Linget, J.-M.; Kirilovsky, J.;
`
Hyafil, F.; Labaudiniere, R. J. Med. Chem. 2003, 46, 4533.
5. Maw, G. N.; Allerton, C. M. N.; Gbekor, E.; Million, W.
A. Bioorg. Med. Chem. Lett. 2003, 13, 1425.
6. Orme, M. W.; Sawyer, J. S.; Schultze, L. M.; Daugan, A.
C. -M.; Gellibert, F., Patent WO 0200656, 2002.
potent than the parent compound and more soluble11
(Table 4).
In summary, we have adapted a method using a cop-
per-I catalyzed arylation to prepare novel analogues of
tadalafil where the methyl group is replaced by various
aryl or heteroaryl groups. Depending on the nature of
the aryl substituent, the unwanted epimerization of the
tetrahydrocarboline system was partially or totally
avoided by lowering both temperature and the amount
of base and increasing the amount of catalyst. The
synthesized compounds were evaluated as PDE5 inhib-
itors and displayed significant activities comparable to
that of tadalafil although weaker. This route can be
used to access unpatented analogues of tadalafil and
could be crucial for the identification of more selec-
tive, more soluble PDE5 inhibitors in a competitive
context.
7. Klapars, A.; Huang, X.; Buchwald, S. L. J. Am. Chem.
Soc. 2002, 124, 7421.
8. Li, Q.; Li, T.; Woods, K. W.; Gu, W.-Z.; Cohen, J.; Stool,
V. S.; Galicia, T.; Hutchins, C.; Frost, D.; Rosenberg, S.
H.; Sham, H. L. Bioorg. Med. Chem. Lett. 2005, 15, 2918.
9. General procedure for arylation of Nortadalafil: (6R,12aR)-
6-Benzo[1,3]dioxol-5-yl-2,3,6,7,12,12a-hexahydro-pyrazi-
no[10, 20:1,6]-pyrido[3,4-b]indole-1,4-dione (Nortadalafil,
6a) (133 mM, 1 equiv), CuI (2 equiv), K3PO4 (2 equiv),
( )-trans-1,2-diaminocyclohexane (4.0 equiv) and iodo
derivatives (1 equiv) were dissolved in anhydrous dioxane
in inerted vessels and stirred at 14 °C for 14 days. The
reaction mixture was diluted with ethyl acetate and washed
with NH3/water (5/95) and water. The organic layer was
dried over MgSO4, evaporated to dryness and purified by
TLC.
10. Compounds were evaluated with a time-resolved fluores-
cence resonance energy transfer-based assay (HTRFÒ
technology) using the cGMP quantification kit (Cisbio
international # 62GMPPEB) and bovine cGMP-specific
PDE5 (Calbiochem # 524715). This quantitation method
relies on the competition between free cGMP and a
conjugate cGMP-fluorophore for the binding to a cGMP-
specific antibody labelled with europium cryptate. Fixed
amounts of bovine PDE5 (2 lL) and cGMP (150 nM) were
incubated 24 h at 37 °C in the presence of varying inhibitors
concentrations. Theses assays were performed in black half
area 96-well microplates (Corning # 3694). Assay volume
was 36 lL and the assay buffer contained 50 mM Tris/HCl,
pH 7.4, and 6 mM MgCl2. At the end of the incubation, the
detection reagents were added according to the manufac-
turer’s protocol. After 1 h incubation at room temperature
in the dark, the HTRFÒ signals were later read using a
Victor3V (Wallac 1420 Multilabel Counter; Perkin-Elmer).
Each inhibition assay was performed in duplicate and each
well read twice. IC50 values were obtained using the curve
fitting software XLfit4.2 (IDBS).
Acknowledgments
We are grateful to the institutions that support our lab-
´
oratory (Inserm, Universite de Lille2 and Institut Pas-
teur de Lille). This work was funded by ‘Region Nord-
´
Pas-de-Calais’ and EC. We acknowledge the expert sup-
port of the ‘Laboratoire d’Application RMN
(LARMN)’ team (U.Lille2) for NMR spectroscopy.
´
We thank Andre Tartar and Helene Gras-Masse for
thoughtful comments and discussions.
´ `
Supplementary data
Supporting information available: Measurement of opti-
cal rotation of compounds 6b, 6c and compound from
Scheme 3. Experimental procedures and analysis of
compounds 4–17. Supplementary data associated with
this article can be found, in the online version, at
doi:10.1016/j.bmcl.2006.10.069.
11. Kinetic solubilities were determined as previously
described in: Poulain, R.; Horvath, D.; Bonnet, B.;
Eckhoff, C.; Chapelain, B.; Bodinier, MC.; Deprez, B.
J. Med. Chem. 2001, 44, 3378.