Comparison of different heterocyclic scaffolds as substrate analog PDE5 inhibitors

Article abstract of DOI:10.1016/j.bmcl.2005.05.090

Several different heterocyclic systems were compared as PDE5 inhibitor scaffolds. In addition to the known 3H-imidazo[5,1-f][1,2,4]triazin-4-ones and pyrazolopyrimidinones, isomeric imidazo[1,5-a][1,3,5]triazin-4(3H)-ones were also shown to be potent and selective PDE inhibitor scaffolds with in vivo activity. SAR trends were elucidated for sulfonamide derivatives with generality across different scaffolds.

Full text of DOI:10.1016/j.bmcl.2005.05.090

Bioorganic & Medicinal Chemistry Letters 15 (2005) 3900–3907
Comparison of different heterocyclic scaffolds
as substrate analog PDE5 inhibitors
Helmut Haning,* Ulrich Niewo¨hner, Thomas Schenke, Thomas Lampe,
Alexander Hillisch and Erwin Bischoff
BAYER HealthCare AG, Business Group Pharma, D-42096 Wuppertal, Germany
Received 14 April 2005; revised 20 May 2005; accepted 25 May 2005
Available online 29 June 2005
Abstract—Several different heterocyclic systems were compared as PDE5 inhibitor scaffolds. In addition to the known 3H-imi-
dazo[5,1-f][1,2,4]triazin-4-ones and pyrazolopyrimidinones, isomeric imidazo[1,5-a][1,3,5]triazin-4(3H)-ones were also shown to
be potent and selective PDE inhibitor scaffolds with in vivo activity. SAR trends were elucidated for sulfonamide derivatives with
generality across different scaffolds.
Ó 2005 Elsevier Ltd. All rights reserved.
Heterocyclic systems of the purinone type play an impor-
tant role in cellular biochemistry. Isosteric heterocycles
have been used as drugs to treat pathological situations
where purinones are involved in various indications,
e.g., antivirals, metabolic disorders (gout) and cancer.1a–c
Therefore, we were interested in bringing out a more
comprehensive comparison of different heterocyclic sys-
tems as PDE inhibitors. Herein, we present our results
with different heterocyclic scaffolds as PDE inhibitors,
as well as some fundamental substituent effects on inhib-
itory potency and selectivity (Fig. 1).
Intracellular levels of purine second messengers cAMP
(1) and cGMP (2) (Fig. 1) are regulated via synthesis
and degradation to AMP and GMP, respectively. Deg-
radative enzymes for these second messengers are phos-
phodiesterases (PDEs). PDEs constitute an enzyme
superfamily with at least 11 members and various iso-
forms differing by their substrate acceptance (either
cAMP, cGMP or both).
Figure 2 gives an overview of the investigated heterocy-
clic scaffolds.
Syntheses of 3H-imidazo[5,1-f][1,2,4]triazin-4-ones as
PDE inhibitors have been published.5 The synthesis of
the corresponding purinone systems was carried out,
as previously reported (Scheme 1).6,7
Previously we have disclosed potent and selective PDE5
inhibitors of the 3H-imidazo[5,1-f][1,2,4]triazin-4-one
type.2 It has been demonstrated that PDE5 inhibitors
carrying this structural fragment are consistently more
potent than congeners of the pyrazolopyrimidine type,
e.g., SildenafilÒ. This finding cannot be explained satis-
factorily on the basis of X-ray crystallograhpic data,
since both PDE5 inhibitors, VardenafilÒ (3) and Silde-
nafilÒ (4), show a similar binding pattern in the pub-
lished X-ray structure3, whereas at least the tenfold
difference in potency has been shown to be due to the
different heterocyclic systems.4
Pyrimidinone ring closure is effected by amino carbox-
amidoimidazoles after acylation with benzoic acid
chlorides under basic conditions. The corresponding 2-
amino-3-cyanoimidazoles can be used also, in these
instances cyclisation is best performed in the presence
of hydrogen peroxide. Alternately, cyclisation can be
achieved from the amino-carboxamido heterocycle and
benzoic acid esters using potassium tert-butoxide.8,9
There are many examples of pyrazolopyrimidine sys-
tems as PDE inhibitors or kinase inhibitors reported
in the literature.10–14 In brief, 2-amino-3-carbamoylpy-
razoles can be generated from hydrazines and alkoxy-
ethylidenemalonodinitrile during a Thorpe cyclisation
and subsequent hydrolysis of the nitrile to the amide
(Scheme 2). Again, acylation and ring closure are
achieved under similar conditions as for the purinones.
Keywords: PDE5 inhibition; Heterocycles.
*
Corresponding author. Tel.: +49 202 364459; fax: +49 202 364160;
e-mail: Helmut.Haning@bayerhealthcare.com
Deceased.
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.05.090

H. Haning et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3900–3907
3901
Figure 1. cAMP (1), cGMP (2), VardenafilÒ (3), and SildenafilÒ (4).
The isomeric pyrazolopyrimidinones, as found in Silde-
nafilÒ, were prepared as previously described (Scheme
3).15,16
Continuing in the series of heterocyclic scaffolds with
two nitrogen atoms in the five-membered part, imi-
dazo[1,5-a][1,3,5]triazin-4(3H)-ones also were prepared
(Scheme 4).
Imidazo[1,5-a][1,3,5]triazin-4(3H)-ones have been de-
scribed in the literature in the context of purinone isos-
teres, albeit only with either hydrogen or heteroatom
substitution at the 2-position.17
However, the previously reported synthesis proved to be
largely applicable to 2-aryl substituted cases as well.
Alkylation of 2-acylamino-ethylcyanoacetate, followed
by reaction with aryl amidines, delivers an aminopyri-
midinone that undergoes an interesting rearrangement
to yield the desired heterocyclic scaffold.18
Figure 2. Overview of investigated heterocyclic PDE inhibitor scaffolds.
Scheme 1. Synthesis of alkoxyphenylpurinones. Reagents and conditions: (a) i. HC(OEt)3, AcCN, RF, ii. R1NH2, RT; (b) Py, toluene, DMAP cat. or
NaH, THF, RT; (c) MeOH, NaOH or K2CO3.

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H. Haning et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3900–3907
Scheme 2. Synthesis of phenylpyrazolopyrimidinones. Reagents and conditions: (a) RF, neat; (b) i. MeOH, NH2NHR1, RF, ii. NH3, H2O2, EtOH,
48 h; (c) Py, toluene, DMAP cat.; (d) MeOH, NaOH, RF.
Scheme 3. Synthesis of phenylpyrazolopyrimidinones. Reagents and conditions: (a) i. N2H4, ii. alkylation, e.g., Me2(SO4), iii. NaOH; (b) i. HNO3,
H2SO4, ii. NH3, iii. SnCl2; (c) DCM, NEt3, DMAP; (d) NaOH, EtOH.
Scheme 4. Synthesis of imidazo[1,5-a][1,3,5]triazin-4(3H)-ones. Reagents and conditions: (a) R1Br, NaOEt; (b) NaOEt; (c) i. Py, TMSCl, RT ii.
HMDS, RF.
Recently, cycloalkyl substituted imidazo[1,5-a][1,3,5]
triazin-4(3H)-ones have been reported as PDE7 inhibi-
tors in the patent literature.19
Finally, two heterocyclic scaffolds with three nitrogen
atoms in the five-membered ring were prepared: 3-al-
kyl-3,6-dihydro-7H-[1,2,3]triazolo[4,5-d]pyrimidin-7-ones,
carrying the heterocyclic core of ZaprinastÒ, were
prepared either by alkylation of ZaprinastÒ or by
treating an appropriately substituted azide with
carbamoylmalononitrile in the presence of base
Members of the isoxazolo[4,5-d]pyrimidin-7(6H)-ones
also were prepared as purinone isosteres with two het-
eroatoms in the five-membered ring (Scheme 5).20

H. Haning et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3900–3907
3903
Scheme 5. Synthesis of phenyl-isoxazolo[4,5-d]pyrimidin-7(6H)-ones. Reagents and conditions: (a) CH3NO2, KF, propanol; (b) K2Cr2O7,
NBu4HSO4, CH2Cl2; (c) NH2OH.H2SO4, Tol/EtOH, RF; (d) ClCOCO2Et, NEt3, Et2O; (e) i. NH3 in MeOH, RT, ii.Zn, NH4Cl, water; (f) i. Py,
DMAP cat., 60 °C, ii. MeOH, Na2CO3, 4d RF.
Scheme 6. Synthesis of 3-alkyl-3,6-dihydro-7H-[1,2,3]triazolo[4,5-d]pyrimidin-7-ones. Reagents and conditions: (a) i. NaOEt; (b) Py, cat. DMAP;
(c) NaOH, EtOH.
Scheme 7. Synthesis of [1,2,4]triazolo[3,4-f][1,2,4]triazin-8-(7H)-ones. Reagents and conditions: (a) R1COOH, 165 °C, neat; (b) H2O2, HOAc, RF;
(c) NaH, dioxane, 90 °C, 16 h; (d) i. NaH, CO(OEt)2, 90 °C, 16 h ii. 2-(OEt)-Ethanol, RF, 16 h.
(Scheme 6), followed by acylation and ring
closure.
azole-3-thiol that in turn is treated with a 2-alkoxy ben-
zoic acid nitrile after desulfurisation. Thermal ring clo-
sure is achieved after acylation with diethylcarbonate.
This scaffold constitutes an overlay of the nitrogen atom
pattern of VardenafilÒ and SildenafilÒ.
Examples of [1,2,4]triazolo[3,4-f][1,2,4]triazin-8-(7H)-
ones have been described in the literature but mostly
with heteroatom substitution in position 2.21 They were
prepared according to Scheme 7.22
For the different heterocyclic systems, only activity data
were generated, no attempt was made to add in vitro PK
parameters to these SAR studies. Table 1 gives PDE
inhibitory activity of several different heterocyclic scaf-
Thiocarbonohydrazide is condensed with the corre-
sponding acid to yield a 4-amino-5-alkyl-4H-1,2,4-tri-

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H. Haning et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3900–3907
Table 1. IC50 data23 for 2-ethoxyphenyl PDE5 inhibitor heterocyclic scaffolds
Compound
Heterocycle
R1
R2
—
PDE1 (IC50, nM)
102
PDE5 (IC50, nM)
6
Propyl
7
8
9
c-Propyl
c-Butyl
c-Pentyl
—
—
—
1000
1000
500
300
100
50
10
Propyl
Me
—
110
11
12
13
Propyl
c-Pentyl
Me
Me
Me
Me
300
30
5
5
1000
200
14
15
Propyl
Propyl
Me
H
79015a
2715a
—
50
16
17
Propyl
Me
Me
200
40
40
20
c-Pentyl
18
19
Propyl
—
—
—
300
500
c-Pentyl
800
(continued on next page)

H. Haning et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3900–3907
3905
Table 1 (continued)
Compound
Heterocycle
R1
R2
—
PDE1 (IC50, nM)
50
PDE5 (IC50, nM)
200
20
21
c-Pentyl
c-Pentyl
—
200
20
Table 2. IC50 data for sulfonamide PDE5 inhibitors
Compound
Heterocycle
R1
R2
—
R3
PDE1 (IC50, nM)
PDE5 (IC50, nM)
22
Propyl
Me
—
102
23
24
25
26
27
28
29
Propyl
Propyl
Propyl
Me
Me
Et
Me
Me
3002
228
>1000
20
8
5
Me
Me
Me
Me
Et
1000
10
10
1
c-Pentyl
c-Pentyl
c-Pentyl
Me
10
1
Me
Me
>1000
7
30
31
Propyl
Me
Me
500
30
10
10
c-Pentyl
(continued on next page)

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H. Haning et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3900–3907
Table 2 (continued)
Compound
Heterocycle
R1
R2
R3
PDE1 (IC50, nM)
1000
PDE5 (IC50, nM)
32
33
34
Propyl
Me
Me
8
Propyl
Et
Et
Me
Me
>1000
>1000
20
40
c-Pentyl
35
36
Propyl
Me
Me
Me
40015a
6.615a
c-Pentyl
10
5
37
38
c-Pentyl
c-Pentyl
—
—
50
30
50
10
folds. 3H-imidazo[5,1-f][1,2,4]triazin-4-ones prove to be
the most potent scaffolds (11–13), followed by pyrazolo-
pyrimidinones (10 and 14, 15). Heterocycles with three
heteroatoms in the five- membered part were less active
(18–20). Cyclopentyl substitution in R1 leads to a higher
PDE1 inhibitory activity (12, 17, 20), a trend that is also
valid for sulfonamide derivatives (vide infra). Selectivity
varies only slightly between the different unfunctional-
ized scaffolds, with the notable exception of [1,2,4]triaz-
olo[3,4-f][1,2,4]triazin-8-(7H)-ones, which show a higher
inhibitory activity for PDE1.
Several interesting substituent effects were observed that
seem to constitute general SAR trends for distinct
scaffolds.
Ethyl substitution the R2 position consistently diminish-
es PDE1 activity and to a lesser extent PDE5 activity
(24, 29, 33, and 34).
Cyclopentyl at the R1 position (27, 31, 36, 37, and 38)
increases PDE1 inhibitory activity however, in combina-
tion with ethyl as R2 (29, 33), this effect is overcome by
the influence of the ethyl group. This effect is reminiscent
of the effect of an ethyl group in a recently published
purine PDE5 inhibitor series.25
Only the most potent scaffolds were selected for further
derivatisation to evaluate substituent effects on activity
and selectivity. Table 2 depicts SAR trends for arylsulf-
onamide derivatives—obtained by chloro-sulfonation
and subsequent reaction with an amine—of several
heterocycles. As previously observed, 3H-imidazo[5,1-
f][1,2,4]triazin-4-ones are the most potent PDE5 inhibi-
tors in our studies (23–29). The isomeric imidazo[1,
5-a][1,3,5]triazin-4(3H)-ones that are less potent (30)
however demonstrate in vitro activity comparable to
those of purinones (22) and pyrazolopyrimidinones
(32, 35) and show in vivo oral efficacy in a rabbit model
of erectile dysfunction.24
The reduction in PDE1 and PDE5 inhibition seen with
the ethyl analogues can be explained examining the pub-
lished X-ray structures for PDE1B26 and PDE5A.27
Docking the R2 ethyl compounds (e.g., 29) into PDE5A
shows potential steric clashes between the R2 substituent
and Ala767. In addition, the H-bonding network
around Tyr612 can become perturbed, leading to a
slight overall loss in activity. In human PDE1A, B and
C and bovine PDE1A, and B the amino acid corre-
sponding to PDE5-Ala767 is a histidine (PDB1B-
His373). The R2 ethyl compounds (e.g., 29) potentially
˚
The potency of [1,2,4]triazolo[3,4-f][1,2,4]triazin-8-(7H)-
ones (37) and isoxazolopyrimidinones (38) did not im-
prove upon addition of sulfonamide substituents.
intrude about 2 A less deep into the binding pocket than
VardenafilÒ to avoid steric clashes with this histidine.
The associated loss of contacts (e.g., to Gln421) explains

H. Haning et al. / Bioorg. Med. Chem. Lett. 15 (2005) 3900–3907
3907
the drastically reduced binding affinity of the R2-ethyl
derivatives to PDE1.
14. Markwalder, J. A.; Arnone, M. R.; Benfield, P. A.;
Boisclair, M.; Burton, C. R.; Chang, C.-H.; Cox, S. S.;
Czerniak, P. M.; Dean, C. L.; Doleniak, D.; Grafstrom,
R.; Harrison, B. A.; Kaltenbach, R. F., III; Nugiel, D. A.;
Rossi, K. A.; Sherk, S. R.; Sisk, L. M.; Stouten, P.;
Trainor, G. L.; Worland, P.; Seitz, S. P. J. Med. Chem.
2004, 47, 5894.
15. (a) Terrett, N. K.; Bell, A. S.; Brown, D.; Ellis, P. Bioorg.
Med. Chem. Lett. 1996, 6, 1819; (b) Bell, A. S.; Brown, D.;
Terrett, N. K. Eur. Pat. Appl. 1992, EP 463756, Chem.
Abstr. 1992, 116, 255626.
16. In the meantime the sildenafil scaffold 5-(2-ethoxyphenyl)-
1-methyl-3-propyl-1,6-dihydro-7H-pyrazolo[4,3-d]-pyrim-
idin-7-one is commercially available.
17. (a) Golankiewicz, B.; Januszczyk, P.; Ikeda, S.; Balzarini,
J.; De Clercq, E. J. Med. Chem. 1995, 38, 3558; (b)
Golankiewicz, B.; Holtwick, J. B.; Holmes, B. N.; Duesler,
E. N.; Leonard, N. J. J. Org. Chem. 1979, 44, 1740; (c)
Holtwick, J. B.; Golankiewicz, B.; Holmes, B. N.; Leon-
ard, N. J. J. Org. Chem. 1979, 44, 3835.
Methyl as R1 has a similar effect as ethyl substitution in
R2, PDE1 and PDE5 activities are reduced (26) puta-
tively due to a loss in hydrophobic interactions, e.g.,
with PDE5A-Leu725 and Phe786 and PDE1B-Met336
and Phe392.
However, the interpretation of binding potency or inhib-
itory activity on the basis of structural data has to be
done with caution especially for closely related deriva-
tives that do not differ by orders of magnitude in their
respective biological activity.
Overall these sulfonamide derivatives followed the same
potency order as the unsubstituted scaffolds. Activity in
most cases is increased with polar sulfonamide residues
on the aromatic ring. Selectivity can be tuned with both
parameters: substitution pattern and heterocyclic
scaffold.
18. Niewoehner, U.; Haning, H.; Lampe, T.; Es-Sayed, M.;
Schmidt, G.; Bischoff, E.; Dembowsky, K.; Perzborn, E.;
Schlemmer, K.-H. PCT Int. Appl. 2001, WO 01047928,
Chem. Abstr. 2001, 135, 92657.
19. Inoue, H.; Murafuji, H.; Hayashi, Y. PCT Int. Appl. 2004,
WO 04111053, Chem. Abstr. 2004, 142, 74617.
20. (a) Laurent, S.; Barbieux-Flammang, M.; Van Haverbeke,
Y.; Flammang, R.; Wentrup, C. Bull. Soc. Chim. Belg.
1994, 103, 181; (b) Niewoehner, U.; Haning, H.; Lampe,
T.; Es-Sayed, M.; Schmidt, G.; Bischoff, E.; Dembowsky,
K.; Perzborn, E.; Schlemmer, K.-H. PCT Int. Appl. 2001,
WO 01047934, Chem. Abstr. 2001, 135, 76888.(c) Gewald,
K.; Bellmann, P.; Jaensch, H. J. Liebigs Ann. Chem. 1980,
10, 1623.
A new PDE5 inhibitor class with oral efficacy—
imidazo[1,5-a][1,3,5]triazin-4(3H)-ones—was identified
during these comparative studies.
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