Phosphodiesterase inhibitors. Part 5: Hybrid PDE3/4 inhibitors as dual bronchorelaxant/anti-inflammatory agents for inhaled administration

Article abstract of DOI:10.1016/j.bmcl.2012.08.121

(-)-6-(7-Methoxy-2-(trifluoromethyl)pyrazolo[1,5-a]pyridin-4-yl) -5-methyl-4,5-dihydropyridazin-3(2H)-one (KCA-1490) exhibits moderate dual PDE3/4-inhibitory activity and promises as a combined bronchodilatory/anti- inflammatory agent. N-alkylation of the pyridazinone ring markedly enhances potency against PDE4 but suppresses PDE3 inhibition. Addition of a 6-aryl-4,5-dihydropyridazin-3(2H)-one extension to the N-alkyl group facilitates both enhancement of PDE4-inhibitory activity and restoration of potent PDE3 inhibition. Both dihydropyridazinone rings, in the core and extension, can be replaced by achiral 4,4-dimethylpyrazolone subunits and the core pyrazolopyridine by isosteric bicyclic heteroaromatics. In combination, these modifications afford potent dual PDE3/4 inhibitors that suppress histamine-induced bronchoconstriction in vivo and exhibit promising anti-inflammatory activity via intratracheal administration.

Full text of DOI:10.1016/j.bmcl.2012.08.121

Bioorganic & Medicinal Chemistry Letters 23 (2013) 375–381
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Phosphodiesterase inhibitors. Part 5: Hybrid PDE3/4 inhibitors as dual
bronchorelaxant/anti-inflammatory agents for inhaled administration
Koji Ochiai a, Satoshi Takita a, Akihiko Kojima a, Tomohiko Eiraku a, Kazuhiko Iwase a, Tetsuya Kishi a,
Akira Ohinata a, Yuichi Yageta a, Tokutaro Yasue a, David R. Adams b, Yasushi Kohno a,
⇑
a Discovery Research Laboratories, Kyorin Pharmaceutical Co., Ltd, 2399-1 Nogi, Nogi-machi, Shimotsuga-gun, Tochigi 329-0114, Japan
b Institute of Chemical Sciences, Heriot-Watt University, Riccarton, Edinburgh EH14 4AS, United Kingdom
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 20 June 2012
Revised 25 July 2012
Accepted 30 August 2012
Available online 9 November 2012
(À)-6-(7-Methoxy-2-(trifluoromethyl)pyrazolo[1,5-a]pyridin-4-yl)-5-methyl-4,5-dihydropyridazin-3(2H)-
one (KCA-1490) exhibits moderate dual PDE3/4-inhibitory activity and promises as a combined bron-
chodilatory/anti-inflammatory agent. N-alkylation of the pyridazinone ring markedly enhances potency
against PDE4 but suppresses PDE3 inhibition. Addition of a 6-aryl-4,5-dihydropyridazin-3(2H)-one
extension to the N-alkyl group facilitates both enhancement of PDE4-inhibitory activity and restoration
of potent PDE3 inhibition. Both dihydropyridazinone rings, in the core and extension, can be replaced by
achiral 4,4-dimethylpyrazolone subunits and the core pyrazolopyridine by isosteric bicyclic heteroaro-
matics. In combination, these modifications afford potent dual PDE3/4 inhibitors that suppress hista-
mine-induced bronchoconstriction in vivo and exhibit promising anti-inflammatory activity via
intratracheal administration.
Keywords:
Asthma
COPD
Phosphodiesterase (PDE)
Dual PDE3/4 inhibitor
Inhaled administration
Crown Copyright Ó 2012 Published by Elsevier Ltd. All rights reserved.
Inhibition of phosphodiesterase 4 (PDE4) as a treatment option
for respiratory disease has become firmly established with the re-
cent launch of roflumilast for management of severe chronic
obstructive pulmonary disease (COPD).1 The utility of PDE4 inhib-
itors for treatment of COPD, and potentially also for asthma, arises
from their pronounced anti-inflammatory activity coupled with
bronchorelaxant properties.2 Inhibition of PDE3 also confers bron-
chorelaxant activity,3 however, and combined inhibition of PDE3
and PDE4 is thought to function additively or synergistically to in-
duce airway smooth muscle relaxation.4 For that reason we have
been exploring the potential of dual PDE3/4 inhibitors as agents
with enhanced efficacy for treatment of respiratory disease,5 and
in this context we have previously disclosed (À)-6-(7-methoxy-
2-(trifluoromethyl)pyrazolo[1,5-a]pyridin-4-yl)-5-methyl-4,5-
dihydropyridazin-3(2H)-one (KCA-1490, Fig. 1)5b as a lead compound
that exhibited promising activity in pharmacological models of
inflammation and bronchoconstriction.
A key challenge that hindered the clinical deployment of first
generation PDE4 inhibitors, of which rolipram is the archetypal
example, was their propensity for emetogenesis at doses required
for effective anti-inflammatory action.6 With second generation
PDE4-selective inhibitors, such as roflumilast, that problem has
been partially addressed, although treatment is still limited by
nausea and emesis at high doses.2c In our initial pharmacological
profiling, we found KCA-1490 to possess an improved therapeutic
window over roflumilast, in that it exhibited a significantly re-
duced emetogenic liability at doses required for potent bronchod-
ilatory and anti-inflammatory action.5b Unfortunately detailed
toxicological evaluation of KCA-1490 in several animal models
subsequently revealed that it was unsuitable as a drug develop-
ment candidate. In particular, cardiovascular effects, that we
attributed to PDE3 inhibition,7 were observed in dogs and rabbits.
Further development of dual PDE3/4 inhibitors for respiratory dis-
ease would be difficult without resolving this issue. One potential
solution to the problem would be to develop a dual PDE3/4 inhib-
itor for direct topical administration to the lung by inhalation. This
could allow dose sparing administration with substantially re-
duced systemic exposure to a compound, and it is precisely this
principle that has driven the successful development of inhaled
corticosteroids (ICS) for chronic application in asthma manage-
ment.8 Thus we set out to evaluate the potential of dual PDE3/4
inhibitors for inhaled delivery rather than oral administration.
KCA-1490 exhibits moderate levels of PDE3 and PDE4 inhibition
(Fig. 1). With the potential for cardiovascular effects, we had hith-
erto been cautious about developing a potent PDE3-inhibitory
component into the compound’s overall profile. However, given
that an ideal compound for inhalation would have reduced sys-
temic bioavailability, we considered that there would now be
scope for improving the in vitro inhibitory activity against both
PDE3 and PDE4. We describe here our strategy for modification
of KCA-1490 that led to the discovery of potent dual PDE3/4
⇑
Corresponding author.
E-mail address: yasushi.kohno@mb.kyorin-pharm.co.jp (Yasushi Kohno).
0960-894X/$ - see front matter Crown Copyright Ó 2012 Published by Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.08.121

376
K. Ochiai et al. / Bioorg. Med. Chem. Lett. 23 (2013) 375–381
PDE3 purine-
scanning Gln
O
N
O
O
X
NH
N
NH
N
N
∗
N
R
established
PDE3
N
N
H
O
inhibitors
CF3
N
N
R2
PDE3-inhibitory
extension
MeO
OR1
PDE4-inhibitory core
Altana dual PDE3/4 inhibitors
PDE4 purine-
scanning Gln
N
KCA-1490 (Š)-form
imazodan R = H
CI-930
PDE3A IC50 (μM): 0.36
PDE4B IC50 (μM): 0.042
R = Me
target compounds: enhanced dual PDE3/4 inhibition
O
O
O
achiral
replacement
N
N
N
N
(CH2)n
O
N
N
achiral
replacement
O
N
N
N
N
H
O
NH
N
H
O
heterobicycle
optimization
R
S
N
N
N
N
CF3
CF3
MeO
N
OMe
OMe
Figure 1. Planned development pathway for inhaled dual PDE3/4 inhibitors derived from KCA-1490.
Table 1
inhibitors exhibiting promising anti-inflammatory activity under
conditions of intratracheal administration.
Our existing structure–activity relationship (SAR) analysis in
the compound series leading to KCA-1490 revealed that N-alkyl-
ation of the pyridazinone ring could substantially improve PDE4-
inhibitory potency but at the expense of PDE3 inhibition, which
was severely compromised.5b,b The reason for this divergent effect
of N-alkylation is that the preferred binding mode for the pyrazol-
opyridine–pyridazinone core is fundamentally different in the cat-
alytic pockets of the two phosphodiesterase families. Thus, as
previously described with our binding models for KCA-1490,5c
the pyrazolopyridine subunit engages the conserved purine-scan-
ning glutamine in the PDE4 catalytic pocket whilst the bound ori-
entation is reversed in PDE3 so that pyridazinone ring fulfills this
role. In the PDE4-bound structure the pyridazinone NH does not
hydrogen bond to the protein but is conveniently positioned so
that N-alkylation extends the surface contact of the inhibitor with
residues around the rim of the catalytic pocket. In contrast, binding
of KCA-1490 to PDE3 exploits the pyridazinone N(1) centre as a key
hydrogen bond donor for engagement of the purine-scanning glu-
tamine, and unsubstituted lactam functionality is critical for PDE3-
inhibitory activity. We decided to investigate the addition of a 6-
aryl-4,5-dihydropyridazin-3(2H)-one extension to the N-alkyl
group, as summarised in Figure 1, on the grounds that this should
facilitate both enhancement of the PDE4-inhibitory component
and restore PDE3 inhibition by reinstating a subunit with the
established PDE3-inhibitory chemotype of imazodan and CI-930.9
A similar strategy has been explored by Altana Pharmaceuticals
in developing orally active phthalazinone–pyridazinone hybrid
PDE3/4 inhibitors.10
b
Compda
R1
n
Inhibition IC50 (nM)
PDE3A
PDE4B
KCA-1490
CF3
CF3
CF3
CF3
CF3
CF3
Et
—
—
2
3
4
5
3
4
360
2380
130
140
15
42
70
KCA-1450c
3a
3b
3c
3d
3e
3f
0.66
3
0.3
0.2
5.9
1.7
72
11
1.2
Et
a
Compounds 3a–3f were evaluated as equimolar mixtures of all four
stereoisomers.
b
Enzyme assays were performed using the core catalytic domains from the
PDE3A and PDE4B isoforms according to previously reported procedures.11,12 Data
reported are the mean of at least three experiments.
c
Racemic form of KCA-1490.
exemplified by compound 4 in Scheme 1, under basic conditions.
In the case of target compounds with an ethylene linker (n = 2),
the route was modified because of a tendency for elimination of
hydrogen bromide from the intermediate (2, n = 2) under basic
conditions, which afforded the corresponding N-vinyl pyridazinone
as a by-product. The modified route is illustrated for compound 3a,
and involved preparation of intermediate 5 from starting material
4 through a 4-step sequence consisting of O-alkylation with (2-
bromoethoxy)(tert-butyl)diphenylsilane, N-Boc protection, re-
moval of the TBDPS group and finally iodination of the alcohol.
The general synthetic route to our primary set of compounds for
evaluation (3a–3p, Tables 1–3) is summarized in Scheme 1. In
brief, for target compounds with trimethylene, tetramethylene
and pentamethylene linkers N-alkylation of the preformed core
(1 R = CF3 or Et)5c with the appropriate
a, x-dibromoalkane gave
2. The target compounds were then completed by O-alkylation of
a (para-hydroxyphenyl)pyridazinone or pyrazolone subunit, as

K. Ochiai et al. / Bioorg. Med. Chem. Lett. 23 (2013) 375–381
377
Table 2
Table 3
Compda
R2
Position
Inhibition IC50 (nM)
b
Compda
R2
R3
R4
Inhibition IC50 (nM)
b
PDE3A
PDE4B
PDE3A
PDE4B
3j
3m
H
F
H
H
270
1520
0.5
1.4
O
3c
3g
p
p
15
0.3
O
3n
3o
H
F
F
F
58
15
0.3
3.1
N
NH
NH
N
N
O
40000
6
3p
F
F
5.1
2.2
O
O
O
NH
N
N
N
NH
NH
NH
a
These compounds were tested in racemic form.
See footnote of Table 1.
b
3h
3i
p
p
360
0.1
20
38,000
the trifluoromethyl substituted series (3a–3d; R1 = CF3) 23- to
350-fold enhancements were seen in PDE4 inhibition relative to
the parent structure (KCA-1450). Moreover, the 6-aryl-4,5-dihyd-
ropyridazin-3(2H)-one extension was clearly effective as a PDE3-
inhibitory subunit. Optimal PDE3 inhibition (IC50 = 15 nM) was
found with the tetramethylene linker (compound 3c), whereas
the PDE4-inhibitory activity was essentially the same with both
pentamethylene and tetramethylene linkers (3d, 3c). We also as-
sessed two ethyl-substituted pyrazolopyridine analogues (3e, 3f)
in this compound subset because during the development of
KCA-1490 both ethyl and trifluoromethyl groups at the pyrazolo-
pyridine 2-position supported activity. With compounds 3e and
3f optimal activity against both PDE3 and PDE4 was seen with
the tetramethylene linker, where 3f showing essentially equipo-
tent inhibition.
O
3j
p
270
0.5
NH
N
N
N
3k
3l
m
m
590
0.5
O
NH
O
1730
0.064
NH
a
Compounds 3c and 3k were evaluated as equimolar mixtures of all four ster-
eoisomers; compounds 3g, 3h, 3i, 3j, and 3l were tested in racemic form.
b
See footnote in Table 1.
Having identified the tetramethylene chain as the most promis-
ing linker, we next explored options for replacement of the termi-
nal pyridazinone unit in the PDE3-inhibitory extension of the
compound series. We had initially retained a chiral 5-methyl-3,4-
dihydropyridazinone in this extension for the compound subset
in Table 1 because this subunit was well established in PDE3 inhib-
itors such as CI-930 (Fig. 1) and had also been successfully
exploited by Merck13 for the development of a series of PDE3-
selective inhibitors. However, we have recently shown that the
5-methyl-3,4-dihydropyridazinone ring of KCA-1490 and analogs
can be replaced effectively by an achiral 4,4-dimethylpyrazolone
subunit.5d We therefore sought to evaluate this and other achiral
pyridazinone-based options in our hybrid PDE3/4 inhibitor series,
compounds 3g–3j (Table 2). Assessment of this series revealed that
replacement of the dihydropyridazinone ring in parent structure
3c by fully unsaturated pyridazinone subunits (3g, 3i) severely re-
duced PDE3-inhibitory potency; with 4,5-dihydropyridazin-3(2H)-
one and 4,4-dimethyl-1H-pyrazol-5(4H)-one replacements (com-
pounds 3j and 3h respectively) activity loss against PDE3 was
somewhat less profound. These latter two compounds retained
similar levels of PDE4-inhibitory activity to the parent compound
(3c), though a significant reduction in PDE4 inhibition was again
observed with the unsaturated pyridazinone subunits (3g, 3i).
Given the loss in PDE3-inhibitory activity observed for 3g–3j,
we decided to assess the impact of repositioning the heterocycle
on the phenoxy ring of the extension. To do this we prepared
compounds 3k and 3l for comparison with 3c and 3j (Table 2).
Use of iodide 5 for N-alkylation of the preformed pyrazolopyri-
dine–pyridazine core (1, R1 = CF3), which in this case was the race-
mic form (KCA-1450) of KCA-1490, gave the required target after
final Boc deprotection. Target compounds 3q–3w (Tables 4 and
5) with tetramethylene linkers were prepared by similar strategies
to the route outlined in Scheme 1, but replacing the pyrazolopyri-
dine–pyridazinone starting material (1) with an alternative pre-
formed core. The synthesis of the appropriate core starting
materials has been described in our earlier work.5c,d
We first needed to select the optimum linker length for our pro-
posed hybrid PDE3/4 inhibitor structures. For this purpose we eval-
uated a series of alkylidine linkers [(CH2)n; n = 2–5] in compounds
3a–3f (Table 1), where a chiral 5-methyl-3,4-dihydropyridazinone
subunit was used in both the core and extension. For simplicity we
constructed these compounds from racemic building blocks and
tested them as equimolar mixtures of all four stereoisomers;
unsurprisingly no diastereoselective discrimination had arisen
during the assembly of the structures. Although the formation of
stereoisomeric mixtures in this compound subset was a complicat-
ing factor, we planned to remove at least one and preferably both
chiral subunits during subsequent development. The compounds
were assessed for inhibitory activity against the core catalytic do-
mains from PDE3A and PDE4B using previously reported proto-
cols.11,12 As expected, the hybrid compounds exhibited
substantial improvement in PDE4-inhibitory potency. Thus, in

378
K. Ochiai et al. / Bioorg. Med. Chem. Lett. 23 (2013) 375–381
Scheme 1. Reagents and conditions: (a) Br(CH2)nBr, NaH/DMF, rt; (b) phenol derivatives, K2CO3, KI/DMF, 60 °C; (c) Br(CH2)2OTBDPS, K2CO3/DMF, rt to 60 °C (80%); (d)
(Boc)2O, DMAP/MeCN, rt (quant.); (e) TBAF/THF, rt (81%); (f) I2, Ph3P, imidazole/THF, rt (98%); (g) 5, NaH/DMF, 60–80 °C (48%); (h) TFA/CH2Cl2, rt (48%).
Repositioning the 5-methyl-3,4-dihydropyridazinone ring in 3c
from the p-position to the m-position (3k) had little impact on
PDE4 inhibition but severely reduced PDE3 inhibition. With the
dimethylpyrazolone, as an achiral surrogate for the 5-methyl-3,4-
dihydropyridazinone, further weakening of PDE3-inhbitory activ-
ity was observed on altering the extension connectivity from p-3j
to m-3l. The latter compound showed further gains in PDE4-inhib-
itory performance, however, and proved to be a very potent inhib-
itor for PDE4 with 4 orders of magnitude selectivity over PDE3.
Based on the data in Table 2, the two most promising rings as
achiral subunits for the PDE3-inhibitory extension of our hybrid
PDE3/4 inhibitor series appeared to be the 4,5-dihydropyridazin-
3(2H)-one and 4,4-dimethyl-1H-pyrazol-5(4H)-one in compounds
3h and 3j. In an effort to restore the PDE3-inhibitory activity of
these compounds to levels comparable to 3c, we next considered
introduction of substituents on the aryl ring of the extension.
Merck had previously demonstrated that fluorination of the aryl
ring in their PDE3-selective 6-aryl-5-methyl-4,5-dihydropyrida-
zin-3(2H)-one series strongly enhanced PDE3 inhibition.13a Given
this precedent we focused on the impact of monofluorination at
the 2- and 3-positions of the phenoxy ring in compound 3j and
on 2,3-difluorination (Table 3). Compound 3m (R3 = F, R4 = H)
showed an activity loss against PDE3 but 3n (R3 = H, R4 = F) was
about fivefold more potent than 3j as a PDE3 inhibitor. The difluo-
rinated compound (3o) was 18-fold more potent than 3j, however,
and proved to be equipotent with 3c as a PDE3-inhibitor. Finally,
replacement of the 4,4-dimethylpyrazolone ring in 3o with the
4,5-dihydropyridazinone (3p) resulted in a further slight gains in
potency against PDE3 (IC50 5.1 nM) whilst retaining respectable
levels of activity against PDE4 (IC50 2.2 nM).
compound subset, we next turned our attention to achiral replace-
ments for the core 5-methyl-3,4-dihydropyridazinone ring. For this
work we used compound 3f as our reference point and compared
the activity of analogs 3q, 3r and 3s (Table 4), possessing, respec-
tively, 4,5-dihydropyridazin-3(2H)-one, pyridazin-3(2H)-one and
4,4-dimethyl-1H-pyrazol-5(4H)-one rings as achiral surrogates
for the core. Pleasingly, both compounds 3q and 3s exhibited com-
parable levels of dual PDE3/4-inhibitory potency to the parent (3f);
compound 3r showed a 12-fold loss in activity against PDE4.
Although ultimately our intention is to reduce systemic exposure
through inhaled delivery, we tested the potent, well-balanced dual
PDE3/4 inhibitors of Table 4 for their capacity to suppress hista-
mine-induced bronchoconstriction in guinea pigs under iv admin-
istration (dosed at 0.1 mg/kg). As expected, 3f and 3s showed
excellent bronchodilatory activity. Compound 3r was moderately
effective but analog 3q, with the 4,5-dihydropyridazin-3(2H)-one
core, displayed comparatively weak bronchodilatory activity. At
present the reason for the poorer performance of 3q is unclear.
With strategies now in place for eliminating both of the chiral
5-methyl-3,4-dihydropyridazinone subunits from our initial com-
pound subset (Table 1), we next focused on bringing the two ap-
proaches together for preparation of potent dual PDE3/4
inhibitors lacking any stereogenic centers (Table 5). Given the
well-balanced dual PDE3/4-inhibitory activity and strong bron-
chodilatory performance of compound 3s, we decided to take the
dimethylpyrazolone ring forward from Table 4 as the subunit for
the core of the structure. From the compounds in Table 3, both
dimethylpyrazolone (3o) and dihydropyridazinone (3p) rings, cou-
pled to a difluorinated phenoxy subunit, were considered as good
candidates for the PDE3-inhibitory extension of the hybrid struc-
ture. However, we also considered replacing the pyrazolo
[1,5-a]pyridine subunit of the core. We had previously shown that
Having developed a successful strategy for eliminating the
chiral subunit in the PDE3-inhibitory extension of our Table 1

K. Ochiai et al. / Bioorg. Med. Chem. Lett. 23 (2013) 375–381
379
Table 4
b
Compda
Top ring
Inhibition IC50 (nM)
Inhibitory effect on bronchoconstriction 0.1 mg/kg ivc (%)
PDE3A
1.2
PDE4B
O
N
N
3f
1.7
2.7
96
28
O
N
N
3q
0.27
O
N
N
3r
3s
2.7
1.7
20
71
92
O
N
N
0.6
a
b
c
Compound 3f was evaluated as an equimolar mixture of all four stereoisomers; compounds 3q–3s were tested in racemic form.
See footnote in Table 1.
The inhibitory activity of compounds on histamine-induced bronchoconstriction in guinea pigs was measured according to an established procedure.14
isosteric imidazopyridine and benzothiazole replacements for the
pyrazolopyridine in combination with a dimethylpyrazolone ring
were particularly effective in achiral analogs of KCA-1490
(Fig. 2).5d Thus, imidazopyridine 7 exhibited well-balanced dual
PDE3/4-inhibitory activity and excellent bronchodilatory activity
under both iv and po administration; benzothiazole 8 was equally
effective in suppressing histamine-induced bronchoconstriction
under iv administration but showed no effect via the oral route.5d
This is significant because for minimal systemic exposure an in-
haled agent should have negligible oral bioavailability. Thus with
ICS 40–90% of administered drug may be deposited in the pharynx,
resulting in ingestion and systemic steroid exposure by absorption
from the gastrointestinal tract.15 We therefore designed and eval-
uated four achiral hybrid PDE3/4 inhibitors (3t–3w) that included
imidazopyridine and benzothiazole subunits as the core heterobi-
cycle (Table 5). (Synthetic details for these compounds16 are pro-
vided in the Supplementary data.)
Compounds 3v and 3w, with the terminal dimethylpyrazolone
subunit, were less effective as PDE3 inhibitors than the cognate
dihydropyridazinones (3t and 3u respectively), though each com-
pound pair showed comparable levels of PDE4-inhibitory activity.
We evaluated the performance of all four compounds in suppress-
ing histamine-induced bronchoconstriction under iv administra-
tion (0.1 mg/kg). Both compounds with the terminal
dihydropyridazinone subunit (3t and 3u) showed potent activity.
In contrast, the corresponding dimethylpyrazolone analogs (3v
and 3w) displayed moderate to low potency, perhaps reflecting a
weaker PDE3-inhibitory component in their activity profiles. Given
the strong bronchodilatory performance of imidazopyridine 3t and
benzothiazole 3u, we took these compounds forward for evalua-
tion of anti-inflammatory activity under conditions of intratracheal
administration (see Supplementary data). Both 3t and 3u were
effective for suppression of LPS-induced inflammation (56%, 51%
at 100 lg/kg, respectively), thus confirming in vivo activity as in-
haled agents.
In conclusion, we have developed a series of potent hybrid
PDE3/4 dual-inhibitors by rational design based on an understanding
of the distinct PDE3- and PDE4-binding modes of lead compound,
KCA-1490. N-alkylation of the pyridazinone ring in KCA-1490
strongly enhances PDE4-inhibitory potency but compromises
PDE3 inhibition by disrupting hydrogen bonded engagement of
the purine-scanning glutamine in the PDE3 catalytic pocket.
However, by incorporation of a PDE3-inhibitory extension into
the N-alkyl group it is possible both to enhance levels of PDE4 inhi-
bition over KCA-1490 and instill potent PDE3 inhibition. Strategies
were developed both to eliminate the chiral 5-methyl-3,4-dihydro-
pyridazinone in the PDE4-inhibitory core of the hybrid structure
and avoid the need for a similar chiral subunit in the PDE3-inhib-
itory extension. In this way we were able to develop achiral hybrid
structures with low nanomolar dual PDE3/4-inhibitory activity
that potently suppress histamine-induced bronchoconstriction
in vivo under iv administration. We have further demonstrated
that two of the compounds, imidazopyridine 3t and benzothiazole
3u, show promising anti-inflammatory activity under conditions of

380
K. Ochiai et al. / Bioorg. Med. Chem. Lett. 23 (2013) 375–381
Table 5
Inhibitory effect on bronchoconstriction 0.1 mg/kg ivb (%)
a
Compd
R
6–5 Fused heteroaromatics
Inhibition IC50 (nM)
PDE3A
PDE4B
6.7
O
3t
5
87
96
N
NH
N
CF3
N
N
S
N
OMe
CF3
3u
6.9
0.77
3.2
OMe
N
O
3v
40
63
76
36
N
CF3
NH
OMe
S
N
CF3
3w
0.38
OMe
a
See footnote in Table 1.
See footnote in Table 4.
b
Supplementary data
O
O
O
NH
N
NH
N
NH
N
Supplementary data (preparation of 3a–3w) associated with
this article can be found, in the online version, at http://
dx.doi.org/10.1016/j.bmcl.2012.08.121.
S
N
N
CF3
CF3
CF3
N
N
N
References and notes
OMe
OMe
OMe
6
7
8
1. (a) Giembycz, M. A.; Field, S. K. Drug Des. Dev. Ther. 2010, 4, 147; (b) Ulrik, C. S.;
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PDE3A
PDE4B
IC50 4130 nM
IC50 110 nM
IC50 140 nM
IC50 150 nM
IC50 540 nM
IC50 63 nM
Inhibitory effect on histamine-induced bronchoconstriction in guinea pig
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0.1 mg/kg iv adiministration
0.1 mg/kg po adiministration
100%
>70%
96%
inactive
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Figure 2. Achiral dimethylpyrazolone analogs of KCA-1490.5d
intratracheal administration, thereby laying a foundation for the
potential development of these compounds as inhaled agents for
respiratory disease. Further evaluation of this hybrid PDE3/4 inhib-
itor series will be reported in due course.
Acknowledgments
We are grateful to Dr. S. J. MacKenzie, Dr. S. Hastings and Dr. R.
A. Clayton of the Kyorin Scotland Research Laboratory for their
many valuable suggestions and performing the PDE inhibition
assays.

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