Selective urokinase-type plasminogen activator (uPA) inhibitors. Part 3: 1-Isoquinolinylguanidines

Article abstract of DOI:10.1016/j.bmcl.2004.03.094

A series of 1-isoquinolinylguanidines are shown to be potent inhibitors of uPA with selectivity over tPA and plasmin. Potency is enhanced by the presence of a 4-halo and a 7-aryl substituent, particularly when substituted by a 3-carboxylic acid group. Compound 13j (UK-356,202) combines excellent potency and selectivity, and has been selected as a candidate for clinical evaluation.

Full text of DOI:10.1016/j.bmcl.2004.03.094

Bioorganic & Medicinal Chemistry Letters 14 (2004) 3227–3230
Selective urokinase-type plasminogen activator (uPA)
inhibitors. Part 3: 1-Isoquinolinylguanidines
Christopher G. Barber,* Roger P. Dickinson and Paul V. Fish
Department of Discovery Chemistry, Pfizer Global Research and Development, Sandwich, Kent CT139NJ, UK
Received 22 December 2003; revised 17 March 2004; accepted 30 March 2004
Abstract—A series of 1-isoquinolinylguanidines are shown to be potent inhibitors of uPA with selectivity over tPA and plasmin.
Potency is enhanced by the presence of a 4-halo and a 7-aryl substituent, particularly when substituted by a 3-carboxylic acid group.
Compound 13j (UK-356,202) combines excellent potency and selectivity, and has been selected as a candidate for clinical evalu-
ation.
Ó 2004 Elsevier Ltd. All rights reserved.
Urokinase-type plasminogen activator (uPA) is a serine
protease that has been implicated as a key mediator of
cellular invasion and tissue remodeling.1 An inhibitor of
uPA may have a therapeutic role in disease situations
where uPA-driven degradation of extracellular matrix,
or uPA-dependent cell migration is thought to be
important––including tumour growth, metastasis, angio-
genesis and chronic wounds.2–7 The closely related en-
zyme tPA also acts via activation of plasminogen to
plasmin and is a key component of the fibrinolytic cas-
cade.8;9 Achievement of adequate selectivity over both
tPA and plasmin is therefore an important requirement
in a therapeutically valuable uPA inhibitor.
tion of various 3-substituents as in 2 gives further in-
creases in potency.10;11
As part of our programme of exploration of substitution
in the pyridine ring, we also investigated benzo-substi-
tution to give quinoline and isoquinoline derivatives.
This communication reports our results leading to
the identification of 1-isoquinolinylguanidines with
enhanced potency and selectivity for uPA.
In most cases, the required isoquinolinylguanidines were
prepared by reaction of guanidine with the corre-
sponding 1-chloroisoquinoline (Scheme 1). The 1-chloro
intermediates were either known compounds (4-CH3,12
5-OCH3,12 7-OCH3,12 4-Cl,13 4-Br,14 5-Br15) or were
prepared from the corresponding isoquinoline by oxida-
tion with either 30% H2O2/AcOH or mCPBA/CH2Cl2,
followed by treatment of the N-oxide with POCl3
(5-C6H5,16 5-OCH2C6H5,17 6-Br,18 7-OCH2C6H519). In
the case of 5-bromo-1-chloro- and 7-bromo-1-chloro-
isoquinolines, a mixture of 5- and 7-bromoisoquino-
line20 was oxidized, the product was treated with POCl3
and the mixture of chloro compounds was separated by
We previously reported that 2-pyridinylguanidine (1) is
a selective inhibitor of uPA, that potency is enhanced by
the presence of a 5-halo substituent, and that introduc-
CO2H
Cl
N
N
N
N
H2N
NH2
a
H2N
NH2
R
R
N
N
1
2
N
NH2
NH2
Cl
3a-q
4a-q
Keywords: uPA; Urokinase plasminogen activator; UK-356202;
1-Isoquinolinylguanidine.
Scheme 1. Reagents and conditions: (a) Guanidine HCl, NaH,
DMSO, 100 °C.
* Corresponding author. Tel.: +44-1304-641182; fax: +44-1304-6519-
87; e-mail: christopher_barber@sandwich.pfizer.com
0960-894X/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2004.03.094

3228
C. G. Barber et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3227–3230
a
b
Br
5-/7-
Br
5-/7-
N
N
O
5
6
Br
c, d
e
+
h
N
N
N
N
N
OHC
HOCH2
Br
Cl
Cl
Cl
Cl
Cl
c, g
f
3e
3j
7
3o
N
N
H3C
HO2C
Ar
OH
Cl
Cl
3p
3 m,n
3q
Scheme 2. Reagents and conditions: (a) 30% H2O2, AcOH, 80 °C; (b) POCl3, CH2Cl2, reflux; (c) n-BuLi, THF/Et2O (1:1), )78 °C; (d) DMF;
(e) NaBH4, MeOH; (f) MeLi, THF; (g) CO2; (h) Ar(BOH)2, tetrakis(triphenylphosphine)palladium(0), Na2CO3, H2O, MeO(CH2)2OMe.
fractional crystallization from THF at )78 °C to give the
individual isomers 3e and 3j (Scheme 2). The 7-bromo
isomer 3j was used for the preparation of further 7-
substituted intermediates. Thus, treatment with n-BuLi
and DMF gave the aldehyde 7, which was converted to
the alcohols 3o and 3p. The acid 3q was prepared by
successive treatment of 3j with n-BuLi and CO2. 7-Aryl
substituted compounds 3m and 3n were prepared via
Suzuki coupling of 3j with the corresponding arylbo-
ronic acids.
treated with N,N0-bis(t-butoxycarbonyl)-S-methyl-
isothiourea in the presence of mercury(II) chloride,
followed by deprotection with trifluoroacetic acid.10
Compounds were tested for their ability to inhibit uPA,
tPA and plasmin as described previously.10 Results for
simple isoquinolinylguanidine derivatives, expressed as a
calculated Ki, are listed in Table 1, together with com-
parative figures for 1.
Benzo-fusion across the 3,4-positions of the pyridine
ring to give the 1-isoquinolinylguanidine 4a leads to a
marked increase in potency compared with 1, and fusion
across the 4,5-positions as in 3-isoquinolinylguanidine
15 is also favourable.30 A clear drop in potency is
observed with 2-quinolinylguanidine 14, which is con-
sistent with our previous finding that a 6-substituent in
the pyridine ring is unfavourable.10 Compound 4a also
shows a reasonable level of selectivity with respect to
tPA and plasmin. Based on these results, further SAR
exploration was carried out in the 1-isoquinolinylgua-
nidine series, and results for a range of analogues are
summarized in Table 2.
An alternative approach was used for the preparation of
7-substituted-4-chloro intermediates (Scheme 3). Con-
version of 4-bromocinnamic acid to the acid chloride
followed by treatment with sodium azide gave the azide
9. This was submitted to a Curtius rearrangement
(CAUTION, risk of explosion)21 with concomitant
cyclization to give the isoquinolin-1-one 10, which was
treated with PCl5 to give the 1,4-dichloroisoquinoline
intermediate 11. This was used for preparation of a
series of 7-aryl derivatives (12a–i) via Suzuki coupling
with a variety of arylboronic acids. Treatment of the 1,4-
dichloroisoquinolines 12a–i with guanidine in DMSO
gave the required isoquinolinylguanidine derivatives
13a–i. Acid hydrolysis of the nitriles 13a–i gave the
corresponding acids 13j–l.
From the results in Table 2 it is apparent that methyl
substitution at the 4-position (4b) is tolerated but a
chloro or bromo substituent is preferred (4c and 4d).
This parallels the situation in the pyridine series where
An alternative approach was used to prepare 2-quino-
linylguanidine 14 and 3-isoquinolinylguanidine 15. In
these cases, the corresponding amino compounds were
5-halo substitution also gave an increase in potency.10
A
small drop in potency results from introduction of
COR
Cl
N
Cl
Cl
Cl
Cl
f
e
c
d
NH
N
Br
N
N
Ar
Br
Br
Ar
O
NH2
NH2
8 : R = OH
9 : R = N3
13a-i
a, b
10
11
12a-i
Scheme 3. Reagents and conditions: (a) SOCl2, 23 °C; (b) NaN3, Me2CO, )10to 0 °C; (c) Ph2O, 270 °C (Caution, risk of explosion); (d) PCl5, 140 °C;
(e) Ar(BOH)2, tetrakis(triphenylphosphine)palladium(0), Na2CO3, H2O, MeO(CH2)2OMe; (f) Guanidine HCl, NaH, DMSO, 100 °C.

C. G. Barber et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3227–3230
Table 1. Enzyme inhibition data for 2-pyridinylguanidine and benzo-fused analogues
3229
NH2
NH2
N
N
N
N
N
N
N
N
NH2
NH2
H2N
NH2
H2N
NH2
15
1
14
4a
uPA Ki (lM)
tPA Ki (lM)
Plasmin Ki (lM)
29
a
209
367
a
9.3
a
1.92
126
68
a
a
(a) <50% inhibition at 1 mM.
Table 2. Enzyme inhibition data for 1-isouinolinylguanidines
Table 3. Enzyme inhibition data for 7-aryl-4-chloro-1-isoquinolinyl-
guanidines
4
5
6
R
7
3
Cl
N
8
N
N
NH2
R'
4
N
NH2
NH2
13
Compd
R
Ki (lM) or % inhibition
NH2
Ki (lM) or % inhibition
uPA
tPA
Plasmin
68
Compd
R0
4a
4b
4c
4d
4e
4f
H
1.92
126.2
a
uPA
tPA
Plasmin
4-CH3
4-Cl
4-Br
1.40a
0.83
0.58
13a
13b
13c
13d
13e
13f
13g
13h
13i
H
4-CH3
0.31
0.13
0.10
0.083
0.20
0.10
0.30
0.49
0.18
0.037
0.082
0.009
a
a
7.30
128.5
159.2
140.1
b
b
19
15.5
b
b
2-OCH3
3-OCH3
4-OCH3
3,4-(OCH2O)
3-CN
14.5
ND
5-Br
5-OCH3
4.93
3.62
237.8
c
b
3.00
3.20
4g
4h
4i
5-OCH2C6H5
5-C6H5
6-Br
4.8013
>100
2.34
a
b
123.9
b
47.5
131.4
50.8
a
b
4.10
82% @ 10 lM
4-CN
b
4j
7-Br
7-OCH3
0.80
0.66
95.1
a
2-OCH3, 5-CN
3-CO2H
4-CO2H
2-OCH3, 5-CO2H
b
a
4k
4l
13j
b
c
7-OCH2C6H5
7-C6H5
7-(3,4-
4.8013
0.40
0.34
a
13k
13l
b
65% @ 3 lM
4.75
4m
4n
123.9
d
47.5
12
b
(a) <50% inhibition at 30 lM; (b) <50% inhibition at 100 lM; (c) <50%
inhibition at 10 lM.
OCH2O)C6H4
7-CH2OH
4o
4p
4q
1.30a
0.64
1.00
d
7-CH(OH)CH3
7-CO2H
a
a
24
a
substitution of the 7-phenyl substituent, which is sum-
marized in Table 3.
(a) <50% inhibition at 100 lM; (b) <50% inhibition at 1 mM; (c) <50%
inhibition at 300 lM; (d) 55% inhibition at 100 lM.
A consistent additive effect upon the potency is seen
with the 4-chloro substituent in the presence of a 7-aryl
group (compare 13a vs 4m, and 13f vs 4n). The intro-
duction of additional alkyl or alkoxy substitution in the
phenyl ring is beneficial (13b–f). However, several of
these compounds show significantly reduced selectivity
with respect to plasmin. The most marked effect on uPA
potency results from introduction of a carboxylic acid
substituent as in 13j–l. A meta-carboxylic acid gives the
highest potency, and the 2-methoxybenzoic acid ana-
logue 13k is the most potent compound in the series.
The structural similarity between 13j and 13l and the
most potent compound in the pyridine series (2, Ki
0.17 lM), suggests that the pyridyls are functioning as
ring-opened isoquinolines.22
5-substituents (4e–g), and 5-phenyl substitution is par-
ticularly detrimental (4h). The most favourable effects
are seen with 7-substitution. Thus, the bromo and
methoxy analogues 4j and 4k show an increase in
potency, and aryl substitution as in 4m and 4n is par-
ticularly beneficial. Polar substitution, as in the alcohol
4o and the carboxylic acid 4q has no significant effect,
but increasing the lipophilicity of the alcohol (4p) is
beneficial. Some activity against tPA and plasmin was
apparent with some compounds but, in most cases,
selectivity for uPA was excellent.
From these results it was concluded that substitution at
the 4-position of the isoquinoline ring with halogens and
at the 7-position with lipophilic substituents, particu-
larly aryl, is favourable. This further mirrored our pre-
vious work with the pyridylguanidines,10 and suggested
that further potency gains could be achieved by the
In summary, we have found that 1-isoquinolinylguani-
dines are selective inhibitors of uPA, and that potency is
enhanced by the presence of a 4-halo and a 7-aryl sub-
stituent. The carboxylic acid analogue 13j (UK-356,202)

3230
C. G. Barber et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3227–3230
18. Nerenz, H.; Meier, M.; Grahn, W.; Resiner, A.; Schmalz-
lin, E.; Stadler, S.; Merrholz, K.; Brauchle, C.; Jones, P.
G. J. Chem. Soc. (P2) 1998;(2), 437.
combines excellent potency and selectivity, and has been
selected for clinical evaluation.
19. Baker, R.; Curtis, N. R.; Kulagowski, J. J.; Leeson, P. D.;
Smith, A. L.; Ridgill, M. P. Int. Patent Application
WO9,420,459; Chem. Abstr. 1994, 121, 300924.
Acknowledgements
20. Tyson, F. T. J. Amer. Chem. Soc. 1939, 61, 183.
21. A warm solution of the azide in diphenyl ether was added
dropwise to stirred diphenyl ether at 270 °C so that
reaction was instantaneous and no significant concentra-
tion of azide was allowed to accumulate.
We thank Y. M. Ailwood, G. Bish, N. Smith and R. S.
Strang for their assistance in preparing the compounds,
M. F. Burslem, G. Easter and B. Williams-Jones for the
biological data.
22. Since completion of this work, X-ray data have been
published23–29, which can help to rationalize the activity of
the compounds described here. Comparison of the X-ray
crystal structures of the uPA inhibitors amiloride and
phenylguanidine bound to re-engineered human uPA23
showed that both bind in the S1 pocket. Nienaber24
proposed that while these two scaffolds do not overlap,
their combination describes a naphthalene ring that could
have good activity, and further, that a 4-Cl would enhance
potency by occupying the S1b subsite. The SAR that we
describe here system supports that prediction.
23. Spraggon, G.; Phillips, C.; Nowak, U. K.; Ponting, C. P.;
Saunders, D.; Dobson, C. M.; Stuart, D. I.; Jones, E. Y.
Structure 1995, 3, 681.
24. Nienaber, V.; Wang, J.; Davidson, D.; Henkin, J. J. Biol.
Chem. 2000, 275, 7239.
25. Zeslawska, E.; Schweinitz, A.; Karcher, A.; Sondermann,
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