Discovery of S-(2-guanidylethyl)-isothiourea (VUF 8430) as a potent nonimidazole histamine H4 receptor agonist

Article abstract of DOI:10.1021/jm060880d

During an in-house database screen, we identified S-(2-guanidylethyl)- isothiourea as a high affinity agonist for the histamine H₄ receptor, with a 33-fold selectivity over the histamine H₃ receptor and negligible affinity for the other histamine receptor subtypes. This nonimidazole ligand is introduced as a useful and complementary pharmacological tool that enables further unraveling of the physiological roles of the H₄ receptor.

Full text of DOI:10.1021/jm060880d

6650
J. Med. Chem. 2006, 49, 6650-6651
Discovery of S-(2-Guanidylethyl)-isothiourea
(VUF 8430) as a Potent Nonimidazole Histamine
H4 Receptor Agonist
Herman D. Lim, Rogier A. Smits, Remko A. Bakker,
Cindy M. E. van Dam, Iwan J. P. de Esch, and Rob Leurs*
Figure 1. Structures of reference H4R antagonists.
Leiden/Amsterdam Center for Drug Research, Department of
Medicinal Chemistry, Vrije UniVersiteit Amsterdam, De Boelelaan
1083, 1081 HV Amsterdam, The Netherlands.
ReceiVed July 26, 2006
Abstract: During an in-house database screen, we identified S-(2-
guanidylethyl)-isothiourea as a high affinity agonist for the histamine
H4 receptor, with a 33-fold selectivity over the histamine H3 receptor
and negligible affinity for the other histamine receptor subtypes. This
nonimidazole ligand is introduced as a useful and complementary
pharmacological tool that enables further unraveling of the physiological
roles of the H4 receptor.
Figure 2. Structures of H4R agonists.
The human histamine H4 receptor (H4R) is a Gi/o protein-
coupled receptor that was identified and cloned in 2000.1 Several
lines of evidence indicate that this receptor plays important roles
in the immune system. Activation of the receptor leads to
chemotaxis of mast cells and eosinophils2,3 and mediates the
production of inflammatory mediators, such as IL-16 and
leukotriene B4.4,5 These data suggest that H4R antagonists have
potential as drugs to treat inflammatory diseases, such as asthma
and allergy.6 To validate the H4R as a drug target, pharmaco-
logical tools such as selective agonists and antagonists are
needed. A few selective H4R antagonists have been published
in literature, most notably 1 and 2 (Figure 1).7,8
The H4R agonist OUP-16 described earlier shows only
moderate affinity for the H4R (pKi ) 6.9).9 Recently, we have
described 4-methylhistamine (4) as a selective human H4R
agonist (pKi ) 7.3) that shows >100-fold selectivity over the
human H1R, H2R, and H3R.10 In the same publication, the H2R
agonist and H3R antagonist dimaprit (5) was also identified as
an H4R agonist with moderate affinity (Figure 2).10 Here we
describe a focused screening effort of close analogues of
dimaprit (5) taken from our proprietary compound collection,
using SK-N-MC cells stably expressing the human H4R.11 The
affinity of the ligands for the human H4R was determined by
displacement of [3H]histamine binding, as described previ-
ously.10
Substitution of the tertiary amine group of dimaprit (5) by a
guanidine group (compound 6, Table 1) results in a dramatic
decrease in affinity. However, shortening the spacer of 6 that
connects the isothiourea and guanidine groups from a propylene
to an ethylene moiety leads to excellent H4R affinity (compound
7, S-(2-guanidylethyl)-isothiourea dihydrobromide, (VUF8430)).
This compound has a pKi of 7.5, which is almost as high as
that of histamine (3; pKi ) 7.9; Figure 3 and Table 1).
The two chemically different basic moieties of 7 are key
for affinity, as the corresponding compound with two isothiourea
groups and the compound with two guanidine groups (8 and 9,
respectively) have almost 10-fold lower affinity. Analogues
separating the guanidine and isothiourea moieties by a longer
carbon spacer have reduced affinity for the human H4R (compare
Table 1. pKi Values of Dimaprit (5) Analogues at the Human H4R, as
Determined by Displacement of [3H]Histamine Bindinga
compoundb
X
n
R
pKi
3 (histamine)
5 (dimaprit)
6
7
8
9
10
11
7.9 ( 0.1
6.5 ( 0.1
5.1 ( 0.1
7.5 ( 0.1
6.6 ( 0.1
6.4 ( 0.1
5.5 ( 0.1
5.4 ( 0.1
S
S
S
S
NH
S
S
3
3
2
2
2
4
6
N,N-dimethyl
guanidine
guanidine
isothiurea
guanidine
guanidine
guanidine
a Data shown are the mean ( SEM of at least three independent
experiments. b Histamine was used as the dihydrochloride salt, all other
compounds in Table 1 were used as the dihydrobromide salts.
10 and 11). This shows that the ethylene spacer of 7 is optimal
for interaction with the human H4R.
Compound 7 was originally derived from the H2R agonist
dimaprit (5), but it is poorly active at the H2R as determined at
the right atrium of the guinea pig (pD2 ) 3.8, R ) 0.4).12 In a
binding assay, 7 shows only minimal inhibition of [125I]-
iodoaminopotentidine binding at the human H2R expressed in
CHO cells (Figure 3A). Likewise, 7 displays minimal inhibition
of [3H]pyrilamine binding to the human H1R expressed in
COS-7 cells (Figure 3A). Furthermore, it shows only moderate
affinity (pKi ) 6.0 ( 0.1) at the human H3R, which is the closest
relative of the human H4R, as determined in a [3H]NR-
methylhistamine displacement binding assay on the human H3R
stably expressed in SK-N-MC cells (Figure 3).
Compound 7 exerts agonistic activity (pEC50 ) 7.3 ( 0.1)
at the human H4R, which is determined as the inhibition of
forskolin-induced cAMP-mediated increase in â-galactosidase
activity. This inhibition reaches the same level as that exerted
by histamine (3). Therefore, 7 is a full agonist (intrinsic activity
R ) 1). Furthermore, the activity of 7 is dose-dependently
shifted rightward by the H4R-antagonist 1 (Figure 3B). Schild
plot analysis of the antagonism of 1 against 7 results in a pA2
value of 7.8, with a slope of 0.95 ( 0.05, which is in accord
with the previously described pA2 of 1 against histamine (3).10
* To whom correspondence should be addressed. E-mail: leurs@
few.vu.nl. Phone: +31(0)205987600. Fax: +31(0)205987610.
10.1021/jm060880d CCC: $33.50 © 2006 American Chemical Society
Published on Web 10/26/2006

Letters
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 23 6651
previously described human H4R agonist 4-methylhistamine (4).
Therefore, these two compounds may complement each other
in their use as H4R pharmacological tools. Additionally, we
report an improved, high-yield synthesis of this ligand that gives
easy access to this novel pharmacological tool. The compound
is currently being used to further characterize the H4R in vivo.
Acknowledgment. We thank Dr. T. W. Lovenberg for
supplying us with SK-N-MC cells stably expressing the human
H3R or H4R. Thanks also to Ben Bruyneel for expert technical
assistance.
Supporting Information Available: Experimental protocols
and characterization data. This material is available free of charge
via the Internet at http://pubs.acs.org.
References
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Figure 3. Pharmacological characteristics of compound 7. (A)
Displacement of radioligands bound at the human H1R, H2R, H3R, and
H4R by different concentrations of 7. (B) Competitive antagonism by
1 of H4R agonism by 7, as measured by the inhibition of forskolin-
induced CRE-â-galactosidase activity.
Scheme 1. Synthesis of S-(2-Guanidylethyl)-isothiourea (7)a
(5) Takeshita, K.; Sakai, K.; Bacon, K. B.; Gantner, F. Critical role of
histamine H4 receptor in leukotriene B4 production and mast cell-
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(8) Terzioglu, N.; van Rijn, R. M.; Bakker, R. A.; de Esch, I. J.; Leurs,
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a Reagents and conditions: (i) ethanol, reflux; (ii) 48% HBr, microwave
130 °C, 20 min (3×); (iii) thiourea, ethanol, microwave 125 °C 15 min.
(9) Hashimoto, T.; Harusawa, S.; Araki, L.; Zuiderveld, O. P.; Smit, M.
J.; Imazu, T.; Takashima, S.; Yamamoto, Y.; Sakamoto, Y.; Kurihara,
T.; Leurs, R.; Bakker, R. A.; Yamatodani, A. A selective human
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Compound 7 also exerts full agonistic activity (pEC50 ) 6.5 (
0.1, R ) 1) at the human H3R in CRE-â-galactosidase assay
performed in SK-N-MC cells. Interestingly, at the highest tested
concentration (100 µM), 7 shows no agonistic activity at the
human H1R and only 50% agonistic activity at the human H2R
(data not shown). The latter agrees with the result reported
previously for H2R activity evaluated in the right atrium of
guinea pig.12
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Pyati, J.; Li, X.; Chai, W.; Carruthers, N.; Lovenberg, T. W. Cloning
and pharmacological characterization of a fourth histamine receptor
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(12) Sterk, G. J.; van der Goot, H.; Timmerman, H. The influence of
guanidino and isothiourea groups in histaminergic compounds on
H2-activity. Agents Actions 1986, 18, 137-140.
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The synthesis of S-(2-guanidylethyl)-isothiourea (7) has been
described in literature.13,14 According to this procedure, ami-
noalcohol 12 (Scheme 1) is treated with S-ethylisothiourea
hydrobromide 13, and in a one-pot procedure, the resulting
alcohol 14 was treated with thiourea and concentrated HBr
(48%). However, the isolated yield of this original procedure
is very poor (10%). By iterative treatment of 14 with HBr under
microwave conditions, isolation of bromide 15 and subsequent
formation of the isothiourea moiety, a considerable increase in
isolated yield (72%) can be obtained, making this compound
readily available.
(14) Shapira, R.; Doherty, D. G.; Burnett, W. T., Jr. Chemical protection
against ionizing radiation. III. Mercaptoalkylguanidines and related
isothiuronium compounds with protective activity. Radiat. Res. 1957,
7, 22-34.
In conclusion, we have discovered a new potent H4R agonist
that shows a different pharmacological profile than that of the
JM060880D