Rigidified 2-aminopyrimidines as histamine H4 receptor antagonists: Effects of substitution about the rigidifying ring

Article abstract of DOI:10.1016/j.bmcl.2010.01.131

Three novel series of histamine H₄ receptor (H₄R) antagonists containing the 2-aminopyrimidine motif are reported. The best of these compounds display good in vitro potency in both functional and binding assays. In addition, represen

Full text of DOI:10.1016/j.bmcl.2010.01.131

Bioorganic & Medicinal Chemistry Letters 20 (2010) 1900–1904
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Bioorganic & Medicinal Chemistry Letters
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Rigidified 2-aminopyrimidines as histamine H4 receptor antagonists:
Effects of substitution about the rigidifying ring
*
John R. Koenig , Huaqing Liu, Irene Drizin, David G. Witte, Tracy L. Carr, Arlene M. Manelli, Ivan Milicic,
Marina I. Strakhova, Thomas R. Miller, Timothy A. Esbenshade, Jorge D. Brioni, Marlon Cowart
Neuroscience Research, Global Pharmaceutical Research and Development, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064-6100, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Three novel series of histamine H4 receptor (H4R) antagonists containing the 2-aminopyrimidine motif
are reported. The best of these compounds display good in vitro potency in both functional and binding
assays. In addition, representative compounds are able to completely block itch responses when dosed ip
in a mouse model of H4-agonist induced scratching, thus demonstrating their activities as H4R
antagonists.
Received 4 December 2009
Revised 26 January 2010
Accepted 28 January 2010
Available online 1 February 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Histamine
Histamine H4 receptor
The histamine H4 receptor (H4R) has attracted considerable
interest from both academia and industry since its cloning several
years ago.1 H4R has been found to be expressed in lymphocytes,
mast cells, and dendritic cells,2 and H4 antagonists have been
shown to block histamine-mediated shape change and chemotaxis
of eosinophils and mast cells.3 Thus, it has been proposed that
antagonists of H4R could be used to treat conditions arising from
immune and inflammatory responses.1c Indeed, H4 antagonists
Scheme 1. General preparation of compounds. Reagents and conditions: (i) NaH, dimethyl carbonate, reflux, 2 h; (ii) guanidine, DMF, 120 °C, overnight; (iii) TsCl, DMAP, Et3N,
CH2Cl2, rt, overnight; (iv) diamine or Boc-protected diamine, Et3N, CH3CN, reflux, 48 h; or hydroxylamine, KOtBu, 0 °C to rt, THF, 48 h; (v) TFA, CH2Cl2, rt, 1 h.
* Corresponding author.
E-mail address: john.r.koenig@abbott.com (J.R. Koenig).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.01.131

J. R. Koenig et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1900–1904
1901
have been reported to be active in in vivo models of H4-agonist in-
duced itch4 and ovalbumin-induced airway inflammation.5 In
addition, the antinociceptive activity of H4 antagonists in models
of inflammatory, postsurgical, osteoarthritic, and neuropathic pain
has been reported by ourselves6 and others.7
The in vitro results at H4R are shown in Tables 1–3, with the
functional assays being run as previously described.6 In an earlier
publication,6b we demonstrated that compounds with a rigidifying
cycloalkyl ring are potent at H4, with six- or seven-membered rings
being preferred and roughly equivalent to each other in terms of
We recently disclosed some results of our studies of H4R ligands
containing the 2-aminopyrimidine moiety. We found that 6-aryl-
2-aminopyrimidines, exemplified by I, can be potent antagonists
of H4 at both human and rat receptors but tend to suffer from prob-
lems with metabolism and off-target activity.6a Rigidification of the
structure by including a new ring junction, such as in compound II,
largely overcomes these issues while retaining much of the original
potency, with the best potencies being obtained when the rigidify-
ing ring is six- or seven-membered.6b We have also demonstrated
that the aryl ring of II could be appended rather than fused to the
rigidifying ring (e.g., 1) with only a modest reduction in potency.
Table 1
Summary of in vitro potency at histamine H4 receptorsa
NH2
N
N
R1
A
R2
n
Compd R1
R2
n
A
Human H4
FLIPR
Rat H4 FLIPR
pKb SEM
or
pKb SEM
or
pEC50 SEM
pEC50 SEM (% eff)
(% eff)
N
N
1
2
Ph
H
H
H
2
2
7.24 0.02
6.57 0.02
7.26 0.07
NH
6.46 0.11
(80%)
NH
H
N
3
4
5
Ph
Ph
Ph
H
H
H
2
1
1
7.88 0.27
7.52 0.11
8.04 0.06
8.24 0.24
H
HN
Intrigued by the in vitro results obtained with 1, and in an effort
to further understand the SAR of such compounds, we prepared a
series of similar rigidified analogs with substitution(s) about the
rigidifying ring. The results of this campaign are presented in this
communication.
Synthesis of the compounds in this study proceeded according
to the general synthetic scheme in Scheme 1. Ketone A, available
commercially or prepared according to Scheme 2 or 3, was con-
verted to the corresponding 1,3-dicarbonyl B with sodium hydride
and dimethyl carbonate. Reaction of B with guanidine with heating
in DMF afforded hydroxypyrimidine C, which upon tosylation and
reaction with a diamine or hydroxylamine yielded compound D.
For those diamines that were Boc-protected, a final deprotection
step with TFA removed the Boc group.
N
7.28 0.21
(49%)
NH
H
N
7.61 0.10
(47%)
H
HN
N
6
Ph
H
1
7.34 0.03
<7.34b
NHMe
N
7
8
Ph
Ph
Ph
H
H
H
H
2
2
2
2
7.88 0.05
7.57 0.05
7.59 0.08
6.31 0.05
7.51 0.08
NHMe
NHMe
N
N
7.26 0.06
(84%)
Ketone A with spiro-cyclopentyl substitution alpha to the car-
bonyl was prepared via pinacol rearrangement of bi(cyclopen-
tane)-1,10-diol, as outlined in Scheme 2. With other spiro-
cycloalkyl substitution, or with bis-alkyl or bis-benzyl substitution,
the required ketones were prepared according to Scheme 3.8
9
7.97 0.20
6.37 0.02
HN
N
N
N
10
NH
NH
CO2Me
CO2H
11
12
H
H
2
2
5.63 0.07
<5.44b
N
N
N
NH
5.37 0.005 <4.91b
Scheme 2. Synthesis of spiro-cyclopentyl compounds A. Reagents and conditions:
(i) (CH3O)3CH, BF3–OEt2, ꢀ20 °C to rt, 2 h.
N
H
H
H
13
14
15
Ph
Ph
Ph
2
2
2
7.17 0.06
6.61 0.05
7.15 0.09
7.37 0.13
HN
N
N
6.84 0.16
7.73 0.15
NH
Scheme 3. Synthesis of compounds A with other spiro-cycloalkyl substitution or
with bis-alkyl or bis-benzyl substitution. Reagents and conditions: (i) KOtBu,
electrophile, tBuOH, rt, overnight (electrophiles: 1-chloro-5-iodopentane, MeI, EtI,
BnBr, or 1,2-bis(bromomethyl)benzene).
NHMe
a
n P 2.
n = 1.
b

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J. R. Koenig et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1900–1904
Table 2
Summary of in vitro potency at histamine H4 receptorsa
NH2
N
N
R
A
R
Compd
R
A
Human H4 FLIPR pKb SEM
Rat H4 FLIPR pKb SEM or
or pEC50 SEM (% eff)
pEC50 SEM (% eff)
N
16
17
18
Me
Me
Et
7.60 0.22
7.09 0.20
7.11 0.08
7.04 0.18 (62%)
<6.26b (64%)
NH
N
N
NH
<6.81b (37%)
NH
N
19
Bn
4.57 0.004 (86%)
4.60 0.03 (129%)
NH2
a
n P 2.
n = 1.
b
Table 3
Summary of in vitro potency at histamine H4 receptorsa
NH2
N
R
N
R
A
m
Compd
20
R
m
1
A
Human H4 FLIPR pKb SEM
or pEC50 SEM (% eff)
Rat H4 FLIPR pKb SEM or
pEC50 SEM (% eff)
N
N
H
H
8.01 0.02
8.07 0.18
<5.32b (77%)
<6.60b (35%)
NH
H
21
1
H
HN
CH3
N
22
H
1
7.66 0.07
6.90 0.03 (84%)
HN
N
COOCH3
NH2
23
24
H
H
1
1
4.90 0.02
<4.67
4.77 0.07 (52%)
<4.68
N
N
COOH
NH2
25
26
27
H
H
H
1
1
1
<4.16
4.63 0.05 (48%)
<6.00b (88%)
NH2
N(CH3)2
8.11 0.08
7.70 0.08
O
CH3
N
<5.48b (86%)
O
N
28
29
H
H
1
1
8.56 0.05
8.54 0.08
5.42 0.28 (87%)
7.48 0.06 (58%)
NH
N
NH

J. R. Koenig et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1900–1904
1903
Table 3 (continued)
Compd
R
m
A
Human H4 FLIPR pKb SEM
Rat H4 FLIPR pKb SEM or
or pEC50 SEM (% eff)
pEC50 SEM (% eff)
N
30
31
32
33
H
H
H
H
1
1
1
1
8.34 0.07
7.60 0.13 (46%)
N
H
N
N
8.57 0.09
7.28 0.10
5.96 0.06
6.76 0.31 (71%)
6.87 0.02 (63%)
5.13 0.05 (53%)
NH2
NH2
H
N
NH
H
H
N
34
35
36
H
H
H
1
2
2
5.99 0.05
7.32 0.06
7.33 0.01
5.65 0.10 (48%)
<4.86b (105%)
<4.96b (84%)
NH
H
N
NH
N
NH2
H
N
37
H
2
7.00 0.10
5.81 0.1 (50%)
H
N
H
N
NH
38
39
1
1
4.57 0.0006 (49%)
6.30 0.02
4.86 0.24 (135%)
6.09 0.08
N
NH2
a
n P 2.
n = 1.
b
potency. We had also demonstrated that such structural rigidifica-
tion was helpful in increasing oral bioavailability and drug-like-
ness, presumably by reducing the number of rotatable bonds. For
compounds 1–15 (Table 1), the marked effect of including an ap-
pended phenyl group alpha to the pyrimidine ring is evident in
comparing the potencies of compounds 1 and 2, suggesting that
some substitution is beneficial about the rigidifying cycloalkyl ring.
However, potencies decrease when the phenyl group is placed at
the beta position instead of the alpha position (potencies of 14 vs
1). Furthermore, it appears that a less lipophilic substitution at this
position may not be optimal, as clearly diminished potencies are
observed for the alpha-pyridyl compounds 10, 11, and 12. As has
been noted earlier with related chemical series,6a many of the com-
pounds showing antagonism in the human H4R FLIPR assay show
partial agonism in the rat H4R FLIPR assay. These results may be
due to the substantial difference in sequences between the human
and rat H4Rs (69% homology).9
weakest of these compounds (19) even showed agonist activity
in both the human and rat H4 FLIPR assays, for reasons that are
not clear. However, it was found that inclusion of a spiroalkyl ring
at the alpha position (in effect, ‘tying back’ the ethyl groups of 18)
was tolerated and in fact led to some quite potent compounds,
such as 28, 29, 30, and 31 (Table 3). Importantly, such spiro-cyclo-
pentyl compounds retain the favorable physicochemical properties
of the appended compounds10 but with generally higher potencies
at H4. It was found that compounds with a spiro cyclohexyl ring or
spiro indenyl group at the alpha position were consistently less po-
tent than the corresponding cyclopentyl analogs (e.g., 36 or 39 vs.
31), in one case even exhibiting weak agonism at both human and
rat H4R (38). Evaluation of two of the most potent compounds from
Table 3 in H4R binding assays confirmed that these compounds
were potent at H4Rs of both human (29: pKi = 8.70 0.18; 30:
pKi = 8.82 0.23) and rat (29: pKi = 8.58 0.05; 30: pKi = 8.84
0.02).
Evaluation of three of the most potent compounds from Table 1
in H4R binding assays, performed as previously described,6 con-
firmed that these compounds indeed display moderately good
potencies at both the human H4R (1: pKi = 7.40 0.11; 9: pKi =
8.15 0.32; 15: pKi = 7.31 0.006) and rat H4R (1: pKi = 8.35
0.05; 9: pKi = 8.57 0.05; 15: 8.49 0.18).
We briefly investigated geminal di-substitution at the alpha po-
sition (compounds 16–19, Table 2), but these compounds were less
potent than the corresponding alpha-phenyl derivatives. The
H4 antagonists are known to reduce H4 agonist-induced itch in
mice.4 In order to assess the in vivo effects of the present classes of
H4 antagonists, the functional activity of compounds 1, 3, and 31
was determined in a mouse H4 FLIPR assay, then the compounds
were tested in a mouse model of H4 agonist (clobenpropit)-induced
scratching, according to the method of Bell et al.4d As can be seen in
Table 4, compound 31, which acts as a partial agonist in the rat H4
FLIPR assay, acts as an antagonist in the mouse H4 FLIPR assay. This
is a surprising result, as there is a fairly high homology (84%) be-

1904
J. R. Koenig et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1900–1904
Table 4
less in a mouse model of itch, thus demonstrating their in vivo effi-
cacies as H4R antagonists.
In vitro and in vivo results of selected H4 antagonists in a model of H4-agonist induced
itch
Compound
Mouse H4 FLIPR pKb SEMa
Blockade of scratching in
Acknowledgments
mouse, ED50, ipb
(lmol/kg)
1
3
31
7.34 0.11
7.90 0.05
7.82 0.07
10
3
1
The authors wish to thank Yau Yi Lau and the HT-ADME team
for log D determinations.
a
n P 2.
References and notes
b
At ED50, the responses of drug-treated animals were significantly (p <0.05)
different from responses of vehicle-treated animals.
1. (a) Engelhardt, H.; Smits, R. A.; Leurs, R.; Haaksma, E.; de Esch, I. J. P. Curr. Opin.
Drug Discovery Dev. 2009, 12, 628; (b) Akdis, C. A.; Estelle, F.; Simons, R. Eur. J.
Pharmacol. 2006, 533, 69; (c) de Esch, I. J. P.; Thurmond, R. L.; Jongejan, A.;
Leurs, R. Trends Pharmacol. Sci. 2005, 26, 462.
2. (a) Damaj, B. B.; Becerra, C. B.; Esber, H. J.; Wen, Y.; Maghazachi, A. A. J.
Immunol. 2007, 179, 7907; (b) Zhang, M.; Thurmond, R. L.; Dunford, P. J.
Pharmacol. Ther. 2007, 113, 594.
3. (a) Ling, P.; Ngo, K.; Nguyen, S.; Thurmond, R. L.; Edwards, J. P.; Karlsson, L.;
Fung-Leung, W. P. Br. J. Pharmacol. 2004, 142, 161; (b) Buckland, K. F.; Williams,
T. J.; Conroy, D. M. Br. J. Pharmacol. 2003, 140, 1117; (c) O’ Reilly, M.; Alpert, R.;
Jenkinson, S.; Gladue, R. P.; Foo, S.; Trim, S.; Peter, B.; Trevethick, M.; Fidock, M.
J. Recept. Signal Trans. 2002, 22(1–4), 431.
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Thurmond, R. J. Allergy Clin. Immunol. 2007, 119, 176; (b) Rees, J.; Murray, C. S.
Clin. Exp. Dermatol. 2005, 30, 471; (c) Thurmond, R. L.; Gelfand, E. W.; Dunford,
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Pharmacol 2004, 142, 374.
Table 5
Summary of in vitro potency for selected compounds at histamine H3 receptorsa
Compd Human H3 binding potency
Rat H3 binding potency
pKi SEM
pKi SEM
1
15
29
30
31
5.65 0.08
6.65 0.13
6.98 0.03
7.20 0.05
6.53 0.03
6.18 0.02
6.98 0.03
7.14 0.04
8.17 0.13
7.10 0.03
a
5. Dunford, P. J.; O’Donnell, N.; Riley, J. P.; Williams, K. N.; Karlsson, L.; Thurmond,
R. L. J. Immun. 2006, 176, 7062.
n P 2.
6. (a) Altenbach, R. J.; Adair, R. M.; Bettencourt, B. M.; Black, L. A.; Fix-Stenzel, S. R.;
Gopalakrishnan, S. M.; Hsieh, G. C.; Liu, H.; Marsh, K. C.; McPherson, M. J.; Milicic,
I.; Miller, T. R.; Vortherms, T. A.; Warrior, U.; Wetter, J. M.; Wishart, N.; Witte, D.
G.; Honore, P.; Esbenshade, T. A.; Hancock, A. A.; Brioni, J. D.; Cowart, M. C. J. Med.
Chem. 2008, 51, 6571; (b) Cowart, M. C.; Altenbach, R. J.; Liu, H.; Hsieh, G. C.;
Drizin, I.; Milicic, I.; Miller, T. R.; Witte, D. G.; Wishart, N.; Fix-Stenzel, S. R.;
McPherson, M. J.; Adair, R. M.; Wetter, J. M.; Bettencourt, B. M.; Marsh, K. C.;
Sullivan, J. P.; Honore, P.; Esbenshade, T. A.; Brioni, J. D. J. Med. Chem. 2008, 51,
6547; (c) Liu, H.; Altenbach, R. J.; Carr, T. L.; Chandran, P.; Hsieh, G. C.; Lewis, L. R.;
Manelli, A. M.; Milicic, I.; Marsh, K. C.; Miller, T. R.; Strakhova, M. I.; Vortherms, T.
A.; Wakefield, B. D.; Wetter, J. M.; Witte, D. G.; Honore, P.; Esbenshade, T. A.;
Brioni, J. D.; Cowart, M. D. J. Med. Chem. 2008, 51, 7094.
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2007, 563, 240; (b) Thurmond, R. L.; Desai, P. J.; Dunford, P. J.; Fung-Leung, W.
P.; Hofstra, C. L.; Jiang, W.; Nguyen, S.; Riley, J. P.; Sun, S.; Williams, K. N.;
Edwards, J. P.; Karlsson, L. J. Pharmacol. Exp. Ther. 2004, 309, 404.
8. Bartelt, R. J.; Weisleder, D.; Momany, F. A. Synthesis 2003, 117.
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121.
tween the rat H4R and the mouse H4R at the protein level.9 The
in vivo assay, meanwhile, revealed that all three of the compounds
studied were able to completely block itch responses after ip
administration and displayed ED50 values of 10 lmol/kg or less.
In addition to displaying H4 antagonist activity, a few of the
studied compounds also showed modest antagonist activity at
the H3 receptor (Table 5), with the binding assay being performed
as described previously.11 Both H4 and H3 receptor antagonists
have been proposed for the treatment of pain,1c,12 so the therapeu-
tic potential of combining these two activities in a single molecule
is intriguing. The data obtained for such compounds as 30 suggest
that achieving such dual activity may be possible.
In summary, we have designed three novel series of drug-like
H4R antagonists containing the 2-aminopyrimidine motif. In gen-
eral, such compounds with an alpha-spiro moiety are more potent
than the corresponding alpha-substituted or alpha-gem-disubsti-
tuted analogs. The best of these compounds display good in vitro
potency in both functional and binding assays. Finally, representa-
10. For example, comparison of log D values measured at pH 7.4—1: 1.73 versus
28: 2.20; 8:2.05 versus 30: 1.60.
11. Esbenshade, T. A.; Krueger, K. M.; Miller, T. R.; Kang, C. H.; Denny, L. I.; Witte, D.
G.; Yao, B. B.; Fox, G. B.; Faghih, R.; Bennani, Y. L.; Williams, M.; Hancock, A. A. J.
Pharmacol. Exp. Ther. 2003, 305, 887.
12. Alguacil, L. F.; Perez-Garcia, C. Curr. Drug Targets CNS Neurol. Disord. 2003, 2,
303.
tive members of these series show ED50 values of 10 lmol/kg or