Influence of imidazole replacement in different structural classes of histamine H3-receptor antagonists

Article abstract of DOI:10.1016/S0928-0987(01)00106-3

The reference compounds for histamine H₃-receptor antagonists carry as a common feature an imidazole moiety substituted in the 4-position. Very recently novel ligands lacking an imidazole ring have been described possessing a N-containing non-aromatic heterocycle instead. In this study we investigated whether imidazole replacement, favourably by a piperidine moiety, is generally applicable to different structural classes of reference compounds, e.g., thioperamide, carboperamide, clobenpropit, FUB 181, ciproxifan, etc. While replacement led to a loss of affinity for many of the compounds, it was successfully applied to some ether derivatives. The piperidine analogues of FUB 181 and ciproxifan, 3-(4-chlorophenyl)propyl 3-piperidinopropyl ether hydrogen oxalate (6) and cyclopropyl 4-(3-piperidinopropyloxy)phenyl methanone hydrogen maleate (7), almost maintained in vitro affinities, pK_i values of 7.8 and 8.4, respectively, and showed high potency in vivo after p.o. administration (ED₅₀ values of 1.6 and 0.18 mg/kg, respectively).

Full text of DOI:10.1016/S0928-0987(01)00106-3

European Journal of Pharmaceutical Sciences 13 (2001) 249–259
www.elsevier.nl/locate/ejps
Influence of imidazole replacement in different structural classes of
histamine H3-receptor antagonists
Galina Meiera, Joachim Apelta, Ulrich Reicherta, Sven Graßmanna, Xavier Ligneaub, Sigurd Elza,
Fabien Leurquinc, C. Robin Ganellinc, Jean-Charles Schwartzd, Walter Schunacka, Holger Starka,e ,
*
a
¨
¨
¨
Institut fur Pharmazie, Freie Universitat Berlin, Konigin-Luise-Strasse 214, 14195 Berlin, Germany
bLaboratoire Bioprojet, 30 Rue des Francs-Bourgois, 75003 Paris, France
cDepartment of Chemistry, Christopher Ingold Laboratories, University College London, 20 Gordon Street, London WC1H 0AJ, UK
d
´
´
´
Unite de Neurobiologie et Pharmacologie Moleculaire (U. 109), Centre Paul Broca de l’INSERM, 2ter Rue d’Alesia, 75014 Paris, France
e
¨
¨
Institut fur Pharmazeutische Chemie, Johann Wolfgang Goethe-Universitat, Marie-Curie-Strasse 9, 60439 Frankfurt, Germany
Received 8 November 2000; received in revised form 5 January 2001; accepted 14 January 2001
th
¨
Dedicated to Univ.-Prof. Dr. Dres. h.c.mult. H. Oelschlager, Jena, on occasion of his 80 Birthday
Abstract
The reference compounds for histamine H3-receptor antagonists carry as a common feature an imidazole moiety substituted in the
4-position.Very recently novel ligands lacking an imidazole ring have been described possessing a N-containing non-aromatic heterocycle
instead. In this study we investigated whether imidazole replacement, favourably by a piperidine moiety, is generally applicable to
different structural classes of reference compounds, e.g., thioperamide, carboperamide, clobenpropit, FUB 181, ciproxifan, etc. While
replacement led to a loss of affinity for many of the compounds, it was successfully applied to some ether derivatives. The piperidine
analogues of FUB 181 and ciproxifan, 3-(4-chlorophenyl)propyl 3-piperidinopropyl ether hydrogen oxalate (6) and cyclopropyl
4-(3-piperidinopropyloxy)phenyl methanone hydrogen maleate (7), almost maintained in vitro affinities, pKi values of 7.8 and 8.4,
respectively, and showed high potency in vivo after p.o. administration (ED50 values of 1.6 and 0.18 mg/kg, respectively).
Elsevier Science B.V. All rights reserved.
2001
Keywords: Histamine; H3 Receptor; Imidazole; Non-imidazole; Antagonist; Medicinal chemistry
1. Introduction
central nervous system (Stark et al., 1996a). Histamine
H3-receptor antagonists have not been introduced to
therapy, but potential therapeutic applications have been
proposed, e.g., memory and learning deficits (Miyazaki et
al., 1995; Blandina et al., 1996; Onodera et al., 1998),
epilepsy (Yokoyama et al., 1993, 1994), schizophrenia
(Schwarz and Arrang, in press), Alzheimer’s disease
(Panula et al., 1995; Morisset et al., 1996), and attention-
deficit hyperactivity disorder (ADHD) (Leurs et al., 1998).
In fact, compound GT 2331 (Ali et al., 1999) has entered
phase II clinical trials for the treatment of ADHD (Gliatech
Corporate Information).
The majority of histamine H3-receptor antagonists re-
ported in the literature to date contain a mono-substituted
4(5)-imidazole moiety. For potential therapeutic use, the
design of histamine H3-receptor ligands devoid of an
imidazole ring is desirable, as a means of providing
compounds with improved pharmacokinetic properties in
The third histamine receptor subtype (H3) has been
identified as a presynaptic autoreceptor located on his-
taminergic nerve endings inhibiting synthesis (Arrang et
al., 1987a) and release (Arrang et al., 1985) of histamine
upon activation. In addition, the histamine H3 receptor also
occurs as a heteroreceptor on non-histaminergic neurons
thereby interacting with a variety of different neurotrans-
mitter systems (Hill et al., 1997). Histamine plays an
important role in the regulation of different physiological
processes, e.g., arousal (Schwartz et al., 1991; Schwartz
and Arrang, in press), and may be implicated in
pathophysiological conditions and diseases affecting the
*Corresponding author. Tel.: 149-69-798 29302; fax: 149-60-798
29258.
E-mail address: h.stark@pharmchem.uni-frankfurt.de (H. Stark).
0928-0987/01/$ – see front matter
PII: S0928-0987(01)00106-3
2001 Elsevier Science B.V. All rights reserved.

250
G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
vivo (Alves-Rodrigues et al., 1996). The development of
non-imidazole histamine H3-receptor antagonists has been
pursued previously (Ganellin et al., 1991, 1998; Kiec-
Kononowicz et al., 1995a,b; Menge et al., 1998;
Walczynski et al., 1999a,b; Linney et al., 2000). Earlier
investigations focused on imidazole replacement by other
nitrogen-containing aromatic heterocycles within known
histamine H3-receptor antagonist structures. However, the
non-imidazole compounds obtained were consistently sub-
stantially inferior to the parent imidazole containing an-
tagonists (Ganellin et al., 1991; Kiec-Kononowicz et al.,
1995a,b). One of the first successful replacements of the
imidazole moiety of a known compound was reported in
1998 and resulted in the development of UCL 1972 (Ki5
39611 nM; ED5051.160.6 mg/kg p.o.) (Ganellin et al.,
1998). Very recent attempts were based on the structural
development of new compounds using low-affinity non-
imidazole histamine H3-receptor ligands as leads (Menge
et al., 1998; Walczynski et al., 1999a,b; Linney et al.,
2000). This led to the discovery of potent non-imidazole
histamine H3-receptor antagonists (Fig. 1), e.g., the ben-
zothiazole A (pA257.7660.13; pKi58.260.2) derived
from sabeluzole (Menge et al., 1998; Walczynski et al.,
1999a,b) and JB 98064 (pKB58.3860.10; pKi5
8.7060.12) derived from dimaprit (Linney et al., 2000).
All of these compounds possess a non-aromatic N-con-
taining heterocycle as a common feature, linked from the
nitrogen atom through an alkylene chain eventually to an
aryl group.
In an effort to reveal general patterns underlying imida-
zole exchangeability, we revived the original idea of
imidazole replacement within different structural classes of
known histamine H3-receptor antagonists. Compounds
containing pyrrolidine or piperidine moieties have been
shown to be potent non-imidazole histamine H3-receptor
antagonists (Ganellin et al., 1998; Linney et al., 2000).
Based on these findings, the piperidino group was chosen
as a general imidazole substitute for our investigation. In
order to assess further moieties suitable for replacement,
one piperidine-containing histamine H3-receptor ligand (5,
Table 1), found to be a potent antagonist in vitro and in
vivo, was further derivatized by replacement of the
piperidino group by azepane-, pyrrolidine-, or diethylamine
moieties, respectively.
pharmacological behaviour of known histamine H3-re-
ceptor antagonists in vitro and in vivo, thereby covering
various structural classes. In the present study we report
the synthesis and pharmacological evaluation of novel
non-imidazole histamine H3-receptor ligands derived from
already established histamine H3-receptor antagonists
(Schwartz et al., 2000). For progress in drug development,
imidazole replacement seems to be of special interest since
many related imidazole-containing compounds are known
to interact with the cytochrome P450 system (Halpert et al.,
1994) and other metabolic enzymes (Karjalainen et al.,
2000).
2. Materials and methods
2.1. Chemistry
2.1.1. General procedures
Melting points (mp) were determined on an Electrother-
¨
mal IA 9000 digital or a Buchi 512 apparatus and are
uncorrected. For all compounds 1H NMR spectra were
recorded on a Bruker AC 400 (400 MHz) spectrometer.
Chemical shifts are reported in ppm downfield from
1
internal trimethylsilane as reference. H NMR signals are
reported in order: Multiplicity (br, broad; s, singlet; d,
doublet; t, triplet; m, multiplet; *, exchangeable by D2O;
Cyhex, cyclohexyl; Ph, phenyl; Pip, piperidino; Pyr,
pyrrolidinyl), number of protons, and approximate cou-
pling constants in Hertz (Hz). Mass spectra were obtained
on a Finnigan MAT CH7A (70 eV, EI spectra) or a
Finnigan MAT CH5DF (FAB1 spectra). All FAB1 spectra
were recorded in Me2SO. Elemental analyses (C, H, N)
were measured on Perkin-Elmer 240 B or Perkin-Elmer
240 C instruments and are within 60.4% of the theoretical
values. Preparative, centrifugally accelerated, rotatory
chromatography was performed using a Chromatotron
7924T (Harrison Research) and glass rotors with 4 mm
layers of silica gel 60 F254 containing gypsum (Merck).
Column chromatography was carried out using silica gel
63–200 mm (Macherey, Nagel) or silica gel 40–63 mm
(Merck) for flash column chromatography. TLC was
carried out on silica gel F254 plates (Merck).
In an attempt to clarify principles and applicability of
imidazole replacement, we investigated the change in
2.1.2. Synthesis
Precursors I (Frankel et al., 1950), III (Doherty et al.,
1957), and IV (Clinton et al., 1949) were prepared as
described in the literature (Fig. 2). Precursor II was
prepared from piperidine and methyl 3-bromopropionate in
acetone with catalytic amounts of potassium iodide under
basic conditions (Fig. 2).
Thiourea derivative 1 (Table 1) was obtained by re-
action from both commercially available 1,49-bipiperidine
with cyclohexyl isothiocyanate. Amide 2 was synthesized
under Schotten–Baumann conditions according to the
procedure described by Stark et al. (1995) from commer-
Fig. 1. Non-imidazole histamine H3-receptor antagonists.

G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
251
both commercially available 1-bromo-3-phenylpropane and
1,3-propanediol to form 3-(3-phenylpropyloxy)propan-1-ol
according to Williamson’s protocol (Williamson, 1851),
conversion of the alcohol to the chloroalkane with thionyl
chloride and alkylation of the corresponding secondary
amines under basic conditions as mentioned before. Syn-
thetic procedures and analytical data for all final com-
pounds are provided below.
2.1.2.1. N-Cyclohexyl-l,49-bipiperidine-19-yl-thiocarbox-
amide hydrogen oxalate (1)
1,49-Bipiperidine (0.84 g, 5 mmol) and cyclohexyl
isothiocyanate (0.7 g, 5 mmol) were dissolved in dry Et2O
(30 ml). The solution was stirred at room temperature for 2
h. The precipitated product was filtered, washed with ether
and crystallized with oxalic acid from Et2O/EtOH: Yield
Fig. 2. Central building blocks and synthesis of compounds 3–11 and 13.
Footnotes: (a) LiAlH4, THF, room temperature, 3 h; (b) arylalkyl halide,
potassium carbonate, potassium iodide, EtOH, 608C, 8 h; (c) N-hydroxy-
4-chlorophenylacetamidine, sodium, MeOH, 08C→room temperature, 1 h;
reflux, 18 h; (d) Frankel et al. (1950); (e) methyl 3-bromopropionate,
potassium iodide, potassium carbonate, acetone, reflux, 4 h; (f) Clinton et
al. (1949); (g) Doherty et al. (1957); (h) 4-chlorobenzylthiourea, EtOH,
potassium iodide, reflux, 6 days; (i) NaH, toluene, room temperature, 12
h; arylalkyl halide or methanesulfonate, 15-crown-5, tetrabutylammonium
iodide, reflux, 4–17 h; (k) 4-cyclopropanecarbonylphenol, triphenylphos-
phine, diethylazodicarboxylate, THF, 08C→room temperature, 16 h; (l)
corresponding isocyanate, acetonitrile, reflux, 2–15 h.
1
70%; mp 2258C; H NMR (Me2SO-d6 ) d 7.34 (d, J57.7
Hz, 1H, NH), 4.76–4.79 (m, 2H, Pip9-2,6He), 4.16 (m, 1H,
Cyhex-1H), 3.30–3.33 (m, 1H, Pip9-4H), 3.08 (m, 4H,
Pip-2,6H2), 2.84–2.90 (m, 2H, Pip9-2,6Ha), 1.97–2.00 (m,
2H, Pip9-3,5He), 1.84 (m, 2H, Cyhex-2,6He), 1.72 (m, 6H,
Pip-3,5H2, Cyhex-3,5He), 1.45–1.60 (m, 6H, Pip-4H2,
Cyhex-4H2, Pip9-3,5Ha), 1.20–1.27 (m, 4H, Cyhex-
2,3,5,6Ha); FAB1-MS m/z (%) 310 (M1H1, 100). Anal.
(C17H31N3S?C2H2O4?0.25H2O) C, H, N.
2.1.2.2. 1-(1,49-Bipiperidine-19-yl)heptane-1-one hydro-
gen oxalate (2)
cially available 1,49-bipiperidine and n-heptanoyl chloride.
Compound 3 was obtained by reduction of the cyano
moiety of I (Frankel et al., 1950) with LiAlH4 according
to published procedures (Nystrom and Brown, 1948). The
clobenpropit analogue 4 was prepared from III in an
adapted preparation sequence described for the imidazole
equivalent (Van der Goot et al., 1992). Ethers 5 and 6 were
synthesized by Williamson reaction (Williamson, 1851)
from intermediate IV (Clinton et al., 1949) and the
corresponding commercially available arylalkyl halide or
the freshly prepared methanesulfonate, respectively. The
aromatic ether 7 was prepared from IV and commercially
available 4-cyclopropanecarbonylphenol by Mitsunobu
type reaction (Mitsunobu, 1981). Carbamates 8 and 9 were
conveniently derived from IV (Clinton et al., 1949) and
commercially available pentyl or phenyl isocyanate as
described for the imidazole analogues (Sasse et al., 1999;
Stark et al., 1996a,b). All reactants being commercially
available, compounds 10 and 11 were obtained by alkyla-
tion of piperidine with the corresponding 1-chloro-v-
phenylalkanes under basic conditions. The benzothiazole
derivative 12 was synthesized from both commercially
available 2-piperidinoethanamine and 2-chlorobenzo-
[d][1,3]thiazole by SNAr reaction. Oxadiazole 13 was
1,49-Bipiperidine (1.68 g, 10 mmol) was dissolved in
water (10 ml) and added dropwise to a solution of n-
heptanoyl chloride (0.74 g, 5 mmol) in dioxane (20 ml) at
08C. The mixture was allowed to warm to room tempera-
ture and stirred for 1 h. The solvent was removed under
reduced pressure. The residue was purified by column
chromatography (eluent: CH2Cl2 /MeOH, NH3-sat.; 95:5)
and crystallized with oxalic acid from Et2O/EtOH: Yield
1
55%; mp 1318C; H NMR (Me2SO-d6 ) d 4.49–4.52 (m,
1H, Pip9-2He), 3.97–4.00 (m, 1H, Pip9-6He), 3.29 (m, 1H,
Pip9-4H), 2.95–3.07 (m, 5H, Pip9-6Ha, Pip-2,6H2), 2.50
(m, 1H, Pip9-2Ha), 2.29 (t, J57.6 Hz, 2H, OCCH2),
1.97–1.99 (m, 2H, Pip9-3,5He), 1.72–1.74 (m, 4H, Pip-
3,5H2), 1.40–1.52 (m, 6H, Pip9-3,5Ha, Pip-4H2,
OCCH2CH2), 1.26 (m, 6H, OC(CH2)2(CH2)3), 0.86 (t,
J57.0 Hz, 3H, CH3); EI-MS m/z (%) 280 (M1, 10).
Anal. (C17H32N2O?C2H2O4) C, H, N.
2.1.2.3. 5-Piperidinopentanamine dihydrochloride (3)
Precursor I (4.17 g, 25.1 mmol) was dissolved in dry
THF (10 ml). A suspension of LiAlH4 (1.5 eq) in dry THF
(20 ml) was added at 08C. The mixture was stirred at room
temperature for 3 h and then carefully treated with EtOH.
A saturated aqueous solution of potassium sodium tartrate
(10 ml) and aqueous sodium hydroxide solution (2 M, 10
ml) were added consecutively. The precipitate was filtered
and the organic layer separated, dried (MgSO4), and
synthesized
from
II
and
N-hydroxy-4-chloro-
phenylacetamidine in alkaline solution in an adapted
preparation sequence according to Clitherow et al. (1996).
Ethers 14, 15, and 16 were obtained in three steps from

252
G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
removed under reduced pressure. The residue was purified
by column chromatography (eluent: CH2Cl2 /MeOH, NH3-
sat.; 90:10). The final product was crystallized with HCl
(5–6 M solution in 2-propanol) from Et2O/2-propanol:
(br, 2H, Pip-4H2); EI-MS m/z (%) 261 (M1, 3). Anal.
(C17H27NO?C2H2O4) C, H, N.
2.1.2.6. 3-(4-Chlorophenyl)propyl
ether hydrogen oxalate (6)
3-piperidinopropyl
1
Yield 50%; mp 187–1908C; H NMR (Me2SO-d6 ) d 10.50
(s*, 1H, Pip-NH1), 8.05 (s*, 3H, NH13 ), 3.36–3.39 (m,
2H, Pip-2,6He), 2.91–2.97 (m, 2H, CH2NH2), 2.75–2.85
(m, 4H, Pip-2,6Ha, Pip-NCH2), 1.68–1.87 (m, 7H, Pip-
NCH2CH2, CH2CH2NH2, Pip-3,4,5He), 1.55–1.62 (m, 2H,
CH2(CH2)2NH2), 1.30–1.41 (m, 3H, Pip-3,4,5Ha), EI-MS
m/z (%) 170 (M1, 4). Anal. (C10H22N2?2HCl?0.5H2O) C,
H, N.
Synthetic procedure was performed as described for 5
starting with IV and 3-(4-chlorophenyl)propyl methane-
sulfonate, except the reaction mixture was heated under
reflux for 17 h. For purification the reaction mixture was
carefully poured into water (50 ml) and extracted with
Et2O. The combined organic layers were washed with
water and dried (MgSO4). The solvent was removed under
reduced pressure and the crude product purified by column
chromatography (first eluent: petroleum ether/ethyl ace-
tate; 3:1; second eluent: CH2Cl2 /MeOH, NH3-sat.; 90:10).
The final product was crystallized with oxalic acid from
2.1.2.4. N-(4-Chlorobenzyl)-S-(3-piperidino-
propyl)isothiourea dihydrochloride (4)
1
Precursor III (1.97 g, 10 mmol) and 4-chloro-
benzylthiourea (Van der Goot et al., 1992) (1.98 g, 10
mmol) were dissolved in dry EtOH (30 ml). Catalytic
amounts of potassium iodide were added. The mixture was
then heated under reflux for 6 days. The solvent was
removed under reduced pressure and the residue purified
by column chromatography (eluent: CH2Cl2 /MeOH; 95:5
to 90:10). The product was crystallized with HCl (5–6 M
solution in 2-propanol) from Et2O/2-propanol: Yield 8%;
Et2O/MeOH: Yield 56%; mp 1488C; H NMR (Me2SO-
d6 ) d 7.32–7.34 (m, 2H, Ph-3,5H), 7.22–7.24 (m, 2H,
Ph-2,6H), 3.40 (t, J56.0 Hz, 2H, N???CH2O), 3.35 (t,
J56.4 Hz, 2H, OCH2???Ph), 3.08 (br, 4H, Pip-2,6H2),
2.97–3.01 (m, 2H, Pip-NCH2), 2.61 (t, J57.6 Hz, 2H,
PhCH2), 1.85–1.92 (m, 2H, Pip-NCH2CH2), 1.74–1.81
(m, 2H, PhCH2CH2), 1.70–1.73 (m, 4H, Pip-3,5H2), 1.52
(br, 2H, Pip-4H2); EI-MS m/z (%) 295 (M1, 4). Anal.
(C17H26ClNO?C2H2O4) C, H, N.
1
mp 1048C; H NMR (Me2SO-d6 ) d 11.03 (br*, 1H, NH),
10.31 (br*, 1H, NH), 9.57 (br*, 2H, NH12 ), 7.46 (m, 4H,
Ph-2,3,5,6H), 4.62 (s, 2H, PhCH2), 3.38 (t, J56.9 Hz, 2H,
SCH2), 3.35 (m, 2H, Pip-2,6He), 3.04 (m, 2H, Pip-NCH2),
2.82 (m, 2H, Pip-2,6Ha), 2.06 (m, 2H, Pip-NCH2CH2),
1.68–1.77 (m, 5H, Pip-4He, Pip-3,5H2), 1.09 (m, 1H,
Pip-4Ha); FAB1-MS m/z (%) 326 (M-H1, 92). Anal.
(C16H24ClN3S?2HCl?H2O) C, H, N.
2.1.2.7. Cyclopropyl
methanone hydrogen maleate (7)
4-(3-piperidinopropyloxy)phenyl
A solution of 4-cyclopropanecarbonylphenol (0.25 g, 1.5
mmol), precursor IV (0.22 g, 1.5 mmol), and tri-
phenylphosphine (0.45 g, 1.7 mmol) in dry THF (20 ml)
was stirred and cooled to 08C under nitrogen. A solution of
diethylazodicarboxylate (0.3 g, 1.7 mmol) in dry THF (20
ml) was added dropwise and the resulting mixture was
allowed to warm to room temperature and stirred under
nitrogen for 16 h. The solvent was removed under reduced
pressure and the residue taken up in ethyl acetate. The
product was extracted with HCl (2N solution) and the
aqueous layer was neutralized with aqueous sodium hy-
droxide. The free base was extracted with CH2Cl2 and the
organic extracts were dried (MgSO4) and the solvent
removed under reduced pressure. The crude product was
purified by column chromatography (eluent: Et2O con-
taining 1% triethylamine). The final product was crys-
tallized either with oxalic acid or maleic acid from EtOH
(analytical data is given for the maleate): Yield 33%; mp
2.1.2.5. 3-Phenylpropyl 3-piperidinopropyl ether hydro-
gen oxalate (5)
Compound IV (2.86 g, 20 mmol) and NaH (suspended
in mineral oil, v560%, 1.00 g, 25 mmol) in dry toluene
(30 ml) were stirred overnight at ambient temperature
under an argon atmosphere. 1-Bromo-3-phenylpropane
(4.38 g, 22 mmol), catalytic amounts of 15-crown-5 and
tetrabutylammonium iodide were added to the solution via
argon counter-current. The reaction mixture was heated
under reflux for 4 h, cooled, filtered, and the solvent
removed under reduced pressure. The residue was purified
by column chromatography (eluent: CH2Cl2 /MeOH, NH3-
sat.; 90:10) and crystallized with oxalic acid from Et2O/
EtOH: Yield 26%; mp 1258C; 1H NMR (Me2SO-d6 ) d
7.23–7.30 (m, 2H, Ph-3,5H), 7.15–7.20 (m, 3H, Ph-
2,4,6H), 3.40 (t, J55.9 Hz, 2H, N???CH2O), 3.36 (t,
J56.4 Hz, 2H, OCH2???Ph), 3.08 (br, 4H, Pip-2,6H2),
2.98–3.02 (m, 2H, Pip-NCH2), 2.61 (t, J57.7 Hz, 2H,
PhCH2), 1.86–1.96 (m, 2H, Pip-NCH2CH2), 1.75–1.82
(m, 2H, PhCH2CH2), 1.71–1.73 (m, 4H, Pip-3,5H2), 1.51
1
928C; H NMR (CF3COOD) d 8.12 (m, 2H, Phe-2,6H),
7.05 (m, 2H, Phe-3,5H), 6.38 (s, 2H, maleic acid), 4.36–
4.39 (t, J55.4 Hz, 2H, OCH2), 3.83–3.90 (m, 2H, Pip-
2,6He), 3.52–3.57 (Pip-NCH2), 3.02–3.12 (m, 2H, Pip-
2,6Ha), 2.80–2.86 (m, 1H, cyclopropyl-1H), 2.42–2.48 (m,
2H, Pip-NCH2CH2), 1.90–2.20 (m, 5H, Pip-3,5H2, Pip-
4Ha), 1.45–1.49 (m, 2H, cyclopropyl-CH2), 1.34–1.38 (m,
2H, cyclopropyl-CH2); EI-MS m/z (%) 287 (M1, 3).
Anal. (C18H25NO2?C4H4O4?0.75 H2O) C, H, N.

G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
253
2.1.2.8. N-Pentyl-3-piperidinopropylcarbamate
chloride (8)
hydro-
Pip-4H2), 1.29–1.30 (m, 4H, Ph(CH2)2(CH2 )2 ); EI-MS
m/z (%) 245 (M1, 8). Anal. (C17H27N?C2H2O4) C, H, N.
Compound IV?HCl (0.79 g, 4.4 mmol) and pentyl
isocyanate (0.58 ml, 4.4 mmol) were heated under reflux
in dry acetonitrile (30 ml) under argon atmosphere for 15
h. The mixture was brought to room temperature. After
evaporation of the solvent, the residue was purified by
column chromatography (eluent: CH2Cl2 /MeOH, NH3-
sat.; 90:10). The final product was crystallized with HCl
(5–6 M solution in 2-propanol) from 2-propanol: Yield
26%; mp 898C; 1H NMR (Me2SO-d6 ) d 10.53 (s*, 1H,
Pip-NH1), 7.13 (t, J55.4 Hz, 1H, NH), 3.98 (t, J56.3
Hz, OCH2), 3.36–3.39 (m, 2H, Pip-2,6He), 2.93–3.03 (m,
4H, NCH2, Pip-NCH2), 2.78–2.87 (m, 2H, Pip-2,6Ha),
1.98–2.05 (m, 2H, CH2CH2O), 1.67–1.86 (m, 5H, Pip-
3,5H2, -4He), 1.36–1.42 (m, 3H, NHCH2CH2, Pip-4Ha),
1.21–1.32 (m, 4H, CH2CH2CH3), 0.86 (t, J57.0 Hz, 3H,
CH3); EI-MS m/z (%) 256 (M1, 3). Anal. (C14H28N2O2?
HCl?0.5H2O) C, H, N.
2.1.2.11. 1-(4-Phenylbutyl)piperidine hydrogen oxalate
(11)
Synthetic procedure and purification as described for 10
starting with piperidine and 1-chloro-4-phenylbutane. The
desired product was crystallized with oxalic acid from
1
2-propanol: Yield 69%; mp 1508C; H NMR (Me2SO-d6 )
d 7.16–7.30 (m, 5H, Ph-2,3,4,5,6H), 2.92 (m, 6H, Pip-
NCH2, Pip-2,6H2), 2.58–2.62 (t, J57.3 Hz, 2H, PhCH2),
1.51–1.72 (m, 10H, Pip-NCH2(CH2 )2, Pip-3,4,5H2); EI-
MS m/z (%) 217 (M1, 6). Anal. (C15H23?C2H2O4) C, H,
N.
2.1.2.12. N-(2-Piperidinoethyl)benzo[d][1,3]thiazole-2-
amine dihydrochloride (12)
2-Piperidinoethanamine (1.29 g, 10 mmol) and 2-chloro-
benzo[d][1,3]thiazole (1.87 g, 11 mmol) were dissolved in
EtOH (50 ml). Triethylamine (3.04 g, 30 mmol) was
added to the solution. The mixture was heated under reflux
for 18 h. After cooling, the solvent was evaporated. HCl
(5–6 M solution in 2-propanol) was added to the crude
product. The resulting crystals were recrystallized from
2.1.2.9. N-Phenyl-3-piperidinopropylcarbamate
chloride (9)
hydro-
Synthetic procedure and purification as described for 8
starting with IV?HCl and phenyl isocyanate, except the
reaction mixture was heated under reflux for 2 h. The final
product was recrystallized from MeOH: Yield 65%; mp
1
MeOH: Yield 88%; mp 2258C; H NMR (Me2SO-d6 ) d
10.54 (br*, 1H, benzothiazolyl-2-NH), 9.87 (br*, 1H, Pip-
NH1), 7.82 (d, J57.8 Hz, 1H, benzothiazolyl-4H), 7.55
(d, J58.0 Hz, 1H, benzothiazolyl-7H), 7.35–7.39 (m, 1H,
benzothiazolyl-6H), 7.18–7.22 (m, 1H, benzothiazolyl-
5H), 3.97 (br, 2H, thiazolyl-NHCH2 ), 3.52 (br, 2H, Pip-
2,6He), 3.37 (br, 2H, Pip-NCH2), 2.97 (br, 2H, Pip-
2,6Ha), 1.81 (br, 4H, Pip-3,5H2), 1.70 (br, 1H, Pip-4He),
1.39 (br, 1H, Pip-4Ha); EI-MS m/z (%) 261 (M1, 1).
Anal. (C14H19N3S?2HCl?0.25H2O) C, H, N.
1
1708C; H NMR (Me2SO-d6 ) d 10.47 (s*, 1H, Pip-NH1),
9.69 (s, 1H, Ph-NH), 7.45–7.47 (m, 2H, Ph-2,6H), 7.26–
7.30 (m, 2H, Ph-3,5H), 6.97–7.01 (m, 1H, Ph-4H), 4.15 (t,
J56.3 Hz, OCH2), 3.39–3.42 (m, 2H, Pip-2,6He), 3.06–
3.10 (m, 2H, Pip-NCH2), 2.83–2.86 (m, 2H, Pip-2,6Ha),
2.07–2.14 (m, 2H, CH2 CH2O), 1.68–1.82 (m, 5H, Pip-
3,5H2, -4He), 1.36–1.42 (m, 1H, Pip-4Ha); EI-MS m/z
(%) 262 (M1, 5). Anal. (C15H22N2O2?HCl?0.1H2O) C, H,
N.
2.1.2.13. 3-(4-Chlorobenzyl)-5-(2-piperidinoethyl)-1,2,4-
oxadiazole hydrogen oxalate (13)
2.1.2.10. 1-(6-Phenylhexyl)piperidine hydrogen oxalate
(10)
Intermediate II (1.0 g, 6 mmol) and N-hydroxy-4-
chlorophenylacetamidine (0.74 g, 4 mmol) were dissolved
in dry MeOH (15 ml) under argon atmosphere. The
solution was cooled to 08C. A freshly prepared solution of
sodium (0.12 g, 5 mmol) in dry MeOH (20 ml) was slowly
added to the reaction mixture. Stirring was continued for 1
h at ambient temperature after which time the solution was
refluxed for 18 h. The mixture was cooled and the solvent
removed under reduced pressure. The residue was sus-
pended in DMF (40 ml) and the suspension stirred for 6 h
at 808C. The solvent was evaporated and the crude product
suspended in water and extracted with CH2Cl2. The
organic layers were combined, washed with water and
dried (MgSO4). The solvent was removed under reduced
pressure and the residue purified by rotatory chromatog-
raphy (eluent: CH2Cl2, NH3-atmosphere). The final prod-
uct was crystallized with oxalic acid from Et2O/EtOH:
Piperidine (4.7 g, 55 mmol), 1-chloro-6-phenylhexane
(5.39 g, 27.5 mmol), potassium carbonate (7.7 g, 55
mmol) and catalytic amounts of potassium iodide were
heated for 8 h at 608C in a mixture of EtOH (25 ml) and
water (5 ml). The solvent was evaporated. The residue was
suspended in water and extracted with CH2Cl2. The
organic layers were combined, dried (MgSO4), and the
solvent was removed under reduced pressure. The crude
product was purified by column chromatography (eluent:
CH2Cl2 /MeOH, NH3-sat.; 90:10) and the final product
crystallized with oxalic acid from Et2O/EtOH: Yield 76%;
mp 1528C; 1H NMR (Me2SO-d6 ) d 7.25–7.29 (m, 2H,
Ph-3,5H), 7.19 (m, 3H, Ph-2,4,6H), 3.05 (m, 4H, Pip-
NCH2, Pip-2,6He), 2.92 (m, 2H, Pip-2,6Ha), 2.56 (t, J5
7.6 Hz, 2H, PhCH2), 1.70–1.72 (m, 4H, Pip-NCH2CH2,
Pip-3,5He), 1.53–1.61 (m, 6H, PhCH2CH2, Pip-3,5Ha,
1
Yield 5%; mp 1528C; H NMR (Me2SO-d6 ) d 7.38–7.40

254
G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
(m, 2H, Ph-3,5H), 7.31–7.34 (m, 2H, Ph-2,6H), 4.09 (s,
2H, PhCH2), 3.29–3.34 (m, 4H, oxadiazolyl-CH2, Pip-
2,6He), 2.98 (br, 4H, Pip-NCH2, Pip-2,6Ha), 1.65 (br, 4H,
Pip-3,5H2), 1.48 (br, 2H, Pip-4H2); FAB1-MS m/z (%)
306 (M1H1, 68). Anal. (C16H20ClN3O?C2H2O4) C, H,
N.
3H, Ph-2,4,6H), 3.42 (t, J55.9 Hz, 2H, N???CH2O), 3.37
(t, J56.4 Hz, 2H, Ph . . . CH2O), 3.01–3.10 (m, 6H,
CH2N(CH2 CH3)2), 2.61 (t, J57.7 Hz, 2H, PhCH2), 1.77–
1.87 (m, 4H, NCH2CH2, PhCH2CH2), 1.17 (t, J57.2 Hz,
6H, (CH3)2); EI-MS m/z (%) 249 (M1, 2). Anal.
(C16H27NO?C2H2O4) C, H, N.
2.2. Pharmacology
2.1.2.14. 3-(Azepane-1-yl)propyl 3-phenylpropyl ether hy-
drogen oxalate (14)
2.2.1. General methods
Synthetic procedure was performed as described for 10
starting with 1-chloro-3-(3-phenylpropyloxy)propane and
hexamethyleneimine. The crude product was purified by
flash column chromatography (eluent: Et2O/petroleum
ether/triethylamine; 66:33:1) to afford the final product as
a light yellow oil, which was crystallized with oxalic acid
from Et2O/EtOH: Yield 58%; mp 1058C; 1H NMR
(CF3COOD) d 7.30–7.34 (m, 2H, Ph-3,5H), 7.20–7.24 (m,
3H, Ph-2,4,6H), 3.87 (t, J55.6 Hz, 2H, azepanyl-N???
CH2O), 3.74 (t, J56.8 Hz, 2H, Ph???CH2O), 3.59–3.65
(m, 2H, azepanyl-2,7He), 3.37–3.42 (m, 2H, azepanyl-
NCH2), 3.20–3.32 (m, 2H, azepanyl-2,7Ha), 2.77 (t, J5
7.4 Hz, 2H, PhCH2), 2.13–2.23 (m, 2H, azepanyl-
NCH2CH2), 2.05–2.12 (m, 4H, azepanyl-3,6He,
PhCH2CH2), 1.89–1.99 (m, 2H, azepanyl-3,6Ha), 1.84 (br,
4H, azepanyl-4,5H2); EI-MS m/z (%) 275 (M1, 3). Anal.
(C18H29NO?C2H2O4?0.25H2O) C, H, N.
2.2.1.1. Histamine H3-receptor antagonist activity on
guinea pig ileum
Histamine H3-receptor antagonist potency was deter-
mined by concentration-dependent inhibition of (R)-a-
methylhistamine-induced relaxation of field-stimulated iso-
lated guinea pig ileum segments (longitudinal muscle with
adhering plexus myentericus) in the presence of the
antagonist according to Vollinga et al. (1992) and Ligneau
et al. (1994). Each experiment was performed at least in
triplicate. Data are presented as mean (S.E.M.#0.15).
Longitudinal muscle strips were prepared from the small
intestine, 20–50 cm proximal to the ileocecal valve. The
muscle strips were mounted between two platinum elec-
trodes (4 mm apart) in 20 ml of Krebs buffer, containing 1
mM mepyramine, connected to an isometric transducer,
continuously gassed with oxygen containing 5% CO2 at
378C. After equilibration of the muscle segments for 1 h
accompanied by washing every 10 min, they were stimu-
lated continuously with rectangular pulses of 15 V and 0.5
ms at a frequency of 0.1 Hz. After 30 min of stimulation, a
cumulative concentration–response curve to (R)-a-
methylhistamine was recorded. Subsequently the prepara-
tions were washed three times every 10 min without
stimulation. The antagonist was incubated 20–30 min
before redetermination of the concentration–response
curve of (R)-a-methylhistamine (Schlicker et al., 1994).
The new antagonists were tested at concentrations that did
not block ileal muscarinic M3 receptors (data not shown).
2.1.2.15. 3-Phenylpropyl 3-(pyrrolidine-1-yl)propyl ether
hydrogen oxalate (15)
Synthetic procedure was performed as described for 10
starting with 1-chloro-3-(3-phenylpropyloxy)propane and
pyrrolidine. The crude product was purified by flash
column chromatography (eluent: Et2O/petroleum ether/
triethylamine; 66:33:1) to afford the final product as a light
yellow oil, which was crystallized with oxalic acid from
Et2O/EtOH: Yield 54%; mp 1068C; 1H NMR
(CF3COOD) d 7.30–7.34 (m, 2H, Ph-3,5H), 7.20–7.24 (m,
3H, Ph-2,4,6H), 3.84–3.86 (m, 4H, Pyr-N???CH2O, Pyr-
2,5He), 3.74 (t, J56.7 Hz, 2H, Phe???CH2O), 3.39–3.44
(m, 2H, Pyr-NCH2), 3.10–3.16 (m, 2H, Pyr-2,5Ha), 2.75
(t, J57.4 Hz, 2H, PhCH2), 2.25–2.34 (m, 2H, Pyr-3,4He),
2.15–2.22 (m, 4H, Pyr-NCH2CH2, Pyr-3,4Ha), 2.03–2.10
(m, 2H, PhCH2CH2); EI-MS m/z (%) 247 (M1, 3). Anal.
(C16H25NO?C2H2O4) C, H, N.
2.2.1.2. Histamine H3-receptor assay on synaptosomes of
rat cerebral cortex
Compounds were examined for their antagonist activity
using an assay where K1-evoked depolarization induces
[3H]histamine release from rat synaptosomes as described
by Garbarg et al. (1992). Each experiment was performed
at least in triplicate. Data are presented as mean (S.E.M.
#0.2). Synaptosomal preparation was obtained by the
method of Whittaker (1966) and preincubated with L-
[3H]histidine (0.4 mM) at 378C for 30 min in a modified
Krebs-Ringer buffer. The synaptosomes were washed
thoroughly, transferred into a fresh 2 mM K1 Krebs-
Ringer solution supplemented with 2 mM or 30 mM K1
(final concentration) and preconditioned for 2 min. Com-
pounds and 1 mM histamine were added 5 min before
depolarization was induced. Incubation was interrupted by
2.1.2.16. 3-(Diethylamino)propyl 3-phenylpropyl ether hy-
drogen oxalate (16)
Synthetic procedure was performed as described for 10
starting with 3-(3-phenylpropyloxy)-1-propylchloride and
diethylamine. The crude product was purified by flash
column chromatography (eluent: Et2O/petroleum ether/
triethylamine; 66:33:1) to afford the final product as a
colourless oil, which was crystallized with oxalic acid
from Et2O/EtOH: Yield: 47%; mp 808C; 1H NMR
(Me2SO-d6 ) d 7.26–7.29 (m, 2H, Ph-3,5H), 7.15–7.20 (m,

G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
255
rapid centrifugation. [3H]Histamine levels were measured
by liquid scintillation spectrometry (Garbarg et al., 1992).
pKi values were calculated based on the Cheng–Prusoff
equation (Cheng and Prusoff, 1973).
hance histamine release in the brain (ED50526 mg/kg
p.o.) (Ligneau et al., 1998). As with the parent drug, 4 did
not exhibit activity in vivo. In vitro, however, some of the
affinity was maintained: 4 is a histamine H3-receptor
antagonist of moderate affinity on guinea pig ileum (pA25
7.4).
2.2.1.3. Histamine H3-receptor antagonist potency in vivo
in the mouse
Imidazole-containing aliphatic ethers of the 3-
(phenylpropyloxy)propyl-type (Stark et al., 1998a,b), have
proven to be potent and selective histamine H3-receptor
antagonists in vitro and in vivo. Previous efforts to develop
non-imidazole histamine H3-receptor antagonists for this
structural type, e.g., by replacing the imidazole group with
pyrazole, azine and diazine moieties (Kiec-Kononowicz et
al., 1995a,b) were if any of limited success. In contrast to
the previous replacements, compounds 5 (pA258.1, pKi5
7.8) and 6 (pA258.3, pKi57.8) displayed equipotent
affinity in vitro with respect to the parent compounds FUB
153 and FUB 181 (Stark et al., 1998a,b). In accordance
with this finding, 5 and 6 were also potent antagonists in
vivo (Table 1). Schild analysis for the interaction of 6 and
the histamine H3-receptor agonist (R)-a-methylhistamine
is shown in Fig. 3.
In vivo testing was performed after oral administration
to Swiss mice according to Garbarg et al. (1992). Brain
histaminergic neuronal activity was assessed by measuring
the main metabolite of histamine, Nt-methylhistamine.
Mice were fasted for 24 h before p.o. treatment. Animals
were decapitated 90 min after treatment, and the cerebral
cortex was isolated. The cerebral cortex was homogenized
in 10 vol of ice-cold perchloric acid (0.4 M). The Nt-
methylhistamine level was measured by radioimmunoassay
(Garbarg et al., 1989). By treatment with 3 mg/kg
ciproxifan the maximal increase in Nt-methylhistamine
level was obtained (Ligneau et al., 1998) and related to the
level reached with the administered drug. The ED50 value
was calculated as mean with S.E.M. (Parker and Waud,
1971; Waud and Parker, 1971).
Potent non-imidazole histamine H3-receptor antagonists
of the aromatic ether type have been reported previously
(Ganellin et al., 1998). Recently, ciproxifan was described
as a highly potent and selective antagonist in vitro (pKi5
9.3) also displaying high efficacy in vivo (ED5050.14
mg/kg p.o.) (Ligneau et al., 1998; Stark et al., 2000). As
with the aliphatic ethers 5 and 6, the aromatic ether 7 also
maintained high potency in vitro and in vivo (Table 1).
The ciproxifan analogue 7 shows only slightly decreased
affinity in vitro (pKi58.4) compared to ciproxifan (Lig-
neau et al., 1998; Stark et al., 2000) and is highly potent in
vivo. These results need to be considered important, since
the non-imidazole histamine H3-receptor antagonist 7 in
vitro resembles the activity of the reference antagonist
thioperamide. In addition, the non-imidazole 7 surmounts
the potency of thioperamide in vivo and is not significantly
different from the efficacy of the parent imidazole-con-
taining compound ciproxifan.
Whereas the aliphatic and aromatic ethers investigated
led to a successful imidazole replacement, aliphatic and
aromatic carbamates 8 and 9, which were also derived
from potent imidazole-containing antagonists (Sasse et al.,
1999; Stark et al., 1996a,b), revealed only poor activity in
vitro and no detectable potency in vivo. This is also true
for the highly lipophilic phenylalkanes 10 and 11 as well
as for the benzothiazole derivative 12, although 11 still
exhibits low potency in vivo. Recently, aromatase inhibi-
tion by the imidazole analogue of 11 has been described,
emphasizing the importance of developing non-imidazole
compounds (Karjalainen et al., 2000). The oxadiazole
derivative 13 showed moderate affinity in vitro being
another interesting lead for further optimization. Although
imidazole replacement also led to a decrease in affinity
when compared to the parent imidazole compound
3. Results and discussion
Compound structures and pharmacological results are
shown in Tables 1 and 2. The affinities of the novel
compounds were determined in vitro on isolated segments
of the guinea pig ileum (Ligneau et al., 1994; Schlicker
and Marr, 1996) and on synaptosomes of rat cerebral
cortex (Garbarg et al., 1992). Results obtained on the two
tests were generally in good agreement, slight differences
for some compounds being potentially attributable to other
experimental errors in two largely distinct settings of
amino acid sequence differences between the guinea pig
and the rat (Ligneau et al., 2000; Tardivel-Lacombe et al.,
2000). In vivo testing was performed on mice brain after
oral administration (Garbarg et al., 1992).
The starting point for this study was the standard
reference antagonist thioperamide, a ligand of high in vitro
affinity (pKi58.4, Arrang et al., 1987b; pA258.3,
Clitherow et al., 1996) as well as a potent antagonist in
vivo (ED5051.0 mg/kg p.o., Ligneau et al., 1998).
Replacement of the imidazole group by a piperidine
moiety leading to 1 was not tolerated and led to a dramatic
loss of potency in vitro and in vivo. The same observation
was made for the carboperamide (Ligneau et al., 1994)
analogue 2 and the impentamine (Vollinga et al., 1995)
analogue 3, with 2 still showing weak antagonist affinity in
vitro.
Another interesting candidate for imidazole replacement
was the isothiourea derivative clobenpropit, a histamine
H3-receptor antagonist of high affinity in vitro (pKB59.9,
Clitherow et al., 1996; pKi59.2, Ligneau et al., 1994). In
vivo, however, clearly higher doses are required to en-

256
G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
Table 1
Histamine H3-receptor antagonist potencies in vitro and in vivo of known imidazole-containing ligands versus novel non-imidazole analogues:
Pip, piperidino; Im, 1H-imidazol-4-yl
No.
R-X
R
In vitro
In vivo
a
X
pA2
pKib
ED506S.E.M. (mg/kg)c
1
Pip
5.6
.10
Im (Thioperamide)
8.3d
8.4e
1.060.5f
2
3
Pip
6.4
.10
Im (Carboperamide)
7.7g
6.4
3.960.8g
Pip
6.0
.10
Im (Impentamine)
8.4h
4
5
Pip
7.4
6.3
.10
Im (Clobenpropit)
9.9d
9.2g
2667f
Pip
8.1
7.3
7.8
3.761.0
1.460.6i
Im (FUB 153)
7.8i
6
Pip
8.3
7.8
1.660.9
0.860.2i
Im (FUB 181)
8.2i
7.9i
7
Pip
7.9
8.4
0.1860.06
0.1460.03j
Im (Ciproxifan)
8.4j
9.3j
8
Pip
6.3
.10
Im (FUB 305)
7.3k
8.1k
0.6960.37k
9
Pip
6.2
6.8
6.6
.10
Im (FUB 138)
7.9f
1.360.6f
10
11
12
Pip
6.5
6.7
.10
Im (FUB 427)
7.7l
7.1m
1.060.3m
Pip
5.7
20
Im (FUB 349)
7.5l
7.3m
7.7n
2.260.7m
Pip
Im
6.6
.10
13
Pip
Im
7.2
6.9
|20
8.1d
8.2d
a Functional H3-receptor assay on guinea pig ileum (Ligneau et al., 1994; Schlicker et al., 1994). b Functional H3-receptor assay on rat cerebral cortex
(Garbarg et al., 1992). c H3-receptor screening after p.o. administration to mice (Garbarg et al., 1992). d pKB value (Clitherow et al., 1996). e Arrang et al.
(1987a). f Stark et al. (1996a,b). g Ligneau et al. (1998). h Vollinga et al. (1995). i Stark et al. (1998a). j Ligneau et al. (1998); Stark et al. (2000). k Sasse et al.
(1999). l De Esch et al. (1999). m Stark et al. (1998b). n Plazzi et al. (1995).

G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
257
Table 2
In vitro and in vivo histamine H3-receptor antagonist potencies of different amino groups in aliphatic ethers, related to 5
a
b
No.
R
pA2
ED506S.E.M.
(mg/kg)
14
15
16
7.8
7.8
6.5
2.360.7
5.562.0
.10
a Functional H3-receptor assay on guinea pig ileum (Ligneau et al., 1994; Schlicker et al., 1994).
b H3-receptor screening after p.o. administration to mice (Garbarg et al., 1992).
(Clitherow et al., 1996), the principle investigated seems
also applicable to this class of histamine H3-receptor
antagonists.
In an attempt to further elucidate moieties suitable for
replacement in the case of 5 and 6, other nitrogen-con-
taining groups were attached to the 3-(phenylpropyl-
oxy)propyl structure resulting in compounds 14, 15, and
16 (Table 2).
compound to be both less lipophilic and less flexible than 5
and 14, it was equipotent in vitro. Unfortunately, the in
vivo potency of 15 was slightly diminished compared to 5
and 14. When a lipophilic and flexible diethylamino moiety
was introduced (16), in vitro and in vivo activity decreased
significantly. Similar results were obtained before in a
series of aromatic ethers (Ganellin et al., 1998).
Enlargement from a six- to a more lipophilic and
flexible seven-membered nitrogen containing ring system
led to 14. This structural change, however, hardly affected
in vitro and in vivo activity compared to 5. A smaller ring
size was achieved by introduction of a pyrrolidine moiety
(15). Although the five-membered ring system causes the
4. Conclusion
In this study non-imidazole histamine H3-receptor an-
tagonists were developed from known H3-receptor ligands
of different structural classes by imidazole replacement.
Exchange of the imidazole ring by a piperidine moiety was
not generally feasible without loss of activity. Histamine
H3-receptor antagonists containing basic or hydrophilic
connecting functionalities with carbonyl groups or related
moieties were less suitable for imidazole replacement.
Nevertheless, the known histamine H3-receptor antagonists
containing aliphatic and aromatic ether structures investi-
gated in this study did tolerate imidazole replacement by
piperidine, resulting in the development of the FUB 181
analogue 6 and the ciproxifan analogue 7, two new potent
non-imidazole histamine H3-receptor antagonists with 7
being also highly potent in vivo. Imidazole exchange by an
azepane or pyrrolidine ring also resulted in histamine
H3-receptor antagonists with high in vitro and in vivo
potency in the case of 14 and 15. Replacement of the
imidazole moiety of other known histamine H3-receptor
antagonists was generally less successful. The inconsisten-
cies of these findings point out distinct sensitivities of
pharmacological properties of the individual compounds in
this study towards imidazole replacement. However, with
compounds 5, 6, 7, and 14 potent non-imidazole histamine
Fig. 3. Cumulative concentration–effect curves of R-(a)-methylhistamine
on guinea pig ileum in the absence (d) and presence of 6: (s) 10 nM,
(.) 30 nM, (,) 100 nM, (j) 300 nM (left) and Schild Plot (right).
Regression analysis yielded a slope of 0.9960.07, which was found not to
be significantly different from unity (competitive antagonism). The pA2
value was 8.2560.04 (95% conf. lim. 8.17–8.33).

258
G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
thioperamide as H3-receptor histamine antagonists. Collect. Czech.
Chem. Commun. 56, 2448–2455.
Ganellin, C.R., Leurquin, F., Piripitsi, A., Arrang, J.-M., Garbarg, M.,
Ligneau, X., Schunack, W., Schwartz, J.-C., 1998. Synthesis of potent
non-imidazole histamine H3-receptor antagonists. Arch. Pharm.
Pharm. Med. Chem. 331, 395–404.
Garbarg, M., Pollard, H., Trung Tuong, M.D., Schwartz, J.-C., Gros, C.,
1989. Sensitive radioimmunoassay for histamine and telemethylhis-
tamine in the brain. J. Neurochem. 53, 1724–1730.
H3-receptor antagonists have been successfully designed
from known antagonists. Moreover, in the light of a
potential therapeutic use, the unparalleled in vivo potency
of the ciproxifan analogue 7 in combination with possible
pharmacokinetic advantages of this compound make it a
promising subject for future investigations.
Garbarg, M., Arrang, J.-M., Rouleau, A., Ligneau, X., Trung Tuong,
M.D., Schwartz, J.-C., Ganellin, C.R., 1992. S-[2-(4-Imidazolyl)-
ethyl]isothiourea, a highly specific and potent histamine H3 receptor
agonist. J. Pharmacol. Exp. Ther. 263, 304–310.
Halpert, J.R., Guengerich, F.P., Bend, J.R., Correia, M.A., 1994. Selective
inhibitors of cytochrome P450. Toxicol. Appl. Pharmacol. 53, 1675–
1683.
Hill, S.J., Ganellin, C.R., Timmerman, H., Schwartz, J.-C., Shankley,
N.P., Young, J.M., Schunack, W., Levi, R., Haas, H.L., 1997. Interna-
tional Union of Pharmacology XIII. Classification of histamine
receptors. Pharmacol. Rev. 49, 253–278.
Acknowledgements
We are indebted to Dominique Dumoulin for technical
help. This work was supported by the Biomedical &
Health Research Programme (BIOMED) of the European
Union and the Fonds der Chemischen Industrie, Verband
der Chemischen Industrie, Frankfurt/Main, Germany.
¨
Karjalainen, A., Kalapudas, A., Sodervall, M., Pelkonen, O., Lammin-
tausta, R., 2000. Synthesis of new potent and selective aromatase
inhibitors based on long-chained diarylalkylimidazole and
diarylalkyltriazole molecule skeletons. Eur. J. Pharm. Sci. 11, 109–
131.
Kiec-Kononowicz, K., Ligneau, X., Schwartz, J.-C., Schunack, W., 1995a.
Pyrazoles as potential histamine H3-receptor antagonists. Arch. Pharm.
(Weinheim) 328, 469–472.
References
Ali, S.M., Tedford, C.E., Gregory, R., Handley, M.K., Yates, S.L., Hirth,
W.W., Phillips, J.G., 1999. Design, synthesis, and structure-activity
relationships of acetylene-based histamine H3 receptor antagonists. J.
Med. Chem. 42, 903–909.
Alves-Rodrigues, A., Leurs, R., Wu, T.-S., Prell, G.D., Foged, C.,
Timmerman, H., 1996. [3H]Thioperamide as a radioligand for the
histamine H3 receptor in rat cerebral cortex. Br. J. Pharmacol. 118,
2045–2052.
Arrang, J.-M., Garbarg, M., Schwartz, J.-C., 1985. Autoregulation of
histamine release in brain by presynaptic H3-receptors. Neuroscience
15, 553–562.
Arrang, J.-M., Garbarg, M., Schwartz, J.-C., 1987a. Autoinhibition of
histamine synthesis mediated by presynaptic H3-receptors. Neuro-
science 23, 149–157.
Kiec-Kononowicz, K., Ligneau, X., Stark, H., Schwartz, J.-C., Schunack,
W., 1995b. Azines and diazines as potential histamine H3-receptor
antagonists. Arch. Pharm. (Weinheim) 328, 445–450.
Leurs, R., Blandina, P., Tedford, C., Timmerman, H., 1998. Therapeutic
potential of histamine H3 receptor agonists and antagonists. Trends
Pharmacol. Sci. 19, 177–183.
Ligneau, X., Garbarg, M., Vizuete, M.L., Diaz, J., Purand, K., Stark, H.,
Schunack, W., Schwartz, J.-C., 1994. [ a new
125I]Iodoproxyfan,
antagonist to label and visualize cerebral histamine H3 receptors. J.
Pharmacol. Exp. Ther. 271, 452–459.
Arrang, J.-M., Garbarg, M., Lancelot, J.-C., Lecomte, J.-M., Pollard, H.,
Robba, M., Schunack, W., Schwartz, J.-C., 1987b. Highly potent and
selective ligands for histamine H3-receptors. Nature (London) 327,
117–123.
Blandina, P., Giorgetti, M., Bartolini, L., Cecchi, M., Timmerman, H.,
Leurs, R., Pepu, G., Giovannini, M.G., 1996. Inhibition of cortical
acetylcholine release and cognitive performance by histamine H3
receptor activation in rats. Br. J. Pharmacol. 119, 1656–1664.
Cheng, Y.C., Prusoff, W.H., 1973. Relationship between the inhibition
constant (Ki) and the concentration of inhibitor which causes 50%
inhibition (I50) of an enzymatic reaction. Biochem. Pharmacol. 22,
3099–3108.
Clinton, R.O., Salvador, U.J., Laskowski, S.C., 1949. Sulfur-containing
amines. VIII. Local anesthetics II. J. Am. Chem. Soc. 71, 3366–3370.
Clitherow, J.W., Beswick, P., Irving, W.J., Scopes, D.I.C., Barnes, J.C.,
Clapham, J., Brown, J.D., Evans, D.J., Hayes, A.G., 1996. Novel
1,2,4-oxadiazoles as potent and selective histamine H3 receptor
antagonists. Bioorg. Med. Chem. Lett. 6, 833–838.
Ligneau, X., Lin, J.-S., Vanni-Mercier, G., Jouvet, M., Muir, J.L.,
Ganellin, C.R., Stark, H., Elz, S., Schunack, W., Schwartz, J.-C., 1998.
Neurochemical and behavioral effects of ciproxifan, a potent histamine
H3-receptor antagonist. J. Pharmacol. Exp. Ther. 287, 658–666.
Ligneau, X., Morisset, S., Tardivel-Lacombe, J., Gbahou, F., Ganellin,
C.R., Stark, H., Schunack, W., Schwartz, J.-C., Arrang, J.-M., 2000.
Distinct pharmacology of rat and human histamine H3 receptors: role
of two amino acids in the third transmembrane domain. Br. J.
Pharmacol. 131, 1247–1250.
Linney, I.D., Buck, I.M., Harper, E.A., Kalindjian, S.B., Pether, M.J.,
Shankley, N.P., Watt, G.F., Wright, P.T., 2000. Design, synthesis, and
structure-activity relationships of novel non-imidazole histamine H3
receptor antagonists. J. Med. Chem. 43, 2362–2370.
Menge, W.M.P.B., Enguehard, C., Romeo, G., Limmen, B., Timmerman,
H., 1998. Synthesis and biological evaluation of a novel class of
non-imidazole histamine H3-receptor antagonists. In: 15th EFMC
International Symposium on Medicinal Chemistry, Edinburgh, Scot-
land, UK.
De Esch, I.J.P., Gaffar, A., Menge, W.M.P.B., Timmerman, H., 1999.
Synthesis and histamine H3 receptor activity of 4-(n-alkyl)-1H-imida-
zoles and 4-(v-phenylalkyl)-1H-imidazoles. Bioorg. Med. Chem. 7,
3003–3009.
Doherty, D.G., Shapira, R., Burnett, Jr. W.T., 1957. Synthesis of amino-
alkylisothiuronium salts and their conversion to mercap-
toalkylguanidines and thiazolines. J. Am. Chem. Soc. 79, 5667–5671.
Frankel, M., Mosher, H.S., Whitmore, F.C., 1950. Addition reactions of
1-cyano-1,3-butadiene. J. Am. Chem. Soc. 72, 81–83.
Mitsunobu, O., 1981. The use of diethyl azodicarboxylate and tri-
phenylphosphine in synthesis and transformation of natural products.
Synthesis, 1–28.
Miyazaki, S., Imaizumi, M., Onodera, K., 1995. Effects of thioperamide,
a histamine H3-receptor antagonist, on a scopolamine-induced learning
deficit using an elevated plus-maze test in mice. Life Sci. 57, 2137–
2144.
Morisset, S., Traiffort, E., Schwartz, J.-C., 1996. Inhibition of histamine
versus acetylcholine metabolism as a mechanism of tacrine activity.
Eur. J. Pharmacol. 315, R1–R2.
Ganellin, C.R., Jayes, D., Khalaf, Y.S., Tertiuk, W., Arrang, J.-M.,
Defontaine, N., Schwartz, J.-C., 1991. Synthesis of pyridyl isosteres of
Nystrom, R.F., Brown, W.G., 1948. Reduction of organic compounds by

G. Meier et al. / European Journal of Pharmaceutical Sciences 13 (2001) 249–259
259
lithium aluminum hydride III. Halides, quinones, miscellaneous nitro-
gen compounds. J. Am. Chem. Soc. 70, 3738–3740.
Schwartz, J.-C., Schunack, W., 1998a. Development of FUB 181, a
selective histamine H3-receptor antagonist of high oral in vivo potency
with 4-(v-(arylalkyloxy)alkyl)-1H-imidazole structure. Arch. Pharm.
Pharm. Med. Chem. 331, 211–218.
Stark, H., Ligneau, X., Arrang, J.-M., Schwartz, J.-C., Schunack, W.,
1998b. General construction pattern of histamine H3-receptor antago-
nists: change of a paradigm. Bioorg. Med. Chem. Lett. 8, 2011–2016.
Onodera, K., Miyazaki, S., Imaizumi, M., Stark, H., Schunack, W., 1998.
Improvement by FUB 181, a novel histamine H3-receptor antagonist,
of learning and memory in the elevated plus-maze test in mice.
Naunyn-Schmiedeberg’s Arch. Pharmacol. 357, 508–513.
Panula, P., Kuokkanen, K., Relja, M., Eriksson, K.S., Sallmen, T., Rinne,
J.O., Kalimo, H., 1995. Significant changes in the human brain
histaminergic system in Alzheimer’s disease. Soc. Neurosci. Abstr. 21,
1977.
Parker, R.B., Waud, D.R., 1971. Pharmacological estimation of drug-
receptor dissociation constants. Statistical evaluation. I. Agonists. J.
Pharmacol. Exp. Ther. 177, 112.
Plazzi, P.V., Bordi, F., Mor, M., Silva, C., Morini, G., Caretta, A.,
Barocelli, E., Vitali, T., 1995. Heteroarylaminoethyl and
heteroarylthioethyl imidazoles. Synthesis and H3-receptor affinity. Eur.
J. Med. Chem. 30, 881–889.
Sasse, A., Kiec-Kononowicz, K., Stark, H., Motyl, M., Reidemeister, S.,
Ganellin, C.R., Ligneau, X., Schwartz, J.-C., Schunack, W., 1999.
Development of chiral N-alkylcarbamates as new leads for potent and
selective H3-receptor antagonists: Synthesis, capillary electrophoresis,
and in vitro and oral in vivo activity. J. Med. Chem. 42, 593–600.
Schlicker, E., Kathmann, M., Reidemeister, S., Stark, H., Schunack, W.,
1994. Novel histamine H3 receptor antagonists: Affinities in an H3
receptor binding assay and potencies in two functional H3 receptor
models. Br. J. Pharmacol. 112, 1043–1048, Erratum: 1994. 113, 657.
Schlicker, E., Marr, I., 1996. The moderate affinity of clozapine at H3
receptors is not shared by its two major metabolites and by structurally
related and unrelated atypical neuroleptics. Naunyn-Schmiedeberg’s
Arch. Pharmacol. 353, 290–294.
Schwartz, J.-C., Arrang, J.-M., in press. Histamine. In: Charney, D.,
Cayle, J., Davis, K., Nemeroff, C. (Eds.), Psychopharmacology: The
Fifth Generation of Progress. Raven Press, New York.
Schwartz, J.-C., Arrang, J.-M., Garbarg, M., Pollard, H., Ruat, M., 1991.
Histaminergic transmission in the mammalian brain. Physiol. Rev. 71,
151.
¨
Stark, H., Sadek, B., Krause, M., Huls, A., Ligneau, X., Ganellin, C.R.,
Arrang, J.-M., Schwartz, J.-C., Schunack, W., 2000. Novel histamine
H3-receptor antagonists with carbonyl substituted 4-(3-(phenox-
y)propyl)-1H-imidazole structures like ciproxifan and related com-
pounds. J. Med Chem. 43, 3987–3994.
Tardivel-Lacombe, J., Rouleau, A., Heron, A., Morisset, S., Pillot, C.,
Cochois,V., Schwartz, J.-C., Arrang, J.-M., 2000. Cloning and cerebral
expression of the guinea pig histamine H3 receptor: evidence for two
isoforms. Neuroreport 11, 755–759.
Van der Goot, H., Schepers, M.J.P., Sterk, G.J., Timmerman, H., 1992.
Isothiourea analogues of histamine as potent agonists or antagonists of
the histamine H3-receptor. Eur. J. Med. Chem. 27, 511–517.
Vollinga, R.C., Zuiderveld, O.P., Scheerens, H., Bast, A., Timmerman,
H.A., 1992. A simple and rapid in vitro test system for the screening
of histamine H3 ligands. Methods Find. Exp. Clin. Pharmacol. 14,
747–751.
Vollinga, R.C., Menge, W.M.P.B., Leurs, R., Timmerman, H., 1995.
Homologs of histamine as histamine H3 receptor antagonists: a new
potent and selective H3 antagonist, 4(5)-(5-aminopentyl)-1H-imida-
zole. J. Med. Chem. 38, 266–271.
Walczynski, K., Guryn, R., Zuiderveld, O.P., Timmerman, H., 1999a.
Non-imidazole histamine H3 ligands. Part 1. Synthesis of 2-(1-
piperazinyl)- and 2-(hexahydro-1H-1,4-diazepin-1-yl) benzothiazole
derivatives as H3 antagonists with H1 blocking activities. Farmaco
541, 684–694.
Walczynski, K., Guryn, R., Zuiderveld, O.P., Timmerman, H., 1999b.
Non-imidazole histamine H3 ligands. Part 2: New 2-substituted
benzothiazoles as histamine H3 antagonists. Arch. Pharm. Pharm.
Med. Chem. 332, 389–398.
Schwartz, J.-C., Arrang, J.-M., Garbarg, M., Lecomte, J.-M., Ligneau, X.,
Schunack, W., Stark, H., Ganellin, C.R., Leurquin, F., Elz, S., 2000.
Non-imidazole alkanamines as histamine H3-receptor ligands and their
therapeutic applications. PCT WO 00/06254.
Stark, H., Lipp, R., Arrang, J.-M., Garbarg, M., Ligneau, X., Schwartz,
J.-C., Schunack, W., 1995. New potent histamine H3-receptor antago-
nists of the amide type. Eur. J. Pharm. Sci. 3, 95–104.
Waud, D.R., Parker, R.B., 1971. Pharmacological estimation of drug-
receptor dissociation constants. Statistical evaluation. II. Competitive
Antagonists. J. Pharmacol. Exp. Ther. 177, 13–24.
Whittaker, V.P., 1966. Some properties of synaptic membranes isolated
from the central nervous system. Ann. NY Acad. Sci. 137, 982–998.
Williamson, A., 1851. About the theory of the formation of ethers. Ann.
Chem. 77, 37–49.
Stark, H., Schlicker, E., Schunack, W., 1996a. Developments of histamine
H3-receptor antagonists. Drugs Future 21, 507–520.
Stark, H., Purand, K., Ligneau, X., Rouleau, A., Arrang, J.-M., Garbarg,
M., Schwartz, J.-C., Schunack, W., 1996b. Novel carbamates as potent
histamine H3 receptor antagonists with high in vitro and oral in vivo
activity. J. Med. Chem. 39, 1157–1163.
Yokoyama, H., Onodera, K., Iinuma, K., Watanabe, T., 1993. Effect of
thioperamide, a histamine H3 receptor antagonist, on electrically
induced convulsions in mice. Eur. J. Pharmacol. 234, 129–133.
Yokoyama, H., Onodera, K., Maeyama, K., Sakurai, E., Iinuma, K.,
Leurs, R., Timmerman, H., Watanabe, T., 1994. Clobenpropit (VUF
9153), a new histamine H3 receptor antagonist, inhibits electrically
induced convulsions in mice. Eur. J. Pharmacol. 260, 23–28.
¨
Stark, H., Huls, A., Ligneau, X., Purand, K., Pertz, H., Arrang, J.-M.,