Investigation of SAR requirements of SR 142801 through an indexed combinatorial library in solution

Article abstract of DOI:10.1016/S0223-5234(99)00210-X

To rapidly gain information on structure-activity relationship (SAR) requirements of the human neurokinin 3 (hNK-3) receptor antagonist SR 142801, an indexed combinatorial library was synthesised in solution and screened on the hNK-3 receptor. SAR considerations drawn from binding affinity of combinatorial mixtures were confirmed through the synthesis and biological evaluation of some individual compounds.

Full text of DOI:10.1016/S0223-5234(99)00210-X

Eur. J. Med. Chem. 34 (1999) 825−835
© 1999 Editions scientifiques et médicales Elsevier SAS. All rights reserved
825
´
Original article
Investigation of SAR requirements of SR 142801 through an indexed
combinatorial library in solution
Luca F. Ravegliaa*, Mauro Vitalic, Marco Articoa, Davide Graziania,
Douglas W.P. Hayb, Mark A. Luttmannb, Renzo Menaa,
Giorgio Pifferic, Giuseppe A.M. Giardinaa
aDepartment of Medicinal Chemistry, SmithKline Beecham S.p.A., Via Zambeletti, 20021 Baranzate, Milano, Italy
bDepartment of Pulmonary Pharmacology, SmithKline Beecham Pharmaceuticals, 709 Swedeland Road, King Of Prussia, PA, 194106,
USA
cIstituto di Chimica Farmaceutica, V.le Abruzzi 42, 20133 Milano, Italy
(Received 3 May 1999; accepted 25 May 1999)
Abstract – To rapidly gain information on structure-activity relationship (SAR) requirements of the human neurokinin 3 (hNK-3) receptor
antagonist SR 142801, an indexed combinatorial library was synthesised in solution and screened on the hNK-3 receptor. SAR considerations
drawn from binding affinity of combinatorial mixtures were confirmed through the synthesis and biological evaluation of some individual
´
compounds. © 1999 Editions scientifiques et médicales Elsevier SAS
tachykinins / neurokinin 3 receptor antagonists / combinatorial chemistry / indexed libraries / structure-activity relationships
1. Introduction
The neurokinin 3 (NK-3) is one of the three receptors
for the family of peptides named tachykinins or neuroki-
nins and belongs to the G protein-coupled receptor
superfamily [1]. In 1996, our group described the identi-
fication of potent and selective non-peptide NK-3 recep-
tor antagonists featuring the 2-phenyl-4-quinoline-
carboxamide framework, including SB 223412 [2, 3];
this is one of the three main chemical classes of non-
peptide NK-3 receptor antagonists reported to date [4, 5].
The other two are the peptide-derived structures reported
Figure 1. Structure of SR 142801.
by Parke-Davis (PD 161182) [6] and the 3,4-dichloro-
phenylpiperidines which were first described by Sanofi
(exemplified by SR 142801, figure 1) [7] and subse-
quently by Merck Sharp and Dohme [8]. Due to our
synthesis of a small library of analogues of SR 142801
involvement in the NK-3 area, we were interested in
was planned via a combinatorial chemistry approach.
investigating the SAR requirements of the 3,4-dichloro-
Since the 3-(3,4-dichlorophenyl)-3-propylpiperidine
framework A (figure 1) of SR 142801 appears to be a key
feature of this class of NK-3 receptor antagonists, able to
drive their selectivity for the NK-3 receptor with respect
to the NK-1 and NK-2 [4, 8], we decided to keep this
phenylpiperidine NK-3 receptor antagonists, only mar-
ginally described in the literature [8]. For this reason, the
*Correspondence and reprints

826
Figure 2. Library design.
moiety fixed and focus our attention on variations in the
basic head X and the acylating group Y.
literature procedure [12], protected at the nitrogen and
activated to nucleophilic substitution by converting the
hydroxy group into the mesylate and this, in turn, into the
more reactive iodo derivative 2 [13]. Displacement of
iodine with the 7 amines X1–X7 (CH3CN, K2CO3, 90 °C,
18 h) afforded the desired intermediate compounds
X1A–X7A, which were purified by flash column chroma-
tography.
In designing the library, 7 amines (X1–X7) and 7
acylating groups (Y1–Y7) were selected, on the basis of
commercial availability of corresponding reagents, to
give a total of 49 compounds (X1–7AY1–7), as all the
possible combinations of X and Y substituents (figure 2).
To simplify the synthetic pathway, it was planned to work
on racemic mixtures.
Solution phase combinatorial chemistry was the tech-
nique of choice for the synthesis of the library because a
suitable point of attachment to the resin could not be
found, ruling out a solid phase approach. Moreover, since
in the synthetic scheme the “combinatorial reaction” was
a well consolidated amide coupling at the last step
(figures 4 and 5), and since the number of compounds to
synthesise was small (49), we reasoned that pools of
compounds could easily be obtained by using mixtures of
reactants, without risk of getting too many by-products.
In particular, to help the structural determination of active
components of the library, the strategy of indexed (or
orthogonal) combinatorial libraries was utilised [9–11].
2.2. Synthesis of the library: set 1
As depicted in figure 4, each intermediate compound
was subsequently deprotected (20% TFA/CH2Cl2, r.t.,
18 h) and reacted with a stoichiometric and equimolar
mixture of the 7 acylating agents Y1–Y7, (CH2Cl2,
K2CO3, r.t., 18 h). Simple washing of the reaction mix-
tures with H2O and evaporation to dryness afforded the
first set (set 1) of desired mixtures (or sublibraries)
X1AY1–7–X7AY1–7 in satisfactory to high yields
(60–80%) [14a] and high purity (95–98%, LC/UV) [14b].
Peak identity was assessed by LC/MS and all expected
molecular weights were detected in each mixture.
It is worth noting that the seven sublibraries of set 1
contain all the desired 49 compounds; each sublibrary has
the X substitution fixed and different from one another,
while containing all possible Y substituents. Since the Y
portion is common to all the sublibraries, binding affinity
data on this set will clarify the SAR related to the X
group.
2. Chemistry
2.1. Synthesis of intermediate compounds
In figure 3, the synthesis of intermediate compounds is
described. Racemic aminol 1 was obtained according to

827
Figure 3. Synthesis of intermediates.
2.3. Synthesis of the library: set 2
with H2O and evaporation to dryness produced the
desired sublibraries X1–7AY1–X1–7AY7 in high yields
(74–95%) [14a] and high purity (90–98%, LC/UV, except
for X1–7AY4: 83%) [14b]. All expected molecular
weights were detected in each mixture by LC/MS.
In figure 5, the synthesis of the second set (set 2) of
sublibraries is described. An equimolar amount of each of
the intermediate compounds X1A–X7A (figure 3) was
mixed into the same reaction vessel and Boc-deprotected;
the mixture was then split into seven equal portions
(X1–7A), and each portion was reacted with a stoichio-
metric amount of acyl chloride/phenylisocyanate/
phenylsulfonyl chloride Y1–Y7 (CH2Cl2, K2CO3, r.t.,
18 h). Again, simple washing of the reaction mixtures
The seven sublibraries of set 2 contain the same 49
compounds of set 1, but in a different combination: in this
case, the Y substituent is fixed and peculiar to any
sublibrary, while all possible X substitutions are common
to all. Binding affinity data on this second set will clarify

828
Figure 4. Synthesis of the library: set 1.
the SAR related to the Y group. By combining informa-
tion coming from the two sets of sublibraries, the best X
(set 1) and the best Y (set 2) substituents should be
identified, thus allowing structural determination of the
most active compounds of the whole library.
pound (20% TFA/CH2Cl2, r.t., 18 h) and reacting it with
the appropriate acylating agent (CH2Cl2, K2CO3, r.t.,
18 h).
3. Pharmacology
2.4. Synthesis of individual compounds
Receptor binding assays were performed with crude
membranes from CHO cells expressing the hNK-3 recep-
tor as detailed previously [3, 15]. For NK-3 receptor
competition binding studies, [125I]-[MePhe7]-NKB bind-
ing to hNK-3-CHO membranes was performed using the
To prove the validity of the technique and the predic-
tions made from binding affinity data of mixtures, the six
individual compounds reported in table II were synthe-
sised, deprotecting the appropriate intermediate com-

829
Figure 5. Synthesis of the library: set 2.
procedure of Sadowski and co-workers [16]. Specific
binding was determined by subtracting total binding from
non-specific binding, which was assessed as the binding
in the presence of 0.5 µM cold [MePhe7]-NKB. Percent
inhibition of specific binding was determined for each
concentration of the compounds and the IC50, defined as
the concentration required to inhibit 50% of the specific
binding, obtained from concentration-response curves.
Values reported in tables I and II are the apparent
inhibition constant (Ki), which was calculated from the
IC50 as described by Cheng and Prusoff [17].
4. Results and Discussion
Human NK-3 receptor binding affinity data for the 14
sublibraries synthesised are reported in table I. Results of
sublibraries of set 1 revealed that pyrrolidinocarbonyl-
methylpiperazine (X7, Ki = 113 nM) should be slightly
more active than phenylpiperidine (X1, Ki = 160 nM)
which, in turn, should be more active than phenylpipera-
zine (X2, Ki = 232 nM). A strongly reduced affinity is
forseen for isopropylpiperazine (X3, Ki = 743 nM) and
morpholine (X4, Ki = 1235 nM).

830
Table I. Binding affinities of library compounds.
Set 1
Set 2
X
Y
hNK-3 Ki
(nM)
X
Y
hNK-3 Ki
(nM)a,b
a,b
Y1-7
160
X1-7
Y1
CH3CO
PhCO
9163
Y1-7
232
X1-7
Y2
Y3
242
Y1-7
743
X1-7
PhCH2CO
394
Y1-7
Y1-7
Y1-7
1235
525
X1-7
X1-7
X1-7
Y4
Y5
Y6
(CH3)2C=CHCO
2,4-(MeO)2PhCO
PhNHCO
503
1835
1497
440
Y1-7
113
X1-7
Y7
PhSO2
570
b
aInhibition of [125I]MePhe7-NKB binding in hNK-3-CHO cell membranes. single determination (n = 1).
As far as the Y substitution is concerned (set 2),
benzoyl (Y2, Ki = 242 nM), also present in SR 142801,
appears to be the best substituent. Increasing the distance
of the phenyl ring from the carbonyl with a methylene
spacer (Y3, Ki = 394 nM) slightly decreases the binding
affinity, while substitution at the phenyl ring with two
methoxy groups (Y5, Ki = 1 835 nM) or replacement of
the phenyl with a methyl (Y1, Ki = 9 163 nM) resulted in
a marked loss of activity. Dimethylacryloyl (Y4, Ki = 503
nM) and phenylsulfonyl (Y7, Ki = 570 nM), although less
potent than the benzoyl (Y2, Ki = 242 nM), maintain a
certain activity, while the urea (Y6, Ki = 1497 nM) does
not appear to be a good replacement for the benzamide.
To test the reliability of the predictions made from
mixtures, six individual compounds (some predicted to
be the most active and some predicted to have medium
activity) were synthesised and screened (table II). The
rank order of potency of individual compounds was
found to be in good agreement with the prediction,
X7AY2 being the most active (hNK-3 binding affinity, Ki
= 35.4 ± 10.4 nM) and X1AY5 the least active (hNK-3
binding affinity, Ki = 513 ± 99.5 nM) amongst individual
compounds prepared. It is worth noting that the best
individual compound synthesised, X7AY2, appears to

831
Table II. Binding affinities of individual compounds.
piperidine NK-3 receptor antagonists with an improved
biological profile compared to SR 142801 [19].
5. Conclusions
In conclusion, an indexed combinatorial library of
analogues of SR 142801 was synthesised in solution and
screened for its hNK-3 receptor binding affinity. Synthe-
sis and biological evaluation of some individual com-
pounds proved the reliability of this very simple, acces-
sible and rather under-utilised technique and allowed us
to gain information on SAR requirements of 3,4-
dichlorophenylpiperidine NK-3 receptor antagonists, sig-
nificantly decreasing the number of reactions and samples
to test.
Individual compounds
X
Y
hNK-3 Ki
(nM)a,b
PhCO
46.2 ± 9.2
513 ± 99.5
50.9 ± 9.2
35.4 ± 10.4
2,4-
(MeO)2PhCO
6. Experimental protocols
PhCO
PhCO
6.1. Chemistry
Melting points were determined with a Büchi 530 hot
stage apparatus and are uncorrected. Proton NMR spectra
were recorded on a Bruker ARX 300 spectrometer at
303 K unless otherwise indicated. Chemical shifts were
recorded in parts per million (δ) downfield from tetra-
methylsilane (TMS); NMR spectral data are reported as a
list. IR spectra were recorded in Nujol mull or neat on
sodium chloride disks or in KBr with a Perkin-Elmer
1420 spectrophotometer; mass spectra were obtained on a
Finnigan MAT TSQ-700 spectrometer. Silica gel used for
flash column chromatography was Kiesegel 60 (230–400
mesh) (E. Merck AG, Darmstadt, Germany). All evapo-
rations were performed at reduced pressure. Combustion
elemental analyses were performed by Redox s.n.c.,
Milan, Italy and analyses indicated by the symbols of the
elements were within ± 0.4% of the theoretical values. All
reagents utilised in figures 3–5 are commercially avail-
able compounds and were used without further purifica-
tion. Racemic aminol 1 and SR 142801 were synthesised
according to Giardina et al. [12] and to Chen et al. [13].
PhCH2CO
289 ± 68.0
175 ± 50.2
1.2 ± 0.3
PhSO2
PhCO SR 142801c,d
aInhibition of [125I]MePhe7-NKB binding in hNK-3-CHO cell
membranes. bmean ± SEM for three determinations (n = 3). in
c
d
house data. SR 142801 is a single enantiomer [18].
have an hNK-3 binding affinity about 30 times lower
than SR 142801 (hNK-3 binding affinity, Ki = 1.2 ±
0.3 nM) [18]. Since X7AY2 differs from SR 142801 only
for the basic head X, this SAR study outlines the
importance of the X substitution in conferring high
binding affinity to the 3,4-dichlorophenylpiperidine NK-3
receptor antagonists. Additional work is needed to under-
stand the requirements of this portion of the molecule and
X7 could be a good starting point for further chemical
modifications in order to obtain novel 3,4-dichlorophenyl-
6.2.
N-tert-butoxycarbonyl-3-(3,4-dichlorophenyl)-3-
(3-iodopropyl)piperidine 2
Racemic aminol 1 [12] (4.7 g, 16.3 mmol) and di-tert-
butyl dicarbonate (4.3 g, 19.6 mmol) were dissolved,
under nitrogen atmosphere, in dry CH2Cl2 (75 mL). The
reaction mixture was stirred at room temperature for 8 h
and left standing overnight. The solvent was evaporated
to dryness and the crude material was purified by flash
column chromatography, eluting with a mixture of

832
CH2Cl2/MeOH 98:2, to obtain 5.4 g (85%) of N-tert-
butoxycarbonyl-3-(3,4-dichlorophenyl)-3-(3-hydroxy-
propyl)piperidine as a pale yellow oil. This compound
(5.4 g, 13.9 mmol) was dissolved in CH2Cl2 (50 mL);
triethylamine (TEA) (2.2 mL, 15.8 mmol) was added and
the solution was cooled to 0 °C. Methanesulfonyl chlo-
ride (1.2 mL, 15.4 mmol), dissolved in CH2Cl2 (7 mL),
was added dropwise and the reaction mixture was al-
lowed to reach room temperature and stirred for addi-
tional 2 h. The reaction was quenched with water and the
extracted organic layer was washed with 10% citric acid,
5% NaHCO3 and brine, dried over MgSO4, filtered and
evaporated to dryness to yield 6.0 g (93%) of N-tert-
butoxycarbonyl-3-(3,4-dichlorophenyl)-3-(3-methane-
sulfonyloxypropyl)piperidine as a pale yellow oil. This
compound (6.0 g, 12.9 mmol) was dissolved in acetone
(120 mL) and KI (3.5 g, 20.9 mmol) was added to the
solution which was refluxed for 16 h. Insoluble material
was filtered off and the filtrate was evaporated to dryness
to yield 6.4 g (100%) of the title compound as a red oil,
which was used in the following reactions without further
purification. IR (neat) 2 940, 2 885, 1 692, 1 470,
11H), 1.49 (s, 9H). ESI-MS (positive, solvent: methanol,
spray 4.5 keV, skimmer: 60 eV, capillary 220 °C) m/z 531
(MH+).
6.3.2. N-tert-butoxycarbonyl-3-(3,4-dichlorophenyl)-3-[3-
(4-phenylpiperazin-1-yl)propyl]piperidine X2A
IR (KBr) 2 942, 2 816, 1 692, 1 602, 1 554,
1
1 502 cm–1. H-NMR (CDCl3) δ 7.44 (d, 1H), 7.38 (d,
1H), 7.24 (dd, 2H), 7.18 (d br, 1H), 6.90 (d, 2H), 6.84 (dd,
1H), 4.05 (m br, 1H), 3.60 (m, br, 1H), 3.20–3.10 (m,
6H), 2.43 (m, 4H), 2.21 (t, 2H), 2.05 (m br, 1H),
1.75–1.10 (m, 7H), 1.46 (s, 9H). ESI-MS (positive,
solvent: methanol, spray 4.5 keV, skimmer: 60 eV,
capillary 220 °C) m/z 532 (MH+), 554 (MNa+).
6.3.3. N-tert-butoxycarbonyl-3-(3,4-dichlorophenyl)-3-[3-
(4-isopropylpiperazin-1-yl)propyl]piperidine X3A
IR (neat) 2 942, 2 812, 1 690, 1 554, 1 470 cm–1.
1H-NMR (CDCl3) δ 7.42 (d, 1H), 7.31 (d, 1H), 7.15 (d br,
1H), 4.05 (m br, 1H), 3.60 (m br, 1H), 3.13 (d, 1H), 3.12
(m, 1H), 2.64 (m, 1H), 2.50 (m, 4H), 2.35 (m, 4H), 2.20
(t, 2H), 2.05 (m, 1H), 1.70–1.00 (m, 7H), 1.45 (s, 9H),
1.00 (d, 6H). ESI-MS (positive, solvent: methanol, spray
4.5 keV, skimmer: 60 eV, capillary 220 °C) m/z 498
(MH+).
1
1 428 cm–1. H-NMR (CDCl3) δ 7.44 (d, 1H), 7.38 (d,
1H), 7.17 (dd, 1H), 3.91 (d, 1H), 3.58–3.47 (m, 1H), 3.31
(d, 1H), 3.31–3.20 (m, 1H), 3.01 (t, 2H), 2.07–1.98 (m,
1H), 1.80–1.40 (m, 7H), 1.49 (s, 9H).
6.3. General procedure for the synthesis of intermediate
compounds X1A–X7A
6.3.4. N-tert-butoxycarbonyl-3-(3,4-dichlorophenyl)-3-[3-
(morpholin-4-yl)propyl]piperidine X4A
K2CO3 (0.303 g, 2.2 mmol) and the appropriate amine
IR (neat) 2 944, 2 858, 2 810, 1 690, 1 590,
(1.1 eq.) were added to
a solution of N-tert-
1
1 554 cm–1. H-NMR (CDCl3) δ 7.44 (d, 1H), 7.35 (d,
butoxycarbonyl-3-(3,4-dichlorophenyl)-3-(3-iodopropyl)
piperidine 2 (0.99 g, 1.98 mmol) in CH3CN (7 mL). The
reaction mixture was heated to 80 °C under magnetic
stirring for 18 h; then it was filtered and the filtrate was
evaporated to dryness. The crude solid obtained was
dissolved in EtOAc and washed with H2O. The organic
layer was dried over MgSO4, filtered and evaporated to
dryness to yield the title compounds which were purified
by flash column chromatography. Yields after chromatog-
raphy were in the range: 75–96%. Spectroscopic data for
compounds X1A–X7A are reported below.
1H), 7.14 (d br, 1H), 4.02 (m br, 1H), 3.63 (m br, 4H),
3.59 (m br, 1H), 3.13 (d, 1H), 3.12 (m, 1H), 2.30 (m, 4H),
2.18 (t, 2H), 2.05 (m br, 1H), 1.71–1.00 (m, 7H), 1.46 (s,
9H). ESI-MS (positive, solvent: methanol, spray 4.5 keV,
skimmer: 60 eV, capillary 220 °C) m/z 457 (MH+), 479
(MNa+).
6.3.5. N-tert-butoxycarbonyl-3-(3,4-dichlorophenyl)-3-[3-
(1,2,3,4-tetrahydroisoquinolin-2-yl)propyl]piperidine
X5A
IR (neat) 2 932, 1 692, 1 554 cm–1. 1H-NMR
(CDCl3–343 K) δ 7.48 (d, 1H), 7.36 (d, 1H), 7.20 (dd,
1H), 7.10–7.03 (m, 3H), 6.94 (m, 1H), 4.01 (d, 1H), 3.58
(dt, 1H), 3.47 (s, 2H), 3.25 (d, 1H), 3.20 (ddd, 1H), 2.83
(t, 2H), 2.59 (t, 2H), 2.34 (t, 2H), 2.09–2.00 (m, 1H),
1.78–1.13 (m, 7H), 1.49 (s, 9H). ESI-MS (positive,
solvent: methanol, spray 4.5 keV, skimmer: 60 eV,
capillary 220 °C) m/z 503 (MH+).
6.3.1. N-tert-butoxycarbonyl-3-(3,4-dichlorophenyl)-3-[3-
(4-phenylpiperidin-1-yl)propyl]piperidine X1A
IR (Nujol) 2 926, 1 690, 1 674, 1 604, 1 554 cm–1.
1H-NMR (CDCl3–343 K) δ 7.47 (d, 1H), 7.37 (d, 1H),
7.30–7.14 (m, 6H), 4.00 (d, 1H), 3.56 (dt, 1H), 3.26 (d,
1H), 3.26–3.18 (m, 1H), 2.90–2.83 (m, 2H), 2.50–2.40
(m, 1H), 2.21 (t, 2H), 2.09–1.91 (m, 3H), 1.80–1.13 (m,

833
6.3.6. N-tert-butoxycarbonyl-3-(3,4-dichlorophenyl)-3-[3-
(1-phenyl-1,3,8-triazaspiro[4,5]decan-4-on-8-yl)propyl]
piperidine X6A
obtain 0.233 g (78%) of the title sublibrary. All seven
compounds forming the sublibrary were characterised by
LC/MS.
IR (neat) 2 938, 2 856, 1 708, 1 690, 1 602,
Purity of the seven sublibraries of set 1, calculated as
the sum of the areas of the seven peaks of the LC/UV
chromatogram attributed by LC/MS to the seven com-
pounds forming each sublibrary, is reported below:
X1AY1–7 98%; X2AY1–7 98%; X3AY1–7 95%;
X4AY1–7 98%; X5AY1–7 97%; X6AY1–7 98%; X7AY1–7
97%.
1
1 502 cm–1. H-NMR (CDCl3–343 K) δ 7.46 (d, 1H),
7.36 (d, 1H), 7.29 (dd, 2H), 7.19 (dd, 1H), 6.99–6.89 (m,
3H), 5.61 (s br, 1H), 4.70 (s, 2H), 3.90 (d br, 1H), 3.56
(m, 1H), 3.29–3.19 (m, 2H), 2.75–2.42 (m, 6H), 2.23 (m,
2H), 2.05 (m, 1H), 1.78–1.05 (m, 9H), 1.47 (s, 9H).
ESI-MS (positive, solvent: methanol, spray 4.5 keV,
skimmer: 60 eV, capillary 220 °C) m/z 601 (MH+), 545.
6.5. General procedure for the synthesis of library (set 2)
compounds X1–7AY1–X1–7AY7
6.3.7. N-tert-butoxycarbonyl-3-(3,4-dichlorophenyl)-3-{3-
[4-(pyrrolidinocarbonylmethyl)piperazin-1-yl]propyl}
piperidine X7A
IR (neat) 2 944, 2 874, 2 812, 1 690, 1 642,
6.5.1. Synthesis of sublibrary X1–7AY1
1
1 554 cm–1. H-NMR (CDCl3–343 K) δ 7.44 (d, 1H),
The synthesis of sublibrary X1–7AY1 is reported as an
example of the synthesis of all sublibraries of the second
set of the library. N-tert-butoxycarbonyl-3-(3,4-di-
chlorophenyl)-3-[3-(4-phenylpiperidin-1-yl)propyl]
piperidine X1A (116 mg, 0.217 mmol), N-tert-
butoxycarbonyl-3-(3,4-dichlorophenyl)-3-[3-(4-phenyl-
7.36 (d, 1H), 7.18 (dd, 1H), 3.90 (d, 1H), 3.55 (dt, 1H),
3.54–3.43 (m, 4H), 3.23 (d, 1H), 3.20 (ddd, 1H), 3.09 (s,
2H), 2.55 (m, 4H), 2.36 (m, 4H), 2.19 (t, 2H), 2.08–1.99
(m, 1H), 1.98–1.80 (m, 4H), 1.78–1.00 (m, 7H), 1.49 (s,
9H). ESI-MS (positive, solvent: methanol, spray 4.5 keV,
skimmer: 60 eV, capillary 220 °C) m/z 567 (MH+).
piperazin-1-yl)propyl]piperidine
X2A
(116 mg,
0.217 mmol), N-tert-butoxycarbonyl-3-(3,4-dichloro-
6.4. General procedure for the synthesis of library (set 1)
compounds X1AY1-7–X7AY1–7
phenyl)-3-[3-(4-isopropylpiperazin-1-yl)propyl]piperidine
X3A (108 mg, 0.217 mmol), N-tert-butoxycarbonyl-
3-(3,4-dichlorophenyl)-3-[3-(morpholin-4-yl)propyl]
piperidine X4A (100 mg, 0.217 mmol), N-tert-
butoxycarbonyl-3-(3,4-dichlorophenyl)-3-[3-(1,2,3,4-
6.4.1 Synthesis of sublibrary X1AY1–7
The synthesis of sublibrary X1AY1–7 is reported as an
example of the synthesis of all sublibraries of the first set
of the library. N-tert-butoxycarbonyl-3-(3,4-dichloro-
phenyl)-3-[3-(4-phenylpiperidin-1-yl)propyl]piperidine
X1A (0.301 g, 0.55 mmol) was dissolved in dry CH2Cl2
(8 mL); TFA (2 mL) was added and the reaction mixture
was stirred at room temperature overnight. After evapo-
ration to dryness, the residue was taken up with CH2Cl2,
washed with 5% NaHCO3, dried over MgSO4, filtered
and evaporated to dryness, to obtain 3-(3,4-dichloro-
phenyl)-3-[3-(4-phenylpiperidin-1-yl)propyl]piperidine.
This compound was dissolved in dry CH2Cl2 (10 mL),
and K2CO3 (165.3 mg, 1.2 mmol) was added; then a
solution of acetyl chloride (564 µL, 7.9 mmol), benzoyl
chloride (918 µL, 7.9 mmol), phenylacetylchloride
(1 047 µL, 7.9 mmol), dimethylacryloyl chloride
(880 µL, 7.9 mmol), 2,4-dimethoxybenzoyl chloride
(1 585 mg, 7.9 mmol), phenylisocianate (859 µL,
7.9 mmol) and phenylsulfonyl chloride (1 013 µL,
7.9 mmol), brought to 100 mL with dry CH2Cl2, was
prepared and 1 mL of this solution was added to the
reaction mixture, which was stirred at room temperature
overnight. The reaction was quenched with H2O (5 mL)
and the extracted organic layer was washed with H2O,
dried over MgSO4, filtered and evaporated to dryness to
tetrahydroisoquinolin-2-yl)propyl]piperidine
X5A
(110 mg, 0.217 mmol), N-tert-butoxycarbonyl-3-(3,4-
dichlorophenyl)-3-[3-(1-phenyl-1,3,8-triazaspiro[4,5]
decan-4-on-8-yl)propyl]piperidine
X6A
(131 mg,
0.217 mmol) and N-tert-butoxycarbonyl-3-(3,4-dichloro-
phenyl)-3-{3-[4-(pyrrolidinocarbonylmethyl)piperazin-
1-yl]propyl}piperidine X7A (124 mg, 0.217 mmol) were
dissolved, under magnetic stirring, in CH2Cl2 (36 mL);
TFA (9 mL) was added and the reaction mixture was
stirred at room temperature overnight. After evaporation
to dryness, the crude material was taken up with CH2Cl2
and washed with 5% NaHCO3, dried over MgSO4,
filtered and evaporated to dryness to yield 0.826 g of a
mixture of Boc-deprotected X1A–X7A. This mixture was
split into seven equal portions (one for each sublibrary)
and K2CO3 (65 mg, 0.47 mmol) and acetyl chloride
(15.49 µL, 0.217 mmol) were added to one of these
portions, dissolved in CH2Cl2 (15 mL). After stirring
overnight, the reaction was quenched with H2O (5 mL),
the organic layer was separated, washed with H2O, dried
over MgSO4, filtered and evaporated to dryness to yield
75 mg (74%) of the title sublibrary. All seven compounds
forming the sublibrary were characterised by LC/MS.

834
Purity of the seven sublibraries of set 2, calculated as
the sum of the areas of the seven peaks of the LC/UV
chromatogram attributed by LC/MS to the seven com-
pounds forming each sublibrary, is reported below:
(m, 4H), 2.90–2.81 (m, 2H), 2.45 (m, 1H), 2.20 (m, 2H),
2.15–2.00 (m, 1H), 2.00–1.60 (m, 9H), 1.30–1.10 (m,
2H). ESI-MS (positive, solvent: methanol, spray 4.5 keV,
skimmer: 60 eV, capillary 220 °C) m/z 595 (MH+). Anal.
C34H41Cl3N2O3 (C, H, N, Cl).
X1–7AY1 98%; X1–7AY2 94%; X1–7AY3 98%;
X
1–7AY4 83%; X1–7AY5 98%; X1–7AY6 94%; X1–7AY7
98%.
6.6.3. N-benzoyl-3-(3,4-dichlorophenyl)-3-[3-(4-phenyl-
piperazin-1-yl)propyl]piperidine dihydrochloride X2AY2
IR (KBr) 3 447, 2 947, 2 403, 1 614, 1 440 cm–1.
1H-NMR (CDCl3–333 K) δ 7.42–7.20 (m, 10H), 6.90 (d,
2H), 6.80 (dd, 1H), 4.25 (m, 1H), 3.53 (d, 1H), 3.53–3.30
(m, 2H), 3.15 (m, 4H), 2.45 (m, 4H), 2.24 (t, 2H),
2.18–2.08 (m, 1H), 1.88 (ddd, 1H), 1.79–1.42 (m, 4H),
1.40–1.10 (m, 2H). ESI-MS (positive, solvent: methanol,
spray 4.5 keV, skimmer: 60 eV, capillary 220 °C) m/z 536
(MH+). Anal. C31H37Cl4N3O (C, H, N, Cl).
6.6. Synthesis of individual compounds reported in
table II
6.6.1. Synthesis of N-benzoyl-3-(3,4-dichlorophenyl)-3-
[3-(4-phenylpiperidin-1-yl)propyl]piperidine hydrochloride
X1AY2
The synthesis of this compound is reported as a general
procedure for the synthesis of all six individual com-
pounds prepared. N-tert-Butoxycarbonyl-3-(3,4-di-
chlorophenyl)-3-[3-(4-phenylpiperidin-1-yl)propyl]
piperidine X1A (0.23 g, 0.43 mmol) was dissolved in
CH2Cl2 (8 mL); TFA (2.2 mL), dissolved in CH2Cl2
(2 mL), was added, and the reaction was stirred at room
temperature overnight. After evaporation to dryness, the
crude material was taken up with CH2Cl2 and washed
with 5% NaHCO3, dried over MgSO4, filtered and
evaporated to dryness to yield crude 3-(3,4-
dichlorophenyl)-3-[3-(4-phenylpiperidin-1-yl)propyl]
piperidine. Half of this crude material (≈ 0.21 mmol) was
dissolved in CH2Cl2 (5 mL), K2CO3 (80 mg, 0.58 mmol)
and benzoyl chloride (24.4 µL, 0.21 mmol) were added
and the reaction was stirred at room temperature over-
night and then quenched with H2O (2 mL). The organic
layer was separated, washed with H2O, dried over
MgSO4, filtered and evaporated to dryness to yield
140 mg (61%) of N-benzoyl-3-(3,4-dichlorophenyl)-3-
[3-(4-phenylpiperidin-1-yl)propyl]piperidine as a yellow
oil. This compound was transformed into its hydrochlo-
ride by dissolving in MeOH and treating with HCl/Et2O.
Evaporation to dryness and trituration with Et2O afforded
the title compound as a solid. IR (KBr) 3 425, 2 937,
6.6.4. N-benzoyl-3-(3,4-dichlorophenyl)-3-{3-[4-(pyrrol-
idinocarbonylmethyl)piperazin-1-yl]propyl}piperidine
dihydrochloride X7AY2
IR (KBr) 3 436, 2 954, 2 437, 1 662, 1 618,
1
1 438 cm–1. H-NMR (CDCl3–333K) δ 7.40–7.10 (m,
8H), 4.40 (m, 1H), 3.52–3.30 (m, 7H), 3.09 (s, 2H), 2.51
(m, 4H), 2.31 (m, 4H), 2.18 (t, 2H), 2.15–2.05 (m, 1H),
1.95–1.80 (m, 5H), 1.70–1.40 (m, 4H), 1.35–1.05 (m,
2H). ESI-MS (positive, solvent: methanol, spray 4.5 keV,
skimmer: 60 eV, capillary 220 °C) m/z 571 (MH+). Anal.
C31H42Cl4N4O2 (C, H, N, Cl).
6.6.5.
N-phenylacetyl-3-(3,4-dichlorophenyl)-3-{3-[4-
(pyrrolidinocarbonylmethyl)piperazin-1-yl]propyl}piperidine
dihydrochloride X7AY3
IR (KBr) 3 435, 2 954, 2 546, 1 653, 1 630,
1
1 451 cm–1. H-NMR (CDCl3–333 K) δ 7.40–7.05 (m,
8H), 4.40 (m, 1H), 3.70 (s, 2H), 3.49 (m, 5H), 3.23 (m,
2H), 3.09 (s, 2H), 2.52 (m, 4H), 2.31 (m, 4H), 2.11 (t,
2H), 2.10–2.00 (m, 1H), 1.98–1.75 (m, 4H), 1.75–1.00
(m, 7H). ESI-MS (positive, solvent: methanol, spray 4.5
keV, skimmer: 60 eV, capillary 220 °C) m/z 585 (MH+).
Anal. C32H44Cl4N4O2 (C, H, N, Cl).
2 532, 1 622, 1 444 cm–1. H-NMR (CDCl3–333 K) δ
1
7.40–7.10 (m, 13H), 4.25 (m, 2H), 3.58–3.30 (m, 4H),
2.90–2.81 (m, 2H), 2.45 (m, 1H), 2.22 (t, 2H), 2.20–2.09
(m, 1H), 2.00–1.55 (m, 9H), 1.35–1.10 (m, 2H). ESI-MS
(positive, solvent: methanol, spray 4.5 keV, skimmer: 60
eV, capillary 220 °C) m/z 535 (MH+). Anal.
C32H37Cl3N2O (C, H, N, Cl).
6.6.6.
N-benzenesulfonyl-3-(3,4-dichlorophenyl)-3-{3-
[4-(pyrrolidinocarbonylmethyl)piperazin-1-yl]propyl}
piperidine dihydrochloride X7AY7
IR (KBr) 3 427, 2 943, 2 582, 1 658, 1 469, 1 340,
1
1 162 cm–1. H-NMR (CDCl3–333 K) δ 7.78 (m, 2H),
6.6.2. N-(2,4-dimethoxy)benzoyl-3-(3,4-dichlorophenyl)-
7.55 (m, 3H), 7.40 (m, 2H), 7.21 (m, 1H), 3.47 (m, 5H),
3.10 (m, 1H), 3.10 (s, 2H), 2.79 (m, 2H), 2.51 (m, 4H),
2.32 (m, 4H), 2.17 (t, 2H), 2.00–1.50 (m, 10H), 1.30–1.10
(m, 2H). ESI-MS (positive, solvent: methanol, spray 4.5
keV, skimmer: 60 eV, capillary 220 °C) m/z 607 (MH+).
Anal. C30H42Cl4N4O3S (C, H, N, Cl).
3-[3-(4-phenylpiperidin-1-yl)propyl]piperidine
hydro-
chloride X1AY5
IR (KBr) 3 447, 2 939, 2 668, 1 607, 1 467 cm–1.
1H-NMR (CDCl3–333 K) δ 7.50–7.20 (m, 9H),
6.50–6.40 (m, 2H), 4.25 (m, 2H), 3.80 (s, 6H), 3.70–3.10

835
[8] Harrison T., Korsgaard M.P.G., Swain C.J., Cascieri M.A., Sadowski
S., Seabrook G.R., Bioorg. Med. Chem. Lett. 8 (1998) 1343–1348.
Acknowledgements
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Stylli C.H., Banks M.N., Tiller P., Bioorg. Med. Chem. Lett. 4
(1994) 2821–2824.
The authors wish to thank Dr Alberto Cerri for NMR
spectroscopic determinations.
[10] Pirrung M.C., Chen J., J. Am. Chem. Soc. 117 (1995) 1240–1245.
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[12] Giardina G.A.M., Grugni M., Rigolio R., Vassallo M., Erhard K.,
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[14] (a) Calculated utilising the average molecular weight. (b) Calculated
as the sum of the areas of the seven peaks of the LC/UV
chromatogram, attributed by LC/MS to the seven compounds
forming each mixture.
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