Structure-activity relationships in 1,4-benzodioxan-related compounds. 11.(1) reversed enantioselectivity of 1,4-dioxane derivatives in α1-adrenergic and 5-HT1A receptor binding sites recognition

Article abstract of DOI:10.1021/jm301525w

5-HT_1A receptor and α₁-adrenoreceptor (α₁-AR) binding sites recognized by the 1,4-dioxanes 2-4 display reversed stereochemical requirements. (S)-2 proved to be a potent 5-HT_1A receptor agonist highly selective over α₁-AR subtypes. Chirality influenced the anticancer activity of 3 and 4 in human prostate cancer cells (PC-3): (R)-4, eutomer at the α_1d-AR subtype, was the most potent. The decreased effect of 4 and (R)-4 in α_1d-AR silenced PC-3 cells confirmed that their anticancer activity was α_1d-AR-dependent.

Full text of DOI:10.1021/jm301525w

Brief Article
pubs.acs.org/jmc
Structure−Activity Relationships in 1,4-Benzodioxan-Related
Compounds. 11.1 Reversed Enantioselectivity of 1,4-Dioxane
Derivatives in α1‑Adrenergic and 5‑HT1A Receptor Binding Sites
Recognition
Alessandro Bonifazi,† Alessandro Piergentili,† Fabio Del Bello,† Yogita Farande,† Mario Giannella,†
Maria Pigini,† Consuelo Amantini,‡ Massimo Nabissi,‡ Valerio Farfariello,‡ Giorgio Santoni,‡
Elena Poggesi,§ Amedeo Leonardi,§ Sergio Menegon,§ and Wilma Quaglia*,†
†Medicinal Chemistry Unit, School of Pharmacy, University of Camerino, Via S. Agostino 1, 62032 Camerino, Italy
‡Experimental Medicine Section, School of Pharmacy, University of Camerino, Via Madonna delle Carceri 9, 62032 Camerino, Italy
§Recordati S.p.A., Drug Discovery, Via Civitali 1, 20148 Milano, Italy
S
* Supporting Information
ABSTRACT: 5-HT1A receptor and α1-adrenoreceptor (α1-AR) binding sites recognized by the 1,4-dioxanes 2−4 display
reversed stereochemical requirements. (S)-2 proved to be a potent 5-HT1A receptor agonist highly selective over α1-AR subtypes.
Chirality influenced the anticancer activity of 3 and 4 in human prostate cancer cells (PC-3): (R)-4, eutomer at the α1d-AR
subtype, was the most potent. The decreased effect of 4 and (R)-4 in α1d-AR silenced PC-3 cells confirmed that their anticancer
activity was α1d-AR-dependent.
Chart 1. Structures of WB 4101 (1) and 2−4
α1-Adrenoreceptors (α1-ARs), belonging to G-protein-coupled
receptor (GPCR) family A, are subdivided into three
pharmacologically distinct subtypes (α1a/A, α1b/B, α1d/D), with
upper and lower case subscript designating the native or
recombinant receptors, respectively.2 α1A- and α1B-AR subtypes
play an important role in cardiac development and/or function
as well as in blood pressure via vasoconstriction.3 Moreover, the
α1A-AR subtype, dominant in the prostate, bladder neck, and
urethra, together with the α1D-AR subtype, located in the
bladder or the spinal cord, mediates lower urinary tract
symptoms (LUTS) caused by benign prostatic hyperplasia
(BPH).4 In addition to their smooth-muscle-relaxing effects, α1-
AR antagonists inhibit the primary tumor growth and
progression to metastasis in human prostate cancer.5 Like the
α1-ARs, 5-HT1A receptor, a member of the serotonin 5-HT1
class, is a GPCR.6 In the past decade, the 5-HT1A subtype has
been the most studied as a major target for agents
therapeutically useful not only in well-established areas such
as depression and anxiety but also in recent perspectives such as
neuroprotection,6 cognition disorders,7 and in pain relief
therapy.8 It is well documented that the transmembrane
amino acid sequence of 5-HT1A receptor exhibits a notably high
degree of homology to α1-ARs (approximately 45%).9 There-
fore, several classes of compounds show high affinity for both
receptor systems, poor specificity, and consequently, adverse
effect problems. Among these classes, the benzodioxane
derivatives are effective ligands of both receptor systems,10,11
and the potent and non-subtype-selective α1-antagonist WB
4101 (1) (Chart 1) is also an effective partial agonist of 5-HT1A
receptor.12
of 5-HT1A receptor and α1-ARs. We have recently demon-
strated that the replacement of the quite planar 1,4-
benzodioxane template of 1 with the less conformationally
constrained 6,6-diphenyl-1,4-dioxane moiety13 afforded 2
(Chart 1), which was able to discriminate between the two
systems, maintaining the high 5-HT1A affinity of 1 but
displaying markedly reduced affinities at all three α1-AR
subtypes. The elimination of one or both ortho-methoxy
substituents of 2, affording 3 and 4 (Chart 1), respectively,
though slightly reducing intrinsic activity, was compatible with
the maintenance of the high affinity for 5-HT1A receptor.
Interestingly, such a modification conferred to these ligands the
ability to potently and preferentially interact with the α1d-AR
subtype with respect to α1a- and α1b-AR subtypes. Compounds
2−4 behaved as antagonists at all three α1-AR subtypes and as
full or partial agonists at 5-HT1A receptor.13 Tested in human
PC-3 prostate cancer cells, they also showed antiproliferative
and cytotoxic activity: 3 and especially 4 proved to be the most
potent, probably owing to their high α1D-AR antagonism.13
Moreover, since serotonin induces proliferation in PC-3 cells,
Many efforts have been devoted to understand the structural
Received: July 27, 2012
requirements of ligands useful to differentiate the binding sites
Published: December 20, 2012
© 2012 American Chemical Society
584
dx.doi.org/10.1021/jm301525w | J. Med. Chem. 2013, 56, 584−588

Journal of Medicinal Chemistry
Brief Article
we demonstrated that the 5-HT1A antagonist (S)-WAY 100135
did not affect the antiproliferative and cytotoxic effects of 2−4,
indicating that their potent 5-HT1A agonist activity was not in
contrast to their anticancer activity.13 Considering that the
interactions with α1-ARs and 5-HT1A receptor are well-known
to be highly stereospecific and stereochemistry can influence
quantitatively and qualitatively the biological profile of ligands,
the enantiomers of 2−4 were prepared and studied to
investigate whether a definite configuration might enhance
the discrimination between the two receptor systems. In
addition, the study of enantioselectivity of chiral 2−4 might
allow us to determine whether their anticancer activity is
triggered by a specific or nonspecific mechanism of action.
Therefore, the evaluation of the cytotoxic activity of the
enantiomers of 2−4 in PC-3 prostate cancer cells and of 4 and
its enantiomers in human PC-3 cells transfected with a control
(siGLO) or silenced α1d-AR gene (siα1d-AR) might allow us to
confirm the previously hypothesized involvement of α1d-AR in
this activity.13,14
analysis of data showed that the configuration of the
stereocenter in position 2 of the 1,4-dioxane nucleus of 2−4
affected affinity and selectivity at α1-ARs and 5-HT1A receptor.
For the interaction with the α1-AR system, (R)-2, (R)-3, and
(R)-4, showing pKi values not significantly different from those
of the corresponding racemic compounds, were the eutomers at
the three α1-AR subtypes. Considering that the enantiomeric
purity of all the enantiomers is 100% with the exception of (S)-
(−)-4, whose value is 98.7%, the eudismic ratios (ER) are >6, 8,
and >13 for the first pair, 15, 19, and >18 for the second pair,
and 3, 3, and >26 for the third pair. In contrast, in the case of
WB 4101 (1) and many of its analogues,17 (S)-enantiomers
were significantly more potent than (R)-enantiomers at α1-ARs.
This observation seems to confirm that, as we previously
hypothesized by structure−activity relationships and docking
studies,13 the 1,4-dioxanes 2−4 do not interact with the same
α1-AR binding sites recognized by 1. Concerning 5-HT1A
receptor, similar to 1 and unlike what was observed for the
α1-AR interaction, the higher affinity resided in the (S)-
configured enantiomers. However, only in the pair of
enantiomers (S)-2/(R)-2, in which the phenoxy group is 2,6-
dimethoxy-substituted, (S)-2 was significantly more affine than
(R)-2 and the eudismic ratio (ER = 81) assumed a high value.
Interestingly, since the eutomers at 5-HT1A receptor were
distomers at α1-AR, a reversal of enantioselectivity was
observed. Consequently, the selectivity ratio 5-HT1A/α1-AR
considerably increased compared to the corresponding racemic
compounds. In particular, (S)-2 showed significantly high 5-
HT1A/α1-AR subtype affinity ratios (5-HT1A/α1a > 1820; 5-
HT1A/α1b = 3020; 5-HT1A/α1d = 1318), which were unusual
because of the notably high degree of homology existing
between these two receptor systems. This result suggests that
the binding sites of the two receptor systems recognized by the
1,4-dioxane derivatives have different stereochemical require-
ments and the (S) configuration is a structural requirement
crucial for the design of novel compounds able to selectively
recognize the 5-HT1A receptor over α1-ARs. Moreover, (S)-2
might be a pharmacologically useful tool to fully activate the 5-
HT1A receptor with reduced hypotensive side effects due to its
low affinities for α1-AR subtypes. The agonist efficacy,
expressed as pD2, of enantiomers (R)- and (S)-2−4 toward
the 5-HT1A receptor was assessed by determining the induced
stimulation of [35S]GTPγS binding in cell membranes from
HeLa cells, using the full agonist 5-hydroxytryptamine (5-HT)
as reference compound (Table 1).13 Data showed that
enantiomers (S)-2−4 were significantly more potent than
(R)-enantiomers (ER = 263, 31, and >23, respectively), with
pD2 values similar to those of the corresponding racemic
compounds and significantly higher than those of the full
agonists 8-OH-DPAT and 5-HT. Moreover, the configuration
of the stereocenter in position 2 proved to be crucial for the
potency but not for the intrinsic activity, both enantiomers of
the three pairs behaving as full [(R)- and (S)-2] or partial
agonists [(R)- and (S)-3; (R)- and (S)-4].
RESULTS AND DISCUSSION
■
To prepare the novel enantiomers (R)- and (S)-2−4, alcohols
(R)- and (S)-515 were treated with mesyl chloride in pyridine to
yield (S)- and (R)-6, respectively, which were aminated with
the suitable amines, 2-(2,6-dimethoxyphenoxy)ethanamine,16
2-(2-methoxyphenoxy)ethanamine, or 2-phenoxyethanamine
(Scheme 1). Enantiomeric purity of amines (R)-(+)- and (S)-
a
Scheme 1
a
Reagents: (a) MsCl, pyridine; (b) 2-(2,6-dimethoxyphenoxy)-
ethanamine or 2-(2-methoxyphenoxy)ethanamine or 2-phenoxyethan-
amine, CH3CH2OCH2CH2OH, Δ.
(−)-2−4, determined by HPLC and by 1H NMR spectroscopy
on addition of the chiral shift reagent (R)-(+)-α-methoxy-α-
(trifluoromethyl)phenylacetic acid [(+)-MTPA] in comparison
with the spectrum of racemic ( )-2−4, was found to be 100%
for all the enantiomers with the exception of (S)-(−)-4, whose
1
value is 98.7%. The HPLC chromatograms and H NMR
spectra of ( )-, (R)-(+)-, and (S)-(−)-4 in the presence of
(+)-MTPA are reported in Supporting Information.
The binding profiles of (R)- and (S)-2−4 were evaluated
using [3H]prazosin and [3H]8-hydroxy-2-(di-n-propylamino)-
tetralin ([3H]8-OH-DPAT) to label cloned human α1-ARs
expressed in Chinese hamster ovary (CHO) cells and 5-HT1A
receptors expressed in HeLa cells, respectively. The potent α1d-
AR antagonist 8-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-
8-azaspiro[4.5]decane-7,9-dione (BMY-7378) and the 5-HT1A
receptor full agonist 8-OH-DPAT were used as reference
compounds.13 The affinity values, expressed as pKi, are
reported in Table 1 and compared with those of WB 4101
(1) and its enantiomers,17 included for useful comparison. An
The in vitro cytotoxic activity of (R)- and (S)-2−4 and 1 in
human PC-3 cells, using 1-(4-amino-6,7-dimethoxy-2-quinazo-
linyl)-4-(1,4-benzodioxan-2-ylcarbonyl)piperazine (doxazo-
sin)18 as reference compound, was carried out using the
sulforhodamine B (SRB) assay according to the National
Cancer Institute protocol, as previously reported.13 All
compounds were active at low micromolar concentration
(Table 2) and chirality affected the antiproliferative and
cytotoxic activity of 3 and 4, highlighting a specific mechanism
585
dx.doi.org/10.1021/jm301525w | J. Med. Chem. 2013, 56, 584−588

Journal of Medicinal Chemistry
Brief Article
Table 1. Affinity Constants (pKi) of 1−4 and Their Enantiomers, BMY-7378, and 8-OH-DPAT for Human Recombinant α1-AR
a
b
c
Subtypes and 5-HT1A Receptor and Agonist Efficacy (pD2) and Relative Efficacy (Emax) on 5-HT1A Receptor in Comparison
to BMY-7378, 8-OH-DPAT, and 5-HT
pKi, human cloned receptor
binding [35S]GTPγS
5-HT1A, pD2 Emax, %
compd
( )-2
R
R1
α1a
α1b
α1d
5-HT1A
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
H
OCH3
OCH3
OCH3
H
6.47
6.78
<6
6.49
6.71
5.78
7.25
7.66
6.38
6.92
7.28
6.77
8.00
7.14
8.24
6.15
<6
7.18
7.25
6.14
8.94
8.86
7.60
8.44
8.49
7.08
9.29
7.98
9.29
8.89
<6
8.85
7.35
9.26
9.18
9.22
9.51
9.23
8.83
9.40
8.68
7.39
8.61
9.43
8.47
8.28
5.88
8.30
9.40
7.70
9.19
9.11
7.82
9.19
106.3
119.5
103.6
81.5
66.6
61.9
77.1
72.7
66.0
(R)-2
(S)-2
( )-3
7.56
7.78
6.59
6.77
6.89
6.36
9.37
7.95
9.39
6.42
<6
(R)-3
H
(S)-3
H
( )-4
H
(R)-4
H
H
(S)-4
H
H
WB 4101 (1)
(R)-1
(S)-1
BMY-7378
8-OH-DPAT
5-HT
9.27
7.60
7.30
26.0
100
100
a
Equilibrium dissociation constants (Ki) were derived from IC50 values using the Cheng−Prusoff equation.20 Each experiment was performed in
b
triplicate. Ki values were from two to three experiments, which agreed within 20%. pD2 values are the negative logarithm of the agonist
concentration required to obtain 50% of the maximal stimulation of [35S]GTPγS binding and were calculated from two to three experiments, which
c
agreed within 20. Maximal stimulation is expressed as a percentage of the maximal 5-HT response.
Table 2. Cytotoxic Activity of 2−4, Their Enantiomers, and
WB 4101 (1) in Comparison to Doxazosin in PC-3 Cells and
Cytotoxic Activity of 4 and Its Enantiomers in siGLO- and
(R)-4 exhibiting the highest potency. This observation
supported the previously hypothesized relationship between
the interaction with α1d-AR subtype and anticancer activity.13
To confirm such a hypothesis, cytotoxic activity of 4 and its
enantiomers in human PC-3 cells transfected with siGLO or
siα1d-AR was evaluated. Silencing of α1d-AR significantly
reduced the anticancer activity of 4 and its enantiomer (R)-4
(Table 2). In particular, for (R)-4 GI50 increased from 0.6 to
10.2 μM (17-fold), TGI from 2.1 to 27.1 μM (12.9-fold), and
LC50 from 4.0 to 52.1 μM (13-fold). No significant differences
in cytotoxic activity were found comparing siGLO-transfected
with untransfected cells. Moreover, silencing of α1d-AR
significantly inhibited (about 76%) the noradrenaline
(−)-NA-induced proliferation compared to siGLO-transfected
PC-3 control cells (Figure 1C in Supporting Information),
whereas no changes were found by comparing siGLO-
transfected and untransfected PC-3 prostate cancer cells.
These results strongly confirm the hypothesis that the
antiproliferative and cytotoxic effects of 4 and (R)-4 are α1d-
AR-dependent. Moreover, it is known that antidepressant drugs
can also be prescribed to prevent symptoms of depression in
prostate cancer patients.19 Therefore, the potentially beneficial
antidepressant and antianxiety effects, due to the agonism at 5-
HT1A receptor, and the reduced hypotensive side effects, due to
the low affinities for α1a and α1b-AR subtypes, make (R)-4 a
promising lead to be optimized for the development of α1d-AR
antagonists potentially useful in the treatment of human
prostate cancer.
a
α1d-AR-Silenced PC-3 Cells
compd
( )-2
PC-3 cell
GI50, μM
TGI, μM
LC50, μM
b
c
c
c
c
c
UT
14.0 0.7
32.0 1.3
16.0 0.9
32.0 1.3
58.0 1.6
30.0 0.6
16.0 0.9
9.0 0.6
45.0 1.2
65.0 1.6
37.0 1.5
45.0 1.4
29.0 0.8
67.0 1.6
b
(R)-2
(S)-2
( )-3
(R)-3
(S)-3
( )-4
UT
b
UT
b
c
UT
9.0 0.8
b
UT
4.0 0.3
b
UT
21.0 1.2
34.0 1.5
b
c
c
c
UT
3.0 0.2
6.0 0.7
8.0 0.5
siGLO
4.1 0.8
7.1 0.6
25.0 0.8
2.1 0.1
2.5 0.4
27.1 0.3
43.0 0.9
41.7 1.3
49.2 0.9
199 1.6
49.0 2.5
10.7 0.5
39.8 0.9
4.0 0.2
5.8 0.2
52.1 1.1
>300
siα1d-AR
23.6 1.2
0.6 0.07
0.9 0.5
b
(R)-4
(S)-4
UT
siGLO
siα1d-AR
10.2 0.6
19.1 1.1
22.8 0.4
20.7 0.7
64.6 0.4
26.9 1.3
b
UT
siGLO
>300
siα1d-AR
>300
b
WB 4101 (1)
UT
>200
b
doxazosin
UT
75.8 3.6
a
Each quoted value represents the mean of quadruplicate determi-
nations standard error (n = 5). Statistical analysis was performed by
comparing the anticancer effect induced by enantiomers (R)- and (S)-
2−4 with respect to the corresponding racemic compounds and by
( )-4, (R)-4, and (S)-4 in α1d-AR-silenced with respect to siGLO-
b
c
transfected PC-3 cells. UT = untransfected. This value is not
significantly different from that previously reported.13
In conclusion, in the present study we demonstrated that the
binding sites of 5-HT1A receptor and α1-ARs recognized by the
1,4-dioxanes 2−4 possessed reversed stereochemical require-
ments. Other novel derivatives and molecular modeling studies,
of action. Interestingly, the eutomers at the α1d-AR subtype
[(R)-3 and (R)-4] were also efficient cytotoxic agents, with
586
dx.doi.org/10.1021/jm301525w | J. Med. Chem. 2013, 56, 584−588

Journal of Medicinal Chemistry
Brief Article
objects of a forthcoming paper, might confirm and justify such
an interesting result. (S)-2 proved to be a potent 5-HT1A
receptor full agonist highly selective over α1-AR subtypes. (R)-3
and (R)-4, eutomers at the α1d-AR subtype, also displayed the
best antiproliferative and cytotoxic effects. The critical role of
chirality in the anticancer activity and the decreased
antiproliferative and cytotoxic effects of 4 and (R)-4 in α1d-
AR silenced PC-3 cells strongly confirmed the involvement of
the α1d-AR subtype in their anticancer properties.
ASSOCIATED CONTENT
* Supporting Information
■
S
Elemental analysis results for enantiomers (R)- and (S)-2−4;
experimental and spectroscopic details for (R)-(+)-3, (S)-
(−)-3, (R)-(+)-4, and (S)-(−)-4; HPLC chromatograms and
1H NMR spectra of ( )-, (R)-(+)-, and (S)-(−)-4 in the
presence of (+)-MTPA; experimental details of binding,
cytotoxic assays, cell proliferation in siGLO- and siα1d-AR-
transfected PC-3 cells treated with (−)-NA (Figure 1C);
quantitative real time PCR (qRT-PCR) and Western blot
analysis demonstrating the silencing of α1d-AR gene in PC-3
cells (Figure 1A,B). This material is available free of charge via
the Internet at http://pubs.acs.org.
EXPERIMENTAL SECTION
General. Melting points were taken in glass capillary tubes on a
Buchi SMP-20 apparatus and are uncorrected. H NMR spectra were
■
̈
1
recorded on a Varian EM-390 instrument. The microanalyses were
performed by the microanalytical laboratory of our department. The
elemental composition of the compounds agreed to within 0.4% of
the calculated value. Optical activity was measured at 20 °C with a
Perkin-Elmer 241 polarimeter. Chromatographic separations were
performed on silica gel columns (Kieselgel 40, 0.040−0.063 mm,
Merck) by flash chromatography. Chemical names were generated
using ChemDraw Ultra (CambridgeSoft, version 9.0). The enantio-
meric purity, determined by HPLC, was found to be 100% for all the
enantiomers with the exception of (S)-(−)-4, whose value is 98.7%.
Instruments used were the following: a gradient capable separation
system fitted with a UV or DAD detector; a Waters alliance 2695
HPLC instrument comprising a quaternary pump, Waters 2996 DAD,
an automatic injector, a degassing system. Column was Daicel
Chiralcel OJ-H 5 μm, 250 mm × 4.6 mm, part no. OJH0CE-FA025.
Elution conditions were the following: solvent, MeOH for ( )-3, (R)-
3, (S)-3, ( )-4, (R)-4, (S)-4; 90% MeOH/10% 2-PrOH for ( )-2,
(R)-2, (S)-2; gradient table, isocratic 100% MeOH; duration, 35 min;
flow, 0.6 mL/min; 18 °C; detection at λ = 205 nm. The purity of the
novel compounds, determined by combustion and HPLC analysis, was
>95%. The procedures for the synthesis of (R)-(+)- and (S)-(−)-2 are
reported below.
AUTHOR INFORMATION
Corresponding Author
*Phone: +390737402237. Fax: +390737637345. E-mail: wilma.
quaglia@unicam.it.
■
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
We thank the MIUR (Rome), the University of Camerino, and
the Monte dei Paschi di Siena Foundation for financial support.
■
ABBREVIATIONS USED
■
α1-AR, α1-adrenoreceptor; GPCR, G-protein-coupled receptor;
LUTS, lower urinary tract symptoms; BPH, benign prostatic
hyperplasia; siα1d-AR, silenced α1d-AR gene; CHO, Chinese
hamster ovary; SRB, sulforhodamine B; GI, growth inhibition;
TGI, total growth inhibition; LC, lethal concentration; Ki,
inhibition or dissociation constant; ER, eudismic ratio; qRT-
PCR, quantitative real time polymerase chain reaction; (−)-NA,
noradrenaline
(S)-(6,6-Diphenyl-1,4-dioxan-2-yl)methyl Methanesulfonate
[(S)-(+)-6]. Et3N (1.0 mL, 7.4 mmol) and mesyl chloride (0.85 g, 7.4
mmol) were added to a stirred solution of (R)-(+)-515 (1.0 g, 3.7
mmol) in CHCl3 (20 mL) at 0 °C. After 2 h at 0 °C, the mixture was
washed with H2O and dried over Na2SO4. After evaporation of the
solvent the residue was purified by column chromatography, eluting
with cyclohexane/EtOAc (9:1) to give (S)-(+)-6 as an oil: 1.2 g (93%
REFERENCES
■
(1) For part 10, see the following: Carrieri, A.; Piergentili, A.; Del
Bello, F.; Giannella, M.; Pigini, M.; Leonardi, A.; Fanelli, F.; Quaglia,
W. Structure−activity relationships in 1,4-benzodioxan-related com-
pounds. 10. Novel α1-adrenoreceptor antagonists related to open-
phendioxan: synthesis, biological evaluation, and α1d computational
study. Bioorg. Med. Chem. 2010, 18, 7065−7077.
(2) Langer, S. Z. History and nomenclature of α1-adrenoceptors. Eur.
Urol. 1999, 36 (Suppl. 1), 2−6.
(3) Tanoue, A.; Koshimizu, T.; Tsujimoto, G. Transgenic studies of
α1-adrenergic receptor subtype function. Life Sci. 2002, 71, 2207−
2215.
(4) Roehrborn, C. G.; Schwinn, D. A. α1-Adrenergic receptors and
their inhibitors in lower urinary tract symptoms and benign prostatic
hyperplasia. J. Urol. 2004, 171, 1029−1035.
(5) Desiniotis, A.; Kyprianou, N. Advances in the design and
synthesis of prazosin derivatives over the last ten years. Expert Opin.
Ther. Targets 2011, 15, 1405−1418.
(6) Nichols, D. E.; Nichols, C. D. Serotonin receptors. Chem. Rev.
2008, 108, 1614−1641.
(7) Schechter, L. E.; Dawson, L. A.; Harder, J. A. The potential utility
of 5-HT1A receptor antagonists in the treatment of cognitive
dysfunction associated with Alzheimer’s disease. Curr. Pharm. Des.
2002, 8, 139−145.
(8) Colpaert, F. C. 5-HT1A receptor activation: new molecular and
neuroadaptive mechanisms of pain relief. Curr. Opin. Invest. Drugs
2006, 7, 40−47.
(9) Trumpp-Kallmeyer, S.; Hoflack, J.; Bruinvels, A.; Hibert, M.
Modeling of G-protein-coupled receptors: application to dopamine,
yield); [α]20 +119.6 (c 1, CHCl3). 1H NMR (CDCl3) δ 3.10 (s, 3H,
D
CH3), 3.59−4.01 (m, 4H, CH2O, cycle), 4.32 (m, 2H, cycle), 4.61 (d,
1H, 5-CH), 7.22−7.57 (m, 10H, ArH).
(R)-(6,6-Diphenyl-1,4-dioxan-2-yl)methyl Methanesulfonate
[(R)-(−)-6]. This was prepared as described for (S)-(+)-6 starting from
(S)-(−)-5.15 An oil was obtained: 96% yield; [α]20 −118.9 (c 1,
D
1
CHCl3). The H NMR spectrum was identical to that of (S)-(+)-6.
(R)-2-(2,6-Dimethoxyphenoxy)-N-((6,6-diphenyl-1,4-dioxan-
2-yl)methyl)ethanamine [(R)-(+)-2]. A solution of (S)-(+)-6 (1.43
g, 4.1 mmol) and 2-(2,6-dimethoxyphenoxy)ethanamine16 (4.0 g, 20.5
mmol) in 2-ethoxyethanol (25 mL) was heated to reflux for 4 h.
Removal of the solvent under reduced pressure gave a residue, which
was dissolved in water. The aqueous solution was basified with NaOH
and extracted with CHCl3. Removal of dried solvents gave a residue,
which was purified by column chromatography, eluting with CHCl3 to
1
give (R)-(+)-2 as an oil: 0.66 g (36% yield). The H NMR spectrum
was identical to that of the corresponding racemic compound.13 The
free base was transformed into the oxalate salt, which was crystallized
from 2-PrOH; mp 151−153 °C; [α]20 +111.4 (c 1, MeOH). Anal.
D
(C27H31NO5·H2C2O4) C, H, N.
(S)-2-(2,6-Dimethoxyphenoxy)-N-((6,6-diphenyl-1,4-dioxan-
2-yl)methyl)ethanamine [(S)-(−)-2]. This was prepared as
1
described for (R)-(+)-2 starting from (R)-(−)-6: 25% yield. The H
NMR spectrum was identical to that of the corresponding racemic
compound.13 The free base was transformed into the oxalate salt,
which was crystallized from 2-PrOH; mp 151−153 °C; [α]20D −109.2
(c 1, MeOH). Anal. (C27H31NO5·H2C2O4·0.5H2O) C, H, N.
587
dx.doi.org/10.1021/jm301525w | J. Med. Chem. 2013, 56, 584−588

Journal of Medicinal Chemistry
Brief Article
adrenaline, serotonin, acetylcholine, and mammalian opsin receptors. J.
Med. Chem. 1992, 35, 3448−3462.
(10) Hibert, M. F.; Gittos, M. W.; Middlemiss, D. N.; Mir, A. K.;
Fozard, J. R. Graphics computer-aided receptor mapping as a
predictive tool for drug design: development of potent, selective and
stereospecific ligands for the 5-HT1A receptor. J. Med. Chem. 1988, 31,
1087−1093.
(11) Quaglia, W.; Pigini, M.; Piergentili, A.; Giannella, M.; Marucci,
G.; Poggesi, E.; Leonardi, A.; Melchiorre, C. Structure−activity
relationships in 1,4-benzodioxan-related compounds. 6. Role of the
dioxane unit on selectivity for α1-adrenoreceptor subtypes. J. Med.
Chem. 1999, 42, 2961−2968.
(12) Sharif, N. A.; Drace, C. D.; Williams, G. W.; Crider, J. Y. Cloned
human 5-HT1A receptor pharmacology determined using agonist
binding and measurement of cAMP accumulation. J. Pharm.
Pharmacol. 2004, 56, 1267−1274.
(13) Quaglia, W.; Piergentili, A.; Del Bello, F.; Farande, Y.; Giannella,
M.; Pigini, M.; Rafaiani, G.; Carrieri, A.; Amantini, C.; Lucciarini, R.;
Santoni, G.; Poggesi, E.; Leonardi, A. Structure−activity relationships
in 1,4-benzodioxan-related compounds. 9. From 1,4-benzodioxane to
1,4-dioxane ring as a promising template of novel α1D-adrenoreceptor
antagonists, 5-HT1A full agonists, and cytotoxic agents. J. Med. Chem.
2008, 51, 6359−6370.
(14) Quaglia, W.; Santoni, G.; Pigini, M.; Piergentili, A.; Gentili, F.;
Buccioni, M.; Mosca, M.; Lucciarini, R.; Amantini, C.; Nabissi, M. I.;
Ballarini, P.; Poggesi, E.; Leonardi, A.; Giannella, M. Structure−activity
relationships in 1,4-benzodioxan-related compounds. 8. {2-[2-(4-
Chlorobenzyloxy)phenoxy)ethyl}-[2-(2,6-dimethoxyphenoxy)ethyl]-
amine (clopenphendioxan) as a tool to highlight the involvement of
α1D- and α1B-adrenoreceptor subtypes in the regulation of human PC-
3 prostate cancer cell apoptosis and proliferation. J. Med. Chem. 2005,
48, 7750−7763.
(15) Del Bello, F.; Barocelli, E.; Bertoni, S.; Bonifazi, A.; Camalli, M.;
Campi, G.; Giannella, M.; Matucci, R.; Nesi, M.; Pigini, M.; Quaglia,
W.; Piergentili, A. 1,4-Dioxane, a suitable scaffold for the development
of novel M3 muscarinic receptor antagonists. J. Med. Chem. 2012, 55,
1783−1787.
(16) Woolley, D. W. Probable evolutionary relationship of serotonin
and indoleacetic acid, and some practical consequences therefrom.
Nature (London) 1957, 180, 630−633.
(17) Pallavicini, M.; Budriesi, R.; Fumagalli, L.; Ioan, P.; Chiarini, A.;
Bolchi, C.; Ugenti, M. P.; Colleoni, S.; Gobbi, M.; Valoti, E. WB4101-
related compounds: new, subtype-selective α1-adrenoreceptor antag-
onists (or inverse agonist?). J. Med. Chem. 2006, 49, 7140−7149.
(18) Cal, C.; Uslu, R.; Gunaydin, G.; Ozyurt, C.; Omay, S. B.
Doxazosin: a new cytotoxic agent for prostate cancer? BJU Int. 2000,
85, 672−675.
(19) Torta, R.; Leombruni, P.; Borio, R.; Castelli, L. Duloxetine for
the treatment of mood disorder in cancer patients: a 12-week case-
control clinical trial. Hum. Psychopharmacol. Clin. Exp. 2011, 26, 291−
299.
(20) Cheng, Y. C.; Prusoff, W. H. Relationship between the
inhibition constant (Ki) and the concentration of inhibitor which
causes 50% inhibition (I50) of an enzymatic reaction. Biochem.
Pharmacol. 1973, 22, 3099−3108.
588
dx.doi.org/10.1021/jm301525w | J. Med. Chem. 2013, 56, 584−588