Evaluation of structural effects on 5-HT2A receptor antagonism by aporphines: Identification of a new aporphine with 5-HT2A antagonist activity

Article abstract of DOI:10.1016/j.bmcl.2014.02.066

A set of aporphine analogs related to nantenine was evaluated for antagonist activity at 5-HT_2A and α_1A adrenergic receptors. With regards to 5-HT_2A receptor antagonism, a C2 allyl group is detrimental to activity. The chiral center of nantenine is not important for 5-HT_2A antagonist activity, however the N6 nitrogen atom is a critical feature for 5-HT_2A antagonism. Compound 12b was the most potent 5-HT_2A aporphine antagonist identified in this study and has similar potency to previously identified aporphine antagonists 2 and 3. The ring A and N6 modifications examined were detrimental to α_1A antagonism. A slight eutomeric preference for the R enantiomer of nantenine was observed in relation to α_1A antagonism.

Full text of DOI:10.1016/j.bmcl.2014.02.066

Bioorganic & Medicinal Chemistry Letters 24 (2014) 1664–1667
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Evaluation of structural effects on 5-HT2A receptor antagonism
by aporphines: Identification of a new aporphine with 5-HT2A
antagonist activity
Shashikanth Ponnala a, Junior Gonzales a,b, Nirav Kapadia a,b, Hernan A. Navarro c, Wayne W. Harding a,b,
⇑
a Chemistry Dept., Hunter College, CUNY, 695 Park Avenue, NY 10065, USA
b The Graduate Center, City University of New York, 365 5th Ave., NY 10016, USA
c Center for Organic and Medicinal Chemistry, Research Triangle Institute, Research Triangle Park, NC 27709, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
A set of aporphine analogs related to nantenine was evaluated for antagonist activity at 5-HT2A and a1A
adrenergic receptors.
With regards to 5-HT2A receptor antagonism, a C2 allyl group is detrimental to activity. The chiral
center of nantenine is not important for 5-HT2A antagonist activity, however the N6 nitrogen atom is a
critical feature for 5-HT2A antagonism.
Received 14 January 2014
Revised 19 February 2014
Accepted 24 February 2014
Available online 4 March 2014
Compound 12b was the most potent 5-HT2A aporphine antagonist identified in this study and has sim-
ilar potency to previously identified aporphine antagonists 2 and 3. The ring A and N6 modifications
examined were detrimental to a1A antagonism. A slight eutomeric preference for the R enantiomer of
nantenine was observed in relation to a1A antagonism.
Keywords:
Aporphine
Nantenine
5-HT2A
Ó 2014 Elsevier Ltd. All rights reserved.
a1A
Antagonist
Structure–activity relationship (SAR)
The tetracyclic aporphine template is a privileged scaffold that is
endowed with several biological activities.1–8 As central nervous
system (CNS) receptor ligands, aporphines have been found to
possess high affinity for a number of dopamine receptors (predom-
Several potent 5-HT2A receptor antagonists are known; in par-
ticular compounds with a mixed D2/5-HT2A antagonist profile
(eg., risperidone, clozapine) are quite prominent and are used clin-
ically to manage schizophrenic symptoms.22–24 However, there are
no highly selective 5-HT2A antagonists (>100-fold selectivity vs all
other common neuroreceptor targets) clinically available. Never-
theless, such promising compounds (eg., eplivanserin) have
recently been or are currently being investigated in clinical trials
as anti-insomnia medications. Thus, the identification of new
highly selective and therapeutically useful 5-HT2A receptor antag-
onists is still of topical interest.
Our research team has engaged structure–activity relationship
(SAR) studies on the aporphine alkaloid nantenine (1, Fig. 1) and
have identified a number of new nantenine analogs with antagonist
activity at the 5-HT2A receptor.25–28 We were guided by previous
SAR studies on nantenine for the present study. Nantenine itself
is a high affinity a1A adrenergic receptor antagonist with moderate
5-HT2A receptor antagonist potency (see Table 1). Compounds 2 and
3 (Fig. 1) are two of the most potent 5-HT2A antagonists we have ob-
tained to date. These compounds lack affinity for the a1A adrenergic
receptor. Our prior investigations have mainly focused on the ring A
portion of nantenine and in general have indicated a reasonable de-
gree of tolerance for other types and patterns of substitution in the
A ring in obtaining high 5-HT2A potency and selectivity versus the
inantly D1 and D2),9–12 serotonin (5-HT) receptors13–15 and
a-adren-
ergic receptors.6,16 Furthermore, aporphines are known with both
agonist and antagonist activity at neuroreceptor sites. The contin-
ued exploration of the aporphine template over the last few decades
has been driven to some extent by the opportunity/promise for
discovery of new ligands with high potency and selectivity for
subtypes of the above-mentioned receptors. Such molecules will
continue to supplement our toolbox of CNS receptor ligands that
will be useful as novel biological probes, as new imaging agents
and as leads for drug discovery efforts relevant to psychiatric disor-
ders and drug abuse.
We are primarily interested in evaluating the potential of apor-
phines as ligands for 5-HT2A receptors. 5-HT2A receptors are impli-
cated in several neuropsychiatric maladies including schizophrenia,
depression, anxiety and insomnia.17,18 5-HT2A receptors are also in-
volved in the actions of some stimulant drugs as recent reports have
revealed.19–21
⇑
Corresponding author. Tel.: +1 212 772 5359; fax: +1 212 772 5332.
E-mail address: whardi@hunter.cuny.edu (W.W. Harding).
http://dx.doi.org/10.1016/j.bmcl.2014.02.066
0960-894X/Ó 2014 Elsevier Ltd. All rights reserved.

S. Ponnala et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1664–1667
1665
2
MeO
MeO
MeO
O
We were interested in examining the extent to which an allyl
group would be accommodated at the C2 position since an allyloxy
group seems to be beneficial for antagonism (see data for com-
pound 2, Table 1). The synthetic practicability of obtaining this
structural feature via a Claisen rearrangement (a reaction rarely
employed with aporphines) supported this impetus.32
MeO
O
A
D
B
NMe
NMe
NMe
1
6
C
O
O
O
O
1
O
3
O
2
To obtain the required ring A analogs Scheme 1 was engaged.
Commercially available amine 4 was coupled to bromoacid 5 to
give amide 6. Bischler–Napieralski cyclization of 6 was followed
immediately by reduction of the dihydroisoquinoline thus formed
to the secondary amine 7. Protection of the amine as the N-ethyl
carbamate gave compound 8. Microwave-assisted direct aryla-
tion33 on 8 afforded compound 9. The benzyl ether 9 was deprotec-
ted revealing the phenol functionality in the key intermediate 10.
Reduction of 10 with lithium aluminium hydride (LAH) gave com-
pound 12a. Compound 12b was prepared from 10 via allylation to
afford compound 11 and subsequent LAH reduction. Claisen rear-
rangement of the allyl ether 11 provided compound 13. Reduction
of 13 gave the phenol analog 14. Analogs 15a–15c were prepared
from 13 in two steps via etherification and reduction as shown.
Nantenine enantiomers (R)-1 and (S)-1 were prepared by resolu-
tion of racemic nantenine as described previously.28 The isochro-
Figure 1. Structures of nantenine (1), compound 2 and compound 3.
a
1A receptor. However, the identity and optimal placement of sub-
stituents requires further research for maximal potency and
selectivity.
To continue to expand our understanding of the structural tol-
erance of aporphines as 5-HT2A receptor antagonists as well as
selectivity vs the a1A receptor, we have synthesized and evaluated
a new set of aporphine analogs. These compounds were prepared
in order to probe three regions in the northern portion of the apor-
phine template, namely: ring A, the chirality center and the nitro-
gen atom. As alluded to earlier, our previous studies have indicated
that substituents on the ring A moiety are important in controlling
the 5-HT2A receptor antagonist activity and selectivity. However,
we have not investigated the effect of the chiral center on 5-HT2A
antagonism before. This is an important task especially since the
existing literature suggests that aporphines exhibit a stereochem-
ical preference for activation of dopamine D1 and D229,30 receptors
as well as the related 5-HT1A receptor31—R enantiomers being pre-
dominantly agonists and S enantiomers being predominantly
antagonists in both cases. With regards to the effect of the N6
nitrogen atom, our prior studies suggest that a basic nitrogen atom
is required since the N-acetamide and N-methylsulfonamide deriv-
atives were devoid of activity.25 We sought herein to obtain exper-
imental proof of the absolute requirement for a nitrogen atom for
antagonist activity.
Along those lines we have: (1) synthesized and evaluated new
ring A analogs containing features of compounds 1, 2 or 3; (2) eval-
uated nantenine enantiomers—in order to begin to probe the effect
of the chirality center of aporphines on receptor antagonism; and
(3) synthesized and evaluated compounds that possess a nitro-
gen—oxygen isosteric replacement—to investigate the importance
of the nitrogen atom on receptor antagonism. Details of these stud-
ies are described henceforth.
man analogs 16a and 16b were prepared via
a sequence
involving oxa Pictet–Spengler cyclization and direct arylation as
presented in a recent report.34
All analogs were screened at 10 lM in multi-well format for
intrinsic (agonist) and antagonist activity at the human 5-HT2A
receptor using Fluorescence Imaging Plate Reader (FLIPR)-based
(Molecular devices, Sunnydale, CA) functional assays that detect
receptor-mediated mobilization of internal calcium with a calcium
sensitive fluorescent dye as reported previously.25–27 A similar set
of assays was performed for the
a1A—adrenergic receptor. Data
from these evaluations are presented in Table 1.
As shown in Table 1, compound 12a lacked any appreciable
activity at either receptor. The placement of an allyloxysubstitu-
tent at C2 (i.e., compound 12b) resulted in a significant increase
in 5-HT2A antagonist activity (>60-fold as compared to the parent
phenol 12a). Antagonism of the a1A receptor also increased
although the magnitude of this increase was less (4-fold increase
as compared to 12b). However, when 12b is compared to
nantenine (1), a significant decrease in a1A antagonist activity
was seen. The above data tends to suggest that the C2 methoxyl
Table 1
Ke values for analogs at 5-HT2A and a1A receptors
Compd
R1
R2
X
Ke SEMa (nM)
Selectivity 5-HT2A/a1A
5-HT2A
>3000
a1A
R2
12a
12b
H
Allyl
H
H
NMe
NMe
2950 457
744 74
<1.01
0.06
47
5
X
14
15a
15b
15c
16a
16b
H
Me
Allyl
Allyl
Allyl
Allyl
Allyl
OMe
OMe
OMe
OMe
OMe
OMe
OMe
—
NMe
NMe
NMe
NMe
O
>3000
>3000
566 112
>3000
>3000
>3000
—
R1O
485 123
1374 405
963 103
>3000
>3000
946 61
657 89
0.85
<0.45
<0.32
—
—
13.5
3.4
23.6
<0.007
<0.007
—
Cyclopropylmethyl
Me
Allyl
Me
Me
Me
Allyl
Cyclopropylmethyl
—
—
O
O
>3000
70 10
O
(R)-1
(S)-1
-(1)b
2c
NMe
NMe
NMe
NMe
NMe
—
196
36
3
850
6
7
70 15
>10000
>10000
1.1 0.4
—
3d
68
—
8
Prazosin
Ketanserinb,e
—
—
32
—
a
b
c
Values represent mean SEM for at least three independent experiments.
Ki, Data from Ref. 28.
Data from Ref. 27.
Data from Ref. 26.
d
e
IC50 determined in the presence of 5-HT EC80
.

1666
S. Ponnala et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1664–1667
b, c
a
d
NH
O
NH2
NH
NCO2Et
BnO
BnO
BnO
O
BnO
O
4
O
CO2H
O
O
O
Br
O
Br
O
Br
Br
5
6
7
8
h
e
NMe
NCO2Et
NCO2Et
NCO2Et
RO
O
HO
O
BnO
O
O
O
h
g
f
12
9
10
i
11
O
O
O
O
12a
12b
: R=H
: R=allyl
NMe
NCO2Et
NMe
RO
O
HO
HO
h
g, h
O
15
O
14
13
O
O
O
15a
: R = methyl
15b
15c
: R = allyl
: R=cyclopropylmethyl
Scheme 1. Synthesis of ring A analogs. Reagents and conditions: (a) 1,10-carbonyldiimidazole (CDI), THF, 0 °C-rt, 5 h, 80%; (b) trifluoromethanesulfonic acid, pyridine, DCM,
0 °C-rt, 4 h; (c) NaBH4, MeOH, 0 °C, 2 h, 88% over two steps; (d) ethyl chloroformate, K2CO3, DCM, rt, 3 h, 85%; (e) Pd(OAc)2, di-tert-butyl(methyl)phosphonium
tetrafluoroborate, K2CO3, (CH3)3CCOOH, DMSO, 135 °C, microwaves, 6 min, 50%; (f) H2/Pd, rt, 8 h, 95%; (g) alkyl bromide, KI, K2CO3, acetone, 70 °C, 6 h, 60–70%; (h) LAH, THF,
0 °C, 10 h, 50–60%; (i) N,N-diethylaniline, 215 °C, microwaves, 6 min, 90%.
substituent of nantenine is not required for 5-HT2A antagonist
activity.
enantiomer is slightly more potent. Interestingly, there seems to
be a reversal of this trend at the a1A receptor; the (R)-enantiomer
seems to be slightly more potent than the (S)-enantiomer at the
a1A receptor (approximately 3-fold).
Compound 14 lacked antagonist activity for both receptors indi-
cating that an allyl substituent at C2 is not well tolerated at either
receptor. A comparison of 15a with 1 reveals a slight improvement
in 5-HT2A antagonism but a decrease in a1A antagonist activity
upon replacement of the C2 methoxyl group with an allyl substitu-
ent. The antagonist activity of 15a was higher at both receptors
than compound 14 which is indicative of a greater tolerance for
an alkoxy substituent than a phenol at C1. Compound 15b had a
diminished affinity at both receptors as compared to 15a. When
15b is compared to compound 12b, a significant decrease in antag-
onist activity of 15b at both receptors manifests. This suggests that
an allyl substituent at C2 is not well accommodated at either
receptor. Compound 15c had activity and selectivity that was sim-
ilar to 15b. If a comparison of the 15b/15c pair is made with the 2/3
pair of compounds it may be surmised that C1 allyloxy and C1
cyclopropylmethyloxy groups endow the aporphine template with
very similar 5-HT2A antagonist potency irrespective of the identity
of the C2 substituent. That is, it appears that the allyl and cyclopro-
pylmethyl functionalities are bioisosteric with respect to 5-HT2A
receptor antagonism. From our previous studies, an allyloxy or
cyclopropylmethyloxy substituent (i.e., compounds 2 and 3 respec-
tively) imparted high 5-HT2A antagonist activity and selectivity to
the nantenine template. The analysis of compounds 15a, 15b and
15c showed a reversal in this trend and again points to a consider-
able lack of tolerance for a C2 allyl group at the 5-HT2A receptor.
Both compounds 16a and 16b (that lack the N6 moiety) were
devoid of antagonist activity. This supports previous SAR evidence
that a basic N6 atom is critical for affinity to both receptors. This is
also in line with previous molecular docking studies which suggest
that the protonated N6 atom is involved in a H-bonding interaction
with an aspartate residue in the 5-HT2A receptor binding pocket.25
Evaluation of (R)-1 and (S)-1 indicates that the chiral center of
nantenine is not critical for 5-HT2A antagonism although the (S)
In summary, this study has revealed some useful qualitative
information concerning the antagonism of aporphines at the 5-
HT2A receptor. The data suggest that the C2 position is not tolerant
of an allyl moiety. However, a C1 allyloxy substituent is well toler-
ated when the C2 substituent is hydrogen implying that the C2
methoxyl group of nantenine is not required for high 5-HT2A antag-
onist potency. This modification also improves selectivity vs the
a1A receptor (though this selectivity is moderate as compared to
that seen in 2 and 3). Of note, the most potent 5-HT2A aporphine
antagonist identified in this study was compound 12b which rivals
2 and 3 in terms of 5-HT2A antagonist potency.
The chiral center of nantenine does not engender any significant
preference for either enantiomer towards 5-HT2A antagonism.
Somewhat unsurprisingly, the N6 nitrogen is critical for antagonist
activity of nantenine analogs at both receptors.
The evaluation of this set of compounds has further expanded
our fundamental knowledge concerning the viability of the apor-
phine template for development as selective 5-HT2A receptor
antagonists. For certain, evaluation of larger series of analogs will
enable a better understanding of the extent to which the SAR infor-
mation extracted up to this point may be generalized. Compound
12b identified herein as well as compounds 2 and 3 identified ear-
lier, are useful starting points for further SAR exploration and opti-
mization studies. We are continuing in this vein and will report our
findings in due course.
Acknowledgments
This publication was made possible by Grant Numbers
1SC1GM092282 and G12RR003037 from the National Institutes
of Health. Its contents are solely the responsibility of the authors

S. Ponnala et al. / Bioorg. Med. Chem. Lett. 24 (2014) 1664–1667
1667
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Supplementary data
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all new compounds, procedure for biological assays and NMR spec-
tral data on all analogs) associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2014.02.
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