Synthesis of novel 5-substituted-2-aminotetralin analogs: 5-HT1A and 5-HT7 G protein-coupled receptor affinity, 3D-QSAR and molecular modeling

Article abstract of DOI:10.1016/j.bmc.2019.115262

The serotonin 5-HT₇ G protein-coupled receptor (GPCR) is a proposed pharmacotherapeutic target for a variety of central and peripheral indications, albeit, there are no approved drugs selective for binding 5-HT₇. We previously reported that a lead analog based on the 5-substituted-N,N-disubstituted-1,2,3,4-tetrahydronaphthalen-2-amine (5-substituted-2-aminotetralin, 5-SAT) scaffold binds with high affinity at the 5-HT₇ GPCR, and can treat symptoms of autism in mouse models; subsequently, the lead was found to have high affinity at the 5-HT_1A GPCR. Herein, we report the synthesis of novel 5-SAT analogs to develop a 3-dimensional quantitative structure—affinity relationship (3D-QSAR) at the human 5-HT₇ receptor for comparison with similar studies at the highly homologous 5-HT_1A receptor. We report 35 new 5-SAT ligands, some with very high affinity (K_i ≤ 1 nM) and stereoselectivity at 5-HT₇ + or 5-HT_1A receptors, several with modest selectivity (up to 12-fold) for binding at 5-HT₇, and, several ligands with high selectivity (up to 40-fold) at the 5-HT_1A receptor. 3D-QSAR results indicate that steric extensions at the C(5)-position improve selectivity for the 5-HT₇ over 5-HT_1A receptor, while steric and hydrophobic extensions at the chiral C(2)-amino position impart 5-HT_1A selectivity. In silico receptor homology modeling studies, supplemented with molecular dynamics simulations and binding free energy calculations, were used to rationalize experimentally-determined receptor selectivity and stereoselective affinity results. The data from these studies indicate that the 5-SAT chemotype, previously shown to be safe and efficacious in rodent paradigms of neurodevelopmental and neuropsychiatric disorders, is amenable to structural modification to optimize affinity at serotonin 5-HT₇ vs. 5-HT_1A GPCRs, as may be required for successful clinical translation.

Full text of DOI:10.1016/j.bmc.2019.115262

Journal Pre-proofs
Synthesis of novel 5-substituted-2-aminotetralin analogs: 5-HT1A and 5-HT7
G protein-coupled receptor affinity, 3D-QSAR and molecular modeling
Charles K. Perry, Austen B. Casey, Daniel E. Felsing, Rajender Vemula,
Mehreen Zaka, Noah B. Herrington, Meng Cui, Glen E. Kellogg, Clinton E.
Canal, Raymond G. Booth
PII:
S0968-0896(19)31605-0
https://doi.org/10.1016/j.bmc.2019.115262
BMC 115262
DOI:
Reference:
To appear in:
Bioorganic & Medicinal Chemistry
Received Date:
Revised Date:
Accepted Date:
19 September 2019
21 November 2019
9 December 2019
Please cite this article as: C.K. Perry, A.B. Casey, D.E. Felsing, R. Vemula, M. Zaka, N.B. Herrington, M. Cui,
G.E. Kellogg, C.E. Canal, R.G. Booth, Synthesis of novel 5-substituted-2-aminotetralin analogs: 5-HT1A and 5-
HT7 G protein-coupled receptor affinity, 3D-QSAR and molecular modeling, Bioorganic & Medicinal Chemistry
(2019), doi: https://doi.org/10.1016/j.bmc.2019.115262
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Synthesis of novel 5-substituted-2-
aminotetralin analogs: 5-HT1A and 5-HT7 G
protein-coupled receptor affinity, 3D-QSAR
and molecular modeling
Leave this area blank for abstract info.
Charles K. Perrya, , Austen B. Caseya, Daniel E. Felsinga, Rajender Vemulaa, Mehreen Zaka a, Noah B.
Herringtonb, Meng Cuia, Glen E. Kelloggb, Clinton E. Canala and Raymond G. Bootha
aCenter for Drug Discovery, Department of Pharmaceutical Sciences, Department of Chemistry & Chemical
Biology, Northeastern University, Boston, Massachusetts 02115, USA
bDepartment of Medicinal Chemistry & Institute for Structural Biology, Drug Discovery and Development,
Virginia Commonwealth University, Richmond, Virginia, 23298, USA

Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com
Synthesis of novel 5-substituted-2-aminotetralin analogs: 5-HT1A and 5-HT7 G
protein-coupled receptor affinity, 3D-QSAR and molecular modeling
Charles K. Perrya, , Austen B. Caseya, Daniel E. Felsinga, Rajender Vemulaa, Mehreen Zakaa, Noah B.
Herringtonb, Meng Cuia, Glen E. Kelloggb, Clinton E. Canala and Raymond G. Bootha
aCenter for Drug Discovery, Department of Pharmaceutical Sciences, Department of Chemistry & Chemical Biology, Northeastern University, Boston,
Massachusetts, 02115, USA
bDepartment of Medicinal Chemistry & Institute for Structural Biology, Drug Discovery and Development, Virginia Commonwealth University, Richmond,
Virginia, 23298, USA
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
The serotonin 5-HT7 G protein-coupled receptor (GPCR) is a proposed pharmacotherapeutic
target for a variety of central and peripheral indications, albeit, there are no approved drugs
selective for binding 5-HT7. We previously reported that a lead analog based on the 5-substituted-
N,N-disubstituted-1,2,3,4-tetrahydronaphthalen-2-amine (5-substituted-2-aminotetralin, 5-SAT)
scaffold binds with high affinity at the 5-HT7 GPCR, and can treat symptoms of autism in mouse
models; subsequently, the lead was found to have high affinity at the 5-HT1A GPCR. Herein, we
report the synthesis of novel 5-SAT analogs to develop a 3-dimensional quantitative structure—
affinity relationship (3D-QSAR) at the human 5-HT7 receptor for comparison with similar studies
at the highly homologous 5-HT1A receptor. We report 35 new 5-SAT ligands, some with very high
stereoselective affinity (Ki ≤ 1 nM) at 5-HT7 and/or 5-HT1A receptors, several with modest
selectivity (up to 12-fold) for binding at 5-HT7, and, several ligands with high selectivity (up to
40-fold) at the 5-HT1A receptor. 3D-QSAR results indicate that steric extensions at the C(5)-
position improve selectivity for the 5-HT7 over 5-HT1A receptor, while steric and hydrophobic
extensions at the chiral C(2)-amino position impart 5-HT1A selectivity. In silico receptor
homology modeling studies, supplemented with molecular dynamics simulations and binding free
energy calculations, were used to rationalize experimentally-determined receptor selectivity and
stereoselectivity affinity results. The data from these studies indicate that the 5-SAT chemotype,
previously shown to be safe and efficacious in rodent paradigms of neurodevelopmental and
neuropsychiatric disorders, is amenable to structural modification to optimize affinity at serotonin
5-HT7 vs. 5-HT1A GPCRs, as may be required for successful clinical translation.
Available online
Keywords:
5-HT7R
5-HT1A
R
Serotonergic Receptor
2-aminotetralin
3D-QSAR
Molecular Modeling
———
 Corresponding author. e-mail: charles.perry@vanderbilt.edu (C.K. Perry)

Our lab previously reported on the 5-SAT (5-substituted-
aminotetralin) chemotype16, and more recently on the novel
1. Introduction
analog,
(2S)-5-(2'-fluorophenyl)-N,N-dimethyl-1,2,3,4-
Serotonin (5-hydroxytryptamine, 5-HT), is implicated in the
etiology of numerous central and peripheral nervous system
disorders1,2. 5-HT acts at 14 receptor subtypes, all of which are
GPCRs, except, the 5-HT3 ligand-gated ion channel. The human
5-HT7 receptor is the most recently cloned 5-HT GPCR3,4, albeit,
25-years later, no 5-HT7 crystal structure has been reported. In the
brain, 5-HT7 is highly expressed in the raphe nucleus, thalamus,
hypothalamus, hippocampus, amygdala, and cortex5,6 and
modulates dopaminergic, GABAergic, glutamatergic as well as
serotonergic neuronal signaling and neuronal morphology7,8. It is
well established that 5-HT7 receptors regulate circadian rhythms,
the sleep/wake cycle9, and body temperature10. Genetic and
pharmacological studies also suggest the 5-HT7 receptor might
also be targeted to treat cognitive impairment, epilepsy, repetitive
behaviors, anxiety, depression, impulsivity, and other symptoms
common to neurodevelopmental and neuropsychiatric
tetrahydronaphthalen-2-amine (5-FPT, 8b). In mouse models, 5-
FPT decreases repetitive behaviors and enhances social
interactions, suggesting therapeutic utility for autism spectrum
disorder (ASD) and fragile
X
syndrome17. Previous
pharmacological studies indicated that 5-FPT stereoselectively
binds 5-HT7 receptors and 5-HT1A receptors, and could act as
partial agonist and competitive antagonist at both receptors17. To
investigate structural modifications of the 5-SAT scaffold that
might impact 5-HT7 and 5-HT1A affinity and selectivity, we
focused here on the synthesis of analogs with substitutions at the
chiral C(2)-amine position and the C(5)-position. From this work
we discovered several 5-HT7-moderately selective, 5-HT1A-
selective, and dual 5-HT7/5-HT1A receptor ligands that could be
used to advance the pathophysiological and pharmacotherapeutic
understanding of 5-HT7 and 5-HT1A receptors.
disorders11,12
.
2. Chemistry
Some approved antipsychotic drugs bind with high affinity at
5-HT7 receptors13, as well as, at a variety of other aminergic
neurotransmitter GPCRs with various functional activities, not all
of which are neurotherapeutic. The lack of selective 5-HT7-
targeting medications is at least partly due to the approximately
40% transmembrane domain homology between the 5-HT7 and 5-
HT1A receptors14,11. There also are no crystal structures reported
for the 5-HT1A receptor. The neurobiological mechanisms
associated with 5-HT7 and 5-HT1A receptors and their potential
therapeutic utility have been probed with a variety of structurally
diverse ligands15, yet, no drugs specifically targeting either
receptor or co-targeting 5-HT7/5-HT1A receptors have been
developed.
Scheme 1 shows the strategy used here to synthesize racemic
5-SATs 8b-d, h-t. Trifluoromethanesulfonic anhydride was used
to convert commercially-available 5-hydroxy-1-tetralone 1 to the
corresponding triflate 2. Triflate substitution at the 5-position
allowed for more facile and efficient purification of products than
our previous method of bromination at the 5-position16. Reduction
of the ketone generated the racemic alcohol 3, which underwent
dehydration using a catalytic amount of p-toluenesulfonic acid to
give the olefin 4. Treatment of 4 with meta-chloroperoxybenzoic
acid (mCPBA) generated the corresponding epoxide 5, followed
by subsequent acid-catalyzed epoxide-opening18 to give the 2-
tetralone analog 6. The racemates of analogs 8e-g, were
synthesized according to our previously published methods16.
O
O
OH
c
b
d
a
OH
OTf
OTf
OTf
1
2
3
4
O
O
O
N
1
4
2
3
g
e
f
5
OTf
R
R
OTf
5
6
7a-d, h-t
8a-d, h-t
R = aryl, heteroaryl,
cycloalkyl
Scheme 1. aReagents and conditions: (a) (CF3SO2)2O, pyridine, CH2Cl2 rt. 0.5 h; (b) NaBH4, MeOH/toluene, 0 oC
to rt, 1 h; (c), pTSA, benzene, 80 oC, 0.5 h; (d) mCPBA, NaHCO3, CH2Cl2, rt, 2 h; (e) pTSA, benzene, 80 oC, 0.5 h;
(f) Pd(PPh3)4, K2CO3, R-B(OH)2, toluene/water, 80 oC, 3 h or Pd(PPh3)4 or K3PO4, KBr, R-B(MIDA ester), 1,4
dioxane/H2O, 105 oC, 6 h; (g) NH(Me)2HCl, NaBH3CN, MeOH/THF, 55 oC, 16 h.

N
N
NH2
N
N
a
c
b
d
(S)
(S)
(S)
(S)
(S)
OTf
R
OCH3
OCH3
OH
11
(2S)-8b R= 2’-F-C6H4
(2S)-8c R= 2’-Cl-C6H4
(2S)-8n R= 1-naphthyl
9
12
10
Scheme 2. aReagents and conditions: (a) CH2O, MeOH, reflux, 1 h then NaBH4, rt, 1 h; (b) BBr3, CH2Cl2, 0 oC to rt, 24 h; (c) N-(2-pyridyl)-bis
(trifluoromethanesulfonimide), DIPEA, CH2Cl2, 0 oC to rt, 20 h; (d) R-(BOH)2, Pd(PPH3)4, K2CO3, DME/H2O, reflux, 3 h.
N
NH2
N
N
a, b
d
c
*
*
*
*
R5
OCH3
9 = (2S) or (2R)
OH
13 = (2S) or (2R)
OTf
14 = (2S) or (2R)
(2R) or (2S)-15 R = 2’-Cl-phenyl
(2R) or (2S)-16 R = phenyl
Scheme 3. aReagents and conditions: (a) HBr (48% aq), 130 oC, 3 h; (b) propionaldehyde, montmorillonite K 10, NaBH3CN, MeOH, RT, 20 h; (c) N-(2-Pyridyl)-bis-
(trifluoromethanesulfonimide), DIPEA, CH2Cl2, 0 oC to rt, 20 h; (d) R-B(OH)2 K3PO4, KBr, Pd[P(Ph)3]4, 1,4-dioxane, 120 oC, 6 h.
The tetralone intermediate 6 allowed for Suzuki-Miyaura
coupling19 at the triflated C(5) position using commercially
available boronic acids to give the 5-substituted-2-tetralone
analogs 7a-d, h-t. The majority of analogs were generated using
standard Suzuki coupling procedures (corresponding boronic acid
or ester, base and Pd(PPh3)4), resulting in moderate to high yields
(45–98%). However, the furan- and thiophene-containing analogs
7s and 7t, respectively, could not be obtained efficiently using the
standard Suzuki coupling conditions. A modified slow-release
cross-coupling method20 with the corresponding N-
methyliminodiacetic acid (MIDA) boronates improved yields (30–
32%) for 7s and 7t. Suzuki coupling with cycloalkyl boronic acids
generally was unproductive, and only the 5-cyclopentyl-
substituted analog 7q was successfully synthesized, albeit in poor
yield. Suzuki coupling with 2', 6'-halogen-disubstituted boronic
acids was unsuccessful, likely, because the strong electron-
withdrawing substituents result in rapid protodeboronation under
aqueous conditions21. The 5-substituted-2-tetralone analogs 7a-d
and 7h-t underwent reductive amination22 with dimethylamine to
give the racemic 5-SAT analogs 8a-d and 8h-t. Preparative chiral
HPLC (polysaccharide-based stationary phase) was used
successfully to separate a number of the racemic analogs.
Unfortunately, we were not able to achieve enantiomeric
separation of several analogs despite using various HPLC
conditions.
chiral HPLC separation of racemic 8n yielded four peaks that
could not be individually resolved (see Supporting Information).
Chiral-HPLC analysis of (2S)-8n yielded two peaks and this result,
along with 13C-NMR data, suggested the presence of atropisomers,
possibly due to hindered rotation of the naphthyl moiety around
the C(5)-axis. This phenomenon also was seen in 13C-NMR spectra
of analogs (8c and 8o) with steric bulk a the 2'-position, suggestion
possible atropisomers in these analogs as well.
In Scheme 2, (2S)-5-methoxy-1,2,3,4-tetrahydronaphthalen-2-
amine 9 underwent a modified Eschweiler-Clarke reaction24 to
give (2S)-5-methoxy-N,N-dimethyl-1,2,3,4-tetrahydronaphthalen
-2-amine 10. The 5-methoxy group of 10 was converted to the
24
corresponding phenol through ether cleavage using BBr3
,
resulting in intermediate 11. Triflation of the 5-hydroxy group of
11 was unsuccessful using triflicanhydride, however, use of 2-
pyridyltriflimide, known to improve yields in the presence of
amines25, was successful. Suzuki-Miyaura cross coupling was then
used to obtain the final (2S)-5-SATs.
Scheme 3 was implemented to synthesize analogs (2S)- and
(2R)-15 and 16, given that enantiomeric separation of the
racemates was not successful via chiral-HPLC. Analogs (2S)- and
(2R)-5-methoxy-1,2,3,4-tetrahydronaphthalen-2-amine
9
underwent cleavage of the methoxy group to give the crude
alcohol. Reductive alkylation using propionaldehyde then gave
intermediate (2S)- or (2R)-13. Yields of the alkylation step were
poor until addition of the acidic clay montmorillonite26 was used
to facilitate imine formation. The alcohol then was triflated to give
intermediates (2S)- or (2R)-14, which underwent Suzuki-coupling
to yield the final chiral analogs (2S)- and (2R)-15 and 16.
To validate the (2R)- or (2S)-configuration and corresponding
5-HT7 and 5-HT1A affinity of selected analogs, a chiral pool
synthetic strategy was implemented, starting with commercially-
available (2S)- or (2R)-5-methoxy-1,2,3,4-tetrahydronaphthalen-
2-amines (Schemes 2 and 3). The rationale for obtaining analogs
with known (2S)-absolute stereochemistry included that we
previously synthesized and separated the optical isomers of (±)-8b
using chiral-HPLC17 and found very high stereoselectivity for
binding at human 5-HT7 receptors17. It was hypothesized that (2S)-
absolute stereochemistry would correspond to the high affinity
enantiomer, based on the stereoselective high-affinity of the
commercially-available 5-HT7 probe, (2S)-N,N-dimethyl-5-(1,3,5-
trimethylpyrazol-4-yl)-1,2,3,4-tetrahydronaphthalen-2-amine
(AS-19, Table 2), and other structurally similar compounds at 5-
HT7 receptors23,24. Analog (2S)-8c also was synthesized, as this
analog is expected to have behavioral activity similar to 8b17.
Analog (2S)-8n was synthesized via Scheme 2, in part, because

Given the in vivo behavioral activity of the 2'-fluorine analog
(2S)-8b to attenuate or abolish symptoms of ASD17, Scheme 4 was
implemented to obtain the corresponding 2-N,N-dipropyl and 2-
pyrrolidine analogs 17 and 18, respectively. The tetralone
intermediate 7b underwent reductive amination with
dipropylamine hydrochloride or pyrrolidine to give the racemic
products. Racemate 17 was successfully resolved by chiral-HPLC
while attempts to resolve 18 were unsuccessful.
was observed for the 2'-F analog (2S)-(–)-8b. Substitutions at the
3'-position (F, Cl, CF3, OCH3; 8d-g) resulted in moderate affinity
racemic ligands at 5-HT7 and 5-HT1A receptors, without
impressive selectivity for binding, albeit, the sterically larger
moieties showed some preference for 5-HT7. In contrast,
substitution at the 4'-position provides moderate affinity and
selectivity for 5-HT1A binding, at least, regarding the halogen
moieties, F and Cl (8h, 8i). Analog 8j (2'-Cl, 4'-F) had moderate
affinity at both receptors, with 2-fold preference for 5-HT7. The
3',5'-di-halogenated (F, Cl, CF3) analogs 8k–8m had low affinity
at both receptors, with the larger Cl and CF3 moieties
compromising affinity especially at 5-HT1A, Analog 8i, with a 5-
(4’-chlorphenyl) moiety, showed a reversal in stereoselectivity for
binding at both receptors, with the (+)-enantiomer having higher
affinity, especially, at 5-HT1A. We explored this experimental
result further in the Computational Chemistry and Molecular
Modeling section below.
N
a
F
O
17
To further explore the steric parameter, the naphthyl analog 8n
was synthesized. Compound (2S)-(–)-8n demonstrated high
affinity at 5-HT7 (Ki=5.2 nM) and was the most selective analog
for binding 5-HT7 over 5-HT1A (12-fold). It was hypothesized that
adding a heteroatom into the naphthyl ring may improve affinity
via new electrostatic interactions. This strategy, however, proved
detrimental to binding as the racemic isoquinoline analog 8o
showed lower affinity at both 5-HT7 and 5-HT1A receptors
compared to racemic 8n. Further increasing the size of the C(5)-
substituent from naphthalene to anthracene (8p) greatly decreased
affinity at 5-HT7 (Ki>400 nM) and abolished relevant binding
(Ki>1,000 nM) at 5-HT1A. In summary, adding large substituents
(i.e., Cl, -CF3, naphthalene, anthracene) to the C(5)-position of the
5-SATs studied here increases selectivity for 5-HT7 over 5-HT1A.
These results suggest the orthosteric binding pocket at 5-HT7 is
more accommodating to steric bulk at the 5-SAT C(5)-position
compared to 5-HT1A, consistent with QSAR results discussed
below.
F
7b
N
b
F
18
Scheme 4. aReagents and conditions: (a) Dipropylamine HCl, NaBH3CN,
MeOH/THF, 55 oC, 16 h; (b) Pyrrolidine, NaBH3CN, MeOH/THF, 55 oC, 16 h.
3. In vitro binding results and discussion.
Table 1 reports the observed Ki values at human 5-HT7 and 5-
HT1A receptors for binding of the 5-SATs synthesized here. Using
the chiral-HPLC system to separate racemic 8a obtained via
Scheme 1, we determined that the (–)-8a demonstrates much
higher affinity compared to the (+)-enantiomer at both 5-HT7 and
5-HT1A receptors. Accordingly, Scheme 2 was implemented to
obtain authentic (2S)-(–)-8b, which, was found to have 150- and
250-fold higher affinity than (+)-8b at 5-HT7 and 5-HT1A receptors
(respectively), indicating the importance of stereochemistry for
high affinity binding at both receptors. (2S)-stereochemistry is the
preferred orientation of the C(2)-amine moiety in this series of 2-
aminotetralins, as higher affinity was demonstrated by (2S)-(–)-
8b, 8c, 8n, 15, and 16 compared to the corresponding racemic or
(2R)-(+)-stereoisomer. These observations are explored below in
the Computational Chemistry and Molecular Modeling section.
To explore the impact of smaller and non-aromatic C(5)
substituents, we synthesized the cyclopentyl analog 8q, which
exhibited low affinity for both receptors. In contrast, analogs 8r-t,
containing small heteroaromatic 5-membered rings at the C(5)
position, had moderate 5-HT7 affinity, with the (–)-enantiomer
(presumably, 2S) being the high affinity enantiomer for each
analog. Small, heteroaromatic rings at the C(5) position
preferentially bound to 5-HT1A receptors. Within this series, the
thiophen-2'-yl analog (–)-8t demonstrated the highest affinity,
consistent with studies that have shown sulfur atoms generally
increase receptor affinity compared to other bioisosteres27.
With regard to the C(2)-amine substituent, it has been
previously reported that N,N-dipropyl substitution of other 2-
aminotetralin compounds can result in high affinity ligands at the
5-HT7 receptor24. In the current series, replacing the dimethyl
groups of analogs (2S)-(–)- 8a, 8b, and 8c
Substitution of the 5-phenyl moiety at the 2'-position with
fluorine (8b) or chlorine (8c) increased affinity (compared to
parent 8a) at the 5-HT7 receptor but did not impact affinity at 5-
HT1A. The 2'-Cl analog (2S)-(–)-8c demonstrated selectivity (7-
fold) for binding 5-HT7 over 5-HT1A, whereas, no such selectivity
Table 1.
Affinity and selectivity of 5-substituted-N,N-substituted-1,2,3,4-tetrahydronaphthalen-2-amines at human 5-HT7
and 5-HT1A receptors.
R2
R1

Compound
R1
R2
Stereochemistry/
Optical Rotation
5-HT7
5-HT1A
Selectivity
(5-HT1A/5-HT7)
(Ki ± SEM; nM)
5,200 ± 800
(Ki ± SEM; nM)
1,100 ± 320
8a
C6H5
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
(+)
0.21
0.66
1.4
C6H5
(–)
9.5 ± 1.5
740 ± 160
5.3 ± 2
6.3 ± 0.3
1,000 ±140
4 ± 0.1
8ba
(2S)-8b
8c
2'-F-C6H4
(+)
2'-F-C6H4
(2S)-(–)
racemic
(2S)-(–)
racemic
racemic
racemic
racemic
racemic
(+)
0.75
3.3
2'-Cl-C6H4
4.2 ± 1.2
1.0 ± 0.12
18 ± 2
14 ± 2.4
7.5 ± 1.5
23 ± 4
(2S)-8c
8d
2'-Cl-C6H4
7.6
3'-F-C6H4
1.3
8ea
3'-Cl-C6H4
26 ± 6
82 ± 19
3.2
8fa
3'-CF3-C6H4
3'-OCH3-C6H4
4'-F-C6H4
63 ± 7
180 ± 34
73 ± 12
2.9
8ga
28 ± 9
2.6
8h
117 ± 17
84 ± 11
160 ± 9
16 ± 3
30 ± 7
0.26
0.23
1.2
8i
4'-Cl-C6H4
19 ± 4
4'-Cl-C6H4
(–)
190 ± 47
33 ± 11
8j
8k
8l
2'-Cl-4'-F-C6H3
3', 5'-diF-C6H3
3', 5'-diCl-C6H3
3', 5'-diCF3-C6H3
3', 5'-diCF3-C6H3
1'-naphthyl
1'-naphthyl
isoquinolin-4'-yl
anthracen-9'-yl
cyclopentyl
racemic
racemic
racemic
(+)
2.1
180 ± 17
400 ± 55
> 10,000
910 ± 260
22 ± 2
230 ± 16
2,200 ± 520
> 10,000
2,230 ± 430
130 ± 36
64 ± 12
1.3
5.5
8m
N/A
2.5
(–)
8n
(2S)-8n
8o
racemic
(2S)-(–)
racemic
racemic
racemic
(+)
5.9
5.2 ± 1.1
97 ± 11
450 ± 110
240 ± 35
600 ± 40
12
650 ± 110
1,400 ± 400
98 ± 7
6.7
8p
3.1
8q
0.41
0.92
8r
1-methyl-1H-
550 ± 43
pyrrol-2'-yl
1-methyl-1H-
N(CH3)2
(–)
33 ± 4
24 ± 0.2
0.72
pyrrol-2'-yl
8s
8t
furan-2'-yl
furan-2'-yl
thiophen-2'-yl
thiophen-2'-yl
2'-Cl-C6H4
2'-Cl-C6H4
C6H5
N(CH3)2
N(CH3)2
N(CH3)2
N(CH3)2
N(C3H7)2
N(C3H7)2
N(C3H7)2
(+)
(–)
2600 ± 15
71 ± 11
180 ± 43
17 ± 4
5,600 ± 2,000
33 ± 3
2.2
0.46
0.83
0.26
0.33
0.11
1.1
(+)
150 ± 14
4.5 ± 0.5
130-± 32
0.64 ± 0.2
210 -± 42
(–)
(2R)-15
(2S)-15
(2R)-16
(2R)-(+)
(2S)-(–)
(2R)-(+)
390 ± 98
5.9 ± 1
190 -± 17

(2S)-16
C6H5
N(C3H7)2
N(C3H7)2
N(C3H7)2
Pyrrolidine
(2S)-(–)
(+)
34 ± 3
740 ± 94
5.3 ± 1
51 ± 8
5.9-± 1.1
92 ± 18
0.17
0.12
17
2'-F-C6H4
2'-F-C6H4
2'-F-C6H4
(–)
1.8 ± 0.2
1.3 ± 0.1
0.34
18
racemic
0.025
17
,24
aPreviously reported 16,
Table 2. Structure and affinity of reference compounds
5-HT7
5-HT1A
Selectivity
(5-HT1A /5-HT7)
Compound
AS-19
(Ki ± SEM; nM)
(Ki ± SEM; nM)
12 ± 0.9
240 ± 1.6
20
(R)-8-OH-DPAT
(S)-8-OH-DPAT
Buspirone
48 ± 12
1,000 ± 44
840 ± 183
7.1 ± 0.5
7.6 ± 0.1
50 ± 5
0.148
0.0076
0.059
i.e., there is essentially no stereoselectivity for binding at 5-HT1A.
We could find no documentation in the scientific literature
regarding affinity of 8-OH-DPAT enantiomers at 5-HT7 receptors.
We report here that 8-OH-DPAT demonstrated high (20-fold)
stereoselectivity for binding at 5-HT7, with Ki values of 1,000 and
48 nM for the S- and R-enantiomer, respectively (Table 2). Thus,
in contrast to 5-SAT analogs 15–17, wherein, high-affinity,
stereoselective binding at 5-HT7 (and 5-HT1A) receptors is
associated with (S)-stereochemistry of the C(2)-N,N-dipropyl
substituent, it is (2R)-stereochemistry that provides for high
affinity binding of 8-OH-DPAT at 5-HT7. In this regard, Seiler and
Markstein showed31 that substitution(s) of the 2-aminotetalin
scaffold (e.g., at the 5- vs. 8-positions) impacts enantioselectivity
at aminergic GPCRs. We also assessed buspirone, an azapirone
approved for anxiety that has complex pharmacology (including at
5-HT2A and dopamine D2 GPCRs), but, is thought to mediate
anxiolytic effects via activity at 5-HT1A receptors, with reported
Ki=21 nM28,29, relatively consistent with our result (Table 2). To
the best of our knowledge, the affinity of buspirone for human 5-
HT7 receptors (Ki=840 nM, Table 2) is reported here for the first
time.
with dipropyl groups in corresponding analogs (2S)-(–)- 15, 16,
and 17, maintained affinity at 5-HT7 receptors while improving
affinity at 5-HT1A receptors. For example, among compounds in
Table 1, the N,N-dipropyl analog (2S)-(–)-15 demonstrated the
highest affinity at 5-HT1A, with 10-fold selectivity over 5-H7,
compared, whereas, the corresponding N,N-dimethyl analog (2S)-
(–)-8c had nearly 8-fold higher affinity at 5-HT7. The N-
pyrrolidine analog 18 demonstrated highest 5-HT1A selective
binding (~40-fold) of all the compounds in Table 1. These results
suggest that substitution of the C(2)-amine position of 5-SATs
with dimethyl provides selectivity for 5-HT7 high-affinity binding,
whereas, substitution with dipropyl favors 5-HT1A selective
binding, consistent with QSAR results discussed below.
As mentioned, there are no approved drugs that selectively bind
5-HT7 receptors (or 5-HT1A receptors). Commercially-available
reference compounds used to study 5-HT7 and 5-HT1A receptors
include
AS-19
and
8-hydroxy-N,N-dipropyl-1,2,3,4-
tetrahydronaphthalen-2-amine (8-OH-DPAT, Table 2). We
evaluated these 2-aminotetralins in our affinity assays for direct
comparison to results obtained with the 5-SATs in Table 1.
AS-19 is reported to have very high affinity (Ki=0.6 nM) and
selectivity for human 5-HT7 receptors (150-fold lower affinity at
5-HT1A)23. In our assays, we observed (Table 2) that AS-19 had
only moderately high affinity (Ki=12 nM) and only 20-fold
selectivity for binding 5-HT7 over 5-HT1A (Ki=240 nM). The 8-
OH-DPAT compound is used primarily as a ligand and radioligand
for 5-HT1A receptors, with reported Ki values of 6.1 and 4.1 nM
for the S- and R-enantiomer, respectively.30 Our affinity results for
8-OH-DPAT affinity at 5-HT1A are consistent with the literature,
4. 3D-QSAR
We performed 3D-QSAR studies using the CoMFA32,33

and CoMSIA34 tools of Sybyl on both the 5-HT7 and 5-HT1A data
sets using alignments based on molecular docking results. The
target variable was the pKi as calculated from Tables 1 and 2. In
order to determine the best field set for modeling the 40-member
data sets, multiple runs were performed by selecting CoMFA steric
and electrostatic fields, individually and together, and CoMSIA
steric, electrostatic and hydrophobic fields, individually and in all
combinations, at both the standard 2 Å as well as at 1 Å grid
spacing. Many field sets were unable to produce internally
validated models (q2 < 0.25), but a number were found with q2 ~
set compounds. More than half (6/10) of such trials produced
models with acceptable (r2 > 0.6) test set predictions.
Predicted pKi vs. actual pKi plots for these two models are
shown in Supplementary Information Figure S1. The
coefficient*standard deviation contour plots are shown in Figure
S2. To characterize the factors that result in ligand selectivity for
5-HT7 (or 5-HT1A), we also performed a 3D-QSAR study using
pKi(5-HT7) – pKi(5-HT1A) as the target variable (Table 3). While
this model is slightly below the generally accepted standard of q2
= 0.5, it has r2 = 0.665, and is informative about serotonin receptor
subtype selectivity. The predicted vs. actual selectivity is shown in
Figure 1 and the stddev*coefficient plots are shown in Figure 2.
Both 2 Å and 1 Å grid spacing, and a variety of field sets, were
Table 3. 3D QSAR Model
5-HT1A 5-HT7 5-HT7/5-HT1A
Grid spacing (Å)
q2 (cross-validated r2)
Components
2
2
1
0.489
4
0.507
3
0.471
2
Conventional r2
Standard error
F-value
0.782
0.479
31.37
0.246
0.754
0.825
0.400
41.18
0.254
0.746
0.665
0.419
36.78
0.592
0.408
Fraction steric
Fraction hydrophobic
used in calculating models, with the best model (steric field only)
being only minimally superior to the steric/hydrophobic model.
The 3D-QSAR contour plots shown in Figure 2 were generated
to indicate features or regions of space that support receptor
subtype selectivity. Thus, blue regions indicate where steric bulk
should enhance 5-HT7 over 5-HT1A selectivity, while the green
contours suggest regions where hydrophobic groups should
enhance this selectivity. On the other hand, the red and purple
contours, respectively, report on 5-HT1A over 5-HT7 selectivity
due to steric and hydrophobic substitutions, respectively.
Substitutions to the C(5) aromatic ring are illustrated by the
collection of contours in the top portion of Figure 2. The results
show that hydrophobic extensions that enhance the steric bulk on
the C(5) ring, on either side, like the 2'-Cl of (2S)-8c, the 1'-
naphthyl of 8n or the anthracen-9'-yl of 8p, are more 5-HT7
selective. The purple contour at the center of this cluster appears
to suggest the opposite–compounds without such extensions are 5-
HT1A selective, e.g., (+)-8i and (2R)-15. However, there is another
interpretation that should be considered: placing a polar, i.e.,
negative hydrophobic, moiety in this location would enhance 5-
HT7 selectivity, as exemplified by AS-19. All CoMSIA contour
information at the other end of the molecule, the C(2) amine,
Figure 1. CoMSIA steric + hydrophobic predicted vs. actual receptor selectivity
for Ki(5-HT7)/Ki(5-HT1A)
0.5, which is considered the benchmark for reasonable, although
not superior, models35. In order to facilitate comparisons between
5-HT1A and 5-HT7 data, we chose the CoMSIA model with 2 Å
grid spacing applying the steric + hydrophobic field types (Table
3). Including the electrostatic field with the steric and hydrophobic
fields, or, using it to replace either of the other two, generated
inferior models, particularly for the 5-HT1A models. Also, the
smaller 1 Å grid spacing yielded no improvement, and, in some
cases, even worsened the models. We tested the ability of these
models to predict “external” data by randomly splitting the data set
into 30-member training sets and 10-member test sets, followed by
rebuilding the models from the training sets and predicting the test
Figure 2: Stddev*coefficient contour plots for CoMSIA 3D QSAR analysis of serotonin subtype selectivity. The right view is rotated 90º
clockwise around the vertical axis. Steric contours plotted at 90% (blue) and 10% (red); hydrophobic contours plotted at 90% (green) and
10% (purple). The two molecules shown are (2S)-(–)-8b (with carbon atoms in magenta) and (2R)-(+)-8b (with carbon atoms in white).

highlights features for 5-HT1A selectivity. The pyrrolidine analog
18 and both isomers of 8-OH-DPAT are relevant cases with
substituents in these positions and 5-HT1A selectivity.
wherein, the (2S)-(–)-enantiomer had higher affinity than the (2R)-
(+)-enantiomer. To help explain the experimental results for 8i
stereoselective binding, ligand docking, MD simulations, and
MMGBSA calculations were conducted (assignments were (+)-2R
and (–)-2S). At the 5-HT7 receptor, results indicate that (+)-8i
realizes enhanced interactions with residues in TMDs 3 and 6 (e.g.,
D1623.32, V1633.33, W3406.48, F3446.52) compared to (–)-8i; notably,
5. Computational Chemistry and Molecular
Modeling.
Using strategies and methods similar to our previous reports36–
we built homology models of the human 5-HT1A and 5-HT7
38
GPCRs based on the 5-HT1B crystal structure (PDB: 4IAQ) to
rationalize the effects of stereochemistry on the binding affinities
of
5-SATs
at
5-HT7
and
5-HT1A
receptors.
Docking studies followed by molecular dynamics (MD)
simulations using analogs and (2S)-(–)- and (2R)-(+)-8b indicated
a low energy pose of (2S)-(–)-8b in complex with the 5-HT7
receptor such that the 2-dimethylamine moiety binds deeper into
the orthosteric pocket than (2R)-(+)-8b (Figure 3). The basic
nitrogen of (2S)-8b (assumed to be protonated at physiological pH)
can form the expected salt bridge (ionic bond) with the 5-HT7
receptor carboxylic acid residue D1623.32 (2.9 Å). The ligand’s
tetrahydronaphthyl and C(5)-fluorophenyl moieties can form
several strong hydrophobic and aromatic interactions with 5-HT7
residues in TMDs 3, 6, and 7, e.g., V1633.33, F3436.51 F3446.52
,
,
W3406.48, Y3747.43 (Table S1). In comparison, (2R)-(+)-8b can
form an ionic bond with D1623.32 (2.8 Å), but, forms weaker free
energy interactions with residues in TMDs 3, 5, 6, and 7 (Table
S2)—notably, there is loss of interaction with W3406.48. MD
simulations followed by molecular mechanics-generalized Born
surface area (MMGBSA) binding free energy calculations yielded
a difference in binding free energies that is 3.51 kcal/mol lower for
(2S)-(–)-8b compared to (2R)-(+)-8b. Thus, both ligand docking
and MMGBSA results are consistent with experimental results
indicating the (2S)-(–)-enantiomer has 140-fold higher affinity at
5-HT7 (Table 1). Results of ligand docking experiments, MD
simulations, and MMGBSA calculations conducted for (–)-(2S)-
and (2R)-(+)-8b interactions with the 5-HT1A receptor model are
given in Supplementary Information (Figure S3, Tables S1 and
S2). Key findings include that MMGBSA binding free energies for
the 5-HT1A receptor complexes indicated (2S)-(–)-8b has lower
overall binding free energy (by 3.4 kcal/mol) compared to (2R)-
(+)-8b, consistent with the observed 250-fold higher affinity of
(2S)-(–)-8b at 5-HT1A (Table 1).
Figure 4. Merged docked poses of (2S)-(–)-8i (light-blue receptor/dark purple
ligand) and (2R)-(+)-8i (pink receptor/red ligand) at the 5-HT7 receptor following
MD simulations.
there appears to be nil interaction of (–)-8i with W3406.48 (Figure
4). MMGBSA binding free energy calculations for 5-HT7 receptor
interactions indicated a lower energy pose of (+)-8i (by 2.15
kcal/mol) compared to (–)-8i (Tables S3 and S4). At the 5-HT1A
receptor (Figure S3, Tables S3 and S4), (+)-8i was able to make
more low-energy interactions than (–)-8i, which, was positioned
further away from residues in TMDs 5 and 6. Overall, there was a
difference of 3.12 kcal/mol lower calculated binding free energy
for (+)-8i compared to (–)-8i, consistent with the 10-fold higher
affinity of (+)-8i at 5-HT1A (Table 1).
The experimentally-determined affinity values for analog 8i
indicated stereoselective binding at 5-HT7 and 5-HT1A receptors
that is reversed from all other enantiomeric pairs in Table 1,
6. Conclusions
Binding properties of the 2-aminotetralin analogs presented
here demonstrate that the 5-SAT scaffold is amenable to
development of high affinity 5-HT7 and 5-HT1A receptor ligands.
Analyses of results from experimental, computational, and
molecular modeling studies elucidated important structural
requirements for 5-HT7 and 5-HT1A receptor affinity, selectivity,
and enantioselectivity. An unsubstituted phenyl moiety, as well as,
2'- and 3'-substituted phenyl groups at the C(5)-position result in
high affinity 5-HT7 ligands. Groups that have steric bulk at the 2'-
and 3'-phenyl positions enhance selectivity for 5-HT7 over 5-HT1A
binding. In contrast, substituents at the 4'-phenyl position appear
to favor 5-HT1A binding. High affinity for 5-HT7 with moderate
selectivity (~12-fold) over 5-HT1A can be achieved with larger
substituents at the C(5)-position, suggesting a difference in steric
tolerance between the orthosteric binding pockets. Small, 5-
membered heterocyclic systems at the C(5)-position show high
affinity for 5-HT1A, while demonstrating moderate affinity at 5-
HT7. Extension of the C(2)-amine substituents from dimethyl to
dipropyl modestly improves affinity at 5-HT1A, but does not affect
affinity at 5-HT7. Finally, pharmacological, computational, and
molecular modeling results indicate that the (2S)-configuration of
Figure 3. Proposed binding mode of (2S)-(–)-8b (light-purple receptor/sky-blue
ligand) and (2R)-(+)-8b (pink receptor/magenta ligand) at the 5-HT7 receptor,
merged following MD simulations, with TMD 5 removed for clarity. The C(2)-
amine moiety of (2S)-(–)-8b juts deeper into the binding pocket than that of the (2R)-
configuration, and, overall, (2S)-(–)-8b has greater interactions with I233, W3406.48
,
and F3446.52., which may explain its higher affinity.

the 5-SAT analogs reported here is the preferred configuration for
high affinity binding of most 5-SATs at both 5-HT7 and 5-HT1A
receptors.
and washed with saturated aqueous sodium bicarbonate, dried over
sodium sulfate and concentrated in vacuo. No further purification
was done yielding 0.9 g (99%) of a red oil. TLC eluents used (9:1
Hexanes: Ethyl Acetate). 1H-NMR (500 MHz; CDCl3): δ 8.08 (d,
J = 7.7 Hz, 1H), 7.45 (d, J = 7.4 Hz, 1H), 7.40 (t, J = 7.9 Hz, 1H),
While modifications of the 5-SAT scaffold produced modest
selectivity for binding at 5-HT7 receptors, nearly 40-fold
selectivity (18, Table 1) was realized for binding at 5-HT1A
receptors. Many analogs synthesized here share high affinity at
both 5-HT7 and 5-HT1A, and such polypharmacology may be more
productive regarding treatment of neuropsychiatric disorders. For
example, (2S)-8b (5-FPT) has equipotent high-affinity at both 5-
HT7 and 5-HT1A receptors is orally bioavailable, enhances social
interaction, and attenuates repetitive behaviors in several mouse
models relevant to autism and other neuropsychiatric disorders17,
albeit, its in vivo functional activities are not clear. We are
currently assessing (2S)-8c as a follow-up, lead medication
candidate, owing to its enhanced affinity at both receptors.
Considering that 5-HT7 and 5-HT1A receptors are implicated in
3.02 (t, J = 6.1 Hz, 2H), 2.69 (t, J = 6.6 Hz, 2H), 2.17 (quin, J =
13
6.4 Hz, 2H). C-NMR (400 MHz, CDCl ): δ 196.33, 147.41,
3
137.20, 135.10, 127.88, 127.47, 126.14, 123.53, 120.34, 117.15,
113.96, 38.55, 23.77, 22.32. Calcd C11H9F3O4S for [M+H]+:
295.0252. Found: 295.0249.
7.1.2. 5-hydroxy-5,6,7,8-tetrahydronaphthalen-
1-yl trifluoromethanesulfonate (3)
Intermediate 2 (0.840 g, 2.84 mmol) was dissolved in 20 mL of
a 1:1 MeOH:toluene solution and cooled to 0 oC. NaBH4 (0.27 g,
7.2 mmol) was added and the reaction stirred for 1 hour at 0 oC. At
completion the solvent was evaporated off and 20 mL of saturated
ammonium chloride was added followed by extraction with ethyl
acetate (3 x 30 mL). Organic fractions were combined, dried over
sodium sulfate and concentrated in vacuo. Purification was done
by flash chromatography (8:2 hexanes:EtOAc) yielding 0.64 g
(75%) of a white solid. TLC eluents used (7:3 Hexanes: Ethyl
Acetate). 1H-NMR (500 MHz; CDCl3): δ 7.52 (d, J = 7.7 Hz, 1H),
7.29 (t, J = 8.0 Hz, 1H), 7.18 (d, J = 8.2 Hz, 1H), 4.81 (t, J = 4.9
Hz, 1H), 2.88 (dt, J = 17.6, 5.9 Hz, 1H), 2.73 (dt, J = 17.7, 6.6 Hz,
several psychiatric disorders with shared symptomologies8,11,39–43
,
the dual-affinity ligands presented here are a valuable resource as
potential therapeutics leads.
7. Experimental section
7.1 Chemistry
1H), 2.07-1.96 (m, 2H), 1.92-1.81 (m, 2H), 1.78 (d, J = 6.0 Hz,
13
1H). C-NMR (400 MHz, CDCl ): δ 147.97, 142.29, 130.61,
3
All commercially available regents and solvents were
purchased and used without purification, unless otherwise
specified. Flash column chromatography was performed with the
use of Agela Technologies 230−400 mesh silica gel. Analytical
thin-layer chromatography (TLC) was carried out on Agela
Technologies silica gel 60 F254 plates. Final compounds were
used as either free base of their corresponding HCl salts, as noted
below, depending on ease of manipulation of the free base. All
spectra were recorded by a Varian 500 MHz or 400 MHz NMR in
CDCl3 or CD3OD as noted and are expressed as chemical shift (δ)
values in parts per million (ppm). Coupling constants (J) are
presented in Hertz. Abbreviations used in the reporting of NMR
spectra include, s = singlet, bs =broad singlet, d = doublet, t =
triplet, q = quartet, quin = quintet, dd = doublet of doublets, qd =
quartet of doublets, dt = doublet of triplets, tt = triplet of triplets,
ddd = doublet of doublets of doublets, ddt = doublet of doublet of
triplets, dtt = doublet of triplet of triplets, m = multiplet). High
resolution mass spectrometry (HRMS) was performed with
Waters Q-TOF Ultima ESI instrument using time of flight (TOF-
MS) and electron spray ionization (ESI) or using a Waters 70-VSE
instrument using time of flight (TOF-MS) and electron ionization
(EI). HPLC separation of (+) and (–) enantiomers was determined
by UV Trace 220/254 nm on a Shimadzu™ instrument equipped
with a semi-preparative (s-prep)-RegisCell™ (5 m, 25 cm x 10
mm i.d.) chiral (polysaccharide-based) column. Optical rotation
was determined using a JACSO (P-2000) polarimeter. Purity of
targeted compounds was > 95% (determined by HPLC) unless
otherwise noted.
128.75, 127.47, 120.32, 117.16, 67.83, 31.76, 23.71, 18.08. Calcd
C11H11F3O4S for [M+H]+: 296.0330. Found: 296.0331.
7.1.3. 7,8-dihydronaphthalen-1-yl
trifluoromethanesulfonate (4)
Intermediate 3 (0.64 g, 2.15mmol) was dissolved in 10 mL
benzene. p-toluenesulfonic acid monohydrate (95 mg, 0.5mmol)
was added and the reaction stirred for 30 min at reflux. The
reaction was quenched with saturated aqueous sodium bicarbonate
and extracted with diethyl ether (3 x 20 mL). Organic fractions
were combined, dried over Na2SO4 and concentrated in vacuo. The
crude product was purified by flash chromatography (99:1
hexanes:EtOAc) to yield 0.53 g (88%) as a clear oil. TLC eluents
used (95:5 Hexanes: Ethyl Acetate). 1H-NMR (500 MHz; CDCl3):
δ 7.20 (t, J = 7.9 Hz, 1H), 7.08 (d, J = 8.3 Hz, 1H), 7.04 (d, J = 7.5
Hz, 1H), 6.50-6.48 (m, 1H), 6.13 (dt, J = 9.4, 4.6 Hz, 1H), 2.88 (t,
13
J = 8.3 Hz, 2H), 2.35 (tdd, J = 8.2, 4.4, 1.8 Hz, 2H). C-NMR
(400 MHz, CDCl ): δ 147.32, 137.04, 130.51, 128.05, 127.74,
3
126.96, 125.90, 123.63, 120.44, 119.85, 117.25, 114.06, 22.26,
21.08. Calcd C11H9F3O3S for [M+H]+: 278.0225. Found:
278.0223.
7.1.4. 1a,2,3,7b-tetrahydronaphtho[1,2-
b]oxiren-4-yl trifluoromethanesulfonate (5)
Intermediate 4 (0.53 g, 1.9 mmol) was dissolved in 20 mL of
dichloromethane. NaHCO3 (0.33 g, 3.9 mmol) was added to the
reaction mixture followed by (0.69 g, 3.9mmol) mg of mCPBA.
The reaction stirred for 2 hours at room temperature and was
quenched with saturate aqueous sodium sulfate followed by
extraction with dichloromethane (3 x 30 mL). Organic fractions
were combined, washed with saturated aqueous sodium chloride,
dried over sodium sulfate and concentrated in vacuo. Purification
was done by flash chromatography (95:5 hexanes EtOAc) yielding
0.5 g (81%) of a colorless oil. TLC eluents used (9:1 Hexanes:
Ethyl Acetate). 1H-NMR (500 MHz; CDCl3): δ 7.44 (d, J = 7.2 Hz,
1H), 7.26 (quin, J = 8.3 Hz, 2H), 3.89 (d, J = 4.1 Hz, 1H), 3.76 (d,
J = 3.0 Hz, 1H), 2.91 (dd, J = 15.9, 5.3 Hz, 1H), 2.61-2.54 (m, 1H),
2.51-2.47 (m, 1H), 1.74 (td, J = 13.8, 5.5 Hz, 1H). 13C-NMR (400
MHz, CDCl3): δ 147.38, 136.27, 129.64, 127.47, 123.58, 121.65,
7.1.1. 5-oxo-5,6,7,8-tetrahydronaphthalen-1-yl
trifluoromethanesulfonate (2)
The 5-hydroxy-1-teralone 1 (0.5 g, 3.08 mmol) was dissolved
in 20 mL dichloromethane. Pyridine, (0.75 mL, 9.3 mmol) was
o
added and the reaction was cooled to 0 C. To the reaction was
added 4.6mL (4.6 mmol) of
a
1
M
solution of
trifluromethanesulfonic anhydride and was stirred for 5 minutes at
0 oC. The reaction was brought to room temperature and stirred for
30 minutes. The reaction was quenched with deionized water
followed by addition of 2N HCl and extracted with
dichloromethane (3 x 40 mL). Organic fractions were combined

7.1.10. 5-(4-fluorophenyl)-3,4-
dihydronaphthalen-2(1H)-one (7h)
121.54, 120.39, 117.19, 114.00, 54.90, 52.05, 21.09, 17.96. Calcd
C11H9F3O4S for [M+H]+: 295.0252. Found: 295.0249.
Obtained from 6 (0.67 mmol) as a light red oil, yield; 58% (0.09
g), purification 95:5 (hexanes:EtOAC): 1H NMR (500 MHz,
CDCl3) δ 7.30-7.27 (m, 3H), 7.20 (m, 1H), 7.16-7.11 (m, 3H), 3.66
(s, 2H), 3.01 (t, J = 6.5 Hz, 2H), 2.43 (t, J = 7.0 Hz, 2H).
7.1.5. 6-oxo-5,6,7,8-tetrahydronaphthalen-1-yl
trifluoromethanesulfonate (6)
Intermediate 5 (0.45 g, 1.5 mmol) was dissolved in 10 mL
benzene and (0.06 g 0.3 mmol) of p-toluenesulfonic acid
monohydrate was added. The reaction stirred at reflux for 45
minutes and was quenched with saturated aqueous sodium
bicarbonate followed by extraction with diethyl ether. Organic
fractions were combined, dried over sodium sulfate and
concentrated in vacuo. Purification was done by flash column
(95:5 hexanes: EtOAc) to yield 0.28 g (62%) of a light red oil. TLC
7.1.11. 5-(4-chlorophenyl)-3,4-
dihydronaphthalen-2(1H)-one (7i)
Obtained from 6 (0.68 mmol) as a light yellow oil, yield: 72%
(0.125 g), purification 95:5 (hexanes:EtOAc): 1H-NMR (500
MHz; CDCl3): δ 7.38 (d, J = 8.3 Hz, 2H), 7.23 (dd, J = 9.0, 5.3
Hz, 3H), 7.16 (d, J = 7.5 Hz, 1H), 7.13 (d, J = 7.6 Hz, 1H), 3.63
(s, 2H), 2.98 (t, J = 6.6 Hz, 2H), 2.40 (t, J = 6.6 Hz, 2H).
1
eluents used (9:1 Hexanes: Ethyl Acetate). H-NMR (500 MHz;
CDCl3): δ 7.31 (t, J = 7.9 Hz, 1H), 7.19 (t, J = 9.3 Hz, 2H), 3.65
(s, 2H), 3.17 (t, J = 6.7 Hz, 2H), 2.57 (t, J = 6.7 Hz, 2H). C-NMR
7.1.12. 5-(4-chloro-2-fluorophenyl)-3,4-
dihydronaphthalen-2(1H)-one (7j)
13
(400 MHz, CDCl ): δ 208.25, 146.97, 137.06, 129.99, 128.38,
3
Obtained from 6 (0.39 mmol) as a light yellow oil, yield; 46%
(0.05 g), purification 97:3 (hexanes:EtOAc): 1H-NMR (500 MHz;
CDCl3): δ 7.29 (t, J = 7.6 Hz, 1H), 7.24 (td, J = 5.8, 2.6 Hz, 2H),
7.19 (d, J = 7.5 Hz, 1H), 7.10-7.05 (m, 2H), 3.66 (d, J = 7.3 Hz,
2H), 2.87 (ddd, J = 15.6, 7.3, 5.9 Hz, 1H), 2.75-2.70 (m, 1H), 2.53-
2.40 (m, 2H).
123.51, 120.31, 119.87, 117.12, 113.93, 44.80, 36.86, 22.17. Calcd
C11H9F3O4S for [M+H]+: 294.0174. Found: 294.0170.
General Suzuki-coupling procedure for analogs 7a-d, h-r
(Scheme 1)
Intermediate 6 was dissolved in a 5:3 toluene:water mixture.
The solution was degassed with N2 and 3 equivalents of potassium
carbonate were added followed by 3 mol % of tetrakis
triphenylphosphine Pd [0] and 1.5 equivalents of the
corresponding boronic acid. The reaction was heated to 90 0C and
stirred for 3 hours. The reaction was quenched with deionized
water and extracted with ethyl acetate (3x). Organic fractions were
combined, washed with saturated sodium bicarbonate, dried over
sodium sulfate and concentrated in vacuo. Purification was done
by flash chromatography (hexanes:EtOAc). TLC eluents used (9:1
Hexanes: Ethyl Acetate) for all intermediates.
7.1.13. 5-(3,5-difluorophenyl)-3,4-
dihydronaphthalen-2(1H)-one (7k)
Obtained from 6 (0.47 mmol) as a light red oil, yield; 66% (0.08
g), purification 95:5 (hexanes:EtOAC): 1H NMR (500 MHz,
CDCl3) δ 7.29 (t, J = 7.0 Hz, 1H), 7.19 (d, J = 7.5 Hz, 2H ), 6.87-
6.81 (m, 3H), 3.67 (s, 2H), 3.02 (t, J = 6.5 Hz, 2H), 2.46 (t, J = 6.5
Hz, 2H);
7.1.14. 5-(3,5-dichlorophenyl)-3,4-
dihydronaphthalen-2(1H)-one (7l)
Obtained from 6 (0.2 mmol) as a light yellow oil, yield 31%
(0.02 g), purification 95:5 (hexanes:EtOAc). (7l was not able to be
7.1.6. 5-phenyl-3,4-dihydronaphthalen-2(1H)-
one (7a)
fully purified from the excess boronate by flash chromatography).
1
Obtained from 6 (0.89 mmol) as a light yellow oil, yield; 98%
(0.19 g) purification 95:5 (hexanes:EtOAc): 1H-NMR (500 MHz;
CDCl3): δ 7.42 (t, J = 7.3 Hz, 2H), 7.36 (d, J = 7.3 Hz, 1H), 7.34-
7.31 (m, 2H), 7.26 (t, J = 7.5 Hz, 1H), 7.21 (d, J = 7.3 Hz, 1H),
7.13 (d, J = 7.3 Hz, 1H), 3.65 (s, 2H), 3.02 (t, J = 6.6 Hz, 2H), 2.41
(t, J = 6.6 Hz, 2H).
H-NMR (500 MHz; CDCl ): δ 7.40 (t, J = 1.8 Hz, 1H), 7.30 (t, J
3
= 7.6 Hz, 1H), 7.23 (d, J = 1.8 Hz, 2H), 7.19 (t, J = 7.2 Hz, 2H),
3.69 (s, 2H), 3.02 (t, J = 6.6 Hz, 2H), 2.48 (t, J = 6.6 Hz, 2H).
7.1.15. 5-(3,5-bis(trifluoromethyl)phenyl)-3,4-
dihydronaphthalen-2(1H)-one (7m)
Obtained from 6 (1.11 mmol) as a clear oil, yield; 63% (0.25
g), purification 98:2 (hexanes:EtOAC): 1H NMR (500 MHz,
CDCl3) δ 7.91(s, 1H), 7.81 (s, 2H), 7.34 (t, J = 7.5 Hz, 1H), 7.25-
7.22 (m, 2H), 3.69 (s, 2H), 3.01 (t, J = 6.5 Hz, 2H), 2.47 (t, J = 6.5
Hz, 2H).
7.1.7. 5-(2-fluorophenyl)-3,4-
dihydronaphthalen-2(1H)-one (7b)
Obtained from 6 (0.68 mmol) as a light red oil, yield; 61% (0.10
g), purification 95:5 (hexanes:EtOAc): 1H-NMR (500 MHz;
CDCl3): δ 7.42-7.39 (m, 1H), 7.34-7.31 (m, 2H), 7.23 (m, 4H),
3.70 (d, J = 5.5 Hz, 2H), 3.03 (dd, J = 13.7, 6.4 Hz, 1H), 2.89 (dd,
J = 13.7, 7.5 Hz, 1H), 2.53 (q, J = 7.4 Hz, 1H), 2.46-2.40 (m, 1H).
7.1.16. 7,8-dihydro-[1,1'-binaphthalen]-6(5H)-
one (7n)
Obtained from 6 (0.5 mmol) as a clear oil, yield; 70% (0.095g),
purification 95:5 (hexanes:EtOAc); 1H-NMR (500 MHz; CDCl3):
δ 7.96 (dd, J = 13.9, 8.2 Hz, 2H), 7.59 (t, J = 7.6 Hz, 1H), 7.55 (d,
J = 7.1 Hz, 1H), 7.52 (d, J = 8.0 Hz, 1H), 7.45 (d, J = 7.2 Hz, 1H),
7.41 (q, J = 6.3 Hz, 1H), 7.37 (d, J = 7.5 Hz, 1H), 7.29 (t, J = 6.7
Hz, 2H), 3.76 (d, J = 3.8 Hz, 2H), 2.73 (t, J = 6.6 Hz, 2H), 2.41 (t,
J = 6.6 Hz, 2H).
7.1.8. 5-(2-chlorophenyl)-3,4-
dihydronaphthalen-2(1H)-one (7c)
Obtained from 6 (0.34 mmol) as a light yellow oil, yield; 64%
(0.06 g), purification 95:5 (hexanes:EtOAC): 1H NMR (500 MHz,
CDCl3) δ 7.49-7.47 (m, 1H), 7.34-7.32 (m, 2H), 7.30-7.25 (m, 2H),
7.19 (d, J = 7.5 Hz, 1H), 7.13 (d, J = 7.5 Hz, 1H), 3.67 (d, J = 7.5
Hz, 2H), 2.92-2.86 (ddd, J = 15.5, 7.5, 5.5 Hz, 1H), 2.76-2.71 (ddd,
J = 15.5, 8.0, 5.5 Hz, 1H), 2.51-2.43 (m, 2H).
7.1.17. 5-(isoquinolin-4-yl)-3,4-
dihydronaphthalen-2(1H)-one (7o)
7.1.9. 5-(3-fluorophenyl)-3,4-
dihydronaphthalen-2(1H)-one (7d)
Obtained from 6 (0.51 mmol) as an orange oil, yield: 49%
1
(0.07g), purification 6:4 (hexanes:EtOAc); H-NMR (500 MHz;
Obtained from 6 (1.02 mmol) as a light red oil, yield; 62% (0.15
g), purification 95:5 (hexanes:EtOAC): 1H NMR (500 MHz,
CDCl3) δ 7.42-7.37 (m, 1H), 7.30-7.25 (m, 1H), 7.21-7.15 (m, 2H),
7.10-7.01(m, 3H), 3.67 (s, 2H), 3.02 (t, J = 8.0 Hz, 2H), 2.44 (t, J
= 8.0 Hz, 2H).
CDCl3): δ 9.31 (s, 1H), 8.44 (s, 1H), 8.08-8.06 (m, 1H), 7.65 (t, J
= 3.9 Hz, 2H), 7.50-7.48 (m, 1H), 7.36 (t, J = 7.6 Hz, 1H), 7.26
(dd, J = 11.0, 7.6 Hz, 2H), 3.72 (s, 2H), 2.76-2.66 (m, 2H), 2.43-
2.34 (m, 2H).

7.1.18. 5-(anthracen-9-yl)-3,4-
bicarbonate, dried over sodium sulfate and concentrated in vacuo.
dihydronaphthalen-2(1H)-one (7p)
Purification
(Dichloromethane:MeOH).
Dichloromethane: MeOH) for all final products.
was
done
by
TLC
flash
eluents
chromatography
used (95:5
Obtained from 6 (1.10 mmol) as a light yellow oil, yield 73%
(0.140 g), purification 95:5 (hexanes:EtOAC); 1H NMR (500
MHz, CDCl3) δ 8.55 (s, 1H), 8.11 (m, 2H ), 7.55-7.48 (m, 4H),
7.45 (t, J = 7.5 Hz, 1H), 7.40-7.34 (m, 3H), 7.28 (m, 1H), 3.79 (s,
2H), 2.51 (t, J = 6.5 Hz, 2H), 2.33 (t, J = 6.5 Hz, 2H).
7.1.23. N,N-dimethyl-5-phenyl-1,2,3,4-
tetrahydronaphthalen-2-amine HCl (8a)
Obtained from 7a (0.5 mmol) as a clear oil, yield; 60% (0.075
g), purification 95:5 (DCM:MeOH). The oil was converted to the
corresponding HCl salt. 1H-NMR (500 MHz; CDCl3): δ 12.64 (s,
1H), 7.41 (t, J = 7.3 Hz, 2H), 7.35 (t, J = 7.3 Hz, 1H), 7.23 (t, J =
8.3 Hz, 3H), 7.15 (d, J = 7.5 Hz, 1H), 7.12 (d, J = 7.4 Hz, 1H),
3.54 (s, 1H), 3.39 (dd, J = 15.4, 3.9 Hz, 1H), 3.20 (dd, J = 14.8,
11.8 Hz, 1H), 2.85-2.79 (m, 8H), 2.40 (d, J = 10.9 Hz, 1H), 1.81
(qd, J = 11.4, 6.6 Hz, 1H). 13C NMR (400 MHz; CDCl3): δ 142.2,
141.0, 132.34, 132.28, 129.1, 128.72, 128.63, 128.4, 127.4, 126.7,
62.3, 39.8, 39.5, 30.1, 27.3, 24.3 HRMS: Calcd C18H21N for
[M+H]+: 252.1752. Found: 252.1756. HPLC (s-prep): Solvent
System: hexanes/EtOH (91:1) 0.1% DEA (modifier) 0.1% TFA
(modifier); flow rate = 3.0 mL/min; t1 = 16.5, t2 = 18.1 (+)-8a HCl:
7.1.19. 5-cyclopentyl-3,4-dihydronaphthalen-
2(1H)-one (7q).
Obtained from 6 (0.58 mmol) as a clear oil, yield 8% (0.01 g),
1
purification 99:1 (hexanes:EtOAC); H NMR (500 MHz, CDCl3):
δ 7.19-7.14 (m, 2H), 6.97 (d, J = 7.1 Hz, 1H), 3.59 (s, 2H), 3.29-
3.22 (m, 1H), 3.12 (t, J = 6.6 Hz, 2H), 2.52 (t, J = 6.6 Hz, 2H),
2.06-2.00 (m, 2H), 1.86-1.81 (m, 2H), 1.75-1.68 (m, 2H), 1.65-
1.57 (m, 2H).
7.1.20. 5-(1-methyl-1H-pyrrol-2-yl)-3,4-
dihydronaphthalen-2(1H)-one (7r)
Obtained from 6 (0.85 mmol) as a light red oil, yield 48%
(0.170 g), purification 95:5 (hexanes:EtOAC); 1H NMR (500
MHz, CDCl3) δ 7.27-7.21 (m, 2H), 7.15 (d, J = 7.3 Hz, 1H), 6.74
(t, J = 2.2 Hz, 1H), 6.23 (t, J = 3.1 Hz, 1H), 6.09 (dd, J = 3.5, 1.8
Hz, 1H), 3.65 (s, 2H), 3.45 (s, 3H), 2.94 (t, J = 6.6 Hz, 2H), 2.44
(t, J = 6.6 Hz, 2H).
25
25
[a]D +36.6 (c 0.2, MeOH), (–)-8a HCl: [a]D – 36.94 (c 0.3,
MeOH).
7.1.24. 5-(2-fluorophenyl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl (8b)
Obtained from 7b (0.42 mmol) as a light yellow oil, yield; 74%
General slow release Suzuki-coupling procedure for
analogs 7s-t20 (Scheme 1) 6 was dissolved in 1, 4-dioxane and the
solution degassed with N2. The corresponding MIDA ester was
added (1.2 equivalents), followed by 1.1 equivalents of potassium
bromide. K3PO4 (7 equivalents) was dissolved in deionized water
to make a 3M solution and was added, followed by 5 mol %
tetrakis triphenylphosphine Pd [0]. The solution was heated to 105
oC and stirred for 6 hours. The reaction was quenched with
deionized water and extracted with ethyl acetate (3x). Organic
fractions were combined, washed with saturated sodium
bicarbonate, dried over sodium sulfate and concentrated in vacuo.
Purification was done by flash chromatography (hexanes:EtOAc).
TLC eluents used (9:1 Hexanes: Ethyl Acetate) for all
intermediates.
(0.08 g), purification 95:5 (DCM:MeOH). The oil was converted
1
to the corresponding HCl salt. H-NMR (500 MHz; CDCl3): δ
12.82 (s, 1H), 7.36-7.31 (m, 1H), 7.23-7.15 (m, 4H), 7.11-7.07 (m,
2H), 3.56-3.47 (m, 1H), 3.38-3.33 (m, 1H), 3.20 (t, J = 12.0 Hz,
1H), 2.84-2.74 (bs, 7H), 2.65-2.56 (m, 1H), 2.38 (dd, J = 11.0, 5.0
Hz, 1H), 1.90-1.76 (m, 1H). 13C-NMR (500 MHz, CD3OD): δ
162.88, 135.80, 135.24, 134.90, 131.56, 129.51, 129.32, 129.25,
128.00, 125.90, 124.19, 115.64, 60.99, 41.33, 32.08, 26.89, 25.70.
Calcd C18H20FN for [M+H]+: 270.1658. Found: 270.1656 HPLC
(s-prep): Solvent System: hexanes/EtOH (85:15) 0.1% DEA
(modifier) 0.1% TFA (modifier); flow rate = 2.0 mL/min; t1 = 17.1,
25
t2 = 18.8. (+)-8b HCl: [a]D +39.82 (c 0.4, DCM), (–)-8b HCl:
[a]D25 – 40.30 (c 0.4, DCM).
7.1.25. 5-(2-chlorophenyl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl (8c)
7.1.21. 5-(furan-2-yl)-3,4-dihydronaphthalen-
2(1H)-one (7s)
Obtained from 7c (0.31 mmol) as a light yellow oil, yield; 49%
Obtained from 6 (0.34 mmol) as a light yellow oil, yield: 32%
(0.26 g), purification 92:8 (hexanes:EtOAC); 1H-NMR (500 MHz;
CDCl3): δ 7.54 (s, 1H), 7.51 (d, J = 7.8 Hz, 1H), 7.27-7.25 (m, 1H),
7.11 (d, J = 7.5 Hz, 1H), 6.52 (t, J = 2.2 Hz, 1H), 6.49 (d, J = 3.2
Hz, 1H), 3.64 (s, 2H), 3.28 (t, J = 6.6 Hz, 2H), 2.51 (t, J = 6.6 Hz,
2H).
(0.04 g), purification 9:1 (DCM:MeOH). The oil was converted to
1
the corresponding HCl salt. H NMR (500 MHz): δ 12.56 (s, 1H),
7.44-7.38 (m, 1H), 7.30-7.27 (m, 2H), 7.21-7.18 (dt, J = 7.0, 2.5
Hz, 1H), 7.15-7.10 (m, 2H), 6.99 (t, J = 6.0 Hz, 1H), 3.52-3.44 (m,
1H), 3.38-3.33 (m, 1H), 3.18 (t, J = 13.5 Hz, 1H), 2.78-2.83 (m,
6H), 2.72-2.45 (m, 2H), 2.34 (dd, J = 9.5, 2.0 Hz, 1H), 1.88-1.79
(m, 1H) 13C NMR (100 MHz): δ 139.7, 139.4, 139.2, 133.4, 132.8,
132.1, 131.9, 131.0, 130.6, 129.6, 129.2, 128.3, 128.0, 127.0,
126.4, 62.3, 39.9, 39.5, 30.1, 26.7, 26.0, 23.9 Calcd for C18H20ClN
[M+H]+: 286.1363. Found: 286.1368.
7.1.22. 5-(thiophen-2-yl)-3,4-
dihydronaphthalen-2(1H)-one (7t)
Obtained from 6 (0.29 mmol) as a light yellow oil, yield: 30%
(0.02 g), purification 94:6 (hexanes:EtOAc); 1H-NMR (500 MHz;
CDCl3): δ 7.38-7.34 (m, 2H), 7.25 (dd, J = 9.0, 6.3 Hz, 1H), 7.15-
7.11 (m, 2H), 7.03-7.02 (m, 1H), 3.65 (s, 2H), 3.19 (d, J = 6.6 Hz,
2H), 2.47 (t, J = 6.6 Hz, 2H).
7.1.26. 5-(3-fluorophenyl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine (8d)
Obtained from 7d (0.30 mmol) as a light yellow oil, yield; 50%
1
(0.04 g), purification 95:5 (DCM:MeOH). H NMR (500 MHz,
General reductive amination procedure for analogs 8a-d, h-
t (Scheme 1).
CDCl3) δ 7.38-7.34 (m, 1H), 7.22-7.14 (m, 2H), 7.07-7.02 (m, 3H),
6.98-6.96 (m, 1H), 3.19-3.15 (dd, J = 15.5, 3.5 Hz, 1H), 3.12-3.06
(m, 1H), 2.99-2.94 (dd, J =15.5, 11.0 Hz, 1H), 2.76-2.72 (m, 2H),
The corresponding 5-substituted-2-tetralone (7b-d, h-t) were
dissolved in
a 3:1 MeOH:THF solution. Dimethylamine
2.62 (s, 6H), 2.20-2.14 (m, 1H), 1.69-1.60 (ddd, J =18.0, 11.0, 6.5
13
hydrochloride (10 equivalents) was added at room temperature and
the solution stirred for 30 minutes. NaBH3CN (5 equivalents) was
added and the reaction was heated to 55 oC and stirred for 16 hours.
At completion solvent was evaporated off. Deionized water was
added followed by extraction with dichloromethane. Organic
fractions were combined, washed with saturated aqueous sodium
Hz, 1H). C-NMR (400 MHz, CDCl ): δ 163.91, 161.45, 143.24,
3
140.87, 132.47, 132.19, 129.91, 129.13, 128.44, 126.76, 124.93,
116.26, 116.04, 114.39, 114.18, 62.29, 39.77, 39.71, 30.16, 27.19,
24.17. Calcd for C18H20FN [M+H]+: 270.1658. Found: 270.1658.

7.1.27. 5-(4-fluorophenyl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl
(8h)
Obtained from 7l (0.33 mmol) as a clear oil, yield; 35% (0.037
g), purification 97:3 (DCM:MeOH). The oil was converted to the
1
corresponding HCl salt. H-NMR (500 MHz; CD OD): δ 7.41
3
(d, J = 1.7 Hz, 1H), 7.22-7.20 (m, 4H), 7.03 (dd, J = 6.0, 2.5 Hz,
1H), 3.65-3.58 (m, 1H), 3.28 (dd, J = 5.4, 1.7 Hz, 1H), 3.07 (dt, J
= 15.6, 7.8 Hz, 1H), 2.90 (s, 6H), 2.81-2.68 (m, 2H), 2.23 (dt, J =
6.1, 3.1 Hz, 1H), 1.76 (qd, J = 11.7, 5.6 Hz, 1H).13C NMR (400
MHz; CDCl3): δ 144.9, 139.1, 136.8, 134.7, 133.9, 129.8, 127.8,
127.15, 127.09, 126.0, 77.5, 77.2, 76.9, 60.5, 42.0, 33.0, 27.9, 26.3
Calcd C18H19Cl2N for [M+H]+: 320.0973. Found: 320.0975.
Obtained from 7h (0.2 mmol) as a yellow oil, yield; 59% (0.03
g), purification 95:5 (DCM:MeOH). The oil was converted to the
1
corresponding HCl salt. H-NMR (500 MHz; CDCl ): δ 12.54 (s,
3
1H), 7.20 (t, J = 6.1 Hz, 3H), 7.15 (d, J = 7.2 Hz, 1H), 7.09 (t, J =
8.8 Hz, 3H), 3.56 (m, 1H), 3.40 (d, J = 14.3 Hz, 1H), 3.23-3.18 (m,
1H), 2.86 (s, 6H), 2.78 (t, J = 3.0 Hz, 2H), 2.42 (d, J = 8.6 Hz, 1H),
13
1.84-1.80 (m, 1H). C-NMR (400 MHz, CDCl ): δ 163.33,
3
160.88, 140.97, 136.87, 132.38, 132.33, 130.66, 130.58, 128.80,
128.54, 126.59, 115.38, 115.12, 62.22, 39.89, 39.69, 30.10, 27.19,
24.20. Calcd C18H20FN for [M+H]+: 270.1658. Found: 270.1658.
7.1.32. 5-(3,5-bis(trifluoromethyl)phenyl)-N,N-
dimethyl-1,2,3,4-tetrahydronaphthalen-2-amine
HCl (8m)
7.1.28. 5-(4-chlorophenyl)-N,N-dimethyl-
Obtained from 7m (0.25 mmol) as a clear oil, yield; 46% (0.045
1,2,3,4-tetrahydronaphthalen-2-amine HCl (8i)
g), purification 95:5 (DCM:MeOH). The oil was converted to the
corresponding HCl salt. 1H-NMR (500 MHz; CD OD): δ 7.91 (s,
Obtained from 7i (0.29 mmol) as a yellow oil, yield: 36%
(0.03g), purification 9:1 (DCM:MeOH). The oil was converted to
the corresponding HCl salt; 1H-NMR (500 MHz; CDCl3): δ 12.69
(s, 1H), 7.38 (d, J = 8.2 Hz, 2H), 7.23 (t, J = 7.5 Hz, 1H), 7.17 (t,
J = 9.1 Hz, 3H), 7.08 (d, J = 7.4 Hz, 1H), 3.53 (s, 1H), 3.42-3.39
(m, 1H), 3.20 (t, J = 13.2 Hz, 1H), 2.85 (s, 6H), 2.79 (s, 2H), 2.41
(d, J = 11.0 Hz, 1H), 1.86-1.78 (m, 1H). 13C NMR (400 MHz;
CDCl3): δ 140.9, 139.4, 133.4, 132.5, 132.2, 130.5, 129.0, 128.62,
128.54, 126.8, 62.3, 39.80, 39.65, 30.1, 27.2, 24.2 Calcd
C18H20ClN for [M+H]+: 286.1363. Found: 286.1370. HPLC (s-
prep): Solvent System: hexanes/EtOH/MeOH/iPrOH/nPrOH
(80:5:5:5:5) 0.1% TEA (modifier); flow rate = 1.0 mL/min; t1 =
16.1, t2 = 18.1. (+)-8i HCl: [a]D25 +3.88 (c 0.3, MeOH), (–)-8i HCl:
[a]D25 – 4.12 (c 0.5 MeOH).
3
1H), 7.81 (s, 2H), 7.24 (d, J = 4.8 Hz, 2H), 7.07 (t, J = 4.4 Hz, 1H),
3.65-3.59 (m, 1H), 3.28 (dd, J = 15.7, 4.0 Hz, 1H), 3.08 (dd, J =
15.5, 11.1 Hz, 1H), 2.88 (s, 6H), 2.77 (ddd, J = 17.0, 11.6, 5.3 Hz,
1H), 2.62 (ddd, J = 16.9, 4.8, 3.6 Hz, 1H), 2.22 (dt, J = 9.4, 2.9 Hz,
13
1H), 1.75 (qd, J = 11.8, 5.2 Hz, 1H). C-NMR (400 MHz,
CDCl ): δ 142.94, 138.81, 132.87, 131.89, 131.83, 129.97,
3
129.17, 128.41, 127.02, 121.23, 61.94, 39.60, 30.03, 27.03, 23.84.
Calcd C20H19F6N for [M+H]+: 388.1500. Found: 388.1500. HPLC
(s-prep): Solvent System: hexanes/EtOH (95:5) 0.1% DEA
(modifier) 0.1% TFA (modifier); flow rate = 1.5 mL/min; t1 = 33.3,
t2 = 36.1. (–)-8m HCl: [a]D25 – 32.54 (c 0.3, CHCl3), (+)-8b HCl:
[a]D25 +30.93 (c 0.3 CHCl3).
7.1.33. N,N-dimethyl-5,6,7,8-tetrahydro-[1,1'-
binaphthalen]-6-amine (8n)
7.1.29. 5-(2-chloro-4-fluorophenyl)-N,N-
dimethyl-1,2,3,4-tetrahydronaphthalen-2-amine
HCl (8j)
Obtained from 7n (0.49 mmol) as a clear oil, yield: 71% (0.11
g), purification 95:5 (DCM:MeOH). 1H-NMR (500 MHz; CDCl3):
δ 7.89 (d, J = 8.2 Hz, 1H), 7.86 (d, J = 8.3 Hz, 1H), 7.53-7.50 (m,
1H), 7.47 (t, J = 7.4 Hz, 1H), 7.41-7.36 (m, 2H), 7.32-7.27 (m,
2H), 7.23 (q, J = 6.3 Hz, 1H), 7.14 (q, J =7.3 Hz, 1H), 3.38-3.26
(m, 2H), 3.14-3.05 (m, 1H), 2.74 (d, J = 7.0 Hz, 6H), 2.55 (ddd, J
= 15.3, 6.0, 3.5 Hz, 1H), 2.47-2.39 (m, 1H), 2.07-2.04 (m, 1H),
1.67 (dd, J = 11.9, 5.3 Hz, 1H). 13C-NMR (400 MHz, CDCl3): δ
140.32, 140.00, 138.82, 138.43, 133.59, 133.44, 133.39, 133.36,
132.24, 132.00, 131.79, 131.66, 129.13, 128.91, 128.87, 128.39,
128.31, 127.80, 127.76, 126.78, 126.49, 126.37, 126.30, 126.24,
125.93, 125.89, 125.69, 125.48, 125.37, 62.32, 39.91, 30.45,
30.43, 26.74, 26.23, 24.30, 24.00. Calcd C22H23N for [M+H]+:
302.1909. Found: 302.1907.
Obtained from 7j (0.19 mmol) as a light yellow oil, yield; 17%
(0.01 g), purification 95:5 (DCM:MeOH). The oil was converted
1
to the corresponding HCl salt. H-NMR (500 MHz; CD OD):
3
δ 7.33 (ddd, J = 9.5, 8.8, 2.6 Hz, 1H), 7.27-7.23 (m, 3H), 7.16
(tdd, J = 8.4, 5.7, 2.7 Hz, 1H), 6.99 (td, J = 4.2, 2.0 Hz, 1H), 3.65
(dtt, J = 14.8, 8.8, 3.1 Hz, 1H), 3.34-3.28 (m, 1H), 3.14 (dt, J =
15.5, 10.0 Hz, 1H), 2.95 (d, J = 2.7 Hz, 6H), 2.73-2.67 (m, 1H),
13
2.63-2.57 (m, 1H), 2.30-2.24 (m, 1H), 1.88-1.78 (m, 1H). C-
NMR (400 MHz, CDCl ): δ 163.39, 160.91, 138.65, 138.43,
3
135.99, 135.96, 135.69, 134.44, 134.34, 133.86, 133.76, 133.23,
133.01, 132.39, 132.14, 131.79, 131.71, 129.63, 128.76, 128.47,
126.77, 126.75, 117.28, 117.15, 117.03, 114.58, 114.55, 114.38,
114.34, 62.50, 62.21, 40.22, 39.90, 39.78, 39.69, 30.30, 30.03,
26.94, 26.19, 24.10, 23.98. Calcd for C18H19ClFN [M+H]+:
304.1268. Found: 304.1272.
7.1.34. 5-(isoquinolin-4-yl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl (8o)
Obtained from 7o (0.25 mmol) as an orange oil, yield; 43%
7.1.30. 5-(3,5-difluorophenyl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine (8k)
(0.03g), purification 85:15 (DCM;MeOH). The oil was converted
1
to the corresponding HCl salt. H-NMR (500 MHz; CD OD):
3
δ 9.17 (s, 1H), 8.15 (d, J = 14.8 Hz, 1H), 8.09 (dd, J = 6.1, 2.7
Hz, 1H), 7.62 (dt, J = 7.6, 3.6 Hz, 2H), 7.40-7.30 (m, 1H), 7.40-
7.30 (m, 1H), 7.20 (dd, J = 6.1, 2.9 Hz, 2H), 6.98 (ddd, J = 13.6,
6.2, 2.5 Hz, 1H), 3.08 (dt, J = 15.8, 4.3 Hz, 1H), 2.85 (dt, J = 16.5,
8.6 Hz, 1H), 2.73 (ddt, J = 10.5, 6.8, 3.6 Hz, 1H), 2.42-2.36 (m,
Obtained from 7k (0.4 mmol) as a clear oil, yield; 41% (0.047
g), purification 95: (DCM:MeOH). 1H-NMR (500 MHz; CDCl3):
δ 7.22 (t, J = 7.5 Hz, 1H), 7.17 (d, J = 7.5 Hz, 1H), 7.06 (d, J =
7.3 Hz, 1H), 6.81-6.76 (m, 3H), 3.50-3.46 (m, 1H), 3.28 (dd, J =
15.6, 4.1 Hz, 1H), 3.07 (dd, J = 15.3, 11.3 Hz, 1H), 2.82 (s, 6H),
7H), 2.26-2.21 (m, 1H), 1.94-1.87 (m, 1H), 1.47-1.41 (m, 1H).
2.78 (dd, J = 8.0, 3.9 Hz, 2H), 2.28 (d, J = 11.7 Hz, 1H), 1.79-1.71
13
13
C-NMR (400 MHz, CDCl ): δ 152.22, 152.18, 152.08, 152.02,
(m, 1H). C-NMR (400 MHz, CDCl ): δ 164.17, 161.69, 158.11,
3
3
143.03, 142.62, 136.28, 136.04, 135.99, 135.79, 135.31, 135.26,
134.83, 134.75, 132.95, 132.60, 130.73, 130.67, 129.78, 129.73,
128.61, 128.41, 128.37, 128.24, 128.04, 127.95, 127.36, 127.32,
126.01, 125.93, 125.08, 124.87, 60.88, 41.57, 41.47, 32.54, 32.40,
27.72, 27.22, 25.95, 25.57. Calcd C21H22N2 for [M+H]+: 303.1861.
Found: 303.1862.
139.87, 132.63, 132.05, 129.59, 128.22, 126.86, 112.38, 112.13,
103.09, 102.85, 102.59, 62.45, 40.22, 40.15, 30.26, 27.02, 24.21.
The oil was converted to the corresponding HCl salt. Calcd
C18H19F2N for [M+H]+: 288.1564. Found: 288.1569.
7.1.31. 5-(3,5-dichlorophenyl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl (8l)
7.1.35. 5-(anthracen-9-yl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl (8p)

Obtained from 7p (0.7 mmol) as a clear oil, yield; 81% (0.2 g),
(ddd, J = 12.2, 5.4, 2.6 Hz, 1H), 1.69 (dt, J = 11.9, 5.9 Hz, 1H).
13
purification 97:3 (DCM:MeOH). The oil was converted to the
C-NMR (400 MHz, CDCl ): δ 142.18, 134.38, 133.92, 133.73,
3
1
corresponding HCl salt. H-NMR (500 MHz; CD OD): δ 8.44 (s,
129.49, 129.37, 127.29, 126.93, 126.44, 125.64, 61.79, 40.61,
31.19, 27.77, 25.15. Calcd C16H19NS for [M+H]+: 258.1316.
Found: 258.1323. HPLC (s-prep): Solvent System: hexanes/EtOH
(9:1) 0.1% DEA (modifier) 0.1% TFA (modifier); flow rate = 2.0
mL/min; t1 = 29.4, t2 = 34.0. (+)-8t: [a]D25 +19.11 (c 0.2, MeOH),
(–)-8t: [a]D25 – 18.75 (c 0.1, MeOH).
3
1H), 7.97 (dd, J = 8.2, 3.9 Hz, 2H), 7.34 (m, 5H), 7.25 (m, 3H),
7.00 (d, J = 6.7 Hz, 1H), 3.56-3.51 (m, 1H), 3.34 (dd, J = 15.5, 2.9
Hz, 1H), 3.15 (dd, J = 14.8, 12.1 Hz, 1H), 2.81 (s, 6H), 2.24-2.11
13
(m, 2H), 1.93-1.90 (m, 1H), 1.60 (qd, J = 11.9, 5.7 Hz, 1H). C-
NMR (400 MHz, CDCl ): δ 138.20, 135.09, 134.06, 132.44,
3
131.49, 131.45, 130.01, 129.99, 129.65, 129.24, 128.74, 128.64,
126.84, 125.99, 125.94, 125.39, 125.31, 62.46, 39.81, 30.62,
26.21, 23.67. Calcd C26H25N for [M+H]+: 352.2065. Found:
352.2061.
7.1.40. (2S)-5-methoxy-N,N-dimethyl-1,2,3,4-
tetrahydronaphthalen-2-amine (10)
(S)-5-methoxy-1,2,3,4-tetrahydronaphthalen-2-amine HCl 9
(0.25 g, 1.2 mmol) was dissolved in 12 mL MeOH. Formaldehyde
(0.12 mL, 4.4 mmol) was added and the reaction stirred at reflux
for 1 hour. The reaction was then cooled to room temperature and
286 mg (7.5 mmol) NaBH4 was added and stirred for 1 hour.
Solvent was concentrated in vacuo and saturated aqueous
ammonium chloride was added. Extraction was done with
dichloromethane (3 x 20 mL) and organic fractions combined and
dried over sodium sulfate. No further purification was done,
yielding 0.2 g (83%) of a clear oil. TLC eluents used (95:5
7.1.36. 5-cyclopentyl-N,N-dimethyl-1,2,3,4-
tetrahydronaphthalen-2-amine (8q)
Obtained from 7q (0.014 mmol) as a clear oil, yield; 26%
1
(0.003 g), purification 98:2 (DCM:MeOH). H-NMR (500 MHz;
CDCl ): δ 7.19-7.14 (m, 2H), 6.96 (d, J = 6.8 Hz, 1H), 3.45 (tdd,
3
J = 11.7, 4.7, 3.3 Hz, 1H), 3.21-3.16 (m, 2H), 3.12-3.03 (m, 2H),
2.85 (s, 6H), 2.44 (ddd, J = 11.9, 5.6, 2.8 Hz, 1H), 2.00 (m, 2H),
1.91-1.78 (m, 3H), 1.73-1.65 (m, 2H), 1.62-1.48 (m, 3H). Calcd
C17H25N for [M+H]+: 244.2065 Found: 244.2066.
1
Dichloromethane: MeOH). H-NMR (500 MHz; CDCl3): δ 7.06
(t, J = 7.9 Hz, 1H), 6.69 (d, J = 7.6 Hz, 1H), 6.62 (d, J = 8.1 Hz,
1H), 3.77 (s, 3H), 2.96 (ddd, J = 17.6, 5.2, 3.3 Hz, 1H), 2.90 (dd,
J = 15.9, 3.2 Hz, 1H), 2.73 (dd, J = 15.8, 10.6 Hz, 1H), 2.55-2.48
(m, 2H), 2.11 (ddd, J = 12.4, 5.6, 2.9 Hz, 1H), 1.54 (qd, J = 11.9,
5.6 Hz, 1H).
7.1.37. N,N-dimethyl-5-(1-methyl-1H-pyrrol-2-
yl)-1,2,3,4-tetrahydronaphthalen-2-amine (8r)
Obtained from 7r (0.62 mmol) as a red oil, yield: 69% (0.11g),
1
purification 9:1 (DCM:MeOH); H-NMR (500 MHz; CDCl ):
3
δ 7.20-7.15 (m, 2H), 7.10 (dd, J = 6.6, 2.1 Hz, 1H), 6.72 (t, J =
2.2 Hz, 1H), 6.23 (dd, J = 3.4, 2.8 Hz, 1H), 6.07 (dd, J = 3.5, 1.8
Hz, 1H), 3.41 (s, 3H), 3.07-3.02 (m, 1H), 2.85 (dd, J = 15.9, 10.7
Hz, 1H), 2.73-2.67 (m, 1H), 2.64 (tdd, J = 10.8, 5.1, 3.1 Hz, 1H),
2.56 (ddd, J = 22.0, 10.7, 4.9 Hz, 1H), 2.39 (s, 6H), 2.10-2.05 (m,
7.1.41. (2S)-6-(dimethylamino)-5,6,7,8-
tetrahydronaphthalen-1-ol (11)
Intermediate 10 (200 mg, 0.97 mmol) was dissolved in 12 mL
anhydrous dichloromethane under N2. The solution was cooled to
13
o
1H), 1.55 (qd, J = 11.9, 5.2 Hz, 1H). C-NMR (400 MHz,
0 C and 0.5 mL (5.1 mmol) of BBr3 was added dropwise. The
CDCl ): δ 136.95, 136.42, 132.96, 132.75, 129.47, 128.86,
solution was then brought to room temperature and stirred for 20
hours. The reaction was quenched with saturated aqueous sodium
bicarbonate and then basified to a pH of 9 using 2N NaOH.
Extraction was done with dichloromethane (3 x 15 mL) and
organic fractions dried over sodium sulfate and concentrated in
vacuo. Crude product was dissolved in hot ethyl acetate and the
remaining white precipitate was filtered and collected, yielding
0.15 g (80%) of a white solid. TLC eluents used (9:1
3
125.36, 121.63, 108.41, 107.27, 60.66, 41.88, 34.20, 32.73, 27.71,
26.25. Calcd C17H22N2 for [M+H]+: 255.1861. Found: 255.1853.
HPLC (s-prep): Solvent System: hexanes/EtOH (85:15) 0.1%
DEA (modifier) 0.1% TFA (modifier); flow rate = 2.0 mL/min; t1
25
= 19.1, t2 = 22.5. (+)-8r: [a]D25 +10.5 (c 0.3, MeOH), (–)-8r: [a]D
–14.3 (c 0.3, MeOH).
7.1.38. 5-(furan-2-yl)-N,N-dimethyl-1,2,3,4-
tetrahydronaphthalen-2-amine HCl (8s)
1
Dichloromethane: MeOH). H-NMR (500 MHz; CD3OD): δ 6.89
(t, J = 7.8 Hz, 1H), 6.55 (dd, J = 11.7, 7.9 Hz, 2H), 3.50 (tdd, J =
11.3, 4.7, 2.9 Hz, 1H), 3.06 (dt, J = 15.2, 2.2 Hz, 1H), 3.01-2.93
(m, 2H), 2.86 (s, 6H), 2.58 (ddd, J = 17.6, 11.3, 6.1 Hz, 1H), 2.25
(ddd, J = 12.1, 5.8, 3.0 Hz, 1H), 1.77 (qd, J = 11.9, 5.8 Hz, 1H).
Obtained from 7s (0.30 mmol) as a clear oil, yield; 61% (0.05
g), purification 9:1 (DCM:MeOH). The oil was converted to the
1
corresponding HCl salt. H-NMR (500 MHz; CDCl3): δ 12.78 (s,
1H), 7.54-7.50 (m, 2H), 7.23 (t, J = 7.7 Hz, 1H), 7.10 (d, J = 7.6
Hz, 1H), 6.50-6.47 (m, 2H), 3.51 (d, J = 8.7 Hz, 1H), 3.37-3.34
(m, 1H), 3.22-3.14 (m, 2H), 3.03 (t, J = 6.1 Hz, 1H), 2.85 (d, J =
7.1.42. (2S)-6-(dimethylamino)-5,6,7,8-
tetrahydronaphthalen-1-yl
trifluoromethanesulfonate (12)
4.6 Hz, 6H), 2.57-2.55 (m, 1H), 1.87 (qd, J = 12.0, 5.2 Hz, 1H).
13
Intermediate 11 (0.15 g, 0.76 mmol) was dissolved in 2.5 mL
anhydrous dichloromethane under N2. 0.21 mL (1.22 mmol) of
N,N-Diisopropylethylamine was added and stirred for 15 minutes
at room temperature. The reaction was cooled to 0 oC and 0.33 g
(0.91 mmol) 2-pyridyltriflimide was added and the reaction stirred
overnight at room temperature. At completion extraction was done
with dichloromethane (3 x 10 mL) and the organic fractions
washed with saturated aqueous sodium chloride before being dried
over sodium sulfate and concentrated in vacuo. Purification was
done by flash chromatography 95:5 (DCM:MeOH) to yield 0.06 g
(25%) of the product as a light red oil. TLC eluents used (9:1
Dichloromethane: MeOH). 1H-NMR (500 MHz; CDCl3): δ 7.18
(t, J = 7.9 Hz, 1H), 7.12 (d, J = 7.5 Hz, 1H), 7.07 (d, J = 8.0 Hz,
1H), 3.10-3.01 (m, 2H), 2.84 (dd, J = 16.0, 10.6 Hz, 1H), 2.77-
C-NMR (400 MHz, CDCl ): δ 152.90, 142.35, 132.58, 131.58,
3
130.60, 129.21, 126.79, 111.55, 109.46, 62.10, 39.99, 39.36,
30.09, 27.70, 24.39. Calcd C16H19NO for [M+H]+: 242.1545.
Found: 242.1545. HPLC (s-prep): Solvent System: HPLC (s-
prep): Solvent System: hexanes/EtOH (85:15) 0.1% DEA
(modifier) 0.1% TFA (modifier); flow rate = 2.0 mL/min; t1 = 21.1,
25
t2 = 25.1.(+)-8s HCl: [a]D +29.38 (c 0.3, MeOH), (–)-8s HCl:
[a]D25 – 30.98 (c 0.2, MeOH).
7.1.39. N,N-dimethyl-5-(thiophen-2-yl)-1,2,3,4-
tetrahydronaphthalen-2-amine (8t)
Obtained from 7t (0.19 mmol) as a green oil, yield: 73% (0.035
1
g), purification 9:1 (DCM:MeOH); H-NMR (500 MHz; CDCl ):
3
δ 7.33 (dd, J = 5.1, 1.1 Hz, 1H), 7.25 (d, J = 7.4 Hz, 1H), 7.18 (t,
J = 7.6 Hz, 1H), 7.13 (d, J = 7.5 Hz, 1H), 7.08 (dd, J = 5.1, 3.5 Hz,
1H), 7.00 (dd, J = 3.5, 1.1 Hz, 1H), 3.21-3.14 (m, 2H), 3.06-2.96
(m, 2H), 2.87 (ddd, J = 17.1, 11.7, 5.4 Hz, 1H), 2.67 (s, 6H), 2.26
2.71 (m, 2H), 2.44 (s, 6H), 2.23-2.18 (m, 1H), 1.67-1.62 (m, 1H).
13
C-NMR (400 MHz, CDCl ): δ 148.23, 139.75, 129.72, 129.54,
3
127.07, 123.54, 120.35, 118.54, 117.17, 113.97, 59.95, 41.94,

32.51, 25.20, 23.62. Calcd C13H17NO3SF3 for [M+H]+: 324.0881.
Found: 324.0083.
11.4, 4.2 Hz, 1H), 2.31 (d, J = 3.8 Hz, 7H), 2.25-2.19 (m, 1H),
1.90-1.82 (m, 1H), 1.46-1.35 (m, 1H). C-NMR (400 MHz,
13
CDCl ): δ 140.33, 139.99, 138.77, 138.36, 133.59, 133.43,
3
7.1.43. (2S)-5-(2-fluorophenyl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl
((2S)-8b)
133.25, 133.20, 132.07, 131.82, 131.76, 131.64, 129.16, 128.92,
128.89, 128.40, 128.30, 127.83, 127.78, 126.78, 126.54, 126.41,
126.35, 126.29, 126.16, 125.92, 125.89, 125.67, 125.44, 125.34,
62.54, 62.46, 46.28, 30.47, 30.36, 26.81, 26.34, 24.07, 23.70.
Calcd C22H23N for [M+H]+: 302.1909. Found: 302.1909. (2S)-8n:
[a]D25 – 59.5 (c 0.5, DCM).
Intermediate 12 (0.13 g, 0.4 mmol) was dissolved in 3 mL
dimethoxyethane and 0.3 mL water. The solution was degassed
with N2 and 0.112 g (0.8 mmol) K2CO3 was added followed by
0.062 g (0.44 mmol) of 2'-F-boronic acid and 0.071 g (0.06 mmol)
Pd[PPh3]4. The reaction stirred for 4 hours at reflux and was
quenched with water and extracted with ethyl acetate (3 x 20mL)
and organic fractions were dried over sodium sulfate. Purification
was done by flash chromatography 95:5 (DCM:MeOH) to yield
0.03 g (23%) as a yellow oil. The oil was converted to the
General conditions for (2S) or (2R)-6-(dipropylamino)-
5,6,7,8-tetrahydronaphthalen-1-ol ((2S)-13 or (2R)-13)
The starting chiral amine (2S)-9 or (2R)-9 was dissolved in HBr
o
(48% aq) and heated to 130 C for 3 hours. The solvent was
evaporated off and the crude salt was dissolved in methanol. 20
equivalents of propionaldehyde were added followed by
montmorillonite K 10 (0.3 mg per mg of crude salt), and 20
equivalents of NaBH3CN. The reaction stirred for 20 hours at room
temperature and at completion solvent was evaporated off. 2N
NaOH was added and extraction was done with ethyl acetate.
Organic fractions were washed with brine and dried over sodium
sulfate. Purification was done by flash chromatography 97:3
(DCM:MeOH). TLC eluents used (95:5 Dichloromethane:
MeOH).
corresponding HCl salt. TLC eluents used (9:1 Dichloromethane:
1
MeOH). H-NMR (500 MHz; CDCl ): δ 11.82 (s, 1H), 7.37 (td,
3
J = 9.3, 4.3 Hz, 1H), 7.27-7.24 (m, 1H), 7.20 (t, J = 7.0 Hz, 3H),
7.13 (t, J = 8.0 Hz, 2H), 3.56-3.53 (m, 1H), 3.38-3.35 (m, 1H),
3.19 (t, J = 12.7 Hz, 1H), 2.86-2.80 (m, 7H), 2.70-2.67 (m, 1H),
13
2.38 (d, J = 10.2 Hz, 1H), 1.85-1.83 (m, 1H). C-NMR (400 MHz,
CD OD): δ 133.52, 131.33, 129.64, 129.55, 129.21, 128.53,
3
126.26, 126.25, 124.36, 124.30, 115.47, 62.54, 39.24, 29.85,
26.45, 23.98. Calcd C18H20FN for [M+H]+: 270.1658. Found:
270.1656. (2S)-8b HCl: [a]D25 – 41.2 (c 0.7, DCM).
7.1.46. (2S)-6-(dipropylamino)-5,6,7,8-
tetrahydronaphthalen-1-ol ((2S)-13)
7.1.44. (2S)-5-(2-chlorophenyl)-N,N-dimethyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl
((2S)-8c)
Obtained from (2S)-9 (0.94 mmol) to yield 0.13 g as a clear oil
(56% two-step yield). 1H-NMR (500 MHz; CD3OD): δ 6.89 (t, J
= 7.8 Hz, 1H), 6.57 (d, J = 7.6 Hz, 1H), 6.53 (d, J = 7.9 Hz, 1H),
3.58 (tdd, J = 11.6, 5.0, 2.7 Hz, 1H), 3.11 (t, J = 8.2 Hz, 4H), 3.05-
2.94 (m, 3H), 2.56 (ddd, J = 17.5, 11.9, 5.7 Hz, 1H), 2.23 (ddd, J
= 12.0, 5.6, 2.7 Hz, 1H), 1.80 (ddt, J = 18.2, 12.2, 6.1 Hz, 1H),
1.71 (dq, J = 16.1, 7.8 Hz, 4H), 0.97 (s, 6H).
Intermediate 12 (0.065 g, 0.2 mmol) was dissolved in 1 mL
dioxane. The solution was degassed with N2 and 0.068 g (0.3
mmol) K3PO4 was added followed by 0.038 g (0.24 mmol) of 2'-
Cl-boronic acid, 0.026 g KBr (0.22 mmol) and 0.023 g (0.02
mmol) Pd[PPh3]4. The reaction stirred for 6 hours at reflux and was
quenched with water and extracted with ethyl acetate (3 x 20mL)
and organic fractions were dried over sodium sulfate. Purification
was done by flash chromatography 95:5 (DCM:MeOH) to yield
0.04 g (61%) as a yellow oil. The oil was converted to the
corresponding HCl salt. TLC eluents used (9:1 Dichloromethane:
MeOH). 1H-NMR (500 MHz; CDCl3): δ 12.49 (s, 1H), 7.45 (m,
1H), 7.32 (dd, J = 8.3, 4.4 Hz, 2H), 7.23 (t, J = 7.0 Hz, 1H), 7.19-
7.14 (m, 2H), 7.03 (dd, J = 6.7, 4.4 Hz, 1H), 3.53-3.49 (m, 1H),
3.39 (s, 1H), 3.22 (t, J = 12.4 Hz, 1H), 2.86 (s, 6H), 2.73-2.52 (m,
7.1.47. (2R)-6-(dipropylamino)-5,6,7,8-
tetrahydronaphthalen-1-ol ((2R)-13)
Obtained from (2R)-9 (1.2 mmol) to yield 0.22 g as a clear oil
(76% two-step yield). 1H-NMR (500 MHz; CD3OD): δ 6.88 (t, J
= 7.8 Hz, 1H), 6.56 (d, J = 7.6 Hz, 1H), 6.53 (d, J = 8.0 Hz, 1H),
3.60 (tdd, J = 11.5, 5.0, 2.6 Hz, 1H), 3.12-3.10 (m, 4H), 3.05-2.94
(m, 3H), 2.56 (ddd, J = 17.5, 11.9, 5.6 Hz, 1H), 2.23 (ddd, J = 11.9,
5.5, 2.7 Hz, 1H), 1.81 (tt, J = 12.1, 6.1 Hz, 1H), 1.73 (tq, J = 14.5,
7.2 Hz, 4H), 0.97 (t, J = 7.4 Hz, 6H).
13
2H), 2.40-2.38 (m, 1H), 1.87 (t, J = 5.4 Hz, 1H). C-NMR (400
MHz, CDCl ): δ 139.71, 139.43, 139.40, 139.16, 133.42, 132.93,
3
General conditions for (2S) or (2R)-6-(dipropylamino)-
5,6,7,8-tetrahydronaphthalen-1-yl trifluoromethanesulfonate
((2S)-14 or (2R)-14)
132.79, 132.72, 132.19, 131.95, 131.01, 130.65, 129.57, 129.53,
129.46, 129.28, 129.20, 129.04, 128.26, 127.97, 126.97, 126.91,
126.44, 62.33, 62.06, 40.03, 39.72, 30.17, 29.97, 26.74, 26.01,
23.94, 23.84. Calcd C18H20ClN for [M+H]+: 286.1363. Found:
286.1368. (2S)-8c HCl: [a]D25 – 45.5 (c 0.5, MeOH).
Either (2S)-13 or (2R)-13 was dissolved in anhydrous DCM,
and 1.5 equivalents of 2-Pyridyltriflimide were added. The flask
was placed under N2 atmosphere and cooled to -78 oC. 3.0
equivalents of N,N-diisopropylethylamine were added and the
reaction slowly warmed to room temperature and stirred for 20
hours. Saturated aqueous ammonium chloride was added, and
extraction done with DCM. Organic fractions were dried over
sodium sulfate and purification was done by flash chromatography
98:2 (DCM:MeOH). TLC eluents used (95:5 Dichloromethane:
MeOH).
7.1.45. (2S)-N,N-dimethyl-5,6,7,8-tetrahydro-
[1,1'-binaphthalen]-6-amine ((2S)-8n)
Intermediate 12 (0.060 g, 0.19 mmol) was dissolved in 1.5 mL
dimethoxyethane and 0.15 mL water. The solution was degassed
with N2 and 0.101 g (0.73 mmol) K2CO3 was added followed by
0.126 g (0.73 mmol) of 1-naphthyl-boronic acid and 0.033 g (0.03
mmol) Pd[PPh3]4. The reaction stirred for 4 hours at reflux and was
quenched with water and extracted with ethyl acetate (3 x 20mL)
and organic fractions were dried over sodium sulfate. Purification
was done by flash chromatography 95:5 (DCM:MeOH) to yield
0.03 g (53%) as a yellow oil. TLC eluents used (9:1
7.1.48. (2S)-6-(dipropylamino)-5,6,7,8-
tetrahydronaphthalen-1-yl
trifluoromethanesulfonate ((2S)-14)
1
Dichloromethane: MeOH). H-NMR 1H-NMR (400 MHz;
Obtained from (2S)-13 (0.53 mmol) to yield 0.13 g (65%) of a
1
CDCl3): δ 7.80 (d, J = 8.2 Hz, 1H), 7.76 (d, J = 8.2 Hz, 1H), 7.44-
7.35 (m, 2H), 7.30-7.27 (m, 1H), 7.22 (dd, J = 13.8, 7.0 Hz, 1H),
7.17-7.11 (m, 2H), 7.00 (t, J = 8.8 Hz, 1H), 3.07-2.99 (m, 1H),
2.83 (dt, J = 16.9, 9.0 Hz, 1H), 2.67-2.58 (m, 1H), 2.39 (td, J =
yellow oil. H-NMR (500 MHz; CDCl ): δ 7.19 (t, J = 7.9 Hz,
3
1H), 7.14 (d, J = 7.5 Hz, 1H), 7.08 (d, J = 8.0 Hz, 1H), 3.17 (t, J =
11.2 Hz, 1H), 3.12-3.02 (m, 2H), 2.91 (t, J = 13.7 Hz, 1H), 2.75
(ddd, J = 17.4, 12.0, 5.5 Hz, 1H), 2.66 (t, J = 7.4 Hz, 4H), 2.22-

2.19 (m, 1H), 1.69 (qd, J = 12.2, 5.3 Hz, 1H), 1.59 (dq, J = 15.0,
7.4 Hz, 4H), 0.93 (t, J = 7.3 Hz, 6H).
salt. 1H-NMR (500 MHz; CDCl3): δ 12.02 (s, 1H), 7.41 (t, J = 7.3
Hz, 2H), 7.35 (t, J = 7.1 Hz, 1H), 7.23 (dd, J = 12.5, 7.6 Hz, 3H),
7.15 (d, J = 7.5 Hz, 1H), 7.12 (d, J = 7.4 Hz, 1H), 3.68 (s, 1H),
3.39-3.36 (m, 1H), 3.28 (t, J = 13.0 Hz, 1H), 3.09 (dd, J = 15.9,
13.0 Hz, 2H), 2.98 (t, J = 5.4 Hz, 2H), 2.80 (d, J = 5.1 Hz, 2H),
2.45 (d, J = 10.9 Hz, 1H), 2.05 (s, 2H), 1.95 (d, J = 4.5 Hz, 2H),
7.1.49. (2R)-6-(dipropylamino)-5,6,7,8-
tetrahydronaphthalen-1-yl
trifluoromethanesulfonate ((2R)-14).
Obtained from (2R)-13 (0.88 mmol) to yield 0.29 g (86%) of a
13
1.86 (dd, J = 19.3, 10.7 Hz, 1H), 1.02 (q, J = 5.3 Hz, 6H). C-
1
yellow oil. H-NMR (500 MHz; CDCl ): δ 7.17 (t, J = 7.8 Hz,
3
NMR (400 MHz, CDCl ): δ 142.19, 141.06, 132.59, 132.53,
3
1H), 7.12 (d, J = 7.5 Hz, 1H), 7.06 (d, J = 8.0 Hz, 1H), 3.09-2.98
(m, 2H), 2.96 (dd, J = 16.3, 3.0 Hz, 1H), 2.87-2.81 (m, 1H), 2.73
(ddd, J = 17.3, 12.0, 5.5 Hz, 1H), 2.55 (t, J = 7.5 Hz, 4H), 2.14 (dt,
J = 10.0, 2.6 Hz, 1H), 1.63 (qd, J = 12.1, 5.3 Hz, 1H), 1.51 (sextet,
J = 7.4 Hz, 4H), 0.90 (t, J = 7.3 Hz, 6H).
129.13, 128.70, 128.58, 128.41, 127.37, 126.62, 59.69, 52.45,
52.09, 30.29, 27.61, 24.53, 18.16, 11.63. Calcd C22H29N for
25
[M+H]+: 308.2378. Found: 308.2376. 22 HCl: [a]D –33.14 (c
0.5, MeOH).
7.1.53. (2R)-5-phenyl-N,N-dipropyl-1,2,3,4-
tetrahydronaphthalen-2-amine ((2R)-16).
General Suzuki-coupling reaction conditions for (2S) or
(2R)-5-(substituted)-N,N-dipropyl-1,2,3,4-
tetrahydronaphthalen-2-amines (2S)- or (2R)-15 and (2S)- or
(2R)-16.
Obtained from (2R)-14 (0.11 mmol) to yield 0.01 g (29%) as a
1
clear oil, purification 98:2 (DCM:MeOH). H-NMR (500 MHz;
CDCl ): δ 7.41-7.38 (m, 2H), 7.36-7.33 (m, 1H), 7.26-7.24 (m,
3
2H), 7.20 (d, J = 7.5 Hz, 1H), 7.14 (d, J = 7.4 Hz, 1H), 7.09 (d, J
= 7.4 Hz, 1H), 3.50-3.44 (m, 1H), 3.20 (dd, J = 15.8, 4.0 Hz, 1H),
3.06 (dd, J = 15.6, 11.5 Hz, 1H), 2.88 (t, J = 8.2 Hz, 4H), 2.78-
The corresponding triflate was dissolved in 1,4-dioxane. The
solution was degassed with N2. The corresponding boronic acid
(1.5 equivalents) was added followed by 0.10 equivalents of
Pd(PPh3)4, 2 equivalents of K3PO4 and 1.4 equivalents of KBr. The
reaction was heated to 120 oC and stirred for 6 hours. The reaction
was quenched with water and extraction done with ethyl acetate.
Organic fractions were dried over sodium sulfate and purification
was done by flash chromatography (DCM:MeOH). TLC eluents
used (95:5 Dichloromethane: MeOH).
2.75 (m, 2H), 2.22-2.20 (m, 1H), 1.81-1.68 (m, 5H), 0.99 (t, J =
13
7.4 Hz, 6H). C-NMR (400 MHz, CDCl ): δ 142.11, 141.31,
3
133.75, 132.97, 129.17, 128.77, 128.35, 128.29, 127.25, 126.40,
59.00, 52.68, 30.84, 27.86, 24.89, 19.37, 11.70. The oil was
converted to the corresponding HCl salt. Calcd C22H29N for
25
[M+H]+: 308.2378. Found: 308.2375. 23: [a]D +32.90 (c 0.13,
MeOH).
7.1.50. (2S)-5-(2’-chlorophenyl)-N,N-dipropyl-
1,2,3,4-tetrahydronaphthalen-2-amine ((2S)-
15)
General
conditions
for
5-(2’-fluorophenyl)-N,N-
substituted-1,2,3,4-tetrahydronaphthalen-2-amines
(Scheme 4)
(17-18)
Obtained from (2S)-14 (0.17 mmol) to yield 0.01g (21%) as a
1
light yellow oil, purification 97:3 (DCM:MeOH). H-NMR (500
Tetralone 7b was dissolved in a 3:1 MeOH:THF mixture. 10
equivalents of the corresponding amine were added followed by
7.5 equivalents of NaBH3CN. The solution was heated to 65 C
and stirred for 16 hours. At completion solvent was evaporated off.
Deionized water was added followed by extraction with
dichloromethane. Organic fractions were combined, washed with
saturated aqueous sodium bicarbonate, dried over sodium sulfate
and concentrated in vacuo. Purification was done by flash
chromatography (Dichloromethane w/ MeOH). TLC eluents used
(95:5 Dichloromethane: MeOH).
MHz; CDCl ): δ 7.46 (m, 1H), 7.31 (m, 2H), 7.22 (m, 1H), 7.18
3
(m, 2H), 7.02 (t, J = 5.6 Hz, 1H), 3.59-3.44 (m, 1H), 3.29-3.15 (m,
2H), 3.00-2.82 (m, 4H), 2.68-2.50 (m, 2H), 2.34-2.23 (m, 1H),
o
1.93-1.77 (m, 5H), 0.98 (d, J = 5.4 Hz, 6H). The oil was converted
13
to the corresponding HCl salt. C-NMR (400 MHz, CDCl ):
3
δ 139.72, 139.41, 139.34, 139.17, 133.44, 133.03, 132.86, 132.78,
132.45, 132.19, 130.99, 130.61, 129.51, 129.39, 129.14, 128.94,
128.17, 127.88, 126.86, 126.82, 126.31, 59.52, 59.36, 52.57,
52.30, 52.27, 52.15, 51.95, 30.43, 30.02, 27.01, 26.23, 24.09,
18.26, 17.81, 11.53. Calcd C22H28ClN for [M+H]+: 342.1989.
Found: 342.1979. 20 HCl: [a]D25 –35.18 (c 0.25, MeOH).
7.1.54. 5-(2-fluorophenyl)-N,N-dipropyl-
1,2,3,4-tetrahydronaphthalen-2-amine (17)
7.1.51. (2R)-5-(2-chlorophenyl)-N,N-dipropyl-
1,2,3,4-tetrahydronaphthalen-2-amine HCl
((2R)-15).
Obtained from 7b (0.26 mmol) to yield 0.02 g (25%) as a light
1
red oil, purification 98:2 (DCM:MeOH). H-NMR (500 MHz;
CDCl3): δ 7.35 (dt, J = 8.5, 4.3 Hz, 1H), 7.19 (dt, J = 14.9, 7.5 Hz,
4H), 7.13 (d, J = 8.8 Hz, 1H), 7.09 (d, J = 7.4 Hz, 1H), 3.48 (t, J =
10.4 Hz, 1H), 3.18 (t, J = 14.3 Hz, 1H), 3.08 (t, J = 13.4 Hz, 1H),
2.87 (s, 4H), 2.68 (dd, J = 50.5, 10.4 Hz, 2H), 2.22 (d, J = 10.2 Hz,
Obtained from (2R)-14 (0.18 mmol) to yield 0.015g (24%) as a
light yellow oil, purification 97:3 (DCM:MeOH). The oil was
1
converted to the corresponding HCl salt. H-NMR (500 MHz;
CDCl ): δ 11.90 (s, 1H), 7.46 (m, 1H), 7.32 (m, 2H), 7.26-7.22
3
13
1H), 1.76 (dt, J = 15.3, 7.6 Hz, 5H), 0.98 (t, J = 7.3 Hz, 6H). C-
(m, 1H), 7.17 (m, 2H), 7.03 (t, J = 6.7 Hz, 1H), 3.69-3.63 (m, 1H),
NMR (400 MHz, CDCl ): δ 137.25, 135.73, 131.67, 129.71,
3
3.35-3.28 (m, 2H), 3.11-2.99 (m, 4H), 2.71-2.53 (m, 2H), 2.44-
129.25, 129.18, 127.63, 125.66, 124.19, 115.70, 56.72, 32.85,
27.91, 26.10, 22.35, 12.20. Calcd C22H28FN for [M+H]+:
326.2284. Found: 326.2275 HPLC (s-prep): Solvent System:
HPLC (s-prep): Solvent System: hexanes/EtOH (93:7) 0.1% DEA
(modifier) 0.1% TFA (modifier); flow rate = 1.3 mL/min; t1 = 28.2,
t2 = 31.6. (–)-24: [a]D25 –37.98 (c 0.9, DCM), (+)-24: [a]D25 +41.47
(c 0.9, DCM).
13
2.40 (m, 1H), 2.03-1.94 (m, 5H), 1.02 (t, J = 5.7 Hz, 6H). C-
NMR (400 MHz, CDCl ): δ 139.91, 139.61, 139.52, 139.35,
3
133.62, 133.22, 133.05, 132.97, 132.69, 132.42, 131.15, 130.79,
129.70, 129.57, 129.32, 129.11, 128.34, 128.04, 127.04, 126.99,
126.49, 126.47, 59.61, 59.42, 52.58, 52.34, 52.20, 52.00, 30.55,
30.17, 27.17, 26.40, 24.20, 18.22, 17.93, 11.66, 11.62. Calcd
C22H28ClN for [M+H]+: 342.1989. Found: 342.1990. 21 HCl:
[a]D25 +35.97 (c 0.4, MeOH).
7.1.55. 1-(5-(2-fluorophenyl)-1,2,3,4-
tetrahydronaphthalen-2-yl)pyrrolidine HCl
(18)
7.1.52. (2S)-5-phenyl-N,N-dipropyl-1,2,3,4-
tetrahydronaphthalen-2-amine HCl ((2S)-16)
(Previously reported by Holmberg24) Obtained from (2S)-14
(0.13 mmol) to yield 0.026 g (67%) as a clear oil, purification 98:2
(DCM:MeOH). The oil was converted to the corresponding HCl
Obtained from 7b (0.36 mmol) to yield 0.04 g (38%) as a light
green oil, purification: 98:2 (DCM:MeOH). The oil was converted
1
to the corresponding HCl salt. H-NMR (500 MHz; CDCl3): δ

12.52 (s, 1H), 7.35 (m, 1H), 7.22 (m, 3H), 7.14 (d, J = 7.6 Hz, 2H),
7.09 (d, J = 7.3 Hz, 1H), 3.87-3.82 (m, 2H), 3.49 (m, 1H), 3.37 (m,
1H), 3.32-3.28 (m, 1H), 2.97-2.89 (m, 2H), 2.75-2.65 (m, 2H),
placed into scintillation vials with 1 mL SX18-4 ScintiVerse™ BD
Cocktail (Fisher Chemical, Fair Lawn, NJ). Scintillation was
detected by a Tri-Carb 2910 TR Liquid Scintillation Analyzer
(Perkin Elmer, Boston, MA). All experimental data were analyzed
with GraphPad Prism 6.05 or higher software (La Jolla, CA), and
fit with the one-site Ki nonlinear regression model.
13
2.27 (m, 3H), 2.16 (m, 1H), 2.07-2.05 (m, 2H). C-NMR (400
MHz, CDCl ): δ 131.36, 131.32, 129.43, 129.35, 129.07, 128.58,
3
126.31, 124.31, 115.29, 61.84, 51.27, 32.65, 25.80, 23.42. Calcd
C20H22FN for [M+H]+: 296.1815. Found: 296.1812.
7.4 Computational Methods
7.4.1. 3D QSAR
7.2. Cell culture, transfection, and membrane
homogenization
Computational experiments were performed using the QSAR
module of Sybyl X2.1 (Tripos, LLC). The 40 compounds in the
data set were aligned as described above; the reported Ki values
were converted to corresponding pKi values and used as the
dependent variable. To identify the most statistically significant
model, all combinations of CoMFA and CoMSIA field sets were
explored with the SAMPLS algorithm. To model selectivity, we
used as the target value, pKi(5-HT7)–pKi(5-HT1A); positive values
represent 5-HT7-selective compounds and negative values
represent 5-HT1A-selective compounds.
Human embryonic kidney 293 cells (HEK293, ATCC no. CRL-
1573) stably expressing the h5-HT7(a) receptor (Bmax=7.7 (0.4)
pmol/mg protein) were previously described (CHTR7beta cell
line)17. HEK293 and HEK293T cells (ATCC no. CRL-3216) were
used to express h5-HT1A receptors. All cells were grown in 10 cm
plates in a humidified incubator at 37 ⁰C and 5% carbon dioxide.
The cells were maintained in Corning cellgro Dulbecco’s modified
Eagle’s medium (DMEM, MT—10-013) supplemented with 10%
regular fetal bovine serum, 1% penicillin—streptomycin (the
CHTR7beta cell line was grown with 400 μg/mL G418). cDNA
encoding the wild-type h5-HT1A receptor was obtained from UMR
cDNA Resource Center (Rolla, MO). Transfections were
performed on HEK293 and HEK293T cells at less than pass 20
and ~80% confluency. Cells were washed with 1 mL 1x phosphate
buffered saline and incubated for 48 hours in a bath containing 15
μg h5-HT1A cDNA, 30 μL TurboFect transfection reagent (Thermo
Scientific, #R0532), 3 mL Opti-MEM (Gibco, Ref. 31985-070),
and 3 mL the complete growth media outlined above. After 24
hours, an additional 2 mL of complete growth media was added to
the plates.
7.4.2. Receptor homology modeling
The homology models of human serotonin 5-HT7 and 5-HT1A
receptors were built based on the 5-HT1B receptor crystal structure
template (PDB code: 4IAQ), which shares sequencing identity of
40% and 46% with the 5-HT7 and 5-HT1A receptors, respectively.
Sequence alignment between the crystal structure template and
target serotonin receptors were generated using Clustalw server
(http://www.genome.jp/tools/clustalw/). The homology models
were generated using the Prime program included in the
Schrodinger 2018-1 software package. PyMOL was used to
generate the docking images 45-47
.
7.4.3. Molecular docking
Cell membranes were collected at ~70% confluency and after
48 hours of transfection for 5-HT7(a) and 5-HT1A expressing cells,
respectively. Cells were dissociated by pipetting in 25 mL of wash
buffer (50 mM tris-HCl, 4 ֯C, pH=7.4) and centrifuged at 12,000
g and 4 ֯C, for 10 minutes. Next, the supernatant was discarded,
and the membrane pellet homogenized in wash buffer using 50
pumps of a handheld tissue grinder. The membrane suspension
was then centrifuged in 25 mL of wash buffer under the same
conditions; this was repeated once more so that each preparation
underwent 3 rounds of centrifugation and was washed with at least
50 mL of wash buffer. Following the last centrifugation, the
supernatant was discarded, and the membrane pellet was frozen at
-80 ֯C.
The 3D structures of compounds in Tables 1 and 2 were built
using the Maestro graphic software from Schrodinger, LLC. The
compound structures were optimized using ab initio quantum
chemistry at the HF/6-31G* level (GAUSSIAN 16 program
(Gaussian, Inc.)), followed by single point calculations to obtain
the molecular electrostatic potential for charge fitting48.
AUTODOCK49 program was used for the flexible receptor and
ligand docking simulations. The final docking poses were selected
based on binding energies and cluster analysis.
7.4.4. Molecular dynamics simulations
The protonation states of the titratable residues of the 5-HT7
and 5-HT1A model structures were determined by the H++ server
(http://biophysics.cs.vt.edu/). The complexes of selected
compounds in Tables 1 and 2 and the receptor structure models
were immersed in an explicit lipid bilayer of POPC, POPE, and
cholesterol, with molecular ratio of 2:2:150, and, a water box, using
7.3. Radioligand binding assays
Radioligand competition assays were performed following
firmly established methods44. Briefly, membrane pellets described
above were vortexed in assay buffer (50 mM tris-HCl, 10 mM
MgCl2, 0.1 mM EDTA, pH=7.4 at 22 ºC) until homogenous.
Serotonin (10 μM) was used to define nonspecific binding of the
CHARMM-GUI
Membrane
Builder
webserver
(http://www.charmm-gui.org/?doc=input/membrane). NaCl (150
mM) and neutralizing counter ions were added to the system. The
PMEMD.CUDA program in AMBER version 16 was used to
conduct the MD simulations. The latest Amber ff14SB, lipid17,
and TIP3P force field in the Amber16 program was used in MD
simulations for T2Rs with lipid and water. The Antechamber
module of AmberTools was used to generate the parameters PAT
compounds using general AMBER force field (GAFF). The partial
charges for compounds were calculated by using ab initio quantum
chemistry at the HF/6-31G* level (GAUSSIAN 16 program). The
CHELPG charge-fitting scheme was used to calculate partial
charges on the atoms [M1]. The topology and coordinate files of
the system were prepared by using the tleap module of Amber16.
The MD simulations were performed with periodic boundary
5-HT1A/5-HT7(a)
receptor
agonist
radiolabel
[3H]-5-
carboxamidotryptamine (37.7 Ci/mmol; Perkin Elmer Life
Sciences, Boston, MA) at 0.6-1 nM. Experiments were performed
in a total well volume of 250 μL, with 3–10 μg of protein per well,
over 10 concentrations of unlabeled ligand (0.03 nM-100 μM in
half log units; depending on the affinity and solubility of the
unlabeled ligand). Each compound was assessed at least two times
in triplicate or greater. Following a 90-minute incubation with the
assay plate covered, at room temperature, shaking at 300 rpm, the
assay was terminated by rapid vacuum filtration via Whatman
GF/B Fired Filters (Brandel Inc., Gaithersburg, MD), presoaked
with cold wash buffer, using an automated Tomtec Harvester 96
(Hamden, CT). Filters were washed again before being dried and

4.
5.
Gellynck E, Heyninck K, Andressen KW, et al. The serotonin 5-HT7
receptors: Two decades of research. Exp Brain Res. 2013;230(4):555-
568. doi:10.1007/s00221-013-3694-y
Martín-Cora FJ, Pazos A. Autoradiographic distribution of 5-HT7
receptors in the human brain using [3H]mesulergine: comparison to
other mammalian species. Br J Pharmacol. 2004;141(1):92-104.
doi:10.1038/sj.bjp.0705576
Varnäs K, Thomas DR, Tupala E, Tiihonen J, Hall H. Distribution of 5-
HT7 receptors in the human brain: A preliminary autoradiographic
study using [3H]SB-269970. Neurosci Lett. 2004;367(3):313-316.
doi:10.1016/j.neulet.2004.06.025
Hauser SR, Hedlund PB, Roberts AJ, Sari Y, Bell RL, Engleman EA.
The 5-HT7 receptor as a potential target for treating drug and alcohol
abuse. Front Neurosci. 2014;8(January):448.
conditions to produce isothermal-isobaric ensembles (NPT,
constant-temperature, constant-pressure ensemble). The pressure
was regulated using the isotropic position scaling algorithm with
the pressure relaxation time set to 2.0 ps. Long range electrostatics
was calculated using the particle mesh Ewald (PME) method51
with a 10 Å cutoff. Prior to production runs, energy minimization
of 2000 steps of the steepest descent followed by 3000 steps of
conjugate gradient were carried out on the system. Subsequently,
the systems were heated from 0 K to 303 K using Langevin
dynamics with the collision frequency of 1 ps-1. During the
heating, the receptor complexes were position-restrained using an
initial constant force of 500 kcal/mol/Å2, and weakened to 10
kcal/mol/Å2, allowing lipid and water molecules to move freely.
Then, the systems went through 5 ns equilibrium MD simulations.
Finally, a total of 100 ns production MD simulations were
conducted, and coordinates were saved every 100 ps for analysis.
6.
7.
8.
9.
doi:10.3389/fnins.2014.00448
Kvachnina E, Liu G, Dityatev A, et al. 5-HT7 receptor is coupled to G
alpha subunits of heterotrimeric G12-protein to regulate gene
transcription and neuronal morphology. J Neurosci. 2005;25(34):7821-
7830. doi:10.1523/JNEUROSCI.1790-05.2005
Adriani W, Travaglini D, Lacivita E, Saso L, Leopoldo M, Laviola G.
Modulatory effects of two novel agonists for serotonin receptor 7 on
emotion, motivation and circadian rhythm profiles in mice.
Neuropharmacology. 2012;62(2):833-842.
7.4.5. Molecular Mechanics-Generalized Born Surface Area
(MMGBSA) binding free energy calculations
The MM-GBSA and normal mode modules, which are
implemented in Amber16, were performed to calculate the binding
free energy of selected compounds in Tables 1 and 2 with the
receptors. The binding free energy can be decomposed into
contributions from individual interacting residues of the receptors,
which consist of four energy terms: non-bonded electrostatic
interaction, van der Waals energy in gas phase, polar, and non-
polar solvation free energy. The polar component is calculated
using Generalized Born (GB) implicit solvation model. The non-
polar component is calculated using solvent accessible surface
area model. The conformational entropy change upon compound
binding for binding free energy calculations is obtained from the
sum of the translational, rotational, and vibrational components,
with the Nmode module. 100 snapshots were extracted from the
last 10 ns along the MD simulation trajectory at an interval of 100
ps for the binding free energy calculations.
doi:10.1016/j.neuropharm.2011.09.012
10. Naumenko VS, Kondaurova EM, Popova NK. On the role of brain 5-
HT7 receptor in the mechanism of hypothermia: Comparison with
hypothermia mediated via 5-HT1A and 5-HT3 receptor.
Neuropharmacology. 2011;61(8):1360-1365.
doi:10.1016/j.neuropharm.2011.08.022
11. Leopoldo M, Lacivita E, Berardi F, Perrone R, Hedlund PB. Serotonin
5-HT7 receptor agents: Structure-activity relationships and potential
therapeutic applications in central nervous system disorders. Pharmacol
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Funding
15. Modica MN, Lacivita E, Intagliata S, et al. Structure-Activity
Relationships and Therapeutic Potentials of 5-HT7 Receptor Ligands:
An Update. Journal of Medicinal Chemistry. 2018;61(19):8475-8503.
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(6)-phenyl-1,2,3,4-tetrahydronaphthalenes. Bioorganic Med Chem.
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This work was funded by Department of Defense
Congressionally Directed Medical Research Programs Peer
Reviewed Medical Research Program awards PR160095 and
PR141869 (R.G.B./C.E.C.) and National Institutes of Health
awards RO1-DA030989, DA047130, and MH081193 (R.G.B.).
Acknowledgments
The authors thank Drs. Spiro Pavlopolous and George O’Doherty
for use of nanodrop and polarimeter instruments, and, Dr.
Munmun Mukherjee for sharing her expertise in synthetic organic
chemistry. Some of the affinity studies reported here formed part
of the Ph.D. thesis research submitted by Daniel Felsing to the
University Florida.
doi:10.1021/acschemneuro.5b00099
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