Structure-activity relationships of 7-substituted dimethyltyrosine-tetrahydroisoquinoline opioid peptidomimetics

Article abstract of DOI:10.3390/molecules24234302

The opioid receptors modulate a variety of biological functions, including pain, mood, and reward. As a result, opioid ligands are being explored as potential therapeutics for a variety of indications. Multifunctional opioid ligands, which act simultaneously at more than one type of opioid receptor, show promise for use in the treatment of addiction, pain, and other conditions. Previously, we reported the creation of bifunctional kappa opioid receptor (KOR) agonist/mu opioid receptor (MOR) partial agonist ligands from the classically delta opioid receptor (DOR) antagonist selective dimethyltyrosine-tetrahydroisoquinoline (Dmt-Tiq) scaffold through the addition of a 7-benzyl pendant on the tetrahydroisoquinoline ring. This study further explores the structure-activity relationships surrounding 7-position pendants on the Dmt-Tiq scaffold. Some analogues maintain a KOR agonist/MOR partial agonist profile, which is being explored in the development of a treatment for cocaine addiction. Others display a MOR agonist/DOR antagonist profile, which has potential to be used in the creation of a less addictive pain medication. Ultimately, we report the synthesis and in vitro evaluation of novel opioid ligands with a variety of multifunctional profiles.

Full text of DOI:10.3390/molecules24234302

molecules
Article
Structure–Activity Relationships of 7-Substituted
Dimethyltyrosine-Tetrahydroisoquinoline
Opioid Peptidomimetics
Deanna Montgomery 1 , Jessica P. Anand , Mason A. Baber , Jack J. Twarozynski ,
2,3
1
2
2
2
2,3
1,3,
Joshua G. Hartman , Lennon J. Delong , John R. Traynor
and Henry I. Mosberg
*
1
Department of Medicinal Chemistry, College of Pharmacy, University of Michigan, Ann Arbor, MI 48109,
USA; dmontg@umich.edu (D.M.); mbaber@rollins.edu (M.A.B.)
Department of Pharmacology, Medical School, University of Michigan, Ann Arbor, MI 48109, USA;
janand@umich.edu (J.P.A.); jactwaro@umich.edu (J.J.T.); hartjosh@umich.edu (J.G.H.);
delongl@umich.edu (L.J.D.); jtraynor@umich.edu (J.R.T.)
2
3
Edward F. Domino Research Center, Medical School, University of Michigan, Ann Arbor, MI 48109, USA
Correspondence: him@umich.edu; Tel.: +1-734-764-8117
*
Academic Editor: Derek J. McPhee
Received: 2 November 2019; Accepted: 21 November 2019; Published: 26 November 2019
ꢀ
ꢁꢂꢀꢃꢄꢅꢆꢇ
ꢀ
ꢁꢂꢃꢄꢅꢆ
Abstract: The opioid receptors modulate a variety of biological functions, including pain, mood,
and reward. As a result, opioid ligands are being explored as potential therapeutics for a variety of
indications. Multifunctional opioid ligands, which act simultaneously at more than one type of opioid
receptor, show promise for use in the treatment of addiction, pain, and other conditions. Previously,
we reported the creation of bifunctional kappa opioid receptor (KOR) agonist/mu opioid receptor
(
MOR) partial agonist ligands from the classically delta opioid receptor (DOR) antagonist selective
dimethyltyrosine-tetrahydroisoquinoline (Dmt-Tiq) scaffold through the addition of a 7-benzyl
pendant on the tetrahydroisoquinoline ring. This study further explores the structure–activity
relationships surrounding 7-position pendants on the Dmt-Tiq scaffold. Some analogues maintain a
KOR agonist/MOR partial agonist profile, which is being explored in the development of a treatment
for cocaine addiction. Others display a MOR agonist/DOR antagonist profile, which has potential to
be used in the creation of a less addictive pain medication. Ultimately, we report the synthesis and
in vitro evaluation of novel opioid ligands with a variety of multifunctional profiles.
Keywords: peptidomimetic; structure-activity; opioids; multifunctional ligands
1
. Introduction
Opioids have one of the longest known histories of any drug class. The use of opium for ritual,
medicinal, and/or recreational purposes dates back to ancient civilizations [
1
,2]. In the early 1800s,
Friedrich Sertürner isolated the primary active ingredient of opium and named it morphine [
1].
This began the chemical exploration of the opiates, and ultimately, led to discovery of the opioid
receptors and their endogenous ligands. It is widely accepted that there are three major types of
opioid receptors—the kappa opioid receptor (KOR), the mu opioid receptor (MOR), and the delta
opioid receptor (DOR). These receptors have high sequence and structural homology, and they are
all Class A GPCRs [
reviewed [ ]. Though most well-known for its role in regulating pain, the opioid system is also
involved in many other biological processes, including mood [ ] and reward [ 10]. As such, opioids
3,4]. The structure and function of this type of receptor have been thoroughly
5,6
7–
9
8,
remain an important and promising class of molecules for the development of therapeutics for a variety
of indications.
Molecules 2019, 24, 4302; doi:10.3390/molecules24234302
www.mdpi.com/journal/molecules

Molecules 2019, 24, 4302
2 of 24
Functions of the opioid receptors are modulated by both endogenous and exogenous opioid
ligands. In the two centuries since the discovery of morphine, many semi-synthetic and synthetic
opioids have been developed for this purpose. As the complex pharmacology of the opioid system
continues to be revealed, it has been posited that unwanted effects and desired effects may result from
the same interaction of an opioid agonist or antagonist with its target. As a result, the development
of selective agents has declined, and the development of multifunctional ligands, compounds that
act simultaneously at multiple opioid receptor types, has gained popularity as a strategy for the
design of therapeutics [11
reviewed [13].
,12]. The current state of multifunctional opioid ligands has recently been
Our group [14] and others [15
–19] have shown that the dimethyltyrosine-tetrahydroisoquinoline
(
Dmt-Tiq) scaffold can be used in the development of multifunctional opioid ligands (Figure 1). This
scaffold, originally developed as a selective DOR antagonist, has been extensively explored through
synthesis of many analogues. However, the confines of traditional peptide synthesis have limited
substitution on the tetrahydroisoquinoline (Tiq) ring. Nearly two decades ago, minor substitutions
were reported at the 6-, 7-, and 8- positions, but all of these compounds displayed a DOR antagonist
profile similar to that of the parent peptide [17,20]. Recently, we reported that installation of a 7-benzyl
pendant on the Tiq could alter the profile of this series to KOR agonism/MOR partial agonism [14].
Figure 1. A variety of pendants were introduced at the 7-position of the tetrahydroisoquinoline of the
dimethyltyrosine-tetrahydroisoquinoline (Dmt-Tiq) scaffold.
KOR agonists have shown potential for use in the treatment of cocaine addiction because of
their reward-modulating properties. Specifically, administration of a KOR agonist can reduce cocaine
self-administration in non-human primates [21
dysphoria, an intense feeling of unease or dissatisfaction [
,
22]. However, KOR agonism is also associated with
]. As a result, selective KOR agonists
7
have limited therapeutic potential. It is well known that MOR agonism is associated with euphoria.
Therefore, a bifunctional KOR agonist/MOR agonist offers a potential alternative to a selective KOR
agonist that may result in a more favorable side effect profile. In fact, there is evidence to suggest that a
KOR agonist/MOR agonist may be useful in the treatment of cocaine addiction [23,24].
The aim of this work was to explore structure–activity relationships around the 7-benzyl pendant
which introduced KOR agonism to the Dmt-Tiq scaffold. Novel analogues reported here reveal
that substitution on the benzyl ring can maintain a KOR agonist/MOR partial agonist profile while
analogues with other 7-position pendants show varied results. Overall, this work demonstrates the
development of novel Dmt-Tiq peptidomimetics that display a range of multifunctional opioid profiles.
2
. Results
A series of novel Dmt-Tiq compounds with substitution at the Tiq 7-position were prepared and
evaluated in vitro for opioid activity.
2
.1. Synthesis
All compounds were prepared from commercial starting materials according to one of the synthetic
routes shown in Scheme 1. In the first route, commercially available carboxylic acid was reduced to the
corresponding secondary alcohol using borane dimethylsulfide. An Appel reaction was performed
to convert alcohol to benzyl bromide . The pendant was then attached via Suzuki coupling of
intermediate with the corresponding boronic acid or S 2 reaction with the corresponding nucleophile.
1
2
2
3
3
N

Molecules 2019, 24, 4302
3 of 24
In the second route, Boc-protected tetrahydroisoquinoline
5
was prepared from commercially available
7
-bromotetrahydroisoquinoline (
appropriate pendant was attached by Suzuki coupling with the corresponding benzyl bromide. In each
case, after the pendant was attached, the Boc group was removed from intermediate 7a with
4). This intermediate was converted to boronic ester 6, and the
–
z
acid, and the deprotected tetrahydroisoquinoline intermediate was coupled with diBoc-protected
dimethyltyrosine. Finally, the Boc groups were removed to yield the final peptidomimetic 8a–z.
Scheme 1. Preparation of Dmt-Tiq peptidomimetics. Reagents and conditions: (i) BH SMe , THF;
3
2
(
ii) CBr4, PPh , DCM; (iii) substituted aryl boronic acid, Pd(dppf)Cl , K CO , 3:1 acetone:water; (iv)
3
2
2
3
amine, K CO , DMF; (v) Boc O, microwave; (vi) bis(pinacolato)diboron, Pd(dppf)Cl , CH CO K,
2
3
2
2
3
2
DMSO; (vii) substituted benzyl bromide, Pd(dppf)Cl , K CO , 3:1 acetone/water; (viii) HCl, 1,4-dioxane
2
2
3
or TFA, DCM; (ix) diBoc-Dmt, PyBOP, 6Cl-HOBt, DIEA, DMF; (x) TFA, DCM.
2
.2. Pharmacological Evaluation
Each novel compound was evaluated for binding to and stimulation of KOR, MOR, and
3
DOR. Binding affinity (K ) was determined by competitive displacement of [ H]-diprenorphine,
i
a non-selective opioid receptor antagonist with similar affinity for each of the three receptors. Potency
(
EC ) and efficacy, expressed as percent stimulation compared to a standard agonist at each receptor,
50
35
were determined by a [ S]-GTPγS binding assay.
Building on our previous work [14], several ortho and meta substituents on the benzyl ring were
investigated as well as the o-,m-dimethyl analogue. The results of the pharmacological evaluation
of these compounds are shown in Table 1. Data for the previously reported 7-benzyl analogue 4c is
shown for comparison. Previously, this compound was evaluated at human KOR, rat MOR, and rat
DOR. The profile shown here differs slightly from that previously reported because all compounds
in this study were evaluated only at human receptors. All ortho- and meta-substituted analogues
reported here display single digit nanomolar or subnanomolar binding at all three opioid receptors.
In general, ortho analogues show the highest affinity for KOR compared to the other receptors, while
most meta analogues show the highest affinity for DOR. Compared to standard agonists, each of these
analogues retains moderate (54%) to high (89%) efficacy at KOR and low (29%) to high efficacy (81%)
at MOR. Most analogues show no DOR agonism, but the ortho trifluoromethyl analogue 8c shows low
DOR efficacy and potency. Potency for these compounds remains primarily in the double or triple
digit nanomolar range. The balance of potencies varies for ortho analogues, while meta analogues and
the disubstituted analogue are consistently more potent at MOR than KOR.

Molecules 2019, 24, 4302
4 of 24
Table 1. Substituted 7-benzyl pendants on the Dmt-Tiq scaffold. 1
Ki (nM)
EC50 (nM)
% Stimulation
Compound
R
KOR MOR DOR KOR MOR DOR KOR MOR DOR
2
0.3)
.3
5.3
(0.5)
2.2
(0.7)
97
(24)
68
5.7
82
(6)
39
(4)
20
(3)
4
c
(
(
(15) (2.0)
3
0.9)
.1
3.7
(0.6)
2.7
(0.7)
130
(41)
92
72
37
8
a
dns *
(26)
dns *
dns *
(11) (8)
0
.32
1.2
(0.1)
2.5
(0.3)
11
(1)
43
89
(6)
60
(5)
8
b
dns *
(13)
(
0.01)
2
0.6)
.6
4.3
(1.3)
4.9
(2)
173
(55)
53
(11)
342
(32)
81
76
24
(3)
8
c
(
(11) (6)
0
0.1)
.5
6.8
(0.2)
3.6
(0.8)
3.7
(0.8)
664
(515)
80
(9)
31
(9)
8
d
dns
dns
(
3
0.9)
.0
2.7
(0.6)
1.7
(0.4)
148
(38)
24
(3)
74
(4)
48
(6)
8
e
dns
dns *
dns
dns
dns *
dns
(
(
(
(
0
0.2)
.8
0.6
(0.2)
1.4
(0.1)
148
(53)
18
(3)
83
(5)
81
(9)
8
f
5
1.1)
.4
4.1
(1.1)
1.6
(0.5)
319
(115) (96)
205
68
(8)
45
(3)
8
g
5
0.9)
.9
3.4
(1.3)
1.4
(0.2)
287
(61)
53
(6)
60
(8)
29
(3)
8
h
dns
dns
5
1.2)
.8
5.2
(1.4)
3.6
(0.6)
1030
(50)
380
(187)
54
(9)
34
(3)
8
i
dns *
dns *
(
1
3
Binding affinity (K
i
) values determined by competitive displacement of [ H]-diprenorphine in membrane
preparations from CHO cells expressing human KOR, MOR, or DOR. Potency (EC50) and efficacy values determined
3
5
by [ S]-GTPγS binding in the same membrane preparations. Efficacy expressed as percent stimulation versus
standard agonists—U69,593 (KOR), DAMGO (MOR), or DPDPE (DOR). All values expressed as mean (SEM) of
three or more separate assays run in duplicate unless otherwise noted. * n = 2; dns = does not stimulate, average
maximal stimulation <10% at concentrations up to 10 µM.
Next, we explored the incorporation of nitrogen into the aromatic ring of the pendant. In place of the
benzyl pendant, 3- and 4-pyridyl pendants were added at the 7-position of the tetrahydroisoquinoline
ring with a methylene spacer (Table 2). Due to well-known synthetic difficulties [25], the 2-pyridyl
analogue was not successfully synthesized. Evaluation of the pyridyl analogues revealed a loss in
binding and a drastic loss of potency at KOR with low efficacy at MOR and no agonism at DOR,
compared to their carbocyclic counterparts.
Non-aromatic pendants were also explored. Table 3 shows pharmacological data for analogues
with saturated, cyclic amine pendants. For this series, all KOR agonism was lost, and only weak
potency and low efficacy at MOR were observed.

Molecules 2019, 24, 4302
5 of 24
Table 2. Pyridyl 7-position pendants on the Dmt-Tiq scaffold. 1
Ki (nM)
EC50 (nM)
% Stimulation
Compound
R
KOR MOR DOR KOR MOR DOR KOR MOR DOR
5
0.9)
6
5.4
(0.3)
1.5
(0.2)
988
(277)
145
(30)
40
(7)
23
(1)
8
j
dns
dns
dns
(
4
12)
4
42
(18)
2.6
(1.0)
1180
(287)
179
(17)
44
(9)
28
(1)
8
k
dns
(
1
3
Binding affinity (K
i
) values determined by competitive displacement of [ H]-diprenorphine in membrane
preparations from CHO cells expressing human KOR, MOR, or DOR. Potency (EC50) and efficacy values determined
3
5
by [ S]-GTPγS binding in the same membrane preparations. Efficacy expressed as percent stimulation versus
standard agonists—U69,593 (KOR), DAMGO (MOR), or DPDPE (DOR). All values expressed as mean (SEM) of
three or more separate assays run in duplicate unless otherwise noted. dns = does not stimulate, average maximal
stimulation <10% at concentrations up to 10 µM.
Table 3. Aliphatic 7-position pendants on the Dmt-Tiq scaffold. 1
Ki (nM)
EC50 (nM)
% Stimulation
Compound
R
KOR MOR DOR KOR MOR DOR KOR MOR DOR
2
11)
9
4.7
(0.9)
87
(15)
845
(97)
34
(5)
8
l
dns
dns
dns
dns
dns
dns
dns
dns
dns *
dns
(
6
9
17
(11)
175
(9)
639 *
(162)
40 *
(22)
8
m
dns *
(3)
4
17)
3
39
(27)
5.6
(1.6)
860
(200)
38
(14)
8
n
dns
(
1
3
Binding affinity (K
i
) values determined by competitive displacement of [ H]-diprenorphine in membrane
preparations from CHO cells expressing human KOR, MOR, or DOR. Potency (EC50) and efficacy values determined
3
5
by [ S]-GTPγS binding in the same membrane preparations. Efficacy expressed as percent stimulation versus
standard agonists—U69,593 (KOR), DAMGO (MOR), or DPDPE (DOR). All values expressed as mean (SEM) of
three or more separate assays run in duplicate unless otherwise noted. * n = 2; dns = does not stimulate, average
maximal stimulation <10% at concentrations up to 10 µM.
To test whether opioid activity could be maintained in the presence of larger pendants, 1- and
-naphthyl analogues were synthesized. These results are shown in Table 4. The 1-naphthyl pendant
2
displays a KOR agonist/MOR partial agonist profile, while the 2-naphthyl pendant results in a drastic
loss in KOR binding and a complete loss of KOR agonism.
Given the MOR/KOR profile of analogue 8o, a nitrogen scan was conducted to further explore
the structure–activity relationships around this pendant (Table 5). Similar to the pyridyl analogues,
synthetic difficulties prevented the synthesis and evaluation of the 1-isoquinolinyl analogue. With few
exceptions, single digit nanomolar or stronger binding is observed at all three receptors, and these
analogues favor binding to DOR over MOR and KOR. All of these compounds show MOR agonism,
and all except analogue 8u display partial to full KOR agonism. However, only compounds 8q and 8v
shows DOR agonism. Ultimately, the addition of a single nitrogen to this ring results in compounds
that show a range of multifunctional opioid profiles.
Finally, bicyclic pendants with one saturated ring and one aromatic ring were explored. This
subset of analogues displays two distinct profiles (Table 6). Compounds 8x and 8y show balanced
affinity and efficacy at KOR and MOR, while compounds 8w and 8z display a loss in KOR binding
and no KOR agonism. However, the latter two compounds show potent, moderate to high efficacy at
MOR and strong binding but no agonism at DOR.

Molecules 2019, 24, 4302
6 of 24
Table 4. Naphthyl 7-position pendants on the Dmt-Tiq scaffold. 1
Ki (nM)
EC50 (nM)
% Stimulation
Compound
R
KOR MOR DOR KOR MOR DOR KOR MOR DOR
4
0.5)
.7
5.2
(1.0)
3.2
(0.6)
349
(112)
132
(76)
83
(14)
59
(1)
8
o
dns
dns
dns
dns
(
1
42
5.8
(1.6)
3.5
(0.7)
176
(59)
51
(9)
8
p
dns
dns
(23)
1
3
Binding affinity (K
i
) values determined by competitive displacement of [ H]-diprenorphine in membrane
preparations from CHO cells expressing human KOR, MOR, or DOR. Potency (EC50) and efficacy values determined
3
5
by [ S]-GTPγS binding in the same membrane preparations. Efficacy expressed as percent stimulation versus
standard agonists—U69,593 (KOR), DAMGO (MOR), or DPDPE (DOR). All values expressed as mean (SEM) of
three or more separate assays run in duplicate unless otherwise noted. dns = does not stimulate, average maximal
stimulation <10% at concentrations up to 10 µM.
Table 5. Nitrogen scan of 7-position 1-naphthyl pendant on the Dmt-Tiq scaffold. 1
Ki (nM)
EC50 (nM)
% Stimulation
Compound
R
KOR MOR DOR KOR MOR DOR KOR MOR DOR
1
9
1.4
(0.3)
0.46
(0.08)
258
(16)
24
(4)
1.2
(0.8)
25
(3)
63
(8)
23
(4)
8
q
(2)
2
0.7)
.7
1.8
(0.3)
0.69
(0.24)
65
(19)
24
(6)
42
(15)
36
(5)
8
r
dns *
dns *
(
1
0
9.7
(1.4)
1.2
(0.2)
609
(226)
43
(21)
24
(2)
28
(8)
8
s
dns
dns
(1)
1
0.5)
.5
4.6
(0.5)
1.2
(0.4)
58
(11)
54
(15)
69
(1)
31
(6)
8
t
dns *
dns *
dns *
dns *
(
(
8
0.4)
.6
6.6
(1.0)
0.79
(0.25)
54
(19)
45
(12)
8
u
dns
dns
0
.73
1.4
(0.2)
0.30
(0.04)
14
(1)
12
(3)
19
(10)
71
(16)
81
(7)
39
(7)
8
v
(
0.03)
1
3
Binding affinity (K
i
) values determined by competitive displacement of [ H]-diprenorphine in membrane
preparations from CHO cells expressing human KOR, MOR, or DOR. Potency (EC50) and efficacy values determined
3
5
by [ S]-GTPγS binding in the same membrane preparations. Efficacy expressed as percent stimulation versus
standard agonists—U69,593 (KOR), DAMGO (MOR), or DPDPE (DOR). All values expressed as mean (SEM) of
three or more separate assays run in duplicate unless otherwise noted. * n = 2; dns = does not stimulate, average
maximal stimulation <10% at concentrations up to 10 µM.

Molecules 2019, 24, 4302
7 of 24
Table 6. Bicyclic aliphatic 7-position pendants on the Dmt-Tiq scaffold. 1
Ki (nM)
EC50 (nM)
% Stimulation
Compound
R
KOR MOR DOR KOR MOR DOR KOR MOR DOR
4
2
0.6
(0.1)
2.3
(0.6)
8.2
(1.0)
85
(9)
8
w
dns
dns *
dns *
dns
dns *
dns *
(8)
7
.1
7.2
(1.6)
6.6
(1.0)
375
(109)
73
(22)
67
(3)
65
(10)
8
x
(
(
1.4)
2
.6
2.2
(0.6)
1.7
(0.5)
106
(20)
42
(10)
66
(8)
65
(5)
8
y
dns *
dns *
dns *
dns *
0.6)
6
23)
5
1.5
(0.2)
1.3
(0.4)
27
(0.3)
63
(10)
8
z
dns
dns
(
1
3
Binding affinity (K
i
) values determined by competitive displacement of [ H]-diprenorphine in membrane
preparations from CHO cells expressing human KOR, MOR, or DOR. Potency (EC50) and efficacy values determined
3
5
by [ S]-GTPγS binding in the same membrane preparations. Efficacy expressed as percent stimulation versus
standard agonists—U69,593 (KOR), DAMGO (MOR), or DPDPE (DOR). All values expressed as mean (SEM) of
three or more separate assays run in duplicate unless otherwise noted. * n = 2; dns = does not stimulate, average
maximal stimulation <10% at concentrations up to 10 µM.
3
. Discussion
Previous work by our group [14] and others [15
–
19] has indicated that the classically DOR
antagonist selective dimethyltyrosine-tetrahydroisoquinoline (Dmt-Tiq) scaffold can be used as a
starting point for the development of multifunctional opioid ligands. Building on our previous work,
this study further explores installation of a 7-position pendant on the tetrahydroisoquinoline ring as
a means of developing ligands with pharmacologically useful, multifunctional profiles. Previously,
we reported that introduction of ortho and meta substituents onto a 7-benzyl pendant could produce
ligands that demonstrate KOR agonism and MOR partial agonism [14], a bifunctional profile which
has shown promise for the treatment of addiction to cocaine and other drugs of abuse. Here, we further
explore the structure–activity relationships surrounding this novel series of opioid ligands and report
compounds with this and other multifunctional opioid profiles.
Based on previously reported initial results from this series [14], we believed ortho and meta
substituents on a 7-benzyl pendant to be promising structural modifications for the development of
KOR agonist/MOR partial agonist ligands. A series of additional ortho and meta substitutions were
evaluated to confirm whether they would exhibit the anticipated profile (Table 1). As expected, ortho
and meta substitutions on the 7-benzyl pendant are favorable for the development of KOR/MOR
ligands. Because di-substitution (compound 8i) results in a notable drop in KOR potency, it shows
no advantage over a single ortho or meta substituent. A few of the ligands in this series, including
analogue 8c, show DOR agonism, which represents a problem for the development of a therapeutically
useful KOR/MOR ligand because DOR agonism is associated with problematic side effects, including
convulsions [
treatment for addiction since it has been shown to lower the addiction potential of MOR agonists in
preclinical models [28 30]. The strong MOR agonism of some compounds in this series (compounds
and 8f) is also a concern for the development of a therapy, as this activity would likely impart
9,26,27]. On the other hand, DOR antagonism may be beneficial for the development of a
–
8
c
greater abuse potential. The most promising compound in this series for the development of a KOR
agonist/MOR partial agonist for treatment of cocaine addiction, analogue 8b, shows high potency and
efficacy at KOR, high potency and low efficacy at MOR, and is devoid of DOR agonism. This compound

Molecules 2019, 24, 4302
8 of 24
also has higher affinity for KOR and MOR than for DOR (eight-fold and two-fold, respectively), making
it a promising candidate for further evaluation.
The introduction of a nitrogen to the 7-benzyl ring was not favorable for the development of
a KOR/MOR ligand (Table 2). Rather, these analogues are selective for DOR over KOR and MOR
and display low potency and efficacy at KOR and MOR. Replacement of the benzyl pendant with
a saturated, cyclic amine pendant likewise decreases MOR potency drastically and eliminates KOR
agonism altogether (Table 3). Unlike many of the compounds reported here, these analogues do not
show particularly useful opioid profiles.
To explore the potential of installing larger pendants at the Tiq 7-position, we first synthesized
analogues with 1- and 2-naphthyl pendants (Table 4). Though the high clogP (5.6) and associated
insolubility of these compounds is a problem for the ultimate development of a therapeutic, they
were prepared as useful probes to further explore what might be tolerated in this series. Based on
our previous observations from ortho, meta, and para substitutions [14], we hypothesized that the
1-naphthyl pendant would be favorable for the development of a KOR agonist while the 2-naphthyl
pendant would not. The 1-naphthyl pendant points in the same direction as ortho and meta substituents,
where there is room for additional steric bulk to be accommodated in the active configuration of the
KOR orthosteric site. The 2-naphthyl pendant, on the other hand, points in the direction of meta and
para substituents, where it clashes with the receptor. As expected, the 1-naphthyl analogue 8o shows
the desired KOR/MOR profile, while the 2-naphthyl analogue 8p shows a drastic decrease in KOR
binding and a complete loss of KOR agonism.
Based on these findings, we conducted a nitrogen scan of the 1-naphthyl pendant. The introduction
of a single nitrogen drops the clogP by approximately 1.5 units, making these analogues much more
promising candidates for use in animal studies and clinical settings. The profile of these analogues
differed based on the placement of the nitrogen. Most of these compounds (analogues 8q–8v) show
some degree of KOR agonism and MOR agonism, but only analogue 8t displays the desired KOR
agonist/MOR partial agonist profile. Notably, this compound is equipotent at KOR and MOR and
is a promising candidate for further study. It is approximately 3-fold selective for KOR and DOR
over MOR which may lower the abuse potential of such a compound. Analogue 8u displays partial
agonism only at MOR but high affinity for DOR, a profile most similar to that of previously reported
Dmt-Tiq compounds. On the other hand, compound 8v has a potent KOR agonist/MOR agonist/DOR
partial agonist profile and is weakly selective for KOR and DOR over MOR. While interesting, this
profile is likely clinically irrelevant.
Finally, we explored bicyclic pendants with a cyclic amine attached to an aromatic ring. The profile
differs, likely due to the placement of the second ring within the receptor binding site. As expected,
those that would most closely mimic the 1-naphthyl pendant, compounds 8w and 8z, exhibit a
KOR agonist/MOR agonist profile. The binding and efficacy profile at KOR and MOR for these two
compounds is remarkably balanced, though they are more potent at MOR (five-fold and two-fold,
respectively). As discussed above, the higher MOR efficacy and potency of these compounds compared
to others would likely impart greater addiction potential. On the other hand, those compounds which
more closely mimic the 2-naphthyl pendant, analogues 8w and 8z, show no KOR agonism, as expected.
However, these analogues exhibit MOR agonism and DOR antagonism, a bifunctional profile being
explored in the development of a less addictive treatment for pain [13,31,32]. Both compounds display
potent MOR agonism and selectivity for MOR and DOR over KOR (18-fold and 43-fold, respectively).
In addition, compound 8z shows balanced affinity at MOR and DOR, a quality previously explored by
our group as a way to mitigate addiction potential [33]. These compounds represent a starting point
for further study for the development of a MOR agonist/DOR antagonist.
In conclusion, this work reports novel opioid ligands with a variety of multifunctional profiles.
We have further elucidated structure–activity relationships surrounding the 7-position pendant on
the tetrahydroisoquinoline ring of the classically DOR antagonist selective Dmt-Tiq scaffold. Ortho
substituted analogues and select bicyclic pendant analogues show promise for the development of a

Molecules 2019, 24, 4302
9 of 24
KOR agonist/MOR partial agonist, a profile being investigated for the treatment of cocaine addiction.
Compounds 8w and 8z exhibit a balanced MOR agonist/DOR antagonist profile and have potential to
be investigated as a treatment for pain with lowered addiction potential. Future work will examine
the pharmacokinetic properties of these compounds and explore the in vivo activity of interesting
compounds from this series.
4
. Materials and Methods
4
.1. Chemistry
The chemical methods used were the same as those previously described [14] with any changes
noted below. Unless otherwise noted, all reagents and solvents were purchased from commercial
sources and used without additional purification. DiBoc-DMT was prepared from commercially
available DMT according to standard procedures as previously reported [14]. Microwave reactions
were performed in a Discover SP microwave synthesizer (CEM Corp., Matthews, NC, USA) in a closed
vessel with maximum power input of 300 W. Column chromatography was carried out on silica gel
cartridges using an Isolera One flash purification system (Biotage AB, Uppsala, Sweden) with a linear
gradient of 100% hexanes to 100% ethyl acetate. Before chromatographic purification, crude reaction
mixtures were analyzed by thin layer chromatography in hexanes/ethyl acetate. Purification of final
compounds was performed using asemipreparative HPLC (Waters Technologies Corp., Milford, MA,
USA) with a Vydac protein and peptide C18 reverse phase column using a linear gradient of 100%
solvent A (water with 0.1% TFA) to 100% solvent B (acetonitrile with 0.1% TFA) at a rate of 1% per
minute with UV absorbance monitored at 230 nm. Purity of final compounds was determined on an
Alliance 2690 analytical HPLC (Waters Technologies Corp., Milford, MA, USA) with a Vydac protein
and peptide C18 reverse phase column using the same gradient with UV absorbance monitored at
1
2
30 nm. Purity of final compounds used for testing was
≥
95% as determined by HPLC. H-NMR data
for intermediates and final compounds in CDCl or CD OD was obtained on a 400 MHz or 500 MHz
3
3
Varian spectrometer (Agilent Technologies Inc., Santa Clara, CA, USA). EIMS data was obtained using
an Agilent 6130 HPLC-MS (Agilent Technologies Inc., Santa Clara, CA, USA) in positive ion mode.
HREIMS data was obtained using an Agilent QTOF HPLC-MS (Agilent Technologies Inc., Santa Clara,
CA, USA) in positive ion mode.
4
.1.1. General Procedure A for Microwave Suzuki Coupling of Benzyl Bromide 3 and Pendant
Boronic Acid
Benzyl bromide
3 (1.0 eq), the appropriate boronic acid (1.5 eq), Pd(dppf)Cl (0.1 eq), and K CO
2 2 3
(
3.0 eq) were combined in a microwave vessel equipped with a teflon stirbar. The system was flushed
with argon. A degassed mixture of 3:1 acetone:water (3 mL) was added, and the reaction was heated in
◦
a microwave to 100 C for 30 min. The product was purified via silica gel chromatography in ethyl
acetate/hexanes.
4
.1.2. General Procedure B for HCl Boc Deprotection, Peptide Coupling, and TFA Boc Deprotection
The appropriate Boc-protected amine intermediate was dissolved in 1,4-dioxane (2–5 mL) and
excess concentrated HCl (100–500 µL) was added. The reaction mixture stirred at room temperature
for 1–3.5 h. The solvent was removed under vacuum to yield the deprotected amine. The amine
intermediate (1.0 eq), diBoc-DMT (1.05 eq), PyBOP (1.0 eq), and 6Cl-HOBt (1.0 eq) were combined,
and the reaction flask was flushed with argon. Dry DMF (3–12 mL) and DIEA (10 eq) were added.
The reaction mixture stirred at room temperature for 6–24 h. The solvent was removed under vacuum,
and the coupled product was purified via silica gel chromatography in ethyl acetate/hexanes. The
Boc-protected compound was dissolved in DCM (2–2.5 mL). An equal volume of TFA was added, and
the reaction mixture stirred at room temperature for 1–1.5 h. The solvent was removed under vacuum,
and the product was purified by semi-preparative HPLC and lyophilized.

Molecules 2019, 24, 4302
10 of 24
4
.1.3. General Procedure C for Microwave Suzuki Coupling of Boronic Ester 6 and Pendant
Benzyl Bromide
The appropriate benzyl bromide (1.5–2.0 eq), intermediate
6 (1.0 eq), Pd(dppf)Cl (0.1 eq), and
2
K CO (3.0 eq) were combined in a microwave vessel equipped with a teflon stirbar. The system was
2
3
flushed with argon. A degassed mixture of 3:1 acetone:water (2–3 mL) was added, and the reaction was
◦
heated in a microwave to 100 C for 30 min. The product was purified via silica gel chromatography in
ethyl acetate/hexanes.
4
.1.4. General Procedure D for Microwave Suzuki Coupling of Boronic Ester 6 and Pendant
Benzyl Bromide
The appropriate benzyl bromide (1.0 eq), intermediate
6 (1.5 eq), Pd(dppf)Cl (0.1 eq), and K CO
2 2 3
(
3.0 eq) were combined in a microwave vessel equipped with a teflon stirbar. The system was flushed
with argon. A degassed mixture of 3:1 acetone:water (2–3 mL) was added, and the reaction was heated
◦
in a microwave to 100 C for 30 min. The product was purified via silica gel chromatography in ethyl
acetate/hexanes.
4
.1.5. General Procedure E for S 2 reaction of Benzyl Bromide 3 and Pendant Amine
N
Benzyl bromide 3 (1.0 eq), the appropriate nucleophile (1.2 eq), and K CO (1.2 eq) were dissolved
2 3
in dry DMF (3 mL) under an inert atmosphere. The reaction stirred at room temperature overnight.
The reaction mixture was partitioned between 2 M NaOH and ethyl acetate. The aqueous layer was
extracted with additional ethyl acetate. Combined organic layers were dried over MgSO filtered, and
4,
concentrated under vacuum to obtain the product.
4
.1.6. General Procedure F for TFA Boc Deprotection, Peptide Coupling, and TFA Boc Deprotection
The appropriate Boc-protected amine intermediate was dissolved in DCM (1–3 mL). An equal
volume of TFA was added, and the reaction mixture stirred at room temperature for 1–1.5 h. The solvent
was removed under vacuum to yield the deprotected amine. The amine intermediate (1.0 eq),
diBoc-DMT (1.05 eq), and PyBOP (1.0 eq) were combined, and the reaction flask was flushed with
argon. Dry DMF (3–12 mL) and DIEA (10 eq) were added. The reaction mixture stirred at room
temperature for 6–24 h. The solvent was removed under vacuum, and the coupled product was
purified via silica gel chromatography in ethyl acetate/hexanes. The Boc-protected compound was
dissolved in DCM (2–2.5 mL). An equal volume of TFA was added, and the reaction mixture stirred at
room temperature for 1–1.5 h. The solvent was removed under vacuum, and the product was purified
by semi-preparative HPLC and lyophilized.
Tert-butyl 7-(hydroxymethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (2). To a solution of compound
1
in dry THF (15 mL), a 2.0 M solution of borane dimethyl sulfide in THF (2.7 mL, 5.41 mmol, 3.0 eq
)
was added dropwise over 15 min under inert atmosphere. The reaction mixture stirred at room
temperature overnight. The reaction was quenched by the addition of methanol (20 mL). The solvent
was removed under vacuum. The crude product was dissolved in ethyl acetate and washed with
saturated aqueous NaHCO and brine. The combined aqueous layers were extracted with ethyl acetate.
3
The combined organic layers were dried over MgSO , filtered, and concentrated under vacuum to
4
1
yield the product as a colorless oil (475 mg, 100%). H-NMR (CDCl , 500 MHz)
δ
7.16 (d, J = 7.9 Hz,
3
1
H), 7.13 (d, J = 5.8 Hz, 1H), 7.12 (s, 1H), 4.65 (s, 2H), 4.57 (s, 2H), 3.63 (t, J = 5.9 Hz, 2H)), 2.82 (t, J =
5
.8 Hz, 2H), 1.49 (s, 9H).
Tert-butyl 7-(bromomethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (3). To a solution of compound
2
(950 mg, 3.61 mmol, 1.0 eq) in DCM (40 mL), CBr (1.32 g, 3.97 mmol, 1.1 eq) and a solution of PPh
4 3
(
1.14 g, 4.33 mmol, 1.2 eq) in DCM (5 mL) were added. The reaction stirred at room temperature for 2 h.
The product was purified via silica gel chromatography in ethyl acetate/hexanes to yield a white solid
1
(
1.08 g, 92%). H-NMR (CDCl , 500 MHz)
δ
7.19 (d, J = 8.3 Hz, 1H), 7.14 (s, 1H), 7.11 (d, J = 7.8 Hz
,
3
1
H), 4.56 (s, 2H), 4.47 (s, 2H), 3.64 (t, J = 6.0 Hz, 2H), 2.82 (t, J = 5.9 Hz, 2H), 1.49 (s, 9H).

Molecules 2019, 24, 4302
11 of 24
Tert-butyl 7-bromo-3,4-dihydroisoquinoline-2(1H)-carboxylate (
5). 7-bromo-1,2,3,4-tetrahydroisoqui
noline (75 µL, 0.50 mmol, 1.0 eq) and di-tert-butyl dicarbonate (120 mg, 0.55 mmol, 1.1 eq) were
4
combined in a microwave vessel equipped with a teflon stirbar. The system was flushed with argon,
◦
and the reaction was heated in a microwave to 100 C for 15 min. The reaction mixture was diluted
with DCM and washed with saturated aqueous NaHCO and brine. The organic layer was dried over
3
MgSO filtered, and concentrated under vacuum to obtain the product as a pale orange oil (145 mg,
4
,
1
9
9%). H-NMR (CDCl , 500 MHz)
δ
7.25 (m, 2H), 6.98 (d, J = 8.0 Hz, 1H), 4.52 (s, 2H), 3.61 (t, J = 5.7 Hz
,
3
2
H), 2.75 (t, J = 6.0 Hz, 2H), 1.48 (s, 9H).
Tert-butyl 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (
6
).
Intermediate
5 (945 mg, 3.03 mmol, 1.0 eq), bis(pinacolato)diboron (1.54 g, 6.06 mmol, 2.0 eq),
Pd(dppf)Cl (222 mg, 0.303 mmol, 0.1 eq), and potassium acetate (892 mg, 9.09 mmol, 3.0 eq) were
2
combined in DMSO (20 mL), and the system was flushed with argon. The reaction was heated to
◦
9
0 C overnight. The reaction mixture was concentrated under vacuum to remove most DMSO. The
remaining mixture was diluted with water and extracted with three portions of DCM. The combined
organic layers were washed with water and brine, dried over MgSO , filtered, and concentrated under
4
vacuum. The product was purified via silica gel chromatography in ethyl acetate/hexanes to yield a
1
pale yellow oil (1.05 g, 96%). H-NMR (CDCl , 500 MHz)
δ 7.59 (d, J = 5.7 Hz, 1H), 7.14 (d, J = 5.5 Hz,
3
1
H), 4.58 (s, 2H), 3.63 (br s, 2H), 2.84 (br s, 2H), 1.48 (s, 9H), 1.34 (s, 12H).
Tert-butyl 7-(2-methoxybenzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7a). Compound 7a was
synthesized following General Procedure A from compound (60 mg, 0.18 mmol, 1.0 eq),
2-methoxyphenyl)boronic acid (42 mg, 0.28 mmol, 1.5 eq), Pd(dppf)Cl (13 mg, 0.02 mmol, 0.1 eq)
3
(
,
2
1
and K CO (76 mg, 0.55 mmol, 3.0 eq) to yield the product as a colorless oil (37 mg, 57%). H-NMR
2
3
(
8
CDCl , 500 MHz)
δ
7.21 (t, J = 7.6 Hz, 1H), 7.07 (d, J = 7.2 Hz, 1H), 7.03 (s, 2H), 6.95 (s, 1H), 6.88 (t, J =
.2 Hz, 2H), 4.52 (s, 2H), 3.93 (s, 2H), 3.83 (s, 3H), 3.63 (s, 2H), 2.79 (t, J = 5.8 Hz, 2H), 1.49 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(2-methoxybenzyl)-3,4-dihydroisoquinolin-2(1H)-yl)
3
(
propan-1-one (8a). Following General Procedure B, intermediate 7a (37 mg, 0.10 mmol) was deprotected
to yield the amine intermediate as a colorless oil. This intermediate was coupled to diBoc-DMT (45 mg,
0
.10 mmol, 1.05 eq) in the presence of PyBOP (54 mg, 0.10 mmol, 1.0 eq), and DIEA (142 µL, 1.04 mmol,
1
0 eq) to yield the product as a brown oil. No 6Cl-HOBt was used. TFA deprotection yielded the
1
product as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
7.17 (t, J = 7.9 Hz, 2H), 7.05 (t, J =
3
7
(
2
(
.4 Hz, 2H), 6.97 (d, J = 7.7 Hz, 1H), 6.94–6.90 (m, 4H), 6.89–6.82 (m, 4H), 6.50 (d, J = 3.9 Hz, 1H), 6.39
s, 2H), 6.33 (s, 2H), 4.59–4.45 (m, 4H), 4.15 (d, J = 15.7 Hz, 1H), 3.86 (d, J = 3.7 Hz, 2H), 3.85 (d, J =
.8 Hz, 2H), 3.78 (d, J = 1.6 Hz, 3H), 3.78 (d, J = 1.2 Hz, 3H), 3.71–3.65 (m, 1H), 3.65–3.58 (m, 1H), 3.33
d, J = 15.7 Hz, 1H), 3.25–3.17 (m, 2H), 3.09 (d, J = 11.4 Hz, 1H), 2.72–2.65 (m, 1H), 2.65–2.58 (m, 1H),
.55–2.45 (m, 1H), 2.24 (s, 6H), 2.18 (s, 6H), 1.98–1.87 (m, 1H). HPLC retention time: 39.1 min. HREIMS
2
m/z 445.2494 (calcd. for C28H32N2O3, 445.2486).
Tert-butyl 7-(2-hydroxybenzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7b). Compound 7b was
synthesized following General Procedure A from compound
3 (60 mg, 0.18 mmol, 1.0 eq), (2-
hydroxyphenyl)boronic acid (38 mg, 0.28 mmol, 1.5 eq), Pd(dppf)Cl (13 mg, 0.02 mmol, 0.1 eq), and
2
1
K CO (76 mg, 0.55 mmol, 3.0 eq) to yield the product as a yellow oil (33 mg, 53%). H-NMR (CDCl ,
2
3
3
5
00 MHz)
δ
7.16–7.10 (m, 2H), 7.08–7.01 (m, 2H), 6.96 (s, 1H), 6.89 (t, J = 7.5 Hz, 1H), 6.79 (d, J = 7.9 Hz,
1
H), 4.51 (s, 2H), 3.95 (s, 2H), 3.62 (s, 2H), 2.78 (t, J = 5.9 Hz, 2H), 1.48 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(2-hydroxybenzyl)-3,4-dihydroisoquinolin-2(1H)-yl)
(
propan-1-one (8b). Following General Procedure B, intermediate 7b (33 mg, 0.10 mmol) was deprotected
to yield the amine intermediate as an off-white solid. This intermediate was coupled to diBoc-DMT
(42 mg, 0.10 mmol, 1.05 eq) in the presence of PyBOP (50 mg, 0.10 mmol, 1.0 eq), and DIEA (132
µL,
0
.97 mmol, 10 eq) to yield the product as a brown oil. No 6Cl-HOBt was used. TFA deprotection
1
yielded the product as a white solid. H-NMR (CD OD, 500 MHz, rotamers) δ 7.01 (ddt, J = 10.9, 5.1,
3
1.8 Hz, 4H), 6.97 (d, J = 7.6 Hz, 3H), 6.92 (d, J = 7.9 Hz, 1H), 6.88 (d, J = 7.9 Hz, 1H), 6.78–6.70 (m, 4H),
6
.55 (s, 1H), 6.40 (s, 2H), 6.33 (s, 2H), 4.61 (d, J = 16.9 Hz, 1H), 4.58–4.49 (m, 2H), 4.47 (d, J = 16.5 Hz,

Molecules 2019, 24, 4302
12 of 24
1
1
1
H), 4.16 (d, J = 15.6 Hz, 1H), 3.87 (s, 2H), 3.86 (s, 2H), 3.75–3.68 (m, 1H), 3.66–3.59 (m, 1H), 3.34 (d, J =
5.8 Hz, 1H), 3.26–3.17 (m, 3H), 3.10–3.05 (m, 2H), 2.72–2.65 (m, 2H), 2.65–2.59 (m, 2H), 2.55–2.45 (m,
H), 2.24 (s, 6H), 2.19 (s, 6H), 2.03–1.95 (m, 2H). HPLC retention time: 32.8 min. HREIMS m/z 431.2337
(calcd. for C27H30N2O3, 431.2329).
tert-Butyl 7-(2-(trifluoromethyl)benzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7c). Compound 7c
was synthesized following General Procedure C from intermediate
6 (100 mg, 0.278 mmol, 1.0 eq),
1
-(bromomethyl)-2-(trifluoromethyl)benzene (133 mg, 0.556 mmol, 2.0 eq), Pd(dppf)Cl (20 mg, 0.028
2
1
mmol, 0.1 eq), and K CO (115 mg, 0.834 mmol, 3.0 eq) to yield a colorless oil (27 mg 25%). H-NMR
2
3
(
7
CDCl , 400 MHz)
δ
7.67 (d, J = 7.9 Hz, 1H), 7.43 (t, J = 7.5 Hz, 1H), 7.31 (t, J = 7.7 Hz, 1H), 7.18 (d, J =
.9 Hz, 1H), 7.06 (d, J = 7.8 Hz, 1H), 6.95 (d, J = 7.0 Hz, 1H), 6.89 (s, 1H), 4.52 (s, 2H), 4.14 (s, 2H), 3.63 (t,
J = 5.4 Hz, 2H), 2.80 (t, J = 5.9 Hz, 2H), 1.48 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(2-(trifluoromethyl)benzyl)-3,4-dihydroisoquinolin-2
1H)-yl)propan-1-one (8c). Following General Procedure B, intermediate 7c (27 mg, 0.069 mmol, 1.0 eq)
3
(
(
was deprotected to yield the amine intermediate as a white solid. This intermediate was coupled
to diBoc-DMT (30 mg, 0.074 mmol, 1.05 eq) in the presence of PyBOP (36 mg, 0.070 mmol, 1.0 eq)
and DIEA (98
µ
L, 0.700 mmol, 10 eq) to yield the product as a brown oil. No 6Cl-HOBt was used.
1
TFA deprotection yielded the product as an off-white solid (24 mg, 57%, 3 steps). H-NMR (CD OD,
3
5
00 MHz, rotamers) δ 7.70–7.66 (m, 2H), 7.54–7.48 (m, 2H), 7.40–7.35 (m, 2H), 7.28–7.22 (m, 2H), 6.98
(
(
d, J = 7.8 Hz, 1H), 6.95–6.91 (m, 2H), 6.89 (s, 1H), 6.86 (d, 1H), 6.47 (s, 1H), 6.41 (s, 2H), 6.31 (s, 2H), 4.61
d, J = 17.0 Hz, 1H), 4.58–4.51 (m, 2H), 4.49 (d, J = 17.0 Hz, 1H), 4.18 (d, J = 15.8 Hz, 1H), 4.12 (s, 2H),
4
.11 (s, 2H), 3.86–3.80 (m, 1H), 3.59–3.52 (m, 1H), 3.39 (d, J = 15.8 Hz, 1H), 3.26–3.18 (m, 3H), 3.11–3.06
(
(
m, 2H), 2.76–2.69 (m, 2H), 2.68–2.63 (m, 1H), 2.56–2.49 (m, 1H), 2.24 (s, 6H), 2.20 (s, 6H), 2.01–1.92
m, 1H). 19F-NMR (CD OD, 470 MHz, rotamers) δ −60.76,
−
77.10. HPLC retention time: 42.2 min.
3
HREIMS m/z 483.2262 (calcd. for C28H29F3N2O2, 483.2254).
tert-Butyl 7-(2-cyanobenzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7d). Compound 7d was
synthesized following General Procedure C from intermediate (75 mg, 0.209 mmol, 1.0 eq),
-(bromomethyl)phenol (78 mg, 0.418 mmol, 2.0 eq), Pd(dppf)Cl (15 mg, 0.021 mmol, 0.1 eq),
6
3
2
1
and K CO (87 mg, 0.627 mmol, 3.0 eq) to yield the product as a colorless oil (26 mg, 37%). H-NMR
2
3
(
6
2
CDCl , 500 MHz)
δ
7.15 (t, J = 7.8 Hz, 1H), 7.04 (d, J = 7.8 Hz, 1H), 6.99 (d, J = 7.3 Hz, 1H), 6.92 (s, 1H),
.79–6.74 (m, 1H), 6.69 (d, J = 8.1 Hz, 1H), 6.65 (s, 1H), 4.52 (s, 2H), 3.87 (s, 2H), 3.62 (t, J = 5.9 Hz, 2H),
.78 (t, J = 5.9 Hz, 2H), 1.49 (s, 9H).
S)-2-((2-(2-amino-3-(4-hydroxy-2,6-dimethylphenyl)propanoyl)-1,2,3,4-tetrahydroisoquinolin-7-yl)me
3
(
thyl)benzonitrile (8d). Following General Procedure B, intermediate 7d (20 mg, 0.057 mmol, 1.0 eq)
was deprotected to yield the amine intermediate as a colorless oil. This intermediate was coupled to
diBoc-DMT (24 mg, 0.059 mmol, 1.05 eq) in the presence of PyBOP (29 mg, 0.056 mmol, 1.0 eq) and
DIEA (80
µ
L, 0.560 mmol, 10 eq) to yield the product as a brown oil. No 6Cl-HOBt was used. TFA
1
deprotection yielded the product as a white solid (3 mg, 10%, 3 steps). H-NMR (CD OD, 500 MHz,
3
rotamers)
δ
7.70 (s, 1H), 7.68 (s, 1H), 7.62–7.57 (m, 2H), 7.45–7.35 (m, 4H), 7.03 (d, J = 8.1 Hz, 1H), 7.00
(s, 1H), 6.99–6.97 (m, 2H), 6.95 (d, J = 7.8 Hz, 1H), 6.56 (s, 1H), 6.39 (s, 2H), 6.29 (s, 2H), 4.62 (d, J =
1
6.9 Hz, 1H), 4.57–4.49 (m, 3H), 4.18 (d, J = 15.9 Hz, 1H), 4.15 (s, 2H), 4.13 (d, J = 3.1 Hz, 2H), 3.58–3.51
(
m, 1H), 3.43 (d, J = 16.2 Hz, 0H), 3.26–3.18 (m, 4H), 3.06 (dd, J = 13.8, 4.2 Hz, 2H), 2.74–2.69 (m, 2H),
2
3
.69–2.62 (m, 1H), 2.56–2.49 (m, 1H), 2.24 (s, 6H), 2.19 (s, 6H), 2.01–1.95 (m, 1H). HPLC retention time:
4.4 min. HREIMS m/z 440.2336 (calcd. for C28H29N3O2, 440.2333).
Tert-butyl 7-(3-methoxybenzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7e). Compound 7e was
synthesized following General Procedure D from 3-methoxybenzyl bromide (42
µ
L, 0.30 mmol, 1.0 eq)
,
,
intermediate (161 mg, 0.45 mmol, 1.5 eq), Pd(dppf)Cl (22 mg, 0.03 mmol, 0.1 eq), and K CO (124 mg
6
2
2
3
1
0
7
.90 mmol, 3.0 eq) to yield the product as a colorless oil (42 mg, 40%). H-NMR (CDCl , 500 MHz) δ
3
.22 (t, J = 7.8 Hz, 1H), 7.06 (d, J = 7.8 Hz, 1H), 7.01 (d, J = 7.9 Hz, 1H), 6.94 (s, 1H), 6.79 (d, J = 7.5 Hz,
1H), 6.77–6.73 (m, 2H), 4.53 (s, 2H), 3.91 (s, 2H), 3.79 (s, 3H), 3.63 (br s, 2H), 2.80 (t, J = 5.9 Hz, 2H), 1.50
(s, 9H).

Molecules 2019, 24, 4302
13 of 24
(
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(3-methoxybenzyl)-3,4-dihydroisoquinolin-2(1H)-yl)
propan-1-one (8e). Following General Procedure B, intermediate 7e (42 mg, 0.119 mmol, 1.0 eq) was
deprotected to yield the amine intermediate as a white solid. This intermediate was coupled to
diBoc-DMT (50 mg, 0.123 mmol, 1.05 eq) in the presence of PyBOP (61 mg, 0.117 mmol, 1.0 eq),
6
Cl-HOBt (20 mg, 0.117 mmol, 1.0 eq), and DIEA (164 µL, 1.17 mmol, 10 eq). Silica gel chromatography
yielded the coupled product as a colorless oil (66 mg, 88%). TFA deprotection yielded the product as a
1
white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ 7.16 (td, J = 7.8, 3.5 Hz, 2H), 6.99 (dd, J = 7.7,
3
1
(
3
.7 Hz, 1H), 6.97–6.93 (m, 3H), 6.91 (d, J = 7.8 Hz, 1H), 6.76–6.70 (m, 6H), 6.49 (s, 1H), 6.40 (s, 2H), 6.32
s, 2H), 4.61 (d, J = 16.9 Hz, 1H), 4.58–4.53 (m, 2H), 4.50 (d, J = 16.9 Hz, 1H), 4.17 (d, J = 15.7 Hz, 1H),
.87 (s, 2H), 3.86 (s, 2H), 3.78–3.76 (m, 1H), 3.75 (s, 3H), 3.74 (s, 3H), 3.66–3.53 (m, 1H), 3.37 (d, J = 15.8
Hz, 1H), 3.26–3.17 (m, 3H), 3.11–3.05 (m, 2H), 2.70 (q, J = 7.3, 6.7 Hz, 2H), 2.67–2.62 (m, 1H), 2.52 (dt, J
16.2, 6.2 Hz, 1H), 2.24 (s, 6H), 2.19 (s, 6H), 2.01–1.94 (m, 1H). HPLC retention time: 37.9 min. EIMS
=
m/z 445.3 (calcd. for C28H32N2O3, 445.24).
tert-Butyl 7-(3-hydroxybenzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7f). Compound 7f was
synthesized following General Procedure C from intermediate
6 (75 mg, 0.209 mmol, 1.0 eq),
3
-(bromomethyl)phenol (78 mg, 0.418 mmol, 2.0 eq), Pd(dppf)Cl (15 mg, 0.021 mmol, 0.1 eq),
2
1
and K CO (87 mg, 0.627 mmol, 3.0 eq) to yield the product as a colorless oil (26 mg, 37%). H-NMR
2
3
(
6
2
CDCl , 500 MHz)
δ
7.15 (t, J = 7.8 Hz, 1H), 7.04 (d, J = 7.8 Hz, 1H), 6.99 (d, J = 7.3 Hz, 1H), 6.92 (s, 1H),
.79–6.74 (m, 1H), 6.69 (d, J = 8.1 Hz, 1H), 6.65 (s, 1H), 4.52 (s, 2H), 3.87 (s, 2H), 3.62 (t, J = 5.9 Hz, 2H),
.78 (t, J = 5.9 Hz, 2H), 1.49 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(3-hydroxybenzyl)-3,4-dihydroisoquinolin-2(1H)-yl)
3
(
propan-1-one (8f). Following General Procedure B, intermediate 7f (26 mg, 0.077 mmol, 1.0 eq) was
deprotected to yield the amine intermediate as a colorless oil. This intermediate was coupled to
diBoc-DMT (33 mg, 0.080 mmol, 1.05 eq) in the presence of PyBOP (40 mg, 0.076 mmol, 1.0 eq) and
DIEA (110 µL, 0.760 mmol, 10 eq) to yield the crude product as a brown oil. No 6Cl-HOBt was used.
The crude product was purified via silica gel chromatography in ethyl acetate/hexanes. Subsequent
1
TFA deprotection yielded the product as a white solid (8 mg, 40%, 3 steps). H-NMR (CD OD,
3
5
00 MHz, rotamers) δ 7.09–7.03 (m, 2H), 6.99 (d, J = 8.5 Hz, 1H), 6.96–6.93 (m, 3H), 6.91 (d, J = 7.8 Hz,
1
1
H), 6.65 (s, 1H), 6.64 (s, 1H), 6.61–6.57 (m, 3H), 6.49 (s, 1H), 6.40 (s, 2H), 6.32 (s, 2H), 4.62 (d, J =
6.9 Hz, 1H), 4.58–4.52 (m, 2H), 4.49 (d, J = 16.7 Hz, 1H), 4.17 (d, J = 15.7 Hz, 1H), 3.82 (s, 2H), 3.81 (s,
2H), 3.79–3.72 (m, 1H), 3.62–3.56 (m, 1H), 3.37 (d, J = 15.8 Hz, 1H), 3.26–3.17 (m, 3H), 3.07 (dd, J = 13.7,
4
.2 Hz, 2H), 2.75–2.67 (m, 2H), 2.67–2.60 (m, 1H), 2.55–2.48 (m, 1H), 2.24 (s, 6H), 2.20 (s, 6H), 2.02–1.95
(m, 1H). HPLC retention time: 30.9 min. HREIMS m/z 431.2331 (calcd. for C27H30N2O3, 431.2329).
Tert-butyl 7-(3-(trifluoromethyl)benzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7g). Compound
7
g
was synthesized following General Procedure D from 3-(trifluromethyl)benzyl bromide (33
µL,
0
.21 mmol, 1.0 eq), intermediate (114 mg, 0.32 mmol, 1.5 eq), Pd(dppf)Cl (15 mg, 0.02 mmol, 0.1 eq),
6
2
and K CO (88 mg, 0.64 mmol, 3.0 eq) to yield the product as a colorless oil (28 mg, 34%). EIMS m/z
2
3
4
14.2 (calcd. for C22H24F3NO2 + Na, 414.17).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(3-(trifluoromethyl)benzyl)-3,4-dihydroisoquinolin-2
1H)-yl)propan-1-one (8g). Following General Procedure B, intermediate 7g (28 mg, 0.072 mmol, 1.0 eq)
(
(
was deprotected to yield the amine intermediate as a yellow oil. The crude product was rinsed with
two 2 mL portions of diethyl ether to yield a white solid. This intermediate was coupled to diBoc-DMT
(56 mg, 0.138 mmol, 1.92 eq) in the presence of PyBOP (68 mg, 0.131 mmol, 1.82 eq), 6Cl-HOBt (22 mg,
0
.131 mmol, 1.82 eq), and DIEA (184 L, 1.31 mmol, 18 eq). Silica gel chromatography yielded the
µ
coupled product as a colorless oil (64 mg, 72%). TFA deprotection yielded the product as a white solid.
1
H-NMR (CD OD, 500 MHz, rotamers)
δ 7.50–7.43 (m, 8H), 7.02–6.97 (m, 3H), 6.96–6.93 (m, 2H), 6.48
3
(
4
(
(
s, 1H), 6.39 (s, 2H), 6.30 (s, 2H), 4.62 (d, J = 16.9 Hz, 1H), 4.59–4.51 (m, 2H), 4.52 (d, J = 17.3 Hz, 1H),
.18 (d, J = 15.8 Hz, 1H), 4.00 (s, 2H), 3.99 (s, 2H), 3.81 (dt, J = 12.9, 5.7 Hz, 1H), 3.59–3.52 (m, 1H), 3.40
d, J = 15.8 Hz, 1H), 3.26–3.17 (m, 3H), 3.08 (dt, J = 13.8, 4.1 Hz, 2H), 2.71 (q, J = 5.4 Hz, 2H), 2.68–2.64
19
m, 1H), 2.56–2.49 (m, 1H), 2.23 (s, 6H), 2.19 (s, 6H), 2.00–1.93 (m, 1H). F NMR (CD OD, 470 MHz,
3

Molecules 2019, 24, 4302
14 of 24
rotamers) δ −64.03 (d, J = 28.1 Hz),
−
77.17. HPLC retention time: 43.3 min. EIMS m/z 483.3 (calcd. for
C28H29F3N2O2, 483.22).
Tert-butyl 7-(3-cyanobenzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7h). Compound 7h was
synthesized following General Procedure D from 3-(bromomethyl)benzonitrile (40 mg, 0.20 mmol,
1
(
5
(
.0 eq), intermediate
84 mg, 0.61 mmol, 3.0 eq) to yield the product as a colorless oil (30 mg, 43%). H-NMR (CDCl3,
00 MHz 7.50 (dt, J = 7.4, 1.5 Hz, 1H), 7.45 (s, 1H), 7.43 (t, J = 8.0 Hz, 1H), 7.39 (t, J = 7.6 Hz, 2H), 7.08
d, J = 7.8 Hz, 1H), 6.96 (d, J = 7.7 Hz, 1H), 6.90 (s, 1H), 4.54 (s, 2H), 3.96 (s, 2H), 3.64 (t, J = 5.9 Hz, 2H),
.81 (t, J = 5.9 Hz, 2H), 1.49 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(3-isocyanobenzyl)-3,4-dihydroisoquinolin-2(1H)-yl)
6 (109 mg, 0.30 mmol, 1.5 eq), Pd(dppf)Cl (15 mg, 0.02 mmol, 0.1 eq), and K CO
2 2 3
1
) δ
2
(
propan-1-one (8h). Following General Procedure B, intermediate 7h (30 mg, 0.086 mmol, 1.0 eq) was
deprotected to yield the amine intermediate as an off-white solid (21 mg, 84%). The crude product
was rinsed with three small portions of diethyl ether. This intermediate was coupled to diBoc-DMT
(
32 mg, 0.077 mmol, 1.05 eq) in the presence of PyBOP (39 mg, 0.074 mmol, 1.0 eq), 6Cl-HOBt (13 mg
,
0
.074 mmol, 1.0 eq), and DIEA (104 L, 0.74 mmol, 10 eq). Silica gel chromatography yielded the
µ
coupled product as a white solid (27 mg, 57%). TFA deprotection yielded the product as a white solid.
1
H-NMR (CD OD, 500 MHz, rotamers)
δ
7.57–7.51 (m, 6H), 7.48–7.43 (m, 2H), 7.03–6.99 (m, 3H),
.97–6.94 (m, 2H), 6.48 (s, 1H), 6.39 (s, 2H), 6.29 (s, 2H), 4.63 (d, J = 17.0 Hz, 1H), 4.59–4.49 (m, 2H), 4.53
d, J = 17.6 Hz, 2H), 4.18 (d, J = 15.8 Hz, 1H), 3.98 (s, 2H), 3.97 (s, 2H), 3.83 (dt, J = 12.3, 5.6 Hz, 1H),
.59–3.51 (m, 1H), 3.41 (d, J = 15.7 Hz, 1H), 3.27–3.18 (m, 3H), 3.07 (dt, J = 13.8, 4.0 Hz, 2H), 2.74–2.70
m, 2H), 2.69–2.63 (m, 1H), 2.53 (dt, J = 16.2, 5.9 Hz, 1H), 2.24 (s, 6H), 2.20 (s, 6H), 1.98 (dt, J = 16.0,
3
6
(
3
(
5
.8 Hz, 1H). HPLC retention time: 35.2 min. EIMS m/z 440.2 (calcd. for C28H29N3O2, 440.23).
Tert-butyl 7-(2,3-dimethylbenzyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7i). Compound 7i was
synthesized following General Procedure C from intermediate
6
(100 mg, 0.278 mmol, 1.0 eq),
1
-(bromomethyl)-2,3-dimethylbenzene (83 mg, 0.417 mmol, 1.5 eq), Pd(dppf)Cl (20 mg, 0.028 mmol,
2
0
.1 eq), and K CO (115 mg, 0.834 mmol, 3.0 eq) to yield the product as a colorless oil (51 mg, 52%).
2 3
1
H-NMR (CDCl , 500 MHz) δ 7.09–7.06 (m, 2H), 7.04 (d, J = 7.6 Hz, 1H), 6.99 (d, J = 6.1 Hz, 1H), 6.93 (d,
3
J = 7.9 Hz, 1H), 6.86 (s, 1H), 4.51 (s, 2H), 3.98 (s, 2H), 3.63 (t, J = 6.8 Hz, 2H), 2.79 (t, J = 6.0 Hz, 2H), 2.30
s, 3H), 2.15 (s, 3H), 1.50 (s, 9H).
S)-2-amino-1-(7-(2,3-dimethylbenzyl)-3,4-dihydroisoquinolin-2(1H)-yl)-3-(4-hydroxy-2,6-dimethylphenyl)
(
(
propan-1-one (8i). Following General Procedure B, intermediate 7i (51 mg, 0.145 mmol) was deprotected
to yield the amine intermediate. This intermediate was coupled to diBoc-DMT (63 mg, 0.153 mmol,
1
.05 eq) in the presence of PyBOP (76 mg, 0.146 mmol, 1.0 eq), and DIEA (199 µL, 1.46 mmol, 10 eq) to
yield the product as a brown oil. No 6Cl-HOBt was used. TFA deprotection yielded the product as a
1
white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
7.04–6.97 (m, 5H), 6.97–6.92 (m, 2H), 6.90–6.88
3
(m, 2H), 6.86–6.82 (m, 2H), 6.41 (s, 1H), 6.40 (s, 2H), 6.30 (s, 2H), 4.58 (d, J = 17.0 Hz, 1H), 4.62–4.48 (m,
2
H), 4.46 (d, J = 16.9 Hz, 1H), 4.14 (d, J = 15.7 Hz, 1H), 3.95 (d, J = 2.8 Hz, 2H), 3.93 (s, 2H), 3.83–3.74 (m,
1
4
(
H), 3.62–3.49 (m, 1H), 3.36 (d, J = 14.8 Hz, 1H), 3.26–3.17 (m, 3H), 3.08 (t, J = 4.9 Hz, 1H), 3.05 (t, J =
.9 Hz, 1H), 2.72–2.61 (m, 3H), 2.54–2.47 (m, 1H), 2.25 (s, 3H), 2.25 (s, 3H), 2.23 (s, 6H), 2.19 (s, 6H), 2.09
d, J = 2.2 Hz, 3H), 2.07 (d, J = 1.6 Hz, 3H), 2.00–1.93 (m, 1H). HPLC retention time: 43.2 min. HREIMS
m/z 443.2699 (calcd. for C29H34N2O2, 443.2693).
Tert-butyl 7-(pyridin-3-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7j). Compound 7j was
synthesized following General Procedure C from intermediate
6 (104 mg, 0.289 mmol, 1.0 eq),
3
-(bromomethyl)pyridine hydrobromide (110 mg, 0.433 mmol, 1.5 eq), Pd(dppf)Cl (21 mg, 0.029
2
mmol, 0.1 eq), and K CO (120 mg, 0.867 mmol, 3.0 eq) to yield the product as a colorless oil (20 mg,
2
3
1
2
(
2
1%). H-NMR (CDCl , 500 MHz)
δ
8.49 (s, 1H), 8.46 (d, J = 4.1 Hz, 1H), 7.46 (d, J = 7.7 Hz, 1H), 7.20
dd, J = 7.8, 4.8 Hz, 1H), 7.07 (d, J = 7.9 Hz, 1H), 6.97 (d, J = 7.9 Hz, 1H), 6.91 (s, 1H), 4.52 (s, 2H), 3.93 (s,
H), 3.62 (br s, 2H), 2.79 (t, J = 6.0 Hz, 2H), 1.48 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(pyridin-3-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-
yl)propan-1-one (8j). Following General Procedure B, intermediate 7j (20 mg, 0.062 mmol, 1.0 eq)
3
(

Molecules 2019, 24, 4302
15 of 24
was deprotected to yield the amine intermediate as a colorless oil. The crude product was rinsed
with several small portions of diethyl ether. This intermediate was coupled to diBoc-DMT (26 mg
,
0
0
.064 mmol, 1.05 eq) in the presence of PyBOP (32 mg, 0.061 mmol, 1.0 eq), 6Cl-HOBt (10 mg,
.061 mmol, 1.0 eq), and DIEA (86
µ
L, 0.61 mmol, 10 eq). Silica gel chromatography yielded the
1
coupled product (8 mg, 21%, 2 steps). TFA deprotection yielded the product as a white solid. H-NMR
(
(
2
(
CD OD, 500 MHz, rotamers)
δ
8.67–8.61 (m, 4H), 8.29 (d, J = 8.1 Hz, 1H), 8.26 (d, J = 8.0 Hz, 1H), 7.87
dd, J = 8.1, 5.5 Hz, 1H), 7.83 (dd, J = 8.0, 5.5 Hz, 1H), 7.09–6.98 (m, 5H), 6.56 (s, 1H), 6.39 (s, 2H), 6.23 (s,
H), 4.64 (d, J = 17.1 Hz, 1H), 4.59–4.52 (m, 2H), 4.53 (d, J = 16.6 Hz, 1H), 4.21 (d, J = 15.8 Hz, 1H), 4.14
s, 2H), 4.12 (d, J = 3.2 Hz, 2H), 3.97 (dt, J = 13.2, 5.3 Hz, 1H), 3.48 (d, J = 16.1 Hz, 1H), 3.43 (dd, J = 13.0,
.5 Hz, 1H), 3.26–3.19 (m, 3H), 3.08 (ddd, J = 13.7, 6.4, 4.1 Hz, 2H), 2.72 (t, J = 6.0 Hz, 2H), 2.69–2.62 (m,
3
6
1
1
H), 2.57–2.49 (m, 1H), 2.23 (s, 6H), 2.20 (s, 6H), 1.99 (dt, J = 16.1, 6.1 Hz, 1H). HPLC retention time:
6.9 min. EIMS m/z 416.2 (calcd. for C26H29N3O2, 416.23).
Tert-butyl 7-(pyridin-4-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7k). Compound 7k was
synthesized following General Procedure C from intermediate
6 (112 mg, 0.312 mmol, 1.0 eq),
4
-(bromomethyl)pyridine hydrobromide (118 mg, 0.468 mmol, 1.5 eq), Pd(dppf)Cl (23 mg, 0.031
2
mmol, 0.1 eq), and K CO (129 mg, 0.936 mmol, 3.0 eq) to yield the product as a colorless oil (20 mg,
2
3
1
2
6
0%). H-NMR (CDCl , 500 MHz)
δ
8.49 (d, J = 5.4 Hz, 2H), 7.13–7.05 (m, 3H), 6.97 (d, J = 7.8 Hz, 1H),
.91 (s, 1H), 4.53 (s, 2H), 3.91 (s, 2H), 3.63 (s, 2H), 2.80 (t, J = 5.0 Hz, 2H), 1.48 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(pyridin-4-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-
3
(
yl)propan-1-one (8k). Following General Procedure B, intermediate 7k (20 mg, 0.062 mmol, 1.0 eq)
was deprotected to yield the amine intermediate as a cloudy, yellow oil. The crude product was
rinsed with several small portions of diethyl ether. This intermediate was coupled to diBoc-DMT
(
26 mg
,
0.064 mmol, 1.05 eq) in the presence of PyBOP (32 mg, 0.061 mmol, 1.0 eq), 6Cl-HOBt (10 mg
,
0
.061 mmol, 1.0 eq), and DIEA (86
µL, 0.61 mmol, 10 eq). Silica gel chromatography yielded the
coupled product (24 mg, 63%, 2 steps). TFA deprotection yielded the product as a white solid.
1
H-NMR (CD OD, 500 MHz, rotamers)
δ 8.69–8.65 (m, 4H), 7.81–7.77 (m, 4H), 7.10–7.07 (m, 2H), 7.06
3
(
(
d, J = 7.8 Hz, 1H), 7.01 (d, 2H), 6.58 (s, 1H), 6.40 (s, 2H), 6.25 (s, 2H), 4.65 (d, J = 17.1 Hz, 1H), 4.58–4.55
m, 2H), 4.53 (d, J = 16.8 Hz, 1H), 4.25–4.22 (m, 3H), 4.20 (d, J = 3.0 Hz, 2H), 3.96 (dt, J = 12.9, 5.3 Hz,
1
H), 3.48 (d, J = 15.0 Hz, 1H), 3.46–3.42 (m, 1H), 3.27–3.19 (m, 3H), 3.08 (ddd, J = 13.7, 6.2, 4.2 Hz, 2H),
2
5
.74 (t, J = 6.1 Hz, 2H), 2.70–2.62 (m, 1H), 2.58–2.51 (m, 1H), 2.24 (s, 6H), 2.21 (s, 6H), 2.00 (dt, J = 16.2,
.8 Hz, 1H). HPLC retention time: 17.0 min. EIMS m/z 416.3 (calcd. for C26H29N3O2, 416.23).
Tert-butyl 7-(piperidin-1-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7l). Compound 7l was
synthesized following General Procedure E from compound
13 L, 0.129 mmol, 1.2 eq), and K CO (18 mg, 0.129 mmol, 1.2 eq) to yield the product as an orange
oil (27 mg, 77%). H-NMR (CDCl , 500 MHz)
3 (35 mg, 0.107 mmol, 1.0 eq), piperidine
(
µ
2
3
1
δ
7.10 (d, J = 8.0 Hz, 1H), 7.09–7.03 (m, 2H), 4.56 (s,
H), 3.63 (s, 2H), 3.42 (s, 2H), 2.80 (t, J = 5.7 Hz, 2H), 2.36 (s, 4H), 1.57 (p, J = 5.5 Hz, 4H), 1.48 (s, 9H),
.47–1.39 (m, 2H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(piperidin-1-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-
3
2
1
(
yl)propan-1-one (8l). Following General Procedure B, intermediate 7l (27 mg, 0.082 mmol, 1.0 eq)
was deprotected to yield the amine intermediate as an orange oil. The crude product was rinsed
with several small portions of diethyl ether. This intermediate was coupled to diBoc-DMT (35 mg,
0
.087 mmol, 1.05 eq) in the presence of PyBOP (43 mg, 0.082 mmol, 1.0 eq), 6Cl-HOBt (14 mg, 0.082
mmol, 1.0 eq), and DIEA (115
µ
L, 0.82 mmol, 10 eq). TFA deprotection yielded the product as a white
1
solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
6.48–6.45 (m, 2H), 6.41 (d, J = 7.8 Hz, 1H), 6.35 (d, J =
3
7
3
.8 Hz, 1H), 6.32 (d, J = 8.3 Hz, 1H), 5.97 (s, 1H), 5.59 (s, 2H), 5.41 (s, 2H), 3.89 (d, J = 17.3 Hz, 1H),
.82–3.76 (m, 3H), 3.46 (d, J = 16.0 Hz, 1H), 3.43–3.32 (m, 4H), 3.24 (dt, J = 13.0, 5.2 Hz, 1H), 2.73 (d, J =
1
=
6.1 Hz, 2H), 2.67–2.56 (m, 5H), 2.46–2.37 (m, 3H), 2.32–2.25 (m, 2H), 2.10 (q, J = 10.9 Hz, 4H), 1.97 (t, J
5.7 Hz, 2H), 1.93–1.85 (m, 1H), 1.79 (dt, J = 16.4, 6.5 Hz, 1H), 1.43 (s, 6H), 1.40 (s, 6H), 1.21 (dt, J =
6.4, 5.4 Hz, 1H), 1.13 (s, 2H), 1.10 (s, 3H), 1.01 (d, J = 14.4 Hz, 2H), 0.92 (q, J = 13.2 Hz, 4H), 0.74–0.66
m, 2H). HPLC retention time: 16.7 min. EIMS m/z 422.3 (calcd. for C26H35N3O2, 422.27).
1
(

Molecules 2019, 24, 4302
16 of 24
Tert-butyl 7-(pyrrolidin-1-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7m). Compound 7m
was synthesized following General Procedure E from compound (29 mg, 0.089 mmol, 1.0 eq),
pyrrolidine (9 L, 0.107 mmol, 1.2 eq), and K CO (15 mg, 0.107 mmol, 1.2 eq) to yield the product as a
dark yellow-orange oil (28 mg, 100%). H-NMR (CDCl , 500 MHz)
3
µ
2
3
1
δ
7.13–7.10 (m, 1H), 7.10–7.06 (m,
H), 4.56 (s, 2H), 3.67–3.61 (m, 2H), 3.58 (s, 2H), 2.80 (d, J = 6.1 Hz, 2H), 2.55–2.47 (m, 4H), 1.79 (p, J =
.0 Hz, 4H), 1.49 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(pyrrolidin-1-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-
3
2
3
(
yl)propan-1-one (8m). Following General Procedure B, intermediate 7m (28 mg, 0.088 mmol, 1.0 eq)
was deprotected to yield the amine intermediate. The crude product was rinsed with several small
portions of diethyl ether. This intermediate was coupled to diBoc-DMT (37 mg, 0.091 mmol, 1.05 eq) in
the presence of PyBOP (45 mg, 0.087 mmol, 1.0 eq), 6Cl-HOBt (15 mg, 0.087 mmol, 1.0 eq), and DIEA
1
(
122 µL, 0.87 mmol, 10 eq). TFA deprotection yielded the product as a white solid. H-NMR (CD OD,
3
5
7
4
3
6
2
4
00 MHz, rotamers) δ 7.31–7.26 (m, 2H), 7.24 (d, J = 7.6 Hz, 2H), 7.17 (d, J = 7.9 Hz, 1H), 7.13 (d, J =
.8 Hz, 1H), 6.79 (s, 1H), 6.41 (s, 2H), 6.23 (s, 2H), 4.70 (d, J = 17.2 Hz, 1H), 4.61 (d, J = 17.0 Hz, 1H),
.62–4.51 (m, 2H), 4.31 (s, 2H), 4.31–4.22 (m, 3H), 4.03 (dt, J = 11.7, 5.2 Hz, 1H), 3.53 (d, J = 16.0 Hz, 1H),
.51–3.41 (m, 5H), 3.29–3.25 (m, 1H), 3.26–3.21 (m, 2H), 3.20–3.13 (m, 4H), 3.12–3.06 (m, 2H), 2.79 (t, J =
.0 Hz, 2H), 2.76–2.67 (m, 1H), 2.59 (dt, J = 16.4, 6.0 Hz, 1H), 2.24 (s, 6H), 2.22 (s, 6H), 2.19–2.16 (m, 4H),
.05–1.97 (m, 5H). HPLC retention time: 15.3 min. HREIMS m/z 408.2649 (calcd. for C25H33N3O2,
08.2646).
tert-butyl 7-(morpholinomethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7n). Compound 7n was
synthesized following General Procedure E from compound
10 L, 0.114 mmol, 1.2 eq), and K CO (16 mg, 0.114 mmol, 1.2 eq) to yield the product as a pale
yellow oil (32 mg, 100%). H-NMR (CDCl , 500 MHz)
3 (31 mg, 0.095 mmol, 1.0 eq), morpholine
(
µ
2
3
1
δ
7.11 (d, J = 7.8 Hz, 1H), 7.08 (s, 2H), 4.56 (s,
H), 3.70 (t, J = 4.7 Hz, 4H), 3.64 (t, J = 6.5 Hz, 2H), 3.45 (s, 2H), 2.81 (t, J = 5.9 Hz, 2H), 2.43 (t, J = 4.6 Hz
H), 1.49 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(morpholinomethyl)-3,4-dihydroisoquinolin-2(1H)-yl)
3
2
4
,
(
propan-1-one (8n). Following General Procedure B, intermediate 7n (28 mg, 0.088 mmol, 1.0 eq) was
deprotected to yield the amine intermediate. The crude product was rinsed with several small portions
of diethyl ether. This intermediate was coupled to diBoc-DMT (37 mg, 0.091 mmol, 1.05 eq) in the
presence of PyBOP (45 mg, 0.087 mmol, 1.0 eq), 6Cl-HOBt (15 mg, 0.087 mmol, 1.0 eq), and DIEA
1
(
122 µL, 0.87 mmol, 10 eq). TFA deprotection yielded the product as a white solid. H-NMR (CD OD,
3
5
00 MHz, rotamers)
δ 7.30–7.28 (m, 2H), 7.24 (d, J = 7.8 Hz, 1H), 7.17 (d, J = 7.6 Hz, 1H), 7.14 (d, J = 7.8
Hz, 1H), 6.81 (s, 1H), 6.40 (s, 2H), 6.22 (s, 2H), 4.70 (d, J = 17.3 Hz, 1H), 4.63–4.55 (m, 3H), 4.31 (s, 2H),
4.29–4.21 (m, 3H), 4.13–4.05 (m, 1H), 4.08–3.99 (m, 4H), 3.74 (q, J = 12.7 Hz, 4H), 3.55 (d, J = 16.0 Hz, 1H),
3.42–3.32 (m, 5H), 3.28–3.25 (m, 1H), 3.25–3.20 (m, 2H), 3.21–3.12 (m, 4H), 3.13–3.06 (m, 2H), 2.79 (t, J =
6.1 Hz, 2H), 2.75–2.66 (m, 1H), 2.60 (dt, J = 16.5, 5.9 Hz, 1H), 2.24 (s, 6H), 2.22 (s, 6H), 2.03 (dt, J = 11.1,
5.7 Hz, 1H). HPLC retention time: 14.0 min. HREIMS m/z 424.2597 (calcd. for C25H33N3O3, 424.2595).
Tert-butyl 7-(naphthalen-1-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7o). Compound 7o
was synthesized following General Procedure C from intermediate
6 (100 mg, 0.278 mmol, 1.0 eq),
1
-(bromomethyl)naphthalene (123 mg, 0.556 mmol, 2.0 eq), Pd(dppf)Cl (20 mg, 0.028 mmol, 0.1 eq),
2
1
and K CO (115 mg, 0.834 mmol, 3.0 eq) to yield the product as a yellow oil (41 mg, 39%). H-NMR
2
3
(
CDCl , 500 MHz)
δ
8.03–7.98 (m, 1H), 7.89–7.86 (m, 1H), 7.78 (d, J = 8.2 Hz, 1H), 7.49–7.45 (m, 2H),
.43 (d, J = 7.7 Hz, 1H), 7.31 (d, J = 7.1 Hz, 1H), 7.03 (s, 2H), 6.95 (s, 1H), 4.50 (s, 2H), 4.41 (s, 2H), 3.63 (s,
H), 2.80 (t, J = 5.9 Hz, 2H), 1.49 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(naphthalen-1-ylmethyl)-3,4-dihydroisoquinolin-2
1H)-yl)propan-1-one (8o). Following General Procedure B, intermediate 7o (41 mg, 0.110 mmol, 1.0 eq
was deprotected to yield the amine intermediate. This intermediate was coupled to diBoc-DMT
47 mg, 0.115 mmol, 1.05 eq) in the presence of PyBOP (57 mg, 0.110 mmol, 1.0 eq), 6Cl-HOBt (57 mg
.330 mmol, 3.0 eq), and DIEA (154 L, 1.1 mmol, 10 eq). Silica gel chromatography yielded the
coupled product (57 mg, 78%, 2 steps). TFA deprotection yielded the product as a white solid.
3
7
2
(
(
)
(
,
0
µ

Molecules 2019, 24, 4302
17 of 24
1
H-NMR (CD OD, 500 MHz, rotamers)
δ
7.98–7.94 (m, 2H), 7.86–7.83 (m, 2H), 7.76 (d, J = 8.2 Hz, 2H),
3
7
2
4
.46–7.39 (m, 6H), 7.35 (d, J = 7.0 Hz, 1H), 7.32 (d, J = 6.9 Hz, 1H), 7.00 (d, J = 7.8 Hz, 1H), 6.97–6.94 (m,
H), 6.92 (s, 1H), 6.88 (d, J = 7.8 Hz, 1H), 6.47 (s, 1H), 6.38 (s, 2H), 6.32 (s, 2H), 4.55 (d, J = 17.1 Hz, 1H),
.52–4.47 (m, 2H), 4.44 (d, J = 16.9 Hz, 1H), 4.38 (s, 4H), 4.10 (d, J = 15.8 Hz, 1H), 3.79 (dt, J = 11.9, 5.6
Hz, 1H), 3.56–3.49 (m, 1H), 3.36 (d, J = 15.8 Hz, 1H), 3.24–3.15 (m, 3H), 3.05 (dt, J = 13.7, 5.1 Hz, 2H),
.68 (t, J = 6.2 Hz, 2H), 2.66–2.60 (m, 1H), 2.49 (dt, J = 16.0, 6.2 Hz, 1H), 2.21 (s, 6H), 2.16 (s, 6H), 1.94 (dt,
2
J = 16.1, 6.0 Hz, 1H). HPLC retention time: 43.3 min. EIMS m/z 465.2 (calcd. for C31H32N2O2, 465.25).
Tert-butyl 7-(naphthalen-2-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7p). Compound 7p
was synthesized following General Procedure C from intermediate
6 (100 mg, 0.278 mmol, 1.0 eq),
2
-(bromomethyl)naphthalene (123 mg, 0.556 mmol, 2.0 eq), Pd(dppf)Cl (20 mg, 0.028 mmol, 0.1 eq),
2
1
and K CO (115 mg, 0.834 mmol, 3.0 eq) to yield the product (42 mg, 40%). H-NMR (CDCl , 500
2
3
3
MHz)
.06 (s, 2H), 6.97 (s, 1H), 4.53 (s, 2H), 4.11 (s, 2H), 3.64 (s, 2H), 2.81 (t, J = 6.0 Hz, 2H), 1.49 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(naphthalen-2-ylmethyl)-3,4-dihydroisoquinolin-2
1H)-yl)propan-1-one (8p). Following General Procedure B, intermediate 7p (42 mg, 0.112 mmol, 1.0 eq)
δ 7.79 (q, J = 8.1, 7.7 Hz, 3H), 7.65 (s, 1H), 7.45 (p, J = 7.1, 6.5 Hz, 2H), 7.32 (d, J = 8.5 Hz, 1H),
7
(
(
was deprotected to yield the amine intermediate. Half of this intermediate (18 mg, 0.058 mmol, 1.0 eq)
was coupled to diBoc-DMT (25 mg, 0.061 mmol, 1.05 eq) in the presence of PyBOP (30 mg, 0.058 mmol,
1
.0 eq), 6Cl-HOBt (10 mg, 0.058 mmol, 1.0 eq), and DIEA (81
µ
L, 0.58 mmol, 10 eq). TFA deprotection
1
yielded the product as a brown solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
7.81–7.72 (m, 6H), 7.63
3
(
d, J = 6.7 Hz, 2H), 7.46–7.38 (m, 4H), 7.29 (d, J = 8.3 Hz, 2H), 7.05 (d, J = 8.5 Hz, 1H), 7.02–6.96 (m, 3H),
6.93 (d, J = 7.8 Hz, 1H), 6.53 (s, 1H), 6.40 (s, 2H), 6.32 (s, 2H), 4.61 (d, J = 16.9 Hz, 1H), 4.57–4.48 (m, 2H),
4.50 (d, J = 16.8 Hz, 1H), 4.16 (d, J = 15.8 Hz, 1H), 4.07 (s, 2H), 4.06 (s, 2H), 3.77 (dt, J = 12.1, 5.7 Hz, 1H),
3.61–3.53 (m, 1H), 3.38 (d, J = 15.8 Hz, 1H), 3.24–3.17 (m, 3H), 3.06 (dt, J = 13.8, 5.2 Hz, 2H), 2.71 (q, J =
5.8 Hz, 2H), 2.67–2.61 (m, 1H), 2.52 (dt, J = 15.7, 6.3 Hz, 1H), 2.22 (s, 6H), 2.18 (s, 6H), 1.98 (dt, J = 16.2,
5.9 Hz, 1H). HPLC retention time: 43.5 min. EIMS m/z 465.3 (calcd. for C31H32N2O2, 465.25).
Tert-butyl 7-(quinolin-8-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7q). Compound 7q
was synthesized following General Procedure A from compound
3 (75 mg, 0.23 mmol, 1.0 eq),
quinolin-8-ylboronic acid (60 mg, 0.345 mmol, 1.5 eq), Pd(dppf)Cl (17 mg, 0.023 mmol, 0.1 eq), and
2
1
K CO (95 mg, 0.69 mmol, 3.0 eq) to yield the product as a colorless oil (18 mg, 21%). H-NMR (CDCl ,
2
3
3
500 MHz)
δ
8.97 (dd, J = 4.2, 1.8 Hz, 1H), 8.15 (dd, J = 8.3, 1.8 Hz, 1H), 7.74–7.65 (m, 1H), 7.46 (s, 1H),
7
1
.44 (s, 1H), 7.41 (dd, J = 8.3, 4.2 Hz, 1H), 7.13 (dd, J = 7.6, 1.8 Hz, 1H), 7.07 (s, 1H), 7.04 (d, J = 7.9 Hz,
H), 4.64 (s, 2H), 4.52 (s, 2H), 3.62 (s, 2H), 2.79 (t, J = 5.3 Hz, 2H), 1.48 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(quinolin-8-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-
(
yl)propan-1-one (8q). Following General Procedure F, intermediate 7q (18 mg, 0.048 mmol) was
deprotected to yield the amine intermediate. This intermediate was coupled to diBoc-DMT (21 mg,
0
.05 mmol, 1.05 eq) in the presence of PyBOP (25 mg, 0.048 mmol, 1.0 eq), and DIEA (84 µL, 0.48 mmol,
1
0 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection yielded the product as a white
1
solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
9.03 (td, J = 4.9, 1.7 Hz, 2H), 8.79 (dd, J = 8.3, 1.7 Hz,
3
1
H), 8.75 (dd, J = 8.3, 1.7 Hz, 1H), 8.04 (dd, J = 8.1, 1.6 Hz, 1H), 8.00 (dd, J = 8.1, 1.4 Hz, 1H), 7.81 (ddd,
J = 13.0, 8.3, 4.9 Hz, 2H), 7.75–7.66 (m, 3H), 7.63 (dd, J = 7.3, 1.4 Hz, 1H), 7.04 (dd, J = 7.8, 1.8 Hz, 1H),
.01 (d, J = 1.7 Hz, 1H), 6.96 (s, 2H), 6.94 (d, J = 7.9 Hz, 1H), 6.58 (s, 1H), 6.38 (s, 2H), 6.24 (s, 2H), 4.61
d, J = 15.8 Hz, 1H), 4.58 (s, 2H), 4.56 (s, 2H), 4.56–4.52 (m, 2H), 4.47 (d, J = 17.0 Hz, 1H), 4.17 (d, J =
5.9 Hz, 1H), 3.84 (dt, J = 12.9, 5.5 Hz, 1H), 3.54–3.47 (m, 1H), 3.38 (d, J = 15.9 Hz, 1H), 3.25–3.16 (m,
H), 3.11–3.05 (m, 2H), 2.69 (q, J = 5.5 Hz, 2H), 2.66–2.59 (m, 1H), 2.52 (dt, J = 16.2, 7.4, 4.8 Hz, 1H), 2.21
7
(
1
3
(
(
s, 6H), 2.18 (s, 6H), 1.98 (ddd, J = 16.2, 6.4, 4.4 Hz, 1H). HPLC retention time: 25.0 min. EIMS m/z 466.3
calcd. for C30H31N3O2, 466.24).
Tert-butyl 7-(isoquinolin-8-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7r). Compound 7r
was synthesized following General Procedure A from compound
3 (77 mg, 0.24 mmol, 1.0 eq),
isoquinolin-8-ylboronic acid (49 mg, 0.283 mmol, 1.2 eq), Pd(dppf)Cl (18 mg, 0.024 mmol, 0.1 eq),
2
1
and K CO (98 mg, 0.71 mmol, 3.0 eq) to yield the product as a pale pink oil (55 mg, 62%). H-NMR
2
3

Molecules 2019, 24, 4302
18 of 24
(
CDCl , 500 MHz)
δ
9.47 (s, 1H), 8.52 (d, J = 5.6 Hz, 1H), 7.73 (d, J = 8.3 Hz, 1H), 7.68–7.60 (m, 2H), 7.40
3
(
1
d, J = 7.0 Hz, 1H), 7.02 (q, J = 8.0 Hz, 2H), 6.94 (s, 1H), 4.48 (s, 4H), 3.61 (s, 2H), 2.77 (t, J = 6.1 Hz, 2H),
.47 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(isoquinolin-8-ylmethyl)-3,4-dihydroisoquinolin-2
1H)-yl)propan-1-one (8r). Following General Procedure F, intermediate 7r (27 mg, 0.072 mmol) was
deprotected to yield the amine intermediate. This intermediate was coupled to diBoc-DMT (31 mg
(
(
,
0.076 mmol, 1.05 eq) in the presence of PyBOP (37 mg, 0.072 mmol, 1.0 eq), and DIEA (125 µL,
0
.72 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection yielded the product
1
as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
9.73 (s, 1H), 9.72 (s, 1H), 8.54 (t, J = 5.0 Hz,
3
2H), 8.38 (d, J = 6.3 Hz, 1H), 8.35 (d, J = 6.2 Hz, 1H), 8.17–8.10 (m, 3H), 8.08 (q, J = 7.6 Hz, 1H), 7.82 (d, J
=
6.6 Hz, 1H), 7.76 (d, J = 6.9 Hz, 1H), 7.07–7.02 (m, 2H), 7.00–6.94 (m, 3H), 6.53 (s, 1H), 6.36 (s, 2H), 6.17
(
s, 2H), 4.61 (s, 2H), 4.60 (s, 2H), 4.57 (d, J = 14.3 Hz, 1H), 4.54 (dd, J = 11.9, 4.1 Hz, 2H), 4.48 (d, J = 17.0
Hz, 1H), 4.16 (d, J = 15.8 Hz, 1H), 3.97 (dt, J = 12.9, 5.3 Hz, 1H), 3.45 (d, J = 15.8 Hz, 1H), 3.38 (dt, J = 13.2,
.7 Hz, 1H), 3.24–3.15 (m, 3H), 3.07 (ddd, J = 13.6, 11.5, 4.1 Hz, 2H), 2.69 (t, J = 6.0 Hz, 2H), 2.62 (ddd, J =
2.5, 7.5, 4.7 Hz, 1H), 2.52 (ddd, J = 16.2, 7.5, 4.8 Hz, 1H), 2.19 (s, 6H), 2.17 (s, 6H), 1.96 (ddd, J = 16.3, 6.9,
6
1
4.7 Hz, 1H). HPLC retention time: 21.4 min. EIMS m/z 466.3 (calcd. for C30H31N3O2, 466.24).
Tert-butyl 7-(isoquinolin-5-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7s). Compound 7s
was synthesized following General Procedure A from compound
3 (73 mg, 0.224 mmol, 1.0 eq),
isoquinolin-5-ylboronic acid (46 mg, 0.269 mmol, 1.2 eq), Pd(dppf)Cl (16 mg, 0.022 mmol, 0.1 eq), and
2
1
K CO (93 mg, 0.672 mmol, 3.0 eq) to yield the product as a yellow oil (48 mg, 57%). H-NMR (CDCl ,
2
3
3
500 MHz)
δ
9.26 (s, 1H), 8.50 (d, J = 6.0 Hz, 1H), 7.88 (d, J = 8.1 Hz, 1H), 7.76 (d, J = 6.0 Hz, 1H), 7.55 (t,
J = 7.6 Hz, 1H), 7.50 (d, J = 7.1 Hz, 1H), 7.04 (d, J = 7.8 Hz, 1H), 6.97 (s, 1H), 6.91 (s, 1H), 4.49 (s, 2H),
.36 (s, 2H), 3.62 (t, J = 5.9 Hz, 2H), 2.78 (t, J = 5.9 Hz, 2H), 1.47 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(isoquinolin-5-ylmethyl)-3,4-dihydroisoquinolin-2
1H)-yl)propan-1-one (8s). Following General Procedure F, intermediate 7s (24 mg, 0.064 mmol) was
deprotected to yield the amine intermediate. This intermediate was coupled to diBoc-DMT (28 mg
4
(
(
,
0
0
.067 mmol, 1.05 eq) in the presence of PyBOP (33 mg, 0.064 mmol, 1.0 eq), and DIEA (111 µL,
.64 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection yielded the product
1
as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
9.67 (d, J = 7.1 Hz, 2H), 8.52 (d, J = 7.0 Hz,
3
2
6
H), 8.44 (t, J = 7.6 Hz, 2H), 8.35 (t, J = 8.9 Hz, 2H), 8.04–7.91 (m, 4H), 7.04–6.92 (m, 5H), 6.51 (s, 1H),
.37 (s, 2H), 6.23 (s, 2H), 4.58 (d, J = 17.4 Hz, 2H), 4.56–4.50 (m, 2H), 4.53 (s, 2H), 4.52 (s, 2H), 4.47 (d, J =
1
7.1 Hz, 1H), 4.15 (d, J = 15.8 Hz, 1H), 3.90 (dt, J = 12.9, 5.4 Hz, 1H), 3.48–3.39 (m, 1H), 3.41 (d, J = 15.8
Hz, 1H), 3.24–3.16 (m, 3H), 3.07 (ddd, J = 13.3, 8.5, 4.1 Hz, 2H), 2.69 (t, 2H), 2.62 (ddd, J = 12.6, 7.5, 4.7
Hz, 1H), 2.51 (ddd, J = 16.2, 7.3, 4.8 Hz, 1H), 2.21 (s, 6H), 2.17 (s, 6H), 1.96 (ddd, J = 16.2, 6.9, 4.9 Hz,
1
H). HPLC retention time: 21.5 min. EIMS m/z 466.3 (calcd. for C30H31N3O2, 466.24).
Tert-butyl 7-(quinolin-5-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7t). Compound 7t was
synthesized following General Procedure A from compound
3 (50 mg, 0.153 mmol, 1.0 eq),
quinolin-5-ylboronic acid (32 mg, 0.184 mmol, 1.2 eq), Pd(dppf)Cl (11 mg, 0.015 mmol, 0.1 eq),
2
1
and K CO (63 mg, 0.459 mmol, 3.0 eq) to yield the product as a yellow oil (36 mg, 63%). H-NMR
2
3
(
CDCl , 500 MHz)
δ
8.90 (dd, J = 4.2, 1.7 Hz, 1H), 8.30 (ddd, J = 8.6, 1.7, 0.9 Hz, 1H), 8.04 (d, J = 8.5 Hz,
3
1
4
H), 7.66 (t, J = 7.8 Hz, 1H), 7.39–7.33 (m, 2H), 7.03 (d, J = 7.6 Hz, 1H), 6.99–6.85 (m, 2H), 4.48 (s, 2H),
.39 (s, 2H), 3.60 (t, J = 6.1 Hz, 2H), 2.77 (t, J = 6.0 Hz, 2H), 1.47 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(quinolin-5-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-
yl)propan-1-one (8t). Following General Procedure F, intermediate 7t (36 mg, 0.096 mmol) was
deprotected to yield the amine intermediate. This intermediate was coupled to diBoc-DMT (41 mg
(
,
0
0
.100 mmol, 1.05 eq) in the presence of PyBOP (49 mg, 0.095 mmol, 1.0 eq), and DIEA (123 µL,
.95 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection yielded the product
1
as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ 9.15 (dd, J = 8.5, 5.5 Hz, 2H), 9.10 (td, J = 5.1,
3
1
.5 Hz, 2H), 8.13 (t, J = 9.1 Hz, 2H), 8.05 (ddd, J = 18.3, 8.7, 7.1 Hz, 2H), 7.93 (ddd, J = 8.7, 7.4, 5.2 Hz,
2H), 7.78 (d, J = 7.1 Hz, 1H), 7.74 (d, J = 7.1 Hz, 1H), 7.04–6.92 (m, 5H), 6.50 (s, 1H), 6.36 (s, 2H), 6.23 (s,

Molecules 2019, 24, 4302
19 of 24
2
(
1
H), 4.59 (d, J = 16.4 Hz, 1H), 4.55 (s, 2H), 4.54 (s, 2H), 4.54–4.52 (m, 2H), 4.47 (d, J = 17.1 Hz, 1H), 4.15
d, J = 15.8 Hz, 1H), 3.90 (dt, J = 12.9, 5.4 Hz, 1H), 3.43 (dt, J = 13.1, 6.6 Hz, 1H), 3.41 (d, J = 15.9 Hz,
H), 3.24–3.16 (m, 3H), 3.07 (ddd, J = 13.3, 8.5, 4.1 Hz, 2H), 2.69 (t, J = 6.0 Hz, 2H), 2.63 (ddd, J = 12.5,
7
4
.4, 4.7 Hz, 1H), 2.50 (ddd, J = 16.2, 7.4, 4.8 Hz, 1H), 2.20 (s, 6H), 2.17 (s, 6H), 1.94 (ddd, J = 16.1, 6.8,
.6 Hz, 1H). HPLC retention time: 21.6 min. EIMS m/z 466.3 (calcd. for C30H31N3O2, 466.24).
Tert-butyl 7-(quinolin-4-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7u). Compound 7u
was synthesized following General Procedure A from compound
3 (50 mg, 0.153 mmol, 1.0 eq),
quinolin-4-ylboronic acid (32 mg, 0.184 mmol, 1.2 eq), Pd(dppf)Cl (11 mg, 0.015 mmol, 0.1 eq), and
2
1
K CO (63 mg, 0.459 mmol, 3.0 eq) to yield the product as a colorless oil (32 mg, 56%). H-NMR
2
3
(
(
2
CDCl , 500 MHz)
δ
8.83 (d, J = 4.4 Hz, 1H), 8.13 (d, J = 8.4 Hz, 1H), 8.03 (dd, J = 8.5, 1.3 Hz, 1H), 7.70
t, J = 7.7 Hz, 1H), 7.54 (t, J = 7.7 Hz, 1H), 7.15 (d, J = 4.2 Hz, 1H), 7.07 (d, J = 7.6 Hz, 1H), 7.03–6.88 (m,
H), 4.50 (s, 2H), 4.40 (s, 2H), 3.63 (t, J = 5.4 Hz, 2H), 2.80 (t, J = 5.8 Hz, 2H), 1.47 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(quinolin-4-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-
yl)propan-1-one (8u). Following General Procedure F, intermediate 7u (32 mg, 0.085 mmol) was
deprotected to yield the amine intermediate. This intermediate was coupled to diBoc-DMT (37 mg
3
(
,
0
0
.089 mmol, 1.05 eq) in the presence of PyBOP (44 mg, 0.085 mmol, 1.0 eq), and DIEA (148 µL,
.85 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection yielded the product
1
as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ 9.07 (dd, J = 9.2, 5.6 Hz, 2H), 8.55 (t, J =
3
9
(
2
1
6
1
.6 Hz, 2H), 8.26 (t, J = 8.8 Hz, 2H), 8.18–8.11 (m, 2H), 8.00–7.93 (m, 2H), 7.78 (d, J = 5.6 Hz, 2H), 7.12
dd, J = 7.7, 1.8 Hz, 1H), 7.10 (s, 1H), 7.05 (s, 2H), 7.01 (d, J = 7.8 Hz, 1H), 6.60 (s, 1H), 6.37 (s, 2H), 6.20 (s,
H), 4.74 (s, 2H), 4.71 (s, 2H), 4.61 (d, J = 17.2 Hz, 1H), 4.55 (dd, J = 12.0, 4.1 Hz, 2H), 4.51 (d, J = 17.1 Hz,
H), 4.20 (d, J = 15.8 Hz, 1H), 3.99 (dt, J = 12.9, 5.3 Hz, 1H), 3.48 (d, J = 15.8 Hz, 1H), 3.40 (dt, J = 13.2,
.7 Hz, 1H), 3.25–3.17 (m, 3H), 3.08 (ddd, J = 13.9, 9.9, 4.1 Hz, 2H), 2.73 (t, J = 6.1 Hz, 2H), 2.65 (ddd, J =
2.5, 7.4, 4.8 Hz, 1H), 2.54 (ddd, J = 16.4, 7.4, 4.9 Hz, 1H), 2.21 (s, 6H), 2.18 (s, 6H), 1.98 (ddd, J = 16.3,
7
.0, 4.7 Hz, 1H). HPLC retention time: 21.2 min. EIMS m/z 466.3 (calcd. for C30H31N3O2, 466.24).
Tert-butyl 7-(isoquinolin-4-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7v). Compound 7v
was synthesized following General Procedure A from compound
3 (50 mg, 0.153 mmol, 1.0 eq),
isoquinolin-4-ylboronic acid (32 mg, 0.184 mmol, 1.2 eq), Pd(dppf)Cl (11 mg, 0.015 mmol, 0.1 eq), and
2
K CO (63 mg, 0.459 mmol, 3.0 eq) to yield the crude product. Silica gel chromatography yielded a
2
3
mixture of products. This mixture was used directly in the next step.
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(isoquinolin-4-ylmethyl)-3,4-dihydroisoquinolin-2
1H)-yl)propan-1-one (8v). Following General Procedure F, intermediate 7v (19 mg, 0.051 mmol) was
(
(
deprotected. The crude product was purified by semi-preparative HPLC to yield the product as a
+
white solid (21 mg, 100%). EIMS calcd. for [C19H18N2 + H] : 275.15, found: 275.2. The amine was
coupled to diBoc-DMT (23 mg, 0.057 mmol, 1.05 eq) in the presence of PyBOP (28 mg, 0.054 mmol
,
1
.0 eq), and DIEA (94
µ
L, 0.54 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA
1
deprotection yielded the product as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
9.65 (d, J =
3
6
7
4
.0 Hz, 2H), 8.50 (t, J = 8.3 Hz, 2H), 8.42–8.35 (m, 4H), 8.22–8.14 (m, 2H), 8.04–7.97 (m, 2H), 7.10 (dd, J =
.8, 1.8 Hz, 1H), 7.07 (s, 1H), 7.02 (s, 2H), 6.97 (d, J = 7.9 Hz, 1H), 6.56 (s, 1H), 6.37 (s, 2H), 6.21 (s, 2H),
.61 (d, J = 16.1 Hz, 1H), 4.57 (s, 2H), 4.56 (s, 2H), 4.55–4.52 (m, 2H), 4.49 (d, J = 17.2 Hz, 1H), 4.17 (d, J
=
15.8 Hz, 1H), 3.95 (dt, J = 12.9, 5.3 Hz, 1H), 3.45 (d, J = 15.5 Hz, 1H), 3.43–3.38 (m, 1H), 3.25–3.17 (m,
H), 3.07 (ddd, J = 13.7, 8.1, 4.1 Hz, 2H), 2.72 (t, J = 6.0 Hz, 2H), 2.65 (ddd, J = 12.7, 7.3, 4.8 Hz, 1H), 2.52
ddd, J = 16.4, 7.1, 4.8 Hz, 1H), 2.21 (s, 6H), 2.17 (s, 6H), 1.95 (dt, J = 16.4, 6.2 Hz, 1H). HPLC retention
3
(
time: 21.5 min. EIMS m/z 466.3 (calcd. for C30H31N3O2, 466.24).
Tert-butyl 7-((3,4-dihydroisoquinolin-2(1H)-yl)methyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7w).
Compound 7w was synthesized following General Procedure E from compound
3
(50 mg, 0.153 mmol,
1
1
.0 eq), 1,2,3,4-tetrahydroisoquinoline (23
.2 eq) to yield the product as a colorless oil (24 mg, 41%). H-NMR (CDCl , 500 MHz)
µL, 0.184 mmol, 1.2 eq), and K CO (25 mg, 0.184 mmol,
2
3
1
δ
7.19 (d, J = 7.5
3
Hz, 1H), 7.15 (s, 1H), 7.12–7.08 (m, 4H), 6.99 (d, J = 7.0 Hz, 1H), 4.57 (s, 2H), 3.66 (br s, 2H), 3.64 (s, 2H),
.63 (s, 2H), 2.90 (t, J = 6.0 Hz, 2H), 2.83 (t, J = 5.9 Hz, 2H), 2.75 (t, J = 5.9 Hz, 2H), 1.49 (s, 9H).
3

Molecules 2019, 24, 4302
20 of 24
(
S)-2-amino-1-(7-((3,4-dihydroisoquinolin-2(1H)-yl)methyl)-3,4-dihydroisoquinolin-2(1H)-yl)-3-(4-hy
droxy-2,6-dimethylphenyl)propan-1-one (8w). Following General Procedure F, intermediate 7w (24 mg,
.063 mmol, 1.0 eq) was deprotected to yield the amine intermediate. This intermediate was coupled
to diBoc-DMT (27 mg, 0.067 mmol, 1.05 eq) in the presence of PyBOP (33 mg, 0.064 mmol, 1.0 eq), and
DIEA (111 L, 0.64 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection
yielded the product as a white solid. 1H-NMR (CD OD, 500 MHz, rotamers)
7.36–7.33 (m, 2H),
.32–7.23 (m, 7H), 7.21–7.13 (m, 4H), 6.85 (s, 1H), 6.41 (s, 2H), 6.21 (s, 2H), 4.72 (d, J = 17.3 Hz, 1H), 4.62
d, J = 17.3 Hz, 1H), 4.64–4.57 (m, 2H), 4.44 (s, 2H), 4.42–4.35 (m, 5H), 4.29 (d, J = 16.0 Hz, 1H), 4.10 (dt,
J = 13.0, 5.0 Hz, 1H), 3.73 (br s, 1H), 3.57 (d, J = 16.1 Hz, 1H), 3.46–3.35 (m, 2H), 3.28–3.26 (m, 1H),
.26–3.15 (m, 6H), 3.15–3.07 (m, 2H), 2.80 (t, J = 6.0 Hz, 2H), 2.76–2.67 (m, 1H), 2.62 (dt, J = 16.5, 6.0 Hz,
H), 2.24 (s, 6H), 2.22 (s, 6H), 2.04 (dt, J = 6.9, 5.0 Hz, 1H). HPLC retention time: 21.3 min. HREIMS m/z
0
µ
δ
3
7
(
3
1
4
70.2799 (calcd. for C30H35N3O2, 470.2802).
Tert-butyl 7-((3,4-dihydroquinolin-1(2H)-yl)methyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7x).
Compound 7x was synthesized following General Procedure E from compound
3
(50 mg, 0.153 mmol
,
1
1
.0 eq), 1,2,3,4-tetrahydroquinoline (23
.2 eq). The reaction mixture was diluted with water. The aqueous layer was extracted with
µL, 0.184 mmol, 1.2 eq), and K CO (25 mg, 0.184 mmol,
2
3
several portions of ethyl acetate. Combined organic layers were dried over MgSO filtered, and
4,
concentrated under vacuum. Silica gel chromatography yielded a mixture of the desired product
and 1,2,3,4-tetrahydroquinoline. The product was partitioned between ethyl acetate and 2 M NaOH.
The aqueous layer was extracted with ethyl acetate. Some 1,2,3,4-tetrahydroquinoline remained.
The mixture was resubmitted to reaction conditions with additional 3 (25 mg, 0.076, 0.5 eq) and K CO
2 3
(
(
25 mg, 0.184 mmol, 1.2 eq). Silica gel chromatography yielded the desired product as a colorless oil
1
24 mg, 28%). H-NMR (CDCl , 500 MHz)
δ
7.08 (s, 1H), 7.02 (s, 1H), 6.99 (t, J = 6.5 Hz, 2H), 6.59 (t, J
7.3 Hz, 1H), 6.51 (d, J = 8.5 Hz, 1H), 4.55 (s, 2H), 4.44 (s, 2H), 3.65 (s, 2H), 3.36 (t, J = 5.7 Hz, 2H),
.92–2.75 (m, 4H), 2.03 (p, J = 6.1 Hz, 2H), 1.50 (s, 9H).
S)-2-amino-1-(7-((3,4-dihydroquinolin-1(2H)-yl)methyl)-3,4-dihydroisoquinolin-2(1H)-yl)-3-(4-hydroxy-
3
=
2
(
2,6-dimethylphenyl)propan-1-one (8x). Following General Procedure F, intermediate 7x (24 mg,
0
.063 mmol, 1.0 eq) was deprotected to yield the amine intermediate. This intermediate was coupled
to diBoc-DMT (27 mg, 0.067 mmol, 1.05 eq) in the presence of PyBOP (33 mg, 0.064 mmol, 1.0 eq), and
DIEA (111
µ
L, 0.64 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection
1
yielded the product as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ 7.09 (d, J = 7.9 Hz, 1H),
3
7
2
.04 (d, J = 8.1 Hz, 2H), 7.01 (d, J = 7.9 Hz, 1H), 6.99–6.89 (m, 5H), 6.64–6.51 (m, 5H), 6.40 (s, 2H), 6.27 (s,
H), 4.62 (d, J = 17.1 Hz, 1H), 4.58–4.53 (m, 2H), 4.51 (d, J = 17.0 Hz, 2H), 4.45 (s, 2H), 4.44 (s, 2H), 4.19
(d, J = 15.8 Hz, 1H), 3.93 (dt, J = 12.9, 5.4 Hz, 1H), 3.51–3.43 (m, 1H), 3.46 (d, J = 15.7 Hz, 1H), 3.40–3.34
(m, 4H), 3.26–3.19 (m, 3H), 3.11–3.05 (m, 2H), 2.81 (q, J = 5.9 Hz, 4H), 2.73 (t, J = 6.2 Hz, 2H), 2.69–2.62
(m, 1H), 2.54 (dt, J = 16.3, 6.1 Hz, 1H), 2.23 (s, 6H), 2.20 (s, 6H), 2.06–1.95 (m, 5H). HPLC retention time:
3
5.9 min. HREIMS m/z 470.2800 (calcd. for C30H35N3O2, 470.2802).
Tert-butyl 7-(indolin-1-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7y). Compound 7y was
synthesized following General Procedure E from compound
3
(50 mg, 0.153 mmol, 1.0 eq), indoline
(
(
21 µL, 0.184 mmol, 1.2 eq), and K CO (25 mg, 0.184 mmol, 1.2 eq) to yield the product as a brown oil
2
3
1
37 mg, 66%). H-NMR (CDCl , 500 MHz) δ 7.18 (d, J = 7.9 Hz, 1H), 7.15–7.08 (m, 3H), 7.07 (t, J = 7.6
3
Hz, 1H), 6.69 (t, J = 7.3 Hz, 1H), 6.52 (d, J = 7.8 Hz, 1H), 4.58 (s, 2H), 4.21 (s, 2H), 3.66 (s, 2H), 3.32 (t, J =
.2 Hz, 2H), 2.99 (t, J = 8.3 Hz, 2H), 2.84 (t, J = 6.0 Hz, 2H), 1.51 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(indolin-1-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-yl)
propan-1-one (8y). Following General Procedure F, intermediate 7y (37 mg, 0.102 mmol, 1.0 eq) was
deprotected to yield the amine intermediate. This intermediate was coupled to diBoc-DMT (43 mg
.105 mmol, 1.05 eq) in the presence of PyBOP (52 mg, 0.100 mmol, 1.0 eq), and DIEA (174 L,
8
(
,
0
µ
1
.0 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection yielded the product
1
as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ 7.24–7.18 (m, 3H), 7.18–7.11 (m, 4H), 7.06 (d,
3
J = 7.9 Hz, 1H), 7.03–6.94 (m, 4H), 6.89 (d, J = 8.5 Hz, 1H), 6.67 (s, 1H), 6.41 (s, 2H), 6.27 (s, 2H), 4.63 (d,

Molecules 2019, 24, 4302
21 of 24
J = 16.9 Hz, 1H), 4.60–4.56 (m, 2H), 4.53 (d, J = 17.2 Hz, 1H), 4.40–4.32 (m, 4H), 4.21 (d, J = 15.8 Hz, 1H),
.89 (dt, J = 12.7, 5.5 Hz, 1H), 3.55–3.50 (m, 1H), 3.50–3.46 (m, 4H), 3.44 (d, J = 15.9 Hz, 1H), 3.28–3.17
m, 3H), 3.13–3.06 (m, 2H), 2.99 (t, J = 7.6 Hz, 4H), 2.74 (t, J = 4.7 Hz, 2H), 2.72–2.65 (m, 1H), 2.56 (dt, J =
6.3, 6.0 Hz, 1H), 2.23 (s, 6H), 2.20 (s, 6H), 1.98 (dt, J = 16.2, 5.9 Hz, 1H). HPLC retention time: 30.1 min.
3
(
1
HREIMS m/z 456.2644 (calcd. for C29H33N3O2, 456.2646).
Tert-butyl 7-(isoindolin-2-ylmethyl)-3,4-dihydroisoquinoline-2(1H)-carboxylate (7z). Compound 7z was
synthesized following General Procedure E from compound
3 (50 mg, 0.153 mmol, 1.0 eq), isoindoline
HCl (29 mg, 0.184 mmol, 1.2 eq), and K CO (25 mg, 0.184 mmol, 1.2 eq) to yield the product as an
2
3
1
orange oil (22 mg, 39%). H-NMR (CDCl , 500 MHz)
δ
7.29 (s, 1H), 7.21 (d, J = 7.8 Hz, 1H), 7.18 (s, 4H),
.12 (d, J = 7.6 Hz, 1H), 4.58 (s, 2H), 3.93 (s, 4H), 3.88 (s, 2H), 3.65 (t, J = 8.2 Hz, 2H), 2.84 (t, J = 6.2 Hz,
H), 1.49 (s, 9H).
S)-2-amino-3-(4-hydroxy-2,6-dimethylphenyl)-1-(7-(isoindolin-2-ylmethyl)-3,4-dihydroisoquinolin-2(1H)-
3
7
2
(
yl)propan-1-one (8z). Following General Procedure F, intermediate 7z (22 mg, 0.060 mmol, 1.0 eq)
was deprotected to yield the amine intermediate. This intermediate was coupled to diBoc-DMT
(26 mg, 0.063 mmol, 1.05 eq) in the presence of PyBOP (31 mg, 0.060 mmol, 1.0 eq), and DIEA (105
µL,
0
.600 mmol, 10 eq) to yield the product. No 6Cl-HOBt was used. TFA deprotection yielded the product
1
as a white solid. H-NMR (CD OD, 500 MHz, rotamers)
δ
7.41–7.38 (m, 8H), 7.37–7.34 (m, 2H), 7.30
3
(
dd, J = 7.9, 1.8 Hz, 1H), 7.19 (dd, J = 7.9, 3.5 Hz, 1H), 7.16 (dd, J = 8.0, 3.7 Hz, 1H), 6.86 (s, 1H), 6.42 (s,
2
4
H), 6.24 (s, 2H), 4.72 (d, J = 17.2 Hz, 1H), 4.70–4.64 (m, 8H), 4.61 (d, J = 15.3 Hz, 1H), 4.65–4.57 (m, 2H),
.56 (s, 2H), 4.53 (d, J = 10.7 Hz, 2H), 4.29 (d, J = 16.0 Hz, 1H), 4.09 (dt, J = 13.2, 4.9 Hz, 1H), 3.57 (d, J =
1
=
5.9 Hz, 1H), 3.44–3.37 (m, 1H), 3.29–3.25 (m, 1H), 3.24 (q, J = 13.0 Hz, 2H), 3.15–3.07 (m, 2H), 2.80 (t, J
4.3 Hz, 2H), 2.75–2.67 (m, 1H), 2.61 (dt, J = 16.2, 5.6 Hz, 1H), 2.25 (s, 6H), 2.23 (s, 6H), 2.07–2.00 (m,
H). HPLC retention time: 20.1 min. HREIMS m/z 456.2645 (calcd. for C29H33N3O2, 456.2646).
1
4
.2. Pharmacology
The pharmacological methods used were the same as those previously described [14], with any
changes noted below. Unless otherwise noted, all tissue culture reagents and radiolabeled ligands
were purchased from commercial sources. Cell lines were provided by Professor Lawrence Toll [34].
4
.2.1. Cell Lines and Membrane Preparations
Membranes prepared from transfected Chinese Hamster Ovary cells stably expressing human
◦
KOR, human MOR, or human DOR were used for all assays. Cells were grown to confluence at 37 C
in 5% CO in 1:1 DMEM:F12 media with 10% v/v fetal bovine serum and 5% v/v penicillin/streptomycin.
2
Membranes were prepared by washing confluent cells three times with ice cold phosphate-buffered
saline (0.9% NaCl, 0.61 mM Na HPO , 0.38 mM KH PO , pH 7.4). Cells were detached from the plates
2
4
2
4
by incubation in warm harvesting buffer (20 mM HEPES, 150 mM NaCl, 0.68 mM EDTA, pH 7.4)
and pelleted by centrifugation at 200 g for 3 min. The cell pellet was suspended in ice-cold 50 mM
Tris-HCl buffer, pH 7.4 and homogenized with a Tissue Tearor (Biospec Products, Inc, distributed by
×
Cole-Parmer, Vernon Hills, IL, USA) for 20 s at setting 4. The homogenate was centrifuged at 20,000
for 20 min at 4 C, and the pellet was rehomogenized in 50 mM Tris-HCl pH 7.4 with a Tissue Tearor
×
g
◦
for 10 s at setting 2, followed by recentrifugation. The final pellet was resuspended in 50 mM Tris-HCl
◦
pH 7.4 and frozen in aliquots at
−
80 C. Protein concentration was determined via Pierce BCA protein
assay kit using bovine serum albumin as the standard.
4
.2.2. Binding Affinity
Binding affinities for all test compounds at KOR, MOR, and DOR were determined by competitive
3
displacement of [ H]-diprenorphine as previously reported [35
–
38]. In a 96-well plate format, cell
3
membranes (5–10
µ
g of protein) and [ H]-diprenorphine (0.2 nM) were incubated in Tris-HCl buffer
◦
(
50 mM, pH 7.4) with various concentrations of test compound at 25 C for 1 h, allowing the mixture to
reach equilibrium. Nonspecific binding was determined using the opioid antagonist naloxone (10 µM),

Molecules 2019, 24, 4302
22 of 24
and total binding was determined using vehicle in the absence of competitive ligand. After incubation,
membranes were filtered through Whatman GF/C 1.2 micron glass fiber filters and washed with 50 mM
Tris-HCl buffer. The radioactivity remaining on the filters was then quantified by liquid scintillation
counting after saturation with EcoLume liquid scintillation cocktail in a Microbeta 2450 (Perkin-Elmer,
Waltham, MA, USA). Binding affinity (K ) values were calculated using the Cheng–Prusoff equation
i
via nonlinear regression analysis using GraphPad Prism software from at least three separate binding
assays performed in duplicate.
35
4
.2.3. Stimulation of [ S]-GTPγS Binding
Agonist stimulation of KOR, MOR, and DOR by all test compounds was determined by
S]-guanosine 5 -O-[γ-thio]triphosphate ([ S]-GTPγS) binding assays as previously reported [35–38].
[35
0
35
In a 96-well plate format, membranes from cells expressing opioid receptors as described above (10 µg
35
of protein), [ S]-GTP
γS (0.1 nM), and guanosine diphosphate (30 µM) were incubated in GTPγS buffer
(
50 mM Tris-HCl, 100 mM NaCl, 5 mM MgCl , 1 mM EDTA, pH 7.4) with various concentrations of
2
◦
test compound at 25 C for 1 h. Basal stimulation was determined by incubation in the absence of any
ligand. After incubation, membranes were filtered through Whatman GF/C 1.2 micron glass fiber filters
and washed with GTPγS buffer with no EDTA. The radioactivity remaining on the filters was then
quantified by liquid scintillation counting after saturation with EcoLume liquid scintillation cocktail in
a Perkin-Elmer Microbeta 2450. Data are reported as percent stimulation compared to the effects of
10 µM standard agonists—U69,593 (KOR), DAMGO (MOR), or DPDPE (DOR). Percent stimulation
and EC50 values were determined via nonlinear regression analysis using GraphPad Prism software
from at least three separate assays performed in duplicate. Efficacy is expressed as percent stimulation
relative to standard agonists.
Author Contributions: Conceptualization, D.M. and H.I.M.; methodology, D.M.; validation, D.M. and J.P.A.;
formal analysis, D.M., J.P.A., M.A.B., J.J.T., J.G.H., L.J.D.; investigation, D.M., J.P.A., M.A.B., J.J.T., J.G.H., L.J.D.;
resources, J.R.T., H.I.M.; data curation, J.P.A.; writing—original draft preparation, D.M.; writing—review and
editing, D.M., H.I.M.; visualization, D.M.; supervision, J.R.T., H.I.M.; project administration, D.M., J.P.A.; funding
acquisition, J.R.T., H.I.M.
Funding: This research was funded by NIH, grant number DA003910. Resources were provided by the Edward F.
Domino Research Center. D.M. was supported by an ACS MEDI Pre-doctoral Fellowship and an AFPE Pre-doctoral
Fellowship. J.P.A. was supported by NIH, grant number DA 048129. M.A.B. was supported by the University of
Michigan Interdisciplinary REU grant (NSF DBI #1560096).
Acknowledgments: The authors would like to thank Ashley C. Brinkel for additional in vitro studies.
Conflicts of Interest: The authors declare no conflict of interest. The funders had no role in the design of the
study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to
publish the results.
References
1.
2.
3.
4.
5.
6.
Brownstein, M.J. A brief history of opiates, opioid peptides, and opioid receptors. Proc. Natl. Acad. Sci. USA
993, 90, 5391–5393. [CrossRef] [PubMed]
Levinthal, C.F. Milk of Paradise/Milk of Hell—The History of Ideas about Opium. Perspect. Biol. Med. 1985
8, 561–577. [CrossRef] [PubMed]
Dreborg, S.; Sundstrom, G.; Larsson, T.A.; Larhammar, D. Evolution of vertebrate opioid receptors. Proc.
Natl. Acad. Sci. USA 2008, 105, 15487–15492. [CrossRef] [PubMed]
1
,
2
Kieffer, B.L.; Evans, C.J. Opioid receptors: From binding sites to visible molecules in vivo. Neuropharmacology
2
009, 56, 205–212. [CrossRef]
Erlandson, S.C.; McMahon, C.; Kruse, A.C. Structural Basis for G Protein–Coupled Receptor Signaling. Annu.
Rev. Biophys. 2018, 47, 1–18. [CrossRef]
Weis, W.I.; Kobilka, B.K. The Molecular Basis of G Protein–Coupled Receptor Activation. Annu. Rev. Biochem.
2
018, 87, 897–919. [CrossRef]

Molecules 2019, 24, 4302
23 of 24
7
8
9
1
1
.
.
.
Chavkin, C.; Koob, G.F. Dynorphin, Dysphoria, and Dependence: The Stress of Addiction.
Neuropsychopharmacol. Rev. 2016, 41, 373–374. [CrossRef]
Lalanne, L.; Ayranci, G.; Kieffer, B.L.; Lutz, P.-E. The kappa opioid receptor: From addiction to depression,
and back. Front. Psychiatry 2014, 5, 170. [CrossRef]
Lutz, P.E.; Kieffer, B.L. Opioid receptors: Distinct roles in mood disorders. Trends Neurosci. 2013, 36, 195–206.
[CrossRef]
0. Le Merrer, J.; Becker, J.A.J.; Befort, K.; Kieffer, B.L. Reward Processing by the Opioid System in the Brain.
Physiol. Rev. 2009, 89, 1379–1412. [CrossRef]
1. Dietis, N.; Guerrini, R.; Calo, G.; Salvadori, S.; Rowbotham, D.J.; Lambert, D.G. Simultaneous Targeting
of Multiple Opioid Receptors: A Strategy to Improve Side-effect Profile. Br. J. Anaesth. 2009, 103, 38–49.
[CrossRef]
1
2. Turnaturi, R.; Arico, G.; Ronsisvalle, G.; Parenti, C.; Pasquinucci, L. Multitarget opioid ligands in pain relief:
New players in an old game. Eur. J. Med. Chem. 2016, 108, 211–228. [CrossRef]
1
3. Anand, J.P.; Montgomery, D. Multifunctional opioid ligands. Handb. Exp. Pharmacol. 2018, 247, 21–51.
1
4. Montgomery, D.; Anand, J.P.; Griggs, N.W.; Fernandez, T.J.; Hartman, J.G.; Sánchez-Santiago, A.A.;
Pogozheva, I.D.; Traynor, J.R.; Mosberg, H.I. Novel Dimethyltyrosine–Tetrahydroisoquinoline
Peptidomimetics with Aromatic Tetrahydroisoquinoline Substitutions Show in Vitro Kappa and Mu Opioid
Receptor Agonism. ACS Chem. Neurosci. 2019. [CrossRef]
1
1
5. Balboni, G.; Guerrini, R.; Salvadori, S.; Bianchi, C.; Rizzi, D.; Bryant, S.D.; Lazarus, L.H. Evaluation of the
Dmt-Tic pharmacophore: Conversion of a potent delta-opioid receptor antagonist into a potent delta agonist
and ligands with mixed properties. J. Med. Chem. 2002, 45, 713–720. [CrossRef]
6. Balboni, G.; Salvadori, S.; Trapella, C.; Knapp, B.I.; Bidlack, J.M.; Lazarus, L.H.; Peng, X.;
Neumeyer, J.L. Evolution of the bifunctional lead mu agonist/delta Antagonist containing the
0
0
2
,6 -dimethyl-L-tyrosine-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid (Dmt-Tic) opioid pharmacophore.
ACS Chem. Neurosci. 2010, 1, 155–164. [CrossRef]
1
1
1
2
7. Santagada, V.; Balboni, G.; Caliendo, G.; Guerrini, R.; Salvadori, S.; Bianchi, C.; Bryant, S.D.; Lazarus, L.H.
Assessment of substitution in the second pharmacophore of Dmt-Tic analogues. Bioorg. Med. Chem. Lett.
2
000, 10, 2745–2748. [CrossRef]
8. Balboni, G.; Salvadori, S.; Guerrini, R.; Negri, L.; Giannini, E.; Bryant, S.D.; Jinsmaa, Y.; Lazarus, L.H. Direct
influence of C-terminally substituted amino acids in the Dmt-Tic pharmacophore on delta-opioid receptor
selectivity and antagonism. J. Med. Chem. 2004, 47, 4066–4071. [CrossRef]
9. Pagé, D.; Naismith, A.; Schmidt, R.; Coupal, M.; Labarre, M.; Gosselin, M.; Bellemare, D.; Payza, K.; Brown, W.
Novel C-terminus modifications of the Dmt-Tic motif: A new class of dipeptide analogues showing altered
pharmacological profiles toward the opioid receptors. J. Med. Chem. 2001, 44, 2387–2390. [CrossRef]
0. Pag
Novel Dmt-Tic dipeptide analogues as selective delta-opioid receptor antagonists. Bioorg. Med. Chem. Lett.
000, 10, 167–170. [CrossRef]
é, D.; McClory, A.; Mischki, T.; Schmidt, R.; Butterworth, J.; St-Onge, S.; Labarre, M.; Payza, K.; Brown, W.
2
2
2
2
1. Mello, N.K.; Negus, S.S. Effects of kappa opioid agonists on cocaine- and food-maintained responding by
rhesus monkeys. J. Pharmacol. Exp. Ther. 1998, 286, 812–824. [PubMed]
2. Negus, S.S.; Mello, N.K.; Portoghese, P.S.; Lin, C. Effects of Kappa Opioids on Cocaine Self-Administration
by Rhesus Monkeys. J. Pharmacol. Exp. Ther. 1997, 282, 44–55. [PubMed]
3. Neumeyer, J.L.; Bidlack, J.M.; Zong, R.; Bakthavachalam, V.; Gao, P.; Cohen, D.J.; Negus, S.S.; Mello, N.K.
Synthesis and opioid receptor affinity of morphinan and benzomorphan derivatives: Mixed kappa agonists
and mu agonists/antagonists as potential pharmacotherapeutics for cocaine dependence. J. Med. Chem. 2000
,
4
3, 114–122. [CrossRef] [PubMed]
2
4. Bowen, C.A.; Stevens Negus, S.; Zong, R.; Neumeyer, J.L.; Bidlack, J.M.; Mello, N.K. Effects of Mixed-Action
kappa/mu Opioids on Cocaine Self-Administration and Cocaine Discrimination by Rhesus Monkeys.
Neuropsychopharmacology 2003, 28, 1125–1139. [CrossRef]
2
2
5. Dick, G.R.; Woerly, E.M.; Burke, M.D. A general solution for the 2-pyridyl problem. Angew. Chemie Int. Ed.
2
012, 51, 2667–2672. [CrossRef]
6. Mabrouk, O.S.; Viaro, R.; Volta, M.; Ledonne, A.; Mercuri, N.; Morari, M. Stimulation of delta Opioid Receptor
and Blockade of Nociceptin/Orphanin FQ Receptor Synergistically Attenuate Parkinsonism. J. Neurosci.
2
014, 34, 12953–12962. [CrossRef]

Molecules 2019, 24, 4302
24 of 24
2
2
7. Dripps, I.J.; Jutkiewicz, E.M. Delta Opioid Receptors and Modulation of Mood and Emotion. Handb. Exp.
Pharmacol. 2017, 247, 179–197.
8. Abdelhamid, E.E.; Sultana, M.; Portoghese, P.S.; Takemori, A.E. Selective Blockage of the Delta Opioid
Receptors Prevents the Development of Morphine Tolerance and Dependence in Mice. J. Pharmacol. Exp.
Ther. 1991, 258, 299–301.
2
9. Rozenfeld, R.; Abul-Husn, N.S.; Gomes, I.; Devi, L.A. An Emerging Role for the Delta Opioid Receptor in the
Regulation of Mu Opioid Receptor Function. Sci. World J. 2007, 7, 64–73. [CrossRef]
0. Martin, T.J.; Kim, S.A.; Cannon, D.G.; Sizemore, G.M.; Bian, D.; Porreca, F.; Smith, J.E. Antagonism of
3
0
Delta(2) Opioid Receptors by Naltrindole-5 -isothiocyanate Attenuates Heroin Self-Administration but not
Antinociception in Rats. J. Pharmacol. Exp. Ther. 2000, 294, 975–982.
3
1. Mosberg, H.I.; Yeomans, L.; Harland, A.A.; Bender, A.M.; Sobczyk-Kojiro, K.; Anand, J.P.; Clark, M.J.;
Jutkiewicz, E.M.; Traynor, J.R. Opioid Peptidomimetics: Leads for the Design of Bioavailable Mixed Efficacy
mu Opioid Receptor (MOR) Agonist/delta Opioid Receptor (DOR) Antagonist Ligands. J. Med. Chem. 2013
6, 2139–2149. [CrossRef] [PubMed]
2. Henry, S.P.; Fernandez, T.J.; Anand, J.P.; Griggs, N.W.; Traynor, J.R.; Mosberg, H.I. Structural Simplification
of a Tetrahydroquinoline-Core Peptidomimetic -Opioid Receptor (MOR) Agonist/ -Opioid Receptor (DOR)
Antagonist Produces Improved Metabolic Stability. J. Med. Chem. 2019, 62, 4142–4157. [CrossRef] [PubMed]
3. Nastase, A.F.; Griggs, N.W.; Anand, J.P.; Fernandez, T.J.; Harland, A.A.; Trask, T.J.; Jutkiewicz, E.M.;
Traynor, J.R.; Mosberg, H.I. Synthesis and Pharmacological Evaluation of Novel C-8 Substituted
Tetrahydroquinolines as Balanced-Affinity Mu/Delta Opioid Ligands for the Treatment of Pain. ACS
Chem. Neurosci. 2018, 9, 1840–1848. [CrossRef] [PubMed]
,
5
3
3
µ
δ
3
4. Toll, L.; Berzetei-Gurske, I.P.; Polgar, W.E.; Brandt, S.R.; Adapa, I.D.; Rodriguez, L.; Schwartz, R.W.;
Haggart, D.; O’Brien, A.; White, A.; et al. Standard binding and functional assays related to medications
development division testing for potential cocaine and opiate narcotic treatment medications. NIDA Res.
Monogr. 1998, 178, 440–466. [PubMed]
5. Traynor, J.R.; Nahorski, S.R. Modulation by mu-opioid agonists of guanosine-5 -O-(3-[35S]thio)triphosphate
0
3
binding to membranes from human neuroblastoma SH-SY5Y cells. Mol. Pharmacol. 1995, 47, 848–854.
[PubMed]
3
3
3
6. Lee, K.O.; Akil, H.; Woods, J.H.; Traynor, J.R. Differential binding properties of oripavines at cloned mu- and
delta-opioid receptors. Eur. J. Pharmacol. 1999, 378, 323–330. [CrossRef]
7. Harrison, C.; Traynor, J.R. The [ S]GTPgammaS binding assay: Approaches and applications in
pharmacology. Life Sci. 2003, 74, 489–508. [CrossRef]
8. Harland, A.A.; Bender, A.M.; Griggs, N.W.; Gao, C.; Anand, J.P.; Pogozheva, I.D.; Traynor, J.R.;
Jutkiewicz, E.M.; Mosberg, H.I. Effects of N-Substitutions on the Tetrahydroquinoline (THQ) Core of
35
Mixed-Efficacy
µ-Opioid Receptor (MOR)/δ-Opioid Receptor (DOR) Ligands. J. Med. Chem. 2016, 59,
4
985–4998. [CrossRef]
Sample Availability: Limited samples of most final compounds are available from the authors. Detailed synthetic
procedures for all novel compounds are included in the manuscript.
©
2019 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access
article distributed under the terms and conditions of the Creative Commons Attribution
CC BY) license (http://creativecommons.org/licenses/by/4.0/).
(