Discovery of novel N-sulfonamide-tetrahydroquinolines as potent retinoic acid receptor-related orphan receptor γt inverse agonists for the treatment of autoimmune diseases

Article abstract of DOI:10.1016/j.ejmech.2019.111984

Targeting the nuclear receptor RORγt is thought to be effective in autoimmune disorders. Tertiary sulfonamide 1 was found to be a potent RORγt inverse agonist previously. However, the high hepatic clearance value limits its druggability. In this study, we designed and synthesized a series of N-sulfonamide-tetrahydroquinolines by molecular modeling and scaffold hopping strategy, aiming at improving the metabolic stabilities. Detailed SAR exploration led to identification of potent RORγt inverse agonists such as 13 with moderate binding affinity and inhibitory activity of Th17 cell differentiation. Binding mode of 13 with RORγt-LBD was revealed by molecular docking. Moreover, 13 showed lower intrinsic clearance in mouse liver microsomes compared with 1 and potent in vivo efficacy and safety in psoriasis models, which can be used as a good starting point for the further optimization.

Full text of DOI:10.1016/j.ejmech.2019.111984

European Journal of Medicinal Chemistry 187 (2020) 111984
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Research paper
Discovery of novel N-sulfonamide-tetrahydroquinolines as potent
retinoic acid receptor-related orphan receptor
the treatment of autoimmune diseases
gt inverse agonists for
Nannan Sun 1, Xiaojun Ma 1, Kaifeng Zhou 1, Chen Zhu, Zhonglian Cao, Yonghui Wang,
Jun Xu**, Wei Fu*
School of Pharmacy & Minhang Hospital, Fudan University, Shanghai, 201301, PR China
a r t i c l e i n f o
a b s t r a c t
Article history:
Targeting the nuclear receptor ROR
fonamide 1 was found to be a potent ROR
clearance value limits its druggability. In this study, we designed and synthesized a series of N-sulfon-
amide-tetrahydroquinolines by molecular modeling and scaffold hopping strategy, aiming at improving
g
t is thought to be effective in autoimmune disorders. Tertiary sul-
t inverse agonist previously. However, the high hepatic
Received 23 November 2019
Received in revised form
11 December 2019
Accepted 17 December 2019
Available online 19 December 2019
g
the metabolic stabilities. Detailed SAR exploration led to identification of potent ROR
such as 13 with moderate binding affinity and inhibitory activity of Th17 cell differentiation. Binding
mode of 13 with ROR t-LBD was revealed by molecular docking. Moreover, 13 showed lower intrinsic
gt inverse agonists
g
Keywords:
N-Sulfonamide-tetrahydroquinolines
Retinoic acid receptor-related orphan
clearance in mouse liver microsomes compared with 1 and potent in vivo efficacy and safety in psoriasis
models, which can be used as a good starting point for the further optimization.
© 2019 Elsevier Masson SAS. All rights reserved.
receptor gt (RORgt)
Inverse agonists
Th17 cells
Metabolic stability
Psoriasis
1. Introduction
to reduce the level of IL-17 [8,9]. The success of IL-17 antibodies
(Secukinumab, Ixekizumab, and Brodalumab) and the progress of
The nuclear receptor (NR) retinoic acid receptor-related orphan
receptor t (RORc, also known as NR1F3) is an important tran-
RORgt inverse agonists in clinical trials established that RORgt is a
valuable drug-target for the treatment of autoimmune diseases,
especially for the treatment of psoriasis [10].
g
scription factor which is indispensable for pro-inflammatory
cytokine interleukin IL-17 to be expressed from the T helper
17 cells (Th17) [1,2]. The IL-17 plays a central role in the patho-
genesis of autoimmune diseases as rheumatoid arthritis (RA),
multiple sclerosis (MS), psoriasis, and inflammatory bowel diseases
(IBD). Based on its influence over multiple inflammatory pathways,
Considerable efforts have been directed toward the discovery of
ROR
series, as represented by compound 1 (Fig. 1), attract our attention
with excellent ROR t selectivity and activity in biochemical assay,
gt inverse agonists [11e28]. Among them, tertiary sulfonamide
g
GAL4 human transcription cell assay and human peripheral blood
mononuclear cell (PBMC) assay, besides, the ligand-lipophilicity
efficiency (LLE) of compound 1 is moderate-to-high [12,13,15,18].
RORgt has been established as a suitable drug target [3e7]. Since
RORgt has a constitutive transcriptional activity, discovery of its
inverse agonists, which bind with ROR
activity below the constitutive level, appeared as a promising way
g
t but decrease the receptor
The ROR
cells, where highly lipophilic ligands are easy to reach. This makes
the LLE as an important criteria to evaluate ROR t inverse agonists,
gt is exclusively expressed inside the nucleus of immune
g
for it depends on its measured potency (EC50) and its chemical
lipophilicity (cLogP). However, compound 1 displayed high CLhep
value, as it was completely consumed during 1-h incubation with
either human liver microsomes (HLM) or rat liver microsomes
(RLM), and therefore, the high clearance rate limits compound 1’s
further development. The N-cyclopropyl sulfonamide group
* Corresponding author. School of Pharmacy & Minhang Hospital, Fudan Uni-
versity, Shanghai, 201301, PR China.
** Corresponding author. School of Pharmacy & Minhang Hospital, Fudan Uni-
versity, Shanghai, 201301, PR China.
E-mail addresses: jun_xu@fudan.edu.cn (J. Xu), wfu@fudan.edu.cn (W. Fu).
These authors contributed equally in this work.
1
https://doi.org/10.1016/j.ejmech.2019.111984
0223-5234/© 2019 Elsevier Masson SAS. All rights reserved.

2
N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
Fig. 1. Structures of typical RORgt inverse agonists from the literatures [15e17,25,26].
seemed to be a metabolic soft spot in the structure of 1 and the
major metabolite of 1 was identified to be sulfonamide N-deal-
kylation afterwards, which confirmed our former assumption
[15,16]. Though scientists in Genentech have developed GNE-3500
with favorable potency and greatly improved metabolic stability,
GNE-3500 might be a big challenge for asymmetric synthesis.
In the present study, we aimed to improve the metabolic sta-
core, we designed compounds (11e17) with different linkers to
match the tetrahydroquinoline core, and found that compound 13
is the best of choice. Next, we tested different substituting group on
the aromatic ring at the right hand side (RHS). Among the new
compounds 13e33, we found the p-C(CH3)3 group is the best. At the
fourth round of design, we focused on the N-acetylpiperazine
moiety in the left hand side (LHS) and designed compounds 34e38
to match the best core structure, the linker and the aromatic group
at RHS, expecting to find new compounds with better activities as
bility of sulfonamide derivatives, while maintaining the RORgt in-
verse agonist profile. During the structural design, we also paid full
attention to the important criteria as cLogP, LLE values to evaluate
our new compounds. Using compound 1 as a starting point and
focusing on the metabolic soft spot in tertiary sulfonamide, we
developed a scaffold-hopping strategy to improve the metabolic
stability of tertiary sulfonamide analogues and performing four
rounds of structural design and optimization. Based on the newly
synthesized compounds, we performed activity and metabolic
stability tests on dual fluorescence resonance energy transfer (dual
FRET) assay, Th17 cell differentiation assay, mouse liver micro-
somes (MLM) assay, and a mice psoriasis model testing. Finally, we
developed a strategy to improve the metabolic stability of tertiary
sulfonamide analogues and discovered a novel compound 13 as a
RORgt inverse agonists (Fig. 2).
2.2. Chemical synthesis
At each round of structural design as described above, we syn-
thesized the new sulfonamide derivatives. Here we depicted the
general approach that we used to synthesize a prototypical com-
pound with the dihydroquinoline group as the core structure. The
following Scheme 1 and Scheme 2 illustrate the overall steps. We
followed the way as described in the literature for the synthesis of
compound 1 [12,15], and all the data of compound 1 were provided
in the part of supporting information.
potent RORgt inverse agonist, with good in vivo efficacy and safety
We started to prepare our newly designed compounds by
reacting a series of amine-containing compounds (2a-10a) with
benzenemethanesulfonylchloride and triethylamine in DCM at
room temperature. We generated the sulfamide intermediate
(2be12b) through this reaction. We then coupled the bromo sub-
stituent of 2be12b to a series of N-containing heterocyclic com-
pounds by Buchwald-Hartwig coupling reaction in the presence of
Palladium (II) acetate, 2-dicyclohexylphosphino-20,60-di-i-por-
poxy-1,10-biphenyl and Cs2CO3. So far, we obtained compounds
(2e12) (Scheme 1). As compounds 9a and 10a are not commercially
available, we prepared these two compounds by using 6-bromo-2-
tetralone, which was dissolved in methylsulfonic acid, and then
adding slowly the sodium azide at room temperature to generate
compounds 9c and 10c, and then using 9c and 10c to react with
borane dimethyl sulfide complex solution in THF to generate 9a and
10a, respectively. The preparation of amide compounds 16 and 17 is
in the same way as that for the sulfonamide derivatives, and the
only difference is that we used benzoyl chloride and phenylacetyl
chloride in amide condensation (Scheme 1).
in our mice psoriasis models.
2. Results and discussion
2.1. Compound design
In a try to predict the metabolic property of compound 1 using
the QikProp module of the academic software schrodinger 3.5
€
(https://www.schrodinger.com/qikprop), the N-cyclobutyl group
was pointed out as a possible metabolic site (Fig. S1). When
observing the structure of compound 1, we also doubted its N-
cyclobutyl group to be vulnerable to the incubation with liver mi-
crosomes. Firstly, we designed compounds (2e10) with different
hydro-benzoheterocycle cores, trying to identify a more suitable
core to replace the N-cyclobutyl group. The binding modes of these
designed compounds were confirmed by docking study, showing
the same binding mode as the lead compound (Fig. S2). After
further tested on biochemistry assay, it turned out that compound 8
has the suitable core structure. At the second round of structural
optimization, based on the identified best tetrahydroquinoline
Reagents and conditions: (a) Acyl chloride or sulfonyl chloride,
triethylamine, dichloromethane, rt., 12h, yield: 65.57% for 5a as

N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
3
Fig. 2. General outline for structural design and optimizations based on the lead compound 1. These designs are: targeting the core structure (2e10), the linker group (11e17), the
aromatic ring (18e33), and modifying the acetylpiperazine moiety (34e38).
example; (b) Palladium(II) acetate, 2-dicyclohexylphosphino-20,60-
di-i-porpoxy-1,10-biphenyl, Cs2CO3, toluene, 90 ꢀC, 12h, yield 75%
for 5 as example; (c) NaN3, methylsulfonic acid, rt., 5h, yield 46%;
(d) BH3, tetrahydrofuran, rt., 3h, reflex, 4h, yield 74%.
Reagents and conditions: (a) Chlorosulfonyl isocyanate, tert-
butanol, triethylamine, dichloromethane, rt., 12h, yield: 67%; (b)
CH3I, K2CO3, N, N-dimethylformamide, 110 ꢀC, 6h, yield 64%; (c)
Palladium (II) acetate, 2-dicyclohexylphosphino-20,60-di-i-por-
poxy-1,10-biphenyl, Cs2CO3, toluene, 90 ꢀC, 12h, yield 75% for 9a as
example; (d) TFA, ethyl acetate, rt., 1h, yield 60%; (e) benzyl bro-
mide, K2CO3, N, N-dimethylformamide, rt., 12h, yield 54% for 13 as
example.
tetrahydroquinolin to prepare compound 8c through a novel two-
step one-pot reaction with chlorosulfonyl isocyanate and tert-
butanol in DCM at room temperature. We then treated compound
8c with CH3I, and then with K2CO3 in DMF to finish its methylation
and generate compound 8d. We used the Buchwald-Hartwig
coulping reaction again to couple compound 8d to a series of N-
containing heterocyclic compounds, and generated the corre-
sponding intermediates (e.g., 18a) by removing the tert-butylox-
ycarboryl group at compound 18a in the presence of trifluoroacetic
acid, we generated the N-methyl sulphonyl methane group of these
intermediate compounds (e.g., 18b). We then dissolved the com-
pound 18b in DMF and made it deprotonated with 2.5 equiv. of
K2CO3, and later made it to react with the respective substituted
benzyl bromide at room temperature, so we generated various final
products as the new sulfonamide derivatives.
We started to synthesize our newly designed compounds 13, 14,
15, 18e38 (Scheme 2) from the synthesized compound 8a (as
shown in Scheme 1). In particular, We used the 6-bromo-1,2,3,4-

4
N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
Scheme 1. Syntheses of newly designed sulfonamide derivatives.
Scheme 2. Syntheses of newly designed N-sulfonamide-tetrahydroquinolines derivatives.

N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
5
2.3. Biological evaluation
started to probe the linker structure. As shown in Table 2, com-
pound 11, one carbon atom less in the linker part compared to 8,
2.3.1. Structure-activity relationship
After we synthesized all the newly designed compounds, we
performed a dual FRET assay to measure how good of these com-
displayed ROR
83.5%. Compound 12, with one more carbon atom at the linker
compared to 8, turned to be potent inverse agonist
M). Compound 12 improved the activity, yet it
gt agonist with EC50 of 4.836 mM and efficacy of
a
pounds binding with human ROR
g
t ligand binding domain (LBD),
(IC50 ¼ 0.906
m
in the presence of a coactivator peptide which was derived from
steroid receptor coactivator (SRC)-1. The dual FRET tests were car-
ried out in a similar way as that in our previous studies. Meanwhile,
we also monitored the LLE values of these new sulfonamide de-
rivatives to see whether or not that their LLE values are comparable
to that of compound 1 (LLE ¼ 4.1) [15].
resulted in a reduction in aqueous solubility (clogP ¼ 2.8). We
explored other variants of this functional linker group such as the
sulfanilamide motif. Compound 13 displayed ROR
activity with approximately 9-fold potent than compound 8
(IC50 ¼ 0.218 M), and had a potential improvement in aqueous
solubility (clogP ¼ 2.0). The improved profile (LLE ¼ 4.6) maintains
efficient binding interactions with the ROR t and avoids the high
gt inverse agonist
m
Table 1 depicts the measured IC50 values for compounds 2e10
with different core structures, i.e. benzo-5-member-saturated ring,
benzo-6-member-saturated ring and benzo-7-member-saturated
ring. Among them, compounds 4, 7, 8 and 10 displayed proper
RORgt potency and efficacy. Compound 8 with the tetrahy-
droquinoline core performed better than other ones, with 50-fold
lower potency but good LLE (3.1) and lower clogP value.
g
lipophilicity. 14 and 15 were synthesized in a further exploration of
the length of the sulfanilamide moiety. Compared with 13, com-
pounds 14 and 15 have longer linker structure, but are much less
potent. Besides of sulfanilamide analogues, acyl analogues were
synthesized such as 16 and 17. The activity of acyl analogues
dropped dramatically indicating that acyl motif is not a proper
linker here. In this round of structural optimization, compound 13
The preferred profile of compound 8 led us adopt the tetrahy-
droquinoline as the ideal core structure. Based on compound 8, we
is the most potent RORgt inverse agonist, we studied the binding
mode of 13 (see binding mode study) to guide the follow-up SAR
research.
Table 1
Based on the inverse agonist 13, we wondered what kind of
substituent at the RHS aromatic ring could help to keep or improve
the profile for these newly designed sulfonamide derivatives. As
shown in Table 3, the position of the substituent is critical to the
potency, since shifting from the meta- or ortho-position had a
negative impact, but the substituent at the para-position of the
Structure activity relationships of core changes.
Table 2
cLogPb
LLEc
a
Compd.
Core
RORgt SRC1 IC50
(
m
M) [%eff]
Structure activity relationships of linker changes.
1
2
Lead compound
0.046 [-106.3]
ND
3.2
2.1
4.1
e
3
4
14.04 [-24.6]
46.44 [-71.8]
2.7
2.1
e
2.2
LLEb
2.8
a
Compd.
11
Linker
ROR
g
t SRC1 XC50
(
m
M) [%eff]
cLogP
2.5
5
ND
2.2
e
4.836 [83.5]
0.906 [-55.1]
0.218 [-50.2]
6
7
ND
2.2
2.5
e
12
13
2.8
2.0
3.1
4.6
6.669 [-103.5]
2.7
14
15
0.91 [41.0]
2.8
3.2
3.2
1.9
8
2.048 [-61.6]
2.6
3.1
7.226 [-14.9]
9
ND
2.5
2.5
e
16
17
ND
ND
2.1
2.6
e
e
10
46.05 [-57.6]
1.8
All assay results are reported as the geometric mean of at least two separate runs.
ND means not determined.
All assay results are reported as the geometric mean of at least two separate runs.
ND means not determined.
a
Inhibition of RORgt-LBD recruitment of the SRC1 co-activator peptide. Mean of a
a
minimum of two independent experiments.
Mean of a minimum of two independent experiments, XC50 means IC50 (% max
b
Calculated Log P (cLogP) value.
LLE was calculated as LLE ¼ (ROR
inhibition) or EC50 (% max activation).
c
b
g
t SRC1 pIC50) ꢁ cLogP.
LLE was calculated the same was as that in Table 1.

6
N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
Table 3
Table 4
Structure activity relationships of R1-group substitutions.
Structure activity relationships of R2-group substitutions.
M) [%eff] cLogP/clogDb LLEc
a
Compd. R2
34
ROR
g
t SRC1 IC50
(
m
Compd.
R1
RORg
t SRC1 XC50
(
m
M) [%eff]
cLogP
LLEb
a
0.300 [-25.7]
4.3
2.2
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
o-F
m-F
p-F
o-CH3
m-CH3
p-CH3
o-CF3
m-CF3
p-CF3
o-Cl
m-Cl
p-Cl
p-C(CH3)3
p-Ph
p-OCF3
p-CN
ND
2.2
2.2
2.2
2.5
2.5
2.5
2.9
2.9
2.9
2.8
2.8
2.8
3.8
3.9
3.1
1.5
e
0.324 [-20.7]
1.65 [-90.6]
0.143 [-37.9]
0.768 [34.8]
2.453 [-64.8]
ND
4.3
3.5
3.9
e
35
0.802 [-66.0]
5.1
1.0
3.6
e
36
37
0.284 [-21.1]
0.637 [-25.7]
3.9
7.0
2.6
ND
e
1.555 [-72.4]
0.086 [-25.1]
0.104 [-20.3]
1.135 [-90.0]
0.731 [-93.0]
2.734 [-74.3]
1.034 [-86.6]
7.461 [-81.4]
2.9
e
e
e
3.0
2.3
1.7
2.9
3.6
38
2.006 [-115.0]
3.8
1.9
All assay results are reported as the geometric mean of at least two separate runs.
ND means not determined.
a
Mean of a minimum of two independent experiments, XC50 means IC50 (% max
All assay results are reported as the geometric mean of at least two separate runs.
ND means not determined.
inhibition) or EC50 (% max activation).
b
LLE was calculated the same was as that in Table 1.
a
Mean of a minimum of two independent experiments.
Calculated Log P/Log D (cLogP/cLogD) value.
b
c
LLE was calculated the same was as that in Table 1.
aromatic ring does help, such as compounds 20, 23, 26 and 29.
When the para-position has a larger group as that in compounds
30e33, they maintained strong inhibition efficacy (%eff. as ꢁ93%
Table 5
Results in mouse Th17 cell differentiation assay.
to ꢁ74%) and high potency (e.g., compound 30’s IC50 of 0.73
mM),
but the solubility of 30 (cLogP: 3.8) lead a lower LLE relative to
compound 13.
Compd.
mTh17 IC50
(
m
M)
Th17 %max. inhibition (%)
1
13
0.67
4.93
90
65
At the fourth round of structural design, we tried to replace the
N-acetylpiperazine moiety in the LHS with other substituents
containing hydrogen bond acceptors (Table 4). As shown in Table 4,
changing R2 with a methyl sulfone piperazinyl (34), morpholinyl
(36) and p-methoxycarbonyaniline (37) all resulted in a lost in
administration. Compound 13 showed acceptable PK properties, in
terms of moderate T1/2 (3.3 h) and Tmax. Overall, compound 13
demonstrated improved metabolic stability both in microsomal
and PK study.
RORgt activity. 35 and 38 with proper RORgt inverse agonist ac-
tivity suggested that the Arg367 pocket could tolerate polar group,
but the LLE of compound 38 is still lower than that of compound 13.
Next, we further evaluated the Th17 inhibition activity of the
2.3.3. In vivo efficacy of compound 13
most potent N-sulfonamide-tetrahydroquinolines RORgt inverse
On the basis of the encouraging metabolic stability and potent
Th17 inhibition activity, compound 13 was selected to further test its
in vivo anti-inflammatory activity using the imiquimod (IMQ)-
induced psoriasis mice model. Dexamethasone (DXMS) was chosen
as the reference and positive control. Compound 13 was adminis-
tered intraperitoneally (i.p.) once daily at 4 doses (0.4, 2, 10 and
50 mg/kg) and DXMS was at 5 mg/kg. As in Fig. 3A, the treatment
with compound 13 brought in a significant reduction in clinical
severity of psoriasis in a dose-dependent manner. At the dose of
50 mg/kg and 10 mg/kg, compound 13 had a significant anti-
inflammatory potency as measured through the ear thickness.
Such potency of compound 13 is comparable to that of DXMS at the
dose of 5 mg/kg (Fig. 3A). Besides, the IMQ-induced loss of the body
weight for those mice treated with compound 13 at each of the
different four doses was not so obvious as that of DXMS (Table 7).
This data of body weight loss indicates that compound 13 is much
safer when it is used to treat psoriasis. In addition, compound 13 also
achieved a better inhibition on the erythema and scaling for the back
skinat thedose of0.4 mg/kgcompared with thatof DXMSat the dose
agonist 13. For this purpose, we performed standard mouse
Th17 cell differentiation assay (Table 5). We are pleased to see that
compound 13 has a moderate Th17 cellular inhibition potencies
(IC50 ¼ 4.93
mM) with a maximum inhibition of 65%. The activities
of the compounds tested in the Th17 cellular assay were essentially
consistent with those in the dual FRET assay.
2.3.2. Metabolic stability and PK study
The N-sulfonamide-tetrahydroquinolines were designed to
address the instability of lead compound 1, so we carried out an
in vitro metabolic stability study of compound 13 to test the hy-
pothesis. To our delight, compared to 1, 13 remarkably improved
the metabolic stability and possesses a more favorable clearance
value with Clint(liver) ¼ 47 mL/min/kg, 57.4% remaining at 60 min in
MLM (Table 6). Compound 13 with favorable LLE value and in vitro
microsomal stability profile, was further progressed into single
dose mice experiments to assess its in vivo PK profiles. The PK of
compound 13 investigated in mice following oral (p.o., 50 mg/kg)

N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
7
Table 6
Experimental results of metabolic stability tests.
Compd.
MLM
In mice plasma, compound upon 50 mg , kgꢁ1, p.o.
T1/2 (min)
CLint(mic)
(
mL/min/mg)
CLint(liver) (mL/min/kg)
T1/2 (h)
Tmax (h)
Cmax (ng/mL)
AUC(0-t) (h*ng/mL)
229.3
1
13
2.3
115.5
612
12
2423.5
47.5
3.3
1
102.73
of 5 mg/kg in this study (Figs. 3A and 4 and Fig. S3). We also
compared compound 13 to the clinically immunomodulatory agent
clobetasol topically (Fig. 3B). The IMQ-induced loss of the body
weight for those mice topically treated with compound 13 (4%,1% or
0.25% in cream) is was not so obvious as that of clobetasol. The skin
inflammation of mice that compound 13 topically treated are
weaker than that of the control group and in a dose-dependent
manner too. Taken all these experimental data together, com-
pound 13 has showed reasonable and notable in vivo potency.
of LCeMS and NMR. 1H NMR spectra were recorded with a 400 MHz
Varian spectrometer and 13C NMR spectra were recorded with a
600 MHz Bruker spectrometer. The following abbreviations are
used: br ¼ broad signal, s ¼ singlet, d ¼ doublet, dd ¼ doublet of
doublets, t ¼ triplet, q ¼ quartet, and m ¼ multiplet. Chemical shifts
are expressed in ppm, and coupling constants (J) are expressed in
Hz. All reactions were performed under nitrogen unless otherwise
stated. High resolution mass spectrometry data were given by AB
5600 þ Q TOF. The purity of all the designed compounds was
analyzed using an Agilent 1200 HPLC system with a G1311B qua-
ternary pump, a G1329B ALS and a G4212B DAD detector. The HPLC
method consisted of the following: Agilent C18 RP column
(250 mm ꢂ 4.6 mm, 5 lm); column temperature 25 ꢀC; inject vol-
ume 2.0 lL; HPLC solvent H2O (0.1% TFA)/CH3OH ¼ 60/40 (v/v); flow
rate of 1.2 mL/min; detector wavelength of 254 nm.
2.4. Binding mode study
The binding mode of
a
typical N-sulfonamide-tetrahy-
t-LBD was
droquinoline-based ROR t inverse agonist 13 with ROR
g
g
showed in Fig. 5. The central tetrahydroquinoline ring in the skel-
eton of molecule 13 forms the preferred intermolecular in-
teractions with the side chain of hydrophobic residues Met365,
4.2. Synthesis
Val376, Phe378 and Phe388 in the active site of ROR
benzyl group of 13 resided in the hydrophobic region of the binding
site and formed stacking interactions with His479, Phe486 and
gt-LBD. The
4.2.1. General procedure A for the synthesis of non-
tetrahydroquinoline sulfonamide analogues (2, 3, 4, 5, 6, 7, 8, 9, 10,
11, 12, 16, 17)
p-p
Trp317. In the other side, the ethanoyl moiety of 13 formed a strong
water-mediated hydrogen bond with Arg367 (2.8 Å) and Arg364
(2.8 Å), which was important to the ROR
g
t binding affinity. The
Step 1.7-bromo-1,3,4,5-tetrahydro-2H-benzo[d]azepin-2-one
and 7-bromo-1,2,4,5-tetrahydro-3H-benzo[c]azepin-3-one (9c
and 10c): To a 500 mL round-bottom flask was added 6-bromo-2-
tetralone (10 g, 44.43 mmol) and methanesulfonic acid (47 mL,
72 mmol) at 0 ꢀC. Sodium azide (3.61 g, 55.5 mmol) was slowly
added to the solution and the reaction was stirred at room tem-
perature for 5 h. The reaction mixture was slowly poured into a
2000-mL beaker containing potassium hydroxide (49.8 g,
888 mmol) in water (800 mL) with vigorous stirring. After the
acid was completely quenched, the aqueous solution was
extracted with EtOAc (3 ꢂ 500 mL). The combined organic layers
were washed with saturated aqueous NaHCO3 (100 mL) and
brine (100 mL) respectively, dried over anhydrous MgSO4 and
concentrated to give a mixture of 7-bromo-1,3,4,5-tetrahydro-
2H-benzazepin-2-one and 7-bromo-1,2,4,5-tetrahydro-3H-
benzazepin-3-one (9c and 10c) as brown solid which was used
without purification. LC-MS (ESI): tR ¼ 4.079 min, m/z ¼ 239.9,
241.9 [MþH]þ, m/z ¼ 261.9, 263.9 [MþNa]þ.
binding mode of 13 with ROR t is similar to that of 1 with ROR
g
gt-
LBD. Overlay of 13 with 1 (Fig. 5C) showed that, though there is a
big structure modification on the core part of 1 and 13, both the
benzyl group in the left part and the acetylpiperazine moiety in the
right part almost occupied the same position, adopting the ca-
nonical inverse agonist pose [29,30]. The activity of 1 and 13 in dual
FRET assay was in accord with the binding mode aforementioned.
3. Conclusions
In summary, we have discovered a series of N-sulfonamide-
tetrahydroquinolines as novel RORgt inhibitors displaying favor-
able metabolic stability and potent RORgt inverse agonist activity in
both dual FRET assay and mouse Th17 cell differentiation assay,
suggesting that our strategy to improve the metabolic stability of
tertiary sulfonamide analogues and maintain potent ROR
agonist profile was successful. Notably, partial ROR
agonist 13 stood out with proper clearance and potent activity at
low concentration in mouse psoriasis model. The potent activity in
biochemical and cellular assays, PK profiles, and dose-dependent
inhibition in IMQ-induced mice psoriasis model support 13 could
be a good starting point in further improvements in ROR
potency and physiochemical properties.
gt inverse
gt inverse
Step 2.7-bromo-2,3,4,5-tetrahydro-1H-benzo[d]azepine and 7-
bromo-2,3,4,5- tetrahydro -1H-benzo[c]azepine (9a and 10a):
To a 1000 mL round-bottom flask was added the mixture of 7-
bromo-1,3,4,5-tetrahydro-2H-benzazepin-2-one and 7-bromo-
1,2,4,5-tetrahydro-3H-benzazepin-3-one (9c and 10c) (13 g,
54.2 mmol) and tetrahydrofuran (50 mL). The solution was
cooled to 0 ꢀC and borane-methylsulfide complex (50 mL,
100 M) was slowly added. The reaction mixthre was stirred at
0 ꢀC for 10 min, warmed to room temperature and stirred for 3 h,
then heated at reflux for 4 h. Upon cooling to room temperature,
the mixture was quenched with MeOH (50 mL) to romove
excess borane. The solvent was concentrated under reduced
pressure. The residue was purified by for 2 h. The solution was
then filtered and the filtrate was concentrated under reduced
pressure. The residue was purified by chromatography
(PE:EA ¼ 10:1) and reverse-phase chromatography to give 7-
gt cellular
4. Experimental section
4.1. Materials and methods
General. All the reagents were purchased from commercial
suppliers and used as received. Column chromatography was per-
formed on silica gel (200e300 mesh, Qingdao, China). Melting
point was recorded by WRS-1B digital instrument. Purity and
characterization of compounds were established by a combination

8
N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
Fig. 3. IMQ-induced skin inflammation in mice phenotypically resembles psoriasis. BALB/c mice were treated daily with IMQ cream or vaseline on the shaved back skin and right
ear. Ear thickness of the right ear was measured on the days indicated. Symbols represent mean ear thickness SD or score SD of seven mice per group. Repeated ANOVA, followed
by Dunnett’s Multiple Comparison Test, *p < 0.05, **p < 0.01, ***p < 0.001. (A) Compound 13 was administered intraperitoneally (i.p.) once daily at 4 doses (0.4, 2, 10 and 50 mg/kg)
and DXMS was at 5 mg/kg. Erythema and scaling of the back skin was scored daily on a scale from 0 to 4 according to Psoriasis Area and Severity Index (PASI). The total score is the
sum of ear erythema, back skin erythema and scaliness scales. (B) Compound 13 was topically treated once daily at 3 doses (4%, 1% or 0.1% in cream) and clobetasol propionate cream
was at 0.02%. The body weight is shown in left and the total score (ear erythema, back skin erythema and scaliness scales) is shown in right.

N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
9
Table 7
Experimental results of IMQ-induced psoriasis in vivo study for DXMS and compound 13a.
Compd.
Dosing schedule (mg/kg)
Mice (n) d0/d12
Body weight (mean SD, g)
Body weight change (mean, g)
d0
d12
13
13
13
13
DXMS
control
50 ꢂ 12d
10 ꢂ 12d
2 ꢂ 12d
7/7
7/7
7/7
7/7
7/7
7/7
19.94 0.78
19.00 0.64
19.89 1.16
19.71 0.92
19.86 1.3
19.74 1.64
19.53 1.12
18.11 0.87
19.06 1.43
18.57 1.13
17.19 1.39
17.93 1.30
ꢁ0.41
ꢁ0.89
ꢁ0.83
ꢁ1.14
ꢁ2.67
ꢁ1.81
0.4 ꢂ 12d
5 ꢂ 12d
vehicle ꢂ 12d
a
Data are expressed as mean SEM. P < 0.001.
Fig. 4. IMQ induced psoriasis-like lesions. A, H&E staining of the back skin of mice. (The black arrow highlights hyperkeratosis, the yellow arrow highlights acanthocyte hyperplasia
and the red arrow highlights abscess). B, H&E staining of the right ear of mice. (the black arrow highlights hyperkeratosis, necrosis and abscission, the yellow arrow highlights
acanthocyte hyperplasia and the red arrow highlights inflammatory cell infiltration). (For interpretation of the references to color in this figure legend, the reader is referred to the
Web version of this article.)
Fig. 5. (A) Binding mode of 13 (cyan) within the binding pocket of ROR
gt-LBD; (B) A two-dimensional plot showing the interactions between 13 and the surrounding amino acids.
Hydrogen bonds are indicated by dashed lines between the atoms involved, while hydrophobic contacts are represented by an arc with spokes radiating towards the ligand atoms
they contact; (C) Structural superimposition of 13 (cyan) and 1 (salmon) in the crystal structure of RORgt-LBD. (For interpretation of the references to color in this figure legend, the
reader is referred to the Web version of this article.)

10
N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
bromo-2,3,4,5-tetrahydro-1H-benzoazepine (9a, 1.9 g, 18.9%)
and 7-bromo-2,3,4,5-tetrahydro-1H-benzoazepineand (10a,
1.1 g, 11.0%), Both of 9a and 10a were colorless oil. 9a: 1H NMR
(400 MHz, CDCl3)
d
7.34 (d, J ¼ 3.0 Hz, 5H), 6.89 (d, J ¼ 8.4 Hz, 1H),
6.77 (d, J ¼ 7.6 Hz, 1H), 6.63 (s, 1H), 4.26 (s, 4H), 3.76 (s, 2H), 3.61 (s,
2H), 3.34 (t, J ¼ 5.7 Hz, 2H), 3.17e3.07 (m, 4H), 2.72 (t, J ¼ 5.2 Hz,
2H), 2.13 (s, 3H). LC-MS (ESI): tR ¼ 5.297 min, m/z ¼ 414.3 [MþH]þ,
m/z ¼ 436.2 [MþNa]þ. HRMS (ESIþ) m/z calcd for C22H27N3O3S
[MþH]þ: 414.1846; found: 414.1848.
(400 MHz, CDCl3)
d
7.30 (d, J ¼ 10.4 Hz, 2H), 7.00 (d, J ¼ 7.8 Hz,
1H), 4.20 (s, 1H), 3.62e3.40 (m, 2H), 3.12e2.73 (m, 4H), 2.68 (dd,
J ¼ 21.2, 9.2 Hz, 2H); LC-MS (ESI): tR ¼ 5.175 min, m/z ¼ 226.0,
228.0 [MþH]þ. 10a: 1H NMR (400 MHz, CDCl3)
d 7.33 (d,
J ¼ 8.1 Hz, 2H), 7.09 (d, J ¼ 7.7 Hz, 1H), 4.13 (d, J ¼ 14.7 Hz, 1H),
3.99 (dd, J ¼ 14.7, 9.7 Hz, 1H), 3.58 (d, J ¼ 14.1 Hz, 1H), 3.43 (s,
1H), 3.10e2.92 (m, 2H), 2.83 (dd, J ¼ 14.8, 6.7 Hz, 1H), 2.16e2.05
(m, 1H), 1.73e1.64 (m, 1H); LC-MS (ESI): tR ¼ 5.096 min, m/
z ¼ 226.0, 228.0 [MþH]þ.
4.2.1.5. 1-(4-(2-(benzylsulfonyl)-1,2,3,4-tetrahydroisoquinolin-7-yl)
piperazin-1-yl) ethan-1-one (6). The title compound was obtained
as a colorless oil, 252 mg, yield: 61.0%. 1H NMR (400 MHz, CDCl3)
d
7.33 (t, J ¼ 3.6 Hz, 5H), 7.01 (d, J ¼ 8.4 Hz, 1H), 6.81 (s, 1H),
6.63e6.39 (m, 1H), 4.27 (s, 4H), 3.77 (s, 2H), 3.62 (s, 2H), 3.36 (t,
J ¼ 5.9 Hz, 2H), 3.19e2.99 (m, 4H), 2.70 (t, J ¼ 5.7 Hz, 2H), 2.14 (s,
3H). LC-MS (ESI): tR ¼ 4.988 min, m/z ¼ 413.9 [MþH]þ, m/z ¼ 435.9
[MþNa]þ. HRMS (ESIþ) m/z calcd for C22H27N3O3S [MþH]þ:
414.1846; found: 414.1830.
Step 3: To the solution of amine (2ae10a) (1 mmol) in DCM
(5 mL), triethylamine (2 mmol) was added followed by ben-
zylsulfonyl chloride, 2-Phenylethanesulfonyl chloride, benzene
sulfonyl chloride, benzoyl chloride or phenylacetyl chloride
(1.5 mmol), and the mixture was stirred overnight at room
temperature. The reaction was filtered, concentrated and solved
by EA (20 mL), then washed with ethyl acetate (3 ꢂ 15 mL). The
organic was combined and was dried over Na2SO4. After filtra-
tion, the filtrate was removed in vacuo. The residue was purified
by silica gel column chromatography to give 2be12b, 16be17b.
Step 4: A mixture of Palladium (II) acetate (6 mg, 0.22 mmol) 2-
4.2.1.6. 1-(4-(1-(benzylsulfonyl)-1,2,3,4-tetrahydroquinolin-7-yl)
cyclohexyl)ethan-1-one (7). The title compound was obtained as a
colorless oil, 220 mg, yield: 53.4%. 1H NMR (400 MHz, DMSO)
d
7.37e7.31 (m, 3H), 7.27 (d, J ¼ 3.4 Hz, 2H), 7.07 (d, J ¼ 9.2 Hz, 1H),
6.71 (d, J ¼ 7.8 Hz, 2H), 4.49 (s, 2H), 3.54 (d, J ¼ 4.4 Hz, 4H),
3.47e3.42 (m, 2H), 3.13e3.03 (m, 2H), 3.03e2.94 (m, 2H), 2.66 (t,
J ¼ 6.6 Hz, 2H), 2.02 (s, 3H), 1.78e1.68 (m, 2H). LC-MS (ESI):
tR ¼ 5.01 min, m/z ¼ 413.9 [MþH]þ, m/z ¼ 435.9 [MþNa]þ. HRMS
(ESIþ) m/z calcd for C22H27N3O3S [MþH]þ: 414.1846; found:
414.1840.
dicyclohexylphosphino-20,6’
-di-i-porpoxy-1,10-biphenyl
(22 mg, 0.044 mmol), and cesium carbonate (0.22 g, 0.68 mmol)
were weighted out and the vial was purged with nitrogen, the
bromo-contained intermediate 2be12b, 16be17b (0.44 mmol)
and 1-acetylpiperazine (90 mg, 0.68 mmol) solved in 1,4-
dioxane (15 mL) were added. The mixture was stirred at
100 ꢀC for 16 h. The reaction was filtered, concentrated and
purified by silica gel column chromatography to give 2e12,
16e17.
4.2.1.7. 1-(4-(1-(benzylsulfonyl)-1,2,3,4-tetrahydroquinolin-6-yl)
piperazin-1-yl)ethan-1-one (8). The title compound was obtained
as a colorless oil, 190 mg, yield: 46.1%. 1H NMR (400 MHz, CDCl3)
d
7.42 (d, J ¼ 9.0 Hz, 1H), 7.38e7.28 (m, 3H), 7.21 (dd, J ¼ 7.4, 1.6 Hz,
2H), 6.76 (d, J ¼ 7.7 Hz, 1H), 6.68 (s, 1H), 4.33 (s, 2H), 3.79 (s, 2H),
3.64 (s, 2H), 3.52e3.34 (m, 2H), 3.22e2.98 (m, 4H), 2.64 (t,
J ¼ 6.6 Hz, 2H), 2.14 (s, 3H), 1.90e1.34 (m, 4H). LC-MS (ESI):
tR ¼ 4.990 min, m/z ¼ 413.9 [MþH]þ, m/z ¼ 435.9 [MþNa]þ. HRMS
(ESIþ) m/z calcd for C22H27N3O3S [MþH]þ: 414.1846; found:
414.1835.
4.2.1.1. 1-(4-(1-(benzylsulfonyl)indolin-6-yl)piperazin-1-yl)ethan-1-
one (2). The title compound was obtained as a white solid, 233 mg,
yield: 58.4%; m.p.: 160.7e163.9 ꢀC. 1H NMR (400 MHz, CDCl3)
d 7.30
(dt, J ¼ 14.1, 6.9 Hz, 3H), 7.19 (d, J ¼ 6.7 Hz, 2H), 7.03 (d, J ¼ 8.2 Hz,
1H), 6.94 (d, J ¼ 1.7 Hz, 1H), 6.54 (d, J ¼ 7.4 Hz, 1H), 4.35 (s, 2H),
3.83e3.72 (t, 2H), 3.68e3.58 (m, 4H), 3.20e3.03 (m, 4H), 2.78 (t,
J ¼ 8.5 Hz, 2H), 2.13 (s, 3H). LC-MS (ESI): tR ¼ 4.870 min, m/z ¼ 400.0
[MþH]þ. HRMS (ESIþ) m/z calcd for C21H25N3O3S [MþH]þ:
400.1689; found: 400.1689.
4.2.1.8. 1-(4-(3-(benzylsulfonyl)-2,3,4,5-tetrahydro-1H-benzo[d]aze-
pin-7-yl)piperazin-1-yl)ethan-1-one (9). The title compound was
obtained as a white solid, 53 mg, yield: 12.4%; m.p.: 105.6e107.5 ꢀC.
1H NMR (400 MHz, CDCl3)
d
7.36e7.28 (m, 5H), 6.97 (d, J ¼ 8.2 Hz,
1H), 6.67 (s, 2H), 4.20 (s, 2H), 3.75 (s, 2H), 3.61 (s, 2H), 3.26e3.00
(m, 8H), 2.82 (dd, J ¼ 9.5, 6.8 Hz, 4H), 2.12 (s, 3H). LC-MS (ESI):
tR ¼ 4.949 min, m/z ¼ 428.3 [MþH]þ. HRMS (ESIþ) m/z calcd for
4.2.1.2. 1-(4-(2-(benzylsulfonyl)isoindolin-5-yl)piperazin-1-yl)ethan-
1-one (3). The title compound was obtained as a white solid,
256 mg, yield: 64.2%; m.p.: 159.3e161.4 ꢀC. 1H NMR (400 MHz,
C
23H29N3O3S [MþH]þ: 428.2002; found: 428.2007.
DMSO)
d
7.32 (d, J ¼ 8.4 Hz, 5H), 7.12 (d, J ¼ 8.2 Hz, 1H), 6.92 (d,
J ¼ 8.0 Hz, 1H), 6.85 (s, 1H), 4.54 (s, 2H), 4.47 (d, J ¼ 12.8 Hz, 4H),
3.57 (d, 4H), 3.13 (s, 2H), 3.06 (s, 2H), 2.04 (s, 3H). LC-MS (ESI):
tR ¼ 4.679 min, m/z ¼ 400.0 [MþH]þ. HRMS (ESIþ) m/z calcd for
4.2.1.9. 1-(4-(2-(benzylsulfonyl)-2,3,4,5-tetrahydro-1H-benzo[c]aze-
pin-7-yl)piperazin-1-yl)ethan-1-one (10). The title compound was
obtained as a colorless oil, 35 mg, yield: 8.2%. 1H NMR (400 MHz,
C
21H25N3O3S [MþH]þ: 400.1689; found: 400.1693.
CDCl3)
d
7.32 (d, J ¼ 7.2 Hz, 4H), 7.18 (s, 3H), 6.60 (d, J ¼ 8.4 Hz, 1H),
4.22 (s, 2 H), 4.06 (s, 2H), 3.68 (s, 2H), 3.48 (s, 2H), 3.27 (s, 4H), 2.86
(s, 2H), 2.15 (s, 3H), 1.72 (s, 2H), 1.26 (t, J ¼ 4.9 Hz, 2H). LC-MS (ESI):
tR ¼ 4.945 min, m/z ¼ 428.3 [MþH]þ. HRMS (ESIþ) m/z calcd for
4.2.1.3. 1-(4-(1-(benzylsulfonyl)indolin-5-yl)piperazin-1-yl)ethan-1-
one (4). The title compound was obtained as a colorless oil, 190 mg,
yield: 47.6%. 1H NMR (400 MHz, CDCl3)
d
7.31 (dd, J ¼ 14.2, 6.8 Hz,
C
23H29N3O3S [MþH]þ: 428.2002; found: 428.1999.
4H), 7.20 (d, J ¼ 6.5 Hz, 2H), 6.79 (s, 2H), 4.32 (s, 2H), 3.79 (s, 2H),
3.64 (t, J ¼ 8.5 Hz, 4H), 3.11 (m, 4H), 2.83 (t, J ¼ 8.4 Hz, 2H), 2.15 (s,
3H). LC-MS (ESI): tR ¼ 4.664 min, m/z ¼ 399.9 [MþH]þ, m/z ¼ 421.9
[MþNa]þ. HRMS (ESIþ) m/z calcd for C21H25N3O3S [MþH]þ:
400.1689; found: 400.1669.
4.2.1.10. 1-(4-(1-(phenylsulfonyl)-1,2,3,4-tetrahydroquinolin-6-yl)
piperazin-1-yl) ethan-1-one (11). The title compound was obtained
as a colorless oil, 306 mg, yield: 76.7%. 1H NMR (400 MHz, DMSO)
d
7.65 (dd, J ¼ 8.7, 4.4 Hz, 1H), 7.53 (d, J ¼ 4.2 Hz, 4H), 7.49 (d,
J ¼ 9.0 Hz, 1H), 6.84 (dd, J ¼ 9.0, 2.9 Hz, 1H), 6.63 (d, J ¼ 2.8 Hz, 1H),
3.70 (dd, J ¼ 6.7, 5.4 Hz, 2H), 3.56 (dd, J ¼ 10.1, 5.1 Hz, 4H), 3.17e3.10
(m, 2H), 3.10e3.02 (m, 2H), 2.28 (t, J ¼ 6.7 Hz, 2H), 2.04 (s, 3H), 1.48
4.2.1.4. 1-(4-(2-(benzylsulfonyl)-1,2,3,4-tetrahydroisoquinolin-6-yl)
piperazin-1-yl) ethan-1-one (5). The title compound was obtained
as a white solid, 290 mg, yield: 67.8%; m.p.: 210.7e212.3 ꢀC. 1H NMR
(dd, J ¼ 12.4, 6.3 Hz, 2H). 13C NMR (151 MHz, DMSO)
d 168.09,

N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
11
(250 mL) was added to the mixture and extracted with EA
(3 ꢂ 100 mL). The organic phase was dried over Na2SO4, filtered,
and concentrated under reduced pressure and purified by
chromatography (PE: EA 10:1) to afford the title compound as a
147.85, 138.78, 132.96, 131.59, 129.18, 127.90, 126.56, 125.19, 115.31,
113.81, 54.74, 48.30, 47.94, 45.94, 45.24, 40.46, 25.93, 21.02, 20.73.
LC-MS (ESI): tR ¼ 4.995 min, m/z ¼ 400.0 [MþH]þ, m/z ¼ 422.0
[MþNa]þ. HRMS (ESIþ) m/z calcd for C21H25N3O3S [MþH]þ:
400.1689; found: 400.1688.
light yellow oil 8d (1.4g, 64%). 1H NMR (400 MHz, CDCl3)
d 7.36
(d, J ¼ 8.3 Hz, 1H), 7.22e7.15 (m, 2H), 3.65e3.54 (m, 2H),
2.79e2.72 (m, 3H), 2.69 (d, J ¼ 6.1 Hz, 2H), 1.92 (dd, J ¼ 12.3,
6.3 Hz, 2H), 1.32 (s, 9H); LC-MS (ESI): tR ¼ 5.175 min, m/
z ¼ 426.9, 428.9 [MþNa]þ.
4.2.1.11. 1-(4-(1-(phenethylsulfonyl)-1,2,3,4-tetrahydroquinolin-6-yl)
piperazin-1-yl) ethan-1-one (12). The title compound was obtained
as a white solid, 306 mg, yield: 76.7%; m.p.: 175.3e177.6 ꢀC. 1H NMR
Step 3: A mixture of Palladium(II) acetate (6 mg, 0.22 mmol) 2-
(400 MHz, DMSO)
d
7.36 (d, J ¼ 9.0 Hz, 1H), 7.22 (dd, J ¼ 17.6, 7.0 Hz,
dicyclohexylphosphino-20,6’
-di-i-porpoxy-1,10-biphenyl
5H), 6.77 (d, J ¼ 9.0 Hz, 1H), 6.71 (s, 1H), 3.63 (d, J ¼ 5.4 Hz, 2H), 3.52
(s, 4H), 3.43e3.34 (m, 2H), 3.07 (s, 2H), 3.01 (s, 2H), 2.94e2.83 (m,
(22 mg, 0.044 mmol), and cesium carbonate (0.22 g, 0.68 mmol)
were weighted out and the vial was purged with nitrogen, the
bromo-contained intermediate 8d (0.44 mmol) and a series of
N-containing heterocyclic compounds (0.68 mmol) solved in
1,4-dioxane (15 mL) were added. The mixture was stirred at
100 ꢀC for 16 h. The reaction was filtered, concentrated and
purified by silica gel column chromatography to give 18a-38a.
Step 4: To the solution of intermediate (18a-38a) (1 mmol) in EA
(5 mL), was added trifluoroacetic acid (1 mmol), and the mixture
was stirred at room temperature for 2 h. A mixture of water
(20 mL) and ethyl acetate (15 mL) was added to the residue and
the resulting solution was then basified with NaOH to pH ¼ 10.
The aqueous layers were then washed with ethyl acetate
(2 ꢂ 15 mL). The organic layers were combined and was dried
over Na2SO4, filtered, evaporated, and purified by silica gel col-
umn chromatography to obtain 18b-38b.
Step 5: To the solution of intermediate (18b-38b) (1 mmol) in
N,N-dimethylformamide (5 mL), benzyl bromide, (2-
Bromoethyl) Benzene or 1-Bromo-3-phenylpropane (3 mmol),
and the mixture was stirred at room temperature for 12 h. A
mixture of water (100 mL) and ethyl acetate (30 mL) was added
to the residue and the aqueous layers were then washed with
ethyl acetate (2 ꢂ 15 mL). The organic layers were combined and
was dried over Na2SO4, filtered, evaporated, and purified by
silica gel column chromatography to obtain 13-15, 18-38.
2H), 2.72 (t, J ¼ 6.5 Hz, 2H), 2.01 (s, 3H), 1.88 (d, J ¼ 5.7 Hz, 2H). 13
C
NMR (151 MHz, CDCl3)
d 168.90, 147.65, 137.81, 130.24, 129.88,
128.71, 128.26, 126.81, 123.85, 117.11, 115.35, 53.13, 49.70, 49.40,
46.34, 46.07, 41.19, 29.48, 27.19, 22.60, 21.22. LC-MS (ESI):
tR ¼ 5.287 min, m/z ¼ 428.0 [MþH]þ, m/z ¼ 450.0 [MþNa]þ. HRMS
(ESIþ) m/z calcd for C23H29N3O3S [MþH]þ: 428.2002; found:
428.2010.
4.2.1.12. 1-(4-(1-benzoyl-1,2,3,4-tetrahydroquinolin-6-yl)piperazin-
1-yl)ethan-1-one (16). The title compound was obtained as a yel-
low oil, 170 mg, yield: 46.8%. 1H NMR (400 MHz, DMSO)
d 7.45e7.25
(m, 5H), 6.77 (d, J ¼ 2.7 Hz, 1H), 6.53 (d, J ¼ 6.4 Hz, 2H), 3.71 (t,
J ¼ 6.3 Hz, 2H), 3.53 (dd, J ¼ 10.5, 6.4 Hz, 4H), 3.13e3.04 (m, 2H),
3.04e2.97 (m, 2H), 2.77 (t, J ¼ 6.6 Hz, 2H), 2.51 (dt, J ¼ 3.6, 1.8 Hz,
3H), 1.92 (p, J ¼ 6.5 Hz, 2H). 13C NMR (151 MHz, DMSO)
d 168.07,
147.29, 136.71, 129.53, 127.85, 125.26, 114.89, 113.11, 48.52, 48.13,
45.28, 40.48, 26.45, 23.48, 21.01. LC-MS (ESI): tR ¼ 4.867 min, m/
z ¼ 364.1 [MþH]þ. HRMS (ESIþ) m/z calcd for C22H25N3O2 [MþH]þ:
364.2020; found: 364.2022.
4.2.1.13. 1-(6-(4-acetylpiperazin-1-yl)-3,4-dihydroquinolin-1(2H)-
yl)-2-phenylethan-1-one (17). The title compound was obtained as
a yellow oil, 190 mg, yield: 50.4%. 1H NMR (400 MHz, DMSO)
d 7.18
(dd, J ¼ 30.6, 24.9 Hz, 6H), 6.76 (d, J ¼ 8.0 Hz, 2H), 3.78 (s, 2H), 3.62
(t, J ¼ 5.7 Hz, 2H), 3.53 (s, 4H), 3.06 (d, J ¼ 25.9 Hz, 4H), 2.54 (s, 2H),
2.01 (s, 3H),1.75 (s, 2H). 13C NMR (151 MHz, DMSO)
d 169.52,168.08,
4.2.2.1. 6-(4-acetylpiperazin-1-yl)-N-benzyl-N-methyl-3,4-
dihydroquinoline-1(2H)-sulfonamide (13). The title compound was
obtained as a white solid, 84 mg, yield: 19.0%; m.p.: 213.7e216.1 ꢀC.
128.81, 128.03, 126.13, 124.98, 115.15, 113.34, 48.52, 45.30, 40.50,
26.30, 23.45, 21.02. LC-MS (ESI): tR ¼ 5.114 min, m/z ¼ 378.1
[MþH]þ, m/z ¼ 400.0 [MþNa]þ. HRMS (ESIþ) m/z calcd for
1H NMR (400 MHz, DMSO)
d
7.32 (ddd, J ¼ 20.4, 13.9, 7.9 Hz, 6H),
C
23H27N3O2 [MþH]þ: 378.2176; found: 378.2178.
6.81 (d, J ¼ 8.9 Hz, 1H), 6.74 (s, 1H), 4.29 (s, 2H), 3.65e3.59 (m, 2H),
3.55 (s, 4H), 3.13e3.01 (m, 4H), 2.76 (t, J ¼ 6.6 Hz, 2H), 2.60 (s, 3H),
4.2.2. General procedure A for the synthesis of tetrahydroquinoline
sulfonamide analogues (13e15, 18e38)
2.04 (s, 3H), 1.99e1.89 (m, 2H). 13C NMR (151 MHz, DMSO)
d 167.87,
135.96, 129.97, 129.35, 128.14, 127.72, 123.32, 115.70, 113.87, 53.29,
48.55, 48.17, 46.26, 45.09, 40.28, 33.97, 26.15, 21.49, 20.83. LC-MS
(ESI): tR ¼ 5.351 min, m/z ¼ 443.0 [MþH]þ. HRMS (ESIþ) m/z
calcd for C23H30N4O3S [MþH]þ: 443.2111; found: 443.2109.
Step 1: To a solution of chlorosulfonyl isocyanate (14 g,10 mmol)
in dichloromethane (20 mL) at 0 ꢀC was added dropwise tert-
butanol (0.74 g, 10 mmol). The mixture was stirred at 0 ꢀC for
5 min, then warm to room temperature for 3 h to give the BOC
protected amino-sulfonyl-chloride. The mixture was then
carefully added over 30 min to a solution of 6-bromo-1,2,3,4-
tetrahydroquinoline (2.1 g, 10 mmol) and triethylamine
(4.2 mL, 30 mmol) in DCM (20 mL) at 0 ꢀC. Stirring was
continued without cooling for additional 12 h. The reaction
mixture was concentrated in vacuo, and the residue was dis-
solved in EtOAc (500 mL) and washed with water (200 mL) and
brine (200 mL). The organic layer was dried over MgSO4 and
concentrated under reduced pressure to give 8c which was used
without purification. LC-MS (ESI): m/z ¼ 412.9, 414.9 [MþNa]þ.
4.2.2.2. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-phenethyl-3,4-
dihydroquinoline-1(2H)-sulfonamide (14). The title compound was
obtained as a colorless oil, 100 mg, yield: 21.9%. 1H NMR (400 MHz,
DMSO)
d
7.33e7.19 (m, 6H), 6.76 (d, J ¼ 9.7 Hz, 1H), 6.73 (s, 1H), 3.57
(s, 6H), 3.50 (d, J ¼ 5.3 Hz, 2H), 3.10 (s, 2H), 3.03 (s, 2H), 2.80 (d,
J ¼ 8.0 Hz, 2H), 2.77 (s, 3H), 2.74 (d, J ¼ 6.6 Hz, 2H), 2.05 (s, 3H), 1.89
(d, J ¼ 4.9 Hz, 2H). 13C NMR (151 MHz, DMSO)
d 168.25, 147.24,
138.61, 130.24, 129.82, 128.73, 128.39, 126.30, 123.68, 116.04, 114.22,
51.76, 48.94, 48.57, 46.50, 45.48, 40.68, 34.78, 33.54, 26.55, 21.82,
21.19. LC-MS (ESI): tR ¼ 5.306 min, m/z ¼ 457.3 [MþH]þ, m/z ¼ 479.2
[MþNa]þ. HRMS (ESIþ) m/z calcd for C24H32N4O3S [MþH]þ:
457.2268; found: 457.2272.
Step 2: To
a
solution of tert-butyl ((6-bromo-3,4-
dihydroquinolin-1(2H)-yl)sulfonyl)carbamate (8c) (2.1 g,
5 mmol) in N,N-dimethyl formamide (20 mL) was added methyl
iodide (0.5 mL, 7.5 mmol) and potassium carbonate (1.4 g,
10 mmol). The mixture was heated at 110 ꢀC for 6 h. Water
4.2.2.3. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-(3-phenylpropyl)-
3,4-dihydroquinoline-1(2H)-sulfonamide (15). The title compound

12
N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
was obtained as a colorless oil, 100 mg, yield: 21.9%. 1H NMR
(400 MHz, DMSO) 7.34e7.09 (m, 7H), 6.93e6.62 (m, 2H),
4.2.2.8. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-(3-methylbenzyl)-
3,4-dihydroquinoline-1(2H)-sulfonamide (22). The title compound
was obtained as a colorless oil, 53 mg, yield: 11.6%. 1H NMR
d
3.75e3.48 (m, 8H), 3.10 (dd, J ¼ 16.8, 8.7 Hz, 4H), 2.99 (d, J ¼ 16.3 Hz,
2H), 2.78e2.65 (m, 5H), 2.03 (s, 3H), 1.95e1.81 (m, 2H), 1.74 (dd,
(400 MHz, DMSO)
d
7.32 (d, J ¼ 9.0 Hz, 1H), 7.20 (t, J ¼ 7.5 Hz, 1H),
J ¼ 11.4, 9.6 Hz, 2H). 13C NMR (151 MHz, DMSO)
d
168.11, 129.93,
7.14e6.95 (m, 2H), 6.78 (d, J ¼ 9.2 Hz, 1H), 6.72 (s, 1H), 4.21 (s, 2H),
3.69e3.56 (m, 2H), 3.53 (s, 4H), 3.08 (s, 2H), 3.01 (s, 2H), 2.73 (t,
J ¼ 6.5 Hz, 2H), 2.57 (s, 3H), 2.24 (s, 3H), 2.01 (s, 3H), 1.96e1.79 (m,
129.68, 128.09, 125.65, 123.24, 115.90, 114.02, 49.63, 48.78, 48.40,
46.44, 45.31, 40.52, 34.46, 31.97, 28.56, 26.41, 21.73, 21.01. LC-MS
(ESI): tR ¼ 3.571 min, m/z ¼ 471.3 [MþH]þ, m/z ¼ 493.2 [MþNa]þ.
HRMS (ESIþ) m/z calcd for C25H34N4O3S [MþH]þ: 471.2424; found:
471.2426.
2H). 13C NMR (151 MHz, DMSO)
d 168.15, 147.11, 137.49, 136.05,
130.14, 129.56, 128.50, 128.17, 125.06, 123.42, 115.91, 114.04, 53.45,
48.74, 48.37, 46.53, 45.32, 40.53, 34.20, 26.41, 21.76, 21.03, 20.83.
LC-MS (ESI): tR ¼ 5.543 min, m/z ¼ 457.0 [MþH]þ, m/z ¼ 479.0
[MþNa]þ. HRMS (ESIþ) m/z calcd for C24H32N4O3S [MþH]þ:
457.2268; found: 457.2269.
4.2.2.4. 6-(4-acetylpiperazin-1-yl)-N-(2-fluorobenzyl)-N-methyl-
3,4-dihydroquinoline-1(2H)-sulfonamide (18). The title compound
was obtained as
a
white solid, 90 mg, yield: 19.6%; m.p.:
7.45e7.28 (m, 3H),
155.6e156.8 ꢀC. 1H NMR (400 MHz, DMSO)
d
4.2.2.9. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-(4-methylbenzyl)-
7.22 (dd, J ¼ 13.0, 6.7 Hz, 2H), 6.79 (d, J ¼ 9.1 Hz, 1H), 6.73 (s, 1H),
4.38 (s, 2H), 3.58 (t, J ¼ 9.3 Hz, 6H), 3.10 (d, J ¼ 4.3 Hz, 2H),
3.06e2.96 (m, 2H), 2.76 (t, J ¼ 6.7 Hz, 2H), 2.66 (s, 3H), 2.04 (s, 3H),
3,4-dihydroquinoline-1(2H)-sulfonamide (23). The title compound
was obtained as
a white solid, 70 mg, yield: 15.4%; m.p.:
162.2e165.5 ꢀC. 1H NMR (400 MHz, DMSO)
d
7.31 (d, J ¼ 9.0 Hz, 1H),
1.94 (dd, J ¼ 12.0, 6.1 Hz, 2H). 13C NMR (151 MHz, DMSO)
d
168.10,
7.13 (s, 4H), 6.78 (d, J ¼ 9.1 Hz,1H), 6.71 (s,1H), 4.21 (s, 2H), 3.64e3.45
(m, 6H), 3.11e3.04 (m, 2H), 3.04e2.96 (m, 2H), 2.74 (t, J ¼ 6.6 Hz, 2H),
2.55 (s, 3H), 2.26 (s, 3H), 2.01 (s, 3H), 1.95e1.80 (m, 2H). 13C NMR
147.20, 130.44,130.20, 129.78,129.46,124.42, 123.59,123.02,115.88,
115.28, 114.05, 48.74, 48.37, 47.35, 46.42, 45.30, 40.49, 34.47, 26.31,
21.65, 21.05. LC-MS (ESI): tR ¼ 5.385 min, m/z ¼ 461.0 [MþH]þ, m/
z ¼ 483.0 [MþNa]þ. HRMS (ESIþ) m/z calcd for C23H29FN4O3S
[MþH]þ: 461.2017; found: 461.2019.
(151 MHz, DMSO)
d 168.11, 147.14, 136.67, 133.05, 130.14, 129.58,
128.92,128.01,123.55,115.91,114.08, 53.24, 48.76, 48.39, 46.46, 45.30,
40.49, 34.01, 26.37, 21.69, 21.06, 20.53. LC-MS (ESI): tR ¼ 5.874 min, m/
z ¼ 457.0 [MþH]þ, m/z ¼ 479.0 [MþNa]þ. HRMS (ESIþ) m/z calcd for
4.2.2.5. 6-(4-acetylpiperazin-1-yl)-N-(3-fluorobenzyl)-N-methyl-
3,4-dihydroquinoline-1(2H)-sulfonamide (19). The title compound
C
24H32N4O3S [MþH]þ: 457.2268; found: 457.2264.
was obtained as
a
white solid, 77 mg, yield: 16.7%; m.p.:
7.37 (dd, J ¼ 14.7,
4.2.2.10. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-(2-(trifluoromethyl)
benzyl)-3,4-dihydroquinoline-1(2H)-sulfonamide (24). The title
compound was obtained as a colorlessoil, 81mg, yield:15.9%.1H NMR
144.9e147.6 ꢀC. 1H NMR (400 MHz, DMSO)
d
7.2 Hz, 1H), 7.31 (d, J ¼ 9.0 Hz, 1H), 7.11 (t, J ¼ 6.7 Hz, 2H), 7.02 (d,
J ¼ 10.0 Hz,1H), 6.78 (d, J ¼ 9.0 Hz,1H), 6.71 (s,1H), 4.30 (s, 2H), 3.58
(d, J ¼ 5.0 Hz, 2H), 3.53 (s, 4H), 3.08 (s, 2H), 3.01 (s, 2H), 2.73 (t,
J ¼ 5.8 Hz, 2H), 2.61 (s, 3H), 2.01 (s, 3H), 1.97e1.79 (m, 2H). 13C NMR
(400 MHz, DMSO)
d 7.78e7.61 (m, 2H), 7.58e7.44 (m, 2H), 7.30 (d,
J ¼ 9.0 Hz, 1H), 6.79 (d, J ¼ 9.0 Hz, 1H), 6.72 (s, 1H), 4.47 (s, 2H),
3.63e3.57 (m, 2H), 3.53 (s, 4H), 3.08 (s, 2H), 3.02 (s, 2H), 2.73
(t,J ¼ 6.5 Hz, 2H), 2.68 (s, 3H), 2.01 (s, 3H),1.97e1.88 (m, 2H). 13C NMR
(151 MHz, DMSO)
d 168.07, 147.20, 139.28, 130.38, 130.33,
130.20 129.43, 123.88, 123.42, 115.93, 114.48, 114.08, 52.95, 48.73,
48.37, 46.53, 45.30, 40.49, 34.41, 26.37, 21.73, 21.06. LC-MS (ESI):
tR ¼ 5.352 min, m/z ¼ 461.0 [MþH]þ, m/z ¼ 483.0 [MþNa]þ. HRMS
(ESIþ) m/z calcd for C23H29FN4O3S [MþH]þ: 461.2017;
found:461.2013.
(151 MHz, DMSO) d 168.16, 147.41, 134.88, 132.81, 130.61, 129.28,
128.41,127.84,125.88,123.66,115.91,114.04,50.24,48.73,48.35,46.54,
45.29, 40.49, 35.09, 26.26,21.80,21.02. LC-MS(ESI):tR¼ 5.649min, m/
z ¼ 510.9 [MþH]þ, m/z ¼ 533.0 [MþNa]þ. HRMS (ESIþ) m/z calcd for
C
24H29F3N4O3S [MþH]þ: 511.1985; found: 511.1981.
4.2.2.6. 6-(4-acetylpiperazin-1-yl)-N-(4-fluorobenzyl)-Nmethyl-3,4-
dihydroquinoline-1(2H)-sulfonamide (20). The title compound was
obtained as a white solid, 50 mg, yield: 10.9%; m.p.: 175.2e178.3 ꢀC.
4.2.2.11. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-(3-(trifluoromethyl)
benzyl)-3,4-dihydroquinoline-1(2H)-sulfonamide (25). The title
compound was obtained as a colorless oil, 62 mg, yield: 12.2%. 1H
1H NMR (400 MHz, DMSO)
d
7.36e7.25 (m, 3H), 7.17 (t, J ¼ 8.7 Hz,
NMR (400 MHz, DMSO)
d
7.67 (d, J ¼ 5.1 Hz,1H), 7.62 (s,1H), 7.59 (d,
2H), 6.79 (d, J ¼ 9.1 Hz,1H), 6.75e6.68 (m,1H), 4.27 (s, 2H), 3.58 (dd,
J ¼ 14.8, 8.5 Hz, 7H), 3.09 (s, 2H), 3.03 (s, 2H), 2.75 (t, J ¼ 6.5 Hz, 3H),
2.59 (s, 3H), 2.03 (s, 3H), 1.97e1.78 (m, 2H). 13C NMR (151 MHz,
J ¼ 6.0 Hz, 2H), 7.33 (d, J ¼ 9.0 Hz,1H), 6.79 (d, J ¼ 9.1 Hz,1H), 6.74 (s,
1H), 4.41 (s, 2H), 3.74e3.59 (m, 2H), 3.56 (d, J ¼ 4.0 Hz, 4H),
3.14e3.08 (m, 2H), 3.06e2.96 (m, 2H), 2.76 (t, J ¼ 6.6 Hz, 2H), 2.64
(s, 3H), 2.04 (s, 3H), 2.01e1.82 (m, 2H). 13C NMR (151 MHz, DMSO)
DMSO)
d 168.09, 132.42, 130.21, 130.05, 129.50, 123.55, 115.91,
115.24, 115.08, 114.08, 52.70, 48.74, 48.37, 46.47, 45.30, 43.94, 40.49,
34.15, 26.35, 21.68, 21.06. LC-MS (ESI): tR ¼ 5.645 min, m/z ¼ 461.0
[MþH]þ. HRMS (ESIþ) m/z calcd for C23H29FN4O3S [MþH]þ:
461.2017; found: 461.2021.
d 168.08,147.19,137.93,131.97,130.21,129.43,124.26,123.38,115.94,
114.00, 52.95, 48.69, 48.34, 46.55, 45.31, 40.51, 34.54, 26.36, 21.74,
21.04. LC-MS (ESI): tR ¼ 5.534 min, m/z ¼ 511.0 [MþH]þ, m/
z ¼ 533.0 [MþNa]þ. HRMS (ESIþ) m/z calcd for C24H29F3N4O3S
[MþH]þ: 511.1985; found: 511.1957.
4.2.2.7. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-(2-methylbenzyl)-
3,4-dihydroquinoline-1(2H)-sulfonamide (21). The title compound
was obtained as a colorless oil, 65 mg, yield: 14.2%. 1H NMR
4.2.2.12. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-(4-(trifluoromethyl)
benzyl)-3,4-dihydroquinoline-1(2H)-sulfonamide (26). The title
compound was obtained as a colorless oil, 41 mg, yield: 8.0%. 1H
(400 MHz, DMSO)
d
7.32 (d, J ¼ 8.9 Hz, 1H), 7.18 (d, J ¼ 10.3 Hz, 4H),
6.78 (d, J ¼ 8.9 Hz, 1H), 6.71 (s, 1H), 4.28 (s, 2H), 3.59 (s, 2H), 3.53 (s,
4H), 3.08 (s, 2H), 3.01 (s, 2H), 2.74 (t, J ¼ 6.5 Hz, 2H), 2.56 (s, 3H),
2.23 (s, 3H), 2.01 (s, 3H), 1.92 (s, 2H). 13C NMR (151 MHz, DMSO)
NMR (400 MHz, DMSO)
d
7.70 (s, 2H), 7.48 (d, J ¼ 6.3 Hz, 2H), 7.30
(d, J ¼ 9.0 Hz, 1H), 6.78 (d, J ¼ 8.1 Hz, 1H), 6.71 (s, 1H), 4.39 (s, 2H),
3.59 (s, 2H), 3.53 (s, 4H), 3.08 (s, 2H), 3.02 (s, 2H), 2.74 (s, 2H), 2.63
d
168.13, 147.20, 136.28, 133.83, 130.23, 129.64, 128.36, 127.45,
(s, 3H), 2.01 (s, 3H),1.92 (s, 2H). 13C NMR (151 MHz, DMSO)
d 168.12,
125.74, 123.63, 115.92, 114.07, 51.70, 48.78, 48.42, 46.52, 45.31,
40.51, 34.29, 26.34, 21.82, 21.03, 18.44. LC-MS (ESI): tR ¼ 5.561 min,
m/z ¼ 457.0 [MþH]þ, m/z ¼ 479.0 [MþNa]þ. HRMS (ESIþ) m/z calcd
for C24H32N4O3S [MþH]þ: 457.2268; found: 457.2272.
147.25, 141.31, 130.36, 129.44, 128.51, 125.27, 123.60, 115.90, 114.06,
53.12, 48.71, 48.35, 46.51, 45.31, 34.61, 26.32, 21.72, 21.03. LC-MS
(ESI): tR ¼ 5.627 min, m/z ¼ 511.0 [MþH]þ, m/z ¼ 532.9 [MþNa]þ.
HRMS (ESIþ) m/z calcd for C24H29F3N4O3S [MþH]þ: 511.1985;

N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
13
found: 511.1983.
135.44,130.24,129.59,128.78,128.55,127.32,126.67,126.56,123.55,
115.91, 114.07, 53.26, 48.76, 48.38, 46.52, 45.32, 40.51, 34.32, 26.39,
21.76, 21.03. LC-MS (ESI): tR ¼ 6.018 min, m/z ¼ 519.3 [MþH]þ, m/
z ¼ 541.3 [MþNa]þ. HRMS (ESIþ) m/z calcd for C29H34N4O3S
[MþH]þ: 519.2424; found: 519.2421.
4.2.2.13. 6-(4-acetylpiperazin-1-yl)-N-(2-chlorobenzyl)-N-methyl-
3,4-dihydroquinoline-1(2H)-sulfonamide (27). The title compound
was obtained as a colorless oil, 33 mg, yield: 6.9%. 1H NMR
(400 MHz, DMSO) d 7.49e7.41 (m, 1H), 7.41e7.26 (m, 4H), 6.78 (dd,
J ¼ 9.1, 2.4 Hz, 1H), 6.71 (d, J ¼ 2.2 Hz, 1H), 4.40 (s, 2H), 3.64e3.56
(m, 2H), 3.53 (d, J ¼ 4.8 Hz, 4H), 3.12e3.05 (m, 2H), 3.03e2.95 (m,
2H), 2.73 (t, J ¼ 6.7 Hz, 2H), 2.66 (s, 3H), 2.01 (s, 3H), 1.98e1.78 (m,
4.2.2.18. 6-(4-acetylpiperazin-1-yl)-N-methyl-N-(4-(tri-
fluoromethoxy)benzyl)-3,4-dihydroquinoline-1(2H)-sulfonamide
(32). The title compound was obtained as a colorless oil, 82 mg,
2H). 13C NMR (151 MHz, DMSO)
d
168.13, 147.27, 133.54, 132.33,
yield: 15.6%. 1H NMR (400 MHz, DMSO)
d
7.36 (dt, J ¼ 12.6, 8.9 Hz,
130.36, 129.40, 129.20, 127.29, 123.65, 115.89, 114.05, 51.32, 48.74,
48.36, 46.52, 45.30, 40.50, 34.88, 26.30, 21.74, 21.03. LC-MS (ESI):
tR ¼ 5.693 min, m/z ¼ 477.0 [MþH]þ, m/z ¼ 499.0 [MþNa]þ. HRMS
(ESIþ) m/z calcd for C23H29ClN4O3S [MþH]þ: 477.1722; found:
477.1723.
5H), 6.80 (d, J ¼ 9.1 Hz, 1H), 6.74 (s, 1H), 4.34 (s, 2H), 3.66e3.46 (m,
6H), 3.10 (s, 2H), 3.04 (s, 2H), 2.76 (t, J ¼ 6.6 Hz, 2H), 2.63 (s, 3H),
2.04 (s, 3H), 1.94 (dd, J ¼ 12.0, 6.2 Hz, 2H). 13C NMR (151 MHz,
DMSO)
d 147.56, 147.22, 135.84, 130.30, 129.76, 129.49, 123.55,
120.92,115.90,114.05, 52.77, 48.73, 48.36, 46.49, 45.32, 40.51, 34.43,
26.35, 21.74, 21.02. LC-MS (ESI): tR ¼ 5.710 min, m/z ¼ 527.2
[MþH]þ, m/z ¼ 549.2 [MþNa]þ. HRMS (ESIþ) m/z calcd for
4.2.2.14. 6-(4-acetylpiperazin-1-yl)-N-(3-chlorobenzyl)-N-methyl-
3,4-dihydro-quinoline-1(2H)-sulfonamide (28). The title compound
was obtained as a colorless oil, 63 mg, yield: 13.2%. 1H NMR
C
24H29F3N4O4S [MþH]þ: 527.1934; found: 527.1938.
(400 MHz, DMSO)
d
7.43e7.17 (m, 5H), 6.78 (dd, J ¼ 9.1, 2.5 Hz, 1H),
4.2.2.19. 6-(4-acetylpiperazin-1-yl)-N-(4-cyanobenzyl)-N-methyl-
3,4-dihydroquinoline ꢁ1(2H)-sulfonamide (33). The title compound
was obtained as a colorless oil, 89 mg, yield: 19.1%. 1H NMR
6.72 (s, 1H), 4.28 (s, 2H), 3.64e3.56 (m, 2H), 3.53 (d, J ¼ 4.7 Hz, 4H),
3.14e3.04 (m, 2H), 3.04e2.97 (m, 2H), 2.73 (t, J ¼ 6.7 Hz, 2H), 2.60
(s, 3H), 2.01 (s, 3H), 1.92 (dd, J ¼ 12.0, 6.1 Hz, 2H). 13C NMR
(400 MHz, DMSO)
d
7.85 (d, J ¼ 8.0 Hz, 2H), 7.49 (d, J ¼ 8.0 Hz, 2H),
(151 MHz, DMSO)
d
168.12, 147.18, 138.92, 133.00, 130.23, 129.40,
7.34 (d, J ¼ 8.9 Hz, 1H), 6.81 (d, J ¼ 9.0 Hz, 1H), 6.75 (s, 1H), 4.43 (s,
2H), 3.66e3.54 (m, 6H), 3.12 (s, 2H), 3.06 (s, 2H), 2.78 (t, J ¼ 6.5 Hz,
2H), 2.67 (s, 3H), 2.06 (s, 3H), 2.00e1.87 (m, 2H). 13C NMR (151 MHz,
127.61, 127.42, 126.54, 123.38, 115.93, 114.03, 52.89, 48.71, 48.34,
46.55, 45.31, 40.52, 34.46, 26.37, 21.75, 21.03. LC-MS (ESI):
tR ¼ 5.620 min, m/z ¼ 477.0 [MþH]þ, m/z ¼ 498.9 [MþNa]þ. HRMS
(ESIþ) m/z calcd for C23H29ClN4O3S [MþH]þ: 477.1722; found:
477.1719.
DMSO)
d 168.08, 147.26, 142.31, 132.30, 130.37, 129.39, 128.59,
123.55, 118.55, 115.89, 114.03, 110.19, 53.25, 48.71, 48.34, 46.52,
45.32, 40.52, 34.74, 26.32, 21.75, 21.02. LC-MS (ESI): tR ¼ 4.922 min,
m/z ¼ 468.3 [MþH]þ, m/z ¼ 490.3 [MþNa]þ. HRMS (ESIþ) m/z calcd
for C24H29N5O3S [MþH]þ: 468.2064; found: 468.2062.
4.2.2.15. 6-(4-acetylpiperazin-1-yl)-N-(4-chlorobenzyl)-N-methyl-
3,4-dihydro-quinoline-1(2H)-sulfonamide (29). The title compound
was obtained as
a
white solid, 55 mg, yield: 11.5%; m.p.:
4.2.2.20. N-(4-(tert-butyl)benzyl)-N-methyl-6-(4-(methylsulfonyl)
155.5e158.1 ꢀC. 1H NMR (400 MHz, DMSO)
d
7.38 (d, J ¼ 8.2 Hz, 2H),
piperazin-1-yl)-3,4-dihydroquinoline-1(2H)-sulfonamide
The title compound was obtained as a yellow solid, 41 mg, yield:
7.7%; m.p.: 173.5e174.4 ꢀC. 1H NMR (400 MHz, DMSO)
7.42 (t,
(34).
7.28 (t, J ¼ 8.1 Hz, 3H), 6.77 (d, J ¼ 9.0 Hz, 1H), 6.71 (s, 1H), 4.26 (s,
2H), 3.59e3.51 (m, 6H), 3.07 (s, 2H), 3.01 (s, 2H), 2.73 (t, J ¼ 6.6 Hz,
2H), 2.58 (s, 3H), 2.01 (s, 3H), 1.96e1.79 (m, 2H). 13C NMR (151 MHz,
d
J ¼ 9.2 Hz, 3H), 7.26 (d, J ¼ 8.2 Hz, 2H), 6.89 (dd, J ¼ 9.1, 2.6 Hz, 1H),
6.82 (d, J ¼ 2.3 Hz, 1H), 4.32 (s, 2H), 3.71e3.62 (m, 2H), 3.28 (dd,
J ¼ 13.6, 5.2 Hz, 8H), 2.99 (s, 3H), 2.83 (t, J ¼ 6.6 Hz, 2H), 2.57 (s, 3H),
DMSO)
d 168.21, 147.20, 135.30, 132.06, 130.27, 129.79, 129.45,
128.33, 123.58, 115.90, 114.07, 52.80, 48.72, 48.35, 46.47, 45.30,
40.51, 34.30, 26.31, 21.67, 21.03. LC-MS (ESI): tR ¼ 5.696 min, m/
z ¼ 477.0 [MþH]þ, m/z ¼ 498.9 [MþNa]þ. HRMS (ESIþ) m/z calcd for
2.05e1.94 (m, 2H), 1.33 (s, 9H). 13C NMR (151 MHz, DMSO)
d 146.77,
133.13, 130.18, 129.86, 127.71, 125.10, 123.51, 116.10, 114.30, 53.21,
48.20, 46.48, 45.17, 34.12, 33.70, 30.95, 26.39, 21.73. LC-MS (ESI):
tR ¼ 6.022 min, m/z ¼ 535.0 [MþH]þ. HRMS (ESIþ) m/z calcd for
C
23H29ClN4O3S [MþH]þ: 477.1722; found: 477.1721.
4.2.2.16. 6-(4-acetylpiperazin-1-yl)-N-(4-(tert-butyl)benzyl)-N-
methyl-3,4-dihydro-quinoline-1(2H)-sulfonamide (30). The title
compound was obtained as a white solid, 45 mg, yield: 9.0%; m.p.:
C
26H38N4O4S2 [MþH]þ: 535.2407; found: 535.2410.
4.2.2.21. N-(4-(tertbutyl)benzyl)-6-(4-(ethylsulfonyl)piperazin-1-yl)-
N-methyl-3,4-dihydroquinoline-1(2H)-sulfonamide (35). The title
compound was obtained as a yellow oil, 66 mg, yield: 12.0%; m.p.:
135.2e137.8 ꢀC. 1H NMR (400 MHz, DMSO)
d
7.31 (t, J ¼ 9.2 Hz, 3H),
7.15 (d, J ¼ 8.1 Hz, 2H), 6.76 (d, J ¼ 9.0 Hz, 1H), 6.70 (s, 1H), 4.21 (s,
2H), 3.63e3.42 (m, 6H), 3.06 (s, 2H), 3.00 (s, 2H), 2.72 (t, J ¼ 6.6 Hz,
2H), 2.56 (s, 3H), 2.00 (s, 3H), 1.94e1.81 (m, 2H), 1.22 (s, 9H). 13C
149.4e156.6 ꢀC. 1H NMR (400 MHz, DMSO)
d
7.26 (t, J ¼ 9.2 Hz, 3H),
7.10 (d, J ¼ 8.2 Hz, 2H), 6.73 (dd, J ¼ 9.1, 2.6 Hz, 1H), 6.66 (d,
J ¼ 2.3 Hz, 1H), 4.16 (s, 2H), 3.58e3.44 (m, 2H), 3.13 (d, J ¼ 5.1 Hz,
5H), 3.10 (d, J ¼ 5.4 Hz, 4H), 2.83 (s, 3H), 2.67 (t, J ¼ 6.6 Hz, 2H), 2.50
(s, 3H), 1.85 (dd, J ¼ 11.9, 6.1 Hz, 2H), 1.17 (s, 9H). 13C NMR (151 MHz,
NMR (151 MHz, DMSO)
d 168.10, 149.85, 147.13, 133.13, 130.18,
129.62, 127.70, 125.09, 123.51, 115.91, 114.07, 53.21, 48.77, 48.40,
46.49, 45.32, 40.52, 34.17, 30.95, 26.39, 21.75, 21.02. LC-MS (ESI):
tR ¼ 6.074 min, m/z ¼ 499.3 [MþH]þ, m/z ¼ 521.3 [MþNa]þ. HRMS
(ESIþ) m/z calcd for C27H38N4O3S [MþH]þ: 499.2737; found:
499.2732.
CDCl3)
d 170.54, 150.23, 146.59, 132.22, 130.73, 129.67, 127.49,
124.90, 123.26, 116.77, 114.98, 59.77, 53.53, 50.23, 49.44, 46.58,
45.17, 43.34, 33.87, 30.69, 26.67, 21.90, 20.40, 13.56, 7.17. LC-MS
(ESI): tR ¼ 6.167 min, m/z ¼ 549.0 [MþH]þ, m/z ¼ 571.0 [MþNa]þ.
HRMS (ESIþ) m/z calcd for C27H40N4O4S2 [MþH]þ: 549.2564;
found: 549.2569.
4.2.2.17. N-([1,10-biphenyl]-4-ylmethyl)-6-(4-acetylpiperazin-1-yl)-
N-methyl-3,4-dihydroquinoline-1(2H)-sulfonamide (31). The title
compound was obtained as a colorless oil, 67 mg, yield: 12.9%. 1H
NMR (400 MHz, DMSO)
d
7.72e7.58 (m, 4H), 7.46 (t, J ¼ 7.5 Hz, 2H),
4.2.2.22. N-(4-(tert-butyl)benzyl)-N-methyl-6-morpholino-3,4-
dihydroquinoline-1(2H)-sulfonamide (36). The title compound was
obtained as a colorless oil, 79 mg, yield: 17.3%. 1H NMR (400 MHz,
7.36 (t, J ¼ 7.7 Hz, 4H), 6.81 (d, J ¼ 8.9 Hz, 1H), 6.74 (s, 1H), 4.35 (s,
2H), 3.70e3.59 (m, 2H), 3.56 (d, J ¼ 4.0 Hz, 4H), 3.07 (d, J ¼ 25.2 Hz,
4H), 2.77 (t, J ¼ 6.5 Hz, 2H), 2.65 (s, 3H), 2.04 (s, 3H), 1.99e1.77 (m,
DMSO)
d
7.31 (dd, J ¼ 11.7, 8.7 Hz, 3H), 7.15 (d, J ¼ 8.1 Hz, 2H), 6.75
2H). 13C NMR (151 MHz, DMSO)
d 168.07, 147.17, 139.58, 139.33,
(dd, J ¼ 9.0, 2.4 Hz, 1H), 6.67 (s, 1H), 4.20 (s, 2H), 3.74e3.62 (m, 4H),

14
N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
3.62e3.50 (m, 2H), 3.06e2.95 (m, 4H), 2.72 (t, J ¼ 6.6 Hz, 2H), 2.55
(s, 3H), 2.08e1.71 (m, 2H), 1.22 (s, 9H). 13C NMR (151 MHz, DMSO)
propanesulfonate, 0.1 mg/mL bovine serum albumin, and 10 mM
dithiothreitol in 384-well plates. The total volume was 25 L/well.
m
d
149.82,147.40,133.15,130.12,129.41,127.70,125.10,123.52,115.05,
The europium-labeled SRC1 solution was prepared by adding an
appropriate amount of biotinylated SRC and europium labeled
streptavidin into assay buffer, with final concentrations of 20 and
10 nM, respectively. The allophycocyanin (APC)-labeled-LBD solu-
tion was prepared by adding an appropriate amount of biotinylated
113.30, 65.94, 53.19, 48.49, 46.49, 34.15, 30.95, 26.40, 21.73. LC-MS
(ESI): tR ¼ 6.540 min, m/z ¼ 458.0 [MþH]þ. HRMS (ESIþ) m/z calcd
for C25H35N3O3S [MþH]þ: 458.2472; found: 458.2475.
RORgt-LBD and APC-labeled streptavidin at final concentrations of
4.2.2.23. Methyl
sulfamoyl)-1,2,3,4-tetrahydro-quinolin-6-yl)amino)benzoate
The title compound was obtained as a white solid, 98 mg, yield:
18.8%; mp137.3e140.8 ꢀC. 1H NMR (400 MHz, DMSO)
8.72 (s, 1H),
4-((1-(N-(4-(tert-butyl)benzyl)-N-methyl-
20 nM and 10 nM, respectively. After 15 min of incubation at room
temperature, a 20-fold excess of biotin was added and incubated for
10 min at room temperature to block the remaining free strepta-
vidin. Equal volumes of europium-labeled SRC and APC-labeled
(37).
d
7.80 (d, J ¼ 8.6 Hz, 2H), 7.44 (d, J ¼ 8.7 Hz, 1H), 7.38 (d, J ¼ 8.0 Hz,
1H), 7.21 (d, J ¼ 8.0 Hz, 2H), 7.09e6.94 (m, 4H), 4.29 (s, 2H), 3.79 (s,
3H), 3.67 (s, 2H), 2.80 (t, J ¼ 6.2 Hz, 2H), 2.64 (s, 3H),1.97 (s, 2H),1.27
RORg
t-LBD were then dispensed into 384-well assay plates at
25
m
L volume/well. The 384-well assay plates had 100 L of test
m
(s, 9H). 13C NMR (151 MHz, DMSO)
d 165.87, 149.89, 148.75, 136.93,
compound in DMSO predispensed into each well. The plates were
incubated for 1 h at room temperature and then read on Envision in
LANCE mode configured for europeum-APC labels.
133.08, 131.83, 130.89, 130.42, 127.74, 125.10, 123.47, 120.05, 118.54,
117.89, 113.49, 53.19, 51.27, 46.55, 34.13, 30.94, 26.24, 21.67. LC-MS
(ESI): tR ¼ 6.960 min, m/z ¼ 522.0 [MþH]þ, m/z ¼ 544.0
[MþNa]þ. HRMS (ESIþ) m/z calcd for C29H35N3O4S [MþH]þ:
522.2421; found: 522.2411.
4.3.2. Mouse Th17 differentiation assay
CD4þ T cells were purified from mouse splenocytes using a
commercial CD4þ T cell negative selection kit (Invitrogen). CD4þ
T
4.2.2.24. 4-((1-(N-(4-(tert-butyl)benzyl)-N-methylsulfamoyl)-
1,2,3,4-tetrahydro-quinolin-6-yl)amino)benzoic acid (38). The title
compound was obtained as a colorless oil, 28 mg, yield: 5.5%. 1H
cells were skewed to Th17 cells by culturing cells in the presence of
anti-CD3 (0.25 g/mL, Bioxcel), anti-CD28 (1 g/mL, Bioxcel), anti-
IFN- (2 g/mL, Bioxcel), anti-IL-4 (2 g/mL, Bioxcel), TGF- (5 ng/
m
m
g
m
m
b
NMR (400 MHz, DMSO)
d
8.63 (s, 1H), 7.78 (d, J ¼ 8.0 Hz, 2H), 7.42
mL, Peprotech) and IL-6 (20 ng/mL, Peprotech) for 4 days before
analysis. Compounds or DMSO control were added to the culture on
day 0 of Th17 differentiation at indicated concentrations. Percent-
age of IL-17 production from CD4þ T cells were analyzed by intra-
cellular staining followed by flow cytometry. Dose-response curves
were plotted to determine half-maximal inhibitory concentrations
(IC50) for the compounds using the GraphPad Prism 5 (GraphPad
Software, San Diego CA, USA).
(d, J ¼ 8.7 Hz, 1H), 7.36 (d, J ¼ 7.6 Hz, 2H), 7.20 (d, J ¼ 7.7 Hz, 2H),
7.12e6.92 (m, 4H), 4.28 (s, 2H), 3.65 (s, 2H), 3.18 (s, 1H), 2.79 (t,
J ¼ 6.0 Hz, 2H), 2.63 (s, 3H), 1.96 (s, 2H), 1.26 (s, 9H). 13C NMR
(151 MHz, DMSO)
d 149.87, 148.02, 137.38, 133.09, 131.46, 130.96,
130.42, 127.74, 125.09, 123.52, 119.53, 117.44, 113.58, 34.19, 34.05,
30.94, 26.24, 21.69. LC-MS (ESI): tR ¼ 6.127 min, m/z ¼ 508.0
[MþH]þ, m/z ¼ 530.0 [MþNa]þ. HRMS (ESIþ) m/z calcd for
C
28H33N3O4S [MþH]þ: 508.2265; found: 508.2265.
4.3.3. Metabolic stability studies in mouse liver microsomes
The metabolic stability of tested compounds was evaluated in
pooled CD1 mouse (male) liver microsomes (XenoTech, Charles
River, Raleigh, NC). The incubation mixture (final incubation
4.2.3. The lead compound (1) was prepared in a multistep process
according to the literature procedure of Fauber et al.l. [1]
The title compound was as a colorless oil. 1H NMR (400 MHz,
DMSO)
d
7.37 (s, 5H), 7.16 (d, J ¼ 8.6 Hz, 2H), 6.89 (d, J ¼ 8.7 Hz, 2H),
volume ¼ 400
mL) was prepared in 0.1 M potassium phosphate
buffer (pH 7.4) containing 0.5 mg/mL microsomal protein, 1 mM
5.74 (s, 1H), 4.36 (s, 2H), 4.16 (s, 2H), 4.05 (t, J ¼ 7.6 Hz, 1H), 3.53 (s,
4H), 3.15e2.97 (m, 4H), 2.01 (s, 3H), 1.92 (dd, J ¼ 20.3, 10.4 Hz, 2H),
NADPH, and 1 mM of the tested compounds. The liver microsomes
were pre warmed at 37 ꢀC for 5 min before the compound was
added. Reactions were initiated with the addition of the NADPH
and test compound. The mixtures were incubated at 37 ꢀC for 0, 5,
10, 20, 40, and 60 min. At each time point, the reaction was
1.77 (d,
J
¼
8.1 Hz, 2H), 1.45e1.23 (m, 2H). LC-MS (ESI):
tR ¼ 5.433 min, m/z ¼ 441.6 [MþH]þ, m/z ¼ 442.0. HRMS (ESIþ) m/z
calcd for C24H21N3O3S [MþH]þ: 442.2159; found: 442.2159.
4.3. Biological assays
quenched with 100 mL of methanol. Samples were centrifuged at
2000 g for 10 min. Supernatant (30 mL) was diluted in water (60 mL)
4.3.1. ROR
g
t dual FRET assay
before LCeMS/MS analysis.
The assays were performed in an assay buffer consisting of
50 mM NaF, 50 mM 3-(N-morpholino)propanesulfonic acid, pH 7.4,
Use equation of first order kinetics to calculate T1/2 and Clint(mic)
Equation of first order kinetics:
.
0.05
mM
3-[(3-cholamidopropyl)
dimethylammonio]
Ct ¼ C0 ꢃ eꢁk ꢃt
e
1
when Ct ¼ C0
2
;
Ln2 0:693
T1=2
¼
¼
ke
ke
0:693
1
CLintðmicÞ
¼
ꢃ
In vitro T1=2 mg=mL microsomal protein in reaction system
mg microsomes
g liver
CLintðliverÞ ¼ CLintðmicÞ ꢃ
ꢃ
kg body weight
g liver

N. Sun et al. / European Journal of Medicinal Chemistry 187 (2020) 111984
15
4.3.4. Imiquimod-induced psoriasis study [31,32]
References
For imiquimod-induced psoriasis induction and treatment, fe-
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appropriate groups (seven mice/group). Mice in treatment group
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commercially available IMQ cream (5%) (MINGXINLIDI; SiChuan-
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Appendix A. Supplementary data
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