Rational design and Structure?Activity relationship of coumarin derivatives effective on HIV-1 protease and partially on HIV-1 reverse transcriptase

Article abstract of DOI:10.1016/j.ejmech.2019.111900

Since dual inhibitors may yield lower toxicity and reduce the likelihood of drug resistance, as well as inhibitors of HIV-1 PR and RT constitute the core of chemotherapy for AIDS treatment, we herein designed and synthesized new coumarin derivatives characterized by various linkers that exhibited excellent potency against PR and a weak inhibition of RT. Among which, compounds 6f and 7c inhibited PR with IC₅₀ values of 15.5 and 62.1 nM, respectively, and weakly affected also RT with IC₅₀ values of 241.8 and 188.7 μM, respectively, showing the possibility in the future of developing dual HIV-1 PR/RT inhibitors. Creative stimulation for further research of more potent dual HIV-1 inhibitors was got according to the molecular docking studies.

Full text of DOI:10.1016/j.ejmech.2019.111900

Journal Pre-proof
Rational design and Structure−Activity relationship of coumarin derivatives effective
on HIV-1 protease and partially on HIV-1 reverse transcriptase
Mei Zhu, Ling Ma, Jiajia Wen, Biao Dong, Yujia Wang, Zhen Wang, Jinming Zhou,
Guoning Zhang, Juxian Wang, Ying Guo, Chen Liang, Shan Cen, Yucheng Wang
PII:
S0223-5234(19)31052-9
DOI:
https://doi.org/10.1016/j.ejmech.2019.111900
EJMECH 111900
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 26 August 2019
Revised Date: 18 November 2019
Accepted Date: 18 November 2019
Please cite this article as: M. Zhu, L. Ma, J. Wen, B. Dong, Y. Wang, Z. Wang, J. Zhou, G. Zhang,
J. Wang, Y. Guo, C. Liang, S. Cen, Y. Wang, Rational design and Structure−Activity relationship of
coumarin derivatives effective on HIV-1 protease and partially on HIV-1 reverse transcriptase, European
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Rational Design and Structure−Activity Relationship of Coumarin
Derivatives effective on HIV-1 Protease and partially on HIV-1
a, 1
a, 1
a
a
Reverse Transcriptase Mei Zhu , Ling Ma , Jiajia Wen , Biao Dong , Yujia
a
b
a
a
a
c
Wang , Zhen Wang , Jinming Zhou , Guoning Zhang , Juxian Wang , Ying Guo ,
b
a,*
a,*
Chen Liang , Shan Cen , and Yucheng Wang
a
Institute of Medicinal Biotechnology, Chinese Academy of Medical Science and
Peking Union Medical College, Beijing 100050, China
b
Lady Davis Institute for Medical Research and McGill AIDS Centre, Jewish General
Hospital, Montreal, Quebec, Canada
c
Institute of Materia Medica, Chinese Academy of Medical Science and Peking Union
Medical College, Beijing 100050, China
ABSTRACT: Since dual inhibitors may yield lower toxicity and reduce the
likelihood of drug resistance, as well as inhibitors of HIV-1 PR and RT constitute the
core of chemotherapy for AIDS treatment, we herein designed and synthesized new
coumarin derivatives characterized by various linkers that exhibited excellent potency
against PR and a weak inhibition of RT. Among which, compounds 6f and 7c
inhibited PR with IC values of 15.5 and 62.1 nM, respectively, and weakly affected
5
0
also RT with IC values of 241.8 and 188.7 µM, respectively, showing the possibility
5
0
in the future of developing dual HIV-1 PR/RT inhibitors. Creative stimulation for
further research of more potent dual HIV-1 inhibitors was got according to the
molecular docking studies.
Keywords: coumarin; HIV-1 PR inhibitors; HIV-1 RT inhibitors; pharmacophore
fusion types
*Corresponding Authors
E-mail adresses: shancen@imb.pumc.edu.cn. (Shan Cen), wangyucheng@imb.pumc.edu.cn.(Yucheng Wang).
1
These authors made equal contributions to this work.

1
. Introduction
Highly active antiretroviral therapy (HAART) plays an important role in the
treatment of AIDS patients. Nevertheless, the adverse effects, the risk of drug
interactions, and the emergence of cross-resistant HIV strains brought by HAART
have promoted to discovering new strategy against HIV-1 [1,2]. Dual inhibitors are
single compounds that are able to inhibit two enzyme activities, which could yield
lower toxicity, simplify dosing, and reduce the likelihood of drug resistance. Dual
inhibitors have been reported in different disease arena, such as Alzheimer [3],
Parkinson [4], inflammation [5], cancer [6−8], as well as virologic [9]. Particularly, an
increasing number of dual inhibitors research has been focused on inhibiting HIV-1
replication, such as dual inhibitors targeting HIV-1 integrase (IN) and reverse
transcriptase (RT) or reverse transcriptase ribonuclease H (RNase H) [10−21],
dual-action against HIV-1 CCR5 and integrase [22], or targeting HIV-1 reverse
transcriptase-associated RNase H and RNA-dependent DNA polymerase (RDDP)
functions [23], and so forth. However, research on dual HIV-1 protease (PR) and
reverse transcriptase inhibitors is inappreciable [24,25], and dual inhibitor is still not
available for clinical use to treat AIDS.
As inhibitors of HIV-1 PR and RT constitute the core of chemotherapy for AIDS
treatment, and dual therapy with a PR inhibitor and RT inhibitors has been confirmed
to be safe and effective [26], our attention has been given to the design and synthesis
of dual inhibitors of HIV-1 PR and RT. HIV-1 PR is responsible for the production of
all viral enzymes and structural proteins necessary to produce mature, virulent virions
[
27]. PR is a homodimer of two 99 amino acid subunits, and two flexible

glycine-dense β-sheets form a flap region over the top of the active site [28,29]. RT is
a 117 kDa heterodimer consisting of p66 and p51 subunits [30], and is responsible for
the conversion of single-stranded viral RNA into double-stranded proviral DNA.
Given that hydrogen bonding interactions are indispensable for both PR and RT to
combining with inhibitors [30−32], it is possible that two scaffolds of RT and PR
inhibitors are merged into one scaffold with dual activities, according to the Designed
Multifunctional Ligands (DMLs) [33] or “portmanteau inhibitors” [21]. In previous
studies, both naturally occurring and synthetic coumarin derivatives show potent RT
inhibition activity, such as Calanolide A (1) [34], fesumtuorin A (2) [35], and 4r (3) in
Figure 1 [36]. Inspired by the above, we introduced the coumarin moieties into the
HIV-1 protease inhibitors, and designed new inhibitors of pharmacophore fusion
types, which contained a coumarin moiety as novel P2 ligands coupled with the
nonpeptide PIs structural scaffolds containing hydroxyethylsulfonamide isosteres
(Figure 2), aiming at obtaining RT inhibition and keeping excellent potency against
PR at the same time.
Figure 1. Structures of coumarin derivatives active against RT.

Figure 2. Design of portmanteau inhibitors against RT and PR, combining coumarin moieties 4
with PR inhibitor 5.
2
. Results and discussion
2
.1.2.1. Chemistry
Synthesis of amine derivatives. Compounds 9-11 were prepared from the
commercially available material (2S, 3S)-1,2-epoxy-3-(boc-amino)-4-phenylbutane
12), as reported in the literature and shown in Scheme 1 [37,38].
(
Scheme 1. Syntheses of Amines 9-11. Reagents and conditions: (a) i-BuNH , CH CN, 80 °C, 6 h;
2
3
(b) Aryl sulfonyl chloride, DIEA, DMAP(Cat.), THF, 0 °C ~ r.t, 3-5 h; (c) CH Cl -CF COOH
2 2 3
(
1:1), 0 °C~r.t, 3 h; (d) H (gas), 50 psi, 10% Pd/C, CH OH, r.t, 2 h.
2
3
Synthesis of target compounds-amide isosteres. The syntheses of inhibitors 6a-8i

shown in Scheme 2 were carried out by coupling coumarin acids 4a-d with amines
-11 under an EDCI/HOBt/DMAP-mediated coupling method [38].
9
Scheme 2. Syntheses of Inhibitors 6a-8i. Reagents and conditions: (e) EDCI, HOBt, DMAP,
anhydrous DMF, Argon, 0 °C~r.t, 3 h.
Synthesis of target compounds-carbamate isosteres. The syntheses of inhibitors
6
j-7k shown in Scheme 3 were synthesized by hydroxycoumarins 4j, 4k and amines
, 10 under the condensing of bis(trichloromethyl) caebonate (BTC) undergoing an
9
one-pot reaction [39].

Ra
O
O
O
+
+
S
Cl CO
3
OCCl3
H N
N
2
O
O
OH
OH
Rc
4
j, 4k
16
9, 10
Ra
O
O
O
f
S
O
N
H
N
O
O
OH
Rc
6j-6k, 7j-7k
Scheme 3. Syntheses of inhibitors 6j-7k. Reagents and conditions: (f) DIEA, anhydrous DCM,
anhydrous THF, 0 °C ~ r.t, 1.5 h.
Synthesis of target compounds-amine isosteres. The syntheses of inhibitors 6-8l
shown in Scheme 4 were synthesized by chlorocoumarins 4l and amines 9-11 by
catalyzed of DIEA under the condition of refluxing [40].
Scheme 4. Syntheses of inhibitors 6-8l. Reagents and conditions: (g) DIEA, anhydrous EtOH,
reflux, 7 h.
2
.2. Anti-PR activity assay
All newly synthesized compounds were tested in vitro PR activity assays against
HIV-1 wild-type protease using a fluorescence resonance energy transfer (FRET)
method [41,42], and the results are summarized in Tables 1, 2, and 3. Among these
compounds, amide derivatives were very potent PR inhibitors, showing IC values in
5
0

the nanomolar range (298.4−0.40 nM), with the exception of 7h and 7i, which
exhibited higher IC values of 557 nM and 563 nM. In particularly, compound 8a
5
0
with
2-oxo-2H-chromene-6-carboxamide
as
the
P2
ligand
and
a
4
-aminophenylsulfonamide as the P2' ligand exhibited four-fold activity with IC50
value of 0.40 nM compared to the reference compound Darunavir (DRV). This might
account for the exposure of both carbonyl and oxygen atom, which could form
strengthen hydrogen bonding interactions with the backbone atoms and residues in the
protease S2 subsite [43]. Meanwhile, compound 6e demonstrated comparable potency
as DRV with IC value of 1.62 nM. Carboxamide substitution in the 3-position led to
5
0
a decrease in the activity (6b−8d, IC50 value of 298.4−27.7 nM), compared with
substitution in the 6-position (6a−8a, IC value of 2.60−0.40 nM). The rigid and
5
0
bulkier hindrance might impose restrictions on the ability of hydrogen to bond with
the protease S2 subsite. But the introducing of hydroxy in the 7-positon showed better
inhibitory activity (6c−8c) than the substitution of hydrogen atoms (6b−8b). It caused
slightly decreasing activity when hydroxy was replaced by methoxyl group (6d−8d).
Acetamide substitution in the 4-position showed activity against PR with IC50
values in the low nanomolar range (55.8−1.62 nM), with the exception of massive
substitution in position 7 (6g−8g, IC50 value of 211.3−75.1 nM), but the shift of
acetamide substitution from position 4 to 3 gave an decreased activity (6h−8i, with
the IC value range of 563−95.4 nM). Just as reported previously, phenylsulfonamide
5
0
derivatives with 4-methoxy (6a−i) and 4-amino (7a−i) groups displayed generally
higher potency than the corresponding substituted compounds with 4-nitro (8a−i)
groups [43−45].
However, carbamate isosteres decreased in the activity, with IC50 values in the
range of 1.53−0.12 µM, as well as phenylsulfonamide derivatives with 4-methoxy
(6j−k) groups displayed generally higher potency than the corresponding substituted
compounds with 4-nitro (7j−k) groups by an order of magnitude. While amine
derivatives showed increasing activity (6l−8l, IC value of 143.5−53.0 nM).
5
0
In brief, amide derivatives exhibited higher potency than those of carbamate and
amine isosteres, which might be due to the conformational restrictions.

On the basis of these results above, selected inhibitors were further evaluated in
cell-based assays. In the assay, virus-producing cells were treated with the compounds
at 10 µM, and the infectivity of the resultant virus was determined, which reflects the
effect of the compounds on the late stage of HIV-1 life cycle including viral
maturation [46]. In particularly, the most active compounds 6a−8a were equipotent as
DRV. Notably, compound 6e showed a slightly decreased antiviral activity (82.94%
inhibition) compared with DRV, although equipotent in biochemical assays in vitro.
While, the amine compounds 6l and 7l exhibited satisfactory active in cell-based
assays with inhibition of 88.80% and 73.17%, respectively. The results might suggest
a correlation between the activity in vitro and in vivo, as well as excellent cell
membrance penetration, as shown in Tables 1 and 3.
To further confirm anti-HIV activity of these compounds, we next investigated
their effect on the growth of wild type HIV-1 in SupT1 cells. Notably, inhibitor 6e
exhibited a promising inhibition ratio of 45.54% against wild-type HIV-1 at a
concentration of 100 nM, compared with the reference compound Nevirapine (NVP)
with 77% inhibition, which was worthy for in-depth study [47].
2
.3. Anti-RT activity assay
The amide derivatives were tested for RT inhibition using in vitro RT-activity assay
[
48], and the results were listed in Table 1. Unfortunately, all the compounds showed
limited potency compared with the reference compound Efavirenz (EFV). Next, we
analyzed antiviral activity of all the compounds in HIV-1 infected cells using a one
round infectivity assay, which was calculated as a percentage of inhibition at a
concentration of 10 µM [18]. Likewise, most of the compounds showed poor
inhibitory effect on the early step of viral replication including reverse transcription of
viral genome by RT.
Surprisingly, compound 8b was confirmed as a weak inhibitor, with IC value of
5
0
7
5.25 µM in in vitro RT activity assay. And 6f and 7c exhibited IC value of 241.8
50
and 188.7 µM, respectively.

Taken together, these results suggest that carboxamide substitution in the 3-position
with 7-hydoxy or 7-methoxy groups led to a decrease in the activity of RT, compared
with unsubstituted compounds in the 7-position, such as 8b vs 8c, and 8d. The rigid
and bulkier hindrance might impose restrictions on the ability of this kind of
derivatives to bind with the reverse transcriptase. In addition, acetamide substitution
in the 4-position with biggish substitution in position 7 showed better activity against
RT than those with minor substitution in the 7-position (6f, 6g vs 6e). The shift of
acetamide substitution from position 4 to 3 led to a decrease in the activity, but
compounds with 7-hydoxy groups displayed higher potency than those with 7-amino
groups (8h vs 8i).
2
.4. Evaluation of biological activity
To better rationalize structure−activity relationships (SARs) of all the amide isosteres,
the IC values in the inhibition of both PR and RT enzyme were plotted against each
5
0
other in correlation plots (Figure 3). The compounds are distributed around two
perpendicular axes crossing the PR IC axis (X axis) at 150 nM and the RT IC axis
5
0
50
(Y axis) at 250 µM (bolded crosshair in the center of each graph). These two axes
spliced the graph into four quarters corresponding to PR/RT dual inhibitors (lower left
quarter), RT selective inhibitors (lower right quarter), PR selective inhibitors (upper
left quarter), and inhibitors of lower potency (upper right quarter). However, these
graphs do not show any particular correlation between RT and PR inhibition. As can
be seen in Figure 3, most of the compounds are distributed in the left half of the
graph, suggesting of critical for PR inhibition but not critical for RT inhibition, which
confirms that the amide compounds are selective PR inhibitors. Taking a
comprehensive view of the activity above, amide isosteres 6f and 7c emerged
possibility of developing dual HIV-1 PR/RT inhibitors, which should be optimized
further.

Figure 3. Scatter plot for the inhibition of RT and PR enzymes. (A) Compounds are categorized
a
according to the nature of their R substitution. (B) Compounds are categorized according to the
b
c
nature of their R substitution. (C) Compounds are categorized according to the nature of their R
d
substitution. (D) Compounds are categorized according to the nature of their R substitution.
Compounds with IC values against PR above 300 nM and IC values against RT above 500 µM
5
0
50
have been arbitrary positioned at the 300 nM and 500 µM value, respectively.
Table 1. Antiviral and Enzymatic Activity of Amide Compounds 6a−8i
O
O
linker
O
O
S
N
N
Ra
H
OH
Rc
Rd
PR
RT
a
c
d
compd.
R
linker
R
R
Inhibi
IC50
Inhibitio
IC50
B
A,F
B
tion
(nM)
n (%)
(µM)
A,
(
%)
E
6a
7a
8a
6b
7b
8b
6c
H
H
6-CO OCH3
H
H
H
H
H
H
H
99.53
98.54
98.72
1.84± 0.53
2.60± 0.40
0.40± 0.11
72.84±7.63
298.4±41.5
155.9±18.7
39.5±16.9
1.26
-
597.4± 5.1
D
6-CO
6-CO
NO2
NH2
nd
H
5.61
44.78
27.06
1.34
12.32
398± 2.6
473.8± 2.5
nd
C
H
3-CO OCH3
-
H
3-CO
3-CO
NO2
NH2
-
-
-
H
75.3±2.5
420.7± 0
7-OH
3-CO OCH3

7
8
6
7
8
6
c
c
d
d
d
e
7-OH
7-OH
3-CO
3-CO
NO2
NH2
H
H
H
H
H
H
73.99
62.1±36.2
27.7±16.9
46.7±16.0
220.9±145.8
102.1±53.9
1.62±0.55
91.04
27.04
8.28
0.45
23.24
-
188.7± 0
554.4± 2.5
370.4± 0
nd
-
-
-
7-OCH3 3-CO OCH3
7-OCH3 3-CO
7-OCH3 3-CO
NO2
NH2
705.7±2.4
458.6±2.6
7-OH
7-OH
7-OH
4-CH OCH3
2CO
82.94
7
e
e
4-CH
2CO
NO2
H
H
H
H
H
H
H
H
73.36
11.1±4.97
37.9±15.3
15.5±2.75
55.8±14.9
15.4±6.89
211.3±84.7
275.6±219.6
75.1±39.1
109.2±57.0
557±194.1
150.6±36.1
222.3±63.5
563±193.1
95.4±56.6
1.72± 0.73
3.91
-
367.0±2.1
nd
8
4-CH
2CO
NH2
17.94
6
7
8
f
f
f
7-OCH3 4-CH OCH3
2CO
-
-
-
-
-
-
-
-
-
-
-
17.72
43.72
7.72
241.8± 2.5
372.8± 7.6
410.6± 0
355.1± 0
nd
7-OCH3 4-CH
2CO
NO2
7-OCH3 4-CH
2CO
NH2
6g
7g
8g
6h
7h
8h
7-N(CH 4-CH OCH3
3)2 2CO
7-N(CH 4-CH
3)2 2CO
7-N(CH 4-CH
43.75
14.65
18.37
31.91
34.41
23.51
19.90
24.80
11.26
NO2
NH2
572.2± 0
nd
3)2
2CO
3-CH OCH3 4-C
7-OH
2CO
H3
4-C
H3
7-OH
7-OH
3-CH
NO2
NH2
375.4± 5.00
450.8± 0
nd
2CO
3-CH
4-C
H3
2CO
6
7
8
i
i
i
7-NH2
7-NH2
7-NH2
3-CH OCH3 4-C
2CO
H3
4-C
H3
3-CH
NO2
NH2
nd
2CO
3-CH
4-C 30.77
H3
nd
2CO
DRV
EFV
99.97
0.091±
0
.008
A
B
All assays were conducted in quadruplicate or triplicate.
IC , half maximal inhibitory concentration. All assays were conducted in triplicate, and the data shown represent
50
mean values (±1 standard deviation) derived from the results of three independent experiments.
C
-: not determined, the inhibition was 0.
D
E
nd: not determined, the value of IC50 exceeding 1000 µM.
the inhibitory effect on virus-producing cell at 10 µM

F
the inhibitory effect on virus-infected cells at 10 µM
Table 2. Enzymatic Activity of Carbamate Compounds 6j−7k
a
c
compd.
R
R
PR IC5
RT IC
50
0
A
A
(µM)
(µM)
B
6
7
j
j
4-CH3
4-CH3
OCH3
NO2
0.16±0.02
1.02±0.56
nd
nd
nd
nd
6
k
k
6,7-CH=CHO
6,7-CH=CHO
OCH3
NO2
0.12±0.02
7
1.53±0.59
DRV
EFV
0.00057±0.00017
0.091± 0.008
A
IC , half maximal inhibitory concentration. All assays were conducted in triplicate, and the data shown represent
50
mean values (±1 standard deviation) derived from the results of three independent experiments.
B
nd: not determined, the value of IC50 exceeding 1000 µM.
Table 3. Antiviral and Enzymatic Activity of Amine Compounds 6l−8l
PR
A
RT
IC50
c
compd.
R
Inhibition (%)
IC50
B
B
(
µM)
(µM)
C
6l
7l
8l
OCH3
NO2
88.80
73.17
-
0.053± 0.02
0.14± 0.06
0.11± 0.05
nd
nd
nd
NH2
DRV
EFV
99.97
0.0017±0.0007
0.091± 0.008
A
B
All assays were conducted in quadruplicate. The inhibitory effect on virus-producing cell at 10 µM.
IC , half maximal inhibitory concentration. All assays were conducted in triplicate, and the data shown represent
50
mean values (±1 standard deviation) derived from the results of three independent experiments.
C
nd: not determined, the value of IC50 exceeding 1000 µM.
2
.5. Molecular modeling studies

To explore the antiviral action of the inhibitors, molecular docking studies were
performed on inhibitor 7e as a representative example, with highly potent activity
against PR as well as less active RT inhibition. The common mode of inhibitory
binding was explored through molecular docking using a HIV PR crystal structure
(PDB-ID: 4mc9) [49], and RT crystal structure (PDB-ID: 2yng) [50]. Significantly,
the inhibitor 7e fits perfectly into the PR binding site through hydrogen bonds and van
der Waals interactions. As can be seen, several hydrogen-bonding interactions are
possible between the residues Arg8 and Gly48, as well as van der Waals interactions
with the outer enzyme atoms, both of which might account for the promising HIV-1
PR inhibitory activity (Figure
4). While, compound 7e with moiety of
2
-oxo-2H-chromen could not be situated right in the narrow pocket of RT by as much
as approximately thirty thousand-fold in contrast with PR potency. Albeit
unsatisfactory, hydrogen-bonding interaction between the residues Lys223 may
provide the weak activity against RT. Overall, in order to better rationalize the activity
of this kind of dual inhibitors, the coumarin moieties for sterically hindering fitting of
the RT active site, could exert their RT inhibitory activity by eliminating the bulkier
furanone ring and replacing by the miniature linker, such as a flexible chain, which
would not affect the activity against PR meanwhile, according to the molecular
docking studies.
Figure 4. (a) Binding mode of compound 7e within the HIV-1 PR model. (b) Binding mode of
compound 7e within the HIV-1 RT model. Ligand exposures are represented as purple spheres.
Amino acid side chains important for the ligand binding are depicted as blue arrows.

2
.6. Correlation for amide isosteres as HIV PR
Further validation was achieved by correlation of the SAR of docked inhibitors 6a,
a, 7b, 8b, 8c, 7e, 6g, 8g and 8h, as shown in Figure 5. The correlation observed
7
between these two sets of IC data (expt vs calcd, correlation coefficiency = 0.86)
5
0
supports our docking model with a common mode of binding as a valid platform for
PR inhibitor design. However, inhibitors should be optimized further for activity
against RT.
Correlation for Amide isosteres as HIV PR
-
-
-
-
-
-
11.5
12.0
12.5
13.0
13.5
2
R = 0.8608
14.0
-
3.0
-2.5
-2.0
-1.5
-1.0
-0.5
0.0
-
PR log(1/IC50) (exp)
Figure 5. Strong correlation of docked amide isosteres analogues supports a common mode of
binding for HIV PR
3
. Conclusion
Since inhibitors of HIV-1 PR and RT constitute the core of chemotherapy for AIDS
treatment, and dual inhibitors might yield lower toxicity and reduce drug resistance,
our attention has been given to the design, synthesis and biological assays of dual
inhibitors of HIV-1 PR and RT. In this paper, we describe a series of new coumarin
derivatives that are characterized by various linkers. All the compounds were tested in
HIV-1 PR and RT activity assay. Amide derivatives were very active PR inhibitors,
showing IC values in the nanomolar range (298.4−0.40 nM), with the exception of
5
0
7
h and 7i (557 nM and 563 nM, respectively), conferring better activity against
carbamate and amine isosteres. In particularly, compound 8a with
-oxo-2H-chromene-6-carboxamide as the P2 ligand and
2
a

4
-aminophenylsulfonamide as the P2' ligand exhibited four-fold activity with IC50
value 0.40 nM compared to clinically available PR inhibitor DRV, and the most active
compounds 6a−8a were equipotent as DRV in cell-based assays with 99.53−98.72%
inhibition. Notably, inhibitor 6e demonstrated comparable potency as DRV with IC50
value of 1.62 nM, and exhibited a promising inhibition ratio of 54.46% against
wild-type HIV-1 at a concentration of 100 nM.
However, these compounds showed poor inhibition activity against RT if compared
to their anti-PR potency. Among which, compound 8b was confirmed as a weak RT
inhibitor, with IC value of 75.25 µM. Compounds 6f and 7c inhibited PR with IC
5
0
50
values of 15.5 and 62.1 nM, respectively, and also weakly affected RT with IC50
values of 241.8 and 188.7 µM, respectively, showing the possibility in the future of
developing dual HIV-1 PR/RT inhibitors. According to the molecular docking studies,
the 2-oxo-2H-chromen of coumarin moieties might be steric hindrance for fitting into
the narrow pocket of RT active site. Eliminating the bulkier furanone ring and
replacing by a flexible, miniature linker may exert their RT inhibitory activity and not
affect the activity against PR meanwhile.
In conclusion, the design of new coumarin derivatives confirm the possibility of
developing dual HIV-1 PR/RT inhibitors and give information for further
development of effective dual HIV-1 inhibitors.
4
. Experimental section
4
.1. Chemistry
All experiments requiring anhydrous conditions were conducted in flame-dried
glassware fitted with rubber septa under a positive pressure of dry argon, unless
otherwise noted. THF was distilled under argon from sodium-benzophenone ketyl and
CH Cl was distilled under argon from calcium hydride. All reactions were monitored
2
2
by thin-layer chromatography on silica gel plates (GF-254) and visualized with the
®
UV light. Flash column chromatography was performed on a CombiFlash Rf 200

system employing silica gel (50-75 µm, Qingdao Haiyang Chemical Co.,Ltd). Melting
points were taken on MP70 Melting Point System with revised. High resolution mass
1
13
spectra were obtained on an Autospee Ultima-TOF spectrometer. H NMR and C
NMR spectra were recorded in CDCl , CD OD, (CD ) CO or DMSO-d on a Bruker
3
3
3 2
6
AVANCE III 400 MHz, 500 MHz or 600 MHz spectrometer (Bruker Inc) with
tetramethylsilane (TMS) as an internal reference. The chemical shifts are given in δ
1
13
(
ppm) referenced to the respective solvent peak (CDCl : H, δ = 7.26 ppm, C, δ =
3
1
13
1
7
7.16 ppm; CD OD: H, δ = 3.31 ppm, C, δ = 49.00 ppm; (CD ) CO: H, δ = 2.05
3 3 2
1
3
1
13
ppm, C, δ = 30, 205 ppm; DMSO-d : H, δ = 2.49 ppm, C, δ = 39.5 ppm), and
6
coupling constants are reported in Hz. All the target compounds were characterized
1
13
by H and C NMRs and HRMS spectra.
4
.1.1.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-methoxyphenyl)sulfonamido)-1-phenylbutan-2
-yl)-2-oxo-2H-chromene-6-carboxamide (6a)
N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDCI, 0.029 g,
0
.15 mmol) and 1-hydroxybenzotriazole (HOBt, 0.015 g, 0.11 mmol) were
sequentially added in batches to stirring solution of
-oxo-2H-chromene-6-carboxylic acid (4a, 0.019 g, 0.10 mmol) and
N-((2R,3S)-3-amino-2-hydroxy-4-phenylbutyl)-N-isobutyl-4-
a
2
methoxybenzenesulfonamide (9, 0.043 g, 0.105 mmol) in dry DMF (1 mL) at 0 °C
under an argon atmosphere. The reaction mixture was stirred for 10 min at 0 °C and
then additional 1 hour at room temperature. 4-Dimethylaminopyridine (DMAP,
0
.0024 g, 0.020 mmol) was added and the reaction mixture was stirred for another 2
hours at room temperature. The solvent was removed under reduced pressure. Water
4 mL) was added to the residue and extracted with ethyl acetate (3 × 4 mL). The
combined organic phases were dried over Na SO , and evaporated, in vacuo. The
(
2
4
residue was purified by chromatography on a silica gel column (30 × 6 cm). Elution
with 1:1 to 2:3 hexanes-ethyl acetate gave 6a as pale yellow crystalline powder: yield
1
0
.050 g (87%); mp 118-120 °C; H NMR (400 MHz, CD OD) δ 7.97 (s, 1H), 7.92 (d,
3

J = 9.6 Hz, 1H), 7.84 (d, J = 8.6 Hz, 1H), 7.66 (d, J = 8.0 Hz, 2H), 7.35 (d, J = 8.6 Hz,
H), 7.28 (d, J = 7.4 Hz, 2H), 7.21 (t, J = 7.4 Hz, 2H), 7.12 (t, J = 7.2 Hz, 1H), 6.93
d, J = 8.0 Hz, 2H), 6.47 (d, J = 9.6 Hz, 1H), 4.25 (dd, J = 9.4, 4.3 Hz, 1H), 4.00 –
1
(
3
3
1
.96 (m, 1H), 3.80 (s, 3H), 3.46 (d, J = 15.0 Hz, 1H), 3.10 (dd, J = 13.3, 8.5 Hz, 1H),
.00 (d, J = 13.4 Hz, 1H), 2.95 – 2.91 (m, 1H), 2.83 – 2.77 (m, 2H), 2.08 – 1.99 (m,
1
3
H), 0.92 (d, J = 6.4 Hz, 3H), 0.85 (d, J = 6.4 Hz, 3H); C NMR (101 MHz, CD OD)
3
δ 168.4, 164.9, 164.5, 161.9, 157.1, 145.1, 140.2, 132.3, 131.8, 130.6, 130.3, 129.3,
1
2
28.8, 127.3, 120.0, 118.1, 117.7, 115.3, 74.7, 59.1, 56.2, 56.1, 54.4, 36.6, 28.0, 20.5,
-
0.4; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 577.2015, found 577.2024.
3
1
34
2
7
4
.1.2.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-nitrophenyl)sulfonamido)-1-phenylbutan-2-yl)
-2-oxo-2H-chromene-6-carboxamide (7a)
The title compound was obtained by 4a which was coupled with 10 through
EDCI/HOBt/DMAP coupling procedure in 78% yield (white powder) as described for
1
6
a: mp 178-180 °C; H NMR (400 MHz, CD OD) δ 8.27 (d, J = 8.0 Hz, 2H), 7.99 (d,
3
J = 8.0 Hz, 2H), 7.93 (d, J = 9.6 Hz, 1H), 7.87 (s, 1H), 7.83 (d, J = 9.6 Hz, 1H), 7.36
d, J = 8.6 Hz, 1H), 7.27 – 7.19 (m, 4H), 7.12 (t, J = 7.2 Hz, 1H), 6.48 (d, J = 9.6 Hz,
H), 4.21 – 4.19 (m, 1H), 3.92 (t, J = 7.6 Hz, 1H), 3.49 (d, J = 15.0 Hz, 1H), 3.24 (dd,
J = 13.4, 10.2 Hz, 2H), 3.15 (dd, J = 15.0, 8.9 Hz, 1H), 2.98 – 2.95 (m, 1H), 2.82 –
(
1
2
.75 (m, 1H), 2.10 – 2.00 (m, 1H), 0.92 (d, J = 6.4 Hz, 3H), 0.86 (d, J = 6.4 Hz, 3H);
1
3
C NMR (101 MHz, CD OD) δ 168.5, 161.9, 157.1, 151.4, 146.7, 145.0, 140.1,
3
1
5
5
32.2, 131.7, 130.3, 129.8, 129.4, 128.9, 127.3, 125.3, 120.1, 118.2, 117.8, 73.8, 58.1,
-
6.3, 53.5, 36.6, 27.8, 20.4, 20.3; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ):
3
0
31
3
8
92.1766, found 592.1772.
4
.1.3.
N-((2S,3R)-4-((4-amino-N-isobutylphenyl)sulfonamido)-3-hydroxy-1-phenylbutan-2-y
l)-2-oxo-2H-chromene-6-carboxamide (8a)
The title compound was obtained by 4a which was coupled with 11 through

EDCI/HOBt/DMAP coupling procedure in 84% yield (pale yellow powder) as
1
described for 6a: mp 155-157 °C; H NMR (400 MHz, CD3OD) δ 7.97 (brs, 2H),
7
.91 (d, J = 9.6 Hz, 1H), 7.84 (d, J = 8.6 Hz, 1H), 7.39 (d, J = 8.0 Hz, 2H), 7.35 (d, J
8.6 Hz, 1H), 7.28 (d, J = 8.0 Hz, 2H), 7.21 (t, J = 7.3 Hz, 2H), 7.12 (t, J = 7.2 Hz,
H), 6.56 (d, J = 8.0 Hz, 2H), 6.46 (d, J = 9.6 Hz, 1H), 4.26 (t, J = 9.2 Hz, 1H), 3.99
t, J = 7.6 Hz, 1H), 3.43 (d, J = 15.0 Hz, 1H), 3.04 (dd, J = 13.3, 8.6 Hz, 2H), 2.91 –
.83 (m, 1H), 2.80 – 2.73 (m, 2H), 2.07 – 1.97 (m, 1H), 0.93 (d, J = 6.4 Hz, 3H), 0.86
=
1
(
2
1
3
(
d, J = 6.4 Hz, 3H); C NMR (101 MHz, CD OD) δ 168.5, 164.9, 157.0, 154.3, 145.1,
3
1
40.3, 132.4, 131.8, 130.4, 130.3, 129.3, 128.8, 127.2, 125.6, 120., 118.1, 117.7,
114.3, 74.9, 59.5, 56.2, 54.7, 36.9, 28.1, 20.6, 20.5; HRMS (ESI) m/z calcd. for
-
C H N O S ([M - H] ): 562.2018, found 562.2043.
30
33
3
6
4
.1.4.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-methoxyphenyl)sulfonamido)-1-phenylbutan-2
-yl)-2-oxo-2H-chromene-3-carboxamide (6b)
The title compound was obtained by 4b which was coupled with 9 through
EDCI/HOBt/DMAP coupling procedure in 98% yield (yellow oil) as described for 6a:
1
mp 152-155 °C; H NMR (500 MHz, CDCl ) δ 8.94 (d, J = 8.3 Hz, 1H), 8.79 (s, 1H),
3
7.74 (d, J = 8.3 Hz, 2H), 7.67 (dd, J = 15.4, 7.9 Hz, 2H), 7.39 (dd, J = 15.8, 8.1 Hz,
2H), 7.29 – 7.26 (m, 4H), 7.20 (d, J = 5.7 Hz, 1H), 6.98 (d, J = 8.3 Hz, 2H), 4.37 –
4.35 (m, 1H), 3.98- 3.97 (m, 1H), 3.86 (s, 3H), 3.25 – 3.17 (m, 2H), 3.11 (d, J = 15.0
Hz, 1H), 3.02- 2.97 (m, 2H), 2.85 (dd, J = 13.3, 6.7 Hz, 1H), 1.86 (dt, J = 13.5, 6.6 Hz,
1
3
1
H), 0.92 (d, J = 6.3 Hz, 3H), 0.86 (d, J = 6.3 Hz, 3H); C NMR (126 MHz, CDCl )
3
δ 162.9, 161.8, 161.1, 154.4, 148.6, 137.5, 134.3, 130.1, 129.8, 129.5, 129.46, 128.5,
1
26.5, 125.3, 118.5, 117.8, 116.7, 114.3, 72.5, 58.5, 55.6, 54.8, 53.4, 35.9, 27.1, 20.1,
-
1
9.9; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 577.2003, found 577.1974.
3
1
34
2
7
4
.1.5.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-nitrophenyl)sulfonamido)-1-phenylbutan-2-yl)
-2-oxo-2H-chromene-3-carboxamide (7b)
The title compound was obtained by 4b which was coupled with 10 through

EDCI/HOBt/DMAP coupling procedure in 96% yield (pale yellow powder) as
1
described for 6a: mp 201-204 °C; H NMR (500 MHz, CD OD) δ 8.71 (s, 1H), 8.34
3
(
d, J = 8.6 Hz, 2H), 8.04 (d, J = 8.6 Hz, 2H), 7.81 (d, J = 7.6 Hz, 1H), 7.75 (t, J = 7.8
Hz, 1H), 7.45 – 7.42 (m, 2H), 7.26 (d, J = 7.2 Hz, 2H), 7.22 (t, J = 7.4 Hz, 2H), 7.14
t, J = 7.0 Hz, 1H), 4.29 (dt, J = 9.5, 5.0 Hz, 1H), 3.81 – 3.78 (m, 1H), 3.54 (dd, J =
(
1
1
1
5.0, 3.0 Hz, 1H), 3.26 – 3.23 (m, 1H), 3.20 – 3.17 (m, 1H), 3.13 (dd, J = 13.9, 4.0 Hz,
H), 3.03 (dd, J = 13.6, 7.0 Hz, 1H), 2.82 (dd, J = 13.9, 9.8 Hz, 1H), 2.07 – 2.01 (m,
1
3
H), 0.93 (d, J = 6.5 Hz, 3H), 0.87 (d, J = 6.5 Hz, 3H); C NMR (151 MHz, CD OD)
3
δ 163.3, 162.5, 155.9, 149.5, 147.0, 144.2, 139.2, 135.7, 131.3, 130.6, 129.8, 129.4,
1
27.5, 126.6, 125.4, 119.8, 119.3, 117.5, 72.5, 57.8, 55.6, 52.8, 36.2, 27.7, 20.3;
-
HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 592.1748, found 592.1739.
3
0
31
3
8
4
.1.6.
N-((2S,3R)-4-((4-amino-N-isobutylphenyl)sulfonamido)-3-hydroxy-1-phenylbutan-2-y
l)-2-oxo-2H-chromene-3-carboxamide (8b)
The title compound was obtained by 4b which was coupled with 11 through
EDCI/HOBt/DMAP coupling procedure in 22% yield (yellow granular solid) as
1
described for 6a: mp 177-179 °C; H NMR (500 MHz, CD OD) δ 8.69 (s, 1H), 7.79
3
(
d, J = 7.5 Hz, 1H), 7.73 (t, J = 7.7 Hz, 1H), 7.60 (d, J = 8.5 Hz, 2H), 7.44 – 7.40 (m,
H), 7.28 (d, J = 7.5 Hz, 2H), 7.22 (t, J = 7.5 Hz, 2H), 7.14 (t, J = 7.5 Hz, 1H), 6.94
d, J = 8.5 Hz, 2H), 4.36 (dt, J = 9.5, 5.0 Hz, 1H), 3.94 – 3.91 (m, 1H), 3.45 (dd, J =
2
(
1
2
6
1
5.0, 3.5 Hz, 1H), 3.21 (dd, J = 13.9, 3.5 Hz, 1H), 3.06 (dd, J = 13.5, 8.0 Hz, 1H),
.95 (dd, J = 15.0, 8.0 Hz, 1H), 2.84 – 2.77 (m, 2H), 2.01 – 1.96 (m, 1H), 0.92 (d, J =
13
.5 Hz, 3H), 0.86 (d, J = 6.5 Hz, 3H); C NMR (151 MHz, CD OD) δ 163.2, 162.5,
3
57.1, 155.8, 149.5, 139.6, 135.6, 131.3, 130.6, 129.8, 129.7, 129.3, 127.4, 126.5,
119.9, 119.2, 117.5, 113.2, 73.5, 59.1, 55.6 54.0, 36.3, 28.1, 20.6, 20.5; HRMS (ESI)
-
m/z calcd. for C H N O S ([M - H] ): 562.2018, found 562.2040.
3
0
33
3
6
4
7
.1.7.
-Hydroxy-N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-methoxyphenyl)sulfonamido)-1-ph

enylbutan-2-yl)-2-oxo-2H-chromene-3-carboxamide (6c)
The title compound was obtained by 4c which was coupled with 9 through
EDCI/HOBt/DMAP coupling procedure in 98% yield (pale yellow oil) as described
1
for 6a: mp 188-190 °C; H NMR (400 MHz, CD OD) δ 8.58 (s, 1H), 7.69 (d, J = 8.0
3
Hz, 2H), 7.59 (d, J = 8.0 Hz, 1H), 7.26 – 7.18 (m, 4H), 7.12 (t, J = 7.2 Hz, 1H), 6.96
(
d, J = 8.0 Hz, 2H), 6.85 (d, J = 8.6 Hz, 1H), 6.75 (s, 1H), 4.32 – 4.29 (m, 1H), 3.89 –
.85 (m, 1H), 3.41 (dd, J = 15.0, 3.2 Hz, 1H), 3.17 (dd, J = 13.8, 3.2 Hz, 1H), 3.09
dd, J = 13.5, 8.0 Hz, 1H), 2.98 – 2.93 (m, 1H), 2.87 – 2.74 (m, 2H), 2.03 – 1.96 (m,
3
(
1
3
1
H), 0.91 (d, J = 6.6 Hz, 3H), 0.85 (d, J = 6.6 Hz, 3H); C NMR (101 MHz, CD OD)
3
δ 165.8, 164.8, 164.5, 164.0, 158.2, 149.8, 139.5, 1323.0, 131.9, 130.6, 130.5, 129.3,
1
2
5
27.4, 115.8, 115.3, 114.1, 114.0, 112.7, 103.1, 73.4, 58.8, 56.1, 55.5, 53.8, 36.9, 28.0,
-
0.5, 20.4; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 593.1964, found
3
1
34
2
8
93.1955.
4
7
.1.8.
-Hydroxy-N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-nitrophenyl)sulfonamido)-1-phenyl
butan-2-yl)-2-oxo-2H-chromene-3-carboxamide (7c)
The title compound was obtained by 4c which was coupled with 10 through
EDCI/HOBt/DMAP coupling procedure in 94% yield (yellow crystalline powder) as
1
described for 6a: mp 252-254 °C; H NMR (400 MHz, CD OD) δ 8.59 (s, 1H), 8.30
3
(d, J = 8.2 Hz, 2H), 8.00 (d, J = 8.2 Hz, 2H), 7.60 (d, J = 8.6 Hz, 1H), 7.23 – 7.17 (m,
4H), 7.11 (t, J = 6.8 Hz, 1H), 6.85 (d, J = 8.6 Hz, 1H), 6.74 (s, 1H), 4.30 – 4.19 (m,
1H), 3.76 – 3.74 (m, 1H), 3.51 (d, J = 15.0 Hz, 1H), 3.25 – 3.14 (m, 2H), 3.09 (dd, J =
13.9, 4.0 Hz, 1H), 3.04 – 3.00 (m, 1H), 2.82 – 2.76 (m, 1H), 2.07 – 1.97 (m, 1H), 0.91
1
3
(
d, J = 6.4 Hz, 3H), 0.85 (d, J = 6.4 Hz, 3H); C NMR (101 MHz, CD OD) δ 165.9,
3
1
1
64.9, 164.0, 158.3, 151.4, 149.9, 146.9, 139.2, 133.0, 130.6, 129.8, 129.3, 127.5,
25.4, 115.8, 114.0, 112.7, 103.1, 72.6, 57.7, 55.4, 52.8, 36.9, 27.7, 20.3; HRMS (ESI)
-
m/z calcd. for C H N O S ([M - H] ): 608.1709, found 608.1700.
3
0
31
3
9
4
.1.9.

N-((2S,3R)-4-((4-amino-N-isobutylphenyl)sulfonamido)-3-hydroxy-1-phenylbutan-2-y
l)-7-hydroxy-2-oxo-2H-chromene-3-carboxamide (8c)
The title compound was obtained by 4c which was coupled with 11 through
EDCI/HOBt/DMAP coupling procedure in 88% yield (pale yellow oil) as described
1
for 6a: mp 267-270 °C; H NMR (400 MHz, CD OD) δ 8.58 (s, 1H), 7.59 (d, J = 8.6
3
Hz, 1H), 7.43 (d, J = 8.0 Hz, 2H), 7.26 – 7.17 (m, 4H), 7.11 (t, J = 7.0 Hz, 1H), 6.84
(d, J = 8.6 Hz, 1H), 6.74 (s, 1H), 6.61 (d, J = 8.0 Hz, 2H), 4.35 – 4.31 (m, 1H), 3.87 (t,
J = 8.6 Hz, 1H), 3.39 (dd, J = 14.9, 3.6 Hz, 1H), 3.17 (dd, J = 13.9, 3.8 Hz, 1H), 3.03
–
1
2.99 (m, 1H), 2.90 (dd, J = 15.0, 8.0 Hz, 1H), 2.80 – 2.72 (m, 2H), 1.99 - 1.93 (m,
1
3
H), 0.90 (d, J = 6.6 Hz, 3H), 0.84 (d, J = 6.6 Hz, 3H); C NMR (101 MHz, CD OD)
3
δ 165.8, 164.9, 158.2, 154.3, 149.8, 139.6, 133.0, 127.3, 125.89, 115.7, 114.4, 114.2,
14.1, 112.7, 103.1, 73.6, 59.1, 55.5, 54.1, 37.0, 28.2, 20.6, 20.5; HRMS (ESI) m/z
1
-
calcd. for C H N O S ([M - H] ): 578.1967, found 578.1991.
3
0
33
3
7
4
.1.10.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-methoxyphenyl)sulfonamido)-1-phenylbutan-2
-yl)-7-methoxy-2-oxo-2H-chromene-3-carboxamide (6d)
The title compound was obtained by 4d which was coupled with 9 through
EDCI/HOBt/DMAP coupling procedure in 68% yield (colorless oil) as described for
1
6
a: mp 177-179 °C; H NMR (600 MHz, CD OD) δ 8.61 (s, 1H), 7.70 (d, J = 8.6 Hz,
3
2
H), 7.66 (d, J = 8.6 Hz, 1H), 7.26 (d, J = 7.4 Hz, 2H), 7.21 (t, J = 7.6 Hz, 2H), 7.13
(t, J = 7.3 Hz, 1H), 6.99 – 6.97 (m, 4H), 4.32 (ddd, J = 10.0, 5.9, 4.2 Hz, 1H), 3.92 (s,
3
1
1
0
1
H), 3.89- 3.86 (m, 1H), 3.79 (s, 3H), 3.43 (dd, J = 15.0, 3.7 Hz, 1H), 3.18 (dd, J =
4.0, 4.0 Hz, 1H), 3.10 (dd, J = 13.6, 8.1 Hz, 1H), 2.98 – 2.95 (m, 1H), 2.87 – 2.84 (m,
H), 2.78 (dd, J = 14.0, 10.0 Hz, 1H), 2.04 – 1.97 (m, 1H), 0.92 (d, J = 6.6 Hz, 3H),
1
3
.85 (d, J = 6.6 Hz, 3H); C NMR (151 MHz, CD OD) δ 166.9, 164.5, 163.7, 162.9,
3
58.1, 149.6, 139.5, 132.5, 132.0, 130.6, 130.5, 129.3, 127.4, 115.3, 115.2, 115.1,
113.5, 101.3, 73.4, 58.8, 56.8, 56.1, 55.5, 53.8, 36.4, 28.1, 20.5, 20.4; HRMS (ESI)
-
m/z calcd. for C H N O S ([M - H] ): 607.2120, found 607.2100.
3
2
36
2
8

4
.1.11.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-nitrophenyl)sulfonamido)-1-phenylbutan-2-yl)
-7-methoxy-2-oxo-2H-chromene-3-carboxamide (7d)
The title compound was obtained by 4d which was coupled with 10 through
EDCI/HOBt/DMAP coupling procedure in 50% yield (yellow crystalline powder) as
1
described for 6a: mp 263-265 °C; H NMR (600 MHz, CD OD) δ 8.65 (s, 1H), 8.32
3
(
d, J = 8.9 Hz, 2H), 8.02 (d, J = 8.9 Hz, 2H), 7.70 (d, J = 8.6 Hz, 1H), 7.24 (d, J = 8.6
Hz, 2H), 7.21 (t, J = 7.2 Hz, 1H), 7.13 (t, J = 7.2 Hz, 1H), 7.01 (dt, J = 7.1, 2.3 Hz,
H), 4.26 (ddd, J = 10.0, 6.0, 4.3 Hz, 1H), 3.76 (ddd, J = 9.0, 6.2, 3.2 Hz, 1H), 3.52
dd, J = 15.2, 3.2 Hz, 1H), 3.24 (dd, J = 13.6, 8.1 Hz, 1H), 3.18 (dd, J = 15.2, 8.7 Hz,
2
(
1
1
H), 3.11 (dd, J = 14.0, 4.2 Hz, 1H), 3.04 – 2.99 (m, 1H), 2.81 (dd, J = 14.0, 9.6 Hz,
13
H), 2.07 – 2.02 (m, 1H), 0.93 (d, J = 6.6 Hz, 3H), 0.87 (d, J = 6.6 Hz, 3H); C NMR
(
151 MHz, CD OD) δ 167.0, 163.8, 163.0, 158.2, 151.4, 149.7, 147.0, 139.2, 132.6,
3
1
30.6, 129.8, 129.4, 127.5, 125.4, 115.3, 115.1, 113.6, 101.3, 72.6, 57.8, 56.8 55.5,
-
5
2.8, 36.3, 27.7, 20.3; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 622.1866,
3
1
33
3
9
found 622.1842.
4
.1.12.
N-((2S,3R)-4-((4-amino-N-isobutylphenyl)sulfonamido)-3-hydroxy-1-phenylbutan-2-y
l)-7-methoxy-2-oxo-2H-chromene-3-carboxamide (8d)
The title compound was obtained by 4d which was coupled with 11 through
EDCI/HOBt/DMAP coupling procedure in 83% yield (yellow crystalline powder) as
1
described for 6a: H NMR (600 MHz, CD OD) δ 8.60 (s, 1H), 7.97 (s, 1H), 7.64 (d, J
3
=
8.6 Hz, 1H), 7.45 (d, J = 8.6 Hz, 2H), 7.27 (d, J = 7.2 Hz, 2H), 7.21 (t, J = 7.6 Hz,
H), 7.12 (t, J = 7.2 Hz, 1H), 6.97 – 6.94 (m, 2H), 6.63 (d, J = 8.6 Hz, 2H), 4.35 (ddd,
J = 10.0, 5.6, 4.2 Hz, 1H), 3.92 – 3.89 (m, 4H), 3.42 (dd, J = 15.0, 4.0 Hz, 1H), 3.19
dd, J = 13.9, 4.0 Hz, 1H), 3.02 (dd, J = 13.5, 8.1 Hz, 1H), 2.93 (dd, J = 14.9, 8.0 Hz,
2
(
1
6
1
H), 2.82 – 2.75 (m, 2H), 2.02 – 1.95 (m, 1H), 0.91 (t, J = 6.6 Hz, 3H), 0.85 (d, J =
1
3
.6 Hz, 3H); C NMR (151 MHz, CD OD) δ 166.9, 164.8, 163.7, 162.9, 158.1, 154.3,
3
49.6, 139.6, 132.5, 130.6, 130.5, 129.3, 127.3, 125.9, 115.2, 114.4, 113.5, 101.3,

7
3.6, 59.2, 56.77 (s), 55.5, 54.1, 36.9, 28.2, 20.6, 20.5; HRMS (ESI) m/z calcd. for
-
C H N O S ([M - H] ): 592.2124, found 592.2092.
31
35
3
7
4
2
.1.13.
-(7-Hydroxy-2-oxo-2H-chromen-4-yl)-N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-metho
xyphenyl)sulfonamido)-1-phenylbutan-2-yl)acetamide (6e)
The title compound was obtained by 4e which was coupled with 9 through
EDCI/HOBt/DMAP coupling procedure in 92% yield (colorless oil) as described for
1
6
a: mp 132-135 °C; H NMR (400 MHz, CD OD) δ 7.62 (d, J = 8.8 Hz, 2H), 7.18 (d,
3
J = 9.2 Hz, 1H), 7.09 – 7.08 (m, 4H), 7.06 – 7.04 (m, 1H), 6.95 (d, J = 8.8 Hz, 2H),
.56 (d, J = 9.2 Hz, 2H), 5.89 (s, 1H), 4.01 – 3.95 (m, 1H), 3.77 (s, 3H), 3.74 – 3.69
6
(m, 1H), 3.31 (dd, J = 14.8, 3.2 Hz, 1H), 3.08 (dd, J = 13.8, 3.2 Hz, 1H), 2.95 – 2.92
(m, 1H), 2.84 – 2.78 (m, 1H), 2.74 – 2.70 (m, 1H), 2.52 (dd, J = 13.8, 11.6 Hz, 1H),
1
3
1
.88 (dq, J = 20.4, 6.8 Hz, 1H), 0.79 (d, J = 6.6 Hz, 3H), 0.73 (d, J = 6.6 Hz, 3H); C
NMR (101 MHz, CD OD) δ 168.8, 163.1, 161.9, 161.4, 155.3, 150.8, 138.4, 130.6,
3
1
5
-
29.2, 128.8, 127.8, 126.2, 125.9, 113.9, 113.0, 111.6, 111.5, 102.1, 72.8, 57.5, 54.8,
4.1, 52.6, 38.7, 35.0, 26.6, 19.1, 19.0; HRMS (ESI) m/z calcd. for C H N O S ([M
3
2
36
2
8
-
H] ): 607.2109, found 607.2132.
4
2
.1.14.
-(7-Hydroxy-2-oxo-2H-chromen-4-yl)-N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-nitrop
henyl)sulfonamido)-1-phenylbutan-2-yl)acetamide (7e)
The title compound was obtained by 4e which was coupled with 10 through
EDCI/HOBt/DMAP coupling procedure in 72% yield (white powder) as described for
1
6
a: mp 223-225 °C; H NMR (400 MHz, CD OD) δ 8.37 (d, J = 8.0 Hz, 2H), 8.02 (d,
3
J = 8.0 Hz, 2H), 7.32 (d, J = 9.2 Hz, 1H), 7.24 – 7.13 (m, 5H), 6.67 – 6.66 (m, 2H),
6
3
.00 (s, 1H), 4.05 – 4.02 (m, 1H), 3.79 – 3.75 (m, 1H), 3.50 – 3.45 (m, 1H), 3.18 –
.09 (m, 3H), 2.99 – 2.95 (m, 1H), 2.62 (t, J = 12.6 Hz, 1H), 2.00 (dq, J = 13.6, 6.8
13
Hz, 1H), 0.89 (d, J = 6.4 Hz, 3H), 0.84 (d, J = 6.4 Hz, 3H); C NMR (101 MHz,
CD OD) δ 170.4, 163.4, 162.9, 156.7, 152.2, 151.4, 147.0, 139.7, 130.2, 129.8, 129.3,
3

1
2
27.6, 127.4, 125.4, 114.4, 113.1, 113.0, 103.6, 73.4, 57.9, 55.7, 53.2, 40.2, 36.4, 27.8,
-
0.3; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 622.1854, found 622.1888.
3
1
33
3
9
4
.1.15.
N-((2S,3R)-4-((4-amino-N-isobutylphenyl)sulfonamido)-3-hydroxy-1-phenylbutan-2-y
l)-2-(7-hydroxy-2-oxo-2H-chromen-4-yl)acetamide (8e)
The title compound was obtained by 4e which was coupled with 11 through
EDCI/HOBt/DMAP coupling procedure in 51% yield (yellow powder) as described
1
for 6a: mp 179-181 °C; H NMR (500 MHz, CD OD) δ 7.76 (brs, 2H), 7.41 (d, J =
3
1
2
1
1
1
2.0 Hz, 1H), 7.31 (d, J = 8.0 Hz, 1H), 7.24 – 7.15 (m, 7H), 6.67 (brs, 2H), 4.11 (m,
H), 3.85 (m, 1H), 3.64 (m, 3H), 3.37 (m, 2H), 2.90 (m, 2H), 2.69 (d, J = 10.0 Hz,
1
3
H), 1.97 (m, 1H), 0.86 (d, J = 10.0 Hz, 6H); C NMR (101 MHz, CD OD) δ 166.2,
3
62.6, 156.4, 152.2, 143.5, 139.6, 134.8, 130.5, 130.3, 130.1, 129.1, 127.5, 127.2,
20.6, 114.3, 112.9, 112.8, 103.3, 73.5, 58.4, 55.54 (s), 53.5, 31.6, 27.8, 20.3, 20.2;
-
HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 592.2112, found 592.2136.
3
1
35
3
7
4
.1.16.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-methoxyphenyl)sulfonamido)-1-phenylbutan-2
-yl)-2-(7-methoxy-2-oxo-2H-chromen-4-yl)acetamide (6f)
The title compound was obtained by 4f which was coupled with 9 through
EDCI/HOBt/DMAP coupling procedure in 48% yield (pale yellow crystalline powder)
1
as described for 6a: mp 115-117 °C; H NMR (400 MHz, CD OD) δ 7.69 (brs, 2H),
3
7.34 (d, J = 5.0 Hz, 1H), 7.16 (m, 5H), 7.02 (brs, 2H), 6.84 (m, 4H), 6.75 – 6.74 (m,
1H), 6.02 (brs, 1H), 4.07 (brs, 1H), 3.84 (s, 6H), 3.59 – 3.54 (m, 2H), 3.42 – 3.38 (m,
1H), 3.17 - 3.14 (m, 1H), 2.97 -2.83 (m, 4H), 2.62 – 2.57 (m, 1H), 1.98 (brs, 1H), 0.83
13
(
d, J = 22.0 Hz, 6H); C NMR (101 MHz, CD OD) δ 170.2, 164.6, 164.5, 163.1,
3
1
1
56.7, 152.0, 139.9, 132.1, 130.6, 130.3, 129.3, 127.4, 127.3, 115.4, 114.0, 113.7,
01.8, 74.2, 59.0, 56.4, 56.2, 55.6, 54.1, 40.1, 36.4, 28.1, 20.5, 20.4; HRMS (ESI) m/z
-
calcd. for C H N O S ([M - H] ): 621.2277, found 621.2276.
3
3
38
2
8

4
.1.17.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-nitrophenyl)sulfonamido)-1-phenylbutan-2-yl)
-2-(7-methoxy-2-oxo-2H-chromen-4-yl)acetamide (7f)
The title compound was obtained by 4f which was coupled with 10 through
EDCI/HOBt/DMAP coupling procedure in 39% yield (white powder) as described for
1
6
a: mp 266-268 °C; H NMR (600 MHz, CD OD) δ 8.36 (d, J = 8.6 Hz, 2H), 8.02 (d,
3
J = 8.6 Hz, 2H), 7.40 (d, J = 8.8 Hz, 1H), 7.19 – 7.14 (m, 5H), 6.86 (s, 1H), 6.78 (d, J
8.8 Hz, 1H), 6.05 (s, 1H), 4.05 – 4.03 (m, 1H), 3.87 (s, 3H), 3.78 – 3.76 (t, J = 5.6
Hz, 1H), 3.57 (dd, J = 32.6, 14.9 Hz, 2H), 3.47 (d, J = 13.2 Hz, 1H), 3.17 – 3.09 (m,
H), 2.98 (dd, J = 12.8, 5.6 Hz, 1H), 2.64 – 2.60 (m, 1H), 2.01 – 1.98 (m, 1H), 0.89 (d,
=
3
1
3
J = 6.6 Hz, 3H), 0.84 (d, J = 6.6 Hz, 3H); C NMR (151 MHz, CD OD) δ 170.3,
3
1
64.5, 163.1, 156.7, 152.0, 151.4, 147.0, 139.7, 130.2, 129.8, 129.3, 127.4, 125.4,
113.9, 113.8, 113.7, 101.8, 73.5, 57.9, 56.4, 55.7, 53.2, 40.2, 36.4, 27.8, 20.3; HRMS
-
(
ESI) m/z calcd. for C H N O S ([M - H] ): 636.2022, found 636.2060.
32 35 3 9
4
.1.18.
N-((2S,3R)-4-((4-amino-N-isobutylphenyl)sulfonamido)-3-hydroxy-1-phenylbutan-2-y
l)-2-(7-methoxy-2-oxo-2H-chromen-4-yl)acetamide (8f)
The title compound was obtained by 4f which was coupled with 11 through
EDCI/HOBt/DMAP coupling procedure in 30% yield (yellow powder) as described
1
for 6a: mp 215-217 °C; H NMR (600 MHz, CD OD) δ 7.46 (d, J = 8.6 Hz, 2H), 7.34
3
(dt, J = 8.8, 3.2 Hz, 1H), 7.18 – 7.16 (m, 4H), 7.14 – 7.11 (m, 1H), 6.85 (d, J = 2.4 Hz,
1
H), 6.76 (dd, J = 8.8, 2.4 Hz, 1H), 6.68 (d, J = 8.6 Hz, 2H), 6.03 (d, J = 2.4 Hz, 1H),
4
.11 – 4.08 (m, 1H), 3.86 (s, 3H), 3.84 – 3.81 (m, 1H), 3.57 – 3.54 (m, 1H), 3.38 (dd,
J = 14.8, 3.6 Hz, 1H), 3.19 (dd, J = 14.0, 3.6 Hz, 1H), 2.97 – 2.95 (m, 1H), 2.87 (d, J
4.8 Hz, 1H), 2.84 (m, 1H), 2.77 (dd, J = 13.8, 6.6 Hz, 1H), 2.61 (dd, J = 14.0, 11.4
Hz, 1H), 1.96 (dt, J = 13.9, 6.6 Hz, 1H), 0.89 (d, J = 6.6 Hz, 3H), 0.83 (d, J = 6.6 Hz,
=
1
3
3
1
5
H); C NMR (151 MHz, CD OD) δ 170.2, 164.9, 164.4, 163.1, 156.6, 154.3, 152.0,
3
39.9, 130.5, 130.3, 129.3, 127.4, 127.3, 114.4, 114.0, 113.9, 113.7, 101.8, 74.4, 59.3,
6.4, 55.5, 54.3, 40.2, 36.9, 28.2, 20.6, 20.5; HRMS (ESI) m/z calcd. for

-
C H N O S ([M - H] ): 606.2280, found 606.2298.
32
37
3
7
4
2
4
.1.19.
-(7-(Dimethylamino)-2-oxo-2H-chromen-4-yl)-N-((2S,3R)-3-hydroxy-4-((N-isobutyl-
-methoxyphenyl)sulfonamido)-1-phenylbutan-2-yl)acetamide (6g)
The title compound was obtained by 4g which was coupled with 9 through
EDCI/HOBt/DMAP coupling procedure in 63% yield (pale yellow crystalline powder)
1
as described for 6a: mp 133-135 °C; H NMR (400 MHz, CD OD) δ 7.70 (d, J = 8.0
3
Hz, 2H), 7.26 (d, J = 9.0 Hz, 1H), 7.19 – 7.18 (m, 4H), 7.15 – 7.13 (m, 1H), 7.03 (d, J
=
8.0 Hz, 2H), 6.56 (d, J = 9.0 Hz, 1H), 6.48 (s, 1H), 5.85 (s, 1H), 4.09 – 4.04 (m, 1H),
.85 (s, 3H), 3.81 (t, J = 7.2 Hz, 1H), 3.51 (d, J = 9.0 Hz, 2H), 3.42 (dd, J = 14.9, 2.8
Hz, 1H), 3.17 (dd, J = 13.9, 2.4 Hz, 1H), 3.03 (s, 6H), 2.92 (dd, J = 14.9, 8.5 Hz, 2H),
3
2
3
1
.83 – 2.79 (m, 1H), 2.66 – 2.60 (m, 1H), 2.04 – 1.92 (m, 1H), 0.87 (d, J = 6.6 Hz,
1
3
H), 0.82 (d, J = 6.6 Hz, 3H); C NMR (101 MHz, CD OD) δ 170.5, 164.9, 164.5,
3
57.1, 154.6, 152.5, 139.9, 132.1, 130.6, 130.3, 129.3, 127.3, 127.0, 115.4, 110.7,
110.5, 109.8, 98.7, 74.2, 58.9, 56.2, 55.6, 54.1, 40.2, 40.1, 36.9, 28.1, 20.5, 20.4;
-
HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 634.2593, found 634.2593.
3
4
41
3
7
4
2
4
.1.20.
-(7-(Dimethylamino)-2-oxo-2H-chromen-4-yl)-N-((2S,3R)-3-hydroxy-4-((N-isobutyl-
-nitrophenyl)sulfonamido)-1-phenylbutan-2-yl)acetamide (7g)
The title compound was obtained by 4g which was coupled with 10 through
EDCI/HOBt/DMAP coupling procedure in 35% yield (yellow powder) as described
1
for 6a: mp 233-235 °C; H NMR (600 MHz, CD OD) δ 8.33 (d, J = 8.4 Hz, 2H), 7.99
3
(
d, J = 8.4 Hz, 2H), 7.34 (d, J = 9.0 Hz, 1H), 7.20 – 7.15 (m, 5H), 6.61 (d, J = 9.0 Hz,
H), 6.51 (s, 1H), 5.88 (s, 1H), 4.03 – 4.00 (m, 1H), 3.78 – 3.75 (m, 1H), 3.53 – 3.45
m, 4H), 3.20 (dd, J = 12.8, 5.2 Hz, 1H), 3.10 (d, J = 4.5 Hz, 1H), 3.05 (s, 6H), 2.98 –
.95 (m, 1H), 2.65 – 2.60 (m, 1H), 1.99 – 1.94 (m, 1H), 0.88 (d, J = 6.6 Hz, 3H), 0.83
1
(
2
1
3
(
d, J = 6.6 Hz, 3H); C NMR (126 MHz, DMSO-d ) δ 167.3, 160.6, 155.3, 152.6,
6
1
49.5, 145.2, 139.0, 129.1, 128.5, 128.1, 127.9,126.0, 125.8, 124.4, 109.4, 108.8,

1
08.1, 97.4, 55.6, 53.5, 51.3, 40.3, 34.8, 25.8, 19.8, 19.7; HRMS (ESI) m/z calcd. for
-
C H N O S ([M - H] ): 649.2338, found 649.2306.
33
38
4
8
4
.1.21.
N-((2S,3R)-4-((4-amino-N-isobutylphenyl)sulfonamido)-3-hydroxy-1-phenylbutan-2-y
l)-2-(7-(dimethylamino)-2-oxo-2H-chromen-4-yl)acetamide (8g)
The title compound was obtained by 4g which was coupled with 11 through
EDCI/HOBt/DMAP coupling procedure in 32% yield (yellow oil) as described for 6a:
1
mp 197-200 °C; H NMR (600 MHz, CD OD) δ 7.45 (d, J = 8.8 Hz, 2H), 7.25 (d, J =
3
9
.0 Hz, 1H), 7.19 – 7.18 (m, 4H), 7.16 – 7.12 (m, 1H), 6.67 (d, J = 8.8 Hz, 2H), 6.56
dd, J = 9.0, 2.4 Hz, 1H), 6.49 (d, J = 2.4 Hz, 1H), 5.85 (s, 1H), 4.08 (ddd, J = 10.8,
.6, 3.6 Hz, 1H), 3.82 (ddd, J = 8.4, 6.9, 3.6 Hz, 1H), 3.55 – 3.47 (m, 2H), 3.37 (dd, J
14.9, 3.6 Hz, 1H), 3.18 (dd, J = 13.9, 3.6 Hz, 1H), 2.99 (s, 3H), 2.94 (dd, J = 13.8,
.4 Hz, 1H), 2.88 – 2.85 (m, 1H), 2.86 (s, 3H), 2.77 (dd, J = 13.6, 6.9 Hz, 1H), 2.62
dd, J = 14.0, 11.4 Hz, 1H), 1.97 – 1.92 (m, 1H), 0.88 (d, J = 6.6 Hz, 3H), 0.83 (d, J =
(
6
=
8
(
1
3
6
1
5
.6 Hz, 3H); C NMR (151 MHz, CD OD) δ 170.5, 164.3, 157.1, 154.7, 154.3, 152.5,
3
39.9, 130.5, 130.3, 129.3, 127.3, 126.9, 125.9, 114.4, 110.7, 110.6, 109.8, 98.7, 74.4,
9.3, 55.6, 54.4, 40.2, 36.9, 36.5, 28.2, 20.6, 20.5; HRMS (ESI) m/z calcd. for
-
C H N O S ([M - H] ): 619.2597, found 619.2621.
33
40
4
6
4
.1.22.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-methoxyphenyl)sulfonamido)-1-phenylbutan-2
-yl)-2-(7-hydroxy-4-methyl-2-oxo-2H-chromen-3-yl)acetamide (6h)
The title compound was obtained by 4h which was coupled with 9 through
EDCI/HOBt/DMAP coupling procedure in 87% yield (pale yellow powder) as
1
described for 6a: mp 196-198 °C; H NMR (400 MHz, CD OD) δ 7.76 (d, J = 7.6 Hz,
3
2
H), 7.53 (d, J = 8.8 Hz, 1H), 7.16 (q, J = 7.4 Hz, 4H), 7.08 – 7.04 (t, J = 8.5 Hz, 3H),
.80 (d, J = 8.8 Hz, 1H), 6.68 (s, 1H), 4.06 (t, J = 5.8 Hz, 1H), 3.86 (s, 3H), 3.80 (dd,
6
J = 10.3, 4.8 Hz, 1H), 3.45 – 3.38 (m, 3H), 3.18 (d, J = 13.6 Hz, 1H), 3.06 – 3.01 (m,
H), 2.92 (dd, J = 15.0, 8.6 Hz, 1H), 2.85 – 2.81 (m, 1H), 2.61 (t, J = 12.5 Hz, 1H),
1

2
.08 (s, 3H), 2.04 – 1.98 (m, 1H), 0.91 (d, J = 6.4 Hz, 3H), 0.85 (d, J = 6.4 Hz, 3H);
1
3
C NMR (101 MHz, CD OD) δ 172.2, 164.5, 164.3, 162.4, 155.3, 152.2, 140.0,
3
1
5
32.0, 130.6, 130.3, 129.2, 127.6, 127.2, 116.7, 115.4, 114.3, 114.2, 103.2, 74.2, 59.1
6.2, 55.4, 54.2, 36.7, 35.1, 28.0, 20.5, 20.4, 15.4; HRMS (ESI) m/z calcd. for
-
C H N O S ([M - H] ): 621.2277, found 621.2300.
33
38
2
8
4
.1.23.
N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4-nitrophenyl)sulfonamido)-1-phenylbutan-2-yl)
-2-(7-hydroxy-4-methyl-2-oxo-2H-chromen-3-yl)acetamide (7h)
The title compound was obtained by 4h which was coupled with 10 through
EDCI/HOBt/DMAP coupling procedure in 63% yield (white powder) as described for
1
6
a: mp 293-295 °C; H NMR (400 MHz, CD OD) δ 8.40 (d, J = 8.0 Hz, 2H), 8.08 (d,
3
J = 8.0 Hz, 2H), 7.55 (d, J = 8.8 Hz, 1H), 7.17 – 7.15 (m, 4H), 7.09 – 7.07 (m, 1H),
6
3
6
0
1
1
2
6
.81 (d, J = 8.8 Hz, 1H), 6.69 (s, 1H), 3.99 (t, J = 6.8 Hz, 1H), 3.76 (t, J = 8.0 Hz, 1H),
.50 – 3.47 (m, 1H), 3.42 (d, J = 13.6 Hz, 2H), 3.20 – 3.06 (m, 3H), 2.98 (dd, J = 13.6,
.4 Hz, 1H), 2.60 (t, J = 12.4 Hz, 1H), 2.13 (s, 3H), 2.03 (dt, J = 13.2, 6.8 Hz, 1H),
1
3
.91 (d, J = 6.4 Hz, 3H), 0.86 (d, J = 6.4 Hz, 3H); C NMR (101 MHz, CD OD) δ
3
72.3, 164.3, 162.5, 155.4, 152.2, 151.4, 146.8, 139.9, 130.3, 129.9, 129.3, 127.7,
27.2, 125.4, 116.7, 114.3, 114.2, 103.3, 73.6, 58.2, 55.6, 53.5, 36.9, 35.1, 27.8, 20.4,
-
0.3, 15.4; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 636.2022, found
3
2
35
3
9
36.2047.
4
.1.24.
N-((2S,3R)-4-((4-amino-N-isobutylphenyl)sulfonamido)-3-hydroxy-1-phenylbutan-2-y
l)-2-(7-hydroxy-4-methyl-2-oxo-2H-chromen-3-yl)acetamide (8h)
The title compound was obtained by 4h which was coupled with 11 through
EDCI/HOBt/DMAP coupling procedure in 58% yield (yellow oil) as described for 6a:
1
mp 232-234 °C; H NMR (400 MHz, CD OD) δ 7.54 – 7.49 (m, 3H), 7.18 – 7.11 (m,
3
4
H), 7.03 (t, J = 7.0 Hz, 1H), 6.80 (d, J = 8.8 Hz, 1H), 6.73 (d, J = 7.8 Hz, 2H), 6.67
(s, 1H), 4.10 – 4.05 (m, 1H), 3.79 (t, J = 7.2 Hz, 1H), 3.43 (brs, 2H), 3.36 (d, J = 13.6

Hz, 1H), 3.17 (d, J = 13.6 Hz, 1H), 2.96 – 2.94 (m, 1H), 2.91 – 2.89 (m, 1H), 2.78 (dd,
J = 13.2, 6.8 Hz, 1H), 2.60 (t, J = 12.4 Hz, 1H), 2.07 (s, 3H), 1.99 – 1.94 (m, 1H),
1
3
0
1
1
2
6
.90 (d, J = 6.4 Hz, 3H), 0.85 (d, J = 6.4 Hz, 3H); C NMR (101 MHz, CD OD) δ
3
72.2, 164.9, 162.4, 155.3, 153.7, 152.3, 140.0, 130.5, 130.4, 129.2, 127.7, 127.1,
26.4, 116.7, 114.9, 114.3, 114.2, 103.2, 74.3, 59.3, 55.3, 54.4, 36.9, 35.1, 28.1, 20.6,
-
0.5, 15.4; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 606.2280, found
3
2
37
3
7
06.2289.
4
2
.1.25.
-(7-Amino-4-methyl-2-oxo-2H-chromen-3-yl)-N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4
-methoxyphenyl)sulfonamido)-1-phenylbutan-2-yl)acetamide (6i)
The title compound was obtained by 4i which was coupled with 9 through
EDCI/HOBt/DMAP coupling procedure in 87% yield (yellow oil) as described for 6a:
1
mp 253-255 °C; H NMR (500 MHz, CD OD) δ 7.74 (d, J = 8.0 Hz, 2H), 7.38 (d, J =
3
8
.5 Hz, 1H), 7.19 – 7.11 (m, 4H), 7.05 – 7.02 (m, 3H), 6.63 (d, J = 8.5 Hz, 1H), 6.49
(brs, 1H), 4.04 – 4.02 (m, 1H), 3.85 (s, 3H), 3.78 (t, J = 6.0 Hz, 1H), 3.42 – 3.35 (m,
3
1
H), 3.15 (dd, J = 13.5, 2.5 Hz, 1H), 3.02 (dd, J = 13.5, 8.0 Hz, 1H), 2.92 (dd, J =
5.0, 8.5 Hz, 1H), 2.84 – 2.81 (m, 1H), 2.62 – 2.57 (m, 1H), 2.04 (s, 3H), 2.01 – 1.97
1
3
(
m, 1H), 0.90 (d, J = 6.5 Hz, 3H), 0.85 (d, J = 6.5 Hz, 3H); C NMR (101 MHz,
CD OD) δ 172.6, 165.0, 164.5, 155.9, 154.0, 152.9, 139.9, 132.0, 130.6, 130.3, 129.2,
3
1
27.4, 127.2, 115.4, 114.1, 113.2, 111.8, 100.4, 74.1, 59.0, 56.2, 55.3, 54.1, 36.7, 35.1,
-
2
8.0, 20.5, 15.3; HRMS (ESI) m/z calcd. for C H N O S ([M - H] ): 620.2437,
3
3
39
3
7
found 620.2452.
4
2
.1.26.
-(7-Amino-4-methyl-2-oxo-2H-chromen-3-yl)-N-((2S,3R)-3-hydroxy-4-((N-isobutyl-4
-nitrophenyl)sulfonamido)-1-phenylbutan-2-yl)acetamide (7i)
The title compound was obtained by 4i which was coupled with 10 through
EDCI/HOBt/DMAP coupling procedure in 97% yield (pale yellow powder) as
1
described for 6a: mp 253-255 °C; H NMR (500 MHz, DMSO-d ) δ 8.39 (d, J = 8.5
6