Synthesis and biological evaluation of novel amprenavir-based P1-substituted bi-aryl derivatives as ultra-potent HIV-1 protease inhibitors

Article abstract of DOI:10.1016/j.bmcl.2012.01.037

A series of P1-substituted biaryl amprenavir derivatives was designed and synthesized. These compounds were evaluated for enzyme inhibition and antiviral activity in vitro. Several compounds showed highly efficient antiviral activity with EC₅₀ values down to 0.10 nM, which are more potent than marketed HIV-1 protease inhibitors. Docking study indicated that 12c has similar binding mode to amprenavir with full occupancy in P1.

Full text of DOI:10.1016/j.bmcl.2012.01.037

Bioorganic & Medicinal Chemistry Letters 22 (2012) 1976–1979
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of novel amprenavir-based
P1-substituted bi-aryl derivatives as ultra-potent HIV-1 protease inhibitors
Jianwei Yan a, Ning Huang b, Shukun Li a, Liu-Meng Yang b, Weiqiang Xing a, Yong-Tang Zheng b,
,
⇑
Youhong Hu a,
⇑
a State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Shanghai 201203, China
b Key Laboratory of Animal Models and Human Disease Mechanisms of the Chinese Academy of Science & Yunnan Province, Kunming Institute of Zoology, Chinese Academy of Sciences,
Kunming, Yunnan 650223, China
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of P1-substituted biaryl amprenavir derivatives was designed and synthesized. These compounds
were evaluated for enzyme inhibition and antiviral activity in vitro. Several compounds showed highly
efficient antiviral activity with EC50 values down to 0.10 nM, which are more potent than marketed
HIV-1 protease inhibitors. Docking study indicated that 12c has similar binding mode to amprenavir with
full occupancy in P1.
Received 26 October 2011
Revised 11 January 2012
Accepted 12 January 2012
Available online 21 January 2012
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
HIV-1 protease inhibitors
Amprenavir
Darunavir biaryl compounds
Docking study
Almost three decades after the discovery of acquired immune
deficiency syndrome (AIDS), the pandemic is still severe.1 The glo-
bal AIDS epidemic has been one of the most challenging problems
in medicine of our time. The discovery of highly active antiretrovi-
ral therapy (HAART) represents a landmark breakthrough in the
clinical management of AIDS.2,3 As one of the crucial ingredients
in HAART, protease inhibitors (PIs) play an important role in pre-
venting the replication of human immunodeficiency virus (HIV).
Although several HIV-1 PIs have been used in clinical practice,
emergence of viral resistance4,5, adverse side effects6–11 and high
costs of the production have limited their utilization. Development
of novel HIV-1 PIs against multi-drug resistant viral strains with
unique profiles is still urgently required for HAART.
disclosed that the modification of P1 or P10 pocket should be a
promising approach toward the design of new drug with improved
resistance profiles. In addition, P10 elongate biaryl protease inhibi-
tors related to atazanavir also exhibited attractive antiviral activi-
ties, which were synthesized by palladium-catalyzed Suzuki and
P1
O
O
O
P2
S
NH2
O
N
H
N
P2'
O
OH
Modification based on the current approved drugs to find novel
potent inhibitors on antidrug-resistant virus is still an efficient way.
Darunavir as a new generation of HIV-1 protease inhibitor is from
amprenavir by increasing the binding of P2 pocket against the
mutant and the wild-type virus.12–14 In this series, the researchers
have pursued high affinity protease inhibitors by the modification
of ligand on P2 and P20 binding pocket.15–18 Two other series of
P1 and P10 chain-extend inhibitors based on darunavir showed
the inhibition of HIV-1 protease on sub-picomolar and low nano-
molar range in cellular antiviral activities.19–21 These investigations
P1'
Amprenavir
Ar
O
O
N
O
S
R
O
N
O
H
OH
⇑
Corresponding authors.
E-mail addresses: zhengyt@mail.kiz.ac.cn (Y.-T. Zheng), yhhu@mail.shcnc.ac.cn
(Y. Hu).
Figure 1. Biaryl HIV-1 protease inhibitors.
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2012.01.037

J. Yan et al. / Bioorg. Med. Chem. Lett. 22 (2012) 1976–1979
1977
Stille coupling reactions.22–25 Herein, we report a new class of P1
elongate biaryl compounds related to amprenavir (Fig. 1) with high
cellular antiviral potencies through the rapid synthesis of Suzuki
coupling (in Scheme 1).
yields. In addition, the intermediates 11 were obtained through
microwave-accelerated Suzuki reaction of 9 with appropriate
boronic acids, and then reduction of 11 gave the desired com-
pounds 12a–c.
The designed compounds in Table 1 were synthesized from
cheap and readily available industrial materials as depicted in
Scheme 1. The direct iodination of phenylalanine 126 in acetic acid
at the presence of I2, NaIO3, and sulfuric acid afforded 4-iodophe-
nylalanine 2. After esterification and Boc-protection of 2, com-
The data of enzyme inhibition and antiviral activities of 16 new
bi-aryl compounds are summarized in Table 1. The HIV-1 protease
inhibition data were assayed using the published method.29 The
antiviral EC50 for HIV-1 virus in vitro were determined in a
C8166 cell line.30 Biaryl compounds 10a–m with the different sub-
stituents have a range of nanomolar binding potencies of HIV-1
protease, which is less potent than amprenavir. However, the
antiviral activities of these compounds are maintained. In order
to increase the solubility of these compounds, compounds 12b
and 12c (R = NH2, Ar = 3-pyridyl or 4-pyridyl) were synthesized
and exhibited stronger enzymatic inhibitory activities at subn-
anomolar level, which are 2–10 times more active than amprena-
vir. Also their antiviral activities are improved up to near 30
times than amprenavir with excellent therapeutic index. From this
investigation, biaryl substitution at P1 site of amprenavir could be
the promising way to improve the potency of HIV-1 protease
inhibition.
pound 3 was converted to
substitution and decarboxylation.27 The reduction of
4 with NaBH4 in ethanol at ꢀ78 °C produced the
-halo alcohol 5.28
a
-halo ketone 4 by nucleophilic
a-halo ketone
a
After achieving epoxide intermediate by treatment of 5 in metha-
nol with K2CO3, regioselectively ring-opening of the epoxide with
isobutyl amine followed by sulfonylation yielded the sulfonamide
6 and 7, respectively. Removal of Boc protection with concentrated
hydrochloric acid in dioxane and then reaction with (S)-(+)-3-
hydroxytetrahydrofuran in the presence of phosgene gave the
key intermediates 8 and 9. By screening the different Pd-catalyzed
Suzuki reaction conditions, compound 8 with various commer-
cially available aryl boronic acids in the present of 5–10 mol % pal-
ladium/active carbon as catalyst, and K2CO3 as
acetonitrile/water gave the desired product 10a–m in 70–80%
a
base in
In order to obtain molecular insight into the putatively adopted
binding mode at the active site of HIV protease, docking study31
I
I
I
b,c
d,e
a
OH
OH
Boc
O
Boc
H2N
H2N
N
H
N
H
Cl
O
O
O
O
1
2
3
4
f
I
I
I
O
O
O
O
O
k,l
g,h,i
S
S
O
O
O
O
N
N
O
N
N
Boc
O
H
H
N
H
Cl
OH
OH
OH
8
6
5
g,h,j
m
Ar
I
I
O
O
O
O
O
O
O
O
k,l
S
S
O
S
O
O
NO2
NO2
O
O
N
N
N
H
N
O
N
H
N
O
H
OH
OH
OH
10a~m
9
7
n
Ar
S
Ar
S
O
O
O
O
O
O
o
NO2
NH2
O
O
N
N
N
H
N
O
O
H
OH
11
OH
12a~c
Scheme 1. Synthesis of compounds 10a–m and 12a–c. Reagents and conditions: (a) I2, NaIO3, H2SO4/HOAc, 71%; (b) SOCl2,CH3OH, reflux, >99%; (c) (Boc)2O, NaHCO3,
THF/CH3OH (4:1), >99%; (d) (i-Pr)2NMgBr, ClCH2CO2Na, MgCl2, THF; (e) 1 N HCl, 75% (for two steps); (f) NaBH4, EtOH, ꢀ78 °C, 83%; (g) K2CO3, CH3OH; (h) isobutyl amine,
EtOH; (i) 4-methoxybenzenesulfonyl chloride, NaHCO3, DCM, H2O, 68% (for three steps); (j) 4-nitrobenzenesulfonyl chloride, NaHCO3,CH2Cl2/H2O, 72% (for three steps); (k)
HCl, dioxane (l) triphosgene, TEA, (S)-(+)-3-hydroxytetrahydrofuran, DCM, 88–90% (for two steps); (m) boronic acids, Pd–C, K2CO3, CH3CN/H2O, 80 °C, 70–80%; (n) boronic
acid, Pd(PPh3)4, K2CO3, toluene/EtOH/H2O, MW, 100 °C, 75–90%; (o) Pd–C, NH2–NH2ꢁH2O, EtOH, 80–90%.

1978
J. Yan et al. / Bioorg. Med. Chem. Lett. 22 (2012) 1976–1979
Table 1
Enzymatic inhibitory and antiviral activity of biaryl protease inhibitorsa
Ar
S
O
O
O
R
O
N
H
N
O
OH
c
b
d
Compound
R
Ar
Ki (nM)
CC50 (mM)
EC50 (nM)
SI (CC50/EC50)
10a
10b
10c
10d
10e
10f
10g
10h
10i
10j
10k
10l
10m
12a
12b
12c
APVe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
OMe
NH2
4-F-Phenyl
5.8
6.4
5.8
7.4
5.2
1.8
1.9
4.3
1.6
8.1
3.5
1.6
0.14
0.23
0.21
0.24
0.17
0.17
0.14
0.041
0.19
0.32
0.16
0.21
0.26
0.032
0.20
0.26
0.23
3.98
4.29
2.32
6.89
6.04
3.63
5.53
4.30
3.46
9.2
4.48
3.08
2.68
2.25
0.10
0.10
2.93
35176
53613
90517
34833
28146
46832
25316
9535
54913
34783
35714
68182
97015
14222
2000000
2600000
78498
4-Cl-Phenyl
4-COMe-Phenyl
4-SMe-Phenyl
4-OCF3-Phenyl
3-Me-4-OMe-Phenyl
4-CO2H-Phenyl
3-OH-Phenyl
3-CO2Me-Phenyl
3-CN-Phenyl
3-Thienyl
3-Pyridyl
4-Pyridyl
Phenyl
3-Pyridyl
3.8
1.98
0.33
0.086
0.84
NH2
NH2
4-Pyridyl
a
b
c
Anti-HIV-1 data represent the mean values of two separate experiments.
Effective concentration required to protect C8166 cells against the cytopathogenicity of HIV-1 by 50%.
Cytostatic concentration required to reduce C8166 cell proliferation by 50% tested by MTT method.
d
e
In vitro therapeutic index; ratio CC50/EC50
.
Amprenavir (APV) was used as positive control.
Figure 2. Superposition of 12c (C-atoms colored in gray) and amprenavir (C-atoms colored in blue).
was performed using the published X-ray crystal structure of HIV-
to Ile50 and Ile150, the amino hydrogen binding to the carbonyl
oxygen of Asp30, the tetrahydrofuran oxygen binding to Asp130
backbone NH, the carbamate NH binding to Gly127, and the biaryl
rings resulted in a full occupancy of the pocket 1.
In summary, a series of P1-substituted biaryl amprenavir deriv-
atives were synthesized. Various biaryl substitutes of amprenavir
exhibit strong antiviral activities. Compounds 12b and 12c are
the most potent with antiviral potencies up to 30 times than that
of amprenavir. In addition, they show significant improvement in
cell toxic properties, displaying excellent in vitro therapeutic in-
dex. Docking study data show that the compound 12c has similar
1 protease complexed with amprenavir (PDB entry code: 3NU332
)
as the template structure. The most potent compound 12c was se-
lected and docked into the crystal structure of the HIV protease–
amprenavir complex. We considered the structural water next to
the flap would mediate hydrogen bonds to both Ile 50NH groups
in the flap. The occupation of 12c subsites P1, P10, P2, and P20 in
a manner similar to that of amprenavir was achieved, as shown
in Figure 2. The important binding interactions were maintained,
for example, the hydroxyl group binding to Asp125, the carboxyl-
ate oxygen and sulfonamide oxygen with a water molecule binding

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Acknowledgments
This work was supported by the National Natural Science Foun-
dation of China (Grant Nos.: 30600775, 81001462), the Eleventh
Five-Year Key Scientific and Technological Program of China
(2009ZX09501-029) and the CAS Knowledge Innovation Project
(KSCX2-YW-R-185). The authors gratefully thank the National Cen-
ter for Drug Screening for the contribution in biological evaluation.
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Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2012.01.037.
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The binding site was defined using the center of the ligand [13.5111, 63.7441,
0
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