Synthesis and activity of N-acyl azacyclic urea HIV-1 protease inhibitors with high potency against multiple drug resistant viral strains

Article abstract of DOI:10.1016/j.bmcl.2005.08.093

As part of our efforts to identify potent HIV-1 protease inhibitors that are active against resistant viral strains, structural modification of the azacyclic urea (I) was undertaken by incorporating acyl groups as P ₁′ ligands. The extensive SA

Full text of DOI:10.1016/j.bmcl.2005.08.093

Bioorganic & Medicinal Chemistry Letters 15 (2005) 5499–5503
Synthesis and activity of N-acyl azacyclic urea HIV-1
protease inhibitors with high potency against multiple drug
resistant viral strains
Chen Zhao,* Hing L. Sham, Minghua Sun, Vincent S. Stoll, Kent D. Stewart, Shuqun Lin,
Hongmei Mo, Sudthida Vasavanonda, Ayda Saldivar, Chang Park, Edith J. McDonald,
Kennan C. Marsh, Larry L. Klein, Dale J. Kempf and Daniel W. Norbeck
GPRD, Abbott Laboratories, Abbott Park, IL 60064-6123, USA
Received 26 April 2005; revised 25 August 2005; accepted 29 August 2005
Available online 3 October 2005
Abstract—As part of our efforts to identify potent HIV-1 protease inhibitors that are active0against resistant viral strains, structural
modification of the azacyclic urea (I) was undertaken by incorporating acyl groups as P1 ligands. The extensive SAR study has
yielded a series of N-acyl azacyclic ureas (II), which are highly potent against both wild-type and multiple PI-resistant viral strains.
Ó 2005 Elsevier Ltd. All rights reserved.
Ph
Ph
Inhibition of human immunodeficiency virus (HIV) pro-
tease is one of the most important approaches for the
intervention of HIV infection.1 A novel series of highly
potent azacyclic urea HIV-1 protease inhibitors (I)2 were
designed and synthesized by Abbott chemists based on
the linear aza protease inhibitors3 and the DuPont
MerckÕs C-2 symmetric cyclic ureas.4 However, the bio-
availability of this series is very low, seldom exceeding
10%. The metabolism studies revealed that efficient
hepatic oxidation of the carbon atoms a to the aza link-
age may be contributing to the poor pharmacokinetics.
To prevent this unfavorable oxidation, an acyl group
H
N
H
N
OH
OH
OH
N
H
N
H
Ph
Ph
C2-symmetric Diol
Core-unit of Ritonavir
O
R2
Ph
H
N
R1
OH
N
N
N
N
H
Ph
Ph
Ph
OH OH
Aza Backbone
Modified Core-unit
R2
O
R1
DuPont Merck's
Cyclic Urea
N
N
N
0
was utilized instead of the benzyl or alkyl group as P1
Ph
Ph
OH
ligands. The extensive SAR study has yielded a series
of N-acyl azacyclic ureas (II), which exhibits excellent
protease inhibition and antiviral activity against wild
type HIV. Furthermore, we have recently found that
this series of compounds also retain high activity against
some highly resistant strains. For example, inhibitor II-
15 holds EC50 values of 54 and 65 nM against the indi-
navir-resistant (Ind-R)5 and lopinavir-resistant (A17)6
viruses. This degree of activity represents a more than
10-fold improvement over lopinavir (see Fig. 1).
I, Aza-cyclic Urea
R1
O
R2
N
N
N
O
R3
Ph
OH
II, N-Acyl Aza-cyclic Urea
Figure 1. Design of rational and structures of ritonavir, azacyclic
ureas, and N-acyl azacyclic ureas.
Keywords: HIV-1 protease inhibitor; Resistant viral strain; N-Acyl
azacyclic urea.
The synthesis of N-acyl azacyclic ureas is outlined in
Scheme 1. Opening of 2S,3R-epoxide 17 with Cbz-pro-
tected benzylhydrazine 28,9 provided compound 3 in
*
Corresponding author. Tel.: +1 847 938 5292; fax: +1 847 937
9195; e-mail: chen.zhao@abbott.com
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.08.093

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C. Zhao et al. / Bioorg. Med. Chem. Lett. 15 (2005) 5499–5503
Ph
Ph
OH
SEMO
CbzNH
O
Ph
CbzNH
Ph
a
b
CbzNH
Ph
N
N
+
NHCbz
NHCbz
HN
74%
65%
NHCbz
Ph
4
2
1
3
O
O
Ph
H
H
H
H
SEMO
NH2
N
N
N
N
N
N
c
f
N
NH2
H
100%
62%
100%
Ph
R1
Ph
Ph
Ph
OSEM
OSEM
5
6
7
O
O
R1
O
O
R2
N
R2
H
R2
H
H
N
N
N
N
N
h
N
N
g
i
N
O
N
N
O
N
O
O
R3
Ph
R3
R3
R3
Ph
Ph
Ph
OSEM
OH
II
OSEM
OSEM
9
8
10
Scheme 1. Reagents and conditions: (a) i-PrOH, reflux, 48 h; (b) SEM-Cl, (i-Pr)2NEt, CH2Cl2, rt, overnight; (c) H2, 10% Pd–C, MeOH, rt, 4 h; (d)
CDI, CH2Cl2, rt, 6 days; (e) H2, 10% Pd(OH)2/C, MeOH, rt, 2 h; (f) R3COCl, pyridine, CH2Cl2, rt; (g) R2X, KOtBu, THF, rt; (h) R1X, NaH, DMF,
rt; (i) 4 N HCl, MeOH, rt.
Table 1. HIV-1 protease inhibitory and antiviral activities of N-acyl azacyclic ureas-R3modifications
MeO
HO
OMe
OH
O
N
N
N
R3
Ph
HO
Compound R3
IC50 (nM) or %
inhibition at
0.5 nM
EC50 (lM)
Compound R3
IC50 (nM) or %
inhibition at
0.5 nM
EC50 (lM)
0% HS 50% HS
0% HS 50% HS
O
OMe
OMe
O
II-1
73%
66%
0.009
0.003
0.027
0.060
II-13
67%
72%
0.004
0.015
0.062
0.047
O
O
II-2
II-14
OMe
O
O
O
II-3
7.9
6.1
0.572
1.49
0.490
3.22
II-15
73%
67%
1.2
0.004
0.004
0.212
0.054
0.003
0.009
0.004
0.004
0.022
0.016
0.056
0.028
0.546
0.353
0.016
0.018
0.056
0.029
0.483
0.067
N
N
O
O
O
NH
II-4
II-16
F
O
O
II-5
76%
3.8
0.009
0.308
0.086
0.929
0.004
0.005
0.061
0.006
0.077
0.576
0.480
5.15
II-17
N
O
O
O
O
II-6
II-18
58%
73%
68%
64%
81%
79%
93%
N
N
O
O
O
N
II-7
2.1
II-19
O
O
O
O
O
II-8
53%
55%
75%
73%
67%
II-20
O
O
II-9
0.028
0.027
0.165
0.048
II-21
S
O
II-10
I-1
O
II-11
Ritonavir
Lopinavir
OH
O
II-12
OMe

C. Zhao et al. / Bioorg. Med. Chem. Lett. 15 (2005) 5499–5503
5501
Table 2. HIV-1 protease inhibitory and antiviral activities of N-acyl azacyclic ureas-R1 and R2 modifications
O
R1
R2
N
N
R3
N
Ph
HO
Compound
R1
R2
R3
% inhibition at 0.5 nM
76a
EC50 (lM) 0% HS
O
O
O
II-22
0.017
0.053
0.045
HO
HO
MeO
OMe
NH
II-23
II-24
88a
73a
H N
2
2
OH
OH
N
N
H2N
O
O
O
O
90a
74a
66a
48a
65b
66b
56b,c
0.074
0.013
0.045
0.050
0.030
0.037
0.150
N
II-25
II-26
II-27
II-28
II-29
II-30
II-31
N
H2N
H2N
H
N
N
N
H
N
H
NH2
N
N
H
O
HO
HO
H2N
H2N
O
O
H
2N
H2N
a At pH 7.0.
b At pH 4.5.
c At 1.0 nM.
74% yield. The secondary hydroxy group was protected
as its SEM-ether 4. After deprotecting the Cbz groups
via hydrogenolysis, the resulting diamine 5 was cyclized
with 1,10-carbonyldiimidazole (CDI) in methylene chlo-
ride to give the azacyclic urea 6 in 62% yield. The N-ben-
zyl group was successfully removed by palladium
hydroxide catalyzed hydrogenolysis in quantitative yield
to afford the key intermediate 7. Acylation of the same
nitrogen provided N-acyl azacyclic urea core 8. Unsym-
metrical alkylation of the urea nitrogens was accom-
plished in two steps. Deprotonation with potassium
tert-butoxide in THF followed by addition of halide
provided mono alkylated product 9. Compound 9 was
then reacted with sodium hydride and second halide in
DMF to afford the diakylated product 10. The com-
pounds with identical R1 and R2 substituents were made
by deprotonation with sodium hydride followed by
addition of 2 equiv of halide. Finally, the SEM ether
was removed with 4 N hydrochloric acid in methanol
at room temperature to give the series of acyl azacyclic
ureas II.12
in Tables 1 and 2. Compared to the most potent azacy-
clic urea (I-1)2, the acyl analogs displayed similar cellu-
lar antiviral activities. Acyl azacyclic ureas retained
excellent activity in the presence of 50% of human ser-
um. For most of acyl azacyclic ureas, the serum effect
is modest and the activity is about 3- to 8-fold decreased
(Table 1).
In addition to antiviral potency against wild type HIV,
the activity against multiple drug resistant viral strains
is a very important criterion for selecting next genera-
tion protease inhibitors. Despite the efficacy of current
protease inhibitors containing antiretroviral regimens,
some failures do occur due to the development of viral
resistance. Selected acyl azacyclic ureas were tested
against multiple drug resistant strains IND-R5 and
A17.6 A number of compounds displayed excellent
activity, with up to 15-fold improved potency over Lop-
inavir (Table 3).
Molecular modeling and X-ray crystal structures were
used to understand the SAR. The crystal structure of
II-11 (Fig. 2, PDB entry 2A4F)11 reveals a binding mode
essentially identical to that of I-1 (PDB entry 1PRO).2b
The acyl azacyclic ureas were tested in enzyme inhibition
and whole cell antiviral assays.10 The results are shown

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C. Zhao et al. / Bioorg. Med. Chem. Lett. 15 (2005) 5499–5503
Table 3. Antiviral activities of N-acyl azacyclic ureas against HIV wild
type and resistant mutant virus
Asp30
Asp25
Compound
EC50 (lM)
WT
INDa
A17b
0.586
0.367
>10
Asp29’
II-1
II-2
0.009
0.003
0.086
0.004
0.006
0.004
0.004
0.003
0.009
0.017
0.053
0.045
0.074
0.013
0.045
0.050
0.030
0.037
0.150
0.004
0.022
0.016
0.094
0.132
1.04
Asp25’
Ile50’
II-7
II-9
0.049
0.088
0.054
0.164
0.081
0.227
0.014
0.504
—
0.102
0.133
0.065
0.189
0.455
0.562
0.192
0.464
2.02
Asp29
II-12
II-15
II-16
II-19
II-20
II-22
II-23
II-24
II-25
II-26
II-27
II-28
II-29
II-30
II-31
I-1
Asp30’
Ile50
Figure 3. Overlap X-ray crystal structures of the N-acyl azacyclic urea
II-11, and azacyclic urea I-1 in complex with HIV-1 protease.
0.058
0.023
0.821
0.102
0.171
—
0.084
0.098
1.07
is solvent exposed, not making direct protein contacts.
These interactions may contribute to the excellent activ-
ity of this series against the resistant viral strains (see
Fig. 3).
0.897
0.312
2.07
—
—
—
—
Ritonavir
Lopinavir
—
0.385
1.24
1.06
Preliminary pharmacokinetic evaluation indicated that
compounds in this series still have relatively low bio-
availability (0–23.2%), even upon co-dosing with ritona-
vir, in spite of the blockage of the putative oxidative N-
dealkylation. The observed poor pharmacokinetics of
the acyl azacyclic ureas may be a consequence of very
low solubility rather than a result of metabolic path-
ways. Further experiments are underway to ascertain
this hypothesis and results will be reported separately.
a IND-R: L10R/M46I/L63P/V82T/I84V.
b A17: L10F/V32I/M46I/I47V/Q58E/I84V.
Asp30
Asp25
Asp29’
Asp25’
References and notes
1. Chrusciel, R. A.; Strohbagh, J. W. Curr. Top. Med. Chem.
2004, 4, 1097.
Ile50’
Asp29
2. (a) Sham, H. L.; Zhao, C.; Marsh, K. C.; Betebenner, D. A.;
Lin, S.; Rosenbrook, W.; Herrin, T.; Li, L.; Madigan, D.;
Vasavanonda, S.; Molla, A.; Saldivar, A.; McDonald, E.;
Wideburg, N. E.; Kempf, D. J.; Norbeck, D. W.; Plattner, J.
J. Biochem. Biophys. Res. Commun. 1996, 225, 436; (b)
Sham, H. L.; Zhao, C.; Stewart, K. D.; Betebenner, D. A.;
Lin, S.; Park, C. H.; Kong, X.; Rosenbrook, W.; Herrin, T.;
Madigan, D.; Vasavanonda, S.; Lyons, N.; Molla, A.;
Saldivar, A.; Marsh, K. C.; McDonald, E.; Wideburg, N.
E.; Denissen, J. F.; Robins, T.; Kempf, D. J.; Plattner, J. J.;
Norbeck, D. W. J. Med. Chem. 1996, 39, 392.
Asp30’
Ile50
Figure 2. X-ray crystal structure of the N-acyl azacyclic urea II-11 in
complex with HIV-1 protease.
3. Sham, H. L.; Betebenner, D. A.; Zhao, C.; Wideburg, N.
E.; Saldivar, A.; Kempf, D. J.; Plattner, J. J.; Norbeck, D.
W. J. Chem. Soc., Chem. Commun. 1993, 1052.
4. Lam, P. Y. S.; Jadhav, P. K.; Eyermann, C. J.; Hodge, C.
N.; Ru, Y.; Bacheler, L. T.; Meek, J. L.; Otto, M. J.;
Rayner, M. M.; Won, Y. N.; Chang, C. H.; Weber, P. C.;
Jackson, D. A.; Sharp, T. R.; Erickson-Vittanen, S.
Science 1994, 263, 380.
5. Resistant viral strain IND-R: L10R/M46I/L63P/ V82T/
I84V. See: (a) Javier Martinez-Picado, J.; Savara, A. V.;
Sutton, L.; DÕAquila, R. T. J. Virol. 1999, 73, 3744; (b)
Condra, J. H.; Schleif, W. A.; Blahy, O. M.; Gabryelski,
L. J.; Graham, D. J.; Quintero, J. C.; Rhodes, A.;
Shivaprakash, M.; Titus, D.; Yang, T.; Teppler, H.;
Squires, K. E.; Deutsch, P. J.; Emini, E. A. Nature 1995,
374, 569.
The 3-OCH3/4-OH makes the same hydrogen bonds
with the main chain N–H groups of Asp29, Asp30,
Asp290, and Asp300. The hydroxyl group of the azacy-
clic urea is oriented between the catalytic aspartates of
Asp25 and Asp250. The carbonyl oxygen of the azacyclic
urea core is hydrogen bonded to the main chain N–H of
Ile50 and Ile500. The phenyl ring makes van der Waals
interactions with Gly490, Val82, and Ile84. The differ-
ence between II-11 and I-1 is the P1 side chain. The acyl
oxygen is not making a direct contact to the protein, it is
˚
3.8 A from the main chain N–H of Ile50, the closest pro-
tein atom. The remainder of the acyl side chain makes
van der Waals contact with Leu230, Val820, and Pro810.
The terminal para-hydroxyl group on the acyl side chain
0

C. Zhao et al. / Bioorg. Med. Chem. Lett. 15 (2005) 5499–5503
5503
6. Resistant viral strain A17: L10F/V32I/M46I/I47V/ Q58E/
I84V. See: Mo, H.; Lu, L.; Dekhtyar, T.; Stewart, K. D.;
sun, E.; Kempf, D. J.; Molla, A. Antiviral Res. 2003, 59, 173.
7. 2S,3R-epoxide (1) was synthesized from Cbz-L-phenyla-
laninol. See Ref. 3.
8. Compound 2 was prepared from benzylhydrazine and N-
(benzyloxycarbonyloxy)succinimide in CH2Cl2 at rt in
69% yield.
Miller, M.; Jaskolski, M.; Rao, J. K. M.; Leis, J.;
Wlodawer, A. Nature 1989, 337, 576.
11. Data were collected at beamline 17-ID in the facilities of
the Industrial Macromolecular Crystallography Associa-
tion Collaborative Access Team (IMCA-CAT) at the
Advanced Photon Source These facilities are supported by
the companies of the Industrial Macromolecular Crystal-
lography Association through a contract with Illinois
Institute of Technology (IIT), executed through IITÕs
Center for Synchrotron Radiation Research and Instru-
mentation. Use of the Advanced Photon Source was
supported by the U.S. Department of Energy, Basic
Energy Sciences, Office of Science under Contract No. W-
31-109-Eng-38.
9. Dutta, A. S.; Morley, J. S. J. Chem. Soc., Perkin Trans. 1
1975, 1712.
10. Protease inhibition assay and cell culture assay see (a)
Sham, H. L.; Zhao, C.; Marsh, K. C.; Betebenner, D. A.;
Lin, S.; Rosenbrook, W.; Herrin, T.; Li, L.; Madigan, D.;
Vasavanonda, S.; Molla, A.; Saldivar, A.; McDonald, E.;
Wideburg, N. E.; Kempf, D. J.; Norbeck, D. W.; Plattner,
J. J. Biochem. Biophys. Res. Commun. 1996, 225, 436; (b)
12. All Compounds reported have NMR, MS, and analytical
data consistent with the structures assigned.