Structure-based design of HIV protease inhibitors: Sulfonamide- containing 5,6-dihydro-4-hydroxy-2-pyrones as non-peptidic inhibitors

Full text of DOI:10.1021/jm960541s

© Copyright 1996 by the American Chemical Society
Volume 39, Number 22
October 25, 1996
Communications to the Editor
One promising possibility to interrupt the viral life cycle
Str u ctu r e-Ba sed Design of HIV P r otea se
In h ibitor s: Su lfon a m id e-Con ta in in g
5,6-Dih yd r o-4-h yd r oxy-2-p yr on es a s
Non -P ep tid ic In h ibitor s¶
is the use of inhibitors of the virally encoded protease,
which is indispensable for viral maturation.1,2 Among
the most potent inhibitors are peptidomimetic com-
pounds containing transition-state inserts in place of
the dipeptidic cleavage sites of the substrates.3-6 The
low oral bioavailability and rapid excretion of peptide-
derived compounds7 have limited their utility as poten-
tial therapeutic agents. Recent advances have resulted
in HIV protease inhibitors with reduced peptidic char-
acter that are more orally bioavailable. An increasing
number of HIV protease inhibitors8-21 are currently
undergoing clinical evaluations, some of which have
already been approved as therapeutic agents for the
treatment of HIV infection.
We previously reported20 the identification of phen-
procoumon (compound 1 in Figure 1, Ki ) 1 µM) from a
broad screening program as an active HIV protease
inhibitory template. It is noted that other independent
studies22-26 have also described 4-hydroxybenzopyran-
2-ones and 4-hydroxypyran-2-ones as inhibitors of HIV
protease. The increasing number of reported crystal
structures of inhibitor/HIV protease complexes have
provided numerous successful examples of structure-
based designs of potent HIV protease inhibitors.27,28 Our
iterative cycles of structure-based design identified
U-96988 (compound 2, Ki ) 38 nM, antiviral IC50 ) 3
µM) as the first clinical candidate.20 Further structure-
based drug design studies led to the discovery of the
sulfonamide-containing U-103017 (compound 3, Ki < 1
nM, antiviral IC50 ) 1-2 µM) as the second generation
clinical candidate.21 More recently, compounds in the
5,6-dihydro-4-hydroxy-2-pyrone template have demon-
strated promising HIV protease inhibitory activity,29,30
and we have previously reported HIV protease inhibi-
tory activity (Ki ) 35 nM) of compound 4.30 We now
describe the structure-activity relationship study of a
new series of sulfonamide-containing 5,6-dihydro-4-
hydroxy-2-pyrones leading to the identification of the
third-generation development candidate.
Suvit Thaisrivongs,*,‡ Harvey I. Skulnick,†
Steve R. Turner,‡ J oseph W. Strohbach,‡
Ruben A. Tommasi,‡ Paul D. J ohnson,†
Paul A. Aristoff,† Thomas M. J udge,‡
Ronald B. Gammill,‡ J eanette K. Morris,‡
Karen R. Romines,‡ Robert A. Chrusciel,
Roger R. Hinshaw,§ Kong-Teck Chong,§
W. Gary Tarpley,§ Susan M. Poppe,§ David E. Slade,§
J anet C. Lynn,| Miao-Miao Horng,| Paul K. Tomich,|
Eric P. Seest,| Lester A. Dolak,| W. J effrey Howe,∇
Gina M. Howard,# Francis J . Schwende,#
Lisa N. Toth,# Guy E. Padbury,# Grace J . Wilson,@
Lihua Shiou,¥ Gail L. Zipp,¥ Karen F. Wilkinson,#
Bob D. Rush,# Mary J . Ruwart,#
Kenneth A. Koeplinger,# Zhiyang Zhao,#
Serena Cole,$ Renee M. Zaya,$ Thomas J . Kakuk,$
Musiri N. J anakiraman,‡ and
Keith D. Watenpaugh‡
Discovery Chemistry, Structural Analytical & Medicinal
Chemistry, Biochemistry, Infectious Diseases Research,
Chemical Biological Screening, Computer-Aided Drug
Discovery, Drug Metabolism Research, Endocrine
Pharmacology & Metabolism, Pharmaceutical Development,
and Preclinical Toxicology, Pharmacia & Upjohn, Inc.,
Kalamazoo, Michigan 49001
Received J uly 25, 1996
The rapid spread of the acquired immunodeficiency
syndrome (AIDS) epidemic has stimulated discovery of
therapeutic agents to arrest the replication of the
causative virus, human immunodeficiency virus (HIV).
¶
Dedicated to Prof. Dieter Seebach on the occasion of his 60th
birthday.
† Discovery Chemistry.
‡ Structural Analytical & Medicinal Chemistry.
Biochemistry.
§ Infectious Diseases Research.
| Chemical Biological Screening.
∇ Computer-Aided Drug Discovery.
The preparation of a representative sulfonamide-
containing 5,6-dihydro-4-hydroxy-2-pyrone V is shown
in Scheme 1. The 5,6-dihydro-4-hydroxy-2-pyrone 530
was condensed with m-nitrobenzaldehyde with alumi-
#
Drug Metabolism Research.
@ Endocrine Pharmacology & Metabolism.
¥
Pharmaceutical Development.
Preclinical Toxicology.
$
S0022-2623(96)00541-9 CCC: $12.00 © 1996 American Chemical Society

4350 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 22
Communications to the Editor
F igu r e 1. Structures of HIV protease inhibitors.
Sch em e 1. Preparation of Sulfonamide-Containing
5,6-Dihydro-4-hydroxy-2-pyronesa
a
(a) m-NO2C6H4CHO, AlCl3, THF; (b) Et3Al, CuBr(CH3)2S,
THF; (c) 10% Pd/C, NH4HCO2, CH3OH; (d) RSO2Cl, pyridine.
F igu r e 2. Crystal structures of the HIV-1 protease triple
mutant (Q7K/L33I/L63I) complexed with inhibitors V (a, top)
and VIIb (b, bottom). The inhibitors are depicted in space-
filling representation, with carbon (gray), oxygen (red), nitro-
gen (blue), sulfur (yellow), and fluorine (purple) atoms as
shown. The enzyme surface at the active site is depicted as
dotted surface, and selected amino acid residues surrounding
the active site are also shown and labeled. See text for
discussion of binding of inhibitors in the active site.
Ta ble 1. Sulfonamide-Containing
5,6-Dihydro-4-hydroxy-2-pyrones
over the substituted phenyl analogues. The crystal
structure of the HIV-1 protease triple mutant (Q7K/
L33I/L63I) complexed with inhibitor V was determined.
This particular protease construct provided an enzyme
with significantly improved stability for structural
studies; its preparation has been described elsewhere.31
Although inhibitor V is a mixture of four diastereomers,
only the 3RR,6R diastereomer could be fitted into the
difference electron density at the active site. The crystal
structure showed nearly symmetrical hydrogen bonding
between the 4-hydroxyl group and the catalytic aspartic
acids. The carbonyl oxygen also made nearly sym-
metrical hydrogen-bonding interactions with the NH
groups of the flap residues Ile 50A and Ile 50B and
replaced the ubiquitous water molecule found in com-
plexes of peptide-derived inhibitors. As shown in Figure
2a, the phenethyl and the propyl groups at C-6 could
be found in the S1′ and the S2′ subsites, respectively.
The C-3R ethyl and phenyl groups occupied the S1 and
the S2 subsites, respectively, with the imidazole sub-
stituent curling into the S3 subsite. The sulfonamide
functionality showed strong hydrogen bonding to a few
residues at the active site.
R
compd K1 (nM) IC50 (µM) IC90 (µM)
phenyl
I
17
8
1.6
1.1
1.2
1.0
0.5
7.3
4.2
4.3
3.0
0.9
4-cyanophenyl
4-fluorophenyl
1-quinolin-8-yl
1-methylimidazol-4-yl
II
III
IV
V
6
3
1
num trichloride catalysis to give the benzylidene inter-
mediate 6. Conjugate addition to that benzylidene
intermediate 6 with triethylaluminum in the presence
of copper(I) bromide dimethyl sulfide complex gave
adduct 7. The nitro functionality was then reduced with
transfer hydrogenolysis using ammonium formate and
10% palladium on carbon as a catalyst. The resulting
amine 8 was reacted with a 1-methylimidazole-4-sulfon-
yl chloride in the presence of pyridine to give the desired
product V.
The compounds in Table 1 are mixtures of four
diastereomers. The R groups of the sulfonamides af-
fected both the HIV protease inhibitory activity and also
the resulting in vitro antiviral activity. The imidazole
sulfonamide (compound V) showed enhanced activity
Since compound V showed promising activity (Ki ) 1
nM, IC50 ) 0.5 µM), the four individual stereoisomers

Communications to the Editor
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 22 4351
Ta ble 2. Individual Diastereomers of Sulfonamide-Containing 5,6-Dihydro-4-hydroxy-2-pyrones
R
compd
stereochemistry
Ki (nM)
IC50 (µM)
IC90 (µM)
1-methylimidazol-4-yl
Va
Vb
Vc
Vd
VIa
VIb
VIc
VId
VIIa
VIIb
VIIc
VIId
3RR,6S
3RR,6R
3RS,6S
3RS6R
3RR,6S
3RR,6R
3RS,6S
3RS,6R
3RR,6S
3RR,6R
3RS,6S
3RS,6R
0.12
0.06
1.0
0.58
0.13
>>1
>>1
0.11
0.04
>>1
0.49
0.14
0.03
1.7
1
0.56
>>1
>>1
0.89
0.26
>>1
1.0
0.3
5-cyano-2-pyridyl
0.04
0.007
0.12
0.10
0.018
0.008
0.22
0.032
5-(trifluoromethyl)-2-pyridyl
0.84
0.10
3.0
0.41
1.8
and VII) showed the most promising activity. Com-
pounds VIb and VIIb, with the 3RR and 6R stereo-
chemistry, were the most active. Both compounds
showed highly potent HIV protease inhibitory activity,
with Ki values below 10 pM, and IC50 values of 30 nM
in the HIV-1IIIB-infected H9 cells. Early safety studies
suggested advantages of compound VIIb over compound
VIb; therefore, compound VIIb (U-140690) was selected
as a candidate.
The crystal structure of the HIV-1 protease triple
mutant (Q7K/L33I/L63I) complexed with inhibitor VIIb
was determined. Difference electron density confirmed
the 3RR,6R stereochemistry, and the inhibitor was
found to bind in one clear orientation (see Figure 2b),
with features which were nearly identical to those
already described for inhibitor V.
F igu r e 3. Time-course blood levels of inhibitor VIIb after
intravenous (opened circles) and oral (closed circles) admin-
istrations to rats. The intravenous dose (n ) 3 rats) was 5 mg/
kg given in 10% propylene glycol and 22.5% (hydroxypropyl)-
â-cyclodextrin at 2 mg/g concentration. The oral dose (n ) 9
rats) was 10 mg/kg given in 0.01 M sodium hydroxide solution
(pH 10) at 2 mg/g concentration. The dotted line denotes the
cell-culture antiviral IC90 value of 0.1 µM in cell culture
medium containing 10% fetal bovine serum, while the dashed
line denotes the IC90 value of 1 µM in cell culture medium
containing 10% fetal bovine serum and 75% human plasma.
U-140690 also inhibited HIV-2 protease with high
potency (Ki < 1 nM). It was shown to be highly selective
for HIV proteases since it inhibited other selected
aspartyl proteases only weakly, with Ki values of 2, 15,
and 9 µM against human pepsin, human cathepsin D,
and human cathepsin E, respectively.35 U-140690 was
also shown to be very effective against a panel of 10
AZT-resistant HIV-1 clinical isolates in primary PBMC,
with a mean IC90 value of 0.15 ( 0.07 µM.36 Since there
are reports indicating that reduction of in vitro antiviral
activity11 and lack of in vivo antiviral activity37 may be
attributed to extensive binding of drug to plasma
proteins, we also evaluated the effect of protein binding
on the in vitro antiviral activity of U-140690. In the
HIV-1IIIB-infected H9 cell assay containing 10% fetal
bovine serum and 75% human plasma, U-140690 still
showed significant antiviral activity with an IC90 value
of 1 µM. Since laboratory ritonavir11 resistant isolates
have been described,38 studies with these HIV-1 iso-
lates,39 which were highly resistant to ritonavir, showed
them to be highly resistant to saquinavir,8 indivanir,12
and viracept,18 with 47- to >125-fold increases in IC90
values as compared to matched, parental isolates.
These isolates, however, remained significantly sensi-
tive to U-140690, with only a 6-fold increase in IC90
value as compared to the wild-type isolate.40 Moreover,
an evaluation of four ritonavir resistant clinical isolates
(32-67-fold increase in IC90 value for ritonavir) indi-
were prepared for biological activity evaluation. The
amine 8 was converted to the corresponding carbo-
benzyloxy analogue, which could then be resolved on a
preparative chiral HPLC column32 into the four indi-
vidual stereoisomers.33 The carbobenzyloxy protecting
group was then removed by catalytic hydrogenolysis,
and the resulting four diastereomers of the amine 8
were individually treated with the sulfonyl chloride to
afford the four separate stereoisomers. The results of
the biological testings are shown in Table 2. Due to the
high enzymatic inhibitory activity of these compounds,
the Ki value determination required the use of a
tandemly linked HIV protease34 since the native dimeric
enzyme dissociated at low enzyme concentration. Al-
though all four diastereomers (compounds Va-d) showed
HIV protease inhibitory activity, the two diastereomers
with the R stereochemistry at the C-3R center are the
more active components. Between compounds Va and
Vb, the diastereomer with the R stereochemistry at the
C-6 center (Vb) showed higher activity. Additional
variation in sulfonamides was then evaluated, and the
examples with substituted pyridyl group (compounds VI

4352 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 22
Communications to the Editor
(13) Lam, P. Y.-S.; J adhav, P. K.; Eyermann, C. J .; Hodge, C. N.;
Ru, Y.; Bacheler, L. T.; Meek. J . L.; Otto, M. J .; Rayner, M. M.;
Wong, Y. N.; Chang, C.-H.; Weber, P. C.; J ackson, D. A.; Sharpe,
T. R.; Erickson-Vitanen, S. Rational Design of Potent, Bioavail-
able, Non-peptide Cyclic Urea as HIV Protease Inhibitors.
Science 1994, 263, 380-384.
(14) Wong, Y. N.; Burcham, D. L.; Saxton, P. L.; Erickson-Vitanen,
S.; Grubb, M. F.; Quon, C. Y.; Huang, S.-M. A Pharmacokinetic
Evaluation of HIV Protease Inhibitors, Cyclic Ureas, in Rats and
Dogs. Biopharm. Drug Disp. 1994, 15, 535-544.
(15) Getman, D. P.; DeCrescenzo, G. A.; Heintz, R. M.; Reed, K. L.;
Talley, J . J .; Bryant, M. L.; Clare, M.; Houseman, K. A.; Marr,
J . J .; Mueller, R. A.; Vazquez, M. L.; Shieh, H.-S.; Stallings, W.
C.; Stegeman, R. A. Discovery of a Novel Class of Potent HIV-1
Protease Inhibitors Containing the (R)-(Hydroxyethyl)urea Isos-
tere. J . Med. Chem. 1993, 36, 288-291.
(16) Bryant, M. HIV Protease Inhibitors: Action to Efficacy. Inhibitor
SC-55389a presented at Session 46 (I) at the 34th ICAAC,
Orlando, FL, 1994.
(17) Kim, E. E.; Baker, C. T.; Dwyer, M. D.; Murcko, M. A.; Rao, B.
G.; Tung, R. D.; Navia, M. A. Crystal Structure of HIV-1 Protease
in Complex with VX-478, a Potent and Orally Bioavailable
Inhibitor of the Enzyme. J . Am. Chem. Soc. 1995, 117, 1181-
1182.
(18) Patick, A. K.; Mo, H.; Markowitz, M.; Appelt, K.; Wu, B.; Musick,
L.; Kalish, V.; Kaldor, S.; Reich, S.; Ho, D.; Webber, S. Antiviral
and Resistance Studies of AG1343, an Orally Bioavailable
Inhibitor of Human Immunodeficiency Virus Protease. Antimi-
crob. Agents Chemother. 1996, 40, 292-297.
(19) Mimoto, T.; Imai, J .; Kisanuki, S.; Enomoto, H.; Hattori, N.;
Akaju, K.; Kiso, Y. Kynostatin KNI-227 and -272, Highly Potent
Anti-HIV Agents: Conformationally Constrained Tripeptide
Inhibitors of HIV Protease Containing Allophenylnorstatine.
Chem. Pharm. Bull. 1992, 40, 2251-2253.
(20) Thaisrivongs, S.; Tomich, P. K.; Watenpaugh, K. D.; Chong, K.-
T.; Howe, W. J .; Yang, C.-P.; Strohbach, J . W.; Turner, S. R.;
McGrath, J . P.; Bohanon, M. J .; Lynn, J . C.; Mulichak, A. M.;
Spinelli, P. A.; Hinshaw, R. R.; Pagano, P. J .; Moon, J . B.;
Ruwart, M. J .; Wilkinson, K. F.; Rush, B. D.; Zipp, G. L.; Dalga,
R. J .; Schwende, F. J .; Howard, G. M.; Padbury, G. E.; Toth, L.
N.; Zhao, Z.; Koeplinger, K. A.; Kakuk, T. J .; Cole, S. L.; Zaya,
R. M.; Piper, R. C.; J effrey, P. Structure-Based Design of HIV
Protease Inhibitors: 4-Hydroxycoumarins and 4-Hydroxy-2-
pyrones as Non-peptidic Inhibitors. J . Med. Chem. 1994, 37,
3200-3204.
(21) Skulnick, H. I.; J ohnson, P. D.; Howe, W. J .; Tomich, P. K.;
Chong, K.-T.; Watenpaugh, K. D.; J anakiraman, M. N.; Dolak,
L. A.; McGrath, J . P.; Lynn, J . C.; Horng, M.-M.; Hinshaw, R.
R.; Zipp, G. L.; Ruwart, M. J .; Schwende, F. J .; Zhong, W.-Z.;
Padbury, G. E.; Dalga, R. J .; Shiou, L.; Possert, P. I.; Rush, B.
D.; Wilkinson, K. F.; Howard, G. M.; Toth, L. N.; Williams, M.
G.; Kakuk, T. J .; Cole, S. L.; Zaya, R. M.; Thaisrivongs, S.;
Aristoff, P. A. Structure-Based Design of Sulfonamide Substi-
tuted Non-peptidic HIV Protease Inhibitors. J . Med. Chem. 1995,
38, 4968-4971.
(22) Bourinbaiar, A. S.; Tan, X.; Nagorny, R. Effect of the Oral
Anticoagulant, Warfarin, on HIV-1 Replication and Spread.
AIDS 1993, 7, 129-130.
(23) Tummino, P. J .; Ferguson, D.; Hupe, L.; Hupe, D. Competitive
Inhibition of HIV-1 Protease by 4-Hydroxy-benzopyran-2-ones
and by 4-Hydroxy-6-phenylpyran-2-ones. Biochem. Biophys. Res.
Commun. 1994, 200, 1658-1664.
(24) Tummino, P. J .; Ferguson, D.; Hupe, D. Competitive Inhibition
of HIV-1 Protease by Warfarin Derivatives. Biochem. Biophys.
Res. Commun. 1994, 201, 290-294.
(25) Vara Prasad, J . V. N.; Para, K. S.; Lunney, E. A.; Ortwine, D.
F.; Dunbar, J . B., J r.; Ferguson, D.; Tummino, P. J .; Hupe, D.;
Tait, B. D.; Domagala, J . M.; Humblet, C.; Bhat, T. N.; Liu, B.;
Guerin, D. M. A.; Baldwin, E. T.; Erickson, J . W.; Sawyer, T. K.
Novel Series of Achiral, Low Molecular Weight, and Potent
HIV-1 Protease Inhibitors. J . Am. Chem. Soc. 1994, 116, 6989-
6990.
(26) Lunney, E. A.; Hagen, S. E.; Domagala, J . M.; Humblet, C.;
Kosinski, J .; Tait, B. D.; Warmus, J . S.; Wilson, M.; Ferguson,
D.; Hupe, D.; Tummino, P. J .; Baldwin, E. T.; Bhat, T. N.; Liu,
B.; Erickson, J . W. A Novel Nonpeptide HIV-1 Protease Inhibi-
tor: Elucidation of the Binding Mode and Its Application in the
Design of Related Analogs. J . Med. Chem. 1994, 37, 2664-2677.
(27) Wlodawer, A.; Erickson, J . W. Structure-based Inhibitors of
HIV-1 Protease. Annu. Rev. Biochem. 1993, 62, 543-585.
(28) Appelt, K. Crystal Structures of HIV-1 Protease-Inhibitor Com-
plexes. Perspect. Drug Discovery Des. 1993, 1, 23-48.
(29) Hamilton, H. W.; Tait, B. D.; Gajda, C.; Hagen, S. E.; Ferguson,
D.; Lunney, E. A.; Pavlovsky, A.; Tummino, P. J . 6-Phenyl-
6-alkylamido-5,6-dihydro-2H-pyran-2-ones: Novel HIV Protease
Inhibitors. Bioorg. Med. Chem. 1996, 6, 719-724.
cated only a 2-3-fold increase in IC90 value for
U-140690.40
After 5 mg/kg intravenous dosing to rats, CLtot ) 0.17
( 0.10 (L/h)/kg, Vss ) 0.51 ( 0.14 L/kg, and t1/2 ) 5.4 (
0.3 h. After 10 mg/kg oral dosing to rats, F ) 30%
(relative to the 5 mg/kg intravenous dosing). Time-
course blood levels of U-140690 after iv and po dosings
in rats are shown in Figure 3. Importantly, the blood
levels of U-140690 exceeded 1 µM (the in vitro IC90 value
in the presence of 10% fetal bovine serum and 75%
human plasma) for 8-12 h. Additional extensive pre-
clinical and safety studies are underway in order to
initiate clinical trials of U-140690 as the third-genera-
tion development candidate for the treatment of HIV
infection.
Su p p or tin g In for m a tion Ava ila ble: Physical data for
compounds 7, 8, VIIa , VIIb, VIIc, and VIId ; descriptions of
the HIV protease inhibition and cell-culture antiviral assays;
crystallization procedure for the recombinant triple mutant
of HIV-1 protease (Q7K/L33I/L63I) complexed with compounds
V and VIIb; data collection and structure refinement descrip-
tion and a table summary of selected diffraction data collection
and refinement statistics for the two crystal structures (7
pages). Ordering information is given on any current mast-
head page.
Refer en ces
(1) Blundell, T. L.; Lapatto, R.; Wilderspin, A. F.; Hemmings, A.
M.; Hobart, P. M.; Danley, D. E.; Whittle, P. J . The 3-D Structure
of HIV-1 Protease and the Design of Antiviral Agents for the
Treatment of AIDS. Trends Biol. Sci. 1990, 15, 425-430.
(2) Debouck, C.; Metcalf, B. W. Human Immunodeficiency Virus
Protease: A Target for AIDS Therapy. Drug. Dev. Res. 1990,
21, 1-17.
(3) Huff, J . R. HIV Protease: A Novel Chemotherapeutic Target
for AIDS. J . Med. Chem. 1991, 34, 2305-2314.
(4) Meek, T. D. Inhibitors of HIV-1 Protease. J . Enzyme Inhib. 1992,
6, 65-98.
(5) Thaisrivongs, S. Chapter 14. HIV Protease Inhibitors. Annu.
Rep. Med. Chem. 1994, 29, 133-144.
(6) Chong, K. T. Recent Advances in HIV-1 Protease Inhibitors. Exp.
Opin. Invest. Drugs 1996, 5, 115-124.
(7) Plattner, J . J .; Norbeck, D. W. Obstacles to Drug Development
from Peptide Leads. In Drug Discovery Technologies; Clark, C.
R., Moos, W. H., Eds.; Ellis Horwood Ltd.: Chichester, 1990;
Chapter 5, pp 92-126.
(8) Martin, J . A. Recent Advances in the Design of HIV Protease
Inhibitors. Antiviral Res. 1992, 17, 265-278.
(9) Kempf, D. J .; Codacovi, L.; Wang, X. C.; Kohlbrenner, W. E.;
Wideburg, N. E., Saldivar, A.; Vasavanonda, S.; Marsh, K. C.;
Bryant, P.; Sham, H. L.; Green, B. E.; Betebenner, D. A.;
Erickson, J .; Norbeck, D. W. Symmetry-based Inhibitors of HIV
Protease. Structure-Activity Studies of Acylated 2,4-Diamino-
1,5-diphenyl-3-hydroxypentane and 2,5-Diamino-1,6-diphenyl-
hexane-3,4-diol. J . Med. Chem. 1993, 36, 320-330.
(10) Kempf, D. J .; Marsh, K. C.; Fino, L. C.; Bryant, P.; Craig-
Kennard, A.; Sham, H. L.; Zhao, C.; Vasavanonda, S.; Kohlbren-
ner, W. E.; Wideburg, N. E.; Saldivar, A.; Green, B. E.; Herrin,
T.; Norbeck, D. W. Design of Orally Bioavailable, Symmetry-
based Inhibitors of HIV Protease. Bioorg. Med. Chem. 1994, 2,
847-858.
(11) Kempf, D. J .; Marsh, K. C.; Denissen, J . F.; McDonald, E.;
Vasavanonda, S.; Flentge, C. A.; Green, B. E.; Fino, L.; Park, C.
H.; Kong, X.-P.; Wideburg, N. E.; Saldivar, A.; Ruiz, L.; Kati,
W. M.; Sham, H. L.; Robins, T.; Stewart, K. D.; Hsu, A.; Plattner,
J . J .; Leonard, J . M.; Norbeck, D. W. ABT-538 is a Potent
Inhibitor of Human Immunodeficiency Virus Protease and has
High Oral Bioavailability in Humans. Proc. Natl. Acad. Sci.
U.S.A. 1995, 92, 2484-2488.
(12) Vacca, J . P.; Dorsey, B. D.; Schleif, W. A.; Levin, R. B.; McDaniel,
S. L.; Darke, P. L.; Zugay, J .; Quintero, J . C.; Blahy, O. M.; Roth,
E.; Sardana, V. V.; Schlabach, A. J .; Graham, P. I.; Condra, J .
H.; Gotlib, L.; Holloway, M. K.; Lin, J .; Chen, I.-W.; Vastag, K.;
Ostovic, D.; Anderson, P. S.; Emini, E. A.; Huff, J . R. L-735,-
524: An Orally Bioavailable Human Immunodeficiency Virus
Type 1 Protease Inhibitor. Proc. Natl. Acad. Sci. U.S.A. 1994,
91, 4096-4100.
(30) Thaisrivongs, S.; Romero, D. L.; Tommasi, R. A.; J anakiraman,
M. N.; Strohbach, J . W.; Turner, S. R.; Biles, C.; Morge, R. R.;
J ohnson, P. D.; Aristoff, P. A.; Tomich, P. K.; Lynn, J . C.; Horng,

Communications to the Editor
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 22 4353
M.-M.; Chong, K.-T.; Hinshaw, R. R.; Howe, W. J .; Finzel, B. C.;
Watenpaugh, K. D. Structure-Based Design of HIV Protease
Inhibitors: 5,6-Dihydro-4-hydroxy-2-pyrones as Effective, Non-
peptidic Inhibitors. J . Med. Chem., in press.
(35) We thank Professor Ben M. Dunn, University of Florida,
Gainesville, FL, for the aspartyl protease assay results.
(36) Experiments performed according to the procedure of J apour et
al. J apour, A. J .; Mayers, D. L.; J ohnson, V. A.; Kuritzkes, D.
R.; Beckett, L. A.; Arduino, J .-M.; Lane, J .; Black, R. J .;
Reichelderfer, P. S.; D'Aquila, R. T.; Crumpacker, C. S. Stan-
dardized Peripheral Blood Mononuclear Cell Culture Assay for
Determination of Drug Susceptibilities of Clinical Human Im-
(31) Mildner, A. M.; Rothrock, D. J .; Leone, J . W.; Bannow, C. A.;
Lull, J . M.; Reardon, I. M.; Sarcich, J . L.; Howe, W. J .; Tomich,
C. C.; Smith, C. W.; Heinrikson, R. L.; Tomasselli, A. G. The
HIV-1 Protease as Enzyme and Substrate: Mutagenesis of
Autolysis Sites and Generation of a Stable Mutant with Retained
Kinetic Properties. Biochemistry 1994, 33, 9405-9413.
(32) The Cbz-8 was resolved on a 0.46 × 25 cm Chiralcel OD-H
column (Chiral Technologies, Inc.); the mobile phase was 35%
2-propanol in hexane with 0.1% TFA at 0.5 mL/min flow rate,
monitored at 238 nm (retention times: 11.8, 17.7, 22.2, and 27.9
min for the four diastereomers).
(33) The absolute stereochemistries of the two chiral centers were
assigned from an X-ray crystal structure of a synthetic inter-
mediate derived from a chiral synthetic procedure. The detail
of the asymmetric synthetic process for the preparation of
U-140690 will be described elsewhere (J udge, T. M.; Phillips,
G.; Morris, J . K.; Lovasz, K. D.; Romines, K. R.; Luke, G. P.;
Tulinsky, J .; Tustin, J . M.; Chrusciel, R. A.; Strohbach, J . W.;
VanderVelde, S. L.; Dolak, L. A.; Seest, E. P.; Mizsak, S. A.;
Watt, W.; Morris, J .; Gammill, R. B. Asymmetric Synthesis of a
New Class of Potent, Nonpeptidic HIV Protease Inhibitors.
Unpublished results).
munodeficiency Virus Type
1 Isolates. Antimicrob. Agents
Chemother. 1993, 37, 1095-1101.
(37) Flexner, C.; Richman, D. D.; Bryant, M.; Karim, A.; Yeramian,
P.; Meechan, P.; Haubrich, R.; Para, M. F.; Fischl, M. A. Effect
of Protein Binding on the Pharmacodynamics of an HIV Protease
Inhibitor. Antiviral Res. 1995 , 26, A282.
(38) Markowitz, M.; Mo, H.; Kempf, D. J .; Norbeck, D. W.; Bhat, T.
N.; Erickson, J . W.; Ho, D. D. Selection and Analysis of Human
Immunodeficiency Virus Type 1 Variants with Increased Resis-
tance to ABT-538, a Novel Protease Inhibitor. J . Virol. 1995,
69, 701-706.
(39) Mo, H.; Markowitz, M.; Ho, D. D. Patterns of Specific Mutations
in HIV-1 Protease that Confer Resistance to a Panel of Protease
Inhibitors. Abstract no. 188, 2nd National Conference on Human
Retroviruses and Related Infections, Washington, D.C., J an 29-
Feb 2, 1995.
(40) Poppe, S. M.; Slade, D. E.; Chong, K.-T.; Hinshaw, R. R.; Pagano,
P. J .; Markowitz, M.; Gorman, R. R.; Tarpley, W. G. HIV-1
Resistant to Multiple Peptidomimetic Protease Inhibitors Lacks
Significant Cross-Resistance to the Dihydropyrone Inhibitors.
Unpublished results.
(34) Cheng, Y.-S. E.; Yin, F. H.; Foundling, S.; Blomstrom, D.;
Kettner, C. A. Stability and Activity of Human Immunodefi-
ciency Virus Protease: Comparison of the Natural Dimer with
a Homologous, Single-Chain Tethered Dimer. Proc. Natl. Acad.
Sci. U.S.A. 1990, 88, 9660-9664.
J M960541S