Design, Biologic Evaluation, and SAR of Novel Pseudo-peptide
of inhibitors 12b-14b and 21b suggested that replacement of
acetamide with bromide resulted in the enhancement of toxicity.
replace isobutyl to enhance the symmetry of inhibitors (12c-20c).
However, the unexpected decreases in potency were observed for
these replacements. This observation indicated that the smaller
isobutyl was the more preferable ligand for HIV-1 protease S1¢
pocket.
Inhibitor 23 exhibited significant ability to inhibit enzyme activity,
with an IC50 value of 5 nM benefiting from the incorporation of 2-
benzoxalinone as P2–P3 ligand. The most active compound 23
showed the lowest docking energy ()13.82 Kcal ⁄ mol). The top-
ranked docked conformation of inhibitor 23 is superimposed with
the experimentally determined binding structure of Vx-478 (ampre-
navir, 1hpv). As seen from this model, it appeared that 2-benzoxa-
zolinone carbonyl oxygen resided within an effective hydrogen
bonding distance to Asp29 and Asp30 (3.3 ꢀ) in the S2 region of
the active site. Additionally, the phenyl ring interacted with the
amino acids residues (Ile47) within the S2 pockets. The hydrogen
bonding interaction between acetamide carbonyl and Asp29¢ could
be formed (distance 3.3 ꢀ). All these interactions contributed to the
high inhibitory potency of inhibitor 23.
Conclusions and Future Directions
A series of novel HIV-1 protease inhibitors based on the (hydroxy-
ethylamino)-sulfonamide isostere incorporating substituted phenyls
and benzheterocycle derivatives bearing rich hydrogen bonding ac-
ceptors as P2 ligands were synthesized. Prolonged chain linking the
benzheterocycle to the carbonyl group resulted in partial loss of
binding affinities. Introduction of a small alkyl substituent with
appropriate size to the -CH2- of P1-P2 linkage as a side chain
resulted in improved inhibitory potency, and in this study, isopropyl
was the best side chain. Replacement of the isobutyl substituent at
P1¢group with phenyl substituent decreased the inhibitory potency.
One of the most potent inhibitors, compound 23 showing high
affinity to HIV-1 protease with an IC50 value of 5 nM, also exhibited
good anti-SIV activity (EC50 = 0.8 lM) with low toxicity
(TC50 > 100 lM). The flexible docking of inhibitor 23 to HIV-1 prote-
ase active site rationalized the interactions with protease. Further
design and chemical modifications of these inhibitors utilizing struc-
ture-based drug design strategies are currently underway.
To explore the optimal length between the P2 group and the car-
bonyl group, we designed several inhibitors with different linkages
between benzheterocycle and the carbonyl group. Compared with
12a, inhibitor 13a and 14a showed attenuated inhibitory activities
with 4-fold and 7.5-fold decrease in IC50, respectively. Similarly, the
inhibitory potency of 16a and 17a was reduced when compared
with that of 15a (IC50 = 20 nM). These reduced inhibition data
might result from the increased length linking the ring to the car-
bonyl group, such change presumably rendered the aromatic portion
not fill well in the S2 pocket and interfered with the interface
between the inhibitor and residues in the protease active site.
Acknowledgments
We also attempted to introduce a methyl substituent to inhibitor
15a at the -CH2- of P1-P2 linkage to yield inhibitor 18a. The intro-
duction of methyl substituent in the linkage resulted in a 1.5-fold
improvement in potency (IC50 = 12 nM). The increased inhibitory
potency might be because of the additional lipophilic interactions
with the residues within S1-S2 subsites by adding methyl substitu-
ent. And 15a and 18a showed equivalent ability to SIV-infected
cells with moderate EC50 value (2.0 lM), which meant the increased
bulk did not impress their permeance into the cells. To extend
this observation, larger alkyl substituents, isopropyl substituent,
and isobutyl substituent were introduced to the -CH2- of P1-P2 link-
age to produce 19a and 20a. However, decreased inhibitory
potency were observed, which might be because the larger alkyl
group within S2 pocket affected the critical hydrogen bond to
Asp 29.
We are grateful for the financial support from the National Natural
Science Foundation of China (NO. 30670415). We also acknowledge
Prof. Arthur J. Olson of the Scripps Research Institute in La Jolla,
CA for his provision of the AutoDock 3.0 program.
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In addition to the screening of P2-P3 ligands, attempts to change
P1¢ and P2¢ ligands were conducted. Replacement of acetamide
group of 12a-21a with bromide produced 12b-21b, which
resulted in more or less decrease in inhibitory potency except 13b
and 21b. The hydrogen bonding (acetamide CO to Asp29 NH), con-
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and 21b showed higher IC50 values, they still exhibited better DG
and EC50 results than 13a and 21a, respectively, so we suspected
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