Crystal-structure-based design and synthesis of novel C-terminal inhibitors of HIV protease.

The X-ray crystal-structure-based design, synthesis, computational evaluation, and activity of a novel class of HIV protease inhibitors are described. The initial lead compounds 2 and 3 were designed by modeling replacement groups for the C-terminal Val-Val-OCH3 of a known hydroxyethylene inhibitor into the active site of the reported crystal structure of HIV protease complexed with MVT-101. The lead compound 2 was found to be a modest inhibitor with a Ki = 1.67 microM. The X-ray crystal structure of compound 2 complexed with HIV protease was solved and used for subsequent design. The lead compound 3 was found to be a more potent inhibitor with Ki = 0.2 microM, and the structure of it complexed with HIV protease was also solved and used for subsequent design. Modification of both the C-terminus and N-terminus of indole 3 resulted in compounds with Ki = 30 nM. Using the crystal structure of compounds 2 and 3 with HIV protease as a starting point, the thermodynamic cycle perturbation molecular dynamics method was applied to a select group of compounds in order to test the accuracy of this type of computation within a series of closely related compounds.

Structure-based design and synthesis of substituted 2-butanols as nonpeptidic inhibitors of HIV protease: secondary amide series.

The design, synthesis, and crystallographic analysis of protein-inhibitor complexes is described for a novel series of nonpeptidic HIV protease (HIV Pr)inhibitors. Beginning with a cocrystal structure of a Phe-Pro peptidomimetic bound to the HIV Pr, design was initiated that resulted in the substituted 2-butanol compound 8 as the lead compound (Ki = 24.5 microM, racemic mixture). Modifications on the initial compound were then made on the basis of its cocrystal structure with HIV Pr and inhibition data, resulting in compounds with enhanced potency against the enzyme (compound 18, Ki = 0.48 microM). These inhibitors were found to bind to the enzyme essentially as predicted on the basis of the original design hypothesis. Stereospecific synthesis of individual enantiomers confirmed the prediction of a binding preference for the S alcohol stereochemistry. Modest antiviral activity was demonstrated for several of the more potent HIV Pr inhibitors in a HIV-1 infected CEM-SS cell line.

Bis tertiary amide inhibitors of the HIV-1 protease generated via protein structure-based iterative design.

A series of potent nonpeptide inhibitors of the HIV protease have been identified. Using the structure of compound 3 bound to the HIV protease, bis tertiary amide inhibitor 9 was designed and prepared. Compound 9 was found to be about 17 times more potent than 3, and the structure of the protein-ligand complex of 9 revealed the inhibitor binds in an inverted binding mode relative to 3. Examination of the protein-ligand complex of 9 suggested several modifications in the P1 and P1' pockets. Through these modifications it was possible to improve the activity of the inhibitors another 100-fold, highlighting the utility of crystallographic feedback in inhibitor design. These compounds were found to have good antiviral activity in cell culture, were selective for the HIV protease, and were orally available in three animal models.

Antiviral and resistance studies of AG1343, an orally bioavailable inhibitor of human immunodeficiency virus protease.

AG1343 ([3S-(3R*,4aR*,8aR*,2'S*,3'S*)]-2-[2' hydroxy-3'-phenylthiomethyl-4'-aza-5'-oxo-5'-(2''-methyl-3''-hydro xy-phenyl) pentyl]-decahydroiso-quinoline-3-N-t-butylcarboxamide methanesulfonic acid) is a selective, nonpeptidic inhibitor of human immunodeficiency virus (HIV) protease (Ki = 2 nM) that was discovered by protein structure-based drug design methodologies. AG1343 was effective against the replication of several laboratory and clinical HIV type 1 (HIV-1) or HIV-2 isolates including pyridinone- and zidovudine-resistant strains, with 50% effective concentrations ranging from 9 to 60 nM. In reversibility studies, inhibition of gag (p55) proteolytic processing in HIV-1 particles from cells treated with AG1343 was maintained for up to 36 h after drug removal. The ability of virus to develop resistance to AG1343 was studied by serial passage of HIV-1 NL4.3 in the presence of increasing concentrations of drug. After 28 passages, a variant with a 30-fold reduction in susceptibility to AG1343 was isolated. Molecular analysis of the protease from this variant indicated a double change from a Met to Ile at residue 46 and an Ile to Val or Ala at residue 84 (M46I+I84V, A). Consistent with these findings, reductions in susceptibility were observed for recombinant viruses constructed to contain the single I84V change or the double M46I+I84V substitutions. Resistance, however, was not detected for recombinant viruses containing other key mutations in HIV-1 protease, including a Val to Ile change at residue 32 or a Val to Ala or Phe at residue 82. The potent anti-HIV activity of AG1343 against several isolates suggests that AG1343 should perform well during ongoing human phase II clinical trials.