Crystal Structures for HIV-1 Reverse Transcriptase in Complexes with Three Pyridinone Derivatives:  A New Class of Non-Nucleoside Inhibitors Effective against a Broad Range of Drug-Resistant Strains

Abstract

In the treatment of AIDS, the efficacy of all drugs, including non-nucleoside inhibitors (NNRTIs) of HIV-1 reverse transcriptase (RT), has been limited by the rapid appearance of drug-resistant viruses. Lys103Asn, Tyr181Cys, and Tyr188Leu are some of the most common RT mutations that cause resistance to NNRTIs in the clinic. We report X-ray crystal structures for RT complexed with three different pyridinone derivatives, R157208, R165481, and R221239, at 2.95, 2.9, and 2.43 A resolution, respectively. All three ligands exhibit nanomolar or subnanomolar inhibitory activity against wild-type RT, but varying activities against drug-resistant mutants. R165481 and R221239 differ from most NNRTIs in that binding does not involve significant contacts with Tyr181. These compounds strongly inhibit wild-type HIV-1 RT and drug-resistant variants, including Tyr181Cys and Lys103Asn RT. These properties result in part from an iodine atom on the pyridinone ring of both inhibitors that interacts with the main-chain carbonyl oxygen of Tyr188. An acrylonitrile substituent on R165481 substantially improves the activity of the compound against wild-type RT (and several mutants) and provides a way to generate novel inhibitors that could interact with conserved elements of HIV-1 RT at the polymerase catalytic site. In R221239, there is a flexible linker to a furan ring that permits interactions with Val106, Phe227, and Pro236. These contacts appear to enhance the inhibitory activity of R221239 against the HIV-1 strains that carry the Val106Ala, Tyr188Leu, and Phe227Cys mutations.

High-resolution structures of HIV-1 reverse transcriptase/TMC278 complexes: Strategic flexibility explains potency against resistance mutations

“…2B). A similar tunnel was seen in the binding of a cyanovinyl-containing iodopyridinone (IOPY) NNRTI (PDB ID 2B5J) (17). In the free TMC278 molecule, the cyanovinyl group is expected to be coplanar with the dimethylphenyl ring.…”

New Pyridinone Derivatives as Potent HIV-1 Nonnucleoside Reverse Transcriptase Inhibitors

“…It is consistent with binding modes experimentally observed for similar derivatives. 15 The pyridinone ring is pinned between Leu100 and Val106, allowing the lactam NH to form a hydrogen bond with Lys101. The hydrophobic 5-Et group faces Tyr181, favoring the conformational change required for efficient inhibition.…”

“…Charged groups appeared clearly unfavorable for activity (14 vs 2), suggesting replacement of the hydrolyzable ester moiety of the initial derivatives. Polar neutral groups were better tolerated in this position (15,18), and compounds displaying weakly polar groups (16,20) (23) were more favorable than a methyl group (21). A selectivity index (cytotoxicity vs antiviral activity) was also measured for the studied compounds.…”

“…Enzyme activities were determined by following the incorporation of [ 32 P]dGTP into a homopolymeric template/primer substrate, poly(rC)-oligo(dG) [12][13][14][15][16][17][18] . The RT assays using recombinant HIV-1 RT (Calbiochem) were performed in a final volume of 50 µL.…”

“…The RT assays using recombinant HIV-1 RT (Calbiochem) were performed in a final volume of 50 µL. Briefly, the reaction mixture contained 50 mM Tris-HCl, pH 8.3, 50 mM KCl, 10 mM MgCl 2 , 2 mM DTT, 10 µg of BSA, 0.01% Triton X100, 20 µg/mL poly(rC)-oligo(dG) [12][13][14][15][16][17][18] (Amersham), a fixed concentration of the labeled substrate (1 µCi per assay [ 32 P]dGTP), a fixed concentration of the inhibitor (10 µM, dissolved in 1 µL of dimethyl sulfoxide). After a 10 min of incubation at 37°C, the reaction mixtures were chilled on ice and collected on a DEAE-Filtermat (Perkin-Elmer).…”