A broad screening program previously identified phenprocoumon (1) as a small molecule template for inhibition of HIV protease. Subsequent modification of this lead through iterative cycles of structure-based design led to the activity enhancements of pyrone and dihydropyrone ring systems (II and V) and amide-based substitution (III). Incorporation of sulfonamide substitution within the dihydropyrone template provided a series of highly potent HIV protease inhibitors, with structure-activity relationships described in this paper. Crystallographic studies provided further information on important binding interactions responsible for high enzymatic binding. These studies culminated in compound VI, which inhibits HIV protease with a Ki value of 8 pM and shows an IC90 value of 100 nM in antiviral cell culture. Clinical trials of this compound (PNU-140690, Tipranavir) for treatment of HIV infection are currently underway.

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Recently, cyclooctylpyranone derivatives with m-carboxamide substituents (e.g. 2c) were identified as potent, nonpeptidic HIV protease inhibitors, but these compounds lacked significant antiviral activity in cell culture. Substitution of a sulfonamide group at the meta position, however, produces compounds with excellent HIV protease binding affinity and antiviral activity. Guided by an iterative structure-based drug design process, we have prepared and evaluated a number of these derivatives, which are readily available via a seven-step synthesis. A few of the most potent compounds were further evaluated for such characteristics as pharmacokinetics and toxicity in rats and dogs. From this work, the p-cyanophenyl sulfonamide derivative 35k emerged as a promising inhibitor, was selected for further development, and entered phase I clinical trials.

A unique strategy for the enhancement of secondary binding of an inhibitor to an enzyme has been demonstrated in the design of new human immunodeficiency virus (HIV) protease inhibitors. When the planar benzene ring of a 4-hydroxycoumarin lead compound (1a, Ki = 0.800 microM) was replaced with medium-sized (i.e., 7-9), conformationally-flexible, alkyl rings, the enzyme inhibitory activity of the resulting compounds was dramatically improved, and inhibitors with more than 50-fold better binding (e.g., 5d, Ki = 0.015 microM) were obtained. X-ray crystal structures of these inhibitors complexed with HIV protease indicated the cycloalkyl rings were able to fold into the S1' pocket of the enzyme and fill it much more effectively than the rigid benzene ring of the 4-hydroxycoumarin compound. This work has resulted in the identification of a promising lead structure for the design of potent, deliverable HIV protease inhibitors. Compound 5d, a small (MW = 324), nonpeptidic structure, has already shown several advantages over peptidic inhibitors, including high oral bioavailability (91-99%), a relatively long half-life (4.9 h), and ease of synthesis (three steps).

Recently, the novel cyclooctylpyranone HIV protease inhibitor 1 was identified in our labs, and an X-ray structure of this inhibitor complexed with HIV-2 protease was obtained. This crystal structure was used to develop two strategies for creating derivatives of 1 with enhanced enzyme inhibitory activity. The first strategy, substitution on the cyclooctyl ring, met with limited success, but provided some interesting information about the conformationally-flexible cycloocytyl ring on the inhibitors. The second strategy, substitution at the meta position of the aryl ring, was far more successful and generated compounds, such as the carboxamide derivatives 41 (Ki = 3.0 +/- 0.4 nM) and 36 (Ki = 4.0 +/- 0.8 nM), which were significantly more active than the corresponding unsubstituted cycloocytlpyranone 2 (Ki = 11.7 +/- 4.7 nM). An X-ray crystal structure of 36 complexed with HIV-1 protease indicated the increase in binding affinity is most likely due to the additional interactions between the amide substituent and the S3 region of the protease.