ABSTRACT
Cathepsin B is a member of the papain superfamily of cysteine proteases and has been implicated in the pathology of numerous diseases, including arthritis and cancer. As part of an effort to identify potent, reversible inhibitors of this protease, we examined a series of dipeptidyl nitriles, starting with the previously reported Cbz-Phe-NH-CH(2)CN (19, IC(50) = 62 microM). High-resolution X-ray crystallographic data and molecular modeling were used to optimize the P(1), P(2), and P(3) substituents of this template. Cathepsin B is unique in its class in that it contains a carboxylate recognition site in the S(2)' pocket of the active site. Inhibitor potency and selectivity were enhanced by tethering a carboxylate functionality from the carbon alpha to the nitrile to interact with this region of the enzyme. This resulted in the identification of compound 10, a 7 nM inhibitor of cathepsin B, with excellent selectivity over other cysteine cathepsins.
Subject(s)
Cathepsin B/antagonists & inhibitors , Dipeptides/chemical synthesis , Enzyme Inhibitors/chemical synthesis , Nitriles/chemical synthesis , Animals , Binding Sites , Crystallography, X-Ray , Dipeptides/chemistry , Drug Design , Enzyme Inhibitors/chemistry , Models, Molecular , Nitriles/chemistry , Rats , Structure-Activity RelationshipABSTRACT
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.
Subject(s)
Anti-HIV Agents , HIV Protease Inhibitors , HIV Protease/metabolism , Pyridines , Pyrones , Animals , Anti-HIV Agents/chemistry , Anti-HIV Agents/metabolism , Anti-HIV Agents/pharmacology , Cell Line , Cell Line, Transformed , Chromatography, High Pressure Liquid , Crystallography, X-Ray , HIV Protease Inhibitors/chemistry , HIV Protease Inhibitors/metabolism , HIV Protease Inhibitors/pharmacology , HIV-1/drug effects , HIV-1/enzymology , Humans , Hydrogen Bonding , Mice , Models, Molecular , Protein Binding , Pyridines/chemistry , Pyridines/metabolism , Pyridines/pharmacology , Pyrones/chemistry , Pyrones/metabolism , Pyrones/pharmacology , Stereoisomerism , Structure-Activity Relationship , SulfonamidesABSTRACT
Potent, non-peptidic, dihydropyrone sulfonamide HIV protease inhibitors have been previously described. Crystallographic analysis of dihydropyrone sulfonamide inhibitor/HIV protease complexes suggested incorporation of a second, C2 symmetry-related sulfonamide group. Selected bis-sulfonamide dihydropyrone analogues display high HIV protease inhibitory activity.
Subject(s)
HIV Protease Inhibitors/chemical synthesis , Pyrones/chemical synthesis , Sulfonamides/chemical synthesis , Dimerization , Drug Design , HIV Protease/metabolism , HIV Protease Inhibitors/chemistry , HIV Protease Inhibitors/pharmacology , HIV-1/enzymology , Indicators and Reagents , Models, Molecular , Molecular Conformation , Molecular Structure , Pyrones/chemistry , Pyrones/pharmacology , Structure-Activity Relationship , Sulfonamides/chemistry , Sulfonamides/pharmacologyABSTRACT
From a broad screening program, the 4-hydroxycoumarin phenprocoumon (I) was previously identified as a lead template with HIV protease inhibitory activity. The crystal structure of phenprocoumon/HIV protease complex initiated a structure-based design effort that initially identified the 4-hydroxy-2-pyrone U-96988 (II) as a first-generation clinical candidate for the potential treatment of HIV infection. Based upon the crystal structure of the 4-hydroxy-2-pyrone III/HIV protease complex, a series of analogues incorporating a 5,6-dihydro-4-hydroxy-2-pyrone template were studied. It was recognized that in addition to having the required pharmacophore (the 4-hydroxy group with hydrogen-bonding interaction with the two catalytic aspartic acid residues and the lactone moiety replacing the ubiquitous water molecule in the active site), these 5,6-dihydro-4-hydroxy-2-pyrones incorporated side chains at the C-6 position that appropriately extended into the S1' and S2' subsites of the enzyme active site. The crystal structures of a number of representative 5,6-dihydro-4-hydroxy-2-pyrones complexed with the HIV protease were also determined to provide better understanding of the interaction between the enzyme and these inhibitors to aid the structure-based drug design effort. The crystal structures of the ligands in the enzyme active site did not always agree with the conformations expected from experience with previous pyrone inhibitors. This is likely due to the increased flexibility of the dihydropyrone ring. From this study, compound XIX exhibited reasonably high enzyme inhibitory activity (Ki = 15 nM) and showed antiviral activity (IC50 = 5 microM) in the cell-culture assay. This result provided a research direction which led to the discovery of active 5,6-dihydro-4-hydroxy-2-pyrones as potential agents for the treatment of HIV infection.
