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Binding energetics analysis shows a unique response of tipranavir to HIV-1 protease mutations associated with drug resistance
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Background: Tipranavir (TPV) is a potent non-peptidic HIV-1 protease inhibitor currently in phase III clinical trials. We studied the inhibitory potency of TPV to the wild type protease and to commonly occurring drug-resistant mutations, including those associated with TPV resistance. Thermodynamics and crystallographic analysis elucidated the mechanisms by which TPV achieves its high affinity and eludes the deleterious effects of mutations.
Methods: The inhibitory potency and binding energetics of TPV and other protease inhibitors were determined by enzyme inhibition and microcalorimetric measurements. The structures of TPV with wild type and mutant proteases were determined by x-ray crystallography.
Results: TPV inhibits the wt protease with a Ki lower than 20 pM and exhibits extremely low loss of potency against known multi-drug resistant mutations. Mutations evaluated here included the active site mutants I50V and V82F/I84V; multidrug resistant L10I/L33I/M46I/I54V/L63I/V82A/I84V/L90M and M46I/A71V/G73S/V82A/I84V/L90M; and the TPV-selected mutant I13V/V32L/L33F/K45I/V82L/I84V. Against the TPV-selected mutant, only lopinavir had a 4-fold better response. Against all other mutants TPV averaged 30-fold lower Ki’s than indinavir, amprenavir, lopinavir and atazanavir. Structurally, TPV establishes a very strong H-bond network with invariant regions, including catalytic Asp25 and backbone of Asp29, Asp30, Gly48 and Ile50. Uniquely, TPV H-bonds directly to Ile50. The binding of TPV to the wt protease is driven by a combination of favorable enthalpic (-0.7 kcal/mol) and entropic contributions (-13.9 kcal/mol). Against mutations, TPV either gains or loses very little enthalpic interactions
Conclusions: Most inhibitors lose affinity against mutations due to combined enthalpy and entropy loses. Good response to mutations is achieved when enthalpy and entropy changes mutually compensate each other. Inhibitors that respond well usually compensate enthalpic loses with entropic gains. TPV is the first inhibitor that has shown the ability to maintain high inhibitory potency by either gaining or sustaining minimal loses in binding enthalpy.
Methods: The inhibitory potency and binding energetics of TPV and other protease inhibitors were determined by enzyme inhibition and microcalorimetric measurements. The structures of TPV with wild type and mutant proteases were determined by x-ray crystallography.
Results: TPV inhibits the wt protease with a Ki lower than 20 pM and exhibits extremely low loss of potency against known multi-drug resistant mutations. Mutations evaluated here included the active site mutants I50V and V82F/I84V; multidrug resistant L10I/L33I/M46I/I54V/L63I/V82A/I84V/L90M and M46I/A71V/G73S/V82A/I84V/L90M; and the TPV-selected mutant I13V/V32L/L33F/K45I/V82L/I84V. Against the TPV-selected mutant, only lopinavir had a 4-fold better response. Against all other mutants TPV averaged 30-fold lower Ki’s than indinavir, amprenavir, lopinavir and atazanavir. Structurally, TPV establishes a very strong H-bond network with invariant regions, including catalytic Asp25 and backbone of Asp29, Asp30, Gly48 and Ile50. Uniquely, TPV H-bonds directly to Ile50. The binding of TPV to the wt protease is driven by a combination of favorable enthalpic (-0.7 kcal/mol) and entropic contributions (-13.9 kcal/mol). Against mutations, TPV either gains or loses very little enthalpic interactions
Conclusions: Most inhibitors lose affinity against mutations due to combined enthalpy and entropy loses. Good response to mutations is achieved when enthalpy and entropy changes mutually compensate each other. Inhibitors that respond well usually compensate enthalpic loses with entropic gains. TPV is the first inhibitor that has shown the ability to maintain high inhibitory potency by either gaining or sustaining minimal loses in binding enthalpy.