Landowski_2006_J.Pharmacol.Exp.Ther_316_572

Carboxylesterases are among the best characterized prodrug-hydrolyzing enzymes involved in the activation of several therapeutic carbamate and ester prodrugs. The broad specificity of these enzymes makes them amenable for designing prodrugs. Porcine liver carboxylesterase 1 specificity for amino acid esters of three nucleoside analogs [floxuridine, gemcitabine, and 2-bromo-5,6-dichloro-1-(beta-D-ribofuranosyl) benzimidazole] was evaluated to assess optimal structural preferences for prodrug design. The amino acid promoiety and the esterification site influenced carboxylesterase hydrolysis rates up to 1164-fold and the binding affinity up to 26-fold. Carboxylesterase (CES) 1 exhibited high-catalytic efficiency hydrolyzing prodrugs containing a phenylalanyl moiety but was over 100-fold less efficient with valyl or isoleucyl prodrugs, regardless of the nucleoside or esterification site. CES1 catalytic efficiency was 2-fold higher with 5' phenylalanyl monoesters than the corresponding 3' esters of floxuridine. This preference was reversed with phenylalanyl gemcitabine prodrugs, evident from a 2-fold preference for 3' monoesters over 5' esters. The newly characterized esterase valacyclovirase was several hundred-fold more efficient (up to 19,000-fold) than carboxylesterase in hydrolyzing amino acid esters but similar in apparent binding affinity. The specific activities of the two enzymes with several amino acid ester prodrugs clearly suggest that initial hydrolysis rates are relatively low for prodrugs with isoleucyl, aspartyl, and lysyl promoieties for both enzymes compared with those with phenylalanyl, valyl, prolyl, and leucyl progroups. The low relative hydrolysis rates of isoleucyl, aspartyl, and lysyl prodrugs may facilitate prolonged systemic disposition of the nucleoside analogs for improved therapeutic action.