Protease-mediated enzymatic hydrolysis and activation of aryl phosphoramidate derivatives of stavudine

• Published in: European journal of medicinal chemistry 2005-05, Vol.40 (5), p.452-466
• Main Authors: Venkatachalam, T.K., Samuel, P., Qazi, S., Uckun, F.M.
• Format: Article
• Language: English
• Subjects: Amides - chemical synthesis; Amides - chemistry; Amides - pharmacokinetics; Antibiotics. Antiinfectious agents. Antiparasitic agents; Antiviral agents; Biological and medical sciences; Biotransformation; Dideoxynucleotides; Distereoisomers; HIV; Hydrolysis; Magnetic Resonance Spectroscopy; Mechanism; Medical sciences; Peptide Hydrolases - metabolism; Pharmacology. Drug treatments; Phosphoric Acids - chemical synthesis; Phosphoric Acids - chemistry; Phosphoric Acids - pharmacokinetics; Prodrugs - pharmacokinetics; Proteases; Spectrophotometry, Ultraviolet; Stampidine; Stavudine - analogs & derivatives; Stavudine - chemical synthesis; Stavudine - metabolism; Stavudine - pharmacokinetics; Stereoisomerism; Thymidine Monophosphate - analogs & derivatives; Thymidine Monophosphate - metabolism
• ISSN: 0223-5234; 1768-3254
• DOI: 10.1016/j.ejmech.2004.11.015

Several proteases are capable of hydrolyzing the aryl substituted phosphoramidate derivatives of stavudine resulting in the formation of the active metabolite, alaninyl d4T monophosphate. Subtilisin Protease A, Subtilisin Griseus, Subtilisin Carlsberg, Papaya, Bacillus were amongst the most effective proteases in hydrolyzing stavudine derivatives and specificity of their activity was confirmed using several protease inhibitors to block the hydrolysis of these phosphoramidate derivatives. We found that these proteases exhibit chiral selectivity at the phosphorus center of stavudine derivatives. Our results indicate that cellular proteases may be responsible for the activation of these phosphoramidate derivatives. In addition, we show that the enzymatic hydrolysis takes place at the carboxymethyl ester side chain of these pro-drugs and the direct attack on the phosphorus center by these enzymes does not occur. Finally, we describe a novel activation pathway hitherto unknown for the activation and viral inhibitory characteristic shown by these phosphoramidate derivatives of stavudine.