DESIGN AND SYNTHESIS OF SUBSTRATE-BASED PEPTIDOMIMETIC HUMAN-IMMUNODEFICIENCY-VIRUS PROTEASE INHIBITORS CONTAINING THE HYDROXYMETHYLCARBONYL ISOSTERE

The human immunodeficiency virus (HIV) codes for an aspartic protease known to be essential for retroviral maturation and replication. The H IV protease can recognize Phe-Pro and Tyr-Pro sequences as the virus-s pecific cleavage site. These features provided a basis for the rationa l design of selective HIV protease-targeted drugs for the treatment of acquired immunodeficiency syndrome (AIDS). HIV protease is formed fro m two identical 99 amino acid peptides. We replaced the two Cys residu es by L-Ala to synthesize [Ala(67,95)]-HIV-1 protease by the solid pha se method and then prepared [Tyr(6,42), Nle(36,46), (NHCH2COSCH2CO)(51 -52), Ala(67,95)] HIV- protease (NY-5 isolate) using the thioester che mical ligation method. Based on the substrate transition state, we des igned and synthesized a novel class of HIV protease inhibitors contain ing an unnatural amino acid, (2S, 3S)-3-amino-2-hydroxy-4-phenylbutyri c acid, named allophenylnorstatine (Apns) with a hydroxymethylcarbonyl (HMC) isostere. Among them, the conformationally constrained tripepti de kynostatin (KNI)-272 (iQoa-Mta-Apns-Thz-NHBu(t)) was a highly selec tive and superpotent HIV protease inhibitor (Ki = 0.0055 nM). KNI-272 exhibited potent antiviral activities against both AZT-sensitive and - insensitive clinical HIV-1 isolates as well as HIV-2 with low cytotoxi city. After i.d. administration, bioavailability of KNI-272 was 42.3% in rats. Also, KNI-272 exhibited in vivo anti-HIV activities in human PBMC-SCID mice. The x-ray crystallography and molecular modeling studi es showed that the HMC group in KNI-272 inter-acted excellently with t he aspartic acid carboxyl groups of HIV protease active site in the es sentially same hydrogen-bonding mode as the transition state. This res ult implies that the HMC isostere is an ideal transition-state mimic a nd contributes to the high activity of KNI-272. (C) 1996 John Wiley & Sons, Inc.