3rd International Workshop on HIV Drug Resistance 2-5 August 1994, Kauai, Hawaii, USA

LIMITED SEQUENCE DIVERSITY OF THE HIV-1 PROTEASE GENE FROM CLINICAL ISOLATES AND IN VITRO SUSCEPTIBILITY TO HIV PROTEASE INHIBITORS

Int Wkshop HIV Drug Res 1994 Aug 2-5;3:18 (abstract no. 17)

D.L. Winslow, R. King, S. Stack, H. Scarnati, and M.J. Otto
DuPont Merck Pharmaceutical Company, Glenolden, Pennsylvania and Wilmington, Delaware (USA)

The genome of HIV-1 encodes viral structural proteins and replicative enzymes that are translated as either p55 gag or p160 gag-pol polyproteins which must be proteolytically processed by the virus-encoded aspartyl protease (PR) in order to form infectious virions of HIV-1. Mutational analysis suggests that only a limited repertoire of mutations can be tolerated by the enzyme. PR sequence data from the few clinical isolates of HIV-1 that have been sequenced suggest that major regions of the PR gene are highly conserved. Proviral DNAs from three laboratory strains and 21 clinical isolates of HIV-1 were extracted from infected cells after proteinase K digestion and the protease gene was PCR amplified and sequenced directly by the Sanger method. In vitro susceptibilities of the virus isolates to protease inhibitors were determined by the ACTG/DoD consensus assay. Four different HIV protease inhibitors were tested including P9941, a C-2 symmetrical diol (DuPont Merck), A80987, an asymmetric mono-ol (Abbott), XM323, a cyclic urea (DuPont Merck), and R031-8959, a hydroxyethylene isostere (Roche). Maximum sequence variation was 10% at the nucleic acid level and 9% at the amino acid level. Purine-purine substitutions were most common. Five noncontiguous regions were conserved across all isolates and corresponded to amino acids 1-9 (amino terminal), 21-32 (active site). 47-56 (“flap” region), 78-88 (substrate binding region), and 94-99 (carboxy terminal). All clinical isolates demonstrated in vitro susceptibility to the protease inhibitors. There was no significant difference between the susceptibility of the reference strains and the clinical isolates. These data suggest that the variable regions of protease, as defined by these isolates, do not contain sites which are critical for interactions with the inhibitors tested.

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1994-08-02
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