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

MULTIPLE SEQUENCE CHANGES WITHIN THE HIV-1 PROTEASE CONFER REDUCED SENSITIVITY TO A SYMMETRIC PROTEASE INHIBITOR

Int Wkshop HIV Drug Res 1994 Aug 2-5;3:6 (abstract no. 5)

Ron Swanstrom, Terri Smith, Steve Pettit, David Irlbeck, Wei Shao, Robert Wehbie, Ravi Sawhney, Lorri Everitt, and John Erickson*
Lineberger Cancer Center, University of North Carolina at Chapel Hill, USA; *NCI-Frederick Cancer Research and Development Center, Frederick, MD, USA

Previously we reported that selection for growth of HIV1 in the presence of the symmetric protease inhibitor A-77003 gave rise to variants with changes in the protease coding domain. The encoded changes include: Va13211e, Va18211e, Arg8Gln, Met46Leu, Met46Phe, and the double mutants Va13211e/Va18211e and Met46Phe/Val82Ala. Recombinant proteases with most of these changes have been made and shown to have an increased K_i for the inhibitor. These mutant enzymes cleave the viral Gag substrate in vitro similarly to the wild type protease, although with minor differences. Some of these changes have been incorporated into the viral genome and mutant viruses generated. The double mutant Va13211e/Va18211e has an approximately 10-fold increase in its IC₅₀. In an effort to define subsite tolerance to sequence change, we have used saturation mutagenesis at each subsite position to identify allowed substitutions.

We have carried out further selections with virus containing the double mutation Va13211e/Va18211e. The mutant virus has been passaged extensively without selection in an effort to generate second-site compensatory mutations. The mutant has also been passaged in the presence of increasing concentrations of A-77003 to select for higher level resistance. Virus has been selected which can grow in the presence of approximately 100 times the IC₅₀ value of inhibitor compared to the unmutated parental virus. Sequence changes associated with these two selections will be presented. These results suggest that virus evolution to higher level resistance can occur.

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1994-08-02
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