Loss of Viral Fitness Associated with Multiple Gag and Gag-Pol Processing Defects in Human Immunodeficiency Virus Type 1 Variants Selected for Resistance to Protease Inhibitors In Vivo

Abstract

We examined the viral replicative capacity and protease-mediated processing of Gag and Gag-Pol precursors of human immunodeficiency virus (HIV) variants selected for resistance to protease inhibitors. We compared recombinant viruses carrying plasma HIV RNA protease sequences obtained from five patients before protease inhibitor therapy and after virus escape from the treatment. Paired pretherapy-postresistance reconstructed viruses were evaluated for HIV infectivity in a quantitative single-cycle titration assay and in a lymphoid cell propagation assay. We found that all reconstructed resistant viruses had a reproducible decrease in their replicative capacity relative to their parental pretherapy counterparts. The extent of this loss of infectivity was pronounced for some viruses and more limited for others, irrespective of the inhibitor used and of the level of resistance. In resistant viruses, the efficiency of Gag and Gag-Pol precursor cleavage by the protease was impaired to different extents, as shown by the accumulation of several cleavage intermediates in purified particle preparations. We conclude that protease inhibitor-resistant HIV variants selected during therapy have an impaired replicative capacity related to multiple defects in the processing of Gag and Gag-Pol polyprotein precursors by the protease.

Structural and kinetic analysis of drug resistant mutants of HIV‐1 protease

“…In addition, crystal structures of V82A, V82D and V82N mutants showed conformational changes compared to the wild‐type structure [32,33], which are likely to reduce enzyme activity. The low catalytic activity is consistent with the reduced infectivity and defects in polyprotein processing that have been observed for virus with mutations V82S or V82A [34].…”

Genetic heterogeneity of the NS3 protease gene in hepatitis C virus genotype 1 from untreated infected patients

“…The polymorphisms outside residues central to NS3 protease activity and substrate binding, even if they arise at random in the absence of selection by inhibitors, may prove to be accessory substitutions of resistance mutations contributing to clinical resistance to HCV protease inhibitor therapy. Moreover, resistance to future NS3 protease inhibitors could occur by mutations arising either in the protease gene itself or, as has been shown for HIV-1, in cleavage sites of the protein [Doyon et al, 1996;Zennou et al, 1998].…”

Crystal structures of HIV protease V82A and L90M mutants reveal changes in the indinavir‐binding site

“…However, single protease mutations and specific combinations can have lower viral infectivity than wild-type HIV. Drug-exposed HIV with multiple protease mutations, including resistant substitutions of M46I/G48V/L90M and F53L/A71V/V82A, produce defects in polyprotein processing and reduced viral infectivity [6]. Therefore, it is important to understand the molecular basis for the altered activity and structural changes of these resistant mutants as compared to the wild-type protease, in order to understand the molecular mechanism of resistance and to develop new antiviral therapies.…”