The Human Immunodeficiency virus type 1 (HIV-1) protease (PR) is a key enzyme in viral replication and a major target for therapeutic intervention. Protease inhibitors (PI) are the backbone of some of the most active combinations of antiretroviral drugs used in the treatment of HIV-infected patients. The long-term efficacy of these compounds, however, is threatened by the emergence of viral resistance and subsequent spread of resistant virus. HIV-1 resistance to PIs is promoted by gradual accumulation of amino-acid substitutions in PR, resulting in altered PI binding [1]–[5]. Resistance-promoting changes in PR generally also decrease viral replicative capacity (RC) due to decreased processing of the natural substrate. Accordingly, additional mutations accumulate in PR over time that mainly compensate for these losses in RC, but may also contribute to resistance directly [1]–[3],[6]. The biological effect of PI resistance mutations thus has to be viewed as the product of their effect on enzyme inhibition (resistance) and on enzyme activity (RC).
Besides mutations directly affecting PR, several mutations in the Gag polyprotein, the main substrate of PR, have been found to play a significant role in the evolution of PI resistance [7]–[13]. These mutations were generally classified as compensatory mutations that restore activity of the mutated PR for its natural substrate [7]–[9],[12],[14],[15]. Particular attention has been turned to mutations in the region surrounding the NC-SP2-P6 cleavage sites at the C-terminus of Gag. Figure 1 gives an overdrive of these mutations and of their position in the NC-SP2-p6 region of HIV-1 Gag. The most frequently observed cleavage site mutations in this region are substitution A431V, located at position P2 of the NC-SP2 cleavage site, mutation L449F at position P1' of the SP2-P6 cleavage site, and mutations K436R and I437V, situated immediately downstream of the NC-SP2 site. Interestingly, some of these mutations appear to depend upon the presence of specific mutations in PR: Mutation A431V is mostly observed in PI-resistant viruses carrying mutations V82A and/or M46I in PR [16]. Mutation L449F is frequently seen in viruses with mutation I84V in PR [16]. Neither of these two substitutions are seen in wild-type, protease inhibitor-naïve viruses. Substitution P453L is a polymorphism found in some inhibitor-naïve viruses. It is, however, seen with significantly higher frequency in resistant viruses carrying the I84V mutation in PR [16] and also specifically seen in resistant viruses carrying the I50V PR mutation, in association with L449F [12].