Retracing the evolutionary pathways of human immunodeficiency virus type 1resistance to protease inhibitors: Virus fitness in the absence and in thepresence of drug

Human immunodeficiency virus type 1 (HIV-1) resistance to protease inhibito rs (P1) is a major obstacle to the full success of combined antiretroviral therapy, High-level resistance to these compounds is the consequence of ste pwise accumulation of amino acid substitutions in the HIV-1 protease (PR), following pathways that usually differ from one inhibitor to another. The s elective advantage conferred by resistance mutations may depend upon severa l parameters: the impact of the mutation on virus infectivity in the presen ce or absence of drug, the nature of the drug, and its local concentration. Because drug concentrations in vivo are subject to extensive variation ol; er time and display a markedly uneven tissue distribution, the parameters o f selection for HIV-1 resistance to PI in treated patients are complex and poorly understood. In this study, we have reconstructed a large series of H IV-1 mutants that early single or combined mutations in the PR, retracing t he accumulation pathways observed in ritonavir-, indinavir-, and saquinavir -treated patients. We have then measured the phenotypic resistance and the drug-free infectivity of these mutant viruses. A deeper insight into the ev olutionary value of HIV-1 PR mutants fame from a novel assay system designe d to measure the replicative advantage of mutant viruses as a function of d rug concentration. By tracing the resultant fitness profiles we determined the range of drug concentrations for which mutant viruses displayed a repli cative advantage over the wild type and the extent of this advantage. Fitne ss profiles were fully consistent with the order of accumulation of resista nce mutations observed in treated patients and further emphasise the key im portance of local drug concentration in the patterns of selection of drug-r esistant HIV-1 mutants.