Background:  Acquired HIV-1 resistance to enfuvirtide (ENF) is primarily associated with mutations within the relatively conserved first heptad repeat (HR1) region of gp41. Viral Env sequences, however, are remarkably variable and could have an important effect on optimal expression of HR1 mutations. To assess the role of the Env genetic background in the development of ENF resistance, we have examined the evolution of whole Env sequences, Env fitness, and susceptibility to ENF in the course of ENF escape in treated patients. 

Methods:  Sequential plasma samples were obtained from 6 patients initiating ENF treatment as part of salvage therapy. Clinical follow-up ranged from 12 to 50 weeks and samples were obtained before, during, and after cessation of ENF treatment. Susceptibility to ENF and Env-mediated viral fitness of plasma-derived virus populations were measured using a recombinant phenotypic assay involving the entire gp120 and the gp41 ectodomain.

Results:  The susceptibility to ENF of baseline viruses was quite variable, ranging between 56 and 756 ng/mL. Regardless of these baseline values, viral populations with increasing phenotypic resistance to ENF were selected under treatment, until a critical threshold value of resistance (> 3000 ng/mL) was reached in patients treated for 20 or more weeks (4 of 6). As expected, ENF-resistant viruses harbored one or more HR1 mutations (positions 36, 38, and 43). Analysis of synonymous Env mutations in sequential samples showed that, in all patients, the HR1 mutations had emerged in the context of Env quasi-species that were different from those of the dominant populations present at baseline. In 4 patients, these novel Env sequences were characterized by increased fitness compared to baseline virus. In all patients, the further evolution of resistance, whether or not accompanied by selection of additional HR1 mutations, involved sequential emergence of populations with new Env genetic backgrounds.

Conclusions:  Mutations conferring ENF resistance are repeatedly selected in different Env genetic backgrounds, leading to the replacement of dominant virus populations over time. The whole Env genetic context thus appears to play a critical role in the expression and selection of HR1 mutations.

Keywords: Entry Inhibitor; Genetic background; Resistance