The new and less toxic protease inhibitor saquinavir–NO maintains anti-HIV-1 properties in vitro indistinguishable from those of the parental compound saquinavir

Although, the antiviral activity, tolerability and convenience of protease inhibitors have improved significantly in recent years, toxicity-associated adverse events including diarrhea, lipid alterations, disturbance of glucose homeostasis and liver enzyme elevations still remain a major concern during treatment of HIV-1 patients. We have recently shown that the covalent attachment of the NO moiety to the HIV-1 protease inhibitor saquinavir (Saq–NO) reduces its toxicity. In this study, we evaluated in vitro the anti-HIV activity of Saq–NO vs. its parental compound Saq. Site directed mutants with the most frequently identified Saq associated resistance mutations and their combinations were generated on proviral AD8-based backbones. Phenotypic assays were conducted using wild type clinical isolates and fully replicating recombinant viruses with Saq and Saq–NO in parallel on purified CD4+ T cells. The following recombinant viruses were generated and tested: L33F, M46I, G48V, I54V, I84V + L90M, M46I + L90M, G48V + L90M, M46I + I54V + L90M, L33F + M46I + L90M. The fold change resistance compared to the wild type viruses was between 1.3 and 7 for all single mutants, between 3.4 and 20 for double mutants and between 16.7 and 28.5 for viruses carrying three mutations for both compounds. The results clearly demonstrate that Saq–NO maintains an anti-HIV-1 profile very similar to that of Saq. The possibility to reduce Saq associated side effects and to increase the concentration of the drug in vivo may allow a higher and possibly more effective dosage of Saq–NO in HIV-1-infected patients and to increase the genetic barrier of this PI thus impairing the selection of resistant clones.

► The covalent addition of a NO group to the protease inhibitor Saq has not modified its antiviral activity.
► Since Saq–NO is less toxic than Saq both in vitro and in vivo, larger doses of Saq–NO could be used for a more effective treatment of HIV-1 patients.
► The reduced toxicity of Saq–NO may limit side effects at the current daily dosage or increase the genetic barrier of the PI based regimen if used at higher dosage.