18th International HIV Drug Resistance Workshop Basic Principles & Clinical Implications June 9–13 2009, Fort Myers, Florida, USA

DRUG CONCENTRATION, SELECTIVE ADVANTAGE AND SELECTION DYNAMICS OF RALTEGRAVIR-RESISTANT MUTANTS

Antivir Ther 2009; 14 Suppl 1:A28 (abstract no. 26)

E Dam, R Quercia and F Clavel
Inserm U941, IUH, Hôpital Saint Louis, Paris, France

BACKGROUND: Emergence of HIV resistance to raltegravir (RAL) in treated patients can follow two main evolutionary pathways that are exclusive of each other. We have examined the mechanisms driving selection of RAL resistance using an approach that assesses the selective advantage of mutant viruses as a function of drug concentration. Because intracellular concentration of antiretroviral drugs is also key to selection of resistance, we also examined the intracellular kinetics of inhibition of HIV replication by RAL in tissue culture.

METHODS: Seven NL4-3-derived HIV-1 clones carrying single or double integrase resistance mutations were evaluated for RAL IC₅₀ , for RC and for their selective advantage profiles, as measured by the ratio of mutant to NL4-3 infectivity as a function of RAL concentration, in a single-cycle assay. To evaluate the intracellular kinetics of RAL activity, target cells were pretreated with RAL for 16 h and inoculated with wild-type HIV with drug still present in the culture or at different times following removal of the drug.

RESULTS: The mean fold-changes in RAL IC₅₀ relative to NL4-3 were E92Q=3.7, G140S=0.7, Q148H=78, N155H=31.5, E92Q+N155H=492, G140S+Q148H =1,436 and Q148H+N155H>30,000. Selective advantage curves revealed that among single mutants, N155H had the highest selective advantage over NL4-3 across concentrations of RAL ranging 1–500 nM. Despite a higher IC₅₀, Q148H showed a markedly lower and narrower selective advantage profile, while mutants E92Q and G140S did not show detectable advantage. Among double mutants, the strongest and widest selective advantage was seen for G140S+Q148H. In the second part of this study, evaluating the intracellular kinetics of RAL activity in tissue culture, we observed that when removed at any time before viral inoculation, RAL was no longer able to inhibit HIV replication, suggesting a short intracellular half-life.

CONCLUSIONS: Selective advantage curves explain why N155H can be selected early in the course RAL resistance evolution in vivo, but is later replaced by genotypes that include Q148HKR. Intracellular half-life of RAL, as measured by its antiviral activity, is short and comparable to that of NNRTIs.

2009-06-09
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