11th Conference on Retroviruses and Opportunistic Infections San Francisco, California - February 8 - 11, 2004

Conf Retroviruses Opportunistic Infect. 2004 Feb 8-11;11th:Abstract No. 621

J Vingerhoets¹, I De Baere¹, H Azijn¹, T Van den Bulcke¹, P McKenna², T Pattery², R Pauwels¹, and M-P de Béthune^1
1Tibotec, Mechelen, Belgium and ²Virco, Mechelen, Belgium

BACKGROUND: TMC125 is a potent next-generation NNRTI, active against wild type and NNRTI-resistant HIV-1. In vitro selection experiments have demonstrated an increased genetic barrier to development of resistance, associated with the presence of multiple mutations in the emerging strains (including changes at positions 100, 179, 181, and 194). Profiling of TMC125 against >5600 clinical isolates demonstrated that some infrequent mutations (K101P, Y181I) were associated with decreased phenotypic susceptibility. To study the effect of these mutations on TMC125 activity, a panel of 82 site-directed mutant viruses with single, double, or triple mutations was assembled. The prevalence of each mutation was determined in the routine clinical sample database at Virco.

METHODS: Mutant PR-RT sequences were constructed with the QuickChange Site-Directed Mutagenesis kit (Stratagene) starting from a pGEM Dnaïvector, containing the HXB2 PR-RT fragment. Site-directed mutant viruses were generated by recombination of the mutant PR-RT amplicon within the genetic background of HXB2. Phenotype and genotype was determined using the Antivirogram and VirtualPhenotype assays, respectively.

RESULTS: The mean wild-type EC₅₀ for TMC125 was 0.9 nM. Among 54 single mutant strains reflecting all currently known NNRTI-resistance mutations, only 3 demonstrated a fold change (FC) in EC₅₀ of >10, compared to wild type: Y181I (12.5 FC, p = 0.5%), Y181V (15.1 FC, p = 0.4%), and F227C (10.1 FC, p = 0.02%). Of 19 double mutants, 1 strain showed a FC >10: V179F+Y181C (129 FC, p = 0.2%). Other, more prevalent, combinations of 2 NNRTI mutations, including L100I, K103N, Y181C and G190A, showed a FC <10 for TMC125, whereas 11/19 showed a FC>10 and 4/19 a FC>100 for efavirenz. 4/9 triple mutants showed a FC >10 but <50 for TMC125. These strains contained L100I+K103N with either Y181C or T386A and K103N+Y181C with either V179I or Y318F. The prevalence of these triple mutants was <0.6%.

CONCLUSIONS: The susceptibility of TMC125 to specific single or multiple mutations, including those observed after in vitro selection or associated with decreased susceptibility in clinical isolates, was tested. The results show that the antiviral activity of TMC125 is not affected by the majority of these mutations, and that only less prevalent mutations are associated with decreased susceptibility to the compound. Long-term antiviral activity in patients with NNRTI-resistant HIV-1 is currently being evaluated in phase 2b trials.

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Copyright © 2004 - Foundation for Retrovirology and Human Health. Reproduction of this abstract (other than one copy for personal reference) must be cleared through the Foundation for Retrovirology and Human Health. Licensed (AIDSLINE) from National Library of Medicine.