16th International HIV Drug Resistance Workshop 12-16 June 2007, Barbados

LOW LEVEL OF BASELINE RESISTANCE TO INTEGRASE INHIBITORS L731,988 AND L870,810 IN RANDOMLY SELECTED SUBTYPE B AND NON-B HIV-1 STRAINS

Antivir Ther. 2007; 12:S7 (abstract no. 5)

K Van Baelen, M Clynhens, E Rondelez, V Van Eygen, P Van den Zegel, H Vermeiren, I Vandenbroucke and LJ Stuyver
Virco BVBA, Mechelen, Belgium

BACKGROUND: HIV-1 integrase inhibitors (INIs) constitute a promising new class of anti-HIV drugs. The effect of naturally occurring polymorphisms in the integrase gene on the susceptibility to INIs L731,988 and L870,810 was investigated using a research phenotypic integrase resistance assay.

METHODS: The RT–RNaseH–IN region (2,898 bp) was PCR-amplified from viral RNA and recombined with an RT–RNaseH–IN-deleted, HXB2D-based backbone. Recombinant viruses (RVS) were produced by nucleofection of plasmids into MT4 cells and tested for their susceptibility to L731,988 and L870,810. Amplicons containing the T66I or E92Q INI resistance-associated mutation, and protease inhibitor TMC114 were used as controls.

RESULTS: In this assay, the IC₅₀ for HXB2 was 1.5 ±0.3 µM for L731,988 and 2.8 ±0.6 nM for L870,810. Phenotypic testing of E92Q and T66I mutants showed, respectively, a ~6-fold and ~2.5-fold decrease in susceptibility to L731,988. Activity of L870,810 towards E92Q was reduced by ~10-fold, while no fold change (FC) was observed towards T66I. From 47 patient samples (26 B – nine therapy-experienced; and 19 non-B – eight therapyexperienced), a total of 90 clonal RVS were derived (1–6 clones/sample). FC values for the 90 RVS on TMC114 were 0.65 ±0.25. RT-IN sequencing of the RVS showed 39% variable positions (111 out of 288 IN amino acids, excluding variability at the 289 stop codon). None of the positions in the HHCC motif, the LEDGF/p75 interaction site, and the catalytic DDE triad were found to be polymorphic. The following resistance-associated IN mutations were detected: V72I, T97A, A128T, V151I, K156N, V165I, V201I, T206S and S230N. These naturally occurring polymorphisms did not have an effect on susceptibility to L731,988 and L870,810. Only one sample (subtype B, RT-sensitive, V72I and L74I), showed a ~sixfold reduced susceptibility to L731,988, but not to L870,810.

CONCLUSIONS: The IC₅₀ and FC values for wild-type and integrase mutant viruses were in concordance with published data. Substantial genotypic variability was detected in clinical samples, but no significant FCs could be assigned. This assay contains the RT, RNaseH and IN genes, allowing the study of clinical significance towards naturally occurring polymorphisms and NRTI-, NNRTI- and INI-selected variants.

2007-06-12
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