Novel Mutational Changes Involved in Delayed Development of Dolutegravir Resistance in HIV-1 B and non-B Subtypes during in vitro Drug Selection
Maureen Oliveira*1, D Moisi1, R-I Ibanescu1, B Spira1, B Brenner1, T Fujiwara2, M Underwood3, and M Wainberg1
1McGill Univ AIDS Ctr, Montreal, Canada; 2Shionogi & Co Ltd, Osaka, Japan; and 3ViiV Hlthcare, NC, US
Background: Despite the superior genetic barrier of dolutegravir (DTG), the outcome of treatment with this second-generation integrase inhibitor remains unknown. In this study, clinical isolates of HIV-1 B and non-B subtypes were subjected to DTG pressure in vitro, to determine whether any subtype-specific mutational profiles might arise.
Methods: Viruses derived from clinical isolates of subtypes B, CRF2_AG, and C were cultured in cord blood mononuclear cells in the presence of DTG and elvitegravir (EGV). The drug concentration was increased gradually to permit the emergence of resistant variants. Viral RNA was extracted from cell culture supernatants and sequenced to reveal changes in the integrase gene. Cell-based phenotypic assays were performed to determine drug sensitivity.
Results: After 20 weeks, isolates selected with EGV adhered to established resistance pathways including mutations conferring high-level resistance. The concentrations of EGV attained were 2.5- to 50-fold higher relative to the EC50 than those obtained with DTG. After 20 weeks of DTG pressure, the R263K mutation arose in all subtype B isolates and 1 AG isolate. The G118R mutation, previously implicated in resistance to other integrase inhibitors, was found in 2 non-B subtypes. Additional changes occurred at positions 151,153, and 288. Genotypic analysis was repeated at week 37, even though further increases in DTG concentrations were minimal. In 1 subtype B virus, a mixture including 138 E/K was detected in addition to R263K. Another B variant displayed a combination of M50I, V151I, and R263K. In the non-B subtypes, the addition of the H51Y mutation was most prevalent. Single mutants containing either the R263K or G118R mutations conferred slight reductions in DTG drug susceptibility (approximately 1.6-fold). Other variants harboring 2 or more mutations displayed higher level resistance (2- to 9-fold).
Conclusions: This study confirms recent observations that the second-generation integrase inhibitor DTG possesses a high genetic barrier for the development of resistance mutations and that subtype differences may play a role in the resulting resistance profiles. In addition, we have observed novel mutational changes in both B and non-B subtypes, concomitant with unique patterns of DTG resistance. While viruses treated with EGV easily escaped drug pressure via predicted pathways, DTG pressure proved more difficult to overcome, resulting in a delayed response.