Abstract
BackgroundRecommended regimens for HIV-positive individuals include the co-administration of dolutegravir (DTG) with two reverse transcriptase inhibitors (RTIs). Although rare, emerging resistance against DTG is often associated with the R263K substitution in integrase. In-vitro-selected R263K was associated with impaired viral replication capacity, DNA integration, and integrase strand-transfer activity, especially when accompanied by the secondary mutation H51Y. Given the reduced fitness of RTI-resistant viruses, we investigated potential impacts on viral replication of combining R263K and H51Y/R263K with major RTI-resistance substitutions including K65R, L74V, K103N, E138K, and M184I/V.ResultsWe combined the R263K or H51Y/R263K with RTI-resistance mutations into the proviral plasmid pNL4.3 and measured the resulting viral infectiousness, replication capacity, and ability to integrate viral DNA into host cells. Infectiousness was determined by luciferase assay in TZM-bl cells. Replicative capacity was monitored over 7 days and viral DNA integration was studied by real-time Alu-qPCR in PM1 cells. We found that viral infectiousness, replication capacities and integration levels were greatly reduced in triple mutants, i.e. H51Y/R263K plus a RT mutation, and moderately reduced in double mutants, i.e. R263K plus a RT mutation, compared to wild-type and single RT-mutant viruses.ConclusionsOur findings help to explain the absence of RTI mutations in individuals who experienced DTG-treatment failure.
Highlights
Recommended regimens for human immunodeficiency virus (HIV)-positive individuals include the co-administration of dolutegravir (DTG) with two reverse transcriptase inhibitors (RTIs)
The addition of DTG‐resistance substitutions reduces viral infectiousness in viruses containing the K65R, L74V, K103N, E138K or M184V/I reverse transcriptase mutations As previously hypothesized, the fitness cost of the H5Y/ R263K combination may have a benefit for patients under DTG treatment in terms of viral load
Our results show that the addition of R263K to the K65R, L74V, K103N, E138K, or M184I/V-harbouring viruses resulted in further moderate decreases in viral infectivity ranging from 1.23-fold to 3.17-fold (P < 0.05), depending on the RT backbone (Fig. 1a–f )
Summary
Recommended regimens for HIV-positive individuals include the co-administration of dolutegravir (DTG) with two reverse transcriptase inhibitors (RTIs). In-vitro-selected R263K was associated with impaired viral replication capacity, DNA integration, and integrase strand-transfer activity, especially when accompanied by the secondary mutation H51Y. Pham et al Retrovirology (2016) 13:31 the appearance of such secondary substitutions as H51Y, M50I, and E138K [7,8,9,10] None of the latter could compensate for the loss in replicative capacity conferred by R263K and, had the effect of further decreasing viral replication, integrase strand-transfer activity and levels of integrated DNA, even though the addition of H51Y to R263K increased resistance to DTG to about sevenfold, as measured by the PhenoSense® Integrase Replication Assay (7–10). Similar impacts on viral replicative capacity and DTG resistance in tissue culture and biochemical assays were observed when the E138K or M50I substitutions were introduced into R263K viruses. The H51Y/R263K and M50I/R263K viruses may confer similar levels of DTG resistance (about sevenfold resistance against DTG as measured in TZM-bl cells by luciferase assay) while the level of resistance is only about 4.3-fold for the E138K/R263K virus [7, 8, 10]
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