A detailed characterisation of drug resistance during darunavir/ritonavir (DRV/r) monotherapy highlights a high barrier to the emergence of resistance mutations in protease but identifies alternative pathways of resistance

Abstract

BackgroundMaintenance monotherapy with DRV/r has yielded variable outcomes and is not recommended. Trial samples offer valuable opportunities for detailed studies. We analysed samples from a 48-week trial in Cameroon to obtain a detailed characterisation of drug resistance. MethodsFollowing failure of NNRTI-based therapy and virological suppression on PI-based therapy, participants were assigned to receive either DRV/r (n=81) or TDF/3TC + LPV/r (n=39). PBMC from study entry underwent bulk protease and RT sequencing. Plasma collected at virological rebound (confirmed or last available HIV-1 RNA >60 copies/ml) underwent ultradeep protease and RT sequencing and bulk gag-protease sequencing. A site-directed mutant with T375A (p2/p7) was phenotypically characterised using a single-cycle assay. ResultsHIV-1 DNA analysis revealed NRTI and NNRTI resistance-associated mutations (RAMs) in 52/90 (57.8%) and 53/90 (58.9%) samples, respectively. In rebound HIV-1 RNA (DRV/r n=21; LPV/r n=2), prevalence was 9/23 (39.1%) and 10/23 (43.5%), respectively, with most RAMs occurring at frequencies [≥]15%. No DRV RAMs were found. Paired HIV-1 DNA and RNA sequences showed partial consistency in resistance patterns. Among 8 participants experiencing virological rebound on DRV/r (n=12 samples), all showed gag mutations associated with PI-exposure, including T375N, T375A (p2/p7), K436R (p7/p1), and mutations in p17, p24, p2 and p6. T375A conferred 10-fold DRV resistance and increased replication capacity. ConclusionsThe study highlights the high resistance barrier of DRV/r while identifying alternative pathways of DRV resistance through gag substitutions. During virological suppression, resistance patterns in HIV-1 DNA reflect treatment history, but due to technical and biological considerations, cautious interpretation is warranted.