Search

Background: Increasing first-line treatment failures in low- and middle-income countries (LMICs) have led to increased use of integrase strand transfer inhibitors (INSTIs) such as dolutegravir. However, HIV-1 susceptibility to INSTIs in LMICs, especially with previous raltegravir exposure, is poorly understood due to infrequent reporting of INSTI failures and testing for INSTI drug resistance mutations (DRMs).Methods: A total of 51 non-subtype B HIV-1 infected patients failing third-line (raltegravir-based) therapy in Uganda were initially selected for the study. DRMs were detected using Sanger and deep sequencing. HIV integrase genes of 13 patients were cloned and replication capacities (RCs) and phenotypic susceptibilities to dolutegravir, raltegravir and elvitegravir were determined with TZM-bl cells. Spearman's correlation coefficient was used to determine cross-resistance between INSTIs.Results: INSTI DRMs were detected in 47% of patients. HIV integrase-recombinant virus carrying one primary INSTI DRM (N155H or Y143R/S) was susceptible to dolutegravir but highly resistant to raltegravir and elvitegravir (>50-fold change). Two patients, one with E138A/G140A/Q148R/G163R and one with E138K/G140A/S147G/Q148K, displayed the highest reported resistance to raltegravir, elvitegravir and even dolutegravir. The former multi-DRM virus had WT RC whereas the latter had lower RCs than WT.Conclusions: In HIV-1 subtype A- and D-infected patients failing raltegravir and harbouring INSTI DRMs, there is high-level resistance to elvitegravir and raltegravir. More routine monitoring of INSTI treatment may be advised in LMICs, considering that multiple INSTI DRMs may have accumulated during prolonged exposure to raltegravir during virological failure, leading to high-level INSTI resistance, including dolutegravir resistance. [ABSTRACT FROM AUTHOR]

Background: Thymidine analogs, namely AZT (Zidovudine or Retrovir™) and d4T (Stavudine or Zerit™) are antiretroviral drugs still employed in over 75% of first line combination antiretroviral therapy (cART) in Kampala, Uganda despite aversion to prescribing these drugs for cART in high income countries due in part to adverse events. For this study, we explored how the continued use of these thymidine analogs in cART could impact emergence of drug resistance and impact on future treatment success in Uganda, a low-income country. Methods: We examined the drug resistance genotypes by Sanger sequencing of 262 HIV-infected patients failing a first line combined antiretroviral treatment containing either AZT or d4T, which represents approximately 5% of the patients at the Joint Clinical Research Center receiving a AZT or d4T containing treatment. Next generation sequencing (DEEPGEN™HIV) and multiplex oligonucleotide ligation assays (AfriPOLA) were then performed on a subset of patient samples to detect low frequency drug resistant mutations. CD4 cell counts, viral RNA loads, and treatment changes were analyzed in a cohort of treatment success and failures. Results: Over 80% of patients failing first line AZT/d4T-containing cART had predicted drug resistance to 3TC (Lamivudine) and non-nucleoside RT inhibitors (NNRTIs) in the treatment regimen but only 45% had resistance AZT/d4T associated resistance mutations (TAMs). TAMs were however detected at low frequency within the patients HIV quasispecies (1-20%) in 21 of 34 individuals who were failing first-line AZT-containing cART and lacked TAMs by Sanger. Due to lack of TAMs by Sanger, AZT was typically maintained in second-line therapies and these patients had a low frequency of subsequent virologic success. Conclusions: Our findings suggest that continued use of AZT and d4T in first-line treatment in low-to-middle income countries may lead to misdiagnosis of HIV-1 drug resistance and possibly enhance a succession of second- and third-line treatment failures. [ABSTRACT FROM AUTHOR]

The role of HIV-1 minority variants on transmission, pathogenesis, and virologic failure to antiretroviral regimens has been explored; however, most studies of low-level HIV-1 drug-resistant variants have focused in single target regions. Here we used a novel HIV-1 genotypic assay based on deep sequencing, DEEPGEN (Gibson et al 2014 Antimicrob Agents Chemother 58∶2167) to simultaneously analyze the presence of minority variants carrying mutations associated with reduced susceptibility to protease (PR), reverse transcriptase (RT), and integrase strand transfer integrase inhibitors (INSTIs), as well as HIV-1 coreceptor tropism. gag-p2/NCp7/p1/p6/pol-PR/RT/INT and env/C2V3 PCR products were obtained from twelve heavily treatment-experienced patients experiencing virologic failure while participating in a 48-week dose-ranging study of elvitegravir (GS-US-183-0105). Deep sequencing results were compared with (i) virological response to treatment, (ii) genotyping based on population sequencing, (iii) phenotyping data using PhenoSense and VIRALARTS, and (iv) HIV-1 coreceptor tropism based on the phenotypic test VERITROP. Most patients failed the antiretroviral regimen with numerous pre-existing mutations in the PR and RT, and additionally newly acquired INSTI-resistance mutations as determined by population sequencing (mean 9.4, 5.3, and 1.4 PI- RTI-, and INSTI-resistance mutations, respectively). Interestingly, since DEEPGEN allows the accurate detection of amino acid substitutions at frequencies as low as 1% of the population, a series of additional drug resistance mutations were detected by deep sequencing (mean 2.5, 1.5, and 0.9, respectively). The presence of these low-abundance HIV-1 variants was associated with drug susceptibility, replicative fitness, and coreceptor tropism determined using sensitive phenotypic assays, enhancing the overall burden of resistance to all four antiretroviral drug classes. Further longitudinal studies based on deep sequencing tests will help to clarify (i) the potential impact of minority HIV-1 drug resistant variants in response to antiretroviral therapy and (ii) the importance of the detection of HIV minority variants in the clinical practice. [ABSTRACT FROM AUTHOR]