Added Value of Next Generation Sequencing in Characterizing the Evolution of HIV-1 Drug Resistance in Kenyan Youth

Abstract

Drug resistance remains a global challenge in children and adolescents living with HIV (CALWH). Characterizing resistance evolution, specifically using next generation sequencing (NGS) can potentially inform care, but remains understudied, particularly in antiretroviral therapy (ART)-experienced CALWH in resource-limited settings. We conducted reverse-transcriptase NGS and investigated short-and long-term resistance evolution and its predicted impact in a well-characterized cohort of Kenyan CALWH failing 1st-line ART and followed for up to ~8 years. Drug resistance mutation (DRM) evolution types were determined by NGS frequency changes over time, defined as evolving (up-trending and crossing the 20% NGS threshold), reverting (down-trending and crossing the 20% threshold) or other. Exploratory analyses assessed potential impacts of minority resistance variants on evolution. Evolution was detected in 93% of 42 participants, including 91% of 22 with short-term follow-up, 100% of 7 with long-term follow-up without regimen change, and 95% of 19 with long-term follow-up with regimen change. Evolving DRMs were identified in 60% and minority resistance variants evolved in 17%, with exploratory analysis suggesting greater rate of evolution of minority resistance variants under drug selection pressure and higher predicted drug resistance scores in the presence of minority DRMs. Despite high-level pre-existing resistance, NGS-based longitudinal follow-up of this small but unique cohort of Kenyan CALWH demonstrated continued DRM evolution, at times including low-level DRMs detected only by NGS, with predicted impact on care. NGS can inform better understanding of DRM evolution and dynamics and possibly improve care. The clinical significance of these findings should be further evaluated.

Keywords: HIV-1 drug resistance; children and adolescents; evolution of drug resistance; minority drug resistance variants; types of drug resistance evolution.

Figure 1

Participants with evolving and reverting NRTI DRMs. This figure presents NRTI DRM evolution and its predicted impact in study participants with short-term evolution (panel (A)), long-term evolution without regimen change (panel (B)), and long-term evolution with regimen change (panel (C)). Each row represents one study participant and includes regimens at earlier (TP1) and later (TP2) time points (first column); identified cumulative DRMs at TP1 and TP2 (second column); and predicted resistance scores (numbers) and levels (colors) at TP1 and TP2, depicted in blocks in the third column on the right, to NRTI drugs abacavir (ABC), zidovudine (AZT), lamivudine and emtricitabine (XTC) and tenofovir disoproxil fumarate (TDF). Identified DRMs are color-coded by evolutionary type according to the legend at the top left, including evolving DRMs in red; reverting DRMs in blue; and all other DRMs (including stable, and increasing/decreasing established/minority DRMs) in black. NGS mutation frequencies for the evolving and reverting DRMs are indicated by the number following the underscores (<1 = undetectable). Predicted resistance levels for each drug are presented as color-coded blocks according to the legend at the top right from green (susceptible) to high-level predicted resistance (red). In panel (C), the first column regimens are at TP1 and the second column regimens are at TP2.

Figure 2

Participants with evolving and reverting NNRTI DRMs. This figure presents NNRTI DRM evolution and its predicted impact in study participants with short-term evolution (panel (A)), long-term evolution without regimen change (panel (B)), and long-term evolution with regimen change (panel (C)). For details and color-coding, see legend to Figure 1.

Figure 3

Difference in DRM prevalence at TP2 among participants taking a drug with penalty score in the short- and long-term studies (Exploratory Analysis 1). The Y-axis shows the estimated difference in TP2 DRM prevalence for participants taking at least one antiretroviral with a positive penalty score versus not taking an antiretroviral with a positive penalty score at TP2. Differences are presented for participants with TP1 DRM prevalence of 0, 15 or 80%. (A) NRTI DRMs at short- and long-term study (see details in Supplementary Table S3). (B) NNRTI DRMs at long-term study (see details in Supplementary Table S4).

Figure 4

Difference in predicted resistance score at TP2 among participants with a minor drug resistance variant at TP1 with penalty score in the short- (left panel) and long-term (right panel) study (Exploratory Analysis 2). The X-axis shows the estimated difference at TP2 in resistance score for participants with at least one minority drug resistance variant at TP1 with positive penalty score versus those without a minority drug resistance variant with a positive penalty score. Contrasts are presented for each drug. The drugs are sorted by the median difference.