Intact HIV Proviruses Persist in Children Seven to Nine Years after Initiation of Antiretroviral Therapy in the First Year of Life

• Published in: Journal of virology Vol. 94; no. 4
• Main Authors: Katusiime, Mary Grace, Halvas, Elias K, Wright, Imogen, Joseph, Kevin, Bale, Michael J, Kirby-McCullough, Bronwyn, Engelbrecht, Susan, Shao, Wei, Hu, Wei-Shau, Cotton, Mark F, Mellors, John W, Kearney, Mary F, van Zyl, Gert U
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 31.01.2020
• Subjects: Anti-Retroviral Agents - therapeutic use; Antiretroviral Therapy, Highly Active - methods; CD4-Positive T-Lymphocytes - virology; Child; Cohort Studies; DNA, Viral - blood; Female; Genome and Regulation of Viral Gene Expression; HIV Infections - genetics; HIV Infections - virology; HIV-1 - genetics; HIV-1 - pathogenicity; Humans; Leukocytes, Mononuclear - virology; Male; Proviruses - genetics; Sequence Analysis, DNA - methods; South Africa; Viral Load - genetics; Viral Load - methods; South Africa; HIV full-length sequencing; HIV persistence children; HIV intact provirus; HIV persistence; human immunodeficiency virus; HIV reservoir decay; HIV early ART children; HIV reservoir; HIV cure; HIV proviral landscape
• ISSN: 0022-538X; 1098-5514; 1098-5514
• DOI: 10.1128/JVI.01519-19

In adults starting antiretroviral therapy (ART) during acute infection, 2% of proviruses that persist on ART are genetically intact by sequence analysis. In contrast, a recent report in children treated early failed to detect sequence-intact proviruses. In another cohort of children treated early, we sought to detect and characterize proviral sequences after 6 to 9 years on suppressive ART. Peripheral blood mononuclear cells (PBMC) from perinatally infected children from the Children with HIV Early antiRetroviral (CHER) study were analyzed. Nearly full-length proviral amplification and sequencing (NFL-PAS) were performed at one time point after 6 to 9 years on ART. Amplicons with large internal deletions were excluded (<9 kb). All amplicons of ≥9 kb were sequenced and analyzed through a bioinformatic pipeline to detect indels, frameshifts, or hypermutations that would render them defective. In eight children who started ART at a median age of 5.4 months (range, 2.0 to 11.1 months), 733 single NFL-PAS amplicons were generated. Of these, 534 (72.9%) had large internal deletions, 174 (23.7%) had hypermutations, 15 (1.4%) had small internal deletions, 3 (1.0%) had deletions in the packaging signal/major splice donor site, and 7 (1.0%) were sequence intact. These 7 intact sequences were from three children who initiated ART after 2.3 months of age, one of whom had two identical intact sequences, suggestive of a cell clone harboring a replication-competent provirus. No intact proviruses were detected in four children who initiated ART before 2.3 months of age. Rare, intact proviruses can be detected in children who initiate ART after 2.3 months of age and are probably, as in adults, maintained by clonal expansion of cells infected before ART initiation. There are limited data about the proviral landscape in children exhibiting long-term suppression after early treatment, particularly in Sub-Saharan Africa where HIV-1 subtype C predominates. Investigating the sequence-intact reservoir could provide insight on the mechanisms by which intact proviruses persist and inform ongoing cure efforts. Through nearly full-length proviral amplification and sequencing (NFL-PAS), we generated 733 NFL-PAS amplicons from eight children. We showed that rare, genetically intact proviruses could be detected in children who initiated ART after 2.3 months of age. The frequency of intact proviruses was lower (  < 0.05) than that reported for HIV subtype B-infected adults treated during early HIV infection. We show that cells harboring genetically intact HIV proviruses are rare in children exhibiting long-term suppression after early treatment and may require the processing of a large number of cells to assess reservoir size. This points to the need for efficient methods to accurately quantify latent reservoirs, particularly in pediatric studies where sample availability is limited.