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Decoding the Somatic Mutation Landscape in Persisting HIV-1 Infected CD4+ T Clones
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CROI 2026 Feb 22-25 Denver
Program Abstract
Marie Armani-Tourret, Ce Gao, Maxime Bellefroid, Chloe Naasz, Toong Seng Tan,
Xu Yu, Mathias Lichterfeld
Ragon Institute of MGH, MIT and Harvard, Cambridge, MA, USA
Background: The long-term persistence of HIV-1 infected CD4+T cell clones is a major barrier to cure strategies. However, mechanisms contributing to their ability to survive long-term and resist immune elimination are poorly understood. Here, we comprehensively characterized the somatic mutation landscape of HIV-1 infected and uninfected CD4+T cell clones isolated from five ART-treated persons living with HIV.
Methods: Single HIV-1 infected cells were sorted based on intracellular P24 protein expression following stimulation with Tat-mRNA. HIV-uninfected CMV-specific tetramer-positive CD4+T cells were sorted for comparison. Single cells from both groups underwent whole-genome amplification; HIV-infected CD4+T clonal cells (n=13 clones) were identified based on proviral sequence identity and integration site analysis, while clonality of uninfected clones (n=10 clones) was assessed by TCR sequencing. Single-cell Whole-Exome Sequencing was performed on a total of 375 single cells (median of 10 cells per infected
clone and 7.5 cells per uninfected clone). Autologous bulk memory CD4+T cells were analyzed as reference populations. Somatic mutations were identified using the SCcaller algorithm on sequencing data generated on the Illumina platform.
Results: We identified 272 high-confidence somatic mutations, including 162 in HIV-1 infected clones and 110 in uninfected clones, that selectively occurred in clonal cell populations but were absent in bulk memory CD4+T cells. Each clone displayed a distinct somatic mutation profile, indicating independent evolutionary trajectories. HIV-1 infected clones harbored significantly more non-synonymous coding variants than uninfected clones (75.9% vs 65.4%, p=0.016), suggesting more effective selection for functionally impactful mutations. An unbiased pathway analysis of the mutated genes showed that in HIV-1 infected clones, non-synonymous mutations were enriched in genes involved in WNT/β-catenin signaling, senescence and DNA repair pathways,
whereas no such enrichment was observed in uninfected clones. Interestingly, we observed a strong mutational bias toward these pathways across multiple independent clones and study persons.
Conclusions: This study provides the first characterization of the somatic mutation landscape in persistent HIV-1 infected CD4+T cell clones. The enrichment of non-synonymous mutations in survival-associated pathways suggests that acquired genetic variations may contribute to the selection and long-term persistence of HIV-1 infected cells.
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