Characterization of Non-subtype B HIV-1 Reservoirs and its Association with Longitudinal Clinical Outcomes

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PROJECT SUMMARY Current HIV antiretroviral treatment successfully controls viral replication and has transformed HIV-infection from a fatal illness to a manageable chronic condition. However, despite suppression of viral replication during treatment, studies have shown that pools of latent viral reservoirs remain detectable, which fuel viral rebound when antiviral suppression treatment is interrupted. These viral reservoirs are established almost immediately upon infection when HIV irreversibly integrates its viral genome into human DNA. Viral reservoirs are extremely durable, not susceptible to therapeutic effects of currently available antiretroviral agents, and have been refractory to recent experimental treatment approaches. HIV infection is also characterized by a high level of intrahost genotypic diversity of viral quasispecies. In addition to genetic diversity associated base substitution mutations, pools of viral DNA genomes recovered from chronically-infected patients under prolonged suppressive therapy often contain high frequencies of genome-truncated and/or hypermutated, non-replication- competent viral DNA genomes. Only a small fraction of proviral genomes in these patients are genome-intact and may lead to productive viral replication and virologic rebound in the absence of treatment. Furthermore, HIV-infected cells infected with both genome-intact and genome-defective proviruses have been shown to clonally expand, serving as a mechanism of HIV persistence. However, our current understanding of HIV reservoirs has been derived almost exclusively from studies on a strain called subtype B HIV-1, the predominate viral subtype affecting first-world nations but only makes up 10% of the global epidemic. In contrast, non-B HIV- 1 subtypes predominate regions such as sub-Saharan Africa where disease burden is the highest globally. Questions remain on whether the remaining 90% of infections by other HIV-1 subtypes differ in reservoir sizes and compositions. To address this question, we will leverage an existing biobank of a previously NIH-funded Ugandan HIV cohort (UARTO), which houses 12360 blood cell samples collected longitudinally over ten years from 500 predominantly subtype A1 and D HIV-1-infected individuals. We will use three cutting-edge technologies (1) FLIP-seq to obtain near-full-length HIV-1 DNA genomes profiles, (2) MIP-seq to co-capture HIV- 1 integration sites and viral genome, and (3) and the Intact Proviral DNA Assay (IPDA) to longitudinally measure the decay/expansion rate of the reservoir. All three technologies allow us to focus on the rare intact viral DNA genomes that is the target for HIV cure strategies. Across subtypes, we will compare reservoir characteristics including absolute genome-intact reservoir sizes, extent of clonal expansion, integration site profiles, viral promoter genotypes, and longitudinal decay/expansion dynamics. We will further investigate demographic, clinical and host factors associated with genome-intact viruses. Overall, we aim identify differences, or the lack of differences, between HIV-1 subtype reservoirs to inform HIV cure research effort on whether a cure strategy should be subtype-specific.