DOI: 10.5061/dryad.5qfttdzhh
title: Multiomic analysis of ART-interruption cohorts identifies cell-extrinsic and -intrinsic mechanisms driving lymphocyte-mediated control of HIV rebound

description: The immunological mechanisms regulating HIV rebound after antiretroviral
 therapy (ART) interruption remain unclear. We examined relationships
 between host factors, HIV reservoir, and HIV time-to-rebound after
 analytical treatment interruption (ATI) by characterizing pre-ATI
 peripheral blood mononuclear cells (PBMCs) from 75 ART-suppressed people
 with HIV (PWH) using high-parameter methods. Across interventional (CLEAR,
 TEACH, REDUC) and non-interventional (A5345) cohorts, delayed rebound was
 not associated with intact HIV. Cohort-specific immune effectors were
 associated with delayed rebound. RNA sequencing of CD4+ T cells from A5345
 revealed that mTOR inhibitor DDIT4 and zinc finger protein ZNF254 were
 associated with delayed rebound. In vitro and in vivo studies demonstrated
 that DDIT4 and ZNF254 suppressed HIV expression. Metformin induced DDIT4
 and suppressed HIV expression in primary cells and cells from
 ART-suppressed PWH, suggesting that this affordable diabetes drug could be
 repurposed to silence HIV. Our results support pursuit of both immune- and
 HIV-silencing strategies to achieve ART-free HIV remission.
description: # Cell extrinsic and intrinsic mechanisms of lymphocyte-mediated post-ART
 control of HIV Contact: Nadia Roan, PhD Gladstone Institutes University of
 California, San Francisco
 [Nadia.roan@gladstone.ucsf.edu](mailto:Nadia.roan@gladstone.ucsf.edu)
 [Nadia.roan@ucsf.edu](mailto:Nadia.roan@ucsf.edu) Dates of collection:
 3/23/19 – 1/16/23 Information about geographic location of data
 collection: Original specimens were from individuals in Denmark enrolled
 in three trials (CLEAR, PMID 26423811; REACH PMID 30932955; and REDUC PMID
 27658863) and individuals in USA enrolled in ACTG A5345 (PMID 38329130).
 CyTOF datasets were generated in San Francisco, California, USA. ## Data
 and File Overview Included are standard flow cytometry (FCS) files
 generated from clinical trial specimens of people with HIV (PWH)
 suppressed on antiretroviral therapy (ART) who underwent analytical
 treatment interruption (ATI). All specimens were obtained before
 analytical treatment interruption, and the primary association of the
 study was the time it took for HIV to rebound to 1,000 copies/ml during
 the ATI. Specimens from four trials are included. Three trials were
 interventional trials performed in Denmark: CLEAR wherein participants
 orally received the latency reversal agent (LRA) panobinostat for 8 weeks
 (n=9 participants), REDUC wherein participants received therapeutic
 vaccination followed by intravenous administration of romidepsin as an LRA
 (n=16 participants), and TEACH wherein participants received the TLR9
 agonist lefitolimod as both an LRA and an immune-boosting agent (n=9
 participants). The fourth trial was the AIDS Clinical Trials Group (ACTG)
 trial A5345, a prospective study wherein individuals underwent ATI in the
 absence of any therapeutic intervention (n=41 participants). All specimens
 were analyzed by CyTOF using separate phenotyping panels dedicated for T
 cell or NK cell analysis. The data files are made available as FCS files,
 within eight zip files: A5345_NK_cells.zip A5345_T_cells.zip
 CLEAR_NK_cells.zip CLEAR_T_cells.zip REDUC_NK_cells.zip REDUC_T_cells.zip
 TEACH_NK_cells.zip TEACH_T_cells.zip The first part of the zip file name
 refers to the cohort (A5345, CLEAR, REDUC, or TEACH), while the second
 part refers to the type of cell analyzed (T cells or NK cells). The FCS
 files within each of these zipped folders includes the cohort identifier
 (“A5345”, “TEACH”, “CLEAR”, or “REDUC”), a de-identified participant ID
 (the PID, a 5-6 digit number), and an indication of whether the data were
 generated using the T cell or NK cell panel (“NKcells” or “Tcells”). T
 cell datasets were pre-gated on live, singlet CD3+CD19- cells. NK cell
 datasets were pre-gated on live, singlet CD3-CD19-CD14-CD33- cells for
 TEACH, CLEAR, and REDUC, and CD3-CD19-CD14-CD33-CD4-CD7- cells for ACTG
 A5345. As standard FCS files, the data can be analyzed using conventional
 flow cytometry software such as FlowJo. Free software to analyze FCS files
 include [https://floreada.io/analysis](https://floreada.io/analysis). ##
 Data-specific information Please see information under “data and file
 overview” and “methodological information”. The associated FCS files were
 generated from PBMCs from ART-suppressed PWH prior to ATI. One primary
 goal of generating these datasets was to identify immune features
 associated with time-to-rebound of HIV after ATI. Hence, we have included
 two tables (uploaded as CSV files, entitled “Table 1” and Table 2” within
 the “data” section) listing the time-to-rebound of HIV viremia to 1,000
 copies/ml (a primary outcome in the study) for each of the analyzed
 specimen. Table 1 provides the rebound time for the CLEAR, TEACH, and
 REDUC trials (in days to viral rebound to &gt;1,000 copies/ml HIV RNA),
 while Table 2 provides the rebound time for the ACTG A5345 trial (in weeks
 to viral rebound to &gt;1,000 copies/ml HIV RNA). For both Tables 1 and 2,
 we provide in the first column the cohort name followed by the PID, and in
 the second column we provide the days or weeks to viral rebound, as
 appropriate. In addition, we have included three tables (Tables 1-3)
 depicting the panels used to generate the CyTOF data. For each table, we
 depict the antigen specificity of the antibody (first column), the metal
 label used (second column), and the antibody clone name (third column).
 Note that antigens labeled with an asterisk were detected at the
 intracellular level. Table 3 lists the antibody panel for T cell analysis
 of CLEAR, TEACH, REDUC, and ACTG A5345. Table 4 lists the antibody panel
 for NK cell analysis of CLEAR, TEACH, and REDUC. Table 5 lists the
 antibody panel for NK cell analysis of ACTG A5345. The included tables:
 Table1_CLEAR_TEACH_REDUC_DaysRebound.csv Table2_ACTGA5345_WeeksRebound.csv
 Table3_Tcells_CLEAR_TEACH_REDUC_ACTGA5345.csv
 Table4_NKcells_CLEAR_TEACH_REDUC.csv Table5_NKcells_ACTGA5345.xlsx ##
 Sharing and access information No licenses or restrictions are placed on
 the data. ## Methodological Information Methods used for generating and
 analyzing the CyTOF datasets generated from clinical specimens have been
 previously described:
 [https://pubmed.ncbi.nlm.nih.gov/35154118/](https://pubmed.ncbi.nlm.nih.gov/35154118/) [https://pubmed.ncbi.nlm.nih.gov/39798087/](https://pubmed.ncbi.nlm.nih.gov/39798087/) ## Human subjects data We have received consent from participants to published the de-identified CyTOF data in the public domain. Each participant specimen was given an ID number that cannot be traced back to the individual.