An unbiased OMICs approach to identify mechanisms of Cocaine regulation of the HIV reservoir
Survival rates of HIV+ substance users are significantly lower compared to non-users, even in settings in which ART is generally available. Among substance-abuse, chronic cocaine use is associated with high incidence of HIV infection.
We apply an unbiased systems biology approach, which combined with the highly characterized subject cohort, identify the pathways triggered by chronic use of cocaine in cART treated HIV infected subjects which lead to HIV persistence and failure to reconstitute the immune system.
In NIDA cohort, the CD4 counts and inducible HIV reservoir (measured through TILDA: Tat-Inducible Limiting Dilution Assay) showed a negative correlation in pooled populations of non-users and chronic cocaine users. Using Sparse Linear regression bioinformatic approaches to integrate results from flow cytometry, transcriptional profiling and plasma metabolomics, we identified pathways associated with exhaustion (TGF-b signaling, fatty-acid metabolism, G2M check, point), exhaustion markers (PD-1 on CD8 TCM, TIGIT on CD8 TCM/TTM, Blimp1 in CD4 TCM/TTM/TEM), and glycolytic metabolites (fructose) were present at significantly higher levels in subjects with high inducible HIV reservoir as monitored by TILDA. In addition, exhaustion markers LAG3 and CD150, along with increased NOTCH signaling pathways, were associated with low CD4 counts. These T cell exhaustion markers, metabolites and pathways appear to drive high inducible HIV reservoir and low CD4 counts in both cocaine users and non-users.
We also utilize high density flow cytometry and tSNE analysis to identify flow cytometry profiles that are associated to high levels of HIV persistence and lack of immune reconstitution. Specific CD4 and CD8 populations that were upregulated in cocaine users and were associated with immune exhaustion were identified. They were positively correlated with the inducible HIV reservoir and negatively correlated with CD4, expressing high PD-1, IRF4, TIGIT, Blimp1, pStat1, pStat3 and pStat5. These data highlight that dysfunctional intracellular signaling in both CD4 and CD8 T cell subsets are associated with high inducible reservoir in cocaine users. This signaling (downstream of surface exhaustion markers like PD1 and TIGIT) is driven by classical anti-inflammatory exhaustion associated molecules like pSTAT3/IRF4 and Blimp1; and by key driver of homeostatic proliferation – pSTAT5. Further exploration of the above described populations using high-throughput (i.e. scRNA-seq) and functional assays could help in determining the role of these populations in cocaine users and pave the way for therapeutic approaches to restrict aberrant clinical outcomes in HIV infected cocaine users.