Elevation of cell-associated HIV-1 RNA transcripts in CSF CD4+ T cells, despite suppressive antiretroviral therapy, is linked to in vivo brain injury

Objective Despite effective antiretroviral therapy (ART), brain injury remains prevalent in people living with HIV-1 infection (PLHIV) possibly due to ART’s lack of direct inhibition of transcription with continued local production of viral transcripts and neurotoxic proteins, such as Tat, rather than cell-free whole virion toxicity. We quantified cell-associated (CA) HIV-1 RNA-transcripts in CSF and blood, in relation to proton Magnetic Resonance Spectroscopy ( 1 H MRS) of major brain metabolites, in well characterised PLHIV. Methods RNA was extracted from cells in 16 paired samples of CSF and blood, from PLHIV on fully suppressive ART. HIV-1 CA-RNA copies were measured using the highly sensitive Double-R assay and normalized /10 6 CD4+ T cells. 18-colour flow cytometry was used to count and analyse CD4+ T cells and monocytes in CSF and blood. The concentrations of major brain metabolites from 1 H MRS in frontal white matter (FWM), posterior cingulate cortex (PCC), and caudate areas were measured. Brain injury in each voxel was defined using a composite score derived by principal component analysis. Results 14/16 CSF cell samples had quantifiable HIV-1 CA-RNA transcripts, at levels significantly higher than in their PBMCs (median 9,266 vs 185 copies /10 6 CD4+ T cells; p<0.0001). Higher levels of CSF transcripts were associated with greater brain injury in the FWM (Std β=-0.73; p=0.007) and PCC (Std β=-0.61; p=0.03). CSF cells were 91% memory T cells, equally CD4+ (median 3,605) cells and CD8+ T cells (3,632), but contained much fewer B cells (0.4 %), NK cells (2.0%) and monocytes (3.1%; 378 cells; >90% CD14+CD16+ phenotype). CXCR3+CD49d+integrin ß7-negative, CCR5+ CD4+ T cells were significantly enriched in CSF, compared with PBMC (p <0.001). Transcriptional activity in CSF cells was highly correlated with levels of transcriptional activity in CD4+ T cells in PBMC (r=0.76; p=0.002). In contrast, HIV-1 RNA in highly purified monocytes from PBMC was detected in only 6/16 samples. Conclusions Elevated HIV-1 transcripts in CSF cells were associated with in vivo brain injury, despite suppressive ART. The cellular source is most likely the predominant CXCR3+ CD49d+ integrinß7-CCR5+ memory CD4+ T cells, not monocytes. Inhibitors of transcription to reduce local production of potentially neurotoxic proteins, should be developed.