Background
Accumulating evidence supports that the establishment and persistence of HIV latent reservoirs despite suppressive ART are intricately linked to the quiescent metabolism of CD4+ memory T cells. We investigated whether reprogramming T cell metabolism towards aerobic glycolysis could facilitate HIV latency reversal from circulating viral reservoirs. We used Zaprinast, a Mitochondrial Pyruvate Carrier inhibitor, to block pyruvate entry into mitochondria and examined its latency reversal capacity on circulating HIV reservoirs from ART-suppressed individuals and its impact on primary CD4 T cells metabolism.
Methods
HIV latency reversal was performed ex vivo on CD4+ T cells or CD8-depleted PBMC from ART-suppressed individuals. HIV gene expression and viral production were measured by RT-qPCR targeting the HIV tat-rev msRNA or HIV gag RNA respectively. Mitochondrial respiration and ATP production were assessed using the Mitostress test and ATP real-time kits (Agilent). Aerobic glycolytic activity was measured using pH Xtra assay (Agilent) and glucose uptake with 2-NBDG. Flow cytometry was used to evaluate mitochondrial mass (MitoView Green, Generon), membrane potential (MitoTracker Red CMXRos, Invitrogen), ROS production (MitoSOX Red, Thermofisher), viability (Viobility 405/452, Miltenyi) and activation (CD69 and CD25 antibodies, Biolegend).
Results
Zaprinast induced viral particles release in 65% (11/17) of CD8-deplected PBMCs isolated from ART-suppressed individuals, achieving 69% of maximal HIV reactivation following TCR activation. Zaprinast also triggered HIV gene reactivation in 50% (4/8) of CD4+ T cell samples isolated from ART-suppressed individuals, confirming that Zaprinast targets CD4+ T cells HIV reservoirs. Metabolic analyses revealed that Zaprinast re-programmed CD4 T cell metabolism in two steps. Initially, Zaprinast transiently inhibited mitochondrial respiration before enhancing both aerobic glycolysis and mitochondrial respiration. This effect was reversible with CD4+ T cells returning to a quiescent metabolism upon removal of the drug. The treatment did not impact cell viability, ROS production, or CD4 T cells activation and proliferation.
Conclusions
This study demonstrates for the first time that MPCi represents a potent new class of LRAs capable of inducing robust HIV latency reversal from circulating viral reservoirs ex vivo. This strongly supports the potential of targeting the quiescent metabolism of CD4+ T cells as a promising approach for Shock and Kill strategies.