Selective reactivation of latent HIV using CyclinT1-Tat-containing virus-like particles

The persistence of HIV reservoirs and their ability to create an active infection after anti-retroviral therapy cessation has prevented development of an HIV cure. Various chemical latency reversal agents (LRAs) have been investigated to promote HIV transcription as part of a kick and kill strategy, but many of these agents lack either potency or specificity and can cause widespread T cell activation and systemic toxicity. We report the development of novel virus-like particles (VLPs), based on HIV itself, that carry a CyclinT1-Tat fusion protein (CycTat) and reactivate HIV from latency both alone and synergistically with the two tested chemical LRAs; a bromodomain inhibitor and a protein kinase C agonist. CycTat resulted in higher reactivation than Tat, although Tat and CycTat delivery were equivalent in some cell lines after co-stimulation with LRAs thought to increase cellular P-TEFb levels. Targeted mutations disrupting key residues in Tat and CycT1 interactions dampened reactivation, suggesting the particles work mechanistically as anticipated. Fusion of VLPs with target cells was required for HIV reactivation, demonstrating that CycTat proteins do not non-specifically cross cell membranes when packaged into VLPs. Additionally, we addressed safety concerns by testing high doses of VLPs on primary CD4⁺ T cells, which resulted in minimal T cell activation. This serves as proof-of-concept for specific reactivation of HIV by delivery of Tat protein by VLPs and shows that additional components, here a truncated CyclinT1, can be engineered into particles to enhance viral reactivation.