Exosome-mediated systemic delivery platform of therapeutic cargo to epigenetically repress HIV-1 infection

Human Immunodeficiency Virus (HIV-1) produces a persistent latent infection. Control of HIV-1 using combination antiretroviral therapy (cART) comes at the cost of life-shortening side effects and development of drug-resistant HIV-1. An ideal and safer therapy should be deliverable in vivo and target the stable epigenetic repression of the virus, inducing a stable “block and lock” of virus expression. Towards this goal, researchers from the City of Hope-Beckman Research Institute developed an HIV-1 promoter-targeting Zinc Finger Protein (ZFP-362) fused to active domains of DNA methyltransferase 3 A to induce long-term stable epigenetic repression of HIV-1. Cells were engineered to produce exosomes packaged with RNAs encoding this HIV-1 repressor protein. The researchers found that the repressor loaded anti-HIV-1 exosomes suppress virus expression and that this suppression is mechanistically driven by DNA methylation of HIV-1 in humanized NSG mouse models. The observations presented here pave the way for an exosome-mediated systemic delivery platform of therapeutic cargo to epigenetically repress HIV-1 infection.

Schematic for mechanism ZPAMt mRNA transfer from producer cells to recipient cells

HEK293T producer cells are transfected with the following 4 plasmids: “Booster” to increase exosome production, cytosolic delivery helper “Connexin43 S368A”, packaging plasmid CD63-L7Ae, and cargo mRNA-C/D_box plasmid. The cargo was intended to be either nLuc reporter, ZFP (non-fusion control), or ZPAMt. Exosomes derived from these producer cells with the ZPAMt-C/D_box mRNA can transfer ZPAMt-C/D_box mRNA in recipient cells and produce the ZPAMt fusion protein. This ZPAMt protein leads to transcriptional epigenetic repression of HIV-1 within the infected cell.