Pharmacologic HIV-1 Nef blockade promotes CD8 T cell–mediated elimination of latently HIV-1–infected cells in vitro
Shariq Mujib, … , Thomas E. Smithgall, Mario A. Ostrowski
Shariq Mujib, … , Thomas E. Smithgall, Mario A. Ostrowski
Published September 7, 2017
Citation Information: JCI Insight. 2017;2(17):e93684. https://doi.org/10.1172/jci.insight.93684.
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Research Article AIDS/HIV Immunology

Pharmacologic HIV-1 Nef blockade promotes CD8 T cell–mediated elimination of latently HIV-1–infected cells in vitro

Abstract

Eradication of the HIV-1 latent reservoir represents the current paradigm to developing a cure for AIDS. HIV-1 has evolved multiple mechanisms to evade CD8 T cell responses, including HIV-1 Nef–mediated downregulation of MHC-I from the surface of infected cells. Nef transcripts and protein are detectable in samples from aviremic donors, suggesting that Nef expression in latently HIV-1–infected CD4 T cells protects them from immune-mediated clearance. Here, we tested 4 small molecule inhibitors of HIV-1 Nef in an in vitro primary CD4 T cell latency model and measured the ability of autologous ex vivo or HIV-1 peptide–expanded CD8 T cells to recognize and kill latently infected cells as a function of inhibitor treatment. Nef inhibition enhanced cytokine secretion by autologous CD8 T cells against latently HIV-1–infected targets in an IFN-γ release assay. Additionally, CD8 T cell–mediated elimination of latently HIV-1–infected cells was significantly enhanced following Nef blockade, measured as a reduction in the frequency of infected cells and Gag protein in cultures following viral outgrowth assays. We demonstrate for the first time to our knowledge that Nef blockade, in combination with HIV-specific CD8 T cell expansion, might be a feasible strategy to target the HIV-1 latent reservoir that should be tested further in vivo.

Authors

Shariq Mujib, Aamir Saiyed, Saleh Fadel, Ardalan Bozorgzad, Nasra Aidarus, Feng Yun Yue, Erika Benko, Colin Kovacs, Lori A. Emert-Sedlak, Thomas E. Smithgall, Mario A. Ostrowski

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Figure 3

Nef blockade enhances CD8 recognition of latently HIV-1–infected CD4 T cells.

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Nef blockade enhances CD8 recognition of latently HIV-1–infected CD4 T c...
Resting HIV-1–infected or mock-infected CD4 T cells were treated with Nef inhibitors as indicated. Ex vivo (A–C) or HIV-1 peptide–expanded autologous CD8 T cells (D–F) were cocultured overnight with latently infected cells at a 1:1 ratio. IFN-γ was quantified in pg/ml via ELISA. Cocultures with Nef inhibitor–treated CD4 T cells resulted in greater IFN-γ secretion by the CD8 T cells, indicating improved recognition of latently HIV-1–infected cells compared with DMSO control. (A) Example of an individual experiment using ex vivo CD8 T cells. Summary data of ex vivo CD8 T cell cocultures with mock (B) and latently HIV-1–infected (C) CD4 T cells are shown. Paired data obtained with inhibitor B9 of ex vivo CD8 T cell cocultures with latently infected targets are shown in C. (D) An example of an individual experiment of cocultures with HIV-1 peptide–expanded CD8 T cells. E and F are summary data of cocultures with expanded CD8 T cells against mock- and latently HIV-1–infected CD4 T cells, respectively. Paired data depicting IFN-γ release of expanded CD8 T cell cocultures between the control and B9 inhibitor–treated cells are shown in F. Summary data of Nef deficient (NL4-3 ΔNef) cocultures with ex vivo (G) or autologous expanded CD8 T cells (H) are reported. No significant differences were observed between the inhibitors and control conditions in these experiments. The dotted horizontal line in each plot indicates the median DMSO control IFN-γ. Each data point represents an individual experiment. All cocultures contained HAART. (A–H) Medians shown; Wilcoxon matched-pairs tests were performed between no inhibitor (NT DMSO) and other wells to determine statistical significance. (B) n = 20 DQBS, B9, IL-15; n = 12 others. (C) n = 32 for DQBS, B9, IL-15; n = 23 others. (E) n = 15. (F) n = 27 DQBS, JZ-96-21, JZ-97-21; n = 28 others. (G) n = 6. (H) n = 11 for NT; n = 10 for other conditions. ND, Not detected. Statistical analysis was adjusted for multiple comparisons using the Benjamini-Hochberg procedure. *P < 0.05, **P < 0.01.