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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

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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 4

Improved killing of latently HIV-1–infected cells by expanded CD8 T cells following Nef blockade.

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Improved killing of latently HIV-1–infected cells by expanded CD8 T cell...
Resting latently HIV-1 NL4-3–infected CD4 T cells treated with Nef inhibitors were cocultured with autologous HIV peptide–expanded CD8 T cells in coculture medium containing HAART. Virus reactivation was induced by adding IL-15 to the cultures in the presence of HAART, along with MHC-I blocking antibody W6/32 to block CD8 T cell–mediated killing for another 6 days. HIV-1 Gag p24 was quantified in the supernatants by ELISA, and an example of an experiment is shown in A; mean and SEM of 2 replicates shown. (B) Paired data between control and inhibitor B9 receiving cocultures are depicted, as well as the summary data of cocultures with each of the indicated treatments. (C) Summary data of cocultures of expanded CD8 T cells with latently infected CD4 T cells generated with Nef-deficient NL4-3 virus (NL4-3 ΔNef) are shown. Medians shown; each symbol represents 1 individual experiment, and the dotted line indicates the median HIV-1 p24 detected in the DMSO control (NT). Wilcoxon matched-pairs tests were performed to determine statistical significance. (B) n = 24 for DQBS, JZ-96-21, and JZ-97-21 and n = 26 for B9 and IL-15; (C) n = 6. Latently infected CD4 T cells were generated with HIV-1 NL4-3 virus for all data reported in A and B, whereas the NL4-3 ΔNef virus was used for data in C. All P values shown have been corrected for multiple comparisons using the Benjamini-Hochberg procedure. *P < 0.05, **P < 0.01, ***P < 0.001.

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