Provirus Activation Plus CD59 Blockage Triggers Antibody-Dependent Complement-Mediated Lysis of Latently HIV-1–Infected Cells

Abstract

Latently HIV-1-infected cells are recognized as the last barrier towards viral eradication and cure. To purge these cells, we combined a provirus stimulant with a blocker of human CD59, a key member of the regulators of complement activation (RCA), to trigger antibody-dependent complement-mediated lysis (ADCML). Provirus stimulants including prostratin and histone deacetylase inhibitors (HDACi) such as romidepsin (RMD) and suberoylanilide hydroxamic acid (SAHA) activated proviruses in the latently HIV-1-infected T cell line ACH-2 as virion production and viral protein expression on the cell surface were induced. RMD was the most attractive provirus stimulant as it effectively activated proviruses at nanomolar concentrations that can be achieved clinically. Antiretroviral drugs including two protease inhibitors (atazanavir and darunavir) and a reverse transcriptase inhibitor (emtricitabine) did not affect the activity of provirus stimulants in the activation of proviruses. However, saquinavir (a protease inhibitor) markedly suppressed virus production, although did not affect the percentage of cells expressing viral Env on the cell surface. Provirus-activated ACH-2 cells expressed HIV-1 Env that colocalized with CD59 in lipid rafts on the cell surface, facilitating direct interaction between them. Blockage of CD59 rendered provirus-activated ACH-2 cells and primary human CD4+ T cells that were latently infected with HIV-1 sensitive to ADCML by anti-HIV-1 polyclonal antibodies or plasma from HIV-1-infected patients. Therefore, a combination of provirus stimulants with RCA blockers represents a novel approach to eliminate HIV-1.

Blockage of CD59 Function Restores Activities of Neutralizing and Nonneutralizing Antibodies in Triggering Antibody-Dependent Complement-Mediated Lysis of HIV-1 Virions and Provirus-Activated Latently Infected Cells

“…However, accumulating data sug-gest that short-term or transient blockage of CD59 may be feasible and tolerable to HIV-1 patients, since CD59 blockage, deficiency, or knockout does not cause severe off-target effects in vitro and in vivo (72)(73)(74)(75)(76)(77)(78)(79)(80). We and others have demonstrated that the use of CD59 blockers in vitro to block CD59 does not cause nonspecific cytolytic effect on cell lines, human RBCs, or PBMCs in the presence of sera/plasma from HIV-1-infected patients (33)(34)(35). Thus, CD59 blockers that are expected to be used short term, not lifelong, will be better tolerated in HIV-1 patients.…”

“…Blockage of hCD59 function with intact BRIC229 molecules allows anti-HIV-1 gp120/gp160 Abs or Abs in the sera/plasma from HIV-1-infected individuals to restore their activity of ADCML to destroy complement-resistant HIV-1 virions and infected cells (30)(31)(32)(33)(34). We have recently reported that blockage of hCD59 renders provirus-activated ACH-2 cells and primary human CD4 ϩ T cells that were latently infected with HIV-1 sensitive to ADCML by anti-HIV-1 polyclonal Abs or plasma from HIV-1-infected patients (35). However, it is unclear whether F(ab=) 2 fragments of BRIC229 would retain hCD59-blocking activity of the intact Ab of BRIC229.…”

“…ADCML of provirus-activated ACH-2 cells by nAbs or nonnAbs was quantified using the calcein release assay, as described in our recent report (35). Briefly, ACH-2 or A3.01 cells treated or untreated with PMA/ionomycin or DMSO were labeled at 37°C for 40 min with 15 M calcein-AM dye (Invitrogen) in 200 l of PBS.…”

“…Blockage of hCD59 function can render complement-resistant HIV-1 virions and infected cells sensitive to ADCML by Abs against HIV-1 Env or sera/plasma from HIV-1-infected patients (30)(31)(32)(33)(34). We have recently reported that blockage of hCD59 renders provirus-activated ACH-2 cells and primary human CD4 ϩ T cells that were latently infected with HIV-1 sensitive to ADCML by anti-HIV-1 polyclonal Abs or plasma from HIV-1-infected patients (35). However, it is unclear whether the enhancement of ADCML is mediated by nAbs, non-nAbs, or both.…”

Antibody-Dependent Cellular Cytotoxicity against Reactivated HIV-1-Infected Cells

“…While several studies have utilized latently infected cell lines to examine reactivation of the latent provirus (39,40) and even elimination of reactivated cells (41), transformed cell lines may not be ideal models of HIV-1 latency and may not be reflective of primary CD4 ϩ T cells as targets for ADCC. Latently infected CD4 ϩ T cells are rare in HIV-1 ϩ subjects on ART (approximately 1 replication-competent virus per million resting CD4 ϩ T cells [38]), and to obtain sufficient HIV-1-infected CD4 ϩ T cell targets, it was necessary to activate the primary cells with mitogens and culture them for several days.…”