Journal
MBIO
Volume 9, Issue 2, Pages -Publisher
AMER SOC MICROBIOLOGY
DOI: 10.1128/mBio.00358-18
Keywords
A32; ADCC; ADCC assay; CD4i Abs; Env; granzyme B assay; HIV-1; luciferase assay; RFADCC; uninfected bystander; bNAbs
Categories
Funding
- CIHR foundation [352417]
- amfAR Innovation grant [109343-59-RGRL]
- FAIR
- NIH [AI100645, AI100663]
- BEAT-HIV Delaney Consortium [UM1 AI 126620]
- National Institutes of Health [AI121135, AI095098, AI098485, AI055332, OD011106]
- DFG [CRC 1279]
- ERC [323035]
- National Health and Medical Research Council [1124680]
- Canada Research Chair on Retroviral Entry [RCHS0235]
- Mathilde Krim Fellowship in Basic Biomedical Research from amfAR
- CIHR Fellowship Awards
- FRSQ postdoctoral fellowship award
- FRSQ Senior Research Scholar Award
- German Research Foundation (DFG)
- ERC
- NIH R01 [AI129769]
- [R01 AI 114266]
- National Health and Medical Research Council of Australia [1124680] Funding Source: NHMRC
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The conformation of the HIV-1 envelope glycoprotein (Env) substantially impacts antibody recognition and antibody-dependent cellular cytotoxicity (ADCC) responses. In the absence of the CD4 receptor at the cell surface, primary Envs sample a closed conformation that occludes CD4-induced (CD4i) epitopes. The virus controls CD4 expression through the actions of Nef and Vpu accessory proteins, thus protecting infected cells from ADCC responses. However, gp120 shed from infected cells can bind to CD4 present on uninfected bystander cells, sensitizing them to ADCC mediated by CD4i antibodies (Abs). Therefore, we hypothesized that these bystander cells could impact the interpretation of ADCC measurements. To investigate this, we evaluated the ability of antibodies to CD4i epitopes and broadly neutralizing Abs (bNAbs) to mediate ADCC measured by five ADCC assays commonly used in the field. Our results indicate that the uninfected bystander cells coated with gp120 are efficiently recognized by the CD4i ligands but not the bNabs. Consequently, the uninfected bystander cells substantially affect in vitro measurements made with ADCC assays that fail to identify responses against infected versus uninfected cells. Moreover, using an mRNA flow technique that detects productively infected cells, we found that the vast majority of HIV-1-infected cells in in vitro cultures or ex vivo samples from HIV-1-infected individuals are CD4 negative and therefore do not expose significant levels of CD4i epitopes. Altogether, our results indicate that ADCC assays unable to differentiate responses against infected versus uninfected cells overestimate responses mediated by CD4i ligands. IMPORTANCE Emerging evidence supports a role for antibody-dependent cellular cytotoxicity (ADCC) in protection against HIV-1 transmission and disease progression. However, there are conflicting reports regarding the ability of nonneutralizing antibodies targeting CD4-inducible (CD4i) Env epitopes to mediate ADCC. Here, we performed a side-by-side comparison of different methods currently being used in the field to measure ADCC responses to HIV-1. We found that assays which are unable to differentiate virus-infected from uninfected cells greatly overestimate ADCC re-sponses mediated by antibodies to CD4i epitopes and underestimate responses mediated by broadly neutralizing antibodies (bNAbs). Our results strongly argue for the use of assays that measure ADCC against HIV-1-infected cells expressing physiologically relevant conformations of Env to evaluate correlates of protection in vaccine trials.
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