1. Mechanisms of HIV-1 Resistance to ADCC David Evans University of Wisconsin-Madison July 26, 2017
“No conflicts of interest to declare”

2. Perspective
HIV-1 has evolved to replicate continuously in the presence of active host immune responses, and has therefore acquired mechanisms to overcome all immune responses in most infected individuals.
The ability of neutralizing antibodies to block HIV-1 infectivity is dependent on their capacity to bind to native conformations of the viral envelope glycoprotein as they exist on the surface of virions.

3. ADCC Assay Antibody Env CD16 HIV-1 Tat NK Cell No plasma No virus
LTR Luc.
0% 100%
Serial antibody dilutions
No virus
No plasma
Similarity to assays for neutralizing antibodies.
Alpert et al. J. Virol. 2012

4. Env Expression on the Surface of HIV-infected
is Tightly Regulated
Gardiner et al. J. Virol. 2016

5. Env Internalization Protects HIV-infected Cells from ADCC
HIV-1 gp41 tail
710
WT: NRVRQGYSPL SFQTHLPIPR
Y710G: G
Y710F: F
% RLU
% MAX
Env
HIVIG (µg/ml)
von Bredow et al. J. Virol. 2015

6. Tetherin
Adapted from Gottlinger Nature

7. Vpu Downregulates and Degrades Tetherin
bTrCP-2
Vpu
Lysosomal
Degradation

8. Neil et al Nature

9. Vpu Protects HIV-infected Cells from ADCC

10. RNAi Knockdown of Tetherin, but not CD4 or NTB-A,
Increases Resistance to ADCC
Tetherin
CD4
NTB-A
Arias et al. PNAS. 2014

11. CD4 Downmodulation by Nef and VpuAb
CD4
Env
CD4
Env
Nef
Vpu
Downregulation
& Degradation

12. CD4 Downmodulation Prevents the Exposure of CD4-Inducible Epitopes

13. Sensitivity of Wildtype versus Nef- and Vpu-Deleted HIV-1 CH77 T/F to Plasma from HIV+ Donors

14. HIV-1 Env Trimer Glycan Shield CD4 Binding Site (CD4bs) V3 gp120 CD4bs
Viral Membrane or
Infected Cell Membrane
Glycan Shield
CD4 Binding Site
(CD4bs)
gp120
gp41 V3
CD4bs
MPER
Burton et al. Cell Host & Microbe. 2012

15. Broadly Neutralizing Antibodies
% RLU
von Bredow et al. J. Virol. 2016
mAb (µg/ml)

16. Non-Neutralizing Antibodies
% RLU
von Bredow et al. J. Virol. 2016
mAb (µg/ml)

17. ADCC Correlates with Neutralization

18. Conclusions
There is little Env on the surface of HIV-infected cells due to trafficking signals in cytoplasmic tail of gp41.
Tetherin antagonism by Vpu protects HIV-infected cells from ADCC by preventing the accumulation of captured virions on the cell surface.
CD4 downmodulation by Nef and Vpu protects HIV-infected cells from ADCC by preventing the exposure of CD4-inducible epitopes in Env.
Many of the same structural features of the Env trimer that afford resistance to neutralizing antibodies also afford resistance to ADCC.

19. Acknowledgements University of Wisconsin University of Montreal
Benjamin von Bredow Andres Finzi
Juan Fernando Arias
Lisa Heyer Scripps Research Institute
Michael Alpert Dennis Burton
Jaye Gardiner New York University
Nathan Sherer Susan Zolla-Pazner
Tulane University
James Robinson
AI121135

21. CD4 Downmodulation by Vpu and Nef Prevents the Exposure
of CD4-Inducible Epitopes on the Surface HIV-infected Cells
Veillette et al. J. Virol. 2014

22. CD4 Downmodulation by Vpu and Nef Protects
HIV-infected Cells from ADCC
Veillette et al. J. Virol. 2015

23. ADCC Correlates with Binding to Env on the Surface of Infected Cells and with Virus Neutralization
von Bredow et al. J. Virol. 2016

25. Tetherin/BST-2/CD317
Klimkait & Orenstein J. Virol.
Kupzig et al Traffic
Tetherin, also known as BST-2 or CD317, was identified a little over 3 years ago by Stuart Neil and Paul Bieniasz, and independently by Nicole Van Damme and John Guatelli, as the interferon-inducible host-cell protein that interferes with the detachment of vpu-deleted HIV-1 from certain restrictive cells.
This factor accounts for the late stage defect in the release of virus particles shown here by electron microscopy. In the absence of Vpu, the virus can complete assembly and budding, note that the viral envelope of these particles is discontiguous with the plasma membrane of the cell. However, the virus remains attached to the surface of the cell.
The explanation for this phenotype was provided with the identification of tetherin. This molecule has a rather unusual topology that includes a an N-terminal cytoplasmic domain, a membrane-spanning domain, an extracellular coiled-coil domain and a site for the addition of a glycosyl phosphatidyl inositol tail at the C-terminus. Based on these structural features that enable both ends of the molecule to be anchored in lipid membranes it was proposed that…
interferon-inducible transmembrane protein that interferes with the release of vpu-deleted HIV-1 from restrictive cells
Neil et al Nature
Van Damme et al Cell Host & Microbe

26. IFNa Treatment of HIV-infected Cells Increases
their Susceptibility to ADCC
Arias et al. PNAS. 2014

27. Vpu Substitutions Differentiate Tetherin Antagonism
from CD4- and NTB-downmodulation
Tetherin
Vpu
A14L X
S52N
S56N X
bTrCP
Sequestration
Degradation
A14L – blocks binding to tetherin,
but does not affect CD4
I46K – blocks CD4 binding
S52N, – prevents tetherin degradation,
S56N but not tetherin sequestration
– does not affect NTB-A
Arias et al. PNAS. 2014

28. CD4 Binding Exposes Surfaces of the Inner Domain of gp120 Recognized by Non-neutralizing Antibodies
Veillette et al. J. Virol. 2014

29. Exceptions…Gp41 MPER-Specific bNAbs and 2G12
% RLU
mAb (µg/ml)

30. Sensitivity of Wildtype versus Nef- and Vpu-Deleted HIV-1 CH77 T/F to Monoclonal Antibodies

31. Mechanisms of HIV-1 Resistance to ADCC
Env internalization
Tetherin antagonism
CD4 downmodulation
Structural features of the Env trimer

32. Antibody-Dependent Cell-mediated Cytotoxicity (ADCC)
Env
SIV or HIV
infected cell
NK cell
CD16
Granules
Perforin
Granzyme

33. Acknowledgements University of Wisconsin University of Montreal
Benjamin von Bredow Andres Finzi
Juan Fernando Arias
Lisa Heyer Scripps Research Institute
Michael Alpert Dennis Burton
Jaye Gardiner New York University
Nathan Sherer Susan Zolla-Pazner
Tulane University
James Robinson
AI121135