1. Neutrophils mediate potent and rapid anti-HIV antibody-dependent functions
Matthew J Worley, Anthony D Kelleher,
Stephen J Kent and Amy W Chung
The University of Melbourne, Australia

2. FcR mediated antibody functions
Antibodies can recruit innate cells via FcR
NK cells can mediate ADCC responses via an Fcγ receptor
Monocytes and neutrophils are able to mediate phagocytosis and ADCC responses via Fcγ receptors

3. FcR mediated antibody functions
Virus
Phagosome
Antibody
Granules
Kill
Fc Receptor
Antigen
Antibody
Infected cell
Fc Receptor
Effector cell

4. FcR mediated antibody functions in HIV
High levels of ADCC antibodies correlate with delayed disease progression with HIV (Baum et al. 1996, Chung et al. 2011, Wren et al. 2013)
The RV144 vaccine trial highlighted the importance of Fc mediated antibody functions
IgG induced ADCC to envelope protein (Haynes et al. 2012)
IgG3 to the V1V2 region reduced risk of infection (Yates et al. 2014)
IgG3 mediated polyfunctional immune responses (Chung et al. 2014)
NK cells and monocytes have been the main focus of recent studies

5. Leukocyte cell prevalence
FcγRI
FcγRIIIB
FcγRIIB
FcγRIIB
FcγRIIA
FcγRIIIA
FcγRI
Neutrophils
~65%
T cells
~20%
NK cells
~5%
B cells
~5%
FcγRIIB
Monocytes
~5%
FcγRIIIA
FcγRIIA

6. Aim Cohort clinical data
Can neutrophils mediate antibody-dependent anti-HIV responses?
Cohort clinical data
HIV
Number of samples 33
CD4 count entry, cells/μl (median, Range)
566 ( )
Viral load RNA copies/ml (median, Range)
26400 ( )

7. Neutrophils are able to mediate anti–HIV antibody-dependent phagocytosis
gp120 protein
Fluorescent bead
Antibody
Fc Receptor
Neutrophil
Phagosome
25ug/ml IgG
Method adapted from Ackerman et al. 2011

8. Neutrophils are able to mediate anti–HIV ADCC responses
gp120
protein
Lysis
Antibody
CFSE viability stain
Fc Receptor
Pkh stained membrane
Loss of CFSE viability stain
25ug/ml IgG
Method adapted from Gómez-Románet al. 2006

9. The ADNP and ADCC responses correlate
25ug/ml IgG

10. Leukocyte cell prevalence
FcγRIIB
FcγRIIIA
FcγRI
FcγRIIA
FcγRIIB
B cells
~5%
FcγRI
FcγRIIIB
NK cells
~5%
Neutrophils
~65%
Monocytes
~5%
FcγRIIIA
T cells
~20%
FcγRIIB
FcγRIIA

11. How do neutrophil ADCC responses compare to other innate cells?

12. PBMC’s mediate enhanced levels of ADCC compared to NK cells at all antibody concentrations

13. Neutrophils perform similar levels of ADCC to PBMC’s

14. Monocytes perform similar levels of ADCC responses to Neutrophils and PBMC’s

15. How do neutrophil ADCC responses compare to other innate cells?

16. PBMC’s mediate enhanced levels of ADCC compared to NK cells

17. Monocytes mediate enhanced levels of ADCC compared to PBMC’s and NK cells

18. Neutrophils mediate rapid ADCC responses to HIV envelope protein

19. Summary and conclusions
Neutrophils can mediate HIV-specific antibody-dependent phagocytosis and ADCC responses
Neutrophils mediate ADCC responses faster than other innate effector cells
Rapid action and high magnitude of ADCC by neutrophils highlights their potential importance early in HIV infections

20. Future directions
Recruit a new cohort of a progressors and controllers
Do neutrophils utilise other antibody-dependent mechanisms to control HIV?
Do the RV144 vaccinees mediate antibody dependent functions with neutrophils?

21. Acknowledgment Peter Doherty Institute/ University of Melbourne
Amy Chung
Stephen Kent
Ivan Stratov
Matthew Parsons
Adam Wheatley
Vijaya Madhavi
Hyon-Xhi Tan
Sarah Lloyd
Fernanda Ana-Sosa Batiz
Kevin John Selva
Wen Shi Lee
Hillary Vanderven
Joshua Glass
Anne Kristensen
Vinca Alcantara
Thakshila Amarasena
Kirby Institute/ University of New South Wales
Anthony Kelleher
Ansari Shaik
Flow cytometry
Tina Luke
Members of the flow core
Funding
National Health and Medical
Research Council