Professeur Patrice DEBREProfesseur Patrice DEBRELaboratoire d’Immunologie Cellulaire et Laboratoire d’Immunologie Cellulaire et TissulaireTissulaireINSERM U945INSERM U945
HHôpital Pitié-Salpêtrière; Parisôpital Pitié-Salpêtrière; Paris
"Vers un vaccin contre le SIDA : "Vers un vaccin contre le SIDA : rôle des cellules NK"rôle des cellules NK"
PathogenPathogen PathogenicityPathogenicity
Measles Hépatitis Polio Rabies
Diphtéria
Tétanos
Cholera
VaccinesVaccines
Vaccine
INFECTION
virus
CD4 infected cells
PATHOGENESIS
CD4 non infected cells
NK cells
PATHOGENESIS
NK cells
CD4 non infected cells
Kill
NK
CD4
+-NKp44
MHC-I NKp44L
NKp44L expression by CD4+ T cells from HIV-infected patients
% N
Kp
44
L
0
10
20
30
40
3- 3-4+ 4+8+ 8+
Control PBMC HIV+ PBMC CD4 / mm3
0 250 500 750 10000
10
20
30
40 p=0.002
% N
Kp
44
L
E/T ratio
0
10
20
30
40
100/150/1 25/112.5/1
44L+
44L-
% N
K L
ysis
Vieillard et al Proc Natl Acad Sci USA (2005)
0.1 1 10 100 1000
viral load(x 1000)
0
10
20
30
40
% N
Kp
44
L
p=0.005
NKp44L expression is induced by a specific gp41 motif
HR1 HR2
NH2
Fusion Peptide TM
517 532 558 595
643 678
3S motif3S motif
HIV-1 gp41 Env
3S : very conserved motif & spécific to HIV-1
0
10
20
30
50/1 25/1 12/1 6/1E/T ratio
+3S+3S
0
10
20
30
40
+3S 1 10 20
anti-3S (g/ml)%
NK
lysis
Vieillard et al Proc Natl Acad Sci USA (2005)Vieillard et al AIDS (2006)
NKp44L
2.7% 17.4%
3SUT
CD
4
Anti-3S Ab : a predictive value for the evolution of the disease ?
0
100
200
300
0 400 800 1200
Anti-3S (U/mL)
CD4 count/mm3
Multivariate study between 40 patients (first quartile) with a slope CD4>2.7/month and 40 patients (last quartile) with slope CD4<-6.9/month
Variable Odds-ratio IC95 p
Sexe 0.372 M 1 W 1.83 0.49-6.87Age (x10y) 1.30 0.76-2.23 0.334CD4 (x100) 0.58 0.42-0.81 0.001Log VL 1.17 0.55-2.51 0.682 Anti-3S Ab 2.89 1.6-5.17 <0.001 (x100)
Vieillard et al AIDS (2006)Unpublished data
1.3: Anti-3S Ab : a predictive value for the 1.3: Anti-3S Ab : a predictive value for the evolution of the disease ?evolution of the disease ?
Vieillard et al AIDS (2006)
363024181260
500
300
100
-100
-300
-500
Time since baseline (months)
Change in CD4 count from baseline
Anti-3S= NEGAnti-3S= NEG(7 CD4 / month)(7 CD4 / month)
Anti-3S = POSAnti-3S = POS
Anti-3SAnti-3S
HIV-1HIV-1
NKp44LNKp44L
CD4CD4 NKNKLysisLysis
Summary-1Summary-1
Conclusion-1Conclusion-1
- NKp44L is specifically expressed on CD4+ T cells from HIV-1 patients;
- NKp44L is induced par a specific peptide from the gp41 HIV-1 protein;
- Anti-3S antibodies are predictive of the evolution of the disease evolution.
No expression of NKp44L in CD4+ T cells infected by HIV-1 : A new escape mechanism
mediated by Nef
Fausther Bovendo et al (In press)
100 101 102 103 104
p24FITC
CD4/AD8.002
100 101 102 103 104
p24FITC
CD4/AD8.002
NKp44L
HIV-p24
HIV viruses
Relative p24+NKp44L+
NKp44L expression
100 101 102 103 1040
100
150
200
50
52.1%
UI
100 101 102 103 104
WT
71.3%
100 101 102 103 104
GAG
68.1%
100 101 102 103 1040
100
150
200
50
67.2%
POL
100 101 102 103 104
73.6%
gp120
100 101 102 103 104
gp4190.4%
104 101 102 103 104104100 101 102 103
11.7%
NEF60.2%
VIF
100100 101 102 1030
100
150
200
50
TAT
72.4%
Rec
ombi
nant
vac
cini
a vi
ruse
s
0
5
10
15
20
WT nef
*
p24+NKp44L+
(flod increased)
WT nef G2A P76/79A L168/169A
HIV Nef variants
0
5
10
15
20 * *
Effect of Nef mutants on NKp44L expression
0
10
20
30
40
50
60
70
100/1 50/1 25/1 12.5/1
E/T ratio
% NK lysis
Effect of Nef mutants on NK cytotoxicity
Conclusion-2Conclusion-2
- NKp44L is non-expressed on HIV-infected CD4+ T cells;
- Retention of NKp44L by Nef HIV protein;
However,
- The mechanism of Nef inhibition remains unknown.
3- Vaccine strategies : Macaque model3- Vaccine strategies : Macaque model« proof of concept »« proof of concept »
3.1 : Macaque model of SHIV infection3.1 : Macaque model of SHIV infection
3.2 : Preventive vaccination3.2 : Preventive vaccination
SHIV162P3-infected macaques model :Relationship between CD4 cell decreases,
NKp44L expression and anti-3S Ab.
Days post-infection
CD
4+ /
mL
1000
1500
2000
2500
2500
500
0 50 100 150 2000
5
10
15
20
25
% N
Kp
44
L
an
ti-3
S (
U /
mL)
0
50
100
150
0 50 100 150 200 2500
5
10
15
20
25
% N
Kp
44L
Vieillard et al AIDS (2008)
Proofs of concept : Immunization of macaques by the 3S peptide
KLH KLH-3S
CD
4 /
mm
3
0
500
1000
1500
2000
0 50 100 150 200
** *
Vir
al lo
ad
1
102
104
106
108
0 50 100 150 200
NS NS
Vieillard et al Proc Natl Acad Sci USA (2008)
0
10
20
30
40
50
0 50 100 150 200
% C
D4
+N
Kp
44
L+
0
500
1000
1500
-400 -200 0 200
an
ti-3
S (
U/m
L)
SHIV162P3
3S3S
3S3S
Days post-infection
Effect of 3S vaccination on CD4 apoptosis, NK activity et T cell-activation
% C
D4
+A
nn
exi n
V+
Days post-infection
0
10
20
30
40
50
UI 14 21 42 LN
*
**
*
**
% N
K lysis
Days post-infection
0
25
50
75
100
0 14 21 42 LN
**
*****
Vieillard et al Proc Natl Acad Sci USA (2008)
0 20 40 60 80
Days post-infection
% C
D4
+C
D6
9+
0
2.5
5
7.5
10
Conclusions1- Pathophysiology : role of NKp44L
- Depletion of non-infected CD4+ cells - In infected cells: Escape mechanism by Nef
2- Immunotherapic interventions
- Preventive vaccine : in macaque model, we have observed :
- high production of anti-3S Ab; - low expression of NKp44L on CD4 cells.
Stable level of the CD4 cells in absence of effect on the
viral load !
Pitié-Salpêtrière Hospital, Paris
France
Laboratory of Cellular and Tissular Immunology
Hugues Fausther Bovendo
Vincent Vieilllard
Patrice Debré
Clinical Epidemiology
Dominique Costagliola
Department of virology
Henry Agut
Daniel Candotti
Acknowlegments
CEA, Fontenay-aux-roses, France
Laboratory of Immuno-Virology
Roger Le Grand
Nathalie Bosquet
Pasteur Institute, Paris, France
Department of virology
Nathalie Sole-Foulon
Olivier Schwartz
Harvard University, Cambridge, MA
Jack Strominger
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