uth rs diabetes dhu adrenal diseases...
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Département Hospitalo-Universitaire
AUToimmune and HORmonal diseaseS
Uth rs
DHU
DIABETES
ADRENAL DISEASES
VASCULITIDES
L’immunomodulation du diabète. Quelles applications? quel avenir ?
TYPE 1 DIABETES MELLITUS
• insulitis
Gepts W 1965, Diabetes
Gianani R 2010, Diabetologia
b cell selectivity
an autoimmune disease
• islet cell autoantibodies
Bottazzo & Doniach 1974, Lancet
anti-GAD
anti-IA2
anti-insulin
anti-ZnT8
increasing incidence ( 3-4%/year) & geographical
incidence variation (> 65/105/year in Finland)
0
20
40
60
1921 50s 1972 1992
incid
en
ce [
105/y
ear]
absence of a unique environmental factor
genes
environment numerous
numerous
Allegheny County
childhood-onset (< 18) T1D registry
diagnosed 1965-79
Secrest et al. Diabetes 2010
acute
renal
cardio
vascular
infections
cancer
n = 1075
age
Controls
mortality rate
/10 3 person-yr
standardized
mortality rate
♂
0-4 1.4880 2.0
5-9 0.1511 3.7
19-14 0.2218 4.6
15-19 0.7166 3.3
20-24 0.8752 2.3
25-29 0.9381 4.5
30-34 1.1388 6.3
35-39 1.4270 3.3
♂
0-4 1.2640 0.0
5-9 0.1550 0.0
19-14 0.1289 4.6
15-19 0.2802 5.8
20-24 0.2927 3.2
25-29 0.3477 4.6
30-34 0.5285 5.1
35-39 0.7086 2.7
Skrivarhaug et al. Diabetologia 2006 [Norway]
T1D MORTALITY
3.9
4.0
increased mortality compared to
the general population
Intermediate disease compared
to high risk autoimmune diseases
TYPE 1 DIABETES: A CHRONIC DISEASE ß
ce
ll m
as
s
diabetes
hyperglycemia
Bingley et al. Diabetologia 2006 heterogeneity
patient duration of
diabetes
diabetes
recurrence Insulitis ICA
1 22 years 60 days CD8+ T cells -
2 17 years 44 days CD8+ T cells -
3 27 years 92 days CD8+ T cells +
b cell
GRZ/perforin
APC
Cl II
Cl I Cl II
TNF-a, IFN-g… T CD4
autoantibodies
NKT
CD1d
IL-4, IFN-g…
NK
NKG2D/MIC
IFN-g…
T reg
IL-10, TGF-b
TYPE I DIABETES
Pancreatic islet
b cell
GRZ/perforin
APC
Cl II
Cl I Cl II
TNF-a, IFN-g… T CD4
TYPE I DIABETES
Pancreatic islet
immunosuppression
b cell mass restoration
graft
regeneration
insulin replacement
therapy
tolerance induction
b cell
GRZ/perforin
APC
Cl II
Cl I Cl II
TNF-a, IFN-g… T CD4
TYPE I DIABETES
Pancreatic islet
cyclosporin A
azathioprine + corticoids
anti-thymoglobulin ± corticoids
anti-CD3
autoantibodies
anti-CD20
[Rituximab]
anti-TNFa
[Etanercept]
IL1 antagonists
[Anakinra]
intensive
Insulin
therapy
nicotinamide
TREATMENT OF DIABETES WITH ANTI-CD3
Herold KC et al. 2002, N Engl J Med
+0.20 ± 1.86 nmol/l
per month
-5.52 ± 1.30 nmol/l
per month
Keymeulen B et al. 2005, N Engl J Med
months months months
n = 40 (placebo) n = 40 (anti-CD3)
TREATMENT OF DIABETES WITH ANTI-CD3
Keymeulen B et al. N Engl J Med 2005
TREATMENT OF DIABETES WITH ANTI-CD3
b cell
GRZ/perforin
APC
Cl II
Cl I Cl II
TNF-a, IFN-g… T CD4
TYPE I DIABETES
Pancreatic islet
insulin
GAD IA2,ZnT8 DiaPep277
T reg
IL-10, TGF-b
extrinsic T cell tolerance
peptides
intrinsic T cell tolerance • anergy
• changes in gene expression profiles
low-dose IL-2
Rapamycin/IL-2
84 228 first and second degree relatives
3152 autoantibody positive
5 year risk > 50% in 372/2103
randomization to
observation (n = 70)
versus
s.c. insulin twice daily 0.25 units/kg (n = 69)
mean follow up 3.7 years
TREATMENT OF PREDIABETES WITH INSULIN
s.c./i.v. & oral insulin (DPT1, 2002, N Engl J Med)
Nasal insulin (Näntö-Salonen, 2008, Lancet)
103391 first and second degree relatives
3483 autoantibody positive
5 year risk 26-50% in 388/2103
randomization to
oral insulin 7.7 mg/24h (n = 186, mean IAA 382 ± 555 nU/ml)
versus
placebo (n = 186, mean IAA 346 ± 336 nU/ml)
TREATMENT OF PREDIABETES WITH ORAL INSULIN
Diabetes Prevention Trial-Type 1
Skyler J et al Diabetes Care 2005
6.4%/year
8.2%/year
IAA ≥ 80 nU/ml
6.2%/year
10.4%/year
Ludvigsson et al, N Engl J Med, 2008
n = 35
n = 34
20 mg GAD-alum days 1 & 30
within 6 months following onset
response to GAD up to 15 months
IL-5/-10/-13/-17/IFNg/TNFa
~ IL-6/-12
GAD-induced expression of
FoxP3 & TNFa
GAD VACCINATION IN RECENT-ONSET T1D
[Intention to treat analysis]
TrialNet intervention studies: within 100 days from diagnosis
• Anti-IL1β [Canakinumab]
• CTLA-4 Ig [Abatacept] in Recent Onset Diabetes
• Glutamic acid decarboxylase (GAD) in New Onset T1DM
• Metabolic Control in New Onset Diabetes
• Rituximab Study (Anti-CD20)
• Mycophenolate mofitil / dacliumab (MMF/DZB) study
http://www.diabetestrialnet.org: accessed 1st March, 2014.
TrialNet prevention studies • Oral insulin for prevention of Type 1 diabetes Study
• Anti-CD3 [Teplizumab] for prevention in relatives
• CTLA4-Ig [Abatacept] for prevention in relatives
• The Nutritional Intervention to Prevent Type 1 diabetes Study
T CD8
b cell
T CD4 TNF-a, IFN-g…
APC
TYPE 1 DIABETES T CELLS TO DIABETES PREVENTION
CHAINE B
MALLVHFLPLLALLALWEPKPTQAFVKQHLCGPHLVEALYLVCGERGFFYTPK SRREVEDPQVEQLELGGSPGDLQTLALEVARQKRGIVDQCCTSICSLYQLENYCN
MALWMRFLPLLALLFLWESHPTQAFVKQHLCGSHLVEALYLCGERGFFYTPM SRREVEDPQVAQLELGGGPGAGDLQTLALEVAQQKRGIVDQCCTSICSLYQLENYCN
B9-23
PEPTIDE C CHAINE A
TRREAED 61LQVGQVELGG 71GPGAGSLQPL 81ALEGSLQKRG 91IVEQCCTSIC101SLYQLENYCN
HUMAN INSULIN
1MALWMRLLPL11LALLALWGPD 21PAAA FVNQHL 31CGSHLVEALY 41LVCGERGFFY 51TPK
PEPTIDE SIGNAL
peptide sequence class I allele
2-11
ALWMRLLPLL
A2
A24
ALWMRLLPLL B8
6-14 RLLPLLALL
A2
RLLPLLALL A26
6-16 RLLPLLALLAL
A2
14-23 LALWGPDPAA
A2
LALWGPDPAA A26
15-24 ALWGPDPAAA
A2
ALWGPDPAAA A26
Toma et al, PNAS 2005; Diabetes 2009
peptide sequence HLA class I
34-42 HLVEALYLV A2
38-46 ALYLVCGER
A3
A11
39-48 LYLVCGERGF A24
41-50
LVCGERGFFY A1
LVCGERGFFY
A3
A11
42-51
VCGERGFFYT A1
VCGERGFFYT A2
VCGERGFFYT B8
VCGERGFFYT B18
44-51
GERGFFYT A1
GERGFFYT B8
45-53 ERGFFYTPK A3
49-57 FYTPKTRRE B8
51-61 TPKTRREAEDL B8
CLASS I ALLELE-SPECIFIC PEPTIDES
peptide binding % CD8+ IFN♂ response
10-4
M 10-6
M long
standing
recent
onset
total
34-42* 58% 37% 2/4 4/10 6/14
6-14* 36% 65% 7/13 1/4 8/17
6-16* 96% 71% 5/15 2/4 7/19
14-23* 44% 36% 7/13 0/4 44%
15-24* 96% 94% 7/13 1/4 8/17
Toma et al, PNAS2005; Toma et al. Diabetes 2009
CD8+ T CELL RESPONSE TO A2-RESTRICTED PREPROINSULIN PEPTIDES [ELISPOTassay]
CD8+ IFN♂ response
long
standing
recent
onset
total
2/4 4/10 6/14
7/13 1/4 8/17
5/15 2/4 7/19
7/13 0/4 7/17
7/13 1/4 8/17
* Val42
, L14
, L16
, A23
& A24
were identified as C-terminal residues
generated by proteasome digestion in vitro.
** 6-14
or
6-16
immunization
anti-6-14
or
anti-6-16
T cell clones
cross-reactives responses
to 6-14 and 6-16
class I preproinsulin peptide
A2.1 2-11
A2.1 6-14
A2.1 15-24
A2.1 30-39
A2.1 33-42
A2.1 34-42
A2.1 42-51
A2.1 101-109
controls PDHase 2008-216 MATA2 58-66 Nef
83-91
b cells
TCD8+
TETRAMER RECOGNITION OF BLOOD CD8+ T CELLS
class I
peptide
biotin
streptavidin
100
101
102
103
104
100
101
102
103
104
Dt1A5 TTM A2 PPIh_Dt1A5 TTM A2 33-42.fcsÉCD3+ CD8+
PerCP-A: CD8 PerCP-A
PE
-A: T
TM
PE
-A
0.810 0.79
99.20
100
101
102
103
104
100
101
102
103
104
Dt1A5 TTM A2 PPIh_Dt1A5 TTM A2 6-14.fcsÉCD3+ CD8+
PerCP-A: CD8 PerCP-A
PE
-A: T
TM
PE
-A
0.0370 0.037
1000
100
101
102
103
104
100
101
102
103
104
25-01-08_D124D 6-14.fcsÉCD3+ CD8+
PerCP-A: CD8 PerCP-A
PE
-A: T
TM
PE
-A
0.60 0.54
99.60
Sandrine Luce
TCD8+
A2.1-restricted peptides
Luce et al. Diabetes 2011
Luce et al. Diabetes 2011
central memory T cells effector memory T cells
single cell PCR
A2.1-restricted peptides
TETRAMER RECOGNITION OF BLOOD CD8+ T CELLS
s.c./i.v. & oral insulin (DPT1, 2002, N Engl J Med)
Nasal insulin (Näntö-Salonen, 2008, Lancet)
NEW PRECLINICAL T1D MODELS
insulin/A2.1/DQ8 mice
YES
Insulin
A2.1
DQ8 genes
OUF
A2/DQ8 Tg Human insulin
x
Luce et al. unpublished
♂ ♂ ♀ ♀
YES
YES Col
Col
Intraperitoneal glucose tolerance test
AU
C m
g.h
/dL
beta cell mass
mg
DIABETES IN RIP-B7 TRANSGENIC YES MICE
T cell
Age (weeks)
dia
be
tes (
%)
% t
ota
l is
lets
non diabetic diabetic
CD4
CD
8
CD11b
CD
11
c
CD19
TC
R♂
dapi anti-glucagon anti-CD3
RIP-B7 YES B7.2
CD28
CD4
MHC
TCR
Luce et al. unpublished
DIABETES IN RIP-B7 TRANSGENIC YES MICE
BrDU proliferation assay CD8+ IFNγ Elispot assay
CD8+ TMr+ expansion assay
Luce et al. unpublished
FROM PALLIATIVE TREATMENT TO PREVENTION OF TYPE 1
DIABETES
diabetes
T CD4
APC
T CD8
NKT NK
T reg
B
damaged
islet of Langerhans
FROM RESEARCH TO CLINIC
diagnosis immunotherapy
YES mice
Luce S et al. Diabetes 2011 Carlier et al. PloSOne 2012
Class II-restricted epitopes
modified by inclusion of a
thiol-disulfide oxidoreductase
motif within flanking residues
Immunotherapy
EU FP7 project
HLA peptide
autoantigen
T diagnostics
prediabetes
Uth rs
DHU
Département Hospitalo-Universitaire
AUToimmune and HORmonal diseaseS
Uth rs
DHU
DIABETES
ADRENAL DISEASES
VASCULITIDES
Andrea Toma Sandrine Luce
Etienne Larger Roberto Mallone
François Lemonnier
Agnès Lehuen
Sylvianne Muller, Jean Paul Briand (CNRS, Strasbourg)
Decio Eizirik (Brussels)
Charbel Masaad (Paris)
Laurent Drouot,
Olivier Boyer (Rouen)
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