Immunogenicity of anti‐TNF agents
Towards personalised treatment
Gertjan Wolbink
Rheumatologist
Reade Rheumatology
Sanquin Immunopathology
G.wolbink@reade.nl
Five anti‐TNF agents are currently approved for the treatment of rheumatoid arthritis
Etanercept
Certolizumab pegol
TNF
receptor
Adalimumab
Golimumab
Infliximab
(Fab’)2
Fc
region
van Schouwenburg PA, et al. Nat Rev Rheumatol. 2013;9:164‒172
Permission obtained from Nature Publishing Group©
van Vollenhoven RF, et al. Nat Rev Rheumatol. 2011;7:205–215
Persistent disease
TNF‐ independent or
incomplete blocking of TNF ?
Amsterdam READE cohort
Long‐term clinical and serological follow‐up of 2000 patients on biologicals*
Ankylosing spondylitis (AS)
Rheumatoid Arthritis (RA) Psoriatic Arthritis (PsA)
*infliximab/adalimumab/etanerceptabatacept/golimumab/tocilizumab/rituximab
Pharmacokinetic assay (drug level test anti‐TNF)
Anti-idiotype
Adalimumab/infliximab/
golumimab/etanercept
TNF
Anti-TNF
Serum trough iInfliximab level (mg/l)
Ankylosing spondylitis:
infliximab levels correlate with response
30
P=0.018
20
10
0
Responders
Non‐responders
Serum trough infliximab level for responders (n=21; 8.2mg/l) and non‐responders
(n=17; 6.3mg/l) according to the ASAS‐20 response criteria, at week 54 (P=0.018)
de Vries MK, et al. Ann Rheum Dis. 2007;66:1252–1254
Copyright ©2007 BMJ Publishing Group Ltd.
Patients with an allergic reaction to infliximab have low serum levels of infliximab
Infliximab concentration
(MG/L)
2 wk
6 wk
14 wk
Mean all patients n=105
23.9
16.0
4.6
Pt S reaction at wk 14
17.4
0.5
0.00
Pt R reaction at wk 14
37.1
2.8
0.00
Wolbink GJ, et al. Ann Rheum Dis. 2005;64:704‒707
Detection of antibody formation
against therapeutic antibodies
Aarden L, et al. Curr Opin Immunol. 2008;20:431‒435
Anti-IgG
MT
Coat: monoclonal therapeutical
Patient IgG
Anti-IgG
Antigen binding test
Wolbink et al:
Arthritis Rheum. 2006
Mar;54(3):711-5
ProtA
IgG
125I-F(ab)
2
Anti-adalimumab
Adalimumab
Adalimumab
Neutralise
pH shift anti-idiotype
Antigen Binding Test
Rabbit-F(ab)anti-idiotype
acid treatment pH 2.5
Detection of anti‐drug‐antibodies (ADA)
Anti-TNF antibody levels
ADA production
Free anti-TNF agent
ADA-drug complexes
Free ADA
ADA detection method
ELISA
–
–
–
+
ABT
–
–
+/–
+
PIA
–
+/–
+
+
++
+
+/–
–
Pharmacokinetic assay
(TNF capture)
van Schouwenburg PA, et al. Nat Rev Rheumatol. 2013;9:164‒172
Generation of human monoclonal antibodies derived from patients producing ADA
Tool to study epitopes involved
“Are there different epitopes involved”
“Is there one dominant immunogenic epitope”
“Do the immunogenic epitopes differ between patients”
“What part of the ADA are neutralising”
Standardisation of assays
van Schouwenburg PA, et al. Ann Rheum Dis. 2013;72:104‒109
Generation of human monoclonal antibodies
Laser
Centrifuge
400 x g
30 minutes
B‐
cell
Sort antigenspecific cells
Nozzle
tip
Point of
analysis
Break-off point
Deflection
plate
+
+
Isolate B cells
Isolate PBMCs
+
+
Nea I
pUC origin
AmC ORF Bam HI
bl2 pronote2
-
Cla I
Xma I
Sma I
Bam III
attB I
CleI
Expression
Vector1
7772 bp
Ampicillin
resistance gene
-
…AGGCATATCGA
LacZ ORF
Xma I
Hias III
Eca RI
Sam I
attIs2
Recombinant expression
monoclonal antibodies
Isolate RNA, determine
sequence for VH/VL
van Schouwenburg PA et al. Ann Rheum Dis 2013;72:104‐9.
Culture 1 cell/well;
screening
ECRF bio‐assay: TNF‐sensitive human endothelial cell line
TNF α
TNF α + adalimumab
No IL-8 production
IL 8 pg/ml
4000
3000
2000
Pg/ml IL-8 production
IL-8 production
2000
1500
1000
500
0
1000
0
0 0.937 1.875 3.75 7.5
15
30
Adalimumab ng/ml
van Schouwenburg PA, et al. Ann Rheum Dis. 2013;72:104‒109
60
-
ADA IFX ADA IFX ADA IFX
1.1
1.2
Anti‐drug antibodies are anti‐idiotypic
and interfere with TNF binding
Anti‐adalimumab
Adalimumab
TNF can be active again
Wolbink GJ, et al. Arthritis Rheum. 2006;54:711–715
Pharmacokinetics
• No anti‐infliximab detected
• Anti‐infliximab detected
infliximab
80
anti-infliximab
infliximab mg/L
infliximab mg/L
infliximab
60
40
20
0
-2 2
6
10 14 18 22 26 30
Weeks
Wolbink, data on file
Adapted from Wolbink GJ, et al. Curr Opin Rheumatol. 2009;21:211–215
80
anti-infliximab
60
40
20
0
-2 2
6 10 14 18 22 26 30
Weeks
Tolerance !!
PK model anti‐infliximab detected transient antibody formation
Infliximab mg/L
80
Infliximab
Anti‐infliximab
60
40
20
0
2
6
10
14
Weeks Wolbink, data on file
Adapted from Wolbink GJ, et al. Curr Opin Rheumatol. 2009;21:211–215
18
22
26
30
Immunogenicity in a long‐term follow‐up cohort of adalimumab
treated rheumatoid arthritis patients
American College of Rheumatology 2010
Bartelds GM, et al. JAMA. 2011;305:1460‒1468
Patients & methods
1
2
3
4
Bartelds GM, et al. JAMA. 2011;305:1460‒1468
Results: baseline characteristics
Total Patient with AAA
Patients without AAA
n=272
n=76
n=196
Age, years
54 ± 12
53 ± 13
54 ± 11
Female, no. (%)
219 (81)
62 (82)
157 (80)
RF, no. (%)
196 (72)
57 (75)
139 (71)
Prior DMARDs
3.1 ± 1.4
3.4 ± 1.5*
3.0 ± 1.3*
MTX use, no. (%)
202 (74)
41 (54)*
161 (82)*
MTX dose (mg/wk)
25 (15‒25)
18 (10‒25)*
25 (15‒25)*
No DMARD, no. (%)
51 (19)
28 (37)*
23 (12)*
Disease duration (years)
8 (3‒17)
12 (5‒18)*
8 (3‐16)*
Erosive disease, no. (%) 201 (74)
63 (83)*
138 (70)*
ESR (mm/h)
23 (11‒42)
35 (18‒60)*
21 (11‒39)*
CRP (mg/L)
12 (5‒29)
19 (7‒46)*
11 (4‒22)*
DAS28 5.2 ± 1.2
5.5 ± 1.1* 5.1 ± 1.3*
Bartelds GM, et al. JAMA. 2011;305:1460‒1468
Sustained remission (DAS28 <2.6) correlates with absence of AAA
AAAAAA+
Remission probability
0.5
0.4
0.3
P<0.0001
0.2
0.1
0.0
0
50
100
150
200
Weeks
After adjustment for confounding variables MTX dosage, ESR and CRP (HR: 3.6; 95% CI 1.8‒7.2, P<0.0001)
Bartelds GM, et al. JAMA. 2011;305:1460‒1468
Percentage of patients developing
anti‐adalimumab antibodies over three years
Antibodies against adalimumab (%)
Percentage of patinets
developing AAA
30
25
20
15
51 of 76 patients (67%) developed AAA during the first 28 weeks of treatment
10
5
0
0
28
56
84
112
Weeks During 156 weeks follow‐up, anti‐adalimumab antibodies
were detected (ASSAY IV:ABT) in 76 (28%) patients
Bartelds GM, et al. JAMA. 2011;305:1460‒1468
140
Methotrexate reduces immunogenicity
in adalimumab‐treated RA
No MTX
Low dose MTX
Intermediate dose MTX
High dose MTX
Percentage of patients developing
AAA
60
50
50%
40
30
35%
20
22%
10
14%
0
0
14
28
42
56
70
84
Weeks Jan van Breemen Research Institute | Reade
EULAR centre of Excellence in Rheumatology
Krieckaert CL, et al. Ann Rheum Dis. 2012;71:1914‒1915
98
112 126 140 154
The accumulative percentage of ADA positive patients depends on assay method used
Antigen binding test
pH‐shift‐anti‐idiotype antigen binding test
% ADA positive
60
40
P<0.0005
20
0
4
16 28
52
78
104
130
156
Weeks Accumulative percentage of patients positive for ADA assessed by
pH‐shift‐anti‐idiotype antigen binding test and antigen binding test
van Schouwenburg PA, et al. Ann Rheum Dis. 2013;72:1680‒1686
Median adalimumab concentration
Free serum adalimumab concentrations
in relation to AAA
No AAA
AAA 13‒100 AU/ml
AAA >100 AU/ml
14
12
10
*
8
P<0.0001*
6
4
2
0
0
16
40
78
Weeks Bartelds GM, et al. JAMA. 2011;305:1460‒1468
130
Mean DDAS28
Adalimumab concentration correlates with treatment efficacy
4.5
4.0
3.5
3.0
2.5
2.0
1.5
1.0
0.5
0.0
‐0.5
‐1.0
‐1.5
2
4
6
8
10
12
14
16
18
Adalimumab concentration (µg/mL)
Mean delta DAS and adalimumab levels per 20 patients at week 28
Pouw M, et al. Ann Rheum Dis. 2013 [ePub]
Anti‐drug antibodies have a large impact on treatment in clinical practice
Lower functional drug levels
Higher disease activity Less remission
Bartelds GM, et al. JAMA. 2011;305:1460‒1468
Implications of immunogenicity
for non‐responders
Jamnitski A, et al. Ann Rheum Dis. 2011;70:284–288
Bartelds GM, et al. Ann Rheum Dis. 2010;69:817–821
RA patients treated with etanercept
First anti‐TNF
n=203 (anti‐TNF naïve)
Second anti‐TNF
n=89 (switchers)
(from adalimumab or infliximab to etanercept)
47 (53%) positive for anti‐inflix/anti‐adalimumab
42 (47%) negative for antidrug antibodies
Outcome
Clinical response (delta DAS28) at 28 weeks
Jamnitski A, et al. Ann Rheum Dis. 2011;70:284‒288
Naïve versus switchers to etanercept
Delta DAS28 at 6 months
Anti‐TNF‐α naïve patients
(n=203)
2.1 (SD1.3)
Switchers without immunogenicity
(n=42, 47%)
1.2 (SD1.3)
**
Switchers with immunogenicity
(n=47, 53%)
2.0 (SD1.3)
**P=0.017
Jamnitski A, et al. Ann Rheum Dis. 2011;70:284–288
Naïve versus switchers to adalimumab
Anti‐ADA
formation
Delta DAS28
Delta DAS28
anti‐ADA
Anti‐TNF‐α naïve patients
(n=183)
32 (18%)
1.7 (SD1.5)
2.0 (SD1.4)
Prior IFX patients without anti‐IFX
(n=19)
3 (16%)
0.9 (SD1.5)
0.9 (SD1.4)
Prior IFX patients with anti‐IFX
(n=33)
11 (33%)
1.2 (SD1.3)
1.6 (SD1.1)
IFX = infliximab
prior IFX patients = patients who were treated with infliximab before adalimumab treatment
Anti‐IFX = anti‐infliximab antibodies
Anti‐ADA = anti‐adalimumab antibodies
Delta DAS2* = Delta DAS28 with patients with anti‐adalimumab excluded from analysis.
Bartelds GM, et al. Ann Rheum Dis. 2010;69:817–821
Conclusions
Amsterdam‐Immunogenicty
(www.biologicals@sanquin.nl)
SANQUIN Immunopathology
• Lucien Aarden
• Theo Rispens
• Diana Wouters
• Pauline van Schouwenburg
• Desiree van der kleij
• Margreet Hart
• Simone Kruithof
• Karin van Schie
• Karien Bloem
READE Rheumatology
• Mike Nurmohamed
• Charlotte krieckaert
• Anna Jamnitski
• Eva Kneepjes
• Mieke Pouw