Clinical implications of immunogenicity of TNF inhibitors · Clinical implications of...
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22-09-08 Clinical implications of immunogenicity Clinical implications of immunogenicity of TNF inhibitors Theo Rispens
Transcript of Clinical implications of immunogenicity of TNF inhibitors · Clinical implications of...
22-09-08
Clinical implications of immunogenicity Clinical implications of immunogenicity of TNF inhibitors
Theo Rispens
DisclosureIn relation to this presentation, I declare the following, real or perceived conflicts of interest:- received fees for lectures from Pfizer, AbbVie, Regeneron
A conflict of interest is any situation in which a speaker or immediate family members have interests, and those may cause a conflict with the current presentation. Conflicts of interest do not preclude the delivery of the talk, but should be explicitly declared. These may include financial interests (eg. owning stocks of a related company, having received honoraria, consultancy fees), research interests (research support by grants or otherwise), organisational interests and gifts.
TNF blockersInflammation in e.g.
• Rheumatoid arthritis• Crohn’s disease• Psoriasis
can be surpressed by blocking TNF:
Infliximab AdalimumabEtanercept Golimumab Certoluzimab20021998 2008 20091998
Specificity: cross-reactive & pre-formed antibodies
complementarity determining regions
framework regions
VL
VH
CH1
idiotopes &
xenotopes
glycans FabregionsCL
CH1
CH2
CH3
allotopes
Fc
van Schie et al. mAbs, 2015, 7:4, 662-671
Immunogenicity therapeutic antibodies
Mouse HumanizedChimeric HumanInfliximab Adalimumab
Immunogenicity:
3-5% mouse30% mouse
Antigen binding test (ABT)
- capture IgG from serum (Prot. A Sepharose)
prot. A
- capture IgG from serum (Prot. A Sepharose)- wash out unbound material- incubate with radiolabeled adalimumab F(ab’)2 - wash out unbound label- measure radioactivity-Does not measure rheumatoid factors
25
30
Adalimumab: Cohort 272 RA patients• 3 year follow-up• Long term measurement of adalimumab levels, ADA and disease activity.
0
5
10
15
20
0 28 56 84 112 140
Antibodies againstadalimumab (%)%
ADA
+
Bartelds et al., JAMA April 2011
Remission (DAS28 < 2.6) is associated with absence of ADA
p<0.00010.3
0.4
0.5AAA -
Remi
ssion
prob
abilit
y
Gecorrigeerd voor MTX dosis, bezinking en CRP (HR:3.6; 95%CI:1.8-7.2, p<0.0001)
0 50 100 150 2000.0
0.1
0.2
AAA+
Time in weeks
Remi
ssion
prob
abilit
y
Bartelds et al., JAMA April 2011
Infliximab RA 22/51 pt (43%) develops antibodies within 1 year
Wolbink et al. Arthritis Rheum 2006
Anti-adalimumab measured in bridging ELISA and antigen binding test
prot. A α-IgG4
� different numbers of ADA+ patients in different assays
7% 13% 22%
bridg. ELISA ABT IgG4-ABT
1
10
100
1000
10000
100000
−−−− −− −− −− −− −− −− −− −
−− −− −− −− −− −− −− −− −− −
−− − − −−− −− − −− − −− −−− −
* x201
* x187 * x168
AU/m
l
7% 13% 22%
Hart et al., JIM 2011
Relation between drug levels and ADA
100
1000
10000
100000
ABT
AU/m
l
10
100
1000
10000
100000
1000000
bridg
ing EL
ISA A
U/ml
� bridging elisa: detects ADA only if no adalimumab is detected
prot. A
10
100
1000
10000
100000
0.001 0.01 0.1 1 10 100adalimumab µg/ml
IgG4-A
BT A
U/ml
190.002
100.001 0.01 0.1 1 10 100
adalimumab µg/ml
120.002
0
10
20
30
40
50
ELISA ABT IgG4-ABT
numb
er of
patie
nts
drug negdrug pos
1
10
0.001 0.01 0.1 1 10 100adalimumab µg/ml
bridg
ing EL
ISA A
U/ml
0.002 2
α-IgG4
Hart et al., JIM 2011
1. acid
2. neutralize / ADL F(ab)2-bt
1. acid
2. neutralize / anti-ADL Fab
ARIA
PIAProt. A
Sepharose
125I
bt bt streptavidin
complex dissociation/competition detection
ADL F(ab)2-bt / 37 oC / 16 hr
1. acid
2. neutralize / ADL-bt / ADL-
sulfo-tag
bt
bt
TRIA
ECL
sulfo-tag
ADA adalimumab (ADL) rabbit anti-ADL Fab
streptavidin
bt bt
Bloem et al, J Immunol Methods 2015
drug-tolerant ADA: analysis in ADL-treated RA patients
ARIA TRIAABT
PIA ECL
ABT ARIA TRIA PIA ECL% 14.9 66.0 57.4 51.1 57.4
Bloem et al, J Immunol Methods 2015
Clinical response vs drug-tolerant ADA assayRA patients/adalimumab
van Schouwenburg et al. ARD 2013
(wk)
NS
PK assay
Anti-idiotype
adalimumab
Anti-TNF
TNF
Drug levels vs anti-drug antibodies: a balance16 or 28 weeks after start adalimumab
10000
100000
ADA (
ABT;
AU/m
L) 10000
100000
ADA (
PIA; A
U/mL
)
ABT vs PK PIA vs PK
10
100
1000
0.001 0.01 0.1 1 10 100adalimumab (ug/mL)
ADA (
ABT;
AU/m
L)
10
100
1000
0.001 0.01 0.1 1 10 100adalimumab (ug/mL)
ADA (
PIA; A
U/mL
)
Detection of anti-drug-antibodies (ADA)
Anti-TNF antibody levelsADA productionFree ADAADA-drug complexesFree anti-TNF agent
van Schouwenburg PA, et al. Nat Rev Rheumatol. 2013;9:164‒172
ADA detection method
ELISA – – – +ABT – – +/– +PIA – +/– + +Pharmacokinetic assay(TNF capture) ++ + +/– –
2.0
2.5
3.0
3.5
4.0
4.5A.
DAS28
Concentration-effect curve (adalimumab/RA)
2 4 6 8 10 12 14 16 18
-1.5
-1.0
-0.5
0.0
0.5
1.0
1.5
2.0
adalimumab concentration (ug/mL)
mea
n∆ D
AS28
Each dot is mean of 20 patients
Pouw et al. ARD 2015
Characterization of anti-adalimumab antibodiesCharacterization of anti-adalimumab antibodies
Making human monoclonal antibodies
Schouwenburg et al., JBC 2014
All monoclonal antibodies are derived from different precursor B-cells
clone isotype V D J R1.1 IgG1 V1-03*01 D2-02*01 J4*02 161.2 IgG4 V1-02*02 D6-13*01R J5*01 91.3 IgG1 V1-18*01 D1-14*01 J4*02 92.1 IgG1 V1-03*01 D3-09*01 J4*02 162.2 IgG4 V1-69*06 D3-10*01 J6*02 152.3 IgG4 V4-31*03 D6-13*01 J3*02 212.4 IgG1 V1-03*01 D3-16*01 J6*02 192.5 IgG4 V4-59*01 D6-13*01 J3*02 162.6 IgG1 V4-39*01 D6-19*01 J4*02 142.7 IgG1 V4-34*01 D1-26*01 J3*02 52.8 IgG4 V1-69*06 DIR1*01R J5*02 212.9 IgG1 V3-48*03 D2-21*02 J6*02 23
adalimumab
Schouwenburg et al., JBC 2014
2.9 IgG1 V3-48*03 D2-21*02 J6*02 232.1 IgG4 V1-03*01 D5-12*01 J5*02 15
2.11 N.D. V1-69*01 D6-19*01 J4*02 142.12 IgG4 V1-18*01 D2-21*01R J6*02 252.13 IgG1 V3-48*03 D5-05*01 J3*02 92.14 IgG1 V3-21*01 D3-16*02 J4*02 13
1.1 N.D. V1-46*01 D3-09*01 J4*02 251.2 IgG1 V4-34*01 D5-05*01 J4*01 101.3 IgG1 V1-18*01 D2-15*01 J3*02 111.4 IgG4 V1-18*01 D2-15*01 J4*02 112.1 IgG1 V1-69*01 D2-02*01 J4*02 82.2 IgG4 V3-74*01 D3-10*01 J3*02 172.3 IgG4 V3-30*01 D5-05*01 J4*02 212.4 IgG4 V3-74*01 D3-10*01 J3*02 20
infliximab
IgG4 anti-adalimumabadalimumab-treated RA patientsin patients without detectable drug levels: significant part is IgG4
IgG (AU)
Schouwenburg et al., J Clin. Immunol 2012; Ann. Rheum. Dis. 2013
Mabs underwent extensive somatic hypermutationA
V H1V H
3V H
4
0
5
10
N
V K1V K
3V K
4
0
5
10
N
1 0- 1 2
1 3- 1 5
1 6- 1 8
1 9- 2 1
0
2
4
6
N
VH gene usage Vκ gene usage VH CDR3 length
VLVH
Fab
Fc1000
10000100000
Kd (p
M)
R S R S R S R S
0
10
20VH VL
Mutat
ion fre
quen
cy (%
)
FR CDR CDRFRR S R S
0
10
20
30 VH VL
no. m
utat
ions
0.11
10100
1000
Kd (p
M)Schouwenburg et al., JBC 2014
mAbs neutralize adalimumab
150
TNF TNF + ADL TNF + ADL + ADA
IL-8 no IL-8 IL-8
T N F
T N F + A D L 1 . 2 1 . 3 2 . 1 2 . 2 2 . 4 2 . 6 2 . 7 2 . 8 2 . 9 2 . 1
02 . 1
2
0
50
100
150IL-
8 (%)
TNF + ADL + ADA
Schouwenburg et al., JBC 2014
Neutralization by ADANeutralization by ADA
Adalimumab: TNF inhibition
105
Anti-adalimumab
40
5050
100
% A
U
125I adalimumab Fab
Adalimumab
85
90
95
100
105
% A
U in
hibi
ted
1 10 100 1000 100000
10
20
30
ng TNF
% A
U
Schie et al., ARD 2015
VLVH
TNF
non-human
Infliximab: potential epitopes
adapted from Liang et al. JBC 2013
infliximab
non-human
Infliximab: TNF inhibition
Anti-infliximab
40
5050
100
% A
U105
Infliximab
1 10 100 1000 100000
10
20
30
ng TNFα
% A
U
85
90
95
100
105
% A
U in
hibi
ted
Schie et al., ARD 2015
10000
100000W
EHI a
ssay
(AU)
correlation WEHI assay – anti-infliximab RIA
100 1000 10000 100000100
1000
10000
RIA (AU)
WEH
I ass
ay (A
U)
Schie et al., ARD 2015
• ADA to anti-TNF antibodies are predominantly neutralizing:
> 98% for adalimumab (21 pt)> 90% for infliximab (34 pt)> 97% for certolizumab (9 pt)
• paratope is immunodominant?• No need for bioassays assessing neutralizing capacity
Immune complexesImmune complexes
Serum analysis anti-adalimumab complexessucrose gradients
0
20
40
% Bin
ding
0
20
40
% Bin
ding
0
20
40
% Bin
ding
adalimumab, no ADA (in ABT)
adalimumab, low ADA (in ABT)
no adalimumab, high ADA (in ABT)
0 5 10 15 200
fraction0 5 10 15 20
0
fraction0 5 10 15 20
0
fraction
� Most sera contain complexes� Complexes are small
� These complexes are not rapidly cleared
0 5 10 15 200
20
40
% Bin
ding
fraction
IgM IgG
Immune complexes are formed upon infliximab administration
ADA
IFX
ADA-positive patients that receive infliximab will develop immune complexes
Various sizes of immune complexes can be formed
70 – 170 µg/mL infliximab
van der Laken et al., Ann Rheum Dis, 2007
Big Small
• Infusion reactions are allergic-like reactions upon infliximab infusion• Symptoms: flushing, chest tightness, dizziness,
bronchospasms
• 7% of patients experience an infusion reaction
Infusion reactions correlate with ADA titer
• We find little anti-IFX IgE in patient sera
• High ADA titer increases chance of infusion reactions
Pascual-Salcedo et al., Rheumatology, 2011
Neutralizing monoclonal antibodies show diversity in complex formation
data unpublished, available upon requist [email protected]
van Schie, unpublished
Immune complex size is concentration dependent
data unpublished, available upon requist [email protected]
70 – 170 µg/mL infliximab
| 36van Schie, unpublished
influence ratio infliximab/anti-infliximab
data unpublished, available upon requist [email protected]
van Schie, unpublished
Immune complex formation: spiking labeled infliximab in ADA+ sera
high ADA titers increase propensity to form large IC
data unpublished, available upon requist [email protected]
van Schie, unpublished
Very large immune complexes can be formed using high concentrations of ADA and drug
data unpublished, available upon requist [email protected]
van Schie, unpublished
Neutrophil activation depends on complex size
data unpublished, available upon requist [email protected]
van Schie, unpublished
measurement of IgE anti-infliximab
data unpublished, available upon requist [email protected]@sanquin.nl
• Small immune complexes can be detected weeks after last administration of adalimumab• Larger immune complexes can be formed in vitrofrom ADA in patients sera
Take homeAnti-drug antibodies (ADA) to therapeutic Abs:• can lead to loss of efficacy• occasionally may induce hypersensitivity
• probably immune complex mediated
• neutralization leads to non-response • Quantity of ADA is relevant: balance ADA/drug
• clinical relevance of ADA is best interpreted in conjunction with PK
Acknowledgements Sanquin ResearchKarin van SchiePauline van SchouwenburgMargreet HartSimone KruithofEls de Groot Gertjan WolbinkDiana Wouters
ReadeGertjan WolbinkMargret de KoningCharlotte KrieckaertMike NurmohamedEva Kneepkens
LUMC, LeidenErik BosRoman KoningBram Koster
AMC, AmsterdamHanke BrandseGeert D’Haens
Lucien AardenKarien Bloem
Sanquin Diagnostics ServicesDesiree van der KleijHenk de VriezeAstrid van LeeuwenSteven Stapel
GenmabRob de JongEsther van BurenTom Vink
Geert D’Haens
LACDR, LeidenStefan RomeijnWim Jiskoot
Unidad de InmunologíaHospital La Paz, SpainChamaida Plasencia RodriguezDora Pascual-Salcedo
