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22-09-08 ADA to ada Functional analysis of the anti-adalimumab response using patient-derived monoclonal antibodies Theo Rispens.

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Presentation on theme: "22-09-08 ADA to ada Functional analysis of the anti-adalimumab response using patient-derived monoclonal antibodies Theo Rispens."— Presentation transcript:

1 22-09-08 ADA to ada Functional analysis of the anti-adalimumab response using patient-derived monoclonal antibodies Theo Rispens

2 Immunogenicity therapeutic antibodies Mouse : HAMAHACAHAHA Immunogenicity : Humanized 3-5% mouse Chimeric 30% mouse Human InfliximabAdalimumab

3 Formation of antibodies to adalimumab Treatment of rheumatoid arthritis with adalimumab: effective in 60-70% of patients Free anti-drug antibodies (ADA) in 17% of patients after 6 months of treatment ADA formation leads to impaired treatment efficacy Bartelds et al 2007 www.biologicals.sanquin.nl

4 ADA issues to address using mAbs -clonality of the response -B cell epitopes -Immune complex formation -Neutralization -Standardization

5 Making human monoclonal antibodies Schouwenburg et al., ARD 2012

6 All monoclonal antibodies are derived from different precursor B-cells

7 All mAbs bind to an overlapping epitope

8 mAbs neutralize adalimumab

9 TNF-  completely inhibits binding of ADA from patient sera Schouwenburg et al., ARD 2012

10 Antibody structure VL VH Fab Fc

11 Adalimumab: possible antigenic determinants FR1-IMGT CDR1-IMGT FR2-IMGT CDR2-IMGT FR3-IMGT CDR3-IMGT FR4-IMGT (1-26) (27-38) (39-55) (56-65) (66-104) (105-117) (118-128) A B BC C C' C'C" C" D E F FG G (1-15) (16-26) (27-38) (39-46) (47-55) (56-65) (66-74) (75-84) (85-96) (97-104) (105-117) (118-128) ——————————————> ——————————> ———————> ————————> ————————> —————————> ———————————> ———————> ——————————> 1 10 15 16 23 26 27 38 3941 46 47 55 56 65 66 74 75 80 84 85 89 96 97 104 105 111112 117 118 128 |........|....| |......|..| |..........| |.|....| |.......| |........| |.......| |....|...| |...|......| |......| |.....|1|....| |.........| adaVH EVQLVESGG.GLVQP GRSLRLSCAAS GFTF....DDYA MHWVRQAP GKGLEWVSA ITWN..SGHI DYADSVE.G RFTISRDNAK NSLYLQMNSLRA EDTAVYYC AKVSYLSTASSLDY WGQGTLVTVSS IGHV3-9*01 EVQLVESGG.GLVQP GRSLRLSCAAS GFTF....DDYA MHWVRQAP GKGLEWVSG ISWN..SGSI GYADSVK.G RFTISRDNAK NSLYLQMNSLRA EDTALYYC AKD YFDY WGQGTLVTVSS (Homo sapiens) A T H D E V V SL |—————————————————————————————————————————————————V-REGION—————————————————————————————————————————————————————————||———N————||———J-REGION——| FR1-IMGT CDR1-IMGT FR2-IMGT CDR2-IMGT FR3-IMGT CDR3-IMGT FR4-IMGT (1-26) (27-38) (39-55) (56-65) (66-104) (105-117) (118-128) A B BC C C' C'C" C" D E F FG G (1-15) (16-26) (27-38) (39-46) (47-55) (56-65) (66-74) (75-84) (85-96) (97-104) (105-117) (118-128) ——————————————> ——————————> ———————> ————————> ————————> —————————> ———————————> ———————> ——————————> 1 10 15 16 23 26 27 38 3941 46 47 55 56 65 66 74 75 80 84 85 89 96 97 104 105 11112 117 118 128 |........|....| |......|..| |..........| |.|....| |.......| |........| |.......| |....|...| |...|......| |......| |.....||....| |.........| adaVL DIQMTQSPSSLSASV GDRVTITCRAS QGI......RNY LAWYQQKP GKAPKLLIY AA.......S TLQSGVP.S RFSGSG..SG TDFTLTISSLQP EDVATYYC QRYNR....APYT FGQGTKVEIK. IIGKV1-27*01 DIQMTQSPSSLSASV GDRVTITCRAS QGI......SNY LAWYQQKP GKVPKLLIY AA.......S TLQSGVP.S RFSGSG..SG TDFTLTISSLQP EDVATYYC QKYNSAP YT FGQGTKLEIK. (Homo sapiens) R A R R V |—————————————————————————————————————————————————————V-REGION——————————————————————————————————————————————————————————————||——J-REGION—|

12 inhibition of anti-ada to wild-type adalimumab by ada Fab variants -single-point mutants of adalimumab -selected for enhanced TNF binding adalimumab ADA adalimumab mutant Fab

13 inhibition of anti-ada to wild-type adalimumab by ada Fab variants. values represent binding relative to uninhibited: 100% = no inhibition = no binding by ada Fab variant = mutation affects binding adalimumab ADA adalimumab mutant Fab

14 mAbs: immune complexes with adalimumab

15 Results gradients Prot A  Most sera contain complexes  Complexes are small  These complexes are not rapidly cleared

16 ELISA and ABT vs affinity prot. A

17 Conclusions -ADA to ada is a diverse response -broad range of unrelated clones -Extensive SHM / affinity maturation -Multiple epitopes on adalimumab -response is also very restricted -All antibodies bind competitively to overlapping epitopes ->98% of antibody response neutralizes TNF-α -formation of small and larger immune complexes

18 Acknowledgements Sanquin Research Pauline van Schouwenburg Margreet Hart Simone Kruithof Els de Groot Marieke van Ham Gertjan Wolbink Diana Wouters Lucien Aarden Sanquin Diagnostics Services Desiree van der Kleij Henk de Vrieze Astrid van Leeuwen Reade Gertjan Wolbink Margret de Koning Charlotte Krieckaert Margret Bartelds Mike Nurmohamed Genmab Rob de Jong Esther van Buren Tom Vink Bayer Christian Votsmeier


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