Transcript MICA - HLA Matchmaker

Antibodies
Against MICA
Epitopes
HLA Epitopes in Clinical Transplantation
Pittsburgh, PA
June 15-16, 2011
Justin W. Mostecki, Ph.D.
MICA
Duquesnoy RJ et al. Hum Immunol 69(12):826, 2008
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Major histocompatibility complex (MHC) class I chain-related protein A
MICA is a ligand for NKG2D
33 polymorphic amino acid positions in the extracellular domain (subset shown above)
MICA antibodies are associated with graft failure
MICA Epitopes
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El-Awar N, Terasaki P, Cai J, Deng, C, Ozawa M, Nguyen A: Epitopes of the HLA-A,
B, C, DR ,DQ and MICA Antigens. Clin Transpl. 175, 2007
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Duquesnoy RJ, Mostecki J, Hariharan J, Balazs I: Structurally based epitope analysis
of major histocompatibility complex class I-related chain A (MICA) antibody specificity
patterns. Hum Immunol 69(12):826, 2008
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Zou Y, Qin Z, Silveus A, Fan Y, Stastny P: Polymorphisms of MICA recognized by
human alloantibodies. Immunogenetics 61(2):91, 2009
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Suarez-Alvarez B, Alonso-Arias R, Bravo-Mendoza C, Lopez-Vazquez A, Ortega T,
Baltar JM, Coto E, Ortega F, Lopez-Larrea C: Identification of epitopes and
immunodominant regions on the MICA protein defined by alloantibodies from kidney
transplant patients. Transplantation 88(3 Suppl):S68, 2009.
Alignment of Polymorphic Extracellular Positions
of LSA-MIC Antigens
Eplet Positions
Eplet
Position
6
14
24
25
26
36
91
114
122
123
125
129
151
156
173
174
175
181
206
208
213
214
215
221
251
271
Surface
Domain
α1
α1
α1
α1
α1
α1
α2
α2
α2
α2
α2
α2
α2
α2
α2
α2
α2
α2
α3
α3
α3
α3
α3
α3
α3
α3
Polymorphic
Visibility
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Positions
6
14
24
24 36
26
36
91
114
122
122 125
125
129
151
156
173
173 175
175
181
206 210
208
213
213 215
215
221
251
271
Duquesnoy RJ et al. Hum Immunol 69(12):826, 2008
38 Eplets
213T
251Q
129M
36C
206GW
215S
174EG
213I
251R
25AC
25AY
129V
173E
14G
175G
175S
214TT
122V
14W
123LE
181R
24A
25TC
6P
271A
151V
114R
156L
156H
221L
125K
91R
125E
208C
251E
156R
208Y
26V
12 Monomorphic at MICB
6R
26G
91Q
114G
122L
151M
173K
181T
206SR
215T
221V
271P
Not covered with LSA-MIC panel
Polymorphisms Shared By Groups of MICA Antigens
El-Awar N et al. Clin Transpl. 175, 2007:
• Antibody eluted from MICA*004 cell line reacted with MICA*004, *009, *027 (defined by
either 36Y, 129V or 173E)
Duquesnoy RJ et al. Hum Immunol 69(12):826, 2008:
a(1-Ep
freq)
x Epfreqmix
Epfreqmix represents eplet
frequency in 2/3 EA &1/3 AA
population mixture
Monomorphic with MICB
and predicted not to illicit
MICA-specific alloantibodies
• Demonstrated CMGWS and AYVE patterns of reactivity
Zou Y et al. Immunogenetics 61(2):91, 2009:
• MICA-G1: 36C, 173K, 206G, 210W and 215S (MICA*001, *002, *007, *012, *017, *018)
• MICA-G2: 206S, 210R and 215T (MICA*004, *006, *008, *009, *019)
• MICA*002 & MICA*008 hybrid recombinants, MICA*008 V129M & MICA*002 K173E
site-directed mutants to map epitopes
AYVE
CMGWS
Supereplets CMGWS and AYVE
AYVE
CMGWS
Do MICA Positive Sera Demonstrate Patterns
Consistent With Monomorphic Eplets?
MFI
MFI
Sample 1: MICA*005-Eluted Antibody Demonstrates Reactivity
Pattern Consistent with 173K Monomorphic Eplet
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MICA*008,*008
typing
MICA*005
Absorption &
elution
Sample reacts with CMGWS antigens, but not AYVE antigens
Sample reactivity to MICA*036 consistent with 36C eplet pattern as determined by
MICA*036 absorption & elution (data not shown)
AYVE
CMKGWS
Sample 1: Antibody Demonstrates 173K Pattern
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CMGWS supereplet re-classified as CMKGWS to include 173K eplet
Sample reactivity to MICA*036 consistent with 36C eplet pattern as determined by
MICA*036 absorption & elution (data not shown)
Sample 2: Shows Pattern Consistent with
206SR/215T Monomorphic Eplets
AYVESRT
CMKGWS
Sample 2: Antibody Demonstrates
206SR/215T Pattern of Reactivity
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No MICA antigens to resolve 206SR and 215T eplets
Fewer antigens with this Beta lot
AYVESRT
CMKGWS
Updated CMKGWS and AYVESRT
Sample 3: AYVESRT
• MICA-reactive sera often contain mixtures of antibody that demonstrate
different patterns of reactivity
Sample 3: No MICA*036 & *037 Reactivity
With MICA*051-eluted Antibody
MICA*051
absorption &
elution
Sample 3: MICA*051-eluted Antibody
Demonstrates 25AY Reactivity Pattern
AYVESRT
CMKGWS
Sample 3: MICA*051-eluted Antibody Shows
Pattern Consistent With 25AY Eplet
Sample 4: AYVESRT With Additional Reactivity
• Positive antigens shown in red contain 125E
• MICA*001 shown in green is 125K
• 25AY-containing antigens ranked at top with manual
eplet analysis
• Next steps:
- Absorption & elution performed with MICA*007
- Absorption & elution performed on MICA*007cleared serum with MICA*036 and MICA*051
Sample 4: MICA*051-eluted Antibody From MICA*007Cleared Serum Demonstrates 25AY Reactivity Pattern
MICA*036 absorption & elution
MICA*051 absorption & elution
• MICA*007 absorbed and eluted antibody shows 125E pattern (data not shown)
AYVESRT
CMKGWS
Sample 4: MICA*051-eluted Antibody From MICA*007Cleared Serum Demonstrates 25AY Reactivity Pattern
Conclusions
• MICA-eluted alloantibody demonstrates 173K eplet pattern to
update supereplet to CMKGWS (similar to MICA-G1, but with 129M)
• Identified samples that demonstrate 206SR/215T pattern to update
supereplet to AYVESRT (similar to MICA-G2, but with 25AY, 129V
and 173E)
• CMKGWS and AYVESRT often contain mixtures of antibody that
demonstrate patterns consistent with a single eplet following
absorption and elution
• MICA*005, *028, *036, *037, *042 and *051 are effective at
resolving CMKGWS and AYVESRT patterns when samples react
with a single eplet
Acknowledgements
University of Pittsburg
University of Texas SW Medical Center
Rene Duquesnoy, Ph.D.
Marilyn Marrari
Yizhou Zou, M.D.
Peter Stastny, M.D.
Gen-Probe Transplant Diagnostics, Inc.
Ivan Balazs, Ph.D.
Christine Heylen, Ph.D.
Jayasree Hariharan
Lioudmila Zaslavskaia
Robert Vorhaben
Clínic Barcelona
Guadalupe, Ercilla, Ph.D.