Early Observations in Patients Antibody Mediated Rejection...
Transcript of Early Observations in Patients Antibody Mediated Rejection...
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Antibody Mediated Rejection: Historical perspectives
and animal models
Robert B. Colvin, M.D.Massachusetts General Hospital
Harvard Medical School
Banff X
August 10, 2009
Banff, Alberta
Early Observations in Patients
1966 Hyperacute rejectionKissmeyer-Nielsen F, Olsen S, Petersen VP, and Fjeldborg O, Hyperacute rejection of kidney allografts, associated with pre-existing humoral antibodies against donor cells. Lancet, 2: 662-5,1966.
1967 Transplant glomerulopathyPorter KA, Dossetor JB, Marchioro TL, Peart WS, Rendall JST, and Terasaki PI, Human renal transplants. I. Glomerular changes. Lab Invest, 16: 153-81,1967.
1968 HLA antibodies post-transplantationMorris PJ, Williams GM, Hume DM, Mickey MR, and Terasaki PI, Serotyping for homotransplantation. XII. Occurrence of cytotoxic antibodies following kidney transplantation in man. Transplantation, 6: 392-9,1968.
Humoral Antibodies in Renal Allotransplantation in ManJeannet, Pinn, Flax, Winn, Russell, NEJM 282:111,1970
Chronic allograft arteriopathy
only in patients with de novo anti-donor antibodies
Dark Ages for Antibody1970-1990
No connection between antibodies and pathology
Antibodies regarded as epiphenomenon
T cells reigned
T Rex
Sunrise in Alberta 1990Antibodies affect outcome of acute rejection
and pathologic pattern
1990Halloran PF, Wadgymar A, Ritchie S, Falk J, Solez K, and Srinivasa NS, The significance of the anti-class I antibody response. I. Clinical and pathologic features of anti-class I-mediated rejection. Transplantation, 49: 85-91,1990.
1992Halloran PF, Schlaut J, Solez K, and Srinivasa NS, The significance of the anti-class I antibody response. II. Clinical and pathologic features of renal transplants with anti-class I-like antibody. Transplantation, 53: 550-5,1992.
1996Trpkov K, Campbell P, Pazderka F, Cockfield S, Solez K, and Halloran PF, Pathologic features of acute renal allograft rejection associated with donor-specific antibody. Analysis using the Banff grading schema. Transplantation, 61: 1586-92,1996.
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Sunrises can be foggy in Canada
C4d Deposition detected in PTC in Rejection
Normal Acute Rejection
Cryostat sections, mAb to C4d, immunoperoxidase detectionFeucht et al 1991
“On the basis of covalent binding and amplification, it appears that staining of complement fragment C4d seems to be a practical tool for the demonstration of in situhumoral immune reactions that are not easily detectable otherwise.”
Helmut Feucht et al, 1991
Connect the dots
DSAC4dPTC
Graft Pathology
DSA = Donor Specific Alloantibody (HLA)
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DSA correlated with C4d+ and polys in caps
C4d and Transplantation Publications by Year
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C4d, Clin --Feucht
Class I Ab, Clin, Path--Halloran
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1990 1991 1992 1993 1994 1995 1996 1997 1998 1999 2000 2001 2002 2003 2004 2005 2006 2007 2008
AHRCollins
CHR Mauiyyedi
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“Chronic humoral rejection” = Transplant glomerulopathy or arteriopathy
+ C4d + DSA
38 pts 61% C4d+ 88% of C4d+ had DSA+
Documented:Transplant glomerulopathy, glomerulitisPTC capillaritis and BM multilamination
Introduced concept of evolution C4d+TG
C4d
Other key observations
C4d Deposition is transient (days)Indicates current antibody activityNickeleit V, Zeiler M, Gudat F, Thiel G, and Mihatsch MJ.
J Am Soc Nephrol, 13: 242-51,2002.
Minami K, Murata K, Lee CY, Fox-Talbot K, Wasowska BA, Pescovitz MD, and Baldwin WM, 3rd, Am J Transplant, 6: 923-32,2006 (rats).
ABOi grafts can have C4d without pathologyFidler ME, Gloor JM, Lager DJ, Larson TS, Griffin MD, Textor SC, Schwab TR, Prieto M, Nyberg SL, Ishitani MB, Grande JP, Kay PA, and Stegall MD, Am J Transplant, 4: 101-7,2004.
Haas M, Rahman MH, Racusen LC, Kraus ES, Bagnasco SM, Segev DL, Simpkins CE, Warren DS, King KE, Zachary AA, and Montgomery RA, Am J Transplant, 6: 1829-40,2006.
Setoguchi K, Ishida H, Shimmura H, Shimizu T, Shirakawa H, Omoto K, Toki D, Iida S, Setoguchi S, Tokumoto T, Horita S, Nakayama H, Yamaguchi Y, and Tanabe K, Am J Transplant, 8: 86-94,2008.
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Sunrise Banff 2001
2005 Chronic Ab Mediated Rejection
Solez K et al, Am J Transplant, 7: 518-26,2007.
2001 Acute Ab Mediated RejectionRacusen et al Am J Transplant, 3: 708-14,2003.
New Banff Categories
Solez K et al, Am J Transplant, 8: 753-60,2008.
2007 C4d deposition without active rejection
Technical Advances
Polyclonal anti-C4d for FFPE tissueUseful for evaluation of glomeruliRegele H, Exner M, Watschinger B, Wenter C, Wahrmann M, Osterreicher C, Saemann MD, Mersich N, Horl WH, Zlabinger GJ, and Bohmig GA. NDT, 16: 2058-66, 2001
Solid Phase Assays for HLA DSADonor cells not needed, sensitive, specificELISAKao KJ, Scornik JC, Small JC. Transplantation 1993; 55: 192–6.
LuminexFulton RJ, McDade RL, Smith PL, Kienker LJ, and Kettman JR.Clin Chem, 43: 1749-56,1997.
Terasaki et al AJT 7:408, 2007
Many studies by Terasaki, Worthington and others showed
de novo HLA antibodies a risk factor for graft loss
C4d has provided a missing link: antibody pathology
A t]
Incubate in fresh normal serumAlloAb
C4d
AntiC4d
C1q
AlloAb
HLA Class I HLA Class II
Complement fixing alloantibodies detected on Ag coated beads (FlowPRA, Luminex)
Wahrmann (Vienna) AJT 7:1033, 2007
FITC
More sensitive than cellular methods ~4x more presens detected
IgG-
IgG+C4d-
IgG+C4d+
C4d fixation by pretransplant sera on Class I beads correlates with outcome
N=338
Wahrmann (Vienna) AJT 7:1033, 2007
Strong correlation with C4d in bx
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Intragraft events Immunoglobulin Gene Transcripts (IGT)vs. time post-transplant
IGT Score (log2)
Einecke (Edmonton) AJT 8:1434, 2008
intragraft synthesis of Ig is a feature of late graft dysfunction
Local Production of DSA by cells in renal allografts
Thaunat et al (Lyon), Transplant 85:1648, 2008
IgG DSALuminex
Perplexing Heterogeneity
Transplant Glomerulopathy (TG)
Antibody (DSA) 70% HLA Class II>I Ab(91% vs 61%)
C4d in PTC 32%PTC multilamination 91%
N=41 pts (3.1%), mean dx 5.5 yrs p-tx
Sis et al (Edmonton) AJT 7:1743, 2007
38% TG + DSA without C4dNon C fix Ab?
26% TG without C4d or DSANon Ab cause (T cell, TMA/CNIT)Residue of past Ab/C
Subsets of CHR revealed bycluster analysis
Ban
ff c
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anff
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TG, Class II DSA+
TG, Class II DSA-
Interstitial Inflammation
Interstitial Fibrosis
All Banff variables, DSA
Brad Farris et al (Boston) Banff 2009 Poster
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Chronic disease evolves over time
Sequential events in CHR
C4d deposition predicts later transplant glomerulopathy
C4d TGBx <12 mo Bx > 12 mo
Neg 6%
Pos 46%
Regele et al (Vienna), JASN 13:2371,2002 (213 bx >12 m)
risk of TG after episode of AHR
Prevalence of TG in 1 yr protocol bxConventional 198 8%Presens HLAi 37 22% p<.015
ABOi 24 13%
• Prior AHR risk factor for TG (OR 17.5, multivariate)• Also for arteriopathy and interstitial fibrosis• Presensitized patients and possibly ABOi increased
risk
Gloor et al (Mayo) AJT 6:1841, 2006
Pathologic sequences observed in repeat biopsies
Collins et al, Mod Pathol, 2009
Postulated Stages of Humoral Rejection
TransplantTime (not to scale)
Blood: de novo antibodies
Graft: C4d
Graft: pathology
Clinical: graft dysfunction
I II III IV
Graft loss
Accommodation RejectionSubclinical ClinicalNo C4d With C4d
Colvin and Smith, Nature Rev Immunol 5:807, 2005
Other Organs
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C4d Publications by Year
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Kidney Heart Liver Lung Pancreas Bowel
281 52 29 21 5 2
Total 390
Kidney Heart Pancreas Lung Liver Bowel
Hyperacute rejection + + + + +
Acute humoral rejection + + +
Chronic humoral rejection +
Accommodation + +
Accepted Organ Specific Criteria for Antibody Mediated Graft Rejection
Lots of blanks!
Cardiac Allograft Vasculopathy (CAV)
C4d+ bx intimaat one year IVUS
Poelzl et al (Innsbruck) Transplant Int 18:2303, 2005
Arteriopathy + C4d + DSA= CHR of heart?
CAV correlates with prior C4d+
80% with C4d CAV27% without C4d CAV
p<0.001
Rodriguez et al (Cleveland) AJT 5:2778, 2005
Even the liver can have AMR
Watson Kozlowski Nickeleit… (Chapel Hill NC) AJT 6:3022, 2006
HLA presensitized patient (HLA-B7): day zero biopsyDeveloped chronic damage (bridging fibrosis, ductular proliferation)Ischemia reperfusion control = neg
C4d
Bellamy (Edinburgh) Histopathol 50:739, 2007
C4d Rosetta Stone
Liver Liver Kidney
Combined kidney and liver transplant from same donor into presensitized recipientBoth rejected on day 18 and both C4d+
Bellamy (Edinburgh) Histopathol 50:739, 2007
Liver is less susceptible to CHR than KidneySimultaneous liver-kidney tx– DSA+
C4d - LiverC4d - Kidney
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Endothelium: Target and Defender Anti-HLA antibodies trigger endothelial responses
Endothelial cell
Colvin, Smith, Nat Immunol Rev 2005
Apoptosis
C4d
Antibody mediated effects on endothelium
MacNK FcR
Phosphorylated signaling proteins In vivo markers of endothelial activation
Normal Rejection
What role do cells play?
Lee CY, Lotfi-Emran S, Erdinc M, Murata K, Velidedeoglu E, Fox-Talbot K, Liu J, Garyu J, Baldwin WM, 3rd, and Wasowska BA, Transplantation, 84: 1324-34,2007.
Endothelial cells in vitro
DSA alone DSA + Macs
MCP-1, KC, IL-6, Rantes, TIMP-1MCP-1, KC
FcRIII KO macrophagesless active, as were F(ab)2 DSA
Macrophages enhance cytokine productionby endothelial cells triggered by anti-MHC I Ab
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Endothelial cells resist effects of antibody/complement
• bcl-xL in renal allografts from patients with donor specific HLA antibody
Salama (Hammersmith) Am J Transplant 1:260, 2001
• muc-1 gene expression/glom ABOiPark et al (Mayo) AJT 3:952, 2003
• CD55 (DAF) in stable vs unstable cardiac graftsGonzalez-Stawinski et al (Cleveland) JHLT 27:357, 2008
C4d+DAF+No dysfunction
C4d+DAF-Acute dysfunction
Gonzalez-Stawinski et al (Cleveland) JHLT 27:357, 2008
Capillary DAF+
identifies C4d+ biopsies with no graft dysfunction
DAF = decay accelerating factor, CD55
Endothelial Gene Expressionin kidneys with rejection and DSA
Transcripts include vWF, caveolin-1,
E-selectin, CD31, CD34…
• Distinct pattern vs DSA- rejection
• Highest levels assoc with C4d+
• Significantly in C4d- DSA+ cases
Sis et al (Edmonton) AJT in press
• Low sensitivity of C4d stain vsnon-C’ fixing effects of DSA
Experimental Studies
Animal Models of AHR/CHR
Xenograft rejectionPig to monkey/baboonRat heart to gal KO mouse
AHRRat (C6) (Baldwin)Mouse heart (Wasowska)
CHRMouse heart allograftMouse with human arteryMonkey renal allograft
AHR depends on complement fixation
Rahimi S, Qian Z, Layton J, Fox-Talbot K, Baldwin WM, 3rd, and Wasowska BA, Non-complement- and complement-activating antibodies synergize to cause rejection of cardiac allografts. Am J Transplant, 4: 326-34,2004.
B10.A hearts B6 Ig KO
Complement fixing IgG2b DSA triggered immediate rejection at 10 days, but non-C’ fix IgG1 did not.
Mouse heart allografts
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AJT 7:57, 2007Control
B10.BR heart B6/RAG1-/-anti-H-2Kk
Chronic Arteriopathy can be caused by DSA Passive Mab to donor class I B10.BR heart B6.RAG1-/- given anti-H-2Kk x28 d
C4d
CD3F4/80(Mac)
Uehara et al AJT 27:5,2007
Non-complement fixing antibody (IgG1) can initiatechronic allograft arteriopathy
Anti-H-2Kk
IgG1(n=8)
No Rx(n=5)
Anti-H-2Kk
IgG2a (Allograft)
(n=10)
Anti-H-2Kk
IgG2a (Isograft)
(n=4)
80
60
40
20
0
100
24.8
5.7
52.3
5.5
Neo
intim
alIn
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p<0.01
p<0.005
p<0.005 p<0.05
Hirohashi et al, submittedMab RAG1-/-B6 recip with H-2Kk heart grafts
Anti-H-2Kk mAb B10BR hearts in B6IgG1 B6.RAG1-/- IgG2a B6.C3-/-RAG1-/-
Control
Hirohashi, ATC 2008
Chronic Allograft Arteriopathyin C3 deficient RAG1-/- recipients
Mab B6.C3-/-RAG1-/- recip with H-2Kk heart grafts Hirohashi et al submitted
DSA-IgG2a No Ab
Alloantibody can cause one feature of chronic humoral rejection without complement (NK/FcR?)
May be relevant to cases with DSA and chronic rejection without C4d
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Human arterial segment transplanted into immunodeficient mouse (SCID/Beige)
Lorber, Tellides, Pober (Yale)Transplant 67:897,1999
Thomsen (Toulouse)-mesenteric
JHLT 25:675, 2006
Galvini (Toulouse)
in press AJT
Anti-HLA class I arteriopathy in 6 wk
Non-human Primate Renal Allografts
Knechtle
Larsen, Kirk
Thomas
Jonker
Wieczorek, Mihatsch, Nickeleit
Cosimi, Kawai
Variants of mixed chimerism protocolAll off immunosuppression after day 30
48% developed anti-donor alloantibodies29% C4d+ 22% TG+
143 Recipients269 biopsies (protocol/indication)
5 nephrectomies143 autopsies
CHR in Cynomolgus Renal AllograftC4d+, DSA+
4498 d371
Smith et al, AJT 6:1790, 2006
Similarity to human CHR
PTC BM multilamination
GBM duplication
Smith et al, (Boston) AJT 8:1662, 2008
Renal allograft loss due to rejection- N=143Non-human primates off immunosuppression
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Sequential Development of CHR-Monkeys
No CHR Stage I Stage II Stage III Stage IV
Days post-transplant
106 182 225 352 371
Smith et al (Boston) AJT 8:1662, 2008M4498
High Noon for AMR
Future Questions
Basic
Regulation of B cell function
Endothelial mechanisms
Mechanism of accommodation
Role of complement and FcR/cells
Clinical
Natural history
Monitoring Ab, intervention
New therapies (Plasma cell, B cell, C’)
Late Graft Failure
AbominableAntibodyT Rex
(this guy is bigger)
Transplant Research MGH
Terry Strom
Mo Sayegh
Harvard Collaborators
CTOT-C
Genomics of Tx
ITN
CTOT
Multicenter Trials
Catherine Adams
Bernard Collins
Nicole Brousiades
Tricia Delle Pella
Diana Taheri
Brad Farris
Sandro Alessandrini
R. Neal Smith
Colvin Lab/Pathology
Donna Fitzpatrick
Susan Saidman
Histocompatibility
Fred Preffer
Megan Sykes
Jay Fishman
Frank Delmonico
Tatsuo Kawai
Win WIlliams
Elliot Heher
Nelson Goes
Nina Tolkoff-Rubin
Joren Madsen
Jim Markman
Ben Cosimi
Clinical
Masa Miyayama
Tommy Hirohashi
Joren Madsen
Cuffy Chase
Paul Russell
Mouse
Kazu Yamada
Akira Shimizu
David Sachs
Pig
Joren Madsen
Gilles Benichou
Svetlan Boskovic
David Sachs
Tatsuo Kawai
Ben Cosimi
Monkey