Yeni Banff Klasifikasyonunun Getirdikleri
Banu Sis, MD, FRCPC
Dept of Laboratory Medicine and Pathology
Alberta Transplant Applied Genomics Centre
University of Alberta,
Edmonton, AB, Canada
3. Ulusal Transplantasyon Immunolojisi ve Genetigi Kongresi
23 Nisan 2011
Banff 2009 Conference on Allograft Pathology
• Banff, AB; 9-14 Agustos, 2009
• 30 ulkeden 263 katilimci (transplant clinicians, pathologists, surgeons, immunologists, researchers)
• Solid organ transplantasyonu ile ilgili cok sayida konu tartisildi:
– Alloantikor yanitlari– Endotel hucrelerinin rejeksiyondaki rolu– Genomik/proteomik yaklasimlar– Rejeksiyonun invaziv olmayan belirliyicileri– Klasifikasyonda guncellemeler
Major topic of discussion was antibody
mediated graft deterioration
• Occurs in various organs• Related mechanisms• Phenotypes• Prognostic factors
Antibody-mediated rejection: An underestimated
problem in allografts
• The hx of ABMR:– First recognized in 1960s (hyperacute rejection)– In early 1990s: Recognition of delayed acute ABMR (Halloran et al)
– Discovery of C4d staining (1991, 1993)– Recognition chronic ABMR and late graft losses due to Ab (early 2000s)
– C4d deposition with no active rejection in ABOi grafts (2007)
– C4d negative ABMR (2009 Edmonton group)
Mul
DSAKidney> 6 mo
post tx
Test once
for HLA
and MICA
ab’s
One year
later
Clinical
?Non-DSA
Heart
Lung
Liver
TX
with
function
200820072006
Multicenter study of anti-HLA antibodies on survival of kidney,
heart, lung, liver allografts in more than 4000 pts
50
60
70
80
90
100
Percent Graft Survival
0 1 2 3 4Years after Testing
Log-rank
p<0.0001
Kidney Allograft Survival
No HLA Antibody (4757)
HLA Antibody (1087)
All HLA Antibodies are De novo
92%
76%
Heart Allograft Survival
50
60
70
80
90
100
Percent Graft Survival
No HLA Antibody (287)
HLA Antibody (141)
Log-rank
p=0.0036
Years after Testing
All HLA Antibodies are De novo
0 1 2 3 4
Lung Allograft Survival
No HLA Antibody (99)
HLA Antibody (32)
Log-rank
p=0.0302
50
60
70
80
90
100
Percent Graft Survival
40
Years after Testing
All HLA Antibodies are De novo
0 1 2 3 4
50
60
70
80
90
100
Percent Graft Survival
No HLA Antibody (269)
HLA Antibody (65)
Log-rank p=NS
(0.1282)
Years after Testing
All HLA Antibodies are De novo
Liver Allograft Survival
ABMR occurs in various organs
Updates in heart and pancreas allografts
C4d Publications by Year
0
10
20
30
40
50
60
70
80
90
2008
2006
2004
2002
2000
1998
1996
1994
1992
1990
Year
Number of Publications
Kidney Heart Liver Lung Pancreas Bowel281 52 29 21 5 2
Total 390Courtesy of Dr Colvin
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!
Courtesy of Dr Colvin
Problems with ISHLT 2004 diagnostic criteria for
ABMR
• A full day session was devoted to revisit criteria for ABMR in heart allografts in partnership with the
ISHLT
• Problems with the current ISHLT criteria for ABMR:– Lack standardization of definitions and grading for C4d and histologic features
– Require “acute graft dysfunction” for diagnosis– Depend on screening by histology
Revisited criteria for acute ABMR in heart
allografts
Consensus was achieved on:
1. Recommending multiple time points to test for DSA2. Performing C4d staining (IF or IHC) on all cardiac allograft biopsies
3. Interpreting C4d staining only in myocardial capillaries
4. Scoring C4d staining as diffuse (>50%), focal (<50%) or negative
5. Only diffuse C4d is accepted as positive
Revisited ABMR criteria in pancreas
allografts
• Perform C4d staining in all pancreas allograft biopsies
• Remove requirement for “graft dysfunction” for the diagnosis of acute ABMR
Acute antibody mediated rejection
1.C4d+
2.Morphologic evidence of tissue injury (interacinar capillaritis, acinar cell
damage, arteritis, etc).
3.DSA testing
Two of 3 criteria necessary for diagnosis
C4d and graft pathology
0%
10%
20%
30%
40%
50%
60%
70%
80%
90%
100%
cg mm g ptc cv PTCBMML C4d Anti-HLA
Ab
% of biopsies with TG
C4d staining
Diffuse positive
Focal positive
Negative
Histologic lesions
and PTCBMML
Severe
Moderate
Mild
Negative
Anti-HLA antibodies
Positive
Negative
Many (50%) Transplant Glomerulopathy cases with Alloantibody
are C4d negative
70% HLA antibodies
36% C4d deposition
91% PTC basement membrane multilayering
35% glomerulitis
70% peritubular capillaritis
Sis et al. AJT 2007; 7: 1743
Gloor et al. AJT 2007; 7: 2124
Miura et al. Clin Transplant 2007; 21: 8
Shimizu et al. Clin Transplant 2009; 23: 39
Early EM findings of TG in longitudinal
analysis of protocol biopsies
cg
Nankivell AJT 2007
Pre-transplant DSA & TG
Cumulative Incidence of TXG
P=0.02
Nankivell Banff 2009
DSA & PTC & glomerular C4d deposition
N =415 biopsies
Biopsies < 1 month are for cause = 98
%%
Nankivell Banff 2009
Role of endothelium in ABMR
Training set
n=81
Validation set
n=82
Incidence of transplant glomerulopathy (*%
)
No Ab
n=30
Ab with no E
n=21
Ab with E
n=30
No Ab
n=31
Ab with no E
n=31
Ab with E
n=20
C4d+ Transplant Glomerulopathy
C4d Negative Transplant Glomerulopathy
0
10
20
30
40
50
60
0
10
20
30
40
50
60
6.7%
19%
43%
6.5%
23%
60%
C4d is negative in 60% of chronic active ABMR biopsies
(=TG with Ab+ E+)
Sis et al. AJT 2009;9:2312-23
60%
C4d negative
Patients with antibody and high ENDAT expression show poor graft survival
Sis et al. AJT 2009;9:2312-23
Training set
N=81
Validation set
N=82
Phenotyping Late Kidney Transplant Loss
What Causes Late Allograft Loss?
• 1,317 kidneys, f/u of ~50 months
• 330 graft losses (25%)
– 138 death with function (41.8%)
– 39 primary nonfunction (11.1%)
– 153 death censored graft loss (46.1%)
• Glomerular: Tg, recurrence, non-
recurrence
• IF/TA—91% with diagnosis (PVN,
Immune mediated, PN, etc)
Am Journal Transplant 2009; 9:527
.
15% death censored graft losses
98% of graft losses attributable to specific causes
Glomerular disease 36%
20% recurrent disease
Most are NOT idiopathic IFTA or CNI toxicity
b) Banff diagnosis and outcome
a) Diagnosis of failed grafts by Banff classification
Survival reflect C4d+ABMR, C4d-veABMR, and GN
Time post Bx1400120010008006004002000
Cum Survival
1.0
0.8
0.6
0.4
0.2
0.0 Other
Glomerulonephritis
T cell mediated rejection or
borderline T cell mediated rejection
C4d+ Antibody mediated rejection
C4d+ Antibody mediated rejection
T cell mediated rejection
or borderline TCMR
Interstitial fibrosis and
tubular atrophy, not
otherwise specified
Other
Glomerulonephritis
p = 0.07
}
6
7
7
61
d) Antibody-associated microcirculation injury and outcome
c) Diagnosis of failed grafts in relationship to antibody
and microcirculation changes
C4d+ Antibody mediated rejection
C4d- Antibody mediated rejection
(PRA+ with microcirculation changes)
PRA-
Glomerulonephritis
PRA+ alone
(without microcirculation changes)
PRA not available
1400120010008006004002000
1.0
0.8
0.6
0.4
0.2
0.0 Glomerulonephritis
No PRA or PRA+ without microcirculation change
C4d+ Antibody mediated rejection
C4d- Antibody mediated rejection
(PRA+ with microcirculation change)
p = 0.01Cum Survival
Time post Bx
p = 0.01
6 7
10
2
1
1
Figure 1A.
a Others (n=6): C4d deposition with no pathology (n=4); Thrombotic microangiopathy (n=1); Inadequate (n=1)b Others (n=4): C4d deposition with no pathology (n=1); T. glomerulopathy (n=1) ; Thrombotic microangiopathy (n=2); Suspicious viral (n=1) c Others (n=2): Obstruction (n=1); Suspicious viral (n=1) d Others (n=9): T. glomerulopathy (n=3) ; Suspicious viral (n=2); Post-transplant lymphoproliferative disorder (n=1); IFTANOS with acute tubular necrosis
(n=1); Acute pyelonephritis (n=1); Inadequate (n=1)e Non-adherence is defined by notes in the records around the time of the biopsy. Patients with multiple biopsies are assigned to the column reflecting their
first recorded non-adherence.
IFTANOS, interstitial fibrosis and tubular atrophy not otherwise specified
Biopsies for clinical indications
Histological diagnosis 0-6 weeks 6 weeks-6 months 6- 12 months >12 months Total
Antibody-mediated rejection 4 (5%) 1 (2%) 1 (2%) 67 (29%) 73 (18%)
Possible ABMR 1 (1%) 0 1 (2%) 12 (5%) 14 (3%)
Mixed rejection 0 1 (2%) 1 (2%) 23 (10%) 25 (6%)
T cell-mediated rejection 7 (9%) 15 (24%) 5 (12%) 8 (3%) 35 (8%)
Borderline 11 (14%) 9 (15%) 9 (22%) 10 (4%) 39 (9%)
Polyoma virus nephropathy 0 3 (5%) 7 (17%) 2 (1%) 12 (3%)
Glomerulonephritis 1 (1%) 2 (3%) 3 (7%) 36 (16%) 42 (10%)
No major abnormalities 49 (62%) 24 (39%) 9 (22%) 37 (16%) 119 (29%)
IFTANOS 0 3 (5%) 3 (7%) 26 (11%) 32 (8%)
Other 6a (8%) 4b (6%) 2c (5%) 9d (4%) 21 (5%)
Total 79 (100%) 62 (100%) 41 (100%) 230 (100%) 412 (100%)
Number of patients Total
Non-adherence e 0 1 0 25 315
Sellares et al 2011, under review
Figure 1B.
Sellares et al 2011, under review
Table 1. Histological diagnosis of the transplants that failed during follow-up period versus those that have not failed during follow
up, using the last biopsy per patient.
Current status of graft during follow-upGrafts that have not failed
after biopsyFailed grafts
Duration of follow-up post biopsy 31.4a (0 – 60.7)b 24.6a (0.03-36.9)b
Histological diagnosis n % n % p-values
Antibody-mediated rejection 37 14% 28 47% 4.66E-07
Possible ABMR 9 4% 2 3% 0.26
Mixed rejection 7 3% 6 10% 0.002
T cell-mediated rejection 17 7% 0 0% 0.04
Borderline 27 11% 1 2% 0.03
Polyoma virus nephropathy 5 2% 1 2% 0.88
Glomerulonephritis 26 10% 11 18% 0.15
No major abnormalities 92 36% 3 5% 2.04E-06
IFTANOS 23 9% 4 7% 0.56
Other 12 5% 4 7% 0.52
TOTAL 255 100% 60 100%
Patients with recorded
non-adherence7 3% 19 32% 0.0001
aMedian and brange shown in months
Sellares et al 2011, under review
Table 3. Non-adherence phenotype in failed grafts
Histologic diagnosis Antibody status at Antibody status at
(last biopsy) time of biopsy time of failure
Non-adherence
documented
before the last biopsy
C4d- ABMR DSA DSA
C4d- ABMR DSA DSA
C4d- ABMR DSA DSA
C4d+ ABMR DSA DSA
C4d+ ABMR DSA DSA
C4d+ ABMR DSA DSA
C4d+ ABMR DSA DSA
C4d+ ABMR DSA DSA
C4d+ ABMR DSA DSA
C4d- ABMR DSA DSA
C4d- ABMR DSA DSA
Possible ABMR NDSA na
Mixed rejection DSA DSA
Mixed rejection DSA DSA
Borderline DSA DSA
GN DSA DSA
Non-adherence
documented
after the last biopsy
TG PRA neg DSA
IFTANOS grade 2 PRA neg DSA
No major abnormalities NDSA na
ABMR, antibody-mediated rejection; IFTANOS, interstitial fibrosis/tubular atrophy not otherwise specified; DSA, donor specific
antibodies; NDSA, PRA positive with no identified DSA; TG, transplant glomerulopathy; GN, glomerulonephritis; na, not available
Sellares et al 2011, under review
Implementation of Banff Working Groups
• To address problematic issues in allograft pathology and Banff
Classification.
• Current Problems with the schema:
– C4d negative ABMR
– Isolated v-lesion : unknown significance (v>0 with no/minimal tubulointerstitial inflammation)
– Borderline cases
– Reproducibility issues
Isolated v-lesion
Working Group
Quality Assurance
Working Group
Glomerular Lesion
Working Group
Polyoma Virus Nephropathy
Working Group
Fibrosis Scoring
Working Group
Molecular Pathology
Working Group
data-driven & validated
refinement of
Banff criteria
Banff Working Groupsestablished in Banff 2009 Meeting
BWGs aim at addressing unmet needs within the Banff Classification to support/refute
potential changes to the classification via conducting multi-center trials.
http://cybernephrology.ualberta.ca/Banff/
Clinical significance of isolated vasculitis:
A multicenter observational study
B Sis, B Lategan, S Bagnasco, M Haas, A Magil, L Cornell, A Herzenberg, I
Gibson, P Randhawa, Y Ozluk, P Halloran, F Cosio, E Kraus
ATC 2011, oral presentation, abstract # 2203
Background
• Microarray analysis of kidney transplant biopsies with TCMR defined by Banff and transcript sets
representing:
– Cytotoxic T lymphocyte activation and infiltration– Interferon gamma-effects– Parenchymal deterioration
• Several outliers for this correlation
Intimal arteritis without significant
tubulo-interstitial inflammation
Microarray study of 143 consecutive kidney transplant biopsies identified
isolated v cases with low inflammatory gene set scores
Mueller et al.
AJT 7:2712-2722, 2007
Isolated v lesion
A biopsy from U of Manitoba
Pictures by Ian Gibson
v3, all other acute Banff scores 0, C4d 0
DSA class II+, graft function N
“isolated v” Lesion
Does this represent a
true acute rejection?
Study question
Study aims
• To determine the clinical significance of isolated v lesion in kidney transplant biopsies
– In terms of graft function –Graft survival
• To determine whether we need to refine Banff criteria to call TCMR based on vasculitis alone
(Banff II/III)
Isolated v-lesion BWG
• To address the clinical significance of isolated v- lesions (with little or no
tubulo-interstitial inflammation) in renal allograft biopsies.
• To determine whether we need to refine Banff criteria to call TCMR Banff
II/III
• N=300 biopsies
• GROUP 1= Isolated v lesion
– v>0 and t<2 and i <2 and C4d-
• GROUP 2= Vasculitis + IA/IB
– v>0 and i>1 and t>1 and C4d-
• GROUP 3= No vasculitis
– v0 and t<2 and i<2 and C4d-
0
5
10
15
20
25
30
35
40
45
Clinical data at biopsy
Group 1 Group 2 Group 3 P-value P-value
Isolated v1 v1 and tubulointerstitial
rejection
v0 and minor
tubulointerstitial
inflammation
Group
1 vs. 2
Group
1 vs. 3
N 117 48 45
Indication for biopsy
Rising creatinine 41 14 12 NS NS
DGF 27 0 0 0.000274 0.000416
Clinical, other 41 33 25 NS 0.017356
Protocol 8 1 8 NS 0.046
Serum Creat at biopsy 4.2 ± 2.9 3.0 ± 1.9 4.4 ± 3.1 0.015 NS
Time post transplant at
biopsy, median
17 33 12 NS NS
Time at biopsy, <6 mo 91 39 39 NS NS
Future function
Group 1 Group 2 Group 3 P-value P-value
Isolated v1 v1 and
tubulointerstitial
rejection
v0 and minor
tubulointerstitial
inflammation
Group
1 vs. 2
Group
1 vs. 3
N
sCreat at 1 mo 2.2 ± 1.6 2.1 ± 0.9 2.3 ± 1.4 NS NS
sCreat at 6 mo 2.0 ± 1.5 1.9 ± 0.8 2.3 ± 1.8 NS NS
Patient, deceased 17/100 pts
(8 unknown)
5/43 pts
(3 unknown)0/45 pts
Death censored
graft failure
20/104 pts
(4 unknown)6/46 pts 7/45 pts
Graft survival
Isolated v1 (v1 and i<1 and t<2)
v1 and i>1 and t>1
v0 and i<2 and t<2
p=0.289
conclusions
• Isolated v1 biopsies are seen early and associated with a high incidence of delayed graft function;
• v1 with or without high tubulointerstitial inflammation is not related to increased graft failure compared to v0.
• Thus, isolated v1 lesions, after the exclusion of antibody-mediated rejection, are of two types: T cell-mediated rejection
and injury, and have no independent prognostic significance.
Antibody-mediated vasculitis
Banff criteria need to be revisited
• Current Banff – only v3 in the presence of DSA and C4d
• However, alloantibody may mediate milder forms of vasculitis intimal arteritis (Edmonton and Paris data)
• Presence of v1/v2 should should not be interpreted as supporting the diagnosis of TCMR unless other features are
present
• DSA testing crucial (C4d is often negative).
Milder forms of intimal arteritis is often associated with
DSA
0
10
20
30
40
50
60
70
80
90
100
v 0 v>0
56%44%
DSA (-)
DSA (+)
0
10
20
30
40
50
60
70
80
90
100
v 0 v>0
C4d negative
C4d focal or diffuse
33%67%
% of biopsies
n=172 n=27 n=189 n=27
p=0.017 p=0.023
Sis et al AJT 2010; 10:421-30
PVN staging BWG
• Volker Nickeleit proposed a classification of PVN includes 3 stages, approved by consensus
• stage-A (early changes, without tubular epithelial
cell necrosis);
• stage-B (active nephropathy with virally induced tubular necrosis);
• stage C (late sclerosing changes)
• Pending for reproducibility studies
Molecular Pathology BWG
• Led by Phil Halloran will facilitate the consensus about how and which molecular markers can be integrated into the Banff
classification.
• RT-PCR or chip (with few genes) based molecular measurements may be incorporated into the existing
histology-based Banff classification
• Discovery of new diagnostic and/or prognostic tissue markers could be feasible with the help of omics technologies.
Other BWGs
• Fibrosis scoring BWG:
• Aim: To standardize fibrosis scoring in renal native and allograft biopsies,
and conduct multi-center reproducibility trials to improve interobserver
agreement.
• Glomerular lesion scoring BWG:
• Aim: To re-examine scoring glomerular double contours (cg), glomerulitis
(g), and mesangial matrix increase (mm) aiming at refinement of these
criteria to increase the interobserver reproducibility.
• Quality assurance BWG:
• Aim: To plan Banff training courses, proficiency tests, and
immunohistochemistry (C4d and BK) multicentre staining trials.
•
Glomerular lesion scoring trial
– in progress
• 23 pathologists participated from 10 countries
• Online virtual microscopy assessment of biopsies– reproducibility studies
– Correlation with clinical outcomes
• Management of sensitized patients• A new concept of C4d negative ABMR• Reports from Banff Working Groups –Significance of isolated vasculitis (Sis et al)–Glomerular Lesion Scoring (Sis et al)–Fibrosis scoring (Colvin et al)–Polyoma virus nephropathy staging (Nickeleit et al)–Quality assurance in transplant pathology (Mengel et al)
• Role of epithelium in allograft deterioration• Molecular Pathology in Transplantation• New insights from Protocol Biopsies
2011 Banff meeting, June 6-10
Paris, France
THANK YOU…
See you in the next Banff Conference in Paris (6-10 June 2011)
http://cybernephrology.ualberta.ca/banff/2011/
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