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/