Antibody-Mediated Rejection in Renal transplantation

1
Antibody-Mediated Rejection in Renal
transplantation
2
Homo transplants differ not only in the strength
of rejection, but also in the nature and location
of the phenomena induced by rejection. This
conclusion must not be regarded as a cause for
discouragement. On the contrary, it should spur
further research. Jean Hamburger
3
Antibody-mediated rejection

• Alloantibodies are now appreciated as important
  mediators of acute and chronic rejection
• Alloantibodies to HLA class I or II and other
  antigens expressed by endothelium cause a variety
  of effects on renal transplants, ranging from
  acute to chronic rejection, and even apparent
  graft acceptance (accommodation).
• It differs in pathogenesis, or nature, from T
  cellmediated rejection.

4
Antibody-mediated rejection

• Alloantibodies preferentially attack a different
  location, namely
• the peritubular and glomerular capillaries, in
  contrast to T cells, which characteristically
  infiltrate tubules and arterial endothelium.
• Antibody-mediated rejection generally has a worse
  prognosis and requires a different form of
  therapy than the usual T cellmediated acute
  rejection

5
Antibody-mediated rejection and C4d

• C4d is a fragment of C4b, an activation product
  of the classic complement pathway
• C4d remains covalently bound in the tissue for
  several days after complement activation
• C4d deposition is strongly associated with
  circulating antibody to donor HLA class I or
  class II antigens
• C4d is currently the best single marker of
  complement-fixing circulating antibodies to the
  endothelium.

Feucht et al Kidney Int 1993 4313331338.
Mauiyyedi S et al. J Am Soc Nephrol 2002 13
779787. Bohmig GA et al. J Am Soc Nephrol 2002
13 10911099
6
Complement activation pathways. The classic
pathway is relevant to antibody-mediated rejection
7
Antibody-mediated rejection

• Four forms of antibody-mediated graft injury have
  been defined
• Hyperacute rejection
• Acute humoral rejection
• Chronic humoral rejection
• Accommodation

8

• Stages I to II represent accommodation, stage III
  represents subclinical humoral rejection, and
  stage IV represents CHR.
• The dashed lines for antibody and C4d deposition
  are meant to reflect the possibility of
  intermittent positivity over time.

• Inevitability of progression is not meant to be
  implied by the term stages. The rate of
  progression is likely to be variable, and the
  early stages, I and II, are reversible. At any
  stage, the antibody/ C4d may become negative, in
  which case the process is inactive.

Colvin RB. J Am Soc Nephrol 18 10461056, 2007
9
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10
Definition
Hyperacute rejection

• Hyperacute rejection arises within minutes or
  hours in presensitized patients who have
  circulating HLA, ABO, or other alloantibody-to-don
  or endothelial surface antigens

11
Hyperacute Rejection

• The pathology of hyperacute rejection overlaps
  completely with AHR.
• Deposition of C4d occurs in PTC and glomeruli,
  just as in AHR
• Early biopsies may be negative for C4d,
  presumably because of lack of access of C4 to the
  site (vasoconstriction) or enough time for
  sufficient amount to be deposited.

12
Hyperacute rejection

• Mediated by Rapid thrombotic occlusion of the
  vasculature of the transplanted allograft
• Occurs within minutes to hours after host blood
  vessels are anastomosed to donor vessels
• Mediated predominantly by IgG antibodies directed
  toward foreign protein molecules, such as MHC
  molecules
• Result from prior exposure to alloantigens from
  blood transfusions, pregnancy, or previous
  transplantation

13
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Acute humoral rejection

• Definition
• Rapid loss of graft function (days), any time
  after transplantation

• An acute loss of graft function that often arises
  in the first few weeks after transplantation
• Cannot be distinguished from cell-mediated
  rejection on clinical grounds

15
Acute humoral rejection

• Antibody-mediated rejection frequently
  undiagnosed
• Lack of typical morphologic characteristics
• C4d immunohistochemistry
• C4 is an element of the complement cascade
• Its degradation product (C4d) adheres to
  endothelial cells
• Can be detected by pathologist

16
Acute humoral rejection

• Classification of rejection as either cellular
  or humoral is flawed
• Significant overlap exists

1 Montgomery R, Zachary AA. Transplantation
200070887894, 2 Becker YT et al , Am J
Transplant 20044996-1001
17
Diagnostic criteria for acute antibody-mediated
rejection (AHR)

• Morphologic evidence of acute tissue injury acute
  tubular injury
• neutrophils and/or mononuclear cells in PTC
  and/or glomeruli and/or capillary thrombosis
  fibrinoid necrosis/intramural or transmural
  inflammation in arteries
• Immunopathologic evidence for antibody action C4d
  and/or (rarely) immunoglobulin in PTC Ig and
  complement in arterial fibrinoid necrosis
• Serologic evidence of circulating antibodies to
  donor HLA or other anti-donor endothelial antigen

Cases that meet only two of the three numbered
criteria are considered suspicious for AHR. Acute
cellular rejection may also be present.
Colvin RB. J Am Soc Nephrol 18 10461056, 2007
18
Patterns of Rejection in ABO Incompatible
Transplants
AMR
Cellular
Accommodation
19
Therapeutic Options For The Treatment Of AMR
Antibody Reduction
Immunomodulation

• Plasmapheresis/IA
• IVIg

• IVIg
• ATG
• IL-2R blockers
• Fk 506, Rapamycin
• MMF/DSG
• CAMPATH?

B-cell Modulation

• Splenectomy
• Anti-CD20
• Cytoxan

20
Antibody Reduction Therapy

• High dose IVIG (1-2 gms/kg)
• Mechanism
• Anti-idiotypic networks probably important
• Many putative immunomodulatory pathways
  identified
• Advantages
• In vitro test for predicting efficacy
• Ease of administration?
• Disadvantages
• Non-responders
• Different techniques required to follow DSA
  titers
• Less rapid Ab removal, unproven for high-titer
  DSA
• Toxicity batch-to-batch variability
• Unproven for ABOi Tx

21
Antibody Reduction Therapy

• Plasmapheresis/Low Dose IVIg (100 mg/kg)
• Mechanism
• Rapid reduction in anti-HLA or isoagglutinin Ab
• Induces donor specific unresponsiveness (HLA) or
  accommodation (ABOI)
• Advantages
• Predictable kinetics of plasmapheresis
• No evidence of nonresponders
• Able to easily follow DSA levels during/after
  therapy
• Disadvantages
• DSA may rebound between treatments or if
  discontinued
• Treatment may be prolonged and immunosuppressive
• Expensive and resource intensive

22
Rituximab

• Monoclonal anti-CD20 antibody (found on B cells)
• May improve outcomes in antibodymediated acute
  rejection episodes

1 Montgomery R, Zachary AA. Transplantation
200070887894, 2 Becker YT et al , Am J
Transplant 20044996-1001
23
B-Cell Modulation

• Anti-CD20
• Mechanism
• Rapid ablation of the peripheral B-cell
  compartment
• Advantages
• Probably reduces precursor cells responsible for
  clonal expansion during AMR
• May produce more effective antibody reduction
  when combined with plasmapheresis or IVIG
• Well-tolerated, little apparent toxicity
• Effect on the immune system is temporary
  (6-months)
• Disadvantages
• Plasma cells persist in the spleen
• May not, on its own, reduce DSA titers during AMR
  
• Immunosuppressive

24
Case Study AMR in () Cytotoxic XM with High
Titer Anti-HLA DSA
DSA titer
Cr 6.5
512
PP/ CMVIg
PRA
128
100
Cr 4.4
100
100
100
98
120
90
Cr 2.1
Cr 1.5
Cr 1.6
80
85
100
70
DSA titer
80
PRA
60
58
64
50
60
Anti-CD20
CD200 CD190
CD2023.7
40
40
30
32
32
Tx
20
16
16
16
16
16
20
8
8
8
8
8
8
10
4
4
4
4
4
2
2
2
1
1
1
0
0
0
-19
-17
-15
-12
-8
-7
-6
-5
-4
-3
-2
-1
0
2
3
4
5
6
7
8
9
11
12
13
16
18
19
Days from Transplant
25
B-cell Modulation

• Splenectomy
• Mechanism
• Reduces plasma cells, precursor cells, B-cell
  immune surveillance capabilities
• Advantages
• Can be performed using minimally invasive
  techniques
• May produce more effective antibody reduction
  when combined with plasmapheresis or IVIG
• Disadvantages
• Life-long risk of sepsis from encapsulated
  bacteria
• Does not appear on its own to reduce DSA titers
• Effect on immune system is permanent

26
The Effect of Splenectomy on Anti-Blood Group Ab
PP/IVIg
512
256
4
Splenectomy
3
Anti-A Titers (1X)
Serum Creatinine (mg/dL)
128
Tx
2
64
1
32
16
0
-28
-23
-21
-18
-16-15
-14
-12
-10 -6
-4
-3
-2
-1
0
3
5
7
8
10
12
14
17
21
25
27
31
32 34
40
42
45
Day With Respect to Transplant
27
Targets of Strategies for Antibody Removal
Plasmapheresis/IVIG
Plasma cells
Splenectomy
Clonal Expansion
B-cells Pre B-cells
Anti-CD20
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Acute De Novo AMR

• Occurs in 4-6 of transplants (80-100 fail)

• By definition the current XM is negative

• Risk factors include historic XM,
• history of sensitizing event(s), high risk
• donor/recipient combination

• Historically suspected only after there is a poor
  
• response to anti-lymphocytic agents

• Diagnosis should be made by histology and
• demonstration of the appearance of DSA

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PP/CMVIg Treatment Protocolfor Acute De Novo AMR
Plasmapheresis single plasma volume exchange
Steroid bolus -OR- a-thymocyte globulin
IVIG 100mg/kg following each PP treatment (CMV
hyperimmune globulin)
PP/Ig
PP/Ig
PP/Ig
PP/Ig
PP/Ig
8
6
4
2
Dx of AMR
Time Relative to Initiation of Therapy (Days)
Heparin D/C FK 506
High Grade
Johns Hopkins InKTP
30
De Novo AMR Renal Allograft Function
plt0.001
1994-2003 22 recipients of deceased or live
donor kidney transplants with AMR by Bx or
DSA treated with PP/IVIg
Mean f/u 5 1/2 years
Median Nadir Cr (IQR) Median Cr at AMR (IQR) Median 1 week Cr (IQR) Median 1 month Cr (IQR) Median Current Cr (IQR)
3.3 (2.2 7.1) 6.4 (3.2 9.3) 4 (2.3 7.9) 1.9 (1.5 3.0) 1.5 (1.2 2.1)
Johns Hopkins InKTP
31
PP/CMVIg Treatment for De Novo AMR
100
100
Allograft Survival
90
90
80
80
70
70
Live Deceased
p NS
60
60
p NS
50
50
1-Year 87.5 85.8
40
40
30
30
20
20
3-Year 87.5 77.1
10
10
0
365
730
1095
0
365
730
1095
Time (days)
Time (days)
5-Year Overall 81.1
Live donor
Deceased donor
Live donor
Deceased donor
Kaplan-Meier Estimate of Graft Survival for
recipients who developed de novo AMR and were
treated with PP/CMVIg therapy
Johns Hopkins InKTP
32
Bx and DSA Proven De Novo AMR
LD DD p
n 5 13
Median Days to AMR (Range) 11 (8-253) 9 (7-50) 0.25
Median Months F/U (Range) 13.6 (4-76) 11.2 (3-89) 0.77
Johns Hopkins InKTP
33
De Novo Renal Function
15
12
9
Serum Creatinine (mg/dL)
6
3
0
LD
DD
Creatinine at Biopsy
Creatinine 1 week
Creatinine 1 mo
Current Cr
PNS for comparison between groups at each
timepoint
Johns Hopkins InKTP
34
De Novo AMR Allograft Survival
100.00
90.00
80.00
70.00
60.00
Survival ()
50.00
40.00
30.00
20.00
10.00
0.00
0
6
12
18
24
30
36
Time (Months)
Live
Deceased
Johns Hopkins InKTP
35
Rejection and Clinical Outcomes Following() XM
and ABOi
POSITIVE CROSSMATCH
ABO INCOMPATIBLE
OF PATIENTS
28
OF PATIENTS
86
1 2 3
4 1 0
1 2 3
31 12 5
Previous Txs
Previous Txs
27/86 (31) 26/86 (30)
AMR CELLULAR REJECTION
AMR CELLULAR REJECTION
3/28 (11) 4/28 (14)
0/28 (0) 7/28 (25)
7/86 (8) 16/86 (19)
SUBCLINICAL AMR SUBCLINICAL CELLULAR
SUBCLINICAL AMR SUBCLINICAL CELLULAR
1-YEAR GRAFT SURVIVAL 3-YEAR GRAFT SURVIVAL
89.8 80.9
92.9 92.9
1-YEAR GRAFT SURVIVAL 3-YEAR GRAFT SURVIVAL
1 death WNE 1 Noncompliance
Bx _at_ 1, 3, 6, 12 mos
Johns Hopkins InKTP
36
() XM vs. De Novo AMR
De Novo Desensitized p
n 18 31
Median Days to AMR (Range) 10 (7-253) 20 (2-634) 0.16
Median Months F/U (Range) 12.4 (3-89) 14.1 (0.6-65) 0.76
Johns Hopkins InKTP
37
() XM vs. De Novo AMR Outcomes
P0.04
P0.01
P0.002
PNS between groups at current timepoint
Johns Hopkins InKTP
38
Allograft Survival After AMR () XM vs. De Novo
100.00
90.00
80.00
70.00
pNS
60.00
Survival ()
50.00
40.00
30.00
20.00
10.00
0.00
0
6
12
18
24
30
36
Time (Months)
() XM
DeNovo
Johns Hopkins InKTP
39
Kaplan-Meier Estimate of Graft Survival () CDC
XM _at_ Time of Tx vs (-) CDC XM
1 Year Graft Survival
pNS
XM _at_ Tx - XM _at_ Tx
N 14 N 32
92.3 87.2
Johns Hopkins InKTP
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Anti-CD20 Rescue Protocol
Inclusion Characteristics

• Failure to Respond to Plasmapheresis/CMVIg Therapy

• Poor or Incomplete Clinical Response

• Persistence of High-Titer DSA

• Persistence of Histologic Evidence of AMR

• Initial Histologic Features That Portend Poor
  Outcome
• and/or Graft Loss (Grade 2-3 AMR)

• Study Group

• Recipients of Deceased or Live Donor Kidneys

• De novo AMR

• AMR After Desensitization

Johns Hopkins InKTP
41
Renal Function Following Anti-CD20 Rescue
p0.0003
p0.01
10
p0.07
9
p0.25
8
7
17 recipients undergoing a-CD20 rescue therapy
for AMR
6
5
4
3
2
1
0
Best
AMR
2 weeks
1 Month
Current
Best Cr
AMR Cr
2 week Cr
1 month Cr
Current Cr
Best AMR 2 weeks 1 month Current
1.8 (1.4 2.1) 4.3 (2.5 6.5) 3.4 (1.9 5.4) 2.1 (1.6 3.3) 1.7 (1.1 2.6)
42
Kaplan-Meier Estimate of Graft Survival for
Anti-CD20 Rescue
100
75
Survival
50
25
0
1
2
3
4
5
6
7
8
9
10
11
12
Months Following Anti-CD20 Treatment
Johns Hopkins InKTP
43
Splenectomy Rescue
N 4
Median Days to AMR (Range) 4 (2-15)
Median Days to Splenectomy Following AMR Dx (Range) 1 (1-4)
Median SCr at Biopsy Dx (Range) 3.4 (1.5 6.0)
Median SCr 1 week (Range) 2.1 (0.8 5.8)
Median SCr 1 month (Range) 1.5 (0.7 2.3)
Median Current SCr (Range) 1.3 (1.2 2.6)
Allograft Survival 100
Median Months Followup (Range) 6.9 (2.2 11.7)
Johns Hopkins InKTP
44
Paired Donation May Reduce the Incidence of AMR
Conventional KPD
Unconventional KPD
ABOi
ABOi
A
B
A
O
() XM
ABOi
B
A
O
A
of KPD 6 (12 patients)
of KPD 5 (13 patients)
Mean PRA 14
Mean PRA 58
6 mos Cr 1.2 mg/dl
6 mos Cr 1.1 mg/dl
Cellular 8
AMR 0
Cellular 23
AMR 0
Patient Survival 100
Patient Survival 100
Graft Survival 91.7
Graft Survival 100
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Other Therapies

• Leflunomide
• An orally administered antimetabolite
• Action against NF-B and JAK3, key mediators in
  cytokine generation
• Inhibits CMV
• FTY720
• Reduces peripheral lymphocyte levels by causing
  sequestration in lymph nodes and Peyer patches
• Efalizumab
• Monoclonal anti-CD11a antibody
• Blocks T cell adhesion
• Alemtuzumab (Campath 1H)
• Monoclonal antibody targeting CD52 (found on all
  lymphocytes)
• Causes profound, prolonged lymphocyte depletion
• Tolerance induction with full immunosuppression
  and bone marrow transplant

46
Summary
47
Algorithm For Approach To AMR
De Novo AMR
AMR after ABOi
AMR after () XM
PP/IVIg
Response
Incomplete Response
Severe AMR
Anti-CD20
Observe
Anti-CD20
Splenectomy
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49
Definition

• Chronic antibody-mediated rejection (CHR)
• Slow, progressive loss of graft function (months
  to years), often with proteinuria, hypertension

Colvin RB. J Am Soc Nephrol 18 10461056, 2007
50
Chronic rejection

• Fibrosis and scarring in transplanted organ
• Multifactorial etiology, not strictly immunologic
• Inflammation, ischemia, and other processes play
  a role
• Episodes of acute rejection are significant risk
  factors

51
Diagnostic criteria for CHRa

• Histologic evidence of chronic injury (need 2 of
  4)
• arterial intimal fibrosis without elastosis
• duplication of glomerular basement membrane
• multilaminated PTC basement membrane
• interstitial fibrosis with tubular atrophy
• Evidence for antibody action/deposition in tissue
• (e.g., C4d in PTC)b
• Serologic evidence of anti-HLA or other
  anti-donor antibody

a-If only two of the numbered criteria are
present, then the diagnosis is considered
suspicious for CHR. b-May be patchy in
distribution.
Colvin RB. J Am Soc Nephrol 18 10461056, 2007
52
Histology

• GBM duplication
• mononuclear cells in glomeruli and PTC
• intimal fibrosis
• tubular atrophy and interstitial fibrosis
• PTC basement membrane multilamination (by EM)

53
Chronic humoral rejection (CHR)
Transplant glomerulopathy With duplication of
the glomerular basement membrane (GBM) and
accumulation of mononuclear cells in glomeruli
(periodic acid-Schiff stain).
Immunofluorescence shows a patch distribution of
C4d positivity in PTC.
Sis B, Mueller T et al ,. Am J Transplant
Suppl 469470, 2006
54
Chronic humoral rejection (CHR)
The PTC have Prominent multilamination of the GBM.
Electron microscopy shows duplication of the GBM
and reactive endothelial cells.
55
C4d

• Positive PTC (often patchy)
• Glomeruli variably positive
• Occasionally only glomeruli positive

56
Serology

• Anti-donor HLA positive in majority, especially
  to MHC class II antigens.

57
CHR Therapy

• Outcome and optimal therapy not yet defined

58
Accommodation

• Transplantation across an ABO barrier, which
  normally precipitates hyperacute rejection, has
  been done successfully using special protocols to
  deplete naturally occurring antiblood group
  antibodies.
• Remarkably, antibody (primarily IgM) returns in
  the circulation, sometimes in high titers, yet no
  obvious graft rejection occurs, a state termed
  accommodation
• At a cellular level, accommodation may occur via
  multiple mechanisms, including internalization,
  downregulation, inactivation, and inhibition of
  the target antigen

59
Accommodation
Diagnosis

• Normal graft function

Histology

• Normal, or minor changes

60
Accommodation
C4d

• Positive PTC variable glomeruli

Serology

• Common with ABO incompatibility
• occasionally with HLA antibodies

J Am Soc Nephrol 18 10461056, 2007
61
Accommodation
Light microscopy is normal
Electron microscopy shows no glomerular
abnormality.
Prominent C4d deposition in PTC is present
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Accommodation

• Outcome and optimal therapy not yet defined

63
Antibody mediated graft rejectionSummary

• The detection of C4d deposition in capillaries of
  allografts has permitted the definition of two
  new forms of antibodymediated rejection, acute
  and chronic, as well as an apparent
  clinicopathologic state in which the endothelium
  undergoes no obvious pathologic change
  (accommodation).

64
Conclusion

• Organ transplantation induces a fundamentally
  abnormal physiologic state
• Steady improvement has been made in both
  prevention and treatment of organ rejection
• The potential for further improvement both
  incremental and revolutionary is great