ABMR pam

Sumanee Prakobsuk
10/07/2012

 Pathophysiology and Pathology.
 Diagnosis criteria.
 Major Histocompatibility Complex (MHC)
molecule .
 Transplantation in the Presence of Antidonor
HLA Antibodies (sensitized patients).
 Treatment Acute ABMR.

 Graft rejection caused by Abtibodies directed
against HLA molecules, ABO antigens or
endothelial cell antigens.

 Most recipients do not have antibodies
against HLA molecules before transplantation
unless they were sensitized by exposure to
alloantigens through
◦ Pregnancy
◦ Blood transfusion,
◦ Previous transplantation.

Donor
Organ 4
Damaged C1
Capillary
Endothelial
Cell
Releases complex

cell platelet
aggregatio C4
Formation n
of factors, cyt 
Antigen- okines
Ab C4b 
complex
C4d
Endothelial
cell
necrosis

C4d is by-product and marker
of complement activation

Schwartz, NEJM 2010

Glomerulitis Peritubular
capillaritis

C4d +

Transplant glomerulopathy
-Thickened of GBM
-Double contours

C4d +
Multilamination of GBM,PTC

Triad
 C4d+

 Presence of circulating antidonor antibodies
 Morphologic evidence of acute tissue injury, such
as (Type/Grade):
◦ I. ATN-like minimal inflammation
◦ II. Capillary and or glomerular inflammation (ptc/g >0)
and/or thromboses
◦ III. Arterial—v3 (tranmural arteritis/fibrinoid necrosis)

 C4d+
 Presence of circulating antidonor antibodies
 Morphologic evidence of chronic tissue injury
◦ Glomerular double contours
◦ Peritubular capillary basement membrane
multilayering
◦ Interstitial fibrosis/tubular atrophy
◦ Fibrous intimal thickening in arteries

 Suspicious for antibody-mediated rejection if
- C4d
or
- Alloantibody
Not demonstrated in the presence of
morphologic evidence of tissue injury.

 Major molecule for self
vs. non-self
determining process

 Very high antigenicity

 In human = human
leukocyte Ag (HLA )

Gabriel M.Danovitch.Hand book of transplantation Fifth Edition

Class I Class II
 On the surface of all  Antigen-presenting cells
nucleated cells (APCs), monocyte, macro
phage, Kuffer
cell, dendritic
 Density of HLA class I cells, alveolar type2
expression cells, renal mesangial
 Plt > B cell > T cell cells, and B lymphocyte
 A, B, C, E, F, G, MICA, MI  DP,DQ, DR, DM,DO
CB
 Present Ag peptides to  Present Ag peptide to
CD8 T cells CD4 T cells


 Increased risk of:
 1. Hyperacute rejection
 2. Memory B cell response leading to early ABMR
 3. Chronic active ABMR

Pretransplant immunologic
evaluation
As always, we are doing three key tests

◦ Tissue typing
 ABO typing
 HLA typing
◦ HLA antibody screening
◦ T and B cell crossmatching

Panel Reactive Antibody
 Determine the state of
pre-sensitization of the transplant
 Predict cross match result
 Predict waiting time

Technologies used to detect HLA
antibodies (sensitization)
2 main methodologies:
Complement-dependent
cytotoxicity (CDC)
Serology
CDC-anti-human globulin (CDC-
AHG)

Enzyme-linked immunosorbent assay (ELISA)

Solid Flow
phase cytometry
Single antigen beads
Luminex

Lymphocytes
(T cells, usually)

Patient
serum

+ rabbit
complement

Red = dead CANNOT
Green = alive
DIFFERENTIATE
IgG FROM IgM

Lymphocytes

Patient
serum
Enhance with
anti-human
globulin

(AHG)

+ rabbit
complement

MCS=median channel shift MFI=mean fluorescence intensity

Luis G. Hidalgo,UAH Histocompatibility Laboratory

Luis G. Hidalgo,UAH Histocompatibility Laboratory

 Most anti-HLA Ab are IgG.

 Donor specific antibody against HLA Class I
or II IgG were clinically relevant conferring
both short and long term risk to the patients.

 IgM HLA ab are not clinically relevant.

 All CDC + IgG (either B or T cell )
contraindication for transplantation
Gebel and Bray. Transplantation Reviews 20: 189-194, 2006

 CDC – and AHG +
◦ no hyperacute rejection but may result in early(1-
2wk) acute rejection and graft loss

 CDC – and FCXM +
◦ High risk in
 Retransplant with previous early graft loss
 PRA >10% both primary and regraft
◦ Low risk in
 Current PRA< 10% both primary and regraft

Gebel and Bray. Transplantation Reviews 20: 189-194, 2006

Year of Waiting List
Registration
Peak 2000 2003 2005 2007 2009
PRA
0-9%

10-
16.5%
79%
OPTN/SRTR 2010 Annual Report

Year of Transplant

Peak 2000 2003 2005 2007 2009
PRA
0-9%

10-
79%
OPTN/SRTR 2010 Annual Report

Peak 3 1 years 5 years 10
 This has led to the problem of determining
PRAthreshold level and characteristics years
the
month (Tx2007 (Tx2003
of
s -2008) -2008) (Tx1998
donor-specific HLAab (DSA HLA-ab) that have
-2008)
(Tx200
a meaningful impact on clinical outcomes
7-2008)
0-9%

10- OPTN/SRTR 2010 Annual Report
79%

 Retrospective review.
 DDKT, 18 centers
 To investigate the relationship between the
pretransplant presence of HLA class I and
classII antibodies and the development of no
immediate function and Acute rejection
episode.

Patients with NIF or ARE were positive for
HLAclass I and II Abtibodies in their
pretransplantation serum than patients
without NIF or ARE

Strong relationship
between the presence
of HLAab and
 poor immediate graft
function
 acute rejection
However, there was reduce graft survival
 no determination of
whether the HLAabs were DSA

 Observational study
 Single-center study of 402 consecutive DDKT.
 Examined the impact of the strength of HLA-DSA
detected on the risk for AMR and graft survival in
DDKT 1998-2006
 DSA HLAab by Luminex single antigen bead assay
 Mean F/U 51.4+- 30.6 months

Carmen Lefaucheur. J Am Soc Nephrol 21: 1398–1406, 2010

AntiHLA+ DSA+61%
AntiHLA- DSA - 84%
AntiHLA+DSA- 93%

The presence of HLA-DSAs on the highest rank
pregraft serum associates with a significantly
decreased graft survival (A),
regardless of whether HLA-DSAs were class I or II (B).

Total Transplant Pt Without ABMR Pt


 long term graft survival was significantly inferior
for patients who had any detectable preexisting
DSA

 Luminex peak MFI predicted AMR and graft
survival.

 MFI > 3000 appeared to the cutoff for significant
decrease in graft survival and whether an episode
of ABMR occurred.


 Proteins other than HLA antigens can also
serve as targets of AMR.

 MICA : MHC Class I chain A.
 Antiangiotensin type I receptor antibody.

NephSAP Transplant, november 2011

 MICA antigens are expressed on endothelial
cells, dendritic cells, fibroblasts, epithelial cells, and
many tumors

 But not on peripheral-blood lymphocytes.

 MICA protein do not associate with B2 micoglobulin
as do MHC class I antigens and not serve to
present antigen to T cell

 They are instead ligands for NK cells.


 Since MICA antigens are not expressed on
lymphocytes,the cells commonly used for
cross-matching
 Antibodies directed against MICA are not
detected with the methods generally used.


 To determine whether an immune response to MICA
antigens might play a role in the failure of kidney
allografts.

 Pretransplantation serum samples from 1910 DDKT.
 Between 1990 and 2004
 20 centers in 13 countries.
 IgG anti-HLA class I & II test : ELISA kits
 Tests for IgG antibodies against MICA antigens
:microbeads (Luminex)
Yizhou Zou, M.D.N Engl J Med 2007;357:1293-300.

93 0.6 %
88.3 2.2 %
P=0.01
11.4 %

 Presensitization of kidney-transplant
recipients against MICA antigens is
associated with an increased frequency of
graft loss and might contribute to allograft
loss among recipients who are well matched
for HLA.

Yizhou Zou, M.D.N Engl J Med 2007;357:1293-300.

 These studies are unable at this time to
provide any absolute thresholds for the
decision to transplat with a given organ or not.

 But do provide data to begin to define level of
risk


Kwaku Marfo.Clin J Am Soc Nephrol 6: 922–936, 2011

PP/low-dose IVIG
author No Pts Inducti F/U AR/AM Pt Graft
on Months R survival survival
(%) (%)
Schweitz 11 OKT3 13 36/27 100 100
we2000

Magee 28 Thymo/ 22 71/39 93 89
2008 Basilixim
ab/Ritux

Thielke 51 Thymo/ 23 33/24 95 93
2009 Ritux
Haririan 41 OKT3 or 47 24/12 78 66
2009 Thymo


High-dose IVIG
author No Pts Induct F/U AR/ Pt Graft
ion Month AMR surviv surviv
s (%) al(%) al(%)

Glotz 13 Thymo 12 8/8 100 93
2002

Jondan 42 Daclizu 24 31/31 98 89
2003 mab

Mai200 20 Thymo 36 50/30 94 89
9
Bachler 37 Thymo 24 38/38 95 87
2010


M.D. Stegalla. American Journal of Transplantation 2006; 6: 346–351

negative crossmatch
Acute ABMR

PP/low-dose IVIG and rituximab
demonstrated more success in abrogating positive
cross-match and lower acute rejection rates

 To investigate the effects of desensitization
protocols using IVIg with or without
plasmapheresis in patients with donor-specific
anti-HLA antibodies on prevention of
antibody-mediated rejection and downregulation
of donor-specific antibodies.

Enver Akalin. Clin J Am Soc Nephrol 3: 1160–1167, 2008

 Pretransplantation DSA, negative CDC cross-
match.
 Anti-HLA antibodies were studied by Luminex
single Beads .
 Biopsies were performed for an increase in
creatinine level and/or proteinuria.


 Induction:Thymoglobulin 1.5 mg/kg per d for 5 d
 Maintenamce: tacrolimus, mycophenolate mofetil,

and a steroid taper.

 All patients received high-dosage IVIG 1.0 g/kg during
transplant surgery and 500 mg/kg on each of
postoperative days 1 and 2.


◦ LRKT candidates with strong class I DSA
 4-8 sessions of pretransplantation PP over 2 to 3 wk
 underwent transplantation after their DSA strength
decreased to moderate or weak.

◦ DDKT recipients with DSA
 3 sessions of PP every other day starting on
postoperative day 1.

Strong MFI> 6000
Moderate MFI 4000 to 5999
Weak MFI 1500 to 3999.


 Group 1 , Seven ( 70%) patients lost DSA completely
 Group 2, four (44%) patients lost DSA Completely
 Group 3, six (43%) patients lost DSA completely


 Kidney transplant recipients with DSA are at higher
risk for developing early acute AMR despite negative
CDC T cell cross-match and require desensitization.

 Not only should the presence of DSA be
documented, but also the strength or titers of the
alloantibodies should be determined to decide the
type of the desensitization protocol.

 Highdosage IVIG alone dose not prevent AMR in
patients with strong DSA

 Aaddition of peritransplantation PP significantly
 decreases the incidence of AMR.

 Assessed the histological lesions at
 3 months and 1 year in patients receiving DDKT, comparing
those with preformed DSA to those without.
 Second, we evaluated the presence and extent of SAMR.
 From January 2002 to March 2007

A. Loupya. American Journal of Transplantation 2009; 9: 2561–2570

Group A (n = 54 )
DSA positive
 Induction :10-day course of ATG a dose of 75 mg/d.

 4 courses of IVIg a dose of 2 g/kg administered over 96 h
◦ first course started before reperfusion,
◦ subsequent courses being given on days 21, 42 and 63.
 Screening onward, the final 18 patients
From 2006 Kidney Bx and measured glomerular
filtrationat day 4(GFR) at 3 months and dose of 375
◦ Received additional prophylactic Rituximab at a
mg/m2
rate 1 year.
 Together with plasmapheresis performed immediately
posttransplant then three times per week for 3 weeks.

Group B (n = 83)
without preformed DSA

20 mg intravenous Basiliximab Day 0,4

At 3 months after transplant
31 % Subclinical AMR in DSA +

At 1 year
 Score higher IF/TA
100 % vs 33 %

 TG 43% vs 0%

 If these findings are comfirmed
in a large series of patients.
 Protocol Biopsies may be
a valuable tool in the management of this
population ??
 Treatment protocol ??


Our review article demonstrates the
importance of the strength of DSAs for
development of AMR.

Currently, we screen all transplant
candidates for anti-HLA antibodies
using Luminex single-antigen beads for
the specificity and the strength of
antibodies Kwaku Marfo.Clin J Am Soc Nephrol 6: 922–936, 2011

Renal Division,
Albert Einstin College of Medicine
MontefioreMedicalCenter,Bronx, New York

C. Wiebea,†, I. W. Gibsonb,c,†,T. D. Blydt-Hansend, M.
Karpinskie, J. Hoe,
L. J. Storsleye, A. Goldbergd, P. E. Birkd,D. N. Rushe and P. W.
Nickersona,c,*

 Sequential evaluation of sera for dnDSA in a
consecutive cohort of kidney transplants.
 Risk factors for dnDSA development
 Correlation of dnDSA with clinical pathologic
and outcome.
C. Wiebe. American Journal of Transplantation 2012; 12: 1157–1167

DSA screening was performed using FlowPRA
beads representing HLA-A,-B, -Cw, -DR, -DQ
and -DP antigens
HLA antibody specificities was performed using
FlowPRA single antigen class I and II beads


 Kidney biopsy
◦ Six-month protocol biopsies.

◦ Newly detected dnDSA patients since January
2008 as standard of care

◦ Clinically indicated allograft biopsy.
 proteinuria was ≥0.5 g/day
 Cr rose ≥25% from baseline without a known cause.


 dnDSA develops in 15% of low risk renal transplant
recipients

 Mean 4.6 +- 3 years posttransplant

 Graft survival at 10 years reduce by 40%
 Independent risk factors for dnDSA development
◦ HLA-DRB1 MM
◦ nonadherence
◦ cellular rejection before dnDSA onset

 The dnDSA typically arises before the onset of
proteinuria or rise creatinie.


 6: TREATMENT OF ACUTE REJECTION

 6.1: We recommend biopsy before treating
acute rejection, unless the biopsy will
substantially delay treatment. (1C)

 6.2: We suggest treating subclinical and
borderline acute rejection. (2D)

BL Kasiske et al.: KDIGO guideline 2009 for kidney transplant recipients

6.4: We suggest treating antibody-mediated acute rejection
with one or more of the following alternatives, with or
without corticosteroids (2C):
 Plasma exchange
 Intravenous immunoglobulin
 anti-CD20 antibody
 lymphocyte-depleting antibody.

6.5: For patients who have a rejection episode, we suggest
 Adding mycophenolate if the patient is not receiving
mycophenolate or azathioprine, or switching azathioprine
to mycophenolate. (2D)

BL Kasiske et al.: KDIGO guideline 2009 for kidney transplant recipients

IAN R. M ACKAY, M.D., N Engl J Med, Vol. 345, No. 10 September 6, 2001

Routinely used
 High dose: 2 gm/kg
 Low dose: 100 mg/kg per session

 Low-dose IVIG is mostly used in combination
with plasmapheresis where it may help replenish
depleted IGs.

 Initial studies used IVIG at high-doses without
plasmapheresis and described a fair degree of
success in desensitization prior to transplant and
also for treating antibody-mediated rejection.
Chethan Puttarajappa, Journal of Transplantation Volume 2012

 Side effects
◦ Aseptic meningitis
◦ Volume overload
◦ AKI possible to high osmotic load


 Plasmapheresis is very effective in reducing the
antibody load but needs to be used in
conjunction with other therapies that target the
antibody producing mechanisms.

 DSAs are monitored along with renal function to
document the effectiveness of the therapy.

 Treatment, if successful, is continued until the
level of antibodies has dropped to safe levels
along with improvement in renal function.


 The mechanism of action of Rituximab in AMR is
not clear, however, the depletion of CD20-positive
subset of B-cells may attenuate
the antibody generation process.

 Side effects
◦ Acute infusion reactions
◦ Reactivation of latent viruses such as hepatitis
B, C, CMV, and TB


 The retrospective 2001-2006
 Compared the outcomes of a PP-based vs. a
PP plus rituximab regimen to treat patients
experiencing AMR and resistant to steroid
plus anti-lymphocyte globulin treatment.

Kaposztas et al.Clin Transplant 2009: 23: 63–73

• At the end of each PP cycle
Rituximab(375 mg/m2).

Induction:
•simulect
•High risk PRA>20%,African,re-transplant:Thymoglobulin

Graft survival rates at 2 years
group A 90%
group B 60%

Beneficial effect was observed with PP in
addition to treatment with rituximab in AMR

Kaposztas et al.Clin Transplant 2009: 23: 63–73

 DDKT
 Biopsy prove AMR & DSA+
 Negative current CDC crossmatch
 All patients received induction with
◦ thymoglobulin (1.5 mg/kg/day × 7–10 doses)
 Maintenance immunosuppression
◦ steroids +Cellcept+tacrolimus /cyclosporine
 Patients with remote positive IgG T- and B-cell CXM received IVIg at
the time of transplantation as prophylaxis against acute rejection (2
g/kg days 0–1, 20–21 and 40–41).

Lefaucheur.American Journal of Transplantation 2009; 9: 1099–1107

 High-dose IVIg regimen (group A)
◦ 12 patients with AMR/DSA+
◦ diagnosed between January 2000 and December 2003
◦ 2 g/kg IVIg, administered over 2 days every 3
weeks, × 4 doses

• Plasmapheresis /IVIg/anti-CD20 regimen (group B)
-12 patients with AMR/DSA+
-diagnosed between January 2004-December 2005.
-daily 1-PV followed by administration of low dose
of IVIg (100 mg/kg) *4
-After the last PP
-high-dose IVIg as described above (2 g/kg every
3 weeks, × 4 doses)
-two weekly doses of rituximab (375 mg/m2)

PP /IVIg/anti-CD20

High-dose IVIg
regimen

Graft survival at 36 months
following the episode of AMR
•50% in group A
•91.7% in group B


High-dose IVIg regimen PP /IVIg/anti-CD20

 PP/IVIg/anti-CD20 leads to improved graft
survival over protocols using IVIg alone.

 Graft survival at 36 months was
◦ 91.7% PP/IVIg/anti-CD20 regimen
◦ 50% in IVIg.

 Diminution of DSAs levels is significantly
greater in patients treated by the association
of PP/IVIg/anti-CD20 as compared to those
treated by IVIg.


 Bortezomib is a novel proteosome inhibitor
that is approved for the treatment of multiple
myeloma.

 Inhibition of proteasomes can lead to decreased
nuclear factor-Kappa B activation, cell cycle
arrest, endoplasmic reticulum stress, and increased
cell apoptosis .

 This action is pronounced in plasma cells likely
because of the high antibody turnover and high
endoplasmic reticulum activity.
Rajeev Raghavan,Journal of Transplantation Volume 2010

Author Center N Complete Results summary
therapy
Idica et al.
2008
- 13 Detail not
apparent
(i) 10 of 13 had significant
decrease (reversal) of DSA
+
(ii) 100% had reduced MFI of
antibodies
Raghavan et
al.
Houston,
TX,
1 (i) 4 cycles
bortezomib, one
(i) Reduced PRA (55% → 30%)
and significant reduction of +
USA dose rituximab, class I antibodies
daily (ii) Successful transplant with
mycophenolate good allograft function at 6-
months

-
Wahrmann Vienna, 2 (i) 2 cycles (i) cPRA mildly decreased in
et al.2010 Austria bortezomib at both patients
intervals of 3- (ii) Overall, no significant
and effect on the levels of
4-months, both antigen-specific IgG or ABO
given with blood group antibodies
steroids


Author Center N Complete therapy Result summary

Walsh et al. Cincinnati 2 (i) 1 cycle bortezomib
(ii) ongoing
(i) Immediate
significant reduction
+
Ohio, USA
plasmapheresis, of DSA
rituximab, (ii) Good allograft
intravenous steroids function at 5- and
(iii) pheresis done at 6-months follow-up
least 72 hours post- (iii) One patient had
bortezomib re-elevation of DSA
which responded to
a second
course of treatment
Sberro-
Soussan
Paris,
France
4 (i) 1 cycle bortezomib
(solo therapy)
(i) No effect on anti-
HLA antibodies -
et al. 2010 within 40
subsequent days,
and at 150
days follow-up.


 case series of four renal transplant recipients in whom
graft biopsy disclosed ABMR , accompanied by
persistent DSA.
 All patients received bortezomib (1.3 mg/m2) on days
1, 4,8 and 11 as sole desensitization therapy without
any modification of their maintenance
immunosuppressive treatment.

American Journal of Transplantation 2010; 10: 681–686

American Journal of Transplantation 2010; 10: 681–686

 Bortezomib
failed to
decrease DSA
intensity within
the 150-day
follow-up
period in all
patients.

They concluded that a single cycle of bortezomib
does not seem to exert an effect on any long-lived
antibody levels (further than 1 year post-transplant)

 20 patients with AMR and DSA + a mean of
19 months after transplantation.

Flechner et al. Transplantation • Volume 90, Number 12, December 27, 2010

1. For acute cellular rejection, Banff scored grade 1A/B
:initial pulse steroid treatment with 15 to 20 mg/kg IV
methylprednisolone given in three divided daily doses
(500 mg three times).

2. Initiation of plasmapheresis twice weekly for two weeks
(total four treatments). Treatments were spaced to
days 1-4-8-11.

3. IV bortezomib given as 1.3 mg/m2 after each
plasmapheresis (total four treatments).

4. When the plasmapheresis was completed, the addition of
2 g/kg IVIG (0.5 g/kg in four divided treatments) was
given to the majority of recipients.

Flechner et al. Transplantation • Volume 90, Number
12, December 27, 2010

For the entire group, patient survival is
100%, and graft survival is 85% with a
mean follow-up of 9.8 months

They found that
• patients with SCr< 3 for combining
There is a rational
mg/dl had better plasmapheresis, as it
Bortezomib with
response more effective in eliminating
may be
plasma cell that produceing high levels
of antibody.
•The mean decrease from peak-
nadir MESF/MFI of the most
dominant DSA was 55% .
•only 25 % had undetectable
DSA after treatment.

Flechner et al. Transplantation • Volume 90, Number 12,
December 27, 2010

 Neurotoxicity 30 %
 Thrombocytopenia 28%
 Neutropenia 11 %
 Nausea 55%
 Diarrhea 44%
 Fatigue 12%

Rajeev Raghavan,Journal of Transplantation
Volume 2010

 Humanized monoclonal antibody directed against
complement protein C5.
 Thereby inhibiting conversion of C5 to C5b and
preventing formation of the membrane attack
complex (C5–9).

 Antibody-mediated rejection is an important cause of
acute and chronic graft failure.

 Improvements in HLA technology
revolutionized the understanding of this important entity.

 Transplantation of sensitized patients remains a difficult
problem.

 However, developments such as paired kidney donation
and desensitization protocols
are continuously improving the rates of transplantation in
this difficult to transplant population.

 Therapies for AMR are still not optimal with high
rates of graft loss leading to poor patient outcomes.

 Newer therapies, such as bortezomib and eculizumab
that target novel pathways in the AMR process are
promising but will need further randomized studies
before becoming widely used.

 Studies will need to be performed to determine the
best use, either alone or in combination, of the
myriad number of therapies currently available

ABMR pam

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