Published Online February 7, 2013
Humoral theory of transplantation: some hot
topics
Junchao Cai1, Xin Qing2, Jianming Tan3, and Paul I. Terasaki1*
1
Terasaki Foundation Laboratory, Los Angeles, CA, USA; 2Department of Pathology,
Harbor-UCLA Medical Center, Torrance, CA, USA, and 3Cell and Organ Transplant Institute,
Fuzhou General Hospital, Fuzhou, Fujian, China
Introduction: Antibody is a major cause of allograft injury. However, it has not
been routinely tested post-transplant.
Sources of data: A literature search was performed using PubMed on the topics
of ‘antibody monitoring’, ‘autoantibody and allograft dysfunction’ and
‘prevention and treatment of antibody-mediated rejection (AMR)’.
Areas of agreement: Donor-specific antibody (DSA) monitoring not only helps to
identify patients at risk of AMR, but also serves as a biomarker to personalize
patient’s maintenance immunosuppression. Development of autoantibody is a
secondary response following primary tissue injury. Some autoantibodies are
directly involved in allograft injury, while others only serve as biomarkers
of tissue injury.
Areas of controversy: It remains controversial whether DSA-positive patients
without symptoms need to be treated. In addition, given the variation in study
designs and patient’s characteristics, there is discrepancy regarding which
treatment regimens provide optimal clinical outcome in preventing/treating
AMR.
Growing points: Efficacy of B-cell and/or antibody-targeted therapies in treating
or preventing AMR would be better measured by the incorporation of antibody
monitoring into current functional and pathological assays.
Areas timely for developing research: Research in B-cell targeted therapies to
prevent and treat AMR is rapidly growing, which includes monoclonal
antibodies against B-cell markers CD20, CD40, CD19, BlyS, etc. It requires
extensive clinical research to determine the best approach to inhibit or delete
antibody and how to balance the drug efficacy with safety.
*Correspondence address.
Terasaki Foundation
Laboratory, 11570 W
Olympic Blvd, CA 90064,
USA. E-mail: terasaki@
terasakilab.org
Keywords: alloantibody/autoantibody/antibody-mediated rejection/c4d/HLA
antibody/rituximab/bortezomib/belimumab/ASKP1240/dacetuzumab/
blinatumomab/eculizumab/C1 inhibitor
Accepted: November 1, 2012
British Medical Bulletin 2013; 105: 139–155
DOI:10.1093/bmb/lds037
& The Author 2013. Published by Oxford University Press. All rights reserved.
For permissions, please e-mail: [email protected]
J. Cai et al.
Donor-specific antibodies may pre-exist in allograft recipients before
transplantation due to previous sensitization history, such as blood
transfusion, transplantation or pregnancy. Cross-match technique, in
which recipient’s serum was used to react with donor’s lymphocytes,
was developed in 1969 and proved to predict graft failure.1 Thereafter,
complement-dependent cytotoxicity (CDC) cross-match and panelreactive antibody (PRA) assays were performed routinely on patients
for kidney and heart transplants. Cell-based flow cytometry technique
(middle 1980s), purified antigen-based ELISA (middle 1990s) and Flow/
Luminex (late 1990s) were introduced to increase the sensitivity and specificity of the antibody tests.2 Probably due to the development of single
antigen flow and luminex beads techniques which allow the detection of
donor-specific antibody at high sensitivity,3 the effect of alloantibody
developed post-transplant gets more attention in recent years compared
with a decade ago. Accumulating evidence has proved the causal relationship between alloantibody and graft rejection, especially chronic rejection in human organ transplantation.4 – 6
In this review, our focus will be on the following topics: (i) the importance of routine antibody monitoring post-transplant; (ii) autoantibody and allograft injury; (iii) prevention and treatment of AMR; (iv)
treatment for donor-specific antibody (DSA)-positive patients with
functioning allografts and (v) problems with current diagnosis of
AMR.
The importance of routine antibody monitoring
post-transplant
Antibody is a major cause of chronic allograft injury
Organ transplantation is the best possible treatment for many patients
with end-stage renal failure, but progressive dysfunction and eventual
allograft loss is associated with increased mortality and morbidity.
Immune injury from acute or chronic rejection and non-immune
causes, such as drug toxicity, ischemia –reperfusion injury, recurrent
disease and infection are potential threats.7
Among all immunological risk factors, alloantibody is a major cause
of chronic allograft injury. Lee et al.8 previously reported that all
chronic rejection failures of kidney transplants were preceded by the
development of HLA antibodies. An international cooperative study of
4763 patients from 36 centers demonstrated that the overall frequency
of HLA antibodies were 20.9% (kidney), 19.3% (liver), 22.8% (heart)
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and 14.2% (lung).9 A 5-year prospective study with the 1014 kidney
patients showed that the presence of DSA was associated with a significantly lower graft survival of 49 vs. 83% in the HLAab-negative group
(P , 0.0001). HLA antibodies produced even late after transplantation
are detrimental to graft outcome. These results indicate that a posttransplant HLA antibody monitoring routine could be appropriate to
improve long-term results.10
DSA is a sensitive biomarker of an active donor-specific immune status
Administration of immunosuppressive drugs is the main method for
most of the transplant recipients to reduce recipient’s immune responses
against allografts. Maintenance immunosuppression for each individual
patient is usually adjusted according to drug levels, allograft function
and biopsy results. Biopsy, although an invasive procedure, is still considered to be the gold standard for the diagnosis of allograft rejection.
In an effort to further improve the outcomes on the basis of individualized treatment, there is an urgent need for non-invasive assays,
as well as biomarkers, to more accurately monitor allogeneic responses
and predict the risk of acute and chronic allograft rejection.11
Accumulated evidence has shown that a high frequency of transplant
recipients had HLA antibodies post-transplant and a causal relationship
between HLA antibody and graft failure has been proved.6 Compared
with other biomarkers which have been used for immune monitoring
of transplant recipients,12 HLA antibody, especially DSA, is more specific and sensitive. Development of DSA requires both humoral and cellular immunity. The presence of donor-specific antibody not only
reflects patient’s uncontrolled humoral immunity but also implies an
active allospecific cellular immunity, since the antibody production and
class-switching need the help of T-cells.
Antibody monitoring helps personalization and minimization of immunosuppression
treatment
As mentioned above, an 20% of the recipients with functioning allograft had de novo HLA antibodies, which suggested that immunosuppressive treatments for these patients were not optimized or strong
enough to inhibit patients’ allospecific antibody responses. Since it has
been proved that alloantibody is a major cause of chronic allograft
injury which results in the final graft loss, attention must be paid to the
positive antibody testing result, and corresponding effective therapeutic
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J. Cai et al.
measures need to be taken to prevent the occurrence of irreversible
antibody-mediated tissue damage.
For the majority of patients without HLA antibodies, maintenance
immunosuppressants seemed to work. However, it is possible
that some of these antibody-negative patients are actually overimmunosuppressed. They may suffer from the side-effects of long-term
immunosuppressive therapy. To avoid side effects of maintenance immunosuppressive treatment, research in transplantation focuses on new
treatment strategies inducing tolerance or allowing drug weaning.
Implementing drug minimization into clinical routine can be only
safely achieved when guided by biomarkers reflecting the individual
immune reactivity.12 Prospective trials have proved the predictive value
of immune monitoring for alloantibody in a long-term allograft outcome. Under the careful monitoring of patients for antibodies, clinicians may identify hyporesponsive allograft recipients who could
benefit from a reduction or even withdrawal of immunosuppression.
Certainly, allo-specific T-cell responses may also need to be monitored,
since the lack of humoral responses does not necessarily imply the
absence of cellular responses, despite the cross-talk between cellular
and humoral responses.
Autoantibody and allograft injury
Autoantibodies are developed following tissue injury
Transplantation-related autoantibody responses usually follow allograft
tissue injury. Tissue injury, such as endothelial damage, not only
results in cell death and denaturation of cell surface antigens, but also
exposes recipient’s T- and B-cells to cryptic self-antigen determinants.
Therefore, both denatured antigens on the endothelium/epithelium and
normally unexposed self-antigens can cause autoantibody responses.
In a murine model, Fukami et al.13 showed that alloantibodies cause
epithelial damage to donor MHC and then induce autoimmunity,
which plays a pivotal role in chronic rejection post-lung transplantation. Nath’s recent study showed that the presence of DSA in AMR
and CAV is significantly associated with the presence of antibodies to
myosin and vimentin in post-HTx patients. Induction of high CD4þ
Th cells specific to cardiac self-antigens that secrete predominantly
IL-5 and IL-17 plays a significant role in the development of Abs to
self-antigens leading to AMR and CAV, respectively.14
However, it is clearly worth considering that development of autoantibody is not a specific sign of allograft rejection; instead, it only suggests a previous or ongoing tissue injury. This tissue injury is not
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Antibody-mediated rejection
necessarily an outcome of immunological rejection. Ischemia and reperfusion, drug cytotoxicity and other potential causes of allograft injury
can all induce autoantibody responses.
Association of autoantibodies with chronic allograft injury
In the past, when autoantibody was found to be associated with
chronic rejection, it was very common that only its tissue target was
identified.15 More specific molecular targets were later identified.
Multiple autoantibodies have been reported to be associated with graft
failure in different organ transplantation. The targets of autoantibodies
may include the components of the basement membrane and connective tissues (collagens, agrin),16 denatured extracellular antigens of the
endothelium (angiotensin type 1 receptor, nucleolin, etc.) and intracellular molecules of tissue cells (vimentin, myosin, tubulin, nucleolin,
etc.). Some of the autoantibodies are reviewed here.
Angiotensin-II type I receptor (AT1R) is an angiotensin receptor,
which mediates the major cardiovascular effect of angiotensin II. It is
an important effector controlling blood pressure and volume in the cardiovascular system. Angiotensin II receptor antagonists (losartan, etc.)
are drugs indicated for hypertension, diabetic nephropathy and congestive heart failure. It was reported that the AT1R-activating antibody
was detected at a high frequency (16/33) in kidney recipients with
refractory vascular rejection. Infusion of human AT1R antibodies
induced acute vascular rejection in transplant-recipient rats, which suggested the pathological effect of the antibody.17 Since AT1R-activating
antibody plays its role through its receptors, authors suggested that
patients with AT1R antibodies might benefit from removal of AT1R
antibodies or from blockade of AT1R receptors. However, there is controversy about the effects of angiotensin II receptor blocker and ACEi
in the long-term graft and patient survivals.18,19 Questions that remain
to be addressed include the following (i) AT1R is a cell surface antigen
on both the endothelial and smooth muscle cells, which is exposed to
recipient’s immune system, why do some but not all patients develop
AT1R antibody? (ii) What are the epitopes of AT1R antibodies,
exposed epitope(s) on intact receptors or cryptic epitopes on denatured
receptors? (iii) Is the effect of antibodies always agonic (as described by
Dragun) or antagonistic?
Vimentin is a type III intermediate filament (IF) protein that is
expressed in mesenchymal cells. IF along with the tubulin-based microtubules and actin-based microfilaments comprise the cytoskeleton.
Vimentin is a major cytoskeletal component of mesenchymal cells and
it is also found in the endothelial cells. Vimentin becomes exposed to
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the immune system only if tissue injury occurs due to surgery or graft
rejection when endothelial cell is broken down. Vimentin Abs has been
found to e associated with chronic allograft failure in the heart20 and
kidney.21 In non-human primates vimentin Abs were also associated
with chronic allograft injury in heart transplant.22
Collagen V is a component of most interstitial tissue. TH17-mediated
Collagen V-specific cellular immunity was first found to predispose
allograft injury in lung transplant.23 Later, Iwata demonstrated in an
animal model that transfer of purified IgG or B-cells from col(V)
immune rats to lung isograft recipients all induced pathology consistent
with primary graft dysfunction (PGD) within 4 days post-transfer; upregulated IFN-gamma, TNF-alpha and IL-1beta locally; and induced
significant reductions in PaO(2)/FiO(2). Examination of the plasma
from patients with or without PGD revealed that higher levels of preformed anti-col(V) Abs were strongly associated with PGD development. This study demonstrates a major role for anti-col(V) humoral
immunity in PGD and identifies the airway epithelium as a target in
PGD.24
Some autoantibodies are directly involved in allograft rejection, while others are only
biomarkers of tissue injury
AT1R is a surface antigen of endothelial and smooth muscle cells.
Purified AT1R antibodies from vascular rejection patients were found
to cause acute vascular rejection in a rat model, which proved its direct
pathological effect in allograft rejection.17 Similarly, autoantibodies
against collagen V, a component of interstitial tissues, were proved to
induce pathology consistent with PGD in lung transplantation.24 These
data clearly showed that autoantibodies to AT1R and collagen are directly involved in allograft injury.
Even though significant association has been established between
graft failure/rejection and other autoantibodies, such as vimentin,20,21,25 myosin,25 tubulin,26 nucleolin,27 there is no experimental
evidence to show their pathological effects in allograft injury. The presence of these autoantibodies might be only an indicator of an ongoing
process of tissue injury or cell death, since their targets are mainly
intracellular molecules.
As mentioned above, besides donor-specific alloimmunity, other
causes may also result in tissue injury, which results in secondary autoimmune responses. It is very likely that autoantibodies may pre-exist in
some of the allograft recipients. That might explain why patients with
only autoantibodies but no HLA-DSA seemed to have no detrimental
effect on graft survival (unpublished data). Further investigation is
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needed to determine the mechanism of development of autoantibody
and its association with alloimmune responses, drug cytotoxicity, etc.
Prevention and treatment of AMR
The prevention and treatment of antibody-mediated rejection (AMR) is
mostly empirical. Multiple agents or procedures are being used in
clinic. The therapeutic strategies of these treatments include (i) the inhibition or depletion of antibody-producing cells; (ii) removal or blockage of antibodies; (iii) prevention of antibody-mediated primary and
secondary tissue injury.
Some of the therapies listed in Table 1 have been reviewed previously.28 Here, we focus on some new therapies, which have been proved
to be effective or may have the potential to treat or prevent AMR.
Rituximab
Rituximab is a chimeric monoclonal antibody against B-cell surface
marker CD20, which deletes B-cells via CDC. It was approved by the
US Food and Drug Administration (FDA) in 1997 to treat CD20þ
B-cell non-Hodgkin lymphoma. In 1998, it was originally introduced
into the clinic of solid organ transplantation to treat post-transplant
lymphoproliferative disorders, which is mainly a B-cell disorder.29
Rituximab was first used to treat humoral rejection in thoracic transplants in 200230,31 and in kidney in 2004.32
In addition to its application in anti-rejection treatment, rituximab
has been used as an induction agent to prevent AMR. It was first used
in combination with rATG,33 and later used alone as induction
therapy.34 – 36 All these studies seemed to confirm its effectiveness and
safety. Most recent study by Kohei et al.37 showed that B-cell targeted
induction treatment with plasmapheresis and rituximab (or splenectomy) was associated with lower frequencies of HLA-DSA and chronic
AMR (C-AMR) in ABO incompatible kidney transplantation. Within 2
years of follow-up time post-transplant, the frequencies of DSA and
C-AMR in ABO incompatible transplant recipients were even lower
than those in ABO compatible renal graft recipients. The success of
this preemptive strike using rituximab implies a central role for B-cells
in graft rejection, and this approach may help to delay or prevent AMR
after solid organ transplantation.
Due to the efficacy of anti-CD20 therapy, new generation of CD20
antibodies has been developed and currently under clinical investigation in the treatment of B-cell lymphoma/leukemia. These antibodies
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Table 1 Therapies for prevention and treatment of AMR
Therapy
Potential mechanism in AMR
1. Inhibition or depletion of antibody-producing cells
Induction/maintenance
Direct or indirect effects in inhibiting or depleting the B-cells
immunosuppressants
Splenectomy
Removal of both T- and B-cells
Rituximab and new generation of Monoclonal antibody against B-cells surface marker CD20,
anti-CD20 Mabs
deletion of B-cells via CDC
Bortezomib
Proteasome inhibitor, depletion of both the short- and
long-lived plasma cells
A human monoclonal antibody that binds the BLyS protein
Belimumaba
(BLSP), and prevents the activation of B-cells
Humanized anti-CD40 mAb binds to CD40 on B-cells and blocks
ASKP1240 and dacetuzumabb
the costimulatory pathway
CD19-targeting therapies
Depleting CD19þ B-cells; CD19 is more widely expressed than
CD20 during B-cell development, from pro-B-cells to early
plasma cell stages
Other CD-targeting Mabs
Inhibit or deplete the B-cell-expressing specific CD markers:
Epratuzumab-CD22, Lumiliximab-CD23, SGN-30-CD30,
IMGN529-CD37,hLL1/Milatuzumab-CD74,IDEC-114/
galiximab-CD80
2. Removal or blockage of antibodies
Plasmapheresis/
Removal of circulating antibodies and other humoral factors
immunoadsorption
(complements, cytokines, etc.)
IvIg
Anti-idiotypic effect (blocking of the antigen-binding sites of
anti-donor antibodies) and others
3. Prevention of antibody-mediated primary and second tissue injury
Glucocorticoid
Anti-inflammatory effects
Anticoagulation
Inhibition of clot formation
Eculizumab
Monoclonal antibody directed against the complement protein
C5, preventing the formation of complement C5b-9 membrane
attack complex
C1 esterase inhibitor
Protease inhibitor, inactivates C1r and C1s proteases in the C1
complex of classical pathway of complement
a
NCT01025193: desensitization with belimumab in sensitized patients awaiting kidney transplant; has
not demonstrated efficacy in primary goal. 2011/3, Benlystaw (belimumab) for the treatment of adult
patients with active, autoantibody-positive SLE who are receiving standard therapy. NCT01536379: A
study of belimumab in the prevention of kidney transplant rejection. This study is not yet open for
participant recruitment.
b
2012 AJT(46), long-term hepatic allograft acceptance based on CD40 blockade by ASKP1240 in
non-human primates. 2012 Feb NCT01279538, complete phase I.
include ofatumumab, GA101, veltuzumab, AMME-133v, etc. They
have higher affinity and increased ADCC activity against CD20þ
B-cells, which may offer better alternatives for transplant clinicians to
use as induction or anti-rejection agent to prevent or treat AMR.
Bortezomib
Bortezomib is a proteasome inhibitor which binds the catalytic site of
the 26S proteasome and inhibits its proteolytic function. The US FDA
granted approval to bortezomib for the treatment of patients with
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mantle cell lymphoma (a subtype of B-cell lymphoma) in 2006 and for
the treatment of patients with multiple myeloma (a cancer of plasma
cells) in 2008. Everly et al.38 reported the first usage of bortezomib in
the treatment of acute antibody- and cell-mediated rejection. Authors
demonstrated a significant reduction in DSA levels post-bortezomib
therapy. Stegall’s39,40 in vivo studies provided evidence to show
that proteasome inhibition causes the apoptosis of plasma cells and
therefore controls the production of antibodies. However, it remains
controversial regarding its efficacy in treating AMR. Current bortezomibbased therapy includes not only bortezomib, but also, plasmapheresis,
IVIG or rituzimab. Successful reduction in the DSA could not be
simply attributed to bortezomib with certainty. It was demonstrated
that when used as the sole agent, bortezomib does not decrease donorspecific antibodies.41 Woodle et al.42 indicated that optimal responses
to bortezomib are obtained when AMR was diagnosed promptly and
early in post-transplant period, within 6 months, but less predictable
results for late AMR, possibly due to the existence of long-live plasma
cells in the bone marrow. Besides that, bortezomib is a rapidly cleared
drug following intravenous administration. Its peak concentrations are
reached at 30 min and drug levels can no longer be measured after
1 h. These pharmacokinetic/pharmacodynamic features may also determine its short-term effect.
Belimumab
Belimumab is a human monoclonal antibody and the first inhibitor
designed to target B-lymphocyte stimulator (BLyS) protein, which prevents the activation of B-cells and reduces antibodies.43 Two clinical
studies involving 1684 patients with lupus demonstrated the safety and
effectiveness of belimumab.44,45 It was approved in 2011 by the US
FDA to treat patients with active, autoantibody-positive systemic lupus
erythematosus (SLE). It was the first drug approved by the FDA since
1955 to treat SLE patients. Due to its effectiveness in treating SLE,
multiple clinical trials of belimumab are conducted to test its safety
and efficacy in other antibody/B-cell-related diseases, including membranous glomerulonephropathy, rheumatoid arthritis, Sjögren’s syndrome, systemic sclerosis, wegener’s granulomatosis, chronic immune
thrombocytopenia, myasthenia gravis, etc. Dr Naji at the University of
Pennsylvania started the first phase II trial in February 2010 with eight
patients enrolled to assess the efficacy of belimumab for decreasing
antibodies levels in the sensitized patients awaiting kidney transplantation. The trial was terminated because it has not demonstrated efficacy in primary goal (NIH clinical trial identifier: NCT01025193). A
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new phase II trial sponsored by GlaxoSmithKline will soon start early
2013 to investigate the efficacy of belimumab in preventing kidney
transplant rejection (NIH clinical trial identifier: NCT01536379).
ASKP1240 and dacetuzumab
ASKP1240 and dacetuzumab are the human monoclonal antibodies
which bind to CD40 on B-cells and blocks co-stimulatory pathway.
Blockade of the CD40– CD154 (CD40 ligand) costimulatory signal is
an attractive strategy for immunosuppression and tolerance induction
in organ transplantation. Treatment with the anti-CD154 monoclonal
antibodies (mAbs) resulted in potent immunosuppression in non-human
primates. Despite plans for future clinical use, further development of
these treatments was halted by complications. As an alternative approach, novel human anti-CD40 mAbs, ASKP1240 and dacetuzumab
were developed. Preliminary data in the primate model demonstrated
that a 6-month ASKP1240 maintenance monotherapy efficiently suppressed both cellular and humoral alloimmune responses and prevented
the rejection on the hepatic allograft.46 A phase I study of ASKP1240 in
de novo kidney transplantation has been completed (NIH clinical trial
identifier: NCT01279538).
CD19-targeted therapies
CD19-targeted therapies: CD19 is a membrane antigen which is more
widely expressed than CD20 during B-cell development, from proB-cells to early plasma cell stages. CD20-targeted immunotherapy with
rituximab depletes mature B-cells through monocyte-mediated
antibody-dependent cellular cytotoxicity, but does not effectively deplete pre-B or immature B-cells. CD19-directed immunotherapy is
expected to treat diverse pre-B-cell-related and plasmablast-related malignancies, and humoral transplant rejection. Moreover, in contrast to
CD20-directed immunotherapies, CD19-targeted therapies could purge
the B-cell repertoire of autoreactive clones and reset the developmental
clock to a point that curtails the extent of emerging self-reactivity, in
addition to reducing autoreactive T-cell activation through the elimination of mature B-cells. CD19-directed immunotherapies could, therefore, offer a new horizon in B-cell depletion for the treatment of
multiple autoimmune diseases.47 Since autoantibody is associated with
graft failure as discussed above, the inhibition of autoantibody using
CD19-targeted therapies may also help to prevent autoantibodymediated allograft ininjury. Currently, 18 active clinical trials are being
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conducted to investigate the effectiveness of CD19-targeted therapies in
treating B-cell malignancies. These agents include Blinatumomab
(MT103, a constructed monoclonal antibody with two binding sites: a
CD3 site for T-cells and a CD19 site for target B-cells; it links T and
B-cells and activates T-cell’s cytotoxic activity against CD19þ B-cells),
SAR3419 (an anti-CD19 antibody-cytotoxin conjugate) and MEDI-551
(a humanized CD19 monoclonal antibody with enhanced ADCC activity), etc. Once any one of these agents is proved to be effective in deleting B-cells, it will soon be introduced into transplant clinics.
Other CD-targeted monoclonal antibodies
Other CD-targeted monoclonal antibodies: multiple B-cell surface markertargeted therapies are currently investigated in clinical trials in treating specific CD marker positive lymphoma or leukemia. These antibodies include
Epratuzumab-CD22, lumiliximab-CD23, SGN-30-CD30, IMGN529CD37, hLL1/milatuzumab-CD74, IDEC-114/galiximab-CD80, etc. The
major mechanism of these therapies is to inhibit or deplete B-cells
which express these surface markers. These agents may soon be used in
transplantation if these trials show positive results.
Eculizumab
Eculizumab is a humanized monoclonal antibody directed against the
complement protein C5. It prevents the formation of complement
C5b-9 membrane attack complex and inhibits the effect of CDC.
Eculizumab was approved by the FDA for patients with paroxysmal
nocturnal hemoglobinuria (PNH) in 2007 and atypical hemolytic
uremic syndrome (aHUS) in 2011. Both PNH and aHUS are characterized by complement-induced hemolysis. Stegall et al.48 reported that
complement inhibition using eculizumab in combination with other desensitization therapies decreases AMR in sensitized renal transplant
recipients. Biglarnia et al.49 presented a study using eculizumab as part
of an anti-rejection protocol and demonstrated a prompt reversal of a
severe complement activation by eculizumab in a patient undergoing
intentional ABO-incompatible pancreas and kidney transplantation.
Multiple controlled clinical trials are undertaken to further investigate
the effect of eculizumab in treating and preventing AMR in kidney
transplantation.
C1 (esterase) inhibitor
C1 (esterase) inhibitor is a protease inhibitor. Its main function is to
inhibit the complement system. The US FDA approved CinryzeTM [C1
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inhibitor (human)] in 2008 for routine prophylaxis against angioedema
attacks in adolescent and adult patients with hereditary angioedema,
also known as C1 inhibitor deficiency. C1 inhibitor has been found to
be effective in preventing AMR in an in vitro 50 and animal models.51 – 53
Two clinical trials are conducted in the USA to investigate the safety,
tolerability of C1 inhibitor therapy to prevent or treat AMR
(NCT01134510, 01147302).
Treating DSA-positive patients with functioning allografts
There is no consensus on the clinical relevance of DSA detected by the
solid phase Luminex assay. It remains controversial whether DSApositive patients without symptoms need to be treated. Roelen et al.54
indicated that the reasons may include problem with the assignment of
positivity, antibody subclasses, the ability of complement fixation,
intact/denatured antigen specificities, etc.
In a multicenter study, patients with functioning transplants were
prospectively tested for HLA antibodies. Four years after testing, graft
survival of patients not having antibodies was 81% compared with
58% for patients with HLA antibodies (P , 0.0001).55 This study
clearly showed that antibody is a major risk factor for long-term graft
survival. Without interventional therapy, 42% of patients with HLA
antibodies lost allografts within 4 years which is significantly higher
than that in the control group (19%). Similar clinical data have been
published and reviewed previously.6,56 Based on these findings, it is
strongly recommended that corresponding treatment needs to be taken
to inhibit or remove antibodies and their producing cells. It will be
crucial in preventing antibody-mediated tissue injury and therefore improving the long-term graft outcome.
Currently, clinical trials are being conducted to investigate the effect
of different interventional DSA-targeted therapies on graft outcome.
Regarding the DSA treatment options, no single immunosuppressive
treatment protocol has been proved to be superior to others in reducing
or deleting antibodies. Villarroel et al.57 demonstrated that by depleting
guanosine and deoxyguanosine nucleotides in T and B lymphocytes,
mycophenolate mofetil/mycophenolic acid inhibits their proliferation
and, hence, immunoglobin (Ig) production. MMF has been approved for
the prophylaxis of allograft rejection after renal, cardiac or liver transplant. Our recent experience in treating 10 DSAþ patients with functioning renal grafts indicated that different patients may respond
differently to treatment. DSAs in eight patients seemed to be easily controlled by adjusting maintenance immunosuppressants (seven cases) or
in combination with bortezomib and pulse steroid (one case), while
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other two patients did not respond to any therapy. Long-term effect of
antibody inhibition or removal in patients with functioning allograft
recipients remains to be seen.
Problems with current diagnosis of AMR
Allograft rejections are classified into either T-cell or AMR based on
coexistence of either cellular or humoral factors with acute or chronic
tissue injury. The Banff 97 classification of renal allograft pathology
has been updated several times since its first publication. Incorporation
of C4d as a marker for humoral rejection is a major addition for Banff
Schema. C4d staining and detection of circulating antidonor antibody
help pathologists to better correlate allograft injury with specific
humoral markers.58,59
T-cell mediated rejection is usually diagnosed based on acute or
chronic features of tissue injury in combination with the evidence of
interstitial infiltrate of mononuclear cell, which include T-cells, B-cells
( plasma cells) and monocytes. Interstitial infiltration is an obvious and
relatively long-term phenomenon. Therefore, based on the current criteria for biopsy diagnosis of rejection, T-cell-mediated rejection (TMR)
is seldom missed.
Comparatively, AMR is more easily underestimated. In an acute rejection case, there was no antidonor antibody. Biopsy showed a negative C4d staining and interstitial mononuclear infiltrate (B-cell 40%).
According to the Banff 07 classification of renal graft pathology,
this case was diagnosed as grade III TMR. However, empirical TMR
treatment with three pulses of methylprednisolone and adjustment of
maintenance immunosuppression did not improve graft function.
Interestingly, TMR was successfully treated with B-cell targeted monoclonal antibody rituximab and plasmapheresis.60 Just as in above case,
diagnosis of AMR was very likely missed. The reasons may include the
following (i) C4d staining is a ‘transient’ marker of complementdependent AMR. The timing of the biopsy determines whether or not
it can be detected. Recent evidence indicates that a considerable fraction of renal allograft biopsies showing antibody-mediated injury are
C4d negative.61 These data suggest that diagnosis of AMR using C4d
as a golden standard may result in missing diagnosis of AMR; (ii)
AMR may not always be complement dependent (C4dþ). It is possible
that some AMR is independent of complement activation;28 (iii) It
might be inappropriate to classify a case as T-cell-mediated rejection
when tissue injury is observed with coexisting interstitial mononuclear
cells infiltrate. Because mononuclear cells include not only T-cells but
also B-cells and monocytes. When specific markers were applied,
British Medical Bulletin 2013;105
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J. Cai et al.
plasma cells were found to be present at a high frequency of 57.5%;62
(iv) Since only limited HLA antigens (usually A, B, and DR) are typed
for most of the transplant patients, it is very common that donor
HLA-A, B and DR-specific antibodies cannot be detected when patient
experiences acute/chronic rejection. Lack of HLA-DSA makes the
AMR diagnosis weak, especially with a negative C4d staining.
However, many patients may turn out to be positive for antidonor antibodies against other mismatched donor antigens, such as DP, DQ or
MICA, etc.
In summary, recipient’s immune responses to an allograft involve both
cellular and humoral immunity which usually work together. Simple
pathological classification of graft rejection into either T- or B-cell
mediated rejection may lead clinicians to a wrong therapeutical strategy,
especially when AMR is often underestimated. To better treat acute rejection and prevent chronic allograft injury, a comprehensive treatment
approach using both T- and B-targeted agents, followed by efficient
maintenance immunosuppression, is strongly suggested. Certainly, efficacy of treatment should be carefully monitored by testing functional
and biological markers, such as DSA.
Funding
This study was funded by Terasaki Family Foundation.
Conflict of interest
PT is a Chairman and major stockholder of One Lambda Inc, one of
the companies produces HLA antibody testing kits.
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