Editorial Commentary
Page 1 of 4
Refractoriness to platelet transfusion in the presence of anti-HLA
antibodies—reassessing the alloantibody hypothesis
Jacqueline N. Poston1,2,3, James C. Zimring1,2,4
1
BloodworksNW Research Institute, Seattle, WA, USA; 2Department of Medicine, Division of Hematology, Seattle, University of Washington
School of Medicine, WA, USA; 3Fred Hutchinson Cancer Research Center, Seattle, WA, USA; 4University of Washington School of Medicine,
Department of Laboratory Medicine, Seattle, WA, USA
Correspondence to: James C. Zimring, MD, PhD. BloodworksNW Research Institute, 1551 Eastlake Ave E, Seattle, WA 98102, USA.
Email: jzimring@bloodworksnw.org.
Provenance: This is an invited article commissioned by the Section Editor Dr. Xixi Tan (Chengdu Blood Center, Chengdu, China).
Comment on: Rijkers M, Saris A, Heidt S, et al. A subset of anti-HLA antibodies induces FcγRIIa-dependent platelet activation. Haematologica
2018;103:1741-52.
Received: 17 March 2019; Accepted: 02 April 2019; Published: 16 April 2019.
doi: 10.21037/aob.2019.04.01
View this article at: http://dx.doi.org/10.21037/aob.2019.04.01
Since platelet products first became routinely available in
the 1970s, platelet transfusions have been a mainstay of the
treatment of thrombocytopenic patients, decreasing the
frequency and severity of bleeding sequelae (1). While the
overall clinical goal is to decrease bleeding, the success of
a platelet transfusion is often determined by an increase in
platelet count post-transfusion [often reported as corrected
count increment (CCI)] (2). The CCI may range from a
substantial increase in platelet count to no effect at all; in
some cases, a decrease in CCI can even be observed posttransfusion. Inadequate CCIs are to be expected by chance
alone with some frequency, on a unit-by-unit basis, given
the significant donor variability in platelet quality. However,
some patients consistently have unusually low CCIs
despite transfusion of multiple platelet units from different
donors—such patients are designated as “refractory” (1).
While substantial progress has been made in recent decades
in understanding different causes of refractoriness, treating
thrombocytopenia in refractory patients remains one of the
major challenges in platelet transfusion therapy.
Humoral alloimmunization is considered to be a major
cause of refractoriness, estimated to be responsible for
approximately 20% of cases (1). The prevailing theory is
that patients develop alloantibodies against alloantigens
on platelets [e.g., Human Leukocyte Antigens (HLA) or
Human Platelet Antigens (HPA)] (1). Such alloantibodies
presumably occur in response to exposure to the
alloantigen(s) during transfusion therapy and/or through
© Annals of Blood. All rights reserved.
antecedent exposure from pregnancy or transplantation.
Once formed, alloantibodies bind to alloantigen(s)
expressed on donor platelets and cause rapid clearance
(called the alloantibody hypothesis herein). This theory is
widely accepted in the field and it is justified by substantial
empirical evidence (1,3,4). Moreover, once a patient is
designated as refractory for immunological reasons, the
administration of HLA matched platelets achieves greater
CCIs than random donor platelets (5-7). Data in canine
and murine models provide additional evidence that
alloantibodies can be responsible for clearance of transfused
platelets (8-11). Thus, there are deducible consequences
of the alloantibody hypothesis that are observed both in
humans and experimental animals.
Numerous studies including the landmark Trial to
Reduce Alloimmunization to Platelets (TRAP) (4) have
noted that a substantial number of patients who develop
alloantibodies do not become refractory. This represents
a large practical problem in patient management, both
in terms of predicting which patients may become
refractory, and also in determining if alloantibodies are
the cause of poor CCIs (as opposed to other reasons). As
such, the field has focused on identifying characteristics
to distinguish refractory causing alloantibodies (RCAs)
from alloantibodies that do not cause refractoriness (nonRCAs). Archived samples from the TRAP trial (12,13), as
well as new samples collected over time (14-18), have been
analyzed in search of properties that distinguish RCAs from
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non-RCAs. A large number alloantibody characteristics
have been considered, including titer of antibody (19), IgG
subtype (20), post-translational modifications (21), and the
specific HLA being recognized (15). Although a correlation
with titer and refractoriness has been reported in one study,
none of the above approaches have revealed a clear answer.
The methods of detecting alloantibodies and the cutoff for
being “positive” have also been scrutinized (12), but has led
to little clarity. Thus, as we reach 25 years of work since the
TRAP trial, it remains entirely unclear what distinguishes
RCAs from non-RCAs. However, the search for a defining
characteristic of RCAs continues as new and more
sophisticated tools of biochemistry and cell biology develop
and as theory of alloantibodies and platelets continue to
mature.
In a highly innovative and rigorous report, Rijkers
et al. have advanced our mechanistic understanding of
alloantibody-platelet interactions (15). They demonstrated
that a subset of anti-HLA alloantibodies can cause activation
of donor platelets and phagocytosis by macrophages in
vitro through a mechanism involving FcγRIIa crosslinking
(15). This resembles similar pathways identified in immune
thrombocytopenic purpura (ITP) (22) and transfusionrelated acute lung injury (TRALI) (23). Rijkers et al.
proceeded to show that alloantibodies specific for the
same HLA molecule have different effects on platelet
activation (15). The mechanisms are unclear, but the data
from Rijkers et al. suggested that epitope recognition, rather
than affinity, determines the alloantibodies’ effects (15).
This group has also reported that geometry of antibody
binding affects how complement is activated (16).
While these findings are of substantial importance in
understanding mechanisms of alloantibody interaction with
platelets, at least from our point of view, the data in hand
seem to argue against the utility in predicting RCAs vs.
non-RCAs. For example, only 30% of patient sera activated
donor platelets in vitro despite all patients being refractory
with detectable anti-HLA alloantibodies (15). It remains
possible that the frequency of activating alloantibodies
is higher in refractory than non-refractory patients since
only sera of patients known to be refractory with antiHLA alloantibodies were tested. Thus, it is unclear if
alloantibody induced platelet activation truly plays a role
in refractoriness. If it does, activation of donor platelets by
recipient sera should predict the CCI for a given platelet
unit. This remains to be tested.
After several decades of effort to identify what
distinguishes RCAs from non-RCAs, it seems fair to ask why
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Annals of Blood, 2019
has so little progress been made. This is not to say that new
understanding about alloantibodies has not been generated—
Beligaswatte et al. have noted that quantity of alloantibodies
as measured by intensity of HLA-coated bead binding
correlates with refractoriness (19). However this has yet to
be validated in a larger clinical context and is in apparent
disagreement with other studies (15). While most guidelines
recommend testing of anti-HLA antibodies in patients with
serial inadequate CCIs, there is no standard for the particular
assay or its interpretation (2). Thus, practically speaking, we
remain dependent on CCIs alone to determine refractoriness
and continue to lack an ability to predict a priori. Of course,
there could be some characteristic of RCAs that is different
from non-RCAs and deeper and deeper characterization over
time will eventually uncover the answer. However, it may also
be time to take a step back and re-assess the supposition that
there must be a difference between RCAs and non-RCAs
that explains why some alloimmunized patients are refractory
and others are not.
There are at least two different ways in which some
patients with alloantibodies can be refractory, and others not,
but with no difference in the alloantibodies between these
two groups. First, alloantibodies may be necessary, but not
sufficient for immune mediated refractoriness. Alloantibody
mediated platelet clearance likely requires in vivo biology
that is not present in a serum/plasma sample (e.g., the
reticuloendothelial system, vascular flow through capillary
beds certain organs, etc.). Recipient genetic polymorphisms
(such as in Fcγ receptors and the complement system) may
determine refractoriness in a patient with alloantibodies.
If true, the answer would never come from analyzing
alloantibodies. Moreover, transfusion medicine faces the
challenge of wide genetic diversity in both the patient
population and the therapy itself. Indeed, Rijkers et al.
showed that the platelets from different donors activated
differently in response to the same alloantibody (15).
This finding is consistent with known variability of platelets
from donor to donor in other assays measuring platelet
activation in response to antibody binding. For example, the
performance of serotonin release assays for heparin induced
thrombocytopenia depends upon the use of specific platelet
donors (24). The search for predictive diagnostics may have
to extend beyond plasma.
Second, the reason a defining characteristic of RCAs
remains elusive may be the validity of the alloantibody
hypothesis itself. Given that antibodies can clear
numerous biological targets, it is reasonable to assume that
alloantibodies clear platelets. Yet, a large number of patients
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Annals of Blood, 2019
have anti-HLA alloantibodies and are not refractory (4).
This could be reconciled by searching for additional
characteristics of alloantibodies, or alternatively, considering
a new hypothesis. That said, an outright rejection of the
alloantibody hypothesis would cause its own problems
including the loss of an explanation of the efficacy of HLA
matched platelets in refractory patients with alloantibodies.
The challenge is to consider if there are any alternative
hypotheses that would predict all of the available data.
One such hypothesis is that CD8+ T cells cause platelet
refractoriness.
Alloantibodies and CD8+ T cell responses tend to go
hand in hand. While there certainly can be a pull and
tug between the arms of the immune system (analogous
to Th1/Th2 paradigms in mice), the issue is usually the
nature rather than the absence of antibody. As such, antiHLA alloantibodies may be indicators of immunity and
correlate with CD8+ T cell mediated platelet clearance.
The role of CD8+ T cells has yet to be assessed in platelet
refractoriness, as there are no clinical laboratory assays
that test CD8+ T cell function regarding platelet targets.
However, there is evidence to support the CD8+ T cell
hypothesis. In a mouse model after alloimmunization,
CD8+ T cells cause platelet refractoriness in the absence
of any alloantibodies (25). Whether this translates into
humans has yet to be established. We are not advocating for
rejecting the alloantibody hypothesis nor are we advocating
for the CD8+ T cell hypothesis in particular. However,
at the very least, we present the CD8+ T cell hypothesis
as an example of an alternative theory to the alloantibody
hypothesis that is equally consistent with the known data.
The CD8+ T cell hypothesis is not in conflict with failure
to identify defining characteristics of RCAs and still explains
efficacy of HLA matched platelets in refractory patients. Of
course, other theories in addition to alloantibodies or CD8+
T cells can be put forth that likewise explain the data, but
space limitations preclude their presentation and discussion
here. In light of difficulties defining RCAs, consideration
of alternative theories is warranted while simultaneously
refining alloantibody characterization.
Acknowledgements
Funding: JN Poston receives support from an institutional
training grant from the National Heart, Lung, and Blood
Institute (T32 HL007093). JN Poston and JC Zimring
would like to recognize Dr. Monica B. Pagano for reviewing
this editorial.
© Annals of Blood. All rights reserved.
Page 3 of 4
Footnote
Conflicts of Interest: JC Zimring is on the scientific advisory
board of Rubius Therapeutics, which is unrelated to the
content of the current work. JN Poston has no conflicts of
interest to declare.
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doi: 10.21037/aob.2019.04.01
Cite this article as: Poston JN, Zimring JC. Refractoriness to
platelet transfusion in the presence of anti-HLA antibodies—
reassessing the alloantibody hypothesis. Ann Blood 2019;4:8.
© Annals of Blood. All rights reserved.
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Ann Blood 2019;4:8