From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
eculizumab therapy.8,9 Therefore, larger
studies are needed to clarify the exact extent of
the C3d(g)-mediated extravascular hemolysis
in PNH patients under eculizumab treatment.
The results of Lin et al presented
here describe a novel mechanism of
erythrophagocytosis based on the CR3-C3d
interaction, which may be operational in
pathophysiologic conditions. If occurring in
vivo, this mechanism provides a rationale for
optimization of the therapy in some patients,
who remain transfusion dependent despite
the eculizumab treatment, using complement
inhibitors acting on the level of C3. This
phenomenon needs attention, because it may
not be restricted to PNH but may also be
relevant to other diseases, characterized by C3d
deposition on cells, including autoimmune
hemolytic anemia, graft rejection, and
atherosclerosis.
Conflict-of-interest disclosure: The author
declares no competing financial interests. n
REFERENCES
1. Lin Z, Schmidt CQ, Koutsogiannaki S, et al. Complement
C3dg-mediated erythrophagocytosis: implications for paroxysmal
nocturnal hemoglobinuria. Blood. 2015;126(7):891-894.
2. Merle NS, Church SE, Fremeaux-Bacchi V,
Roumenina LT. Complement system part I: molecular
mechanisms of activation and regulation. Front Immunol.
2015;6:262.
3. Merle NS, Noe R, Halbwachs-Mecarelli L, FremeauxBacchi V, Roumenina LT. Complement system part II:
role in immunity. Front Immunol. 2015;6:257.
l l l THROMBOSIS AND HEMOSTASIS
Comment on Gunasekera et al, page 895
Game, set, match for
factor
VIII mismatch?
----------------------------------------------------------------------------------------------------Connie H. Miller
CENTERS FOR DISEASE CONTROL AND PREVENTION
In this issue of Blood, Gunasekera et al provide evidence that the high rate of
factor VIII (FVIII) inhibitors seen in black hemophilia A (HA) patients is not due
to a mismatch between the structure of treatment products and FVIII genotypes
common in blacks.1
B
lack HA patients have consistently been
shown to have an almost twofold higher
frequency than white patients of inhibitors,
the neutralizing antibodies that can develop
against FVIII treatment products and limit
their usefulness in stopping or preventing
bleeding.2,3 The mismatch hypothesis to
explain this disparity was an innovative
concept proposed by Viel et al based on
their observation that the pattern of 4
nonsynonymous single-nucleotide
polymorphisms (ns-SNPs) coding for
amino acid changes in normal FVIII
structure could be used to define 6 F8
haplotypes, the frequencies of which vary
by race.4 The 2 most common haplotypes,
H1 and H2, are represented in different
full-length recombinant FVIII products
used for HA treatment. Among 76 African
Americans, 25% had haplotypes other than
H1 and H2, and this group had increased
odds of having an inhibitor (odds ratio, 3.6;
95% confidence interval, 1.1-12.3; P 5 .04).4
Subsequent studies of small populations of
patients of African ancestry have failed to
confirm this finding.5-7 The haplotypes
investigated are rare in white populations,
and no correlation with inhibitors was found
in substantially sized groups of white
patients.5,8
Gunasekera et al present the most
comprehensive study to date to address this
4. Ross GD, Lambris JD. Identification of a C3bi-specific
membrane complement receptor that is expressed
on lymphocytes, monocytes, neutrophils, and erythrocytes.
J Exp Med. 1982;155(1):96-110.
5. Bajic G, Yatime L, Sim RB, Vorup-Jensen T, Andersen
GR. Structural insight on the recognition of surface-bound
opsonins by the integrin I domain of complement receptor 3.
Proc Natl Acad Sci USA. 2013;110(41):16426-16431.
6. Hillmen P, Young NS, Schubert J, et al. The
complement inhibitor eculizumab in paroxysmal
nocturnal hemoglobinuria. N Engl J Med. 2006;
355(12):1233-1243.
7. Peffault de Latour R, Fremeaux-Bacchi V,
Porcher R, et al. Assessing complement blockade in patients
with paroxysmal nocturnal hemoglobinuria receiving
eculizumab. Blood. 2015;125(5):775-783.
8. Hill A, Rother RP, Arnold L, et al. Eculizumab
prevents intravascular hemolysis in patients with
paroxysmal nocturnal hemoglobinuria and unmasks
low-level extravascular hemolysis occurring through
C3 opsonization. Haematologica. 2010;95(4):567-573.
9. Risitano AM, Notaro R, Marando L, et al.
Complement fraction 3 binding on erythrocytes as
additional mechanism of disease in paroxysmal nocturnal
hemoglobinuria patients treated by eculizumab. Blood.
2009;113(17):4094-4100.
DOI 10.1182/blood-2015-07-653428
© 2015 by The American Society of Hematology
Risk factors for development of inhibitors (neutralizing antibodies) against treatment products used to stop or prevent bleeding
in hemophilia patients include 3 major categories. The causative mutation in the gene for FVIII or FIX has been shown to be
the most significant risk factor, as whether a gene product is produced or its structure determines how the immune system
recognizes the infused protein. The treatment product itself, how much exposure has occurred, and conditions of that
exposure provide the trigger for the immune response. Determinants within the immune system control the response through
a multiplicity of genes producing a wide phenotypic variability. Interaction of these 3 components, illustrated by the
overlapping circles of the Venn diagram, provides each person with hemophilia with a highly individualized risk of developing
this complication of treatment at some point during his or her lifetime. Professional illustration by Ken Probst, Xavier Studio.
BLOOD, 13 AUGUST 2015 x VOLUME 126, NUMBER 7
829
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
question, using 3 different approaches.1 First,
statistical analysis using 174 African American
and 198 white HA patients confirmed the
increased inhibitor frequency in African
Americans but showed no correlation of
inhibitor status with ns-SNPs or haplotypes.
The only statistically significant finding
was a higher inhibitor frequency in
patients “potentially exposed to ‘sequencemismatched’ FVIII” than in those not exposed.
As the authors note, this should be interpreted
with caution. The exposed group included any
patient who had ever received a blood product
or plasma-derived factor, including FEIBA.
Because FEIBA is used primarily to treat
inhibitor patients, its inclusion may bias the
results. Second, binding affinities of peptides
containing the relevant ns-SNPs to HLADRB1 alleles were measured to identify
SNP/allele combinations that might increase
inhibitor risk. Weak or no binding was
observed in 85% of these assays. Among
subjects with combinations that did
bind, .50% had not developed inhibitors.
Binding was far less frequent than predicted
by computer algorithms. Third, cultured
CD4 T cells from a small number of patients
infused with mismatched products were
examined by tetramer-guided epitope
mapping to determine reactivity with FVIII
peptides containing the ns-SNP sequences.
Using methods that have successfully
demonstrated T-cell epitopes in mild hemophilia
patients with high-risk mutations resulting in
single amino acid changes, they found no highavidity binding. The authors conclude that the
small number of patients potentially reactive
to the neoepitopes presented by mismatched
products could not account for the high inhibitor
rate seen in African Americans.
If FVIII mismatch is not the answer,
where do we go from here? Risk factors for
development of inhibitors are complex and
interrelated (see figure). The causative gene
mutation is the primary determinant of
inhibitor risk, controlling whether the gene
produces a product, and, if so, how different
that product is from the normal protein.
More than 2500 unique mutations causing
HA have been reported.9 This heterogeneity
makes inclusion of mutation in risk factor
analysis problematic. African American
HA patients have not been found to have
differences from white Americans in the type
830
and frequency of mutations,4,5 but mutation
type has not been included in all analyses.
The use of patient groups with the common
intron-22 inversion to control for mutation
presents an interesting conundrum. Studies
have now shown that the inverted gene does
produce 2 products, which include ns-SNPs
and remain intracellular; they may result in
immune tolerance.10 The uniformity of these
products across all intron-22-inversion
patients has yet to be demonstrated.
Study of immune response genes is
similarly daunting, although it presents
perhaps the most likely area for identification
of racial differences. Study of 13 331 SNPs
in 833 subjects yielded 13 candidate genes for
further investigation.11 This large population,
however, included only 48 black subjects.
Larger numbers of black patients and
Hispanics, who also have increased inhibitor
risk,2,3 will be required to assess whether
their immune risk factors differ from those
in whites. Functional studies of the type
conducted by Gunasekera et al1 will be
necessary to evaluate the validity of any
genetic risk factors identified.
Product exposure as a risk factor is an
area of intense interest. Inhibitor risk is
increased during the first few exposures in
patients with severe hemophilia, although
patients with mild or moderate disease may
have a lifelong risk. Inhibitors may be triggered
by intense treatment episodes, inflammation,
or other challenges to the immune system
during therapy. Product structure differences
and product switching remain areas of concern.
The problem in risk factor analysis is
documenting and quantifying exposure.
With US reliance on multiple treatment
products, it is difficult to identify patients
who have received only a single factor product,
other than previously untreated patients,
who number ,400 affected neonates per year.
Assessment of the immunogenicity of
individual products or treatment methods
requires large numbers. Fewer than 1500
black and 1500 Hispanic patients have been
enrolled in surveillance programs in the
United States, and recruitment of sufficient
numbers for studies has been difficult.
Indeed, we have hypothesized that the
increased numbers of inhibitors seen
may result from attendance at hemophilia
treatment centers and study enrollment
by only the more severely affected inhibitor
patients in these population groups.5
An assessment of all 3 components of risk
is required to provide an individual patient
with treatment designed to minimize the
chance of an inhibitor occurring. Very large
studies, perhaps conducted internationally,
will be required to quantify these risks
and to assess the unique characteristics
of population subgroups such as
blacks and Hispanics that result in their
disproportionate risk.
Conflict-of-interest disclosure: The author
declares no competing financial interests. n
REFERENCES
1. Gunasekera D, Ettinger RA, Nakaya Fletcher S, et al.
Factor VIII gene variants and inhibitor risk in African American
hemophilia A patients. Blood. 2015;126(7):895-904.
2. Aledort LM, Dimichele DM. Inhibitors occur more
frequently in African-American and Latino haemophiliacs.
Haemophilia. 1998;4(1):68.
3. Carpenter SL, Soucie JM, Sterner S, Presley R;
Hemophilia Treatment Center Network (HTCN)
Investigators. Increased prevalence of inhibitors in Hispanic
patients with severe haemophilia A enrolled in the
Universal Data Collection database. Haemophilia. 2012;18(3):
e260-e265.
4. Viel KR, Ameri A, Abshire TC, et al. Inhibitors of
factor VIII in black patients with hemophilia. N Engl J
Med. 2009;360(16):1618-1627.
5. Miller CH, Benson J, Ellingsen D, et al; Hemophilia
Inhibitor Research Study Investigators. F8 and F9 mutations
in US haemophilia patients: correlation with history of
inhibitor and race/ethnicity. Haemophilia. 2012;18(3):375-382.
6. Schwarz J, Astermark J, Menius ED, et al; Hemophilia
Inhibitor Genetics Study Combined Cohort. F8 haplotype
and inhibitor risk: results from the Hemophilia Inhibitor
Genetics Study (HIGS) Combined Cohort. Haemophilia.
2013;19(1):113-118.
7. Lochan A, Macaulay S, Chen WC, Mahlangu JN,
Krause A. Genetic factors influencing inhibitor
development in a cohort of South African haemophilia
A patients. Haemophilia. 2014;20(5):687-692.
8. Lozier JN, Rosenberg PS, Goedert JJ, Menashe I.
A case-control study reveals immunoregulatory gene
haplotypes that influence inhibitor risk in severe
haemophilia A. Haemophilia. 2011;17(4):641-649.
9. Centers for Disease Control and Prevention. CDC
Hemophilia A Mutation Project (CHAMP). Available at: http://
www.cdc.gov/hemophiliamutations. Accessed February 5, 2015.
10. Pandey GS, Yanover C, Miller-Jenkins LM,
et al; PATH (Personalized Alternative Therapies for
Hemophilia) Study Investigators. Endogenous factor
VIII synthesis from the intron 22-inverted F8 locus
may modulate the immunogenicity of replacement therapy
for hemophilia A. Nat Med. 2013;19(10):1318-1324.
11. Astermark J, Donfield SM, Gomperts ED, et al;
Hemophilia Inhibitor Genetics Study (HIGS) Combined
Cohort. The polygenic nature of inhibitors in hemophilia
A: results from the Hemophilia Inhibitor Genetics
Study (HIGS) Combined Cohort. Blood. 2013;121(8):
1446-1454.
DOI 10.1182/blood-2015-02-625574
BLOOD, 13 AUGUST 2015 x VOLUME 126, NUMBER 7
From www.bloodjournal.org by guest on July 31, 2017. For personal use only.
2015 126: 829-830
doi:10.1182/blood-2015-02-625574
Game, set, match for factor VIII mismatch?
Connie H. Miller
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