Case report 651
A novel type 2N VWF gene mutation: a case report
Matthew S. Evansa and M. Elaine Eysterb
Men and boys who present with bleeding associated with low
factor VIII levels and normal von Willebrand studies are
assumed to have hemophilia A until proven otherwise.
However, routinely available coagulation assays cannot
easily distinguish mild hemophilia A from the 2N variant of
von Willebrand disease. We present a case that highlights the
difficulties of recognizing this diagnosis, the role of genetic
testing, and the identification of a 2N variant that has not been
previously described. Blood Coagul Fibrinolysis 29:651–652
Copyright ß 2018 Wolters Kluwer Health, Inc. All rights
reserved.
Blood Coagulation and Fibrinolysis 2018, 29:651–652
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Introduction
Men and boys who present with bleeding associated with
low factor VIII (FVIII) levels, and normal von Willebrand
studies are assumed to have hemophilia A until proven
otherwise. However, routinely available coagulation
assays cannot easily distinguish mild hemophilia A from
the 2N variant of von Willebrand disease (VWD). We
present a case that highlights the difficulties of recogniz-
ing this diagnosis, the role of genetic testing, and the
identification of a 2N variant that has not been
previously described.
Case report
In 1988, a 5-year-old boy was diagnosed with mild hemo-
philia A after presenting with epistaxis and soft tissue
bleeding following minor trauma. Initial evaluation
revealed a FVIII activity (FVIII:C) of 8% with normal
von Willebrand studies including a von Willebrand factor
antigen (VWF:Ag) of 80%, Ristocetin cofactor
(VWF:RCo) of 60%, and a normal pattern of von Will-
ebrand factor (VWF) multimer. Supporting a diagnosis of
hemophilia A was a family history of a younger brother
who was diagnosed with mild HA with similar FVIII:C,
VWF:Ag, and VWF:RCo results, and a sister with normal
VWF levels and a FVIII:C of 44% thought to be indica-
tive of a hemophilia A carrier. No other family members
were known to have a bleeding disorder.
At age 31, he underwent an open reduction and internal
fixation of a finger fracture. A preoperative inhibitor
screen was negative. FVIII:C recovery studies at time
of surgery revealed a suboptimal peak response to recom-
binant FVIII (rFVIII), with 8-h trough levels returning to
his baseline level. Unfortunately, he required prolonged
treatment with rFVIII and had a poor outcome with pain
and a residual deformity of his finger. In retrospect, many
0957-5235 Copyright ß 2018 Wolters Kluwer Health, Inc. All rights reserved.
Copyright © 2018 Wolters Kluwer Health, Inc. All rights reserved.
Keywords: blood coagulation disorder, genotype, type 2N von Willebrand
disease
a
Department of Medicine, Division of Hematology/Oncology, Hemophilia
Treatment Center of Central Pennsylvania, Penn State Health Milton S. Hershey
Medical Center and bDistinguished Professor, Penn State Health-Milton S.
Hershey Medical Center, Department of Medicine, Division of Hematology/
Oncology, Pennsylvania. USA
Correspondence to Matthew S. Evans, MD, Assistant Professor, Penn State
Health-Milton S. Hershey Medical Center, Department of Medicine, Division of
Hematology/Oncology, Hershey, PA 17033, USA.
Tel: +1 717 531 0003 ext 28 3405; fax: +1 717 531 0647;
e-mail: mevans1@pennstatehealth.psu.edu
Received 16 March 2018 Revised 7 June 2018
Accepted 5 July 2018
of his bleeding events as a child and early adulthood were
hematomas that required an extended duration of rFVIII
due to slow resolution of pain and swelling.
In 2014, at age 31, he enrolled in the My Life, Our Future
study for patients with hemophilia A. Genotyping by
Bloodworks Northwest at the Puget Sound Blood Center
revealed no F8 variant using Next Gen screening, by
Sanger sequence screening of all 26 exons of his F8 gene,
and by a multiplex ligation-dependent probe affinity
assay (MLPA-1) assay for large gene duplications or
deletions. Subsequently, a VWF:FVIII binding assay
revealed decreased binding of FVIII to VWF (ratio of
0.19), suggesting a diagnosis of type 2N VWD. Nucleo-
tide sequencing of exons 18–20 of VWF gene through
Quest Diagnostics revealed a homozygous variant
c2390T>G in exon 18 leading to a pLeu797Arg substi-
tution. This specific variant is predicted to be deleterious,
but has not been previously reported in the von Will-
ebrand factor variant database.
A repeat finger surgery was performed with a plasma
derived VWF:FVIII concentrate with excellent FVIII
recovery and improved surgical outcome.
Discussion
Factor VIII clotting factor circulates as an inactive plasma
protein tightly associated with VWF [1,2]. In response to
blood vessel injury, FVIII is cleaved between its A1 and
A2 domains, resulting in an unstable heterotrimeric
FVIIIa molecule which interacts with factor IX in a
subsequent, well characterized series of reactions that
result in blood clot formation [3]. VWF is a central
component of this hemostatic process protecting FVIII
from proteolytic degradation, and also an adhesive link
between platelets and injured blood vessels. In patients
DOI:10.1097/MBC.0000000000000761
652 Blood Coagulation and Fibrinolysis 2018, Vol 29 No 7
with VWD, the half-life of endogenous or infused FVIII
is shortened leading to bleeding complications.
Type 2N VWD is an uncommon disorder characterized
by missense mutations in the FVIII-binding domain at
the N-terminus of VWF leading to defective FVIII–
VWF binding. This leads to instability of FVIII, resulting
in low FVIII plasma levels. The decrease in FVIII in
the circulation causes a phenotype that is very similar to
mild hemophilia A with isolated FVIII deficiency in the
presence of normal VWF antigen, activity, and multimer
studies.
Type 2N VWD was first described in 1990 in a woman
from Normandy, France, with a mild bleeding disorder,
who was eventually found to have a missense mutation
(2372C>T causing a Thr791Met change) in the FVIII
binding region of the VWD gene [4]. Since then, 36
missense mutations have been reported in EAHAD Coag
factor variant database as of December 2017. The major-
ity of mutations are missense mutations, most commonly
2561G>A(Thr791Met) and 2446C>T (Arg854Gln). Our
patient was initially diagnosed with mild hemophilia A in
1988, two years prior to the description of 2N VWD.
Ultimately, gene sequencing identified a homozygous
variant c2390T>G in exon 18 leading to a pLeu797Arg
substitution. Although this particular mutation has not
been previously described, given the location in exon 18,
it correlates to a mutation in his VWF FVIII binding site,
predicted to be deleterious and consistent with a diagno-
sis of type 2N VWD.
Phenotypically, the bleeding patterns between mild
hemophilia A and type 2N VWD can be very similar,
making this a challenging diagnostic dilemma. The diffi-
culty in recognizing this disorder was highlighted by a
recent study carried out by the CDC Hemophilia Inhibi-
tor Research Group that revealed 37 of 1027 (3%) patients
previously identified as having mild hemophilia A lacked
hemophilia mutations [5]. Four of those 37 patients had
VWF:FVIII binding assay of less than 80%, consistent
with a diagnosis of type 2N VWD.
From a diagnostic perspective, the 2N binding assay
should be obtained in any person thought to have mild
hemophilia A, who does not have an antecedent pedigree,
clearly indicative of an autosomal inheritance pattern or
who does not respond well to recombinant FVIII in the
absence of a FVIII inhibitor. If VWF:FVIII binding is
decreased, targeted sequence analysis of VWF exons 17–
21 and 24–27 should be performed to demonstrate a
mutation that correlates to the FVIII binding domain.
Additionally, women who have FVIII levels below the
normal range should not be assumed to be carriers of
hemophilia A until proven by genetic testing. In fact,
with increasing availability of next-generation (NextGen)
sequencing, sequencing strategies to simultaneously
Copyright © 2018 Wolters Kluwer Health, Inc. All rights reserved.
assess factor 8 and VWF genes have successfully been
shown to delineate between these disorders [6,7] As seen
in this case, it is essential to distinguish type 2N VWD
from mild hemophilia A to ensure that appropriate treat-
ment is given. This is an especially important in patients
who were diagnosed as mild hemophilia A prior to the
recognition of VWF Normandy more than 25 years ago.
Conclusions
Routinely available coagulation assays cannot be relied
upon to distinguish mild hemophilia A from the 2N
variant of VWD. This case highlights the difficulties in
recognizing 2N VWD, which can now be avoided by
utilizing a multigene sequencing panel that analyzes both
hemophilia A and VWF genes simultaneously in the
initial evaluation of patients with a mild FVIII deficiency
who do not have a clear antecedent family history of x-
linked transmission. Additionally, the early identification
of patients with 2N variants should result in a cost savings
realized from the effective treatment and prevention of
bleeding episodes as the cost of NextGen sequencing
decreases.
Furthermore, the previously undescribed deleterious
variant listed in this case provides additional insight into
the molecular mechanisms of type 2N VWD. As addi-
tional causative variants are identified, genotypic corre-
lation with bleeding phenotypes will improve our
understanding of FVIII/VWF interactions and bleeding
risk.
Acknowledgment
Livingston Trout and Mellinger Medical Research Fund.
Conflicts of interest
The authors declare that they have no conflict of interest.
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