Major differences in bleeding symptoms between factor VII deficiency and hemophilia B
ABSTRACT Background: The autosomally-inherited factor VII (FVII) deficiency and X-linked hemophilia B offer an attractive model to investigate whether reduced levels of FVII and FIX, acting in the initiation and amplification of coagulation respectively, influence hemostasis to a different extent in relation to age and bleeding site. Methods: Hemophilia B patients (n = 296) and FVII-deficient males (n = 109) were compared for FVII/FIX clotting activity, F7/F9 genotypes and clinical phenotypes in a retrospective, multi-centre, cohort study. Results: Major clinical differences between diseases were observed. Bleeding occurred earlier in hemophilia B (median age 2.0 years, IR 0.9–5.0) than in FVII deficiency (5.2 years, IR 1.9–15.5) and the bleeding-free survival in FVII deficiency was similar to that observed in ‘mild’ hemophilia B (P = 0.96). The most frequent disease-presenting symptoms in hemophilia B (hematomas and oral bleeding) differed from those in FVII deficiency (epistaxis and central nervous system bleeding). Differences were confirmed by analysis of FVII-deficient women. Conclusions: Our data support the notion that low FVII levels sustain hemostasis better than similarly reduced FIX levels. On the other hand, minute amounts of FVII, differently to FIX, are needed to prevent fatal bleeding, as indicated by the rarity of null mutations and the associated life-threatening symptoms in FVII deficiency, which contributes towards shaping clinical differences between diseases in the lowest factor level range. Differences between diseases are only partially explained by mutational patterns and could pertain to the specific roles of FVII and FIX in coagulation phases and to vascular bed-specific components.
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ABSTRACT: Individuals with inherited factor VII (FVII) deficiency display bleeding phenotypes ranging from mild to severe, with 30% of patients having always been asymptomatic (non-bleeding). In 626 FVII-deficient individuals, by analysing data from the International Factor VII (IF7) Registry and the Seven Treatment Evaluation Registry (STER), we determined whether bleeding type at disease presentation and FVII coagulant activity (FVIIc) predict ensuing bleeds. At disease presentation/diagnosis, 272 (43.5%) individuals were non-bleeding, 277 (44.2%) had minor bleeds, and 77 (12.3%) had major bleeds. During a median nine-year index period (IP) observation, 87.9% of non-bleeding individuals at presentation remained asymptomatic, 75.1% of minor-bleeders had new minor bleeds, and 83.1% of major-bleeders experienced new major bleeds. After adjusting for FVIIc levels and other clinical and demographic variables, the relative risk (RR) for ensuing bleedings during the IP was 6.02 (p <0.001) and 5.87 (p <0.001) in individuals presenting with major and minor bleeds, respectively. Conversely, compared to non-bleeding individuals, a 10.95 (p = 0.001) and 28.21 (p <0.001) RR for major bleedings during the IP was found in those with minor and with major bleeds at presentation, respectively. In conclusion, in FVII deficiency, the first bleeding symptom is an independent predictor of the risk of subsequent major bleeds.Thrombosis and Haemostasis 03/2013; 109(6). · 5.76 Impact Factor
- British Journal of Haematology 08/2014; · 4.94 Impact Factor
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ABSTRACT: Bleeding symptoms are frequently reported even in otherwise healthy subjects, and differentiating a normal subject from a patient with a mild bleeding disorder (MBD) can be extremely challenging. The concept of bleeding rate, that is, the number of bleeding episodes occurring within a definite time, could be used as the unifying framework reconciling the bleeding risk observed in congenital and acquired coagulopathies into a single picture. For instance, primary prevention trials have shown that the incidence of non-major bleeding symptoms in normal subjects is around five per 100 person-years, and this figure is in accordance with the number of hemorrhagic symptoms reported by normal controls in observational studies on hemorrhagic disorders. The incidence of non-major bleeding in patients with MBDs (e.g. in patients with type 1 VWD carrying the C1130F mutation) is also strikingly similar with that of patients taking antiplatelet drugs, and the incidence in moderately severe bleeding disorders (e.g. type 2 VWD) parallels that of patients taking vitamin K antagonists. The severity of a bleeding disorder may therefore be explained by a bleeding rate model, which also explains several common clinical observations. Appreciation of the bleeding rate of congenital and acquired conditions and of its environmental/genetic modifiers into a single framework will possibly allow the development of better prediction tools in the coming years and represents a major scientific effort to be pursued.Journal of Thrombosis and Haemostasis 06/2013; 11(s1). · 6.08 Impact Factor
Major differences in bleeding symptoms between factor VII
deficiency and hemophilia B
F. BERNARDI,* A. DOLCE,? M. PINOTTI,* A. D. SHAPIRO,? E. SANTAGOSTINO,§ F. PEYVANDI,§
A. BATOROVA,– M. LAPECORELLA,** J. F. SCHVED,?? J. INGERSLEV?? and G. MARIANI** FOR THE
INTERNATIONAL FACTOR VII DEFICIENCY STUDY GROUP
*Department of Biochemistry and Molecular Biology, University of Ferrara, Ferrara; ?National Institute of Statistics, Rome, Italy; ?Indiana
Hemophilia and Thrombosis Center, Indianapolis, IN, USA; §Hemophilia and Thrombosis Center, IRCCS Maggiore Hospital, Mangiagalli and
Regina Elena Foundation and University of Milan, Milan, Italy; –The National Hemophilia Center, Institute of Hematology and Blood Transfusion,
University Hospital, Bratislava, Slovakia; **Internal Medicine & Hematology, Department of Internal Medicine and Public Health, L? Aquila
University, L? Aquila, Italy; ??Hemophilia Centre, CHU Montpellier, France; and ??Centre for Hemophilia and Thrombosis, Aarhus University
Hospital, Skejby, Aarhus N, Denmark
To cite this article: Bernardi F, Dolce A, Pinotti M, Shapiro AD, Santagostino E, Peyvandi F, Batorova A, Lapecorella M, Schved JF, Ingerslev J,
Mariani G, for the International Factor VII Deficiency Study Group. Major differences in bleeding symptoms between factor VII deficiency and
hemophilia B. J Thromb Haemost 2009; 7: 774–9.
Summary. Background: The autosomally-inherited factor VII
model to investigate whether reduced levels of FVII and FIX,
acting in the initiation and amplification of coagulation
to age and bleeding site. Methods: Hemophilia B patients
(n = 296) and FVII-deficient males (n = 109) were compared
for FVII/FIX clotting activity, F7/F9 genotypes and clinical
phenotypes in a retrospective, multi-centre, cohort study.
Results: Major clinical differences between diseases were
observed. Bleeding occurred earlier in hemophilia B (median
age2.0 years,IR0.9–5.0)thaninFVIIdeficiency(5.2 years,IR
1.9–15.5) and the bleeding-free survival in FVII deficiency was
similartothatobservedin?mild?hemophiliaB(P = 0.96).The
most frequent disease-presenting symptoms in hemophilia B
(hematomas and oral bleeding) differed from those in FVII
deficiency (epistaxis and central nervous system bleeding).
Differences were confirmed by analysis of FVII-deficient
women. Conclusions: Our data support the notion that low
FVII levels sustain hemostasis better than similarly reduced
FIX levels. On the other hand, minute amounts of FVII,
differently to FIX, are needed to prevent fatal bleeding, as
indicatedby therarity ofnull mutationsandtheassociatedlife-
threatening symptoms in FVII deficiency, which contributes
towards shaping clinical differences between diseases in the
lowest factor level range. Differences between diseases are only
partially explained by mutational patterns and could pertain to
thespecific rolesof FVIIandFIX incoagulation phases and to
vascular bed-specific components.
Keywords: bleeding, disease-presenting symptoms, FVII defi-
ciency, hemophilia B.
Knowledge of blood coagulation has substantially increased
during the last decades [1,2], and fostered dramatic improve-
ment in the management of inherited bleeding disorders [3–6]
as well as innovative approaches for gene therapy [7–9].
However, comparative analysis of bleeding symptoms among
hemorrhagic disorders remains fragmentary [10–12], and has
been virtually produced only for X-linked hemophilia A and B
, both caused by deficiency of factors acting as cofactor
(FVIII) and enzyme (FIX) in the amplification phase of
The clinical features of hemophilia  have previously been
compared with those of the recessively inherited coagulation
disorders, pulled together because of their rarity. However, this
approach did not allow weighting the differential effect of
reduced levels of each clotting factor on the clinical phenotype
of the corresponding deficiency.
This information is important for bridging genetic and
biochemical findings with the clinical aspects of these
disorders, which would provide models for molecular and
cellular studies focused on clinical observations and manage-
The F7 and F9 genes are characterized by very similar
size and organization [14,15], and encode for coagulation
and amplification phases of coagulation , respectively.
Increased FVII and FIX levels have been associated with
Correspondence: Francesco Bernardi, Department of Biochemistry
and Molecular Biology, University of Ferrara, Via Fossato di Mortara
74, 44100 Ferrara, Italy.
Tel.: +39 0532 974425; fax: +39 0532 974484.
Received 13 November 2008, accepted 7 February 2009
Journal of Thrombosis and Haemostasis, 7: 774–779DOI: 10.1111/j.1538-7836.2009.03329.x
? 2009 International Society on Thrombosis and Haemostasis
the risk of myocardial infarction  and venous thrombosis
[17,18], respectively, which supports vascular bed-specific
alteration of hemostasis . Local regulation of coagulation
might also explain, in inherited coagulopathies, the prepon-
derance of hemorrhagic manifestations in specific tissues/
Detailed mutational patterns have been reported in auto-
socmally-inherited FVII deficiency [20,21], and X-linked
database.html)  but qualitative and quantitative compari-
son of their clinical features with reference to coagulation
phenotypes is not available.
Here, we have evaluated cohorts of patients virtually
covering the complete clinical and biochemical spectrum of
hemophilia B and FVII deficiency to unravel whether
reduced levels of FVII and FIX influence hemostasis to a
different extent, particularly in relation to age and bleeding
Genotype and phenotype were compared in 296 hemophilia B
and 109 FVII-deficient patients, all of them males. Data from
114 females with FVII deficiency were also available.
The age distribution in hemophilia B patients (median age
32.4, interquartile range 13.2–50.1) did not differ (P = 0.19)
from that in FVII-deficient males (median age 29, interquartile
The review boards of the participating institutions approved
the research protocol. A common form for the patients?
informed consent and study information was developed and
used. Data collection was carried out by using specifically
developed questionnaires (see Appendix S1, supporting mate-
rial). Questionnaires were validated beforehand by examina-
tion of records from four centres from the IRF7 Study Group.
Attending physicians in all centres were asked to fill in blank
files, and individual subject files were reviewed and afterwards
validated by an internal adjudication committee (GM, BF,
AD). Twenty-two records, not suitable for analysis, were
Thirty-seven hemophilia centers from Europe, Asia and the
Americas contributed data for this analysis (see Appendix S2,
The questionnaire-derived data base was completed with
information about genotypes, most of which were determined
in the centre registering the patients or in national reference
centres. F7 and F9 gene mutations were investigated by DNA
sequencing as previously described [21,23]. The results of
mutation analysis were available for 74 males with FVII
deficiency (41 homozygotes and 33 double heterozygotes) and
177 hemophilia B patients.
FIX and FVII activity levels were assayed using standard
methods [21,24] in the laboratory of each participating
hemophilia centre (declared lowest detection limit, 1% of
After testing the normal distribution of the main variables
(Kolmogorov-Smirnov test), the non-normal distribution of
values was described by ordinal statistics (median and inter-
quartile range [IR]) and reported as histograms and box-and-
whisker plots. Kaplan Meier curves (KM) were used to
compare the age at disease-presenting symptom, and non-
parametric statistical analyses (Kruskall–Wallis test) to com-
pare the median values of the main variables. Differences in
the KM curves were evaluated by the Log-rank test. Values
exceeding the upper quartile plus three times the inter-
quartile range were defined as outliers in accordance with
Tukey . All reported P values are two-sided. Analyses
were carried out using the SAS/STAT?Software and S-PLUS
2000 (SAS Institute Inc., Cary, NC, USA).
To estimate the detrimental effect of missense mutations we
exploited the SIFT (Sorting Intolerant From Tolerant) pro-
gram available at the http://blocks.fhcrc.org/sift/ . This
computational tool presumes that important amino acids will
be conserved in the protein family. Changes at well-conserved
positions tend to be predicted as deleterious as indicated by
scores close to zero.
The main coagulation phenotype and genotype features in
hemophilia B and FVII-deficient patients are described in
Patients were classified in accordance with the plasma FVII/
FIX levels using intervals previously indicated for hemophilias
[27,28]. Beside the ?severe? and ?moderate? deficiency classes (£1
and >1–5%) we defined a third category for ?mild? deficiency
of findings of uncertain significance in the assessment of
coagulation/clinical phenotype relationships. The upper limit
of the third class (20%) virtually excludes heterozygotes from
the cohort of FVII-deficient subjects, as confirmed by geno-
typed patients, who were homozygous (n = 41) or doubly
heterozygous (n = 33) for FVII gene mutations.
The distribution of symptomatic subjects within level classes
was similar in FVII deficiency and hemophilia B. The lowest
(110/114, 96.5%) and in FVII (30/33, 90.9%) deficiency by a
high proportion of symptomatic subjects.
Mutation pattern in FVII deficiency differed (P < 0.001)
(nonsense changes, gene insertion/deletions) were rare in FVII
deficiency, particularly in the homozygous condition (1 FVII
Bleeding symptoms in HB and FVII deficiency 775
? 2009 International Society on Thrombosis and Haemostasis
deficient vs. 23 hemophilia B patients). Conversely, splicing
mutations, which are usually compatible with very reduced but
detectable factor levels, were more represented in FVII
Missense changes were found to be by far the most frequent
cause of both diseases, and represented 78% and 85% of the
affected alleles in FVII deficiency and hemophilia B, respec-
tively. The degree of severity of aminoacid substitutions in
FVII and FIX proteins was predicted by a sequence-homol-
ogy-based tool, the SIFT program. Although the results of this
analysis have to be cautiously considered, the lowest probabil-
ity of being tolerated in theproteinstructure (score<0.01) was
assigned to 70% of the changes found in hemophilia B as
compared with 50% in FVII deficiency.
The study of FVII gene polymorphisms able to modulate
FVII levels  did not show appreciable influence on FVII
levels of patients homozygous or doubly heterozygous for
Symptoms related to bleeding (Fig. 1) were analyzed in two
different sets: (i) all bleeds contributing to the overall clinical
status (surgery-related included) and (ii) disease-presenting
bleeding symptoms, evaluated with reference to the residual
plasma factor level classes. The incidence of the most severe
manifestations in hemophilia B (hemarthrosis, hematomas)
and in FVII deficiency (CNS and GI bleeding, hemarthrosis)
deficiencies within the lowest class was purposely avoided
because the evaluation of very low levels requires sensitive
approaches [30,31] rather than standard coagulation labora-
The median age of the first bleeding symptom was 2.0 years
(IR 0.9–5.0) for hemophilia B and 5.2 years (IR 1.9–15.5) for
FVII deficiency (P < 0.001). For patients with plasma levels
>5–20% the median age values were 6 (IR 2.0–15.0) and 6.75
(IR 3.2–11.4) years, respectively.
Table 1 Coagulation and clinical phenotypes, and mutation type distribution in FVII deficiency and hemophilia B
*The allelic frequencies of null, missense and splicing mutations are shown. Null mutations include nonsense mutations, gene deletions and
insertions causing frameshift.
Fig. 1. Clinical features and factor level classes in FVII deficiency and
also reported in relation to factor level classes. CNS, central nervous
system; GI, gastro-intestinal.
776 F. Bernardi et al
? 2009 International Society on Thrombosis and Haemostasis
The bleeding-free survival analysis was conducted in both
diseases (Fig. 2). Surgery-related hemorrhages (Fig. 1, ?sur-
gery?) were not included in this analysis because they do not
provide affordable information about the spontaneous bleed-
ing tendency, a main focus of our study. The bleeding-free
survival in hemophilia B differed significantly (P < 0.001)
from that in FVII deficiency. Further, the curve observed for
hemophilia B patients belonging to the highest FIX level class
(>5–20%) was indistinguishable from that observed in FVII-
deficient patients (Fig. 2, P = 0.96).
severity of the diseases at presentation, the age distribution of
the most frequent disease-presenting symptoms was analysed
and is graphically shown in Fig. 2. CNS hemorrhage threatens
the life of unweaned babies affected by FVII gene mutations
(nonsense mutations and changes affecting the conserved GT
donor splice site) causing barely detectable residual levels.
Differently, epistaxis characterizes the bleeding phenotype in
the first decade of life, particularly in ?moderate? and ?mild?
Hematomas and oral bleeding, but not CNS and GI
bleeding, are main clinical findings in hemophilia B, particu-
larly in the first years of life. Joint bleeding, present in
approximately half of hemophilia B patients, was shown to
occur at a later age after hematomas and oral bleeding, when
children start to crawl and walk, resulting less frequently as a
disease-heralding symptom. Worthy of note is that this
symptom occurred at an earlier age in hemophilia B than in
Hematuria was not found as a presenting symptom in either
Our analysis is based on the incidence of disease-presenting
symptoms, which are not influenced by treatment or prophy-
laxis and thus suitable for highlighting the differences between
diseases and disease subsets. This approach has not been
previously used to compare, neither collectively nor separately,
therarebleedingdisorders withhemophilias.Major differences
were observed in the clinical spectrum of FVII deficiency and
hemophilia B, both in the disease-presenting and overall
The clinical picture was more polymorphic in FVII defi-
ciency than in hemophilia B.
the preponderance of epistaxes and hematomas as first
symptoms in FVII deficiency and hemophilia B, respectively,
support the presence of tissue and vascular bed specific
components interacting with the regulation of coagulation
phases, eventually contributing to local hemostasis. Neverthe-
less, this analysis revealed a strong influence of the coagulation
phenotype in determining the earliest and/or most severe
bleeding phenotype in both diseases.
The bleeding-free survival analysis, particularly useful for
comparing bleeding syndromes, has been used to infer that
the influence of prothrombotic risk factors in children with
severe hemophilia A . Our analysis (Fig. 2) indicates that
B compared with FVII deficiency, even when the same
symptom is matched. Roughly half of the patients with
hemophilia B have experienced an episode of hemarthrosis
within the sixth year of age, whereas in FVII deficiency the first
hemarthrosis may occur later, even in teenagers. The curve
characterizing the vast majority of FVII-deficient males,
including a high proportion of patients with markedly reduced
FVII activity, becomes indistinguishable from that of the
milder forms of hemophilia B (>5–20% FIX levels). The
relatively late onset of bleeding symptoms in FVII deficiency
was further confirmed in females with FVII deficiency, in
whom the median age of the first bleeding symptom was
8.5 years (IR 3.8–21.5). Taken together these data support the
theory that low factor levels in FVII deficiency and hemophilia
B produce differential clinical effects, and suggest that reduced
levels of FVII are still able to ensure hemostasis for a more
extended lifespan. One would argue that higher clotting levels
of FIX, acting in the amplification of the coagulation cascade,
are required to delay the onset of the disease or even to prevent
bleeding in comparison with FVII, initiating the entire process.
The relative physiological concentrations of these proteins in
Fig. 2. Bleeding-free survival analysis in FVII deficiency and hemophilia
B. Bleeding-free survival curves (Kaplan?Meier analysis) in FVII defi-
B patients characterized by FIX levels >5?20%. The age distribution of
the most frequent presenting symptoms in all FVII-deficient and hemo-
philia B patients is shown in the lower part of the figure. The dots indicate
values exceeding the upper quartile plus three times the inter-quartile
range. CNS, central nervous system; GI, gastro-intestinal.
Bleeding symptoms in HB and FVII deficiency 777
? 2009 International Society on Thrombosis and Haemostasis
plasma, and the ex vivo observations indicating that even low
FVII levels may ensure an appreciable thrombin generation
, are consistent with the observed differences.
However, a small subset of patients with FVII deficiency
was characterized by a very precocious and severe bleeding
tendency, which shapes the bleeding-free survival curve in the
first year of life, and could make it even steeper in FVII
deficiency than in severe hemophilia B. The observed
association of neonatal life-threatening symptoms (i.e. CNS)
with the most detrimental (nonsense and +1 splice site)
mutations indicates that minute amounts of FVII are needed
to initiate coagulation and prevent fatal bleeding. This
hypothesis is further supported by (i) the rarity in FVII
deficiency, in our study as well as in other data bases , of
potentially null mutations and the absence of homozygous
insertion/deletions predicting the complete absence of circu-
lating protein and (ii) observations in animal models indicat-
ing the occurrence of fatal perinatal bleeding in FVII but not
FIX knockout mice [34,35]. Indeed, mutation pattern differ-
ences contribute towards shaping phenotype differences
between FVII deficiency and hemophilia B in the lowest
factor level range.
Several clinically relevant observations, stemming from the
comparative evaluation of disease-presenting bleeding symp-
toms and of their age distribution, provide elements to design
therapeutic strategies, with some differences between diseases.
First, the very early occurrence of severe and life-endangering
long-term prophylactic replacement therapy in a relatively
small but clinically relevant subset of children (10%) with FVII
deficiency. Instead, those FVII-deficient patients with a less
severe clinical picture (epistaxis, easy bruising), mimicking that
of a ?platelet disorder?, may be treated on demand.
In hemophilia B, hematomas and oral cavity hemorrhages
may require an early replacement therapy strategy, even based
on prophylaxis, especially if the disease manifests itself with
early bleeding into joints/muscles.
multicentre studies may be biased by differential efficiency in
reporting, but there is no alternative to this approach in rare
disorders. Secondly, some symptoms have been arbitrarily
pooled (i.e. gum bleeding, traumatic gum bleeding, bleeding
following dental eruption and oral bleeding have been
collapsed for analysis as oral bleeding, and as such reported
in the paper); the same holds for hematomas in the limbs as
imaging indicating the tissue involved (subcutaneous, muscle)
was seldom available. However, the presenting symptom
category contains only first symptoms. Thirdly, we do not
present information about mutational patterns in one-third of
patients; however, this does not seem to affect the analysis and
conclusion of the study.
In conclusion, our observations indicate that major clinical
differences between hemophilia B and FVII deficiency are only
partially explained by their mutational patterns, and may
pertain to the specific roles of FVII and FIX in the coagulation
cascade. The data stemming from this comparative analysis
bleeding disorders, to implement disease-suited therapeutic
strategies and to identify subsets of patients with different
F. Bernardi, A. Dolce, M. Pinotti and G. Mariani conceived
and designed the study. F. Bernardi, A. Dolce, M. Pinotti, J.
Ingerslev and G. Mariani analysed and interpreted the data. A.
Dolce performed statistical analysis. F. Bernardi, A. Dolce, M.
Pinotti and G. Mariani drafted the manuscript, and conceived
and designed the study. F. Bernardi, A. Dolce, M. Pinotti, F.
Peyvandi, E. Santagostino, J. Ingerslev and G. Mariani
critically revised the article for important intellectual content.
A.D. Shapiro, E. Santagostino, A. Batorova, M. La Pecorella,
J.F. Schved and G. Mariani collected and assembled the data
from patients. All authors have read and approved the final
version of the manuscript.
This work was supported by a grant from Telethon (GGP
05214) (FB) and Fondazione CARIFE (MP). The authors
acknowledge 37 centers contributing FVII-deficient and hemo-
philia B patients (see Appendix S2, supporting material).
We thank D. Etro (University of Ferrara) for her helpful
Disclosure of conflict of interests
The authors state that they have no conflict of interest.
version of this article:
Appendix S1. Enrolment form used to collect information
about FVII/FIX deficient patients.
or functionality of any supporting materials supplied by the
authors. Any queries (other than missing material) should be
directed to the corresponding author for the article.
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