T R E A T M E N T O F H E M O P H I L I ADECEMBER 2006 • NO 41

MILD HEMOPHILIA

Sam SchulmanDepartment of MedicineMcMaster UniversityHamilton, Ontario

Published by the World Federation of Hemophilia (WFH)

© World Federation of Hemophilia, 2006

The WFH encourages redistribution of its publications for educational purposes by not-for-profit hemophiliaorganizations. In order to obtain permission to reprint, redistribute, or translate this publication, pleasecontact the Communications Department at the address below.

This publication is accessible from the World Federation of Hemophilia's web site at www.wfh.org.Additional copies are also available from the WFH at:

World Federation of Hemophilia1425 René Lévesque Boulevard West, Suite 1010Montréal, Québec H3G 1T7CANADATel. : (514) 875-7944Fax : (514) 875-8916E-mail: wfh@wfh.orgInternet: www.wfh.org

The Treatment of Hemophilia series is intended to provide general information on the treatment andmanagement of hemophilia. The World Federation of Hemophilia does not engage in the practice ofmedicine and under no circumstances recommends particular treatment for specific individuals. Doseschedules and other treatment regimes are continually revised and new side effects recognized. WFHmakes no representation, express or implied, that drug doses or other treatment recommendations inthis publication are correct. For these reasons it is strongly recommended that individuals seek the adviceof a medical adviser and/or to consult printed instructions provided by the pharmaceutical companybefore administering any of the drugs referred to in this monograph.

Statements and opinions expressed here do not necessarily represent the opinions, policies, or recommendations of the World Federation of Hemophilia, its Executive Committee, or its staff.

Treatment of Hemophilia MonographsSeries EditorDr. Sam Schulman

Table of Contents

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

Classification . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .1

Epidemiology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .1

Life expectancy . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .1

Diagnosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .2

Laboratory diagnosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .2

Differential diagnosis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .2

Molecular basis for mild hemophilia . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .3Hemophilia A . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .3Hemophilia B . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .3

Treatment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4Hemophilia A . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4Hemophilia B . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4Follow-up . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .4

Inhibitors . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .5

Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .5

References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .5

IntroductionIn this monograph I will discuss the mild forms ofhemophilia A and hemophilia B, from epidemiologyand molecular basis, via diagnosis, to the treatmentoptions available. The emphasis is on features thatare characteristic of the mild form. Therefore, forexample, treatment with factor concentrates willonly be briefly mentioned.

ClassificationThe severity of hemophilia has been defined by atraditional classification into three forms:

• Severe form – factor level <0.01 IU/mL (<1% ofnormal);

• Moderate form – factor level 0.01 to 0.05 IU/mL(1 to 5% of normal); and

• Mild form – factor level >0.05 to 0.40 IU/mL(more than 5 to 40% of normal).

This definition has been published by the subcommitteeon Factor VIII and Factor IX of the Scientific andStandardization Committee of the InternationalSociety on Thrombosis and Haemostasis.[1]

However, particularly the upper limit for mildhemophilia is vague and may in different publicationsvary from 0.25 IU/mL (25%) up to 0.50 IU/mL(50%), which is the lower limit of the normal range.The wider the range for definition of mild hemophilia,the greater the proportion of females will be. Forexample, in one study, using the definition “5 to50%” to define mild hemophilia, the proportion offemales was 10 per cent.[2] Females, who are generallycarriers of hemophilia, have the same risk of bleedingas a male with mild hemophilia at the correspondingfactor level.

EpidemiologyThe prevalence of hemophilia in general in countrieswhere diagnostic tools are readily available is about1 in 10,000 people. The proportion of those with themild form of hemophilia varies between countries,over time in the same country, and between the twotypes of hemophilia. This variation to a considerableextent depends on the resources available and theawareness of hemophilia among physicians. In the

World Federation of Hemophilia Annual GlobalSurvey 2004, the proportion of patients diagnosedwith mild hemophilia (34 per cent) was close to theproportion with severe hemophilia (43 per cent) incountries with a gross natural product (GNP) percapita of more than US $10,000. In countries with aGNP below US $2,000 per capita, the proportionwith mild hemophilia (18 per cent) was clearlylower than that of the severe form (50 per cent).

In a Swedish survey of all patients with hemophilia,it was observed that the proportion with knownmild hemophilia increased from 35 per cent in 1960to 54 per cent in 1980.[3] In this case the change wasdue to increased awareness and family investigationsrather than to a change in GNP.

In the Canadian Hemophilia Registry of February 2006(data provided by Dr. Irwin Walker) the proportionof patients with the mild form was 62.3 per cent forhemophilia A and 42.7 per cent for hemophilia B,out of a total population of 3,284 patients.

In small countries, the best example probably beingIceland, the distribution can be extremely skewed.[4]

Life expectancyThe course of the disease is obviously less dramaticin mild compared to severe hemophilia, and the lifeexpectancy is thus very close to that of the normalpopulation. Whereas an impressive improvement inlife expectancy was described in Swedish patientswith severe hemophilia in the period from 1960to1980 (57 years) compared to the period from 1831to 1920 (11 years), no such comparison could bemade for mild hemophilia due to lack of data fromthe early period. However, during the period of1960-1980, life expectancy was 72 years for patientswith mild hemophilia, compared to 75.5 years inthe normal population.[5] This positive picture wastragically changed by the emerging transfusion-transmitted viral infections, which affected patientswith all forms of hemophilia. In the United Kingdom,the death rate during the period from 1977 to 1984of 4 per 1,000 with mild or moderate hemophiliaincreased to 85 per 1,000 in HIV-seropositivepatients in 1991-1992.[6]

Mild HemophiliaSam Schulman

DiagnosisThe diagnosis of mild hemophilia is often made aspart of a family investigation, accounting for examplefor 64 per cent of cases at two centres in the UnitedStates.[2] In the other cases in this study, the diagnosiswas made after one or several bleeding episodesand at a mean age of 5.5 years. In a recent Frenchstudy, the diagnosis of mild hemophilia was madeat an average age of 2.4 years.[7] However, fromtime to time elderly persons will be diagnosed withmild hemophilia, as a result of an investigation trig-gered by bleeding complications after surgery ortooth extraction.

Bleeding is rarely spontaneous in these patients. Infact, in the above-mentioned U.S. investigation, 92per cent of bleeding events were precipitated bytrauma.[2] The type of bleeding was less often inthe joint (30 per cent) than in soft tissues (53 per cent).

Laboratory diagnosisPatients investigated for a bleeding diathesis aretypically first screened for platelet count, bleedingtime, activated partial thromboplastin time (aPTT),and prothrombin time. Only the aPTT may beabnormally prolonged, but this depends of the sen-sitivity of the reagent and on the deficient factorlevel. Standardization of the method is crucial toobtain reliable and reproducible results.[8] Manyhemophilia treatment centre laboratories missed thediagnosis of mild hemophilia using aPTT.[9]

A factor assay must therefore be performed if thereis clinical suspicion, even in the absence of a pro-longed aPTT. Unfortunately, even with a routinefactor VIII (FVIII) assay, the resulting clinical diag-nosis can vary. In a study in the United Kingdomwith plasma samples from three untreated patientswith hemophilia A, distributed to a large number ofcentres, one sample with a median level of 5.8%yielded a range from 1.5% to 36%.[10] The mostaccurate results were obtained from comprehensivecare centres. It is thus important for every diagnosticlaboratory to participate in a quality assurance pro-gram for the assays of factor VIII and IX, and alsoto react and improve their performance if the resultis far from the expected number. The WFH plays anactive part in this respect with support for partici-pation in such a quality assurance scheme for centresparticipating in a WFH-sponsored project, theInternational External Quality Assessment (IEQAS)scheme.

There are additional difficulties in the diagnosis ofmild hemophilia using a factor assay in the case of

2 Mild Hemophilia

certain molecular defects, as described below (see‘Molecular basis for mild hemophilia’).

Differential diagnosisMild hemophilia A has to be differentiated fromcertain forms of von Willebrand disease (vWD).The latter is often also characterized by a reducedFVIII level, but in addition there may be a prolongedbleeding time, a prolonged closure time in the PFA-100 instrument, and of course a reduction of thevon Willebrand factor (vWF) level in plasma, whichcan be measured as antigen, ristocetin co-factor, col-lagen binding activity, FVIII binding capacity, mul-timer formation, etc. The most challenging differentialdiagnosis is the von Willebrand Normandy variant(vWD type 2N).[11] As in mild hemophilia A, theFVIII level is typically between 5% and 30% withouta prolonged bleeding time or reduced plasma vWFlevel and the clinical picture may be similar.

The differences between the two entities are shownbelow:

Obviously, a similar problem can occur if the defectis at the other side of the binding, i.e., on the FVIIImolecule at its vWF-binding site in the C1 domain.In the following table are such variants shownaccording to the mutation and the resulting effecton the binding to vWF:

Mutation in Reduction ofFVIII gene affinity for vWFIle2098Ser 8-foldSer2119Tyr 80-foldArg2150His 3-fold

Mild hemophilia A

X-linked inheritance

Mutation in FVIII geneCapacity of vWF to bindFVIII: normalResponse to DDAVP:good

Response to FVIII concen-trate: good

von Willebrand disease type 2NAutosomal recessiveinheritanceMutation in vWF gene*Capacity of vWF to bindFVIII: reducedResponse to DDAVP:shorter effectResponse to FVIII concen-trate: Good only if vWFpresent in concentrate

*The mutation results in amino acid substitution in theN-terminal part of the vWF molecule, where the FVIIIbinding site is located.

Mild Hemophilia 3

These patients also have reduced secretion of func-tional FVIII and thus by definition they have hemo-philia A.

Another differential diagnosis is combined deficiencyof factor V (FV) and FVIII. Again, the FVIII levelsare as in mild hemophilia A, but the inheritance isautosomal. The defect is neither in the FV gene norin the FVIII gene, but in one of the “chaperone”proteins required for the post-translational processingand cellular secretion of these two structurally similarcoagulation factors. The diagnosis is verified byalso measuring the FV level. Typically, both the FVand FVIII levels are in the 5 to 50% range.

Mild hemophilia B must be differentiated from con-ditions characterized by combined deficiency of thevitamin K dependent factors (in addition to FIX alsofactors VII, X, and prothrombin [II]). This can beacquired due to vitamin K deficiency, liver disease,or the use of vitamin K antagonist drugs such aswarfarin; or it can be congenital due to a mutation inthe gene for γ-glutamyl-carboxylase or for vitamin Kepoxide reductase.

Molecular basis for mild hemophiliaHemophilia AIn a study of 101 patients with mild or moderatehemophilia A, most of whom were unrelated,Schwaab et al described that underlying missensemutations accounted for 86 per cent of the patients.[12]A minority of the patients have a much more pro-nounced defect in FVIII activity than the amount ofcirculating FVIII antigen and this is called CrossReacting Material (CRM) positive. One explanationfor this phenomenon was found in patients withmutations causing a change in the electrical chargeof the A2 domain of the FVIII molecule. This wasresponsible for a more rapid degradation of activeFVIII[13], so that although normal amounts of FVIIIare produced and secreted, the activity is diminished.This mutation also appeared to be associated withdiscrepancies in the one-stage and two-stage clottingassays used for measuring FVIII. A few identifiedmutations and the effect on the assays are listed below:

In certain geographical regions one mutation candominate among patients with mild hemophiliadue to a common founder effect. Such an effect hasbeen described in Iceland.[4] Also, in NorthernItaly, 32 per cent of the patients have a duplicationof exon 13 in the FVIII gene, which results in noactivity of the molecule and should result in severehemophilia. However, due to a phenomenon called“exon skipping,” one of the exon 13 twins is some-times not read (alternative splicing) resulting in afew normal FVIII molecules and thus a phenotypeof mild hemophilia.[16]

Hemophilia BIt has been estimated that in 97 per cent of patientswith mild hemophilia B, the underlying defect is amissense mutation.[17] These mutations can causereduced interaction with FVII-tissue factor andtherefore reduced activation of FIX[18], decreasedactivity due to reduced affinity for FVIII[19], orreduced activity if the amino acid substitution is inthe catalytic domain[20], a common mutation in theAmish population. Mutations in the carboxyterminalportion of FIX (residues 403-415) result in impairedsecretion of the factor from the hepatocyte[21], butthe secreted molecules have normal function.

The most spectacular type of mutation in the FIXgene results in low FIX levels until puberty, andthereafter a rise by 5% per year up to a maximumof about 60%. This was first described in 1982 by aDutch group and is called hemophilia B Leyden.[22]It should be noted that even in the normal child thereis an increase of the FIX activity in parallel with thematuration of the liver, but here the baseline is about50% and a major rise occurs already during the firstfive years of life with a second rise during puberty.

Figure 1Increase of the factor IX level with age in the normalpopulation and in patients with hemophilia B Leyden.

The point mutations associated with hemophilia BLeyden have been identified in the promoter regionat nucleotide -20, -6, and –5, and in exon 1 at nucleotide+8 and +13. The promoter region of FIX contains a

Mutation FVIIIdomain

One-stageclot

Two-stageclot

Ala284Glu14

Ser289Leu14

Arg531His13

Arg698Leu14

Arg698Trp14

His1954Leu15

Leu1932Phe14

A2A2A2A2A2A3A3

38%33%36%30%

28-35%106%19%

10%9%19%6%

10-15%18%7%

020406080

100120

0 5 10 15 20 25 30 35 40

Normal

Hemophilia BLeyden

binding site for hepatocyte nuclear factor 4 (HNF-4),a transcription factor in the steroid hormone receptorsuperfamily.[23] In hemophilia B Leyden the bindingof HNF-4 to this promoter region is impaired andthereby also the transactivation of FIX. This is partlyovercome by the increasing level of testosterone inpuberty.

TreatmentThe use of factor concentrates is vital in patientswith the mild form of hemophilia in cases of majorsurgery or trauma. The principles are the same asfor patients with severe hemophilia with identicaltarget levels and need for frequent bolus doses, atleast initially. The dose required to reach this targetlevel, whether given as bolus doses or as continuousinfusion, obviously decreases with increasing baselinelevel of the deficient factor. However, for patientswith mild hemophilia, there are additional treatmentoptions. For minor procedures in patients with factorlevels in the upper range of “mild hemophilia”(approximately 20% factor level) treatment with aninhibitor of fibrinolysis may be sufficient. Thisapproach, for example using tranexamic acid (20mg/kg orally or 10 mg/kg intravenously everyeight hours), should be used for any surgery involv-ing mucosal membranes. For dental extractions,mouth rinse (“swish and swallow”) with tranexamicacid is efficient and safe (see WFH Treatment ofHemophilia Monograph No. 40, Guidelines forDental Treatment of Patients with Inherited BleedingDisorders).

Hemophilia AIn mild hemophilia A, an important alternative tofactor concentrates is desmopressin (DDAVP).Thirty years ago, Dr. Pier Mannucci discovered thatDDAVP elevates the FVIII level about three-foldover baseline and is a useful strategy to providehemostasis during surgical procedures in thesepatients.[24] The consequent and consistent use of thisalternative mode of treatment saved many Italianpatients with mild hemophilia A from transfusion-transmitted viral infections in the late 1970s andearly 1980s.

The dose (0.3 µg/kg) is preferably given subcuta-neously one hour before the procedure and mayhave to be repeated every eight to 24 hours,depending on the extent of the surgery. However,repeated doses may cause fluid retention withhyponatremia, and in sensitive subjects (childrenand women at the time of delivery) seizures. Theeffect may also decrease after several doses — a

4 Mild Hemophilia

phenomenon known as tachyphylaxis. Since therise of FVIII is quite variable between patients (butreproducible from time to time), a test should bedone in each patient to evaluate the response. In apopulation of 62 patients with mild hemophilia, 47per cent responded to DDAVP with a doubling ofthe factor VIII level and a peak level above 30%.[25]Predictors for a good response were a higher base-line level and older age. Intranasal administration is also possible but requireshigher doses.

DDAVP may be useful in patients with mild hemo-philia A and an inhibitor[26], since the antibodiesmay not inhibit the endogenous FVIII or not to thesame extent as exogenous FVIII.

Hemophilia BA small but certain benefit of DDAVP has also beenreported in patients with mild hemophilia B. Althoughthe FIX level did not increase much (1.4 times), therewas a shortening of the aPTT and dental surgerycould be performed with good result.[27] This isprobably a result of compensation from higher levelsof FVIII and vWF. It should be emphasized that thiseffect of DDAVP is by no means as well document-ed as that in hemophilia A and it is of a muchsmaller magnitude in hemophilia B.

In patients with hemophilia B Leyden, attemptshave been made to anticipate the effect of pubertyby using anabolic steroids[28], but this is not a rec-ommendable alternative due to many side effects.

Follow-upPatients with mild forms of hemophilia should notbe left without follow-up after the diagnosis hasbeen established. Although the intervals betweenvisits can be longer than with the severe form, areview of the patient every two to three years isvital. At this visit a bleeding history is taken; physicalexam (especially to identify arthropathy) is performed;laboratory tests to identify complications of previoustransfusions (viral markers, factor inhibitor) are done;and information is given to the patient and/or hisparents. This information should include signs andsymptoms typical for routine and dangerous typesof hemorrhage, as well as appropriate response andtreatment intervention. Self-injection of DDAVP isuseful to teach patients with mild hemophilia Awho bleed at least once per year.

Mild Hemophilia 5

InhibitorsInhibitors occur also in mild hemophilia, most typicallyafter intensive exposure to factor concentrates.[29]A suspicion was raised that continuous infusion offactor concentrates may be more risky in thisrespect than bolus injections.[29] However, thisremains to be proven.

The cumulative incidence of inhibitors in patientswith mild hemophilia A has been reported to be 3to 13 per cent by the age of 33 in some populations.These patients often have missense mutations withconformational changes at the site of an antigenicepitope on the surface of the molecule. The mostcommon mutations that generate an inhibitor areArg593Cys and Trp2229Cys. Patients with thesemutations have a 40 per cent risk of developing aninhibitor.[30] In 60 per cent of these patients, toleranceoccurs spontaneously about nine months afterappearance of the inhibitor, but in the meantimethe inhibitor can cause severe bleeding and evendeaths. Inhibitors in patients with mild hemophilia Bare extremely uncommon.

Conclusion Mild hemophilia is a neglected diagnosis. Patientswith mild forms of hemophilia have generally notreceived the same degree of medical attention asthose with severe forms. Although justified to a cer-tain extent, this has not infrequently resulted inserious neglect. One of the first reports was in 1964of an 18-year-old male with a delayed diagnosisafter trauma-induced hemarthrosis, eventuallyresulting in infection, osteomyelitis, and life-threat-ening sepsis, necessitating amputation of the leg atthe hip joint.[31]

A review of all Swedish patients with hemophiliain 1982 demonstrated the ironic situation that themortality in bleeding in the central nervous systemwas higher in mild than in severe hemophilia.[32]Recently, in a Belgian-French study, intracranialhemorrhages were identified in 83 patients withsevere hemophilia, with fatal outcome in 17 (20 percent); and in 40 patients with mild hemophilia, withfatal outcome in 10 (25 per cent).

Carriers of hemophilia with factor levels in the rangeof mild hemophilia should receive the diagnosis of“mild hemophilia,” since there is a widespread con-cept that males have the disease and females are“only” carriers.

It is of utmost importance to disseminate knowledgeabout mild hemophilia, to keep patients informed

and updated by regular visits, and to implementappropriate diagnostic tools and treatment modalities.

References1. White GC 2nd, Rosendaal F, Aledort LM, et al.

Definitions in hemophilia. Recommendation ofthe scientific subcommittee on factor VIII andfactor IX of the scientific and standardizationcommittee of the International Society onThrombosis and Haemostasis. Thromb Haemost.2001; 85:560.

2. Venkateswaran L, Wilimas JA, Jones DJ, et al.Mild hemophilia in children: prevalence, com-plications, and treatment. J Pediatr HematolOncol 1998; 20:32-35.

3. Larsson SA, Nilsson IM, Blombäck M. Currentstatus of Swedish hemophiliacs: A demographicsurvey. Acta Med Scand 1982; 212:195-200.

4. Jensson O, Stenbjerg Bernvil S, Jonsdottir S, etal. Mild haemophilia A in Iceland: clinicalgenetic studies of three families with the samemutation. J Intern Med 1994; 235:443-450.

5. Larsson SA. Hemophilia in Sweden: Studies ondemography of hemophilia and surgery inhemophilia and von Willebrand’s disease. ActaMed Scand Suppl 1984; 684:1-72.

6. Darby SC, Ewart DW, Giangrande PL, et al.Mortality before and after HIV infection in thecomplete UK population of haemophiliacs. UKHaemophilia Centre Directors’ Organisation.Nature 1995; 377:79-82.

7. Chambost H, Gaboulaud V, Coatmelec B, et al.What factors influence the age at diagnosis ofhemophilia? Results of the French hemophiliacohort. J Pediatr 2002; 141:548-552.

8. Ingram GI, O’Brien PF, North WR. TheICTH/WFH study of the partial thromboplastintime in mild haemophilia. Scand J HaematolSuppl 1980; 37:64-72.

9. Preston FE. Laboratory diagnosis of hereditarybleeding disorders: external quality assessment.Haemophilia 1998; 4 Suppl 2:12-18.

10. Preston FE, Kitchen S, Jennings I, et al.SSC/ISTH classification of hemophilia A: canhemophilia center laboratories achieve the newcriteria? J Thromb Haemost 2004; 2:271-274.

11. Mazurier C, Goudemand J, Hilbert L, et al.Type 2N von Willebrand disease: clinical man-ifestations, pathophysiology, laboratory diagnosisand molecular biology. Best Pract Res ClinHaematol 2001; 14:337-347.

12. Schwaab R, Oldenburg J, Schwaab U, et al.Characterization of mutations within the factorVIII gene of 73 unrelated mild and moderatehaemophiliacs. Br J Haematol 1995; 91:458-464.

13. Pipe SW, Eickhorst AN, McKinley SH, et al.Mild hemophilia A caused by increased rate offactor VIII A2 subunit dissociation: evidencefor nonproteolytic inactivation of factor VIIIain vivo. Blood 1999; 93:176-183.

14. Rudzki Z, Duncan EM, Casey GJ, et al.Mutations in a subgroup of patients with mildhaemophilia A and a familial discrepancybetween the one-stage and two-stage factorVIII:C methods. Br J Haematol 1996; 94:400-406.

15. Keeling DM, Sukhu K, Kemball-Cook G, et al.Diagnostic importance of the two-stage factorVIII:C assay demonstrated by a case of mildhaemophilia associated with His1954—Leusubstitution in the factor VIII A3 domain. Br JHaematol 1999; 105:1123-1126.

16. Acquila M, Pasino M, Lanza T, et al.Duplication of exon 13 causes one third of thecases of mild hemophilia A in northern Italy.Haematologica 2004; 89:758-759.

17. Sommer SS, Bowie EJ, Ketterling RP, et al.Missense mutations and the magnitude offunctional deficit: the example of factor IX.Hum Genet 1992; 89:295-297.

18. Wu PC, Hamaguchi N, Yu YS, et al.Hemophilia B with mutations at glycine-48 offactor IX exhibited delayed activation by thefactor VIIa-tissue factor complex. J ThrombHaemost 2000; 84:626-634.

19. Lefkowitz JB, Nuss R, Haver T, et al. Factor IXDenver, ASN 346—>ASP mutation resulting ina dysfunctional protein with defective factorVIIIa interaction. J Thromb Haemost 2001;86:862-870.

20. Ketterling RP, Bottema CD, Koeberl DD, et al.T296—M, a common mutation causing mildhemophilia B in the Amish and others:founder effect, variability in factor IX activityassays, and rapid carrier detection. Hum Genet1991; 87:333-337.

21. Kurachi S, Pantazatos DP, Kurachi K. The car-boxyl-terminal region of factor IX is essentialfor its secretion. Biochemistry 1997; 36:4337-4344.

22. Briet E, Bertina RM, van Tilburg NH, et al.Hemophilia B Leyden: a sex-linked hereditarydisorder that improves after puberty. N Engl JMed. 1982; 306:788-790.

6 Mild Hemophilia

23. Reitsma PH, Bertina RM, Ploos van Amstel JK,et al. The putative factor IX gene promoter inhemophilia B Leyden. Blood. 1988; 72:1074-1076.

24. Mannucci PM, Ruggeri ZM, Pareti FI, et al. 1-Deamino-8-d-arginine vasopressin: a newpharmacological approach to the managementof haemophilia and von Willebrand’s diseases.Lancet 1977; 1:869-872.

25. Revel-Vilk S, Blanchette VS, Sparling C, et al.DDAVP challenge tests in boys with mild/moderate haemophilia A. Br J Haematol 2002;117:947-951.

26. Robbins D, Kulkarni R, Gera R, et al.Successful treatment of high titer inhibitors inmild hemophilia A with avoidance of factorVIII and immunosuppressive therapy. Am JHematol 2001; 68:184-188.

27. Ehl S, Severin T, Sutor AH. DDAVP (desmo-pressin; 1-deamino-cys-8-D-arginine-vasopressin)treatment in children with haemophilia B. Br JHaematol 2000; 111:1260-1262.

28. Briet E, Wijnands MC, Veltkamp JJ. The pro-phylactic treatment of hemophilia B Leydenwith anabolic steroids. Ann Intern Med. 1985;103:225-226.

29. Sharathkumar A, Lillicrap D, Blanchette VS, etal. Intensive exposure to factor VIII is a riskfactor for inhibitor development in mild hemo-philia A. J Thromb Haemost 2003; 1:1228-1236.

30. Hay CR. Factor VIII inhibitors in mild andmoderate-severity haemophilia A. Haemophilia1998; 4:558-563.

31. Pappas AM, Barr JS, Salzman EW, et al. Theproblem of unrecognized “Mild Hemophilia”.Survival of a patient after disarticulation of thehip. JAMA 1964; 187:772-774.

32. Larsson SA, Wiechel B. Deaths in Swedishhemophiliacs, 1957-1980. Acta Med Scand 1983;214:199-206.