The rare coagulation disorders-review with guidelines for management from the …

GUIDELINES

The rare coagulation disorders – review with guidelines formanagement from the United Kingdom Haemophilia CentreDoctors’ Organisation

P. H. B. BOLTON-MAGGS,* D. J. PERRY,� E. A. CHALMERS,� L. A. PARAPIA,§ J. T. WILDE,–

M. D. WILLIAMS,** P. W. COLLINS,�� S. KITCHEN,�� G. DOLAN§§ and A. D. MUMFORD––

*Department of Clinical Haematology, Manchester Royal Infirmary, Manchester; �Haemophilia Centre and Haemostasis

Unit, Royal Free and University College Medical School, London; �Department of Haematology, Royal Hospital for Sick

Children, Yorkhill NHS Trust, Glasgow; §Haemophilia Centre, Bradford Royal Infirmary, Bradford; –Department of

Haematology, University Hospital Birmingham, Edgbaston, Birmingham, West Midlands; **Department of Clinical and

Laboratory Haematology, Birmingham Children’s Hospital, Birmingham; ��Arthur Bloom Haemophilia Centre,

University Hospital of Wales, Heath Park, Cardiff; ��Department of Coagulation, Sheffield Haemophilia and Thrombosis

Centre, Royal Hallamshire Hospital, Sheffield; §§Department of Haematology, University Hospital, Queens Medical

Centre, Nottingham; and ––Bristol Haemophilia Centre, Bristol Haematology and Oncology Centre, Bristol, UK

Summary. The rare coagulation disorders are her-itable abnormalities of haemostasis that may pre-sent significant difficulties in diagnosis andmanagement. This review summarizes the currentliterature for disorders of fibrinogen, and deficien-cies of prothrombin, factor V, FV + VIII, FVII, FX,the combined vitamin K-dependent factors, FXIand FXIII. Based on both collective clinical experi-ence and the literature, guidelines for managementof bleeding complications are suggested with spe-cific advice for surgery, spontaneous bleeding,

management of pregnancy and the neonate. Wehave chosen to include a section on Ehlers-Dan-los Syndrome because haematologists may beconsulted about bleeding manifestations in suchpatients.

Keywords: rare coagulation disorders, fibrinogen,prothrombin, FV, FVII, FX, FXI, vitamin K depen-dent factors, FXIII, Ehlers Danlos syndrome, FactorV+VIII deficiency, molecular genetics2

Introduction

The rare coagulation disorders are heritable abnor-malities of haemostasis that may present significantdifficulties in diagnosis and management to haemo-philia centre clinicians. The common feature sharedby these disorders is that their overall populationfrequency is low (with the exception of factor XIdeficiency). Consequently, diagnosis and monitoringof affected individuals may require specialist phen-otypic and molecular investigations that are not

widely available. There may be considerable interin-dividual variation in bleeding phenotype amongstaffected individuals resulting at least in part from themolecular heterogeneity of the rare coagulationdisorders. The bleeding risks in affected individualsmay therefore be difficult to assess. Since there arefew long-term prospective studies of large cohorts ofpatients, reliable information about clinical manage-ment is often scarce. Coagulation factor support mayrequire the prescription of unlicensed treatmentproducts that are not readily available.

Although the rare coagulation disorders areuncommon, most haemophilia centres will have ahandful of individuals with one or more disorders.Some centres may have significant number of affec-ted individuals because of the prevalence of thesedisorders in populations in which consanguineousmarriage is common. The objective of this guideline

Correspondence: Dr Paula Bolton-Maggs, Chairman of the

UKHCDO rarer haemostatic disorders working party, Department

of Clinical Haematology, Manchester Royal Infirmary, OxfordRoad, Manchester M13 9WL, UK.

E-mail: [email protected].

Accepted 12 May 20042

Haemophilia (2004), 10, 593–628 DOI: 10.1111/j.1365-2516.2004.00944.x

� 2004 Blackwell Publishing Ltd 593

document is therefore to provide a concise review ofeach rare coagulation disorder and where possible toprovide evidence-based guidelines about diagnosisand management. Particular emphasis is placed onprophylaxis and the management of surgical pa-tients, pregnant mothers and affected neonates. Inview of the limitations of the literature describing therare coagulation disorders, unless specified in thetext, the evidence given below is level III (derivedfrom well-designed non-experimental descriptivestudies, such as case studies) or IV (based on expertcommittee reports or opinions and experiences ofrespected authorities), and the recommendationstherefore grade C (an absence of directly applicableclinical studies of good quality). Details of bloodproducts available for each disorder are not listed inthese guidelines because this information has recentlybeen published in the updated treatment guidelinesfrom the UKHCDO [1].

We have chosen to include Ehlers-Danlos Syn-drome (EDS) because it is a rare disorder andaffected individuals may present to haematologistwith bleeding symptoms.

General recommendations

General recommendations based on establishedmanagement protocols for the more common hae-mostatic disorders are also applicable to the rarecoagulation disorders. Affected individuals should beregistered with a haemophilia centre, and all invasiveprocedures should take place at a centre directlyserved by the haemophilia centre.

Affected individuals and family members should beoffered genetic counselling and screening. It shouldbe recognized that, with the exception of some of thefibrinogen (FI) disorders, the inheritance of the rarecoagulation disorders is usually autosomal recessive.Therefore, an affected proband may be part of akindred with very few affected individuals so thatpredictive information on phenotype may be scarce.It is, therefore, paramount to determine the moleculardefect within affected kindreds to enable compar-ison with affected individuals from other kindredsreported in case series or in electronic databases.

Best practice is for pregnancy in women withsevere rare disorders to be managed in an obstetricunit in a hospital that has a haemophilia centre. Ifthis is not possible, close collaboration between theobstetric unit and haemophilia centre is required.This good communication is also important to ensureappropriate investigation and management of apotentially affected neonate, for example, wherethe parents are related and already have one affected

child or are known to be carriers for one of thesedisorders. Several of the severe disorders are associ-ated with a significant risk of intracranial haemor-rhage in the first week of life.

Paediatricians and neonatologists need to be awareof the increased risk of the rare severe coagulationdefects presenting in offspring of consanguineousparents. It is very important that neonates whopresent with unexpected bleeding be investigatedurgently, and then treated vigorously to correct anysevere defect. Inadequate or delayed treatment ofneonatal intracranial haemorrhage leads to a lethal orpoor outcome, with significant long-term disability.

All affected individuals should be vaccinatedagainst hepatitis A and B by the s.c. route if notalready immune, and should generally be advised toavoid treatment with medication that interferes withplatelet function (salicylates and other non-steroidalanti-inflammatory agents). Paracetamol is a saferanalgesic to use.

Search strategy

Each section has been written by one or moremembers of the group who have particular experi-ence (clinical and/or research) of the disorder dis-cussed. In addition to personal archives, the medicalliterature was searched using appropriate key wordsto obtain as much published evidence as possible.

Abbreviations

PT, prothrombin time;APTT, activated partial thromboplastin time;TT, thrombin time;LMWH, low-molecular weight heparin;Units, we have used IU dL)1, where Internationalstandards are available (FII, FVII, FX) and U dL)1

where no International standard was established atthe time of preparation (FV, FXI and FXIII). Wehave used percentage when referring to a publicationwhich reported in this way. In these instances 100%is equal to 100 U dL)1.

Laboratory investigation of rare bleedingdisorders

Methodological aspects

Introduction In the investigation of bleeding disordersthe results of laboratory tests can be affected by thecollection and processing of blood samples and by theselection, design, quality control and interpretation ofscreening tests and specific assays. These effects have

594 P. H. B. BOLTON-MAGGS et al.

Haemophilia (2004), 10, 593–628 � 2004 Blackwell Publishing Ltd

important diagnostic and therapeutic implications.The purpose of this section of the guideline is todescribe these variables and their potential effectsand where possible to make recommendations aboutmethodological aspects of the laboratory investiga-tion of rarer bleeding disorders, although many willbe applicable to all disorders.

Sample collection and processing

Sample collection For tests of blood clotting venousblood should be collected gently but rapidly using asyringe or an evacuated collection system whenpossible. Application of a tourniquet to facilitatecollection does not normally affect the results ofmost tests for bleeding disorders, providing excessivestasis is avoided. By choice the needle should be notmore than 21 gauge (for infants a 22 or 23 gaugeneedle may be necessary). Collection through per-ipheral venous catheters [2] or non-heparinizedcentral venous catheters [3] can be successful forprothrombin time (PT) and activated partial throm-boplastin time (APTT) testing. Local discard proce-dures should be used where necessary in order toavoid contamination of samples with heparin.

If there is any delay between collection and mixingwith anticoagulant or delay in filling of collectionsystem, the blood must be discarded because ofpossible activation of coagulation. Once blood andanticoagulant are mixed the container should besealed and mixed by gentle inversion five times, evenfor evacuated collection systems, and vigorous sha-king should be avoided. Any difficulty in venepunc-ture may affect the results obtained, particularly fortests of platelet function. Prior to analysis the sampleshould be visually inspected and discarded if there isevidence of clotting or haemolysis.

Anticoagulant, sample filling and container The recom-mended anticoagulant for collection of blood forinvestigations of blood clotting is normally trisodiumcitrate [4,5]. Trisodium citrate (0.105–0.109 m) hasbeen recommended for blood used for coagulationtesting in general, including factor assays [5]. Onevolume of anticoagulant is mixed with nine volumesof blood. Although 0.129 m trisodium citrate hasbeen considered acceptable in the past [6] this is notcurrently recommended – the results of PT and APTTcan be up to 10% longer in the higher strengthanticoagulant depending on the reagents used [7].The samples must be filled correctly, avoiding bothunder- and overfilling to give accurate results [8].If the patient has a haematocrit >0.55 L L)1 resultsof PT and APTT can be affected and the volume of

anticoagulant should be adjusted to take account ofthe altered plasma volume [5].

The inner surface of the sample containeremployed for blood sample collection can influencethe results obtained, particularly for screening tests.The tubes should be inert and should not inducecontact activation (non-siliconized glass is inappro-priate). For factor assays there is evidence that resultson samples collected in a number of different sampletypes are essentially interchangeable [9].

There are specific differences in the haemostaticinvestigation of neonates and infants when comparedwith that of older children and adults. The BritishCommittee for Standards in Haematology hasrecently produced guidelines on the investigationand management of neonatal haemostasis andthrombosis, detailing sampling procedures, laborat-ory investigation and interpretation of test results[10].

Processing and storage of samples prior to analysis Forpreparation of platelet-rich plasma to investigateplatelet function, samples should be centrifuged atroom temperature (18–25 �C) at 150–200 g for 10–15 min, and analysed within 2 h of collection. Formost other tests related to bleeding disorders samplesshould be centrifuged at a speed and time whichproduces samples with residual platelet counts wellbelow 10 · 109 L)1, for example, using 2000 g forat least 10 min. Centrifugation at a temperature of18–25 �C is acceptable for most clotting tests. Aftercentrifugation, prolonged storage at 4–8 �C shouldbe avoided as this can cause cold activation, increas-ing FVII activity [11] and shortening of the PT orAPTT

Samples for APTT should be analysed within 4 hof collection [5] and although the PT of somesamples may be stable for 24 h or more [12] thestability of test results in subjects with moleculardefects is unknown. All investigations should there-fore be completed within 4 h of sample collection orplasmas should be deep frozen within this time forfuture analysis. Clotting factor assay results aregenerally stable for samples stored at )24 �C orlower for up to 3 months [13] and for samples storedat )74 �C for up to 18 months (results within 10%of baseline defined as stable). Storage in domesticgrade )20 �C freezers is normally unacceptable. Iffrozen samples are shipped on dry ice care must betaken to avoid exposure of the plasma to carbondioxide, which may affect the pH [14]. Prior toanalysis, frozen samples must be thawed rapidly at37 �C for 3–5 min. Thawing at lower temperatures isnot acceptable as cryoprecipitation may occur.

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� 2004 Blackwell Publishing Ltd Haemophilia (2004), 10, 593–628

Screening tests The sensitivity of PT and APTT to thepresence of clotting factor deficiencies is dependanton the test system employed [15–18]. The degree ofprolongation in the presence of a clotting factordeficiency can vary dramatically between reagents.The level of clotting factor deficiency, which shouldbe detected (as an abnormal result) by screening testshas not been defined. The use of different activatorsin APTT can influence sensitivity [18] and in additionthere is marked variation between reagents in respectof phospholipid profile [19]. As for APTT, reagentvariation will also influence the sensitivity of PTmethods for detecting abnormalities, particularly ofFVII or FX. It is useful to repeat borderline results ona fresh sample. It should be noted that the within-subject variation of PT and APTT over time might be6–12% [20].

For both PT and APTT the degree of prolongationmay be small in the presence of mild deficiency andtherefore there is a need for adequate quality controlprocedures and for carefully established accuratenormal/reference ranges. In view of the limitations ofscreening tests it is important that results areinterpreted in conjunction with all relevant personaland family history details. Normal screening tests donot always exclude the presence of mild deficiencystates.

Reference ranges In order to interpret the result ofany clotting test it is important to have data onresults of the test in healthy normal subjects. Healthis not well-defined and is often a relative term. Anage-and sex-matched normal range is the idealcontrol group. However, such careful selection isnot essential for many coagulation tests associatedwith the investigation of rare bleeding disorders.Samples from normal subjects should be collected,processed and analysed locally using identical tech-niques to those for patient samples. The literatureand reagent manufacturer’s information should onlybe used as a guide.

In the case of screening tests the reference valueswill be affected by the test system employed and thepossibility that a new batch/lot of reagent from thesame manufacturer has a different normal range thanprevious batches should be considered. For factorassays, results in normal subjects should ideally besimilar by different techniques, although this is notalways the case. The most suitable assay techniquesare those for which the locally established normalrange is similar to those reported elsewhere.

The number of normal subjects selected for ana-lysis should not be <25, which is probably adequatefor most tests of haemostasis related to investigation

of rare bleeding disorders. The results should show anormal distribution. Clear or statistical outliers areprobably aberrant results and it is usually acceptableto exclude these. The most frequently used conven-tion is that the reference range should include thecentral 95% of values, calculated as the mean ± 2 SDas the upper and lower limits respectively. For non-normally distributed values alternative calculationsmay be used (see Walker [21] for review).

Any reference range should only be used as a guideand aid to clinical interpretation.

Clotting factor assay design The most commonlyperformed assays for clotting factors for many yearshave been one-stage clotting assays based on theAPTT in the case of FXI and based on the PT in thecase of FII, FV, FVII or FX. Guidelines for the assayof clotting factors have been published [22]. Anumber of general features of the design of one-stage clotting assays, which are necessary to ensureaccurate, reliable and valid results are discussedbelow. In each case the test depends upon the abilityof a sample containing the factor under investigationto correct or shorten the delayed clotting of plasmacompletely deficient in that factor. Such deficientplasmas must contain <1 U dL)1 of the clottingfactor under investigation and normal levels of allother clotting factors which could influence theresults obtained.

It is important that the amount of factor present inthe mixture of deficient and reference/test plasma israte-limiting in its influence on the clotting timemeasured by APTT or PT. This requires dilution of areference or standard plasma of known concentra-tion. Preparation of several different dilutions of thereference plasma allows construction of a calibrationcurve in which the clotting time response depends onthe dose (concentration) of factor present. At lowerplasma dilutions/higher factor concentrations thefactor under investigation may not be rate limitingand the assay is no longer specific and thereforeinvalid. It may be necessary to extend the calibrationcurve by testing additional dilutions when analysingtest plasmas with concentrations below 10 U dL)1.At very low concentrations of an individual factorthe clotting time of the deficient plasma may not beeven partially corrected by addition of diluted testplasmas. Dilutions are selected so that there is alinear relationship between concentration (logarith-mic scale) and the response in clotting time (log-arithmic or linear scale). The reference curve shouldbe prepared using at least three different dilutionsand a calibration curve should be included each timethe assay is performed unless there is a clear evidence



that the responses are so reproducible that a calib-ration curve can be stored for use on other occasions.The reference plasma should be calibrated by a routetraceable back to WHO international standardswherever possible. Test plasmas should be analysedusing three dilutions so that it is possible to confirmthat the dose–response curve of the test plasma islinear and parallel to the dose–response curve of thereference plasma. It is not acceptable to test a singletest dilution as this reduces the accuracy substantiallyand may lead to important underestimation of thetrue concentration when inhibitors are present. If adose–response curve of a test plasma is not parallel tothe reference curve, and the presence of an inhibitorsuch as an antiphospholipid antibody is confirmed orsuspected then the estimate of activity obtained fromthe highest test plasma dilution is likely to be closestto the real concentration, but it should be noted thatthe criteria for a valid assay cannot be met andresults must be interpreted with caution.

Quality assurance

Laboratory tests of blood coagulation require carefulapplication of quality assurance (QA) procedures toensure reliability of results. QA is used to describe allthe measures that are taken to ensure the reliabilityof laboratory testing and reporting. This includes thechoice of test, the collection of a valid sample fromthe patient, analysis of the specimen and the record-ing of results in a timely and accurate manner,through to the interpretation of the results, whereappropriate, and communication of these results tothe referring clinicians.

Internal quality control (IQC) and external qualityassessment (EQA) are complementary components ofa laboratory QA programme.

Quality assurance is required to check that theresults of laboratory investigations are reliableenough to be released to assist clinical decision-making, monitoring of therapy and diagnosis ofhaemostatic abnormalities.

Internal quality control The IQC is used to establishwhether a series of techniques and procedures areperforming consistently over a period of time (preci-sion). It is therefore deployed to ensure day-to-daylaboratory consistency. It is important to recognizethat a precise technique is not necessarily accurate,accuracy being a measure of the closeness of anestimated value to the true value.

Quality control procedures should be applied toensure immediate and constant control of resultgeneration. Within a laboratory setting the quality

of results obtained is influenced by maintenance ofan up-to-date manual of standard operational pro-cedures; use of reliable reagents and referencematerials; selection of automation and adequatemaintenance; adequate records and reporting systemfor results; an appropriate complement of suitablytrained personnel.

For screening tests it is important to includeregular and frequent testing of QC material thatshould include a normal material and at least 1 levelof abnormal sample. For factor assays a QC sampleshould be included with each group of tests. Patientresults should only be released if QC results remainwithin acceptable target limits.

External quality assessment The EQA is used to iden-tify the degree of agreement between one laboratory’sresults and those obtained by other centres (accuracy).The primary function of EQA is proficiency testing ofindividual laboratory testing, but larger EQA schemescan also provide information concerning the relativeperformance of analytical procedures, including themethod principle, reagents and instruments. Contin-ued participation in EQA schemes has been linked toimproved laboratory performance [23]. This has beenseen not only in the overall performance, evidenced bya reduction of the variability of results betweenlaboratories but also in respect of individual laborat-ories. Centres undertaking investigations for rarebleeding disorders should participate in an accreditedEQA programme where available.

Recommendations

1 Nine parts blood should be mixed with one part0.105–0.109 m trisodium citrate.

2 Underfilled, haemolysed or partially clottedsamples should not be analysed.

3 The collection system used should not inducecontact activation.

4 For preparation of platelet poor plasma, samplesshould be centrifuged at a speed and time thatproduces plasmas with residual platelet countsbelow 10 · 109 L)1 (e.g. 2000 g for 10 min).

5 Analysis should be completed within 4 h ofsample collection. Alternatively plasmas can befrozen at )24 �C for up to 3 months or )74 �Cfor up to 18 months.

6 The sensitivity of screening tests are influenced byreagents and only techniques that are capable ofdetecting clinically important abnormalitiesshould be employed.

7 Reference ranges should be established locallyusing samples from normal subjects, which have



been collected, processed and analysed usingidentical techniques to those for patientsamples. This is essential for screening tests inparticular.

8 The most suitable assay techniques are those forwhich the locally established reference range issimilar to those reported elsewhere.

9 Quality control samples should be analysedregularly and frequently for screening tests, andwith each group of factor assays, and concurrentpatient test results only released to clinicianswhilst QC results remain within an acceptabletarget range.

10 Centres should participate in accredited EQAprogrammes for all tests in the investigation ofbleeding disorders, where available.

11 One-stage clotting factor assays should be calib-rated with reference plasmas traceable back toWHO standards where available.

12 In factor assays Test samples should be analysedusing three dilutions and results are only validwhen the dose–response curve of the test is linearand parallel to the reference curve.

Inherited abnormalities of fibrinogen

Background

Fibrinogen is a 340 kDa protein that is synthesized inthe liver. It has a plasma concentration of approxi-mately 1.5–3.5 g L)1 and a half-life of around4 days. The FI molecule is a homodimer, each halfconsisting of three non-identical polypeptide chainstermed Aa, Bb and c. The genes for all three chainsare located on the long arm of chromosome 4.

Fibrin is produced by proteolytic cleavage of FIby thrombin with the release of fibrinopeptidesA and B and generation of insoluble fibrin monomerfollowed by polymerization. FI is also importantin primary haemostasis for normal platelet aggre-gation.

Definitions

1 Afibrinogenaemia refers to the total absence of FImeasured by an antigenic assay.

2 Hypofibrinogenaemia is a decreased level of nor-mal FI.

3 Dysfibrinogenaemia is characterized by structuralabnormality of the FI molecule resulting in alteredfunctional properties. Classically the functionalassay of FI yields low levels compared withimmunological assay but levels may be concordantand the functional level may even be normal.

The definition and classification of abnormalitiesof FI have changed with increasingly sophisticatedmethods of analysis. Thus, many cases originallyreported as afibrinogenaemia have actually beenshown to be cases of dysfibrinogenaemia and manycases of hypofibrinogenaemia are also dysfunctional(hypodysfibrinogenaemia).

There are many acquired causes of dysfibrinogen-aemia and hypofibrinogenaemia. These will not becovered in this guideline.

Incidence

Afibrinogenaemia has an estimated prevalence1:1 000 000 [24]. Data from the UKHCDO from1999 showed 115 patients registered with 10 havingreceived treatment during that year, three patientswere on home treatment.

Geographical variation in prevalence reflects thehigh incidence in children of consanguineous parentsin Muslim countries. Comparative data from regis-

tries in Iran, Italy and the UK highlights thisgeographical variation (patients with levels £10%only registered) [24].

The incidence of dysfibrinogenaemia is unknownbut is probably under-diagnosed and under-reported.

Clinical phenotypes

Afibrinogenaemia/hypofibrinogenaemia

Bleeding – Afibrinogenaemia is associated with ableeding tendency of variable severity including life-threatening, spontaneous events, but long periodswithout problems are also not uncommon. Thepublished literature consists mostly of case reportswith two larger case series of afibrinogenaemicpatients [25,26]. In the series from Iran [25],bleeding and thrombotic manifestations were repor-ted for 55 patients with afibrinogenaemia. Umbilicalbleeding and mucosal bleeding were the most com-mon features. Musculoskeletal bleeding was lesscommon than in haemophilia but still occurredrelatively frequently. Three of 55 (5.5%) had central

Prevalence of severe fibrinogen deficiency (level <10% of normal)

in different countries.

Country Number Percentage

Iran 70 1.5*

Italy 10 0.2*

UK 11 0.2*

*Percentage of total number of patients registered with inherited

coagulation disorders in each country.



nervous system (CNS) bleeding but the age of onset isnot stated. In the other series of 10 patients from twolarge sibships from Israel, there was again a highincidence of umbilical bleeding with CNS and intra-abdominal bleeding also fairly common. Otherauthors have also noted the relative infrequency ofmusculoskeletal bleeding and there are a number ofother case reports of CNS bleeding and intra-abdominal bleeding related to splenic rupture [27–29]. In some patients there have been recurrentevents.

In hypofibrinogenaemia the bleeding pattern issimilar but appears to follow a milder course andbleeding may follow invasive procedures.Pregnancy-related complications – Afibrinogenaemiaand hypofibrinogenaemia are associated with recur-rent miscarriage, and also with both antepartum andpostpartum haemorrhage [30,31]. In one series of 13pregnancies in six women with afibrinogenaemia,seven ended in spontaneous abortion at 6–7 weeksgestation. This suggests that FI may be important forimplantation and this is supported by studies in micewith afibrinogenaemia where failure of embryoimplantation has been documented [32].Thrombosis – Paradoxically for a disorder associatedwith a significant bleeding tendency there are reportsof thrombotic events in patients with afibrinogenae-mia. Thrombotic events were recorded in two of55 (4%) of cases in one series, including one case ofsinovenous thrombosis in a 5-year-old boy andischaemic gangrene in a 15-year-old girl [25]. Theseevents occurred unrelated to FI replacement therapyor other obvious risk factors and the mechanism ofthrombosis is obscure. There are also a small numberof case reports of thrombotic events in patients withapparent afibrinogenaemia and hypofibrinogenaemia[33,34]. The details of some of these events are notwell-defined and others have been reclassified asdysfibrinogenaemias, therefore, the overall incidenceof thrombosis in these disorders is unclear.Other – Impaired wound healing postsurgery hasalso been reported in afibrinogenaemia.

Dysfibrinogenaemia The literature relating to dysfi-brinogenaemia consists predominantly of collectionsof case reports of patients seen at specialist centres.Most work has been carried out on the molecularanalysis of dysfibrinogenaemia with relatively littleon clinical aspects. This may have introduced bias, aspatients with clinically more severe disease will beover represented.

The clinical phenotype of dysfibrinogenaemia isunpredictable. A compilation of over 250 patientswith dysfibrinogenaemia revealed that 53% were

asymptomatic, 26% had haemorrhage and 21% hadthrombosis, some of whom also had haemorrhage[35]. There is some correlation between moleculardefects and clinical phenotype.Asymptomatic – Patients may present through rout-ine laboratory tests or family studies.Bleeding – Patients may have bleeding postpartumor with surgery and dental extraction. Bleeding fromthe umbilical stump and into the CNS and softtissues are associated with lower levels. Patientsmay also experience delayed wound healing anddehiscence.Thrombosis – The ISTH SSC Subcommittee on FIhas received reports on 51 cases of thrombosisassociated with dysfibrinogenaemia [36]. Only 26mutations were considered to be unambiguouslyassociated with thrombosis as defined by the fol-lowing strict criteria: known molecular defect,thrombosis at early age in two or more familymembers, no other known risk factors, if defectfound in another family this family should alsoexhibit thrombosis.

In the 26 index patients dysfibrinogenaemia wasassociated with deep vein thrombosis, thrombophle-bitis and pulmonary embolus at a young age (mean:32 years). Severe bleeding in this group was onlynoted postpartum. In 187 family members, 20 of99 people with and none of 88 people withoutdysfibrinogenaemia had had a thrombotic event[36]. The thrombotic risk is increased with acquiredrisk factors and coinheritance of other thrombo-philias [37].

Skin necrosis has been described with some dysfi-brinogenaemias (e.g. in those homozygous for FIMarburg). Arterial thrombosis occurs but much lesscommonly than venous thrombosis [36].Pregnancy – Of the 15 women in the ISTH databaseall had had at least one pregnancy and seven had hada postpartum thrombosis. There were 24 sponta-neous abortions and six stillbirths with 34 normaldeliveries [36]. Pregnant women with dysfibrino-genaemia are at risk of bleeding following vaginaldelivery, Caesarean section and with regional anal-gesia. Bleeding may not correlate well with level of FIor thrombin time but women with undetectablefunctional FI predictably bleed at delivery and have ahigh rate of miscarriage.

Diagnosis

Fibrinogen assays should be performed in accordancewith BCSH guidelines [38] (http://www.bcshguide-lines.com/). A FI level derived from the prothrombintime should not be used.



Tests should be interpreted with regard to possiblecauses of acquired dysfibrinogenaemia. The patient’sage, drug history and liver function tests should beknown. Family studies may be helpful.

Afibrinogenaemia There is a marked prolongation ofthe PT, APTT and thrombin time (TT). The bleed-ing time is often prolonged. FI levels are undetect-able by both functional (Clauss) and antigenicassays.

Hypofibrinogenaemia Coagulation tests are pro-longed in proportion to the FI deficiency. TT is themost sensitive test. The total clottable and immuno-genic FIs are both reduced to a similar level as thefunctional FI. It is important to exclude acquiredcauses of hypofibrinogenaemia and family studiesmay be helpful.

Dysfibrinogenaemia The TT is usually the most sen-sitive screening test for dysfibrinogenaemia. TT isusually prolonged but in rare cases may be normal orshortened. It is important to exclude the effect ofheparin and interference with fibrin polymerization,for example, by3 fibrin/fibrinogen degradation pro-ducts (FDPs) or paraproteins, and thrombin functionby a thrombin inhibitor. The reptilase time is usuallyprolonged but may be shortened or sometimesnormal. In some cases the reptilase time is moresensitive than TT. The PT and APTT may beprolonged but are less sensitive than the TT. Thesensitivity of coagulation tests to dysfibrinogenaemiaare dependent on the specific mutation, reagents andtechnique [39].

The functional FI by Clauss method will be low.Antigenic measurements may show a decrease inproportion to the Clauss assay or be significantlyhigher.

A definitive diagnosis requires the demonstrationof a molecular defect. However, as most dysfibri-nogenaemias are dominantly inherited, family stud-ies are useful to confirm inherited rather thanacquired dysfibrinogenaemia.

In patients with dysfibrinogenaemia and a historyof thrombosis it may be helpful to perform athrombophilia screen to exclude coexisting pro-thrombotic defects.

Molecular basis for inherited abnormality offibrinogen

The three FI subunits (Aa, Bb and c) are encoded bythree different genes clustered in a region of 50 kb onchromosome 4 (q28–30), FGA, FGB and FGG.

The FGA is 5.4 kb in length and consists of sixexons. It encodes two different transcripts byalternative splicing at the 3¢-end. The major isoform(a), which represents 99% of mRNA, is coded byexons 1–5. The extended aE variant is produced bythe addition of 236 amino acids encoded by exon 6.

The FGB consists of eight exons and is 8.2 kb inlength. The synthesis of the Bb chain is the rate-limiting step in the production of mature FI and thereis much research into the effect of polymorphismswithin the FGB promoter region on FI levels.

The FGG is 8.4 kb long and has 10 exons. Thec-chain occurs in two isoforms (c and c¢), which areproduced by alternative splicing. The c¢ isoformconstitutes about 15% and contains an additionalbinding site for FXIII and for thrombin.

Congenital afibrinogenaemia Congenital afibrinogen-aemia is an autosomal recessive condition that hasbeen shown to be due to defective FI synthesis.

Although mutations causing afibrinogenaemiahave been detected in all three genes, the majorityfound to date are in FGA. These mutations aremainly deletions, frameshift, nonsense or splicingmutations and several recurrent mutations have beendetected, including an 11 kb deletion in the FGAgene and a donor splice mutation in intron 4 + 1G>T in FGA.

Dysfibrinogenaemia Overall 300 abnormal FIs havebeen described and over 100 different structuraldefects identified. These are mainly associated withmissense mutations affecting FI structure or function.These mutations are registered on a database [35],which is available on-line. The majority (180) ofthese mutations are in FGA of which 74 are at(Aa 16). Twenty-seven mutations have been reportedin the FGB and 75 in the FGG of which 25 affect c275. These mutations affect regions associated withall aspects of FI function.

Mutation detection in cases of dysfibrinogenaemiahas led to the realization that previously discretecases share the same molecular abnormality. This hasallowed pooling of clinical information concerningany given mutation and thus provides a useful guidefor decision-making. For example, FI Bremen (AaGly17 fi Val) is associated with a bleeding disorderand delayed wound healing. Conversely, the major-ity of individuals with dysfibrinogenaemia [AaArg16 fi His (Bern IV and Milano XI)] were asymp-tomatic or associated with only a mild bleedingtendency. Individuals (65%) with FGG mutationsare asymptomatic with only 5% having bleedingproblems and 30%, thrombosis. Certain mutations



such as Arg554 fi Cys (Chapel Hill III, Paris V andDusart) are associated with thrombosis but notbleeding [35,40].

While molecular identification of dysfibrinogenae-mia is useful step in the assessment of each patient,more accurate and clinically robust information isrequired for all mutations registered on the databaseto maximize the utility of this resource.

A mutation database can be viewed at http://www.geht.org/databaseang/fibrinogen/

Treatment principles

The patient’s personal and family history of bleedingand thrombosis are important guides for manage-ment. If FI replacement is required a virally inacti-vated concentrate should be used (grade Brecommendation based on level IIb evidence). Ingeneral a FI level below 0.5 g L)1 is associated withan increased risk of microvascular bleeding [41].

Treatment products

There is no licensed FI concentrate available in theUK. The products that are available in Europe havebeen listed in the treatment guidelines [1].

A suggested amount of FI required is:Dose (g) ¼ Desired increment in g L)1 · plasma

volume, where the plasma volume is 0.07 ·(1 ) haematocrit) · weight (kg)

Therefore, to raise the FI concentration by 1 g L)1

a dose of about 30 mg kg)1 would be required.The half-life of infused FI is 3–5 days [42] and, in

the absence of consumption, treatment is unlikely tobe needed more often than on alternate days. Thesepharmacokinetic data were established in studies ofadult patients and so may not be applicable tochildren.

Although cryoprecipitate is a good source of FIit should not usually be used, as it is not virallyinactivated. Its use may be considered in an emer-gency situation if no suitable alternative is available.

Fibrin glue may be useful to treat superficialwounds or following dental extraction. There arereports of patients developing anti-FI antibodies aftertheir use [1].

Tranexamic acid can be useful to treat mucosalbleeding or prevent bleeding for procedures such asdental extraction, avoiding the need for bloodproducts. Its use may, however, increase the riskof thrombosis and should be used with cautionin people with a personal or family history ofthrombosis and avoided when other risk factors suchas pregnancy, surgery or immobilization are present.

Treatment

Management of spontaneous bleeding

Afibrinogenaemia – Fibrinogen concentrate is thetreatment of choice for significant bleeding. Theaim should be to increase the FI level to 1 g L)1 andmonitor the clinical response. Patients may requirerepeat infusions dependent on the clinical responseand laboratory monitoring. The FI level should bemaintained above 1 g L)1 until haemostasis issecure and above 0.5 g L)1 until wound healing iscomplete.

Indications for prophylaxis Based on the limitedavailable data it is not possible to make a recom-mendation on the use of primary prophylaxis.Secondary prophylaxis may be appropriate in caseswhere there has been potentially life-threateningbleeding because of the risk of recurrence, e.g.intracranial haemorrhage. The frequency and doseof FI concentrate should be adjusted to maintain atrough level above 0.5 g L)1. Antifibrinolytic agentsmay be useful for mucosal bleeding and oestrogen/progesterone preparations have been used in menor-rhagia [43]. Antifibrinolytic medication should beused with caution in patients with a personal orfamily history of thrombosis.Dysfibrinogenaemia – There are few data on themanagement of bleeding in patients with dysfibrin-ogenaemia. The abnormal FI in the patient may bedysfunctional and may interfere with the function ofinfused FI. Furthermore, abnormal FI may interferewith laboratory assays in patients who have receivedFI infusions. Raising the measured FI level in thepatient to 1 g L)1 may, theoretically, not provideadequate haemostasis.

With these considerations in mind, patients whoare bleeding are likely to require FI concentratealthough in some cases topical fibrin glue ortranexamic acid may be sufficient for superficial ormucosal bleeding avoiding the use of pooled bloodproducts. The functional FI level should be raised to1 g L)1 above baseline level and the clinicalresponse observed. Patients may need repeat infu-sions dependent on the clinical response and labor-atory monitoring. The FI level may need to bemaintained above 1 g L)1 until wound healing iscomplete [40].

Management of surgery

Afibrinogenaemia – In the Iranian series [25] postop-erative bleeding was reported in 23 of 55 (40%) ofuntreated cases. It is, therefore, recommended that FI



levels are increased to 1 g L)1 and maintained at thislevel until haemostasis is secure and above 0.5 g L)1

until wound healing is complete.Dysfibrinogenaemia – Patients with a known bleedingphenotype should be treated with a FI concentratepreoperatively to raise and maintain the FI level to1 g L)1 above their baseline level until haemostasis issecure and 0.5 g L)1 above baseline until woundhealing is complete. Venous thrombosis may beprecipitated by FI concentrate and so the use ofcompression stockings is necessary and prophylacticheparin should be considered.

Patients with a thrombotic phenotype should betreated with compression stockings and low-molecu-lar weight heparin (LMWH) dependent on personaland family history in accordance with previousguidelines on inherited thrombophilia [44].

Patients with both a bleeding and thromboticphenotype are complicated and are likely to requirereplacement of FI to maintain a level 1 g L)1 abovebaseline and use of LMWH.

There are a number of options for asymptomaticpeople or those who have not had a previous surgicalchallenge. These are:1 Observe and only give FI concentrate if bleeding

occurs.2 Treat with FI concentrate as above (if family his-

tory is of bleeding).3 Use an antifibrinolytic agent, e.g. tranexamic acid.

The decision will depend on the level of functionalFI (patients <0.5 g L)1 are more likely to bleed), pasthistory of haemostatic challenges, family history ofbleeding and thrombosis.

The use of tranexamic acid, for example, to coverdental extraction may avoid the need for pooledblood products but increases risk of venous throm-bosis during surgery and should be avoided if there isa personal or family history of venous thrombosis.

Management of pregnancy The management dependson the FI level and the personal and family history ofbleeding or thrombosis.Afibrinogenaemia – Prophylaxis with FI concentrate isreported to improve pregnancy outcome and preventpostpartum haemorrhage [30,31]. Regular FI infu-sions are required throughout pregnancy in womenwith afibrinogenaemia and should be commenced assoon as possible to prevent early fetal loss. OptimalFI levels are not defined but fetal loss and bleedinghave been reported in women with FI levels of>0.5 g L)1, therefore, levels above 1 g dL)1 may berequired, particularly toward the end of pregnancy.Dysfibrinogenaemia – During delivery it shouldassumed that the neonate has dysfibrinogenaemia

and invasive monitoring and procedures should beavoided, particularly if the family phenotype is ofbleeding. If tested at birth the interpretation may becomplicated by physiological or acquired dysfibrin-ogenaemia particularly in preterm infants.

Asymptomatic women should be observed closelybut no specific treatment is required unless bleedingoccurs or the family history suggests bleeding islikely. Standard measures for venous thromboproph-ylaxis should be followed [45].

Women with a personal or family history ofthrombosis should be offered antenatal prophylaxiswith LMWH. The delivery should be closely mon-itored and FI replacement therapy given only ifbleeding occurs. The risk of regional anaesthesia isdifficult to assess and should be avoided as thewoman may be on heparin and haemorrhagic com-plications cannot be excluded. General anaesthesia,however, increases risk of venous thrombosis and aplan should be made in advance after discussion withthe patient and an obstetric anaesthetist.

There are a number of management options forwomen who have a bleeding phenotype. The decisionwill depend on the person and family history ofbleeding.

Vaginal delivery can often be managed by obser-vation and infusion of a FI concentrate given to raisethe FI level to 1 g L)1 above baseline only if bleedingoccurs. If the personal or family history of bleeding isstrong a vaginal delivery may need to be coveredwith FI concentrate prophylactically to raise the FIlevel 1 g L)1 above baseline.

If FI concentrate is needed the FI level should bemaintained 0.5 g L)1 above baseline until woundhealing has occurred.

Caesarean section is likely to require FI replace-ment therapy as described for other surgical proce-dures. Thromboprophylaxis with compressionstockings and prophylactic doses of LMWH will berequired. The risk of epidural haematoma isincreased in patients with a bleeding phenotype bothat the time of insertion and removal of an epiduralcatheter. This type of anaesthesia should therefore beavoided.Miscarriage – There are no studies on the manage-ment of women with dysfibrinogenaemia who haverecurrent miscarriages. The options are to use pro-phylactic LMWH and if this fails consider the use ofFI replacement. In the absence of any data regardingdosing maintaining a FI level 1 g L)1 above baselinecould be tried.Management of thrombosis – Patients with dysfibrino-genaemia who suffer thrombosis should be managedaccording to BSCH guidelines [44].



Factor II – prothrombin deficiency

Molecular biology

Prothrombin (FII) is a 72 kDa single chain glycopro-tein which is synthesized by hepatocytes. It is one ofthe vitamin K-dependent coagulation factors andrequires post-translational carboxylation to becomefunctionally active. Prothrombin consists of fourdomains – Gla domain, kringle 1 and kringle 2domains and a serine protease domain. FXa activatesprothrombin on the surface of platelets in thepresence of FV and calcium. During cleavage ofprothrombin the activation peptide fragment 1 + 2 isreleased.

Incidence

Prothrombin deficiency is probably the rarest inher-ited bleeding disorder with an estimated prevalenceof 1:2 000 000 in the general population [46]. Aswith many rare haemostatic disorders the mode ofinheritance is autosomal recessive and the conditionis therefore seen more frequently where consanguin-eous marriages are common.

Clinical phenotypes

Two clinical phenotypes are recognized – hypopro-thrombinaemia (type I deficiency), in which pro-thrombin antigen and activity levels are reducedconcomitantly, and dysprothrombinaemia (type IIdeficiency) in which prothrombin activity is reducedbut antigen levels are normal. Occasional combineddefects (compound heterozygotes) have also beenreported.

Complete deficiency of prothrombin is not des-cribed suggesting that this is incompatible with life.This is in keeping with mouse knockout modelswhere experimental inactivation of the prothrombingene results in partial embryonic lethality andneonatal death [47].

Published data on this disorder are very limitedand the largest case series reported is from Iran [46].Prior to the publication of this series only 26 cases ofprothrombin deficiency and 22 cases with otherprothrombin abnormalities had been described in theworld literature [48].

Fourteen patients were included in the Iranianseries, 11 of whom had hypoprothrombinaemia withlevels of 4–10% and three with dysprothrombinae-mia. Haemarthrosis and muscle haematomas werethe most frequent severe bleeding manifestations inthis group of patients. There was one case of

intracranial bleeding and life-threatening umbilicalbleeding was reported in two neonates. Postoperativebleeding was also reported in the absence of replace-ment therapy but there were no cases of postpartumhaemorrhage recorded. Mucosal bleeding was alsofrequently reported but was not severe.

Bleeding manifestations in the 26 cases of hypo-prothrombinaemia reviewed by Girolami were sim-ilar to the Iranian series and again mucosal bleeding,soft tissue bleeding and haemarthrosis were relativelycommon [48]. There were also three reported casesof intracerebral bleeding. Two other cases of intra-cranial haemorrhage in hypoprothrombinaemia havebeen reported more recently in infants aged 4 and7 months [49].

Bleeding symptoms in patients with dyspro-thrombinaemias appear to be much more variableand many cases are asymptomatic or have onlyrelatively mild bleeding problems.

Diagnosis

Both the PT and APTT may be prolonged in FIIdeficiency, depending on the reagent, but it shouldalso be noted that the degree of abnormality may beminimal and results can be within the normal range.A specific FII assay may therefore be required in thepresence of clinical suspicion or appropriate familyhistory and normal screening tests. The one-stageclotting assay employing thromboplastin and basedon the PT is suitable and is the most widely usedtechnique. There are a number of assays, whichemploy different snake venoms to convert prothrom-bin to thrombin. These include Echis Carinatusvenom assays that do not require phospholipidsand do not differentiate between fully and partiallycarboxylated forms of FII. Other assays employingTaipan or Textarin venom are phospholipid-dependent.

In hypoprothrombinaemia results of all assays arereduced essentially in parallel [48]. In three subjectsconsidered to be homozygous for hypoprothrombin-aemia, FII activity levels were 9–16 U dL)1 by PT-based assay, compared with levels of 43–75 U dL)1

in heterozygous subjects. Normal subjects related tothese kindreds and unrelated normal subjects had FIIlevels in the range 84–130 U dL)1 [48]. This issimilar to the normal range of 84–132 IU dL)1

(mean ± 2 SD) in normal subjects tested withhuman recombinant thromboplastin PT-based assays(S. Kitchen, personal communication).

In dysprothrombinaemias results of PT-based andsome venom assays are substantially lower than resultsof antigen and some other assays. In one recent report



of a family with dysprothrombinaemia FII by EchisCarinatus assay was 2 U dL)1 with results of 26–49 U dL)1 by other FII assay techniques.

Acquired deficiency of prothrombin may rarely beassociated with the presence of antibodies and lupusanticoagulant effects. These patients may experiencebleeding.

The diagnosis of mild prothrombin deficiency maybe especially difficult in premature or young neonateswhere vitamin K deficiency may complicate assess-ment. Reassessment after vitamin K replacement maybe necessary.

Molecular defects

Prothrombin is encoded by a gene on chromosome11. To date at least 32 different mutations have beenidentified in patients with prothrombin deficiencyand these have recently been reviewed [50]. Seven-teen mutations have been described in dysprothrom-binaemia, all of which are missense mutations. Themutations in hypoprothrombinaemia are also pre-dominately missense, although nonsense mutationshave also been found.

In dysprothrombinaemia the mutations result inamino acid substitutions within the cleavage sites forFXa and the serine protease region of prothrombin,whereas the mutations in hypoprothrombinaemia areoften close to the Gla and kringle domains and the Achain.

Management

Treatment options There are no specific prothrombinconcentrates available and prothrombin complexconcentrates are therefore treatment of choice. Themajority of these products are 3-factor concentratescontaining therapeutic quantities of FII, FIX and FX[1]. The 4-factor concentrates, which in additioncontain FVII, are also available [1]. These concen-trates contain known quantities of these coagulationfactors although the potency of the vial is usuallyexpressed in terms of the FIX content. There isusually approximately 1 unit of prothrombin perunit of FIX and this can be used as a basis for dosagecalculations. In the absence of an appropriate 3- or 4-factor concentrate, virally inactivated fresh frozenplasma (FFP) is an alternative source of prothrombin.Management of bleeding and surgery – It should benoted that there is only very limited published dataavailable on which to base therapeutic decisions.

It is estimated that 1 unit of prothrombin will raisethe plasma prothrombin level by 1 IU dL)1. Relat-ively low levels of prothrombin (20–30 IU dL)1) are

thought to be required for normal haemostasis anddoses of 20–30 IU kg)1 have been used previouslyand seem to be effective. Higher doses may, however,be required in the event of life-threatening bleedingor major surgery, and monitoring of prothrombinlevels should be performed. The half-life of pro-thrombin is around 72 h, which facilitates relativelyinfrequent dosing, usually every 2–3 days.Management of pregnancy – Other than a small num-ber of reports of postpartum haemorrhage there is nopublished data on the management or outcome ofpregnancy in prothrombin deficiency. It is thereforedifficult to make firm recommendations on preg-nancy management but it would seem reasonable toincrease the prothrombin level to above 25 IU dL)1

prior to delivery.Management of neonates and children – There does notappear to be a high incidence of severe bleedingduring the neonatal period although umbilicalbleeding has been reported. Prophylactic replace-ment therapy is therefore not routinely recommen-ded for this age group.

The use of prophylaxis in older children should bebased on the frequency and type of bleeding. Whererecurrent joint bleeding is a feature prophylaxisshould be used to prevent the development of achronic arthropathy.

Factor V deficiency

Molecular biology

Clotting FV is a large glycoprotein of MW 249 kDaencoded for by a gene on chromosome 1, which issynthesized by hepatocytes and megakaryocytes. FVand FVIII have approximately 40% amino acidhomology in their respective A and C domains andtheir overall protein structures are identical. Plateletscontain approximately 20% of total circulating FV.FV is activated by thrombin and the resultingheterodimer FVa acts as a cofactor for FXa in theconversion of prothrombin to thrombin. At the timeof writing 26 separate mutations have been charac-terized [51]. Twelve of these are located in the exon(exon 13), which encodes the entire B domain. Themajority of defects appear to give rise to a quanti-tative disorder whereas qualitative defects have beenreported in approximately a quarter of cases [46,52].

Clinical phenotype

Hereditary FV deficiency is a very rare autosomalrecessive condition. Owren described the first case, aNorwegian lady, in 1947 [53]. The prevalence of the



homozygous state is approximately 1 per million[54]. Parental consanguinity is often present. It hasbeen reported that in homozygote individuals FVlevels range from <1 to 10 U dL)1 with a normalrange of 71–125 U dL)1 [55]. Homozygous defici-ency is associated with a moderately severe bleedingdisorder [55]. It usually presents in childhood witheasy bruising and mucous membrane bleeding, inparticular epistaxes and oral cavity bleeding. Hae-marthroses and muscle haematomas are less of afeature than in FVIII deficiency and are often relatedto trauma rather than being spontaneous. Gastroin-testinal bleeding and haematuria may occur rarely.Postoperative, postdental extraction and postpartumbleeding have been reported. There have beenoccasional reports of intracranial bleeding especiallyin the antenatal and neonatal periods [56–59]. Twoof these cases were complicated by the developmentof FV inhibitors following plasma infusions and oneof the children died from repeated intracranialhaemorrhage.

Diagnosis

The FV deficiency is associated with prolongation ofboth the PT and APTT but a normal TT. Both PTand APTT are corrected by mixing with normalplasma. Deficiency of FV is confirmed by performinga prothrombin time-based FV assay or by immuno-logical assessment of FV levels. Individuals withreduced FV levels should also have a FVIII assayperformed to exclude combined FV and FVIII defi-ciency.

Management

There is no FV concentrate available for the treat-ment of FV deficiency. The only blood productavailable for FV replacement is FFP. It is recommen-ded that a virally inactivated preparation is used [1]preferably sourced from a country that has noreported case of variant CJD in its donor populationor BSE in its cattle. At the time of writing two virallyinactivated FFP products are available in the UK.These are methylene blue-treated single unit FFP(MB FFP), prepared by the National Blood Service(England and Wales) and the Scottish NationalBlood Transfusion Service (SNBTS) and a commer-cial product [60], a pooled plasma product virallyinactivated using a solvent detergent technique (SD-plasma). The mean FV level in units of MB FFP is80 U dL)1 (NBA, personal communication). As SD-plasma is a pooled product individual units of eachbatch have the same FV level overcoming the

problem of variability of single unit FFP enabling amore consistent dosing strategy. Lot release criteriaof this product require that each batch contains atleast 70 U dL)1 of clotting FV, FVII, FX, FXI andFXIII.

Spontaneous bleeding episodes The minimum circula-ting level of FV that has to be achieved for effectivehaemostasis is likely to vary from individual toindividual but it has been reported to be not less than15 U/dL)1 [46]. In order to achieve this level it issuggested that a dose 15–20 mL kg)1 of FFP isadministered to patients presenting with a bleedingepisode. Use of agents such as tranexamic acidshould also be considered.

It is recommended that the FV level be measuredfollowing FFP administration to ensure that theminimum haemostatic level of 15 U dL)1 has beenachieved. If not, a further dose of FFP should begiven. If bleeding continues despite achieving anadequate FV level a further dose of FFP should beconsidered. Close monitoring of FV levels is recom-mended to determine the FV half-life in individualpatients and guide further FFP administration whenlevels fall below 15 U dL)1 if clinically indicated.

In a recent review of the use of SD-plasma in themanagement of patients with single hereditary factordeficiency a dose of 15 mL kg)1 was used to treat 51bleeding episodes in 12 patients with single factordeficiencies. This included 34 episodes in six FV-deficient individuals [60]. Bleeds included haemart-hoses, haematomas, dental, gastrointestinal andgynaecological haemorrhages. Effective haemostasiswas achieved in 49 of 50 episodes assessed (98%)with an average dose of SD-plasma of 4.7 units(940 mL). It has to be appreciated that although ahaemostatically effective circulating level of FV maybe obtained with a particular dose of FFP this levelmay not be sufficient to normalize the PT. In oneneonatal case an FFP dose of 30 mL kg)1 twice dailywas required to achieve correction of the PT [57]. Ithas been suggested that in cases of severe bleeding notcontrolled with FFP replacement platelet transfusionsmay be considered, although proven efficacy of thistreatment modality is lacking in the literature [61].Platelets provide a concentrated supply of FV, which,following a-granule release upon platelet activation,can presumably bind immediately to surface recep-tors optimizing prothrombinase complex activity.The use of recombinant activated factor VII (rFVIIa)should also be considered in patients not respondingto FFP. This product has recently been used success-fully in a FV-deficient patient [62]. However, inpatients with FV levels of <1 U dL)1 the FV level may



be a major rate limiting factor in thrombin generationand rFVIIa may not be effective.

Surgery A trial of procedure may be considered inindividuals with a partial deficiency of FV undergo-ing surgical or dental procedures in whom there is noprior history of spontaneous or surgery-relatedbleeding. FFP should be administered if excessivebleeding occurs. In patients with partial deficiency ofFV who have a bleeding history, and in patients withlevels of <1 U dL)1, FFP should be administeredimmediately prior to the procedure. FFP dosagesshould be those recommended for bleeding episodes.FV levels should be measured following FFP andfurther units administered prior to surgery if theminimum haemostatic level has not been achieved.The FV level should be observed closely followingthe procedure and further doses of FV administeredto maintain a factor level above 15 U dL)1 untilwound healing is established. Close monitoring ofcardiovascular function is advised postoperativelybecause of the possible risk of fluid overload asso-ciated with repeated FFP doses, a problem describedin some surgical cases [55].

In the study reported by Horowitz and Pehta [60]SD-plasma at an initial dose of 15 mL kg)1 providedeffective surgical prophylaxis in seven procedures onfive FV-deficient patients. Data available for six ofthese episodes showed an average increase in FV levelof 13 U dL)1 (range: 9–17 U dL)1). Unfortunatelypatient baseline FV levels and follow-up FFP dosageschedules postsurgery were not reported.

Pregnancy There are no available data on the man-agement of FV deficiency in pregnancy. In patientswith <1 U dL)1 FV we recommend the administra-tion of FFP at the dosage suggested above for surgeryas soon as the patient is in established labour, withclose monitoring of FV levels. Further doses of FFPshould be administered if and when required tomaintain minimum haemostatic FV levels until afterdelivery. If Caesarean section is performed FFPdosing should be continued until wound healing isestablished. Patients with a partial deficiency and nohistory of bleeding during invasive procedures couldbe managed expectantly.

Neonates Severe and moderate FV deficiency can bediagnosed using cord or peripheral blood: FV levelsdo increase in the first month of life and milddeficiencies may need to be confirmed when the childis older.

Bleeding in the neonatal period appears to beuncommon [55] although it must be emphasized that

FV deficiency can be a cause of intracranial haemor-rhage in this age group (see above). A small numberof single case reports have described this occurrence,with FV levels of the affected neonates varyingbetween 2 and 8 U /dL)1 [52,56–58,63]. This diag-nosis should therefore be considered for babiespresenting with intracranial haemorrhage and abnor-mal coagulation. Treatment of bleeding in neonateswith FV deficiency requires the administration ofvirucidally treated FFP at a dose of 15–20 mL kg)1

in order to increase the FV level to >15 U dL)1. Fluidoverload is a potential problem, particularly whererepeated infusions are required and plasma exchangemay be necessary in such cases. Platelets provide analternative source of FV and platelet transfusion maybe a valuable therapeutic addition to FFP. As yet,there have been no reports of rFVIIa used in thetreatment of neonatal haemorrhage secondary to FVdeficiency, although adult experience would suggestthis to be worth considering, particularly where fluidbalance problems exist [61].

Babies found to have a FV deficiency of<15 U dL)1 should have a cranial ultrasound per-formed in the first few days of life to excludeintracranial haemorrhage. There is no evidence tosuggest that prophylactic infusion of FFP is of valuein asymptomatic FV-deficient neonates. Vitamin Kcan be given i.m. to babies with FV levels>15 U dL U dL)1, oral or i.v. vitamin K can begiven to more severely affected individuals.

Factor V inhibitors

Development of alloantibodies to FV in FFP is apotential complication of hereditary FV deficiency.Although very few cases have been reported in theliterature this may represent under-reporting. Theoccurrence of inhibitors, especially transient ones oflow level, following FFP replacement therapy maynot be uncommon. In the event of a bleedingproblem it has been suggested that low-level inhib-itors can be neutralized using large amounts of FFP[61]. However, as in the treatment of surgical casesthere are concerns over fluid overload in thissituation and close cardiovascular monitoring isadvised. Intravenous immunoglobulin may be effect-ive in eradicating the FV inhibitor [64].

It has been shown that in acquired FV deficiencydue to the development of FV autoantibodies,platelet FV is relatively protected from antibodyneutralization [65]. Platelet infusions have beenreported to be effective in acquired FV deficiency[66] and may therefore also be an effective treatmentalternative in patients with hereditary deficiency



complicated by inhibitors. Prophylactic use ofweekly platelet infusions was effective in one inhib-itor patient in stabilizing a subdural haematoma[57]. Use of rFVIIa concentrate should also beconsidered.

Combined deficiency of factors V and FVIII

Molecular biology

Combined FV and FVIII deficiency is a rare auto-somal recessive disorder which usually arises as aconsequence of consanguinity [67]: Oeri first repor-ted it in 1954 [68] and affected individuals havereduced plasma levels of both FV and FVIII. Studiesof the inheritance pattern indicated that it was likelyto be due to a single gene defect rather than due tocoinheritance of separate defects of the FV and FVIIIgenes. This has been confirmed, the gene beinglocated on the long arm of chromosome 18. The geneencodes a resident protein of the endoplasmic reti-culum, the golgi intermediate compartment, termedthe ERGIC-53 protein [69–74]. This protein has beenidentified as playing a major role in intracellulartrafficking of certain proteins including FV and FVIII[75–77]. Although it would appear that withinhepatocytes FV and FVIII are synthesized normally,defective ERGIC-53 function results in disturbanceof the passage of the factors through the cell andimpaired release into the circulation. A number ofdifferent mutations have been described in this geneleading to combined FV and FVIII deficiency [69,71].

Clinical phenotype

Although mild bleeding symptoms such as easybruising and epistaxes are not uncommon in affectedindividuals, circulating levels of FV and FVIII areusually sufficient to prevent more severe spontaneousbleeding episodes. However, bleeding is commonfollowing surgery, dental extraction and trauma;menorrhagia and postpartum haemorrhage arecommonly seen in affected women [78,79]. Thebleeding severity observed in affected individuals hasbeen reported as similar to that in individuals withdeficiencies of either factor alone at similar levels[79].

Diagnosis

The combined deficiency disorder is associated witha prolongation of both the PT and APTT, with theAPTT prolongation disproportionate to that of thePT. Both test times are corrected by mixing studies

using normal plasma. APTT-based activity assaysand antigen assays reveal levels of between 5 and20 IU dL)1 for both FV and FVIII [67].

Management

Bleeding episodes The treatment of bleeding episodesis dependant on the nature of the bleed and theaffected individual’s FV and FVIII levels. Sponta-neous bleeding episodes occurring in patients withcombined FV and FVIII deficiency should be treatedwith both FVIII concentrates and FFP (as a source ofFV). For minor bleeding episodes FVIII levels shouldbe raised to at least 30 IU dL)1 and to at least50 IU dL)1 for more severe bleeds with rFVIIIconcentrate being the treatment of choice. As forpatients with FV deficiency, FFP should be adminis-tered in order to increase the FV level to at least25 U dL)1. The FFP product of choice is virallyinactivated FFP.

Surgical procedures These should be covered withFVIII concentrates administered 12 hourly to main-tain FVIII levels above 50 IU dL)1 and 12 hourlyFFP to achieve minimum levels of FV of 25 U dL)1

until wound healing is established.

Pregnancy There are no published data on the man-agement of pregnant women with this combineddeficiency. FV levels in pregnancy do not consis-tently increase or decrease whereas FVIII levels willincrease throughout pregnancy: any possible bleed-ing is therefore likely to be dependent on the FV levelduring labour and postdelivery. In keeping withgeneral recommendations for the management ofbleeding disorders in pregnancy, affected womenshould be managed by an obstetric unit in closeliaison with a haemophilia centre. FV and FVIIIlevels should be confirmed in the third trimester sothat the delivery can be planned with regard tohaematological intervention. FV levels should bemaintained above the haemostatic level of 15 U dL)1

during labour, using virucidally treated plasma as thesource of the factor. FVIII levels should remain at>50 IU dL)1 throughout this period. If Caesareansection is carried out, it would be prudent tocontinue FV replacement until wound healing hashealed in women with FV levels of <15 U dL)1.Epidurals can be performed if the woman has FVlevels of >15 U dL)1 and FVIII >50 IU dL)1.

Neonates Combined FV and FVIII deficiency can bediagnosed in the neonatal period using cord orperipheral blood. Affected babies should receive oral



rather than i.m. vitamin K and go on to receive s.c.childhood vaccinations. They should be managedexpectantly and there is no indication for routineprophylaxis with plasma and FVIII. Neonatal intra-cranial haemorrhage has not been described in thiscondition.

Factor VII deficiency

Molecular biology

Factor VII is a vitamin K-dependent glycoproteinwith a MW of approximately 50 kDa and circulatesin plasma in two forms – the majority in a singlechain inactive form with a concentration of 10 nmo-les L)1 (0.5 lg mL)1) and a much smaller amount(approximately 10–110 pmoles L)1) as the activetwo-chain form. The FVII gene maps to chromosome13 at 13q34, spans approximately 12 kb of DNAand consists of nine exons encoding a mature proteinof 406 amino acids. FVII through its interaction withtissue factor is fundamental to the initiation ofcoagulation.

Plasma factor VII levels

Factor VII plasma levels are determined by bothenvironmental and genetic factors with the latteraccounting for up to one-third of the variation inplasma FVII. Amongst environmental factors, dietaryfat intake and the levels of plasma triglycerides arepositively correlated with FVII:C levels but age,obesity, diabetes and in women, the use of sexhormones can all influence FVII levels.

Five and possibly six polymorphisms within thehuman F7 gene have been shown to affect bothplasma FVII:C levels and plasma VIIa levels [re-viewed in 80]. Of these the most important is theArg353Gln polymorphism within exon 7, whichaffects approximately 20% of the UK population.Heterozygosity for this polymorphism is associatedwith an approximately 25% reduction in FVII:C andVII:Ag levels and homozygosity, an approximately50% reduction in circulating plasma FVII.

Inherited factor VII deficiency

Factor VII deficiency is the most common of the �rareinherited coagulation disorders�. Severe FVII defici-ency (FVII:C <2 IU dL)1) has an estimated preval-ence of 1:300 000–1:500 000. FVII deficiency isinherited in an autosomal recessive manner and itsfrequency is significantly increased in countrieswhere consanguineous marriage is practised. There

is a relatively poor correlation between absolute FVIIlevels and the risk of bleeding with some individualswith very low FVII levels exhibiting very fewsymptoms whilst others with much higher levels,have a significant bleeding diathesis.

Clinical phenotype

The spectrum of bleeding problems in FVII defici-ency is very variable [81]. Epistaxes, gum bleeding,menorrhagia and other mucous membrane-typebleeding are common. Menorrhagia and chroniciron deficiency are frequently seen in women withFVII deficiency. Joint bleeds have been reported inpatients with severe FVII deficiency although this isnot a consistent finding. In patients with severeFVII deficiency (FVII:C <2 IU dL)1), bleeding intothe central nervous system is common and reportedin between 15 and 60% of cases [82]. Such casesoften present shortly after birth and this presenta-tion is associated with a high morbidity andmortality.

Diagnosis

The diagnosis of FVII deficiency is suspected follow-ing the finding of a prolonged PT, which corrects,unless an inhibitor is present, in a 50:50 mix withnormal plasma. The APTT, TT and FI concentrationare normal. FVII levels are low at birth and age/gestation-related ranges for FVII are essential if adeficiency is suspected [10].

It is important to exclude vitamin K deficiency orother acquired causes of a clotting disorder beforethe diagnosis of FVII deficiency is made. A thera-peutic trial of vitamin K may be of value. In somecases, family studies may be helpful in establishingthe diagnosis of FVII deficiency. The diagnosis ofFVII deficiency may be especially difficult in prema-ture or young neonates where vitamin K deficiencymay complicate assessment. Reassessment after vita-min K replacement may be necessary.

Functional FVII assays Functional FVII activity(FVII:C) is measured using a one-stage PT-basedassay. In some unusual cases of FVII deficiency, theremay be a discrepancy in the levels of FVII:Cdepending upon the source of thromboplastin [83].The use of a human thromboplastin is advised on thebasis that the results are more likely to reflect in vivoFVII levels. In the functional FVII assay, there islimited conversion of FVII–FVIIa and FVII:C assays,therefore, measure both the inactive zymogen andpreformed FVIIa. Blood samples for determination of



FVII:C should not be stored at 4 �C because this canlead to cold activation of FVII and lead to substantialoverestimation of the true FVII level [84].

Immunological FVII assays FVII antigen (FVII:Ag) isfrequently measured using an enzyme-linked immu-nosorbent assay (ELISA) or4 Immunoradiometricassay (IRMA) assay and either monoclonal or poly-clonal antibodies. Such assays can detect as little as0.01 IU dL)1 of FVII. Immunological assays shouldnot be used in preference to a functional FVII assay.

Factor VIIa assays In samples where there has beenno activation of FVII, assays for FVII:C and FVII:Agshould be equivalent. Two direct assays for FVIIahave been reported but for diagnosing FVII defici-ency [85,86] there is little if any value in measuringbasal FVIIa levels in patient with FVII deficiency.

FVII deficiency and thrombosis

Thrombosis in association with FVII deficiency hasbeen reported although the mechanism is unclear[80].

Inherited factor VII deficiency

The FVII mutation website (http://www.193.60.222.13/index.htm) lists 120 mutations through-out the FVII gene. In common with many diseases,numerous examples of repeated mutations within theFVII gene have been reported.

Management

Current therapeutic options to manage patients withFVII deficiency include fibrinolytic inhibitors, plas-ma, intermediate purity FIX concentrates (prothrom-bin complex concentrates), FVII concentrates andrFVIIa. Plasma FVII has a short in vivo half-life ofapproximately 5 h although this may be shorterduring a bleeding episode [87]. For severely affectedindividuals (FVII:C <2 IU dL)1), rFVIIa is the treat-ment of choice [1] although it is not licensed, as yet,for this indication.

Currently available treatments

Plasma: Fresh Frozen Plasma (FFP) has been widelyused in the management of FVII deficiency althoughthere is little information available on its efficacy,and it should not be used unless there is noalternative. FFP has been used to manage patientsundergoing various surgical operations either by

itself or in combination with FVII concentrate[88,89]. In cases in which prolonged administrationis required, it is difficult to administer sufficient FFPquickly enough and problems with fluid overloadmay be encountered. FFP as a source of FVII is notrecommended for managing patients with FVIIdeficiency.Factor IX concentrates and prothrombin complex

concentrates: Intermediate purity FIX concentratescontain variable amounts of FVII and have beensuccessfully used to manage patients with FVIIdeficiency [90–92]. The amount of FVII in theseconcentrates is very variable but usually indicated bythe manufacturer. These concentrates also containactivated forms of FVII, FIX and FX and should beused with caution, as there are reports of bothvenous and arterial thromboses in association withtheir use. They should be avoided in patients withliver disease, in patients with major trauma, inpatients with antithrombin deficiency and in theneonate whose liver is relatively immature. Pro-thrombin complex concentrates are not recommen-ded for managing patients with FVII deficiency.Factor VII concentrates: Plasma-derived FVII

concentrates (5 BPL Elstree, UK and Baxter) have beensuccessfully used to manage patients with FVIIdeficiency with spontaneous bleeds; for patientsundergoing a variety of surgical procedures and forprophylaxis against bleeds in children with severeFVII deficiency [93–95]. These products are virallyinactivated to reduce the risk of viral transmission.Recombinant factor VIIa: Patients with FVII

deficiency can be safely managed using rFVIIa [96–102] and this is the treatment of choice [1] althoughthere is limited published data. The rFVIIa probablyhas a shorter half-life than that of plasma FVIIparticularly in children where an increased clearancehas been demonstrated and in pregnancy.Fibrin glue: May be effective in facilitating local

haemostasis. Currently two products are licensed foralthough one of these is only licensed for use in liversurgery [1].Tranexamic acid: In practise, 10 mL of a 5%

solution of tranexamic acid is used as a mouthwashevery 8 hourly. However, this is not commerciallyavailable and will require to be formulated by thePharmacy Department. In women with menorrhagia,tranexamic acid 15 mg kg)1 8 hourly (in practise1 g, 6–8 hourly) may be effective when taken for theduration of the menstrual period.

Acute bleeding episodes Efficient haemostasis can beachieved with levels of FVII:C in the range of10–15 IU dL)1. rFVIIa is the recommended choice



for patients with FVII deficiency requiring replace-ment therapy.Factor VII concentrates – Factor VII concentrates havebeen successfully used to treat a variety of acutebleeding problems:1 Mucosal bleeding: A correction to 40–100%

resulted in cessation of bleeding within 15 min inall cases [103].

2 Intracranial bleeding: An 8-year-old boy receiveda dose of 37 IU kg)1 FVII concentrate every 6 hwith peak FVII levels of approximately 100% andtroughs as low as 4% over the 11-day treatmentperiod. A 37-year-old adult male with intracranialbleeding received alternating doses of 16 and8 IU kg)1 every 6 h for 10 days with peak FVIIlevels in the upper 30s (%). The peak FVII levelduring surgical coverage with FVII concentratewas approximately 100% in all cases, with troughlevels ranging from 8 to 65% over treatmentperiods of 24 h to 16 days using treatment inter-vals of 6–12 h.

3 Prophylaxis: FVII concentrates have been success-fully used for prophylaxis against bleeds in chil-dren with severe FVII deficiency. Suggestedguidelines for the administration of FVII for long-term prophylaxis are in the range of 10–50 IU kg)1

one to three times a week (grade B recommenda-tions based on level III evidence). Although illo-gical when once considers the short half-life ofFVII, in practice it appears successful [103].

rFVIIa – In the study of Mariani et al., rFVIIa wasused to treat 17 patients (FVII:C <1% in ninepatients and <5% in an additional five patients) with27 spontaneous bleeding episodes who in additionalso underwent seven major and 13 minor surgicalprocedures [104]. Fifteen haemarthroses were treatedwith only a single dose of rFVIIa (14–30 lg kg)1)and in only one case was rFVIIa ineffective.

Management of surgery The minimum level of VII:Crequired for haemostasis is not known with certaintybut levels of 10–15 IU dL)1 are generally sufficientto achieve haemostasis. Recent work using thethrombin generation test (TGT) has suggested thatlevels of 2 U dL)1 are sufficient to normalize theTGT although this has not been used to monitorreplacement therapy [105].Factor VII concentrate – For surgery, doses of FVIIconcentrate ranging from 8 to 40 U kg)1 every 4–6 hhave been successfully used [89,106–109]. Carefulmonitoring of patients receiving FVII concentrates isessential to prevent excessively high peaks andtroughs. For major surgery trough FVII levels shouldnot fall below 20 IU dL)1.

rFVIIa – Mariani et al. have reported their experi-ence with rFVIIa in treating 17 patients with 27spontaneous bleeding episodes who in addition alsounderwent seven major and 13 minor surgical pro-cedures [104]. Nine of these patients had FVII levelsof <1% and a further five, FVII levels <5%. Fifteenhaemarthroses were treated with only a single doseof rFVIIa (14–30 lg kg)1) and in only one case wasrFVIIa ineffective. Seven major surgical procedureswere performed in severely affected patients undercover of rFVIIa and no bleeding occurred eitherduring or after surgery. In the case of surgery,rFVIIa was given at 2–3 h intervals for the first 24 hfollowed by longer intervals (3–8 h) for theremaining postoperative period. All the surgicalpatients also received tranexamic acid alongwith the rFVIIa. In one patient rFVIIa becameineffective following the development of an anti-FVII antibody.

Other groups have reported the successful use ofrFVIIa with a variety of clinical problems[96,99,101,102]. A dose of 20–25 lg kg)1 admini-stered every 4–6 h appears to be successful intreating the majority of patients with FVII defici-ency who are either bleeding or who requiretreatment prior to and following surgery (grade Brecommendations based on level III evidence). Thisshould be combined with tranexamic acid10 mg kg)1 every 8 h i.v. or 25 mg kg)1 every 8 horally. For dental surgery, 10 mL of a 5% solutionof tranexamic acid can be used as a mouthwashevery 8 h for 5–7 days.

In patients receiving 20 lg kg)1 of rFVIIa every6 h, this leads to peak FVII levels of 430–830 IU dL)1 and trough levels of 30 IU dL)1

(although the results are highly dependent upon thethromboplastin used to assay the FVII). Inhibitorsfollowing treatment with rFVIIa have been reportedbut appear rare [110].

Pregnancy There are no published data available onthe management of FVII deficiency in pregnancy.Continuous infusion of rFVIIa has been used in onepatient with FVII deficiency (VII:C 3.7 U dL)1l) toprovide haemostatic cover for an elective Caesareansection [111]. Pharmacokinetic studies performedprior to the infusion revealed a very high clearancerate (0.208 L h)1 kg)1) and a short half-life(0.884 h), which the authors attributed to the preg-nancy. The patient received an initial bolus or rFVIIaof 13.3 lg kg)1 followed by a continuous infusioninitially at a rate of 3.3 lg kg)1 h)1 for 48 h andthen at 1.66 lg kg)1 h)1 for a further 48 h. PlasmaFVII and FVIIa levels were maintained between 100



and 150 U dL)1 and no bleeding problems wereobserved.

The neonate Factor VII deficiency may be difficult todiagnose in the neonatal period because of the lowphysiological levels in the newborn. For thesereasons age- and gestation-related reference rangesmust be used [10]. In cases where FVII levels arelower than expected, screening the parents may be ofbenefit in establishing a diagnosis.

Families in which both parents are known to haveFVII deficiency, pregnancy and delivery should bemanaged in such a way as to minimize the potentialrisk of bleeding both to the mother and baby. Thisrequires close liaison with the obstetric unit includingobstetric anaesthetists and a management planshould be prepared for the delivery and subsequentinvestigation of the neonate. FVII levels may riseduring pregnancy. A FVII assay should be performedprior to delivery. At birth a cord blood sampleshould be taken for FVII assay.

Cranial ultrasound should be undertaken inseverely affected neonates because of the increasedrisk of intracranial haemorrhage.

Prophylaxis during the neonatal period may benecessary in severely affected neonates although evenwith appropriate replacement therapy, haemorrhagemay occur.

Factor X deficiency

Molecular biology

Factor X occupies a unique position in the coagula-tion cascade, as the first enzyme in the commonpathway of thrombus formation. The gene for FX is22 kb long and located at 13q34-ter, 2.8 kb down-stream of the FVII gene and encodes a protein of MW59 kDa. FX synthesis occurs in the liver and follow-ing secretion into plasma, circulates at a concentra-tion of 10 lg mL)1 as a two-chain molecule.Activation of FX–FXa occurs during the initiationof coagulation by a complex consisting of tissuefactor, FVIIa, calcium ions and a suitable phosphol-ipid membrane. However, activation can also takeplace through FIXa, FVIIIa, calcium ions and acidicphospholipid surfaces. Activation of FX in vitro canoccur by the direct action of a metalloproteinasefound in the venom of Vipera russelli. Physiologically,FXa is the most important activator of prothrombin.The presence of the prothrombinase complex (FXa,FVa, Ca2+ and a suitable negatively charged phosp-holipid membrane) accelerates the conversion ofprothrombin to thrombin 280 000-fold.

Inherited factor X deficiency

Severe (homozygous) FX deficiency is inherited asan autosomal recessive disorder with an incidenceof 1:1 000 000 in the general population. Theprevalence of FX deficiency is greater amongstpopulations in which consanguineous marriage iscommon. The prevalence of heterozygous FX defi-ciency is about 1:500, but individuals are usuallyclinically asymptomatic [112]. However, someheterozygotes do have a significant bleeding ten-dency and this may be due to either insufficientenzymatic activity by wild-type FX or inhibition ofone of the reactions in the coagulation pathway bya mutant protein.

Clinical phenotype

Although FX deficiency produces a variable bleedingtendency, patients with severe deficiency [FX activity(FX:C) <1 IU dL)1] tend to be the most seriouslyaffected patients with rare coagulation defects [113].

Patients with FX deficiency may present at any ageand severely affected individuals (FX:C <1 IU dL)1)can present in the neonatal period, with umbilical-stump bleeding. The most frequent symptom in FXdeficiency is epistaxis and is seen with all severitiesof deficiency. Other mucosal-type bleeding is lessfrequent and occurs mainly in patients with severedeficiencies. Menorrhagia occurs in half the womenof reproductive age. Haemarthroses, severe postop-erative haemorrhage and central nervous systemhaemorrhage have been reported. Recurrent hae-marthroses may result in severe arthropathy. Mod-erately affected patients (FX:C 1–5 IU dL)1) maybleed only after haemostatic challenge, e.g. traumaor surgery. Mild FX deficiency (FX:C 6–10 IU dL)1)may be identified incidentally during routine screen-ing or family studies. Patients who are only mildlyaffected may experience easy bruising or menorrha-gia.

Diagnosis

The diagnosis of FX deficiency is suspected followingthe finding of a prolonged PT and APTT whichcorrects, unless an inhibitor is present, in a 50:50 mixwith normal plasma. The diagnosis of FX deficiencyis confirmed by measuring plasma FX levels. Fivedifferent assays are available for measuring plasmaFX levels – the one-stage PT- and APTT-basedassays, a chromogenic assay, an assay employingRussell viper venom (RVV) and finally an immuno-logical assay. The one-stage PT- or APTT-based



assay is sufficient for the diagnosis of FX deficiency.In some cases the results obtained using a PT-basedFX assay may differ according to the type ofthromboplastin used. Chromogenic assays can gen-erate normal values with some dysfunctional FXvariants (type II variant: normal immunological FXbut reduced functional activity) and should not beused as a screening test for FX deficiency. The fullspectrum of tests is only required for detailed proteincharacterization.

Factor X levels are low at birth and age/gestation-related ranges for FX are essential if a deficiency issuspected [10].

It is important to exclude vitamin K deficiency orother acquired causes of a clotting disorder beforethe diagnosis of FX deficiency is made. A therapeutictrial of vitamin K may be of value. In some cases,family studies may help in establishing the diagnosisof FX deficiency.

The diagnosis of mild FX deficiency may beespecially difficult in premature or young neonateswhere vitamin K deficiency may complicate assess-ment. Reassessment after vitamin K replacement maybe necessary.

Inherited factor X deficiency

Factor X mutations are thought to be rare because ofthe central role of FX in the coagulation cascade. TheFX knockout mouse has a lethal phenotype, withdeath occurring either in utero or within a few daysof life and this is consistent with the hypothesis that acomplete absence of FX is a lethal disorder [114]. Amutational website for FX mutants does not exist butapproximately 45 mutations within the FX gene havebeen reported [115].

Management

Inherited FX is a rare disorder and there are nogenerally agreed guidelines for the management ofthis disorder. Current therapeutic options to managepatients with FX deficiency include fibrinolyticinhibitors, plasma and intermediate purity FIX con-centrates (prothrombin complex concentrates). rVIIahas been used successfully to treat acquired FXdeficiency secondary to amyloidosis [116].

The need for replacement therapy is guided by theparticular haemorrhagic episode. The biological half-life of FX is 20–40 h [117], so an adequate level canbe achieved with repeated infusions. Factor levels of10–20 IU dL)1 are generally sufficient for haemos-tasis, even in the immediate postoperative period[118].

Treatment products

Tranexamic acid: Particularly useful for mucosalbleeding. In practise 10 mL of a 5% solution oftranexamic acid is used as a mouthwash every 8 h.However, this is not commercially available and willrequire to be formulated by the Pharmacy Depart-ment.Fibrin glue: May be effective in facilitating local

haemostasis.FFP: A virally inactivated plasma should be used.

20 mL kg)1 followed by 3–6 mL kg)1 twice daily,aiming to keep X:C trough levels above 10–20 IU dL)1.Prothrombin complex concentrates (for details see

[1]): The calculated required dosage for treatment isbased on the empirical finding that 1 IU FX kg)1

bodyweight (bw) raises the FX level by 1.5% ofnormal. Tranexamic acid should not be used con-currently with prothrombin complex concentratesbecause of the risk of thrombosis. The half-life of FXis 60 h and daily treatment is not usually required.However, in cases where replacement therapy isgiven, levels should be monitored on a daily basis.Prothrombin complex concentrates should be usedwith caution, if at all, in patients with concomitantliver disease, large haematomas, major trauma, theneonate and antithrombin deficiency [119].

Concentrates are heat-treated, which significantlyreduces the risk of viral transmission.

Mucosal bleeding Tranexamic acid is particularlyuseful (see above). In women with menorrhagia,tranexamic acid 15 mg kg)1 8 hourly (in practise1 g, 6–8 hourly) may be effective when taken for theduration of the menstrual period.

Management of the acute bleed No specific concen-trates are available and prothrombin complex con-centrates are the treatment of choice.Fibrin glue: May be effective in facilitating local

haemostasis.Plasma: In cases where prothrombin complex

concentrates are not available or are contraindicated,FFP can be used (details as above).Prothrombin complex concentrates: Prothrombin

complex concentrates have been used as regularprophylaxis in patients with severe FX deficiency[120]. Kouides et al. reported the case of a patientwho received 30 units kg)1 of Profilnine adminis-tered twice weekly as part of a home treatmentprogramme. If breakthrough bleeding occurred,another dose was administered but no more thantwo doses in 24 h or on more than 3 consecutive



days. A trough level drawn 48 h postinfusion showedan FX level of 30 U dL)1. In a 12-month follow-upthere were no bleeding episodes reported. A recentpaper [121] reported four cases of FX deficiency inwhich primary prophylaxis was successfully under-taken using a PCC commencing in the case of onechild at the age of 1 month. The four cases reportedall had severe FX deficiency and presented within24–72 h of birth with a severe bleeding diathesis.

Unpublished data at the Royal Free Hospital in apatient with severe FX deficiency (FX:C <1 IU dL)1)has shown that 10 U kg)1 of HT Defix (SNBTS)given every third day provides effective prophylaxisagainst bleeds.rFVIIa: rFVIIa has been used to treat amyloid-

associated FX deficiency [116] but the data on its usein other patients with FX deficiency is limited.Adequate levels of FX appear to be important forthe action of rFVIIa and therefore, in severe FXdeficiency rFVIIa may be ineffective [122].

Surgery Surgery in individuals with severe FX defici-ency (FX <1 IU dL)1) has been successfully per-formed following infusion of either FFP or PCCs. Inthe case of FFP, a level of 35 U dL)1 was achievedprior to surgery and FX levels were maintainedabove 20 U dL)1 in the postoperative period with nobleeding reported [118]. A FX level of 20 U dL)1

appeared to be sufficient for efficient haemostasis.However, a recent report has shown that a FX levelabout 0.05 IU mL)1 (5 IU dL)1) achieves adequatehaemostasis [121]. Furthermore, this report alsofound a variation in FX clearance in affected malesthat affected the frequency with which they receivedtreatment.

Pregnancy Factor X levels increase during pregnancy[123], but women with severe FX deficiency and ahistory of adverse outcome in pregnancy may benefitfrom aggressive replacement therapy. However, thepotential for thrombosis associated with replacementtherapy must be carefully evaluated.

Neonates Factor X deficiency may be difficult todiagnose in the neonatal period because of the lowphysiological levels in the newborn, although severedeficiency should not give rise to doubt. Age- andgestation-related reference ranges must be used [10].In cases where FX levels are lower than expected,screening the parents may be of benefit in establishinga diagnosis.

In families in which both parents are known to haveFX deficiency, pregnancy and delivery should bemanaged in such a way as to minimize the potential

risk of bleeding both to the mother and baby. Thisrequires close liaison with the obstetric unit includingobstetric anaesthetists and a management plan shouldbe prepared for the delivery and subsequent investi-gation of the neonate. Maternal FX levels may riseduring pregnancy. A FX assay should be performedprior to delivery in a heterozygous woman who maybe at risk of excessive bleeding. At birth a cord bloodsample should be taken for FX assay of the neonate.

Cranial ultrasound should be undertaken inseverely affected neonates because of the increasedrisk of intracranial haemorrhage. Prophylaxis duringthe neonatal period may be necessary in severelyaffected neonates although even with appropriatereplacement therapy, haemorrhage may occur.

Management of factor X deficiency: X:C >2 IU dL)1

Patients with FX levels >10 IU dL)1 or a lower leveland no significant bleeding history (despite haemo-static challenge) do not require replacement therapy.However, the nature of the surgery and any bleedinghistory in relation to previous haemostatic challengesmust be considered.

Inherited deficiency of the vitamin-K dependentclotting factors

Molecular biology

Coagulation FII, FVII, FIX and FX require c-carb-oxylation of critical glutamic acid residues withintheir Gla domains to enable binding of calciumand attachment to phospholipid membranes. Thec-carboxylation reaction is catalysed by thehepatic enzyme c-glutamyl carboxylase that requiresreduced vitamin K (KH2) as a cofactor. During thec-carboxylation reaction, KH2 is converted to vita-min K epoxide (KO), which is recycled to KH2 by thevitamin K epoxide reductase enzyme complex(VKOR) [124]. Heritable dysfunction of c-glutamylcarboxylase or of the VKOR complex results in thesecretion of poorly carboxylated FII, FVII, FIX andFX that have poor haemostatic function. c-Carboxy-lation is also required for the secretion of functionalproteins C, S and Z and the skeletal proteinsosteocalcin and matrix Gla protein.

Incidence and inheritance

Hereditary combined deficiency of the vitaminK-dependent clotting factors (VKCFD) has beenreported as single case reports in <20 kindredsworldwide. Inheritance is autosomal recessive.



Clinical phenotype

There is wide variation in phenotype in the reportedkindreds with VKCFD. Severely affected individualsmay present at birth with spontaneous intracranialhaemorrhage or umbilical stump bleeding [125–127] or in infancy and childhood with spontaneoushaemarthroses and retroperitoneal, soft tissue orgastrointestinal bleeds. Otherwise, presentation maybe later in life with easy bruising and mucocutane-ous or postsurgical bleeding [128–130]. Severebleeding is usually associated with activities of theVKCFDs of <5 U dL)1 [125,126]. The phenotype ofVKCFD may be more severe if there is also acquiredvitamin K deficiency and less severe if patients havereceived therapeutic vitamin K1. This may presentparticular difficulties in establishing a diagnosis ofVKCFD in neonates.

Severely affected children may show skeletalabnormalities such as nasal hypoplasia, distal digitalhypoplasia, epiphyseal stippling and mild conductivehearing loss [131,132]. These abnormalities aresimilar to those of warfarin embryopathy and areattributed to dysfunction of the vitamin K-dependentproteins osteocalcin and matrix Gla protein.Although proteins C and S are also abnormal inVKCFD, there are no reports of venous thrombosis.

Diagnosis

Affected individuals show prolongation of the PTand APTT associated with variable reductions inspecific activities of FII, FVII, FIX and FX. Careshould be taken, particularly in neonates, to excludecauses of acquired vitamin K deficiency or coumarinexposure. The demonstration of normal fastingserum KH2 usually allows exclusion of these diag-noses if there is clinical uncertainty. Serum warfarinassays may sometimes be necessary to excludefactitious coumarin ingestion. VKCFD is associatedwith comparatively preserved antigenic assays forFII, FVII, FIX and FX and with elevated des-c-carboxyprothrombin (PIVKA-II) in serum. Elevatedserum KO may distinguish VKCFD individualswith a molecular defect in the VKORC complex(designated VKCFD type 2) from those with defec-tive c-glutamyl carboxylase activity (VKCFD type 1).Reduced activities of proteins C and S have beendemonstrated in some kindreds [125,133].

Molecular defects

Two missense mutations in the c-glutamyl carboxy-lase gene have been identified in two consanguineous

kindreds with severe phenotype VKCFD type 1[125,127,134]. A locus responsible for VKCFD type2 has been mapped to 16p12-q21 [135]. Individualswith type 2 deficiency have causative mutations inthe recently identified gene encoding a component ofthe VKOR enzyme complex called VKORC-1 [136].

Management

Vitamin K Most previously reported individuals withVKCFD show partial or complete improvement inclotting factor activity, normalization of the PT andAPTT and resolution of bleeding symptoms with oralor parenteral vitamin K1 [125,126,130–132,134,137].However, in other kindreds vitamin K was ineffective[129,133,138]. There is no clear relationship from theliterature between responsiveness to vitamin K andeither severity or molecular basis of VKCFD.

Treatment with oral vitamin K1 (phytomenadione)is therefore indicated in all patients at diagnosis. Inpatients who show insufficient response, there islimited experience of weekly parenteral vitamin K1

10 mg [125,134].

Management of bleeding or surgery In the rare patientswho have responded poorly to vitamin K1, whorequire surgery or who have an acute bleed, factorreplacement is necessary. There is limited experienceof the use of FFP in the treatment of acute bleeds[125,133,137] and a virally inactivated product istherefore the agent of choice. Since FVII has theshortest half-life of the VKCFDs [87], therapy shouldbe monitored using the PT or FVII activity assay.

The 4-factor prothrombin complex concentratesare an alternative to plasma but as these agents havebeen associated with thrombosis or disseminatedintravascular coagulation and should be used withcaution in some individuals [119].

Management of pregnancy There is a single report of apregnancy progressing to term in an individual withsevere VKCFD managed with oral vitamin K1 15 mgdaily throughout pregnancy. Bleeding from an episi-otomy wound in this case required FFP [137].

Factor XI deficiency

Molecular biology

Factor XI is a dimeric serine protease whose functionin the coagulation pathway was recently clarified.Although usually identified as a �contact factor� it isnow realized that the contact pathway is non-physiological. Coagulation is triggered by activation



of the tissue factor–factor VII pathway resulting inthrombin activation. As tissue factor pathway inhib-itor modulates the tissue factor pathway, thrombincan activate FXI and thereby recruit the �intrinsic�pathway of coagulation [139–141]. These observa-tions may explain why FXI is less essential tohaemostasis than FVIII and FIX, and perhaps whyother factors may influence the bleeding tendency inFXI deficiency.

Factor XI deficiency is an autosomally inheritedcondition, which is particularly common in Ashke-nazi Jews in whom the heterozygote frequency is 8%[142]. FXI deficiency has been described in all racialgroups, but in general, the incidence of severedeficiency (FXI:C level <10 U dL)1, although thereis no precise definition) is very low (estimated at1:1 million) [143].

The FXI gene is 23 kb in length. In the AshkenaziJewish population most FXI deficiency is related totwo mutations: type II, a stop codon in exon 5, andtype III, a missense mutation in exon 9 leading toreduced expression of the FXI molecule. Thesemutations occur in equal frequency in Jews leadingto type II homozygotes who typically have undetect-able FXI levels and a more severe bleeding pheno-type; type III homozygotes who have a low level ofFXI (about 10 U dL)1) while compound heterozy-gotes have levels and clinical expression betweenthese two [144]. Several other mutations have beendescribed, usually in single families; however, com-mon founder effect mutations have been described inFrench Basques [145] and in 11 UK families withC128X in exon 5 [146].

Clinical phenotype

In comparison with haemophilia A and B, bleedingmanifestations in FXI deficiency are much lesspredictable, even in severe deficiency. Spontaneoushaemorrhages are very rare. The condition is notrecessive, bleeding occurs in people with heterozy-gous deficiency and is not related to the FXI:C level[147,148]. Bleeding is most commonly provoked byinjury or surgery, particularly in areas of the bodywhere there is high fibrinolytic activity (the mouth,nose and genitourinary tracts). Tonsillectomy, dentalextractions and sinus surgery are well recognized toresult in excessive blood loss. The same individualmay bleed after one operation and not after another.There is disagreement about whether the bleedingmanifestations breed �true�.

Women with FXI deficiency (including heterozy-gotes) are at risk of menorrhagia [149,150] andbleeding in relation to childbirth.

Bleeding manifestations may depend upon otherassociated factors such as mild von Willebrand’sdisease [151,152]. Associated haemophilia A or B, orplatelet disorders have been occasionally reported.However, these observations do not explain all theobserved variation in bleeding pattern. This clinicalvariability makes the management of FXI deficiencypotentially difficult.

As with any inherited coagulation disorder it isimportant to take a full family history and to test anyindividual at risk (parents, siblings and children).Individuals with very low levels (<10 U dL)1) areeither homozygous or compound heterozygous. Itshould be noted that in the Ashkenazi Jewishcommunity as the heterozygote frequency is so high,there is a higher risk of severe deficiency.

Diagnosis

The diagnosis depends upon determination of a FXIlevel below the reference range. The APTT willusually be prolonged, as most reagent-machinecombinations are sensitive to mild contact factordeficiencies. It should be noted that the referencerange used by many laboratories is incorrect; severalauthors have noted the lower limit of normal torange from 63 to 80 U dL)1 [147–150] so that a cut-off of 50 U dL)1 is too low. Bleeding has beenreported in individuals with levels between 50 and70 U dL)1. Each laboratory should establish its ownreference range. The mean FXI level in a group ofnormal blood donors was 99 U dL)1 with a range of60–139 U dL)1 (S. Kitchen and E. Preston, 1995,personal communication).

In the investigation of possible FXI deficiency it isimportant to recognize the limitation of somescreening tests. The APTT became prolonged at alevel below 40 U dL)1 in one study [153] but at50–70 U dL)1 in another [18]. In another study,which employed samples from patients with previ-ously identified deficiencies, normal APTT resultswere obtained in the presence of FXI levels of 22–50 U dL)1 with one reagent [17]. Data frompublished studies and from EQA programmessuggest that most widely used APTT reagents willhave prolonged APTT results in samples frompatients with FXI below 20–25 U dL)1, whereasthere is a more mixed pattern of normal andabnormal results when FXI is in the region 25–60 U dL)1. The lower limit of normal range forFXI activity is probably between 60 and 70 U dL)1

[17,148]. Thus, a normal APTT does not alwaysexclude the presence of a mild FXI deficiency,particularly if marked elevation of FVIII is present



which may normalize APTT where other factors arereduced [17].

Management

Most individuals with FXI deficiency have fewproblems. However, it is important that surgeryand trauma are managed appropriately. As with mildhaemophilia A and B, these individuals tend to forgettheir potential bleeding risk which results in anincreased risk of suboptimal management. In addi-tion, few doctors are familiar with FXI deficiency.

Treatment options in factor XI deficiency

Patients with severe FXI deficiency (FXI:C <10–20 U dL)1) usually bleed in relation to surgery andthese patients, should therefore receive treatment toincrease their FXI levels. Patients with FXI levelsbetween 20 and 70 U dL)1 may bleed and in thesepatients a bleeding history, especially in relation toany haemostatic challenges and the nature of theproposed surgery, will guide the need for therapy.Some of these individuals may have low von Wille-brand factor levels, and it is advisable to measurebaseline levels [148].Fresh frozen plasma: FFP has been used success-

fully to treat patients with FXI deficiency. However,large volumes of FFP may require to be administeredto achieve therapeutic levels, which can lead to fluidoverload. FFP given as a dose of 15–20 mL kg)1

appears effective [154,155]. Monitoring of theresponse is important and a virally inactivatedproduct is recommended. Levels of FXI in patientswith severe FXI deficiency (XI:C <15 U dL)1) areunlikely to rise above 30 U dL)1 with FFP.Factor XI concentrates: FXI concentrates are

available to treat FXI deficiency but only on anamed-patient basis (for details see [1]). There areconcerns about the potential thrombogenicity of FXIconcentrates [156,157] and for these reasons, peakFXI levels should not exceed 70 U dL)1. In addition,tranexamic acid should be avoided in patientsreceiving FXI concentrates.rFVIIa: rFVIIa has been used successfully to

manage adult patients with FXI deficiency undergo-ing surgery [158], although it is not licensed for thisindication.Tranexamic acid: Tranexamic acid is widely used

in FXI deficiency and may be of value in patientswith milder forms of the disorder or to supplementthe use of plasma and rFVIIa. Tranexamic acid isalso of value in the management of women with FXIdeficiency and menorrhagia.

Fibrin glue: May be effective in facilitating localhaemostasis.

Mucosal bleeding

Tranexamic acid: In practise 10 mL of a 5% solutionof tranexamic acid is used as a mouthwash every 8 h.However, this is not commercially available andwill require to be formulated by the PharmacyDepartment.

In women with menorrhagia, tranexamic acid15 mg kg)1 8 hourly (in practise 1 g, 6–8 hourly)may be effective when taken for the duration of themenstrual period.

Spontaneous bleeding It does not usually occur in FXIdeficiency.

Surgery Surgery, however minor, should be carried outunder the supervision of or in consultation with ahaematologist with experience of FXI deficiency. Forpatients with severe FXI deficiency this will usually bein a haemophilia centre. Several strategies may be used,and the treatment needs to be tailored to the individualcircumstances. For patients with severe deficiency,FXI levels of >30 U dL)1 are considered haemostaticfor minor surgery and >45 U dL)1 for major surgery[61]. In practise one should aim to obtain FXI levelsin the region of 70 U dL)1 prior to major surgery.FXI has a half-life of approximately 52 ± 22 h anddaily dosing may not, therefore, be necessary. Dailymonitoring of FXI levels is recommended.

Dental extractions may be carried out with oraltranexamic acid alone (1 g q.d.s. starting the daybefore and continuing for 7 days) even in peoplewith severe deficiency [159].

People with mild FXI deficiency are more difficultto manage because of the variability and unpredict-ability of the bleeding tendency [148]. Oral tranex-amic acid, with or without a virally inactivated FFPshould be considered. For major surgery in youngerpeople factor concentrates may be required.Factor XI concentrates: For patients with severe

FXI deficiency (XI:C <10–20 U dL)1) or for thosewith levels between 20 and 70 U dL)1 but undergo-ing major surgery, FXI concentrate is advised. Inindividuals with FXI levels near the upper end of thisrange the decision will depend upon the bleedinghistory and the nature of the surgery. The doseshould not exceed 30 U kg)1 and peak levels shouldnot exceed 70 U dL)1. Tranexamic acid shouldprobably not be used in patients receiving FXIconcentrate because of concerns about the throm-botic risk. Similarly, FXI concentrates should be used



with caution, if at all, in patients with a history ofcardiovascular or cerebrovascular disease. FXI levelsshould be checked daily.

Pregnancy Women with FXI deficiency are at risk ofmenorrhagia and may be diagnosed as a consequenceof this [160–163]. Their quality of life is oftenreduced. Bleeding in childbirth and postpartum hae-morrhage are potential hazards. Observations of FXIlevels in pregnancy are contradictory but changes aregenerally not clinically significant. Women with FXIlevels in the heterozygous range may bleed at delivery.Vaginal delivery: In women with FXI levels

between about 15 and 70 U dL)1 and no bleedinghistory but previous haemostatic challenges, a policyof �watch and wait� is justified.

For women with FXI levels between about 15 and70 U dL)1 and a significant bleeding history or noprevious haemostatic challenges, tranexamic acid isoften used for 3 days with the first dose beingadministered during labour.

For women with severe FXI deficiency (FXI:C<10–20 U dL)1) FXI concentrate should be givenduring labour.Caesarean section: For women with severe FXI

deficiency (XI:C <10–20 U dL)1) FXI concentrateshould be given. In patients with FXI levels betweenabout 20 and 70 U dL)1, treatment is dependentupon the bleeding history and the FXI level.Epidural anaesthesia: This is generally discouraged

in women with FXI deficiency but may be used in wo-men who have received FXI concentrate and in whoman adequate response (FXI level) has been documented.

Neonates Spontaneous bleeding in the neonatal per-iod has not been reported. No instances of neonatalintracranial haemorrhage resulting from FXI defici-ency have been reported.Circumcision: This may result in serious bleeding

and may be the first presentation of severe FXIdeficiency. For religious circumcision if FXI levels atbirth (cord blood) are <10 U dL)1, the procedureshould be delayed and the FXI levels checked at6 months. If the level remains <10 U dL)1, circum-cision should be performed in hospital under cover ofeither FXI concentrate or FFP. However, the input ofa Mohel to perform the circumcision is important tofulfil religious requirements and this may requireprior coordination with the hospital.

For boys with FXI levels >10 U dL)1 the proce-dure should be carried out under cover of tranexamicacid (15 mg kg)1, 8 hourly for 3 days). The i.v.preparation is given orally in this situation althoughthis is not a licensed use of the product.

Management of factor XI inhibitors

The development of inhibitors to FXI in patientswith FXI deficiency following treatment with FXI israre. rVIIa may be of value in treating these rarepatients [158,164].

Factor XIII deficiency

Molecular biology

Factor XIII (fibrin stabilizing factor) deficiencyinherited (autosomal recessive) or acquired is a veryrare bleeding disorder with a high prevalence ofconsanguinity in affected families. In the UnitedKingdom, the estimated prevalence is one case per1 million [46]. It is much higher in areas with higherconcentration of South Asian immigrants. Similarly,it is much higher in areas of the world with highincidence of consanguinity. The first family, from aconsanguineous marriage, with congenital FXIIIdeficiency was described in 1961 [165].

Clinical phenotype

Patients with FXIII <1 U dL)1 are at greatest riskfrom severe spontaneous bleeding. Those with levelsbetween 1 and 4 U dL)1 are likely to have moderateor severe bleeding. Occasionally patients with levelsabove 5 U dL)1 may bleed [166]. It should be notedthat difficulties in the assay of FXIII complicate therelationship between the FXIII level and clinicalfeatures.

Umbilical bleeding, which occurs a few days afterbirth, is reported in 80% of cases and is verysuggestive of the disorder. Thereafter, patientsexperience a lifelong tendency to severe bruising,muscle haematomas (32%), haemarthroses (24%),intracranial haemorrhage (30%), miscarriages, post-natal bleeding and bleeding after surgery and trauma[167]. FXIII deficiency also leads to a delay inhealing of wounds [168]. In the majority of cases thebleeding diathesis is severe, although there are a fewcases with mild symptoms reported.

Diagnosis

The range of FXIII activity within the normalpopulation is very wide, ranging from 53.2 to221.3% (mean ± SD: 105 ± 28.56%), of the stand-ard normal plasma value in one report [167] and 51–152 U dL)1 in another study [169].

The bleeding time, PT and APTT are normal. Theclot solubility test is a qualitative screening test in



which test plasma is clotted with calcium, thrombinor a combination of both and exposed to a chemicalchallenge, which lyses the clot unless sufficient FXIII-dependant cross-linking has occurred. The clot maybe lysed with 5 mol L)1 urea, 2% acetic acid, 1%monochloroacetic acid or similar. There is no con-sensus on the relative sensitivity of the differentcombinations to low levels of FXIII. In a recent studyby Jennings et al. [170] clotting with calcium andlysing with urea produced abnormal results whenFXIII levels were below 5 U dL)1 (equivalent to5%). Clotting with 30 U mL)1 thrombin and sus-pension in 2% acetic acid produced abnormal resultswhen FXIII levels were 10 U dL)1 or less. Werecommend use of thrombin/acetic acid as thiscombination was associated with the highest pro-portion of abnormal results in the presence ofmarked FXIII deficiency [170], although the samestudy identified important effects of the types ofreagents used in FXIII screening tests.

Several types of FXIII assay have been reportedand at least two are commercially available activityassays. These are based on the cross-linking ofglycine-ethylester into a specific peptide [171] or onincorporation of an amine substrate into FI [172],both of which employ thrombin activation of FXIII.ELISA assays have been used to determine antigeniclevels of FXIII, but are not widely available. Agree-ment between results of FXIII assays may be poorand in a series of proficiency testing exercises theresults of FXIII assays on the same test sample variedwidely between centres, for example, from 3.0 to130 U dL)1 for one sample [170]. These data indi-cate that the accurate determination of FXIII iscurrently problematic. Taken with the variability ofscreening tests [170] it is clear that the detection andcharacterization of FXIII deficiency is difficult, andthis makes the relationship between clinical severityand the degree of FXIII deficiency unclear.

Specific assays should always be used to confirmabnormal screening test results and can be used formonitoring purposes.

Prophylactic factor replacement In view of the highincidence of cerebral haemorrhage it is recommendedthat all patients with severe FXIII deficiency(<1 U dL)1) should receive prophylactic replacementtherapy with FXIII concentrate from the time ofdiagnosis. It should also be considered in those withlevels <4 U dL)1. Some patients with levels morethan 4 U dL)1 may also benefit [166].

The FXIII has a long circulating half-life of 7–10 days and needs to be given at 4–6 weeklyintervals [166,173]. It has been suggested that levels

of 3–10 U dL)1 are sufficient to prevent spontaneoushaemorrhage [166].

It is recommended that 10 units kg)1 bw be givenat intervals of 4 weeks. This appears to preventrecurrent clinical problems and bleeding episodes.

Acute bleeding episodes Whole blood, FFP, storedplasma and cryoprecipitate have all been usedsuccessfully in treatment and are adequate sourcesof FXIII. Plasma-derived FXIII concentrates aresuperior to FFP or cryoprecipitate as these providereliable and high concentrations of FXIII in mini-mum volume, have fewer contaminating substances,and are virally inactivated [173]. It is recommendedin the UK that patients receive a plasma-derivedpasteurized FXIII concentrate until a recombinantproduct is available [1]. Platelets contain FXIII, andin a haemorrhagic emergency platelet transfusionsmay be helpful [169].

It is recommended that 10–20 units kg)1 bwshould be administered, the plasma levels monitoredand kept in the normal range until the bleeding hasstopped.

It is recommended that ideally FXIII levels should bemonitored and in cases of major operation and severehaemorrhage the aim is to obtain normal values.

There are no absolute contraindications. However,the manufacturers advise caution in cases of recentthrombosis and allergic reactions.

Surgery Adults should receive 10–20 units kg)1 bwimmediately before the operation and ideally, theplasma levels monitored. Further treatment can begiven if necessary to keep the level in the normalrange for the following 5 days or until the wound hashealed completely.

Pregnancy The widely held view that women withFXIII deficiency will have inevitable miscarriages, andthat affected men are sterile has not been substan-tiated in the literature [174]. FXIII has an essentialrole in placental implantation and the continuationof pregnancy [175]. Up to 50% of severely affectedpregnant women may miscarry without appropriatetreatment. All severely affected girls should be star-ted on monthly infusions of FXIII concentrate fromthe time of diagnosis and these should be continuedthroughout pregnancy. FXIII levels in plasmafall throughout pregnancy and should be monitoredaiming to keep the trough level >3 U dL)1.

Neonates Factor XIII deficiency is a cause of life-threatening haemorrhage in the neonate and babieswith factor levels of <3 U dL)1 often present with



bleeding in this period. Persistent bleeding from theumbilical cord is the commonest reported symptomin individuals with severe FXIII deficiency [176].Intracranial haemorrhage is another well recognizedpresenting complication of this disorder, as arecephalhaematoma and bleeding after circumcision:the diagnosis should be suspected in any neonatepresenting with such symptoms in whom routinecoagulation screening tests are normal. FXIII defici-ency can be diagnosed from cord or peripheral bloodsamples using the previously described screening testand confirmed by FXIII assay.

Treatment of an acute bleeding episode requiresthe administration of FXIII concentrate at a dose of20 units kg)1. Further treatment should be given asrequired, determined by FXIII levels, if possible, untilthe bleeding has stopped: if intracranial haemorrhagehas occurred then FXIII levels should be maintainedin the normal range for a minimum of 2 weeks,which may require a frequency of up to alternate daydosage before reducing treatment to a prophylacticregimen. In the absence of assay data, some clinicianswould use daily treatment because of the risk of long-term morbidity if bleeding continues. Further studiesare required.

All neonates (and older children) who have beendiagnosed as having a FXIII level of <3 U dL)1

should commence treatment with prophylactic FXIIIconcentrate 10 U kg)1 given once every 4 weeks.Subsequent dosage and dosage intervals can bevaried depending on the pretreatment (>3 U dL)1)or 1 h post-treatment FXIII level (>60 U dL)1).

Close collaboration and careful investigation isrequired where there is unexpected severe bleeding inthe neonatal period, particularly where the parentsare consanguineous.

Inhibitors

The FXIII inhibitors are very rare in congenital FXIIIdeficiency. Rarely FXIII inhibitors arise in the courseof other diseases, and often in relation to chronictherapy with a variety of drugs [167]. Bleeding inthese cases may be severe. Several cases have died ofcerebral haemorrhage [177].

There is no consensus on management. Treatmentsattempted include immunosuppression with steroidsand cyclophosphamide, administration of large dosesof FXIII, and plasma immunoadsorption.

Ehlers-Danlos Syndrome

The EDS is a heterogeneous group of inheritableconnective tissue disorders characterized by joint

hyperextensibility, skin extensibility and tissue fra-gility. It is thought that EDS has a prevalence of onein 5000 to one in 10 000. It is known to affect bothmales and females and all racial and ethnic back-grounds [178].

It is possible that Hippocrates was referring to EDSin his description of the lax joints and multiple scarsnoted in the Nomads and Scythians. However, theearliest unequivocal account was by Van Meek’ren, aDutch surgeon, in 1657. It reappears in the medicalliterature in 1892 when Tschernogobow, after whomthe syndrome is known in Russia, described twocases and proposed that the primary abnormalitywas in the connective tissue. Ehlers, a Danishdermatologist, published a case report of anotherpatient in 1901. This was rapidly followed by areport by Danlos, a French Physician, and thecondition began to be recognized as a distinct entity[179]. A series of further reports were then drawntogether by Weber, the �doyen of British syndromicdiagnosticians�, and the eponym, EDS was coined[180].

The heterogeneity of the condition was soonevident. From the 1960s attempts were made tocategorize it and these culminated in the Berlinnosology in 1988. This has now been superseded bythe revised nosology agreed in Villefranche in 1997based on the cause of each type [181].

Classification

There are six major types of EDS (Table 1). Differenttypes of EDS are classified according to the manifes-tations of signs or symptoms. Rarely a skin biopsy iscarried out including more detailed biochemicalanalysis.

Molecular biology

Patients with EDS exhibit connective tissue abnor-malities of cutaneous strength, elasticity, and healingproperties. Collagen proteins are multimeric, occur-ring in trimers. A minimum of 29 genes contributesto the collagen structure, and the genes occur on 15different chromosomes. There are 19 identifiabledifferent forms of collagen molecules. EDS is causedby variety of abnormalities in the synthesis andmetabolism of collagen [182].

Diagnosis

The diagnosis of EDS is based upon clinical findingsand on the family history. Since many patients do notfit neatly into one of the specific types of EDS, the



diagnosis is often delayed or overlooked. The com-monest presentation to the haematologist will bewith bruising. The PT, APTT and TT are normal.The skin bleeding time is usually normal but on rareoccasions may be prolonged [183]. Platelet abnor-malities have occasionally been reported [184].

Skin hyperextensibility should be tested at aneutral site. It is measured by pulling up the skinuntil resistance is felt. In young children it is difficultto assess because of abundance of s.c. fat [185,186].

Joint hypermobility should be assessed using theBeighton scale. Joint hypermobility depends on age,gender, family and ethnic background. A score of 5/9or greater defines hypermobility [186]. The totalscore is obtained by:1 Passive dorsiflexion of the little fingers beyond 90�;

one point for each hand.

2 Passive apposition of the thumbs to the flexoraspect of the forearm; one point for each hand.

3 Hyperextension of the elbows beyond 10�; onepoint for each elbow.

4 Hyperextension of the knees beyond 10�; onepoint for each knee.

5 Forward flexion of the trunk with knees fullyextended so that the palms of the hand rest flat onthe floor; one point.Easy bruising manifests as spontaneous ecchy-

moses, frequently recurring in the same areas, andcausing characteristic brownish discoloration. Easybruising may be the presenting symptom in earlychildhood. Child abuse should be considered in thedifferential diagnosis.

Tissue fragility manifests as easy bruising and thepresence of dystrophic scars. Scars are found mostly

Table 1. The classification of Ehlers-Danlos Syndromes (EDS) [181] Reprinted by permission of Wiley-Liss, Inc., a subsidiary of John

Wiley & Sons, Inc.

Type Inheritance Basic defect Clinical features

Classical (was EDS I gravis +

II mitis type)

AD Abnormality of type V collagen

encoded by COL5A1 + COL5A2 genes

Major: skin laxity, typical scars, joint

hypermobility

Minor: velvety skin, molluscoid

pseudotumours, muscle hypotonia,

easy bruising, hernias

Hypermobility (was EDS III

hypermobile type)

AD Unknown Major: skin laxity, velvet skin, joint

hypermobility

Minor: recurrent joint dislocations,

chronic limb and joint pain

Vascular (was EDS IV

arterial or ecchymotic type)

AD Structural defects of type III collagen

encoded by the COL3A1 gene

Major: arterial/intestinal/uterine fragility

or rupture, easy bruising, typical

facial appearance

Minor: hypermobility of small joints,

tendon and muscle rupture, club feet,

varicose veins

Kyphoscoliosis (was EDS VI

ocular or scoliosis type)

AR Deficiency of lysyl hydroxylase,

a collagen modifying enzyme

Major: joint laxity, severe muscle

hypotonia in infants scoliosis from

birth, scleral fragility

Minor: tissue fragility, easy bruising,

arterial rupture, Marfanoid habitus,

microcornea, osteopenia

Arthrochalasia (was included

in EDS VII)

AD Deficiency of chains in type I collagen

due to skipping exon 6 in COL1A1

or COL1A2 gene

Major: severe joint hypermobility

with dislocations, congenital

bilateral hip dislocations

Minor: skin laxity, tissue fragility

and scarring, easy bruising, muscle

hypotonia, kyphoscoliosis

Dermatosparaxis (was

included in EDS VII)

AR Deficiency of procollagen I N-terminal

peptidase in collagen type I

Major: severe skin fragility, sagging

redundant skin

Minor: soft, dough skin texture,

premature rupture of fetal membranes,

easy bruising, hernias

Three rare forms of EDS, with the classical phenotype have been recognized, although their syndromic status is unclear: X-linked EDS,

described in a single large UK family (was known as EDS type V); periodontal EDS with prominent gum fragility, AD inheritance (was

known as EDS type VIII); EDS with abnormal platelet aggregation, inheritance possibility AR (was known as EDS type X).

Two entities previously thought to be EDS have now been removed from the EDS classification: EDS type IX has been reclassified as

occipital horn syndrome, an X-linked disorder of copper metabolism; EDS type XI is now termed familial joint hypermobility.



on pressure points (knee, elbow, forehead, chin) andhave a thin, atrophic papyraceous appearance. Fre-quently the scars become wide and discoloured;wound healing is impaired [185,186].

Mitral valve prolapse and proximal aortic dilata-tion should be diagnosed by echocardiography,computed tomography (CT) or magnetic resonanceimaging (MRI). Mitral valve prolapse is a commonmanifestation, but aortic dilation is uncommon.Chronic joint and limb pain is common but skeletalradiographs are normal.

Kyphoscoliosis, arthrochalasia and dermatospar-axis types are considerably less common than theclassical, hypermobility, and arterial types [185,186].

Diagnosis

Confirmatory laboratory tests are not generallyrequired in haematological practice. Mutant formsof genes can be detected in cultured skin fibroblastsby biochemical studies. Molecular genetic studies arehelpful in Types IV, VI, and for some forms of typeVII [187].

Management

There is no specific treatment for bruising orbleeding but measures can be taken to minimize lossof blood and discomfort. Gaping skin wounds,which are common in several types of EDS, shouldbe approached with care. Proper repair of thesewounds is necessary to prevent cosmetic disfigure-ment. The surgeon should be informed and extrasutures may be added and the length of period thesutures are left prolonged by a few days. Tranexamicacid postoperatively has been used to reduce epi-sodes of re-bleeding.

Specialized orthopaedic care is needed for care ofthe joints and the orthopaedic surgeon may usebracing to stabilize joints. Surgical repair of jointsmay be necessary at some time. A physiotherapistmay prescribe exercises to strengthen muscles andteach the patient how to properly use and preservetheir joints. Ascorbic acid up to 4 g daily has beenprescribed but there are no controlled studies toshow its benefit.

All patients should be offered genetic counsellingparticularly prior to pregnancy as there is anincreased risk of uterine rupture [188].

Cardiac problems may be progressive and shouldbe dealt with by a cardiologist.

Consultation with an ophthalmologist (myopia,retinal tears and keratoconus) and a dentist (peri-dontitis) may be necessary.

Patients with EDS have reported that local anaes-thetic is often ineffective. The diagnosis of EDSshould be considered in any patient who complainsunexpectedly of pain during their procedure, partic-ularly when an adequate volume of local anaesthetichas been given.

There are reports of correction of the bleeding timeor improvement in symptoms in patients treated withdesmopressin (DDAVP) [189,190]. There is also acase report of the use of prophylactic DDAVP inlabour to prevent haemorrhage in EDS [191].Although no specific therapies are available to delayor treat the complications of EDS, knowledge of thediagnosis may influence the management of preg-nancy, surgery, bleeding and genetic counselling.There are no controlled studies on therapeuticoptions in relation to bleeding episodes. There is aneed for well-conducted clinical studies at every levelof management.

Mortality

People with type IV disease may die suddenly becauseof spontaneous rupture of medium sized arteries.

Useful contacts

Ehlers-Danlos Support Group, 1 Chandler Close,Richmond N, Yorks DL105QQ, UK.Ehlers-Danlos National Foundation, 6399 WilshireBlvd, Ste 510, Los Angeles, CA 90048, USA.

Acknowledgements

Niamh O’Connell contributed to the section onFXI deficiency, Adrian Minford contributed to thesections on FXIII deficiency and Ehlers-Danlos syn-drome, and authors thank Paul Harrison for helpfulcomments on the manuscript.

Disclaimer

While the advice and information in these guidelinesis believed to be true at the time of going to press,neither the authors, nor the UKHCDO nor thepublishers accept any legal responsibility or liabilityfor the content of these guidelines.

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