Acquired von Willebrand syndrome: A rare disorder of heterogeneous etiologyP Kasatkar, K Ghosh, S Shetty
Department of Hemostasis, National Institute of Immunohaematology (ICMR), KEM Hospital, Parel, Mumbai, Maharashtra, India
Correspondence Address: Source of Support: The authors gratefully acknowledge the partial financial
assistance received from Lady Tata contingency Grant in carrying out this
work, Conflict of Interest: None DOI: 10.4103/0022-3859.113816
Source of Support: The authors gratefully acknowledge the partial financial assistance received from Lady Tata contingency Grant in carrying out this work, Conflict of Interest: None
Context: Acquired von Willebrand syndrome (AVWS) is a rare bleeding disorder that mimics the inherited form of von Willebrand disease (VWD) in terms of laboratory findings and clinical presentation. Aims: To study the etiology of acquired VWD. Settings and Design: The patients referred from various hospitals in and out of Mumbai were included in the study. Materials and Methods: Six patients with AVWS diagnosed at this center over the last 10 years were analyzed against 171 patients with inherited VWD. The differential diagnosis of AVWS was made based on reduced levels of von Willebrand antigen and von Willebrand ristocetin cofactor, decrease in ristocetin induced platelet aggregation, absence of correction in mixing studies with no prior history of bleeding problems and a negative family history for bleeding disorders. Results: In three patients, the disease was associated with systematic lupus erythematosus, out of which one was also associated with Kikuchi lymphadenitis and second with autoimmune hemolytic anemia. Fourth case was associated with hypothyroidism and fifth was a case of dermatitis and vitiligo. The last patient was a case of hemophilia A with Burkitts lymphoma, who developed autoantibodies to von Willebrand factor. Except two patients, all other patients responded to immune suppressive therapy with corticosteroids, while the patient with hypothyroidism responded to oral thyroxine. Conclusion: AVWS is a rare condition and may often be missed or diagnosed as inherited disease associated with heterogeneous disease conditions.
Keywords: Acquired von Willebrand syndrome, autoimmune hemolytic anemia, hypothyroidism, lymphoproliferative disorders, Kikuchi disease, systematic lupus erythematosus
Acquired von Willebrand syndrome (AVWS) results from quantitative and/or qualitative defects in von Willebrand factor (VWF), which is a complex multimeric glycoprotein found in plasma, platelet α granules and subendothelial connective tissue. VWF plays two critical roles in hemostasis: It facilitates platelet adhesion to damaged endothelium and is also a carrier molecule for factor VIII (FVIII). , AVWS is a rare and an underestimated bleeding disorder characterized by a phenotype and clinical manifestation similar to inherited von Willebrand disease (VWD), however, it differs in age of presentation, absence of personal and family history of bleeding and its association with various pathologies, which include, that is, autoimmune disorders. , lymphoproliferative disorders,  cardiovascular disorders,  myeloproliferative disorders,  hypothyroidism,  and may also be drug induced. ,, Inherited VWD is common with an estimated prevalence of 1%, ,, while AVWS is rare and has an estimated prevalence of 0.04%. 
Here, we report six cases of AVWS diagnosed at our center over the last 10 years (2000-2010), where 171 patients with inherited VWD were also diagnosed during the same period. The clinical manifestations, laboratory parameters, underlying pathologies, the diagnostic dilemmas, and management options of cases are discussed in the present report.
This is a retrospective evaluation of all the patients of bleeding disorders recorded in our predesigned proforma, where detailed clinical history of bleeding, nature of bleeding, blood product infusion, medications used to control the bleeding, and family history of bleeding were recorded. All other investigations leading to the final diagnosis were also recorded.
Nine milliliter of venous blood sample was collected in 3.13% sodium citrate (1:9 anti-coagulant to blood) bulbs, 2 ml blood in plain bulb, and 2 ml in ethylene diamine tetra acetic acid (EDTA) container.
Screening coagulation tests included prothrombin time, activated partial thromboplastin time (APTT), and thrombin time. Platelet aggregation studies using Platelet Aggregometer (Chrono-log, Havertown, PA, USA) showed no aggregation with 1.25 mg/ml of ristocetin whereas aggregation with other agonists adenosine diphosphate (ADP), arachdonic acid and collagen) was normal. Factor VIII coagulant (FVIII: C) activity was measured by one stage assay and von Willebrand antigen (VWF: Ag) by enzyme linked immunosorbent assay (ELISA) (Diagnostica Stago, Asnieres, France). Screening for inhibitor to FVIII was carried out by mixing studies by incubating normal pooled plasma and patient's plasma at 37°C for 2 h by APTT based assay. Patients with positive screening results were evaluated by Bethesda assay. von Willebrand ristocetin cofactor activity was assayed according to manufacturer's instructions (commercial kits from Helena Laboratories, Beaumont, TX, USA).
Detection of inhibitors to VWF
The circulating autoantibodies to VWF were assayed by mixing studies of patient's plasma with normal pooled plasma. Samples from VWD patients with similar levels of VWF and negative for inhibitors were taken as negative controls. Various combinations (1:1, 1:3, and 3:1) of plasmas from patients and controls were incubated at 37°C for 1 h and then tested by (i) aggregation with normal "O" group platelets using 1.25 mg/ml of ristocetin and (ii) measurement of VWF: Ag levels by ELISA, and (iii) multimer analysis [Figure 1]. All these are semi quantitative assays to measure the percent of inhibition of VWF in normal plasma by anti-VWF antibody present in patient's sample when compared to control value in different dilutions. The multimeric pattern of VWF was analyzed by sodium dodecyl sulphate (SDS) thin-layer agarose gel electrophoresis, using the discontinuous buffer system. VWF bands were identified by reaction with 125 I-labeled affinity-purified antibody and subsequent autoradiography. 
Routine laboratory investigations included complete blood count, liver function tests, renal function tests, anti-nuclear antibodies (ANA), anti-double stranded (anti-ds) DNA antibodies, anti-cardiolipin antibodies, lupus anticoagulants, hormonal assays i.e. follicle stimulating hormone (FSH), adrenocorticotropic hormone (ACTH), luteinizing hormone (LH), thyroid hormones T and T4, thyroid stimulating hormone (TSH), lactate dehydrogenase (LDH) and complement proteins C3 and C4. Lymph node biopsy was carried out in patients with suspected tumor
The criteria for diagnosis of AVWS were reduced levels of VWF, decrease in ristocetin induced platelet aggregation (RIPA), aberrant multimer profile with no prior history of bleeding problems and a negative family history for bleeding disorders.
Six cases of AVWS were diagnosed and the mean age of the patients was 30.5 years (range 9-52 years); 4 were females and 2 males. All the three cases of SLE were strongly positive for anti-nuclear and anti-double stranded DNA antibodies (cases 1, 2, and 3). Except case 4 who had reduced levels of T3, T4, and TSH, all others had normal levels of thyroid hormones. Liver function and renal function tests were within normal limits and complement levels (C3 and C4) were low in case 1. Except in cases 1 and 6, the remaining patients had normal LDH levels. Four cases had severely reduced FVIII: C and VWF: Ag levels (cases 1, 2, 5, and 6); the remaining two patients (cases 3 and 4) had mild to moderately reduced FVIII: C and VWF: Ag levels. The VWF: RCoF studies in these two patients (cases 3 and 4) showed an activity of 15% and 38% respectively, which corresponded to the antigen levels. Immediate mixing studies (APTT based) using 1:1 mix of patient's plasma and normal pooled plasma showed correction of APTT in cases 1, 3, 4, and 6 whereas, no correction was seen in case 2 and 5. Mixing studies for the presence of anti-VWF antibodies showed inhibition of VWF in normal plasma in all the samples except case 4 by both RIPA and ELISA; case 4, was a case of AVWS associated with hypothyroidism [Table 1].
Menorrhagia, gum bleeding and easy bruisability were the most common symptoms in female patients, while case 5 presented with hemarthrosis; he also had seborrhic dermatitis and vitiligo. Case 1 had an associated diagnosis of Kikuchi lymphadenitis with SLE, while case 6 was a case of inherited hemophilia A, who later developed Burkitt's lymphoma and subsequently AVWS.
AVWS is still a major challenge to the laboratory personnel and the clinicians all over the world. The onset of bleeding in AVWS is usually late in life with neither a past history of bleeding nor any family history of a coagulopathy. , The pathogenic mechanisms involved are increased proteolytic degradation of VWF (myeloproliferative syndromes, drug induced and cardiovascular disorders), antibodies to VWF (monoclonal gammopathy, SLE) or immunoadsorption of VWF onto malignant clones of cells or activated platelets (neoplasia, myeloproliferative disorders) or decreased VWF synthesis (hypothyroidism). None of these proposed mechanisms appears to be specific for any underlying disorder known to be associated with AVWS.
The diagnosis of AVWS is still very challenging and it is often difficult to differentiate AVWS from the inherited VWD. The laboratory parameters closely resemble those of inherited VWD, which include a prolonged bleeding time, prolonged APTT, decreased FVIII: C, VWF: Ag, and RIPA or aberrant multimers. One of the common screening tests for inhibitors to coagulation factors, that is, APTT using 1:1 mix of patient's plasma and normal pooled plasma shows complete correction of APTT even in the presence of inhibitors to VWF, as VWF is not directly involved in any plasmatic coagulation pathway. As a result, often these cases are misdiagnosed as inherited VWD and subsequently when the clinical manifestations of the underlying pathology become evident, repeat laboratory investigations confirm the diagnosis of AVWS. The only differential laboratory diagnosis is the estimation of VWF propeptide, which is normal in AVWS cases except in cases of hypothyroid related AVWS. 
Among the 186 AVWS cases that qualified for the AVWS registry,  the associated pathologies were as follows: Lymphoproliferative (48%) and myeloproliferative disorders (15%), neoplasia (5%), immunological (2%), cardiovascular (21%), and miscellaneous disorders (9%). However, in the present series, except case 4, all the other cases had an immunological cause and all were symptomatic at the time of presentation.
Case 1 was initially diagnosed as a case of inherited VWD with severe deficiency of FVIII: C and VWF. Her routine screening tests for inhibitors to FVIII was negative. She did not have any symptoms of SLE or any autoimmune disorder. Hence, she was not investigated for her autoimmune profile. Subsequently she developed fever, lymphadenopathy and arthralgia. She was investigated for ANA as well as anti-ds DNA and was diagnosed as SLE. Her cervical lymph node biopsy showed Kikuchi-Fujimoto disease More Details (KFD), a condition known to be associated with SLE ,,,, and antiphospholipid antibodies. In the present case, the histopathological features were characterized by presence of necrosis with histiocytes and cellular debris confirming the diagnosis SLE associated with KFD. Patient deteriorated, developed altered sensorium, respiratory distress, and oligouria. She did not respond to immunosuppressive therapy and supportive care and subsequently died of cerebral hemorrhage.
Case 2 is interesting in that she presented as a classical case of VWD without any underlying primary pathology but after 2 years developed symptoms of SLE. On re-evaluation, her hemostatic profile was found to be positive for antibodies to FVIII and VWF. She was treated with corticosteroids, that is, 15 mg/day and prednisolone 60 mg/day. Patient is still on prednisolone 7.5 mg/day with normalized hemostatic profile; her VWF levels have increased to 112% and FVIII level of 108%, which confirms the diagnosis of SLE, associated with AVWS. Case 3 had primarily presented with AIHA with recurrent spontaneous abortions. Who subsequently evolved into AVWS and finally was diagnosed as SLE. On administration of corticosteroids, anti VWF antibodies disappeared and she improved in all her clinical parameters. The patient had two successful deliveries following the therapy. Case 4 was a case of AVWS associated with hypothyroidism who responded to oral thyroxine. Case 5 was associated with seborrhic dermatitis and vitiligo; though vitiligo is a disorder that causes depigmentation of patches of skin, the cause of vitiligo is unknown; however, may arise from autoimmune, genetic, neural, oxidative stress, or viral mechanism. In vitro experiments by mixing studies followed by VWF multimers analysis showed disappearance of low molecular weight multimers. High prevalence of AVWS in patients with thyroid diseases is reported. In majority of AVWS cases, VWF synthesis by megakaryocytes and endothelial cells, as well as its release into the blood are normal, however, the removal of VWF from plasma is accelerated in most patients, except for those with hypothyroidism characterized by decreased VWF synthesis or release.
Case 6 was a case of hemophilia A, who developed Burkitt's lymphoma and developed inhibitors against VWF. The poor response to FVIII concentrate substitution therapy provided the clue for the presence of inhibitors to FVIII; however, the patient was negative for FVIII inhibitors but was positive for VWF inhibitors and died due to gastrointestinal bleeding post-operatively. The present case shows that antibody to VWF should be considered as a possible reason for an increased FVIII requirement in such patients.
In the present series, three out of six patients with AVWS were associated with SLE, but what is noteworthy is that there are no reports of symptomatic bleeding due to AVWS in large series of SLE patients. Bleeding in SLE is often multifactorial and is related to acquired inhibitors to several coagulation factors, thrombocytopenia and vasculitides. It is possible that many patients with SLE may have inhibitors to VWF but this may be subclinical and VWF being a very large and multifunctional protein, unless specific investigations for VWF inhibitors are performed, it is likely that all asymptomatic SLE associated AVWS may be undetected. Normalization of coagulation was achieved in four patients by treatment of the underlying disease.
In conclusion, this report on six patients with AVWS illustrates the complexity of AVWS and its multifactorial etiology. AVWS is underestimated in clinical practice and further prospective studies on its diagnosis, frequency, and management are required. Clinical and laboratory evaluation for AVWS is very complex and there is no single test that can be used to screen for the presence of AVWS. Successful treatment of the associated illness can reverse the clinical and laboratory manifestations. In most patients, AVWS diagnosis was made on an already recognized malignancy or endocrine disorder, but in few cases, the finding of AVWS led to a subsequent diagnosis of an underlying problem. Diagnosis of inherited VWD should be excluded by evaluating family members and by analysis of the past bleeding history of patient. All confirmed AVWS cases should be reported to the National or International registries for better understanding of this complex disorder.
The histopathological diagnosis of Kikuchi-Fujimoto Disease done by Gujral S, Tata Memorial Hospital is gratefully acknowledged.