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Thromb Haemost 2013; 109(06): 999-1006
DOI: 10.1160/TH12-11-0808
Blood Coagulation, Fibrinolysis and Cellular Haemostasis
Schattauer GmbH

C2362F mutation gives rise to an ADAMTS13-resistant von Willebrand factor

Alessandra Casonato
1   University of Padua Medical School, Department of Cardiologic, Thoracic and Vascular Sciences, Second Chair of Internal Medicine, Padua, Italy
,
Elena Pontara
1   University of Padua Medical School, Department of Cardiologic, Thoracic and Vascular Sciences, Second Chair of Internal Medicine, Padua, Italy
,
Monica Battiston
2   Department of Laboratory Diagnostics and Cellular Therapy, IRCCS, C.R.O., Aviano, Pordenone, Italy
,
Margherita Morpurgo
3   Pharmaceutical Chemistry and Pharmacology Department, University of Padua, Italy
,
Maria Grazia Cattini
1   University of Padua Medical School, Department of Cardiologic, Thoracic and Vascular Sciences, Second Chair of Internal Medicine, Padua, Italy
,
Elisabetta Casarin
3   Pharmaceutical Chemistry and Pharmacology Department, University of Padua, Italy
,
Giorgia Saga
1   University of Padua Medical School, Department of Cardiologic, Thoracic and Vascular Sciences, Second Chair of Internal Medicine, Padua, Italy
,
Viviana Daidone
1   University of Padua Medical School, Department of Cardiologic, Thoracic and Vascular Sciences, Second Chair of Internal Medicine, Padua, Italy
,
Luigi De Marco
2   Department of Laboratory Diagnostics and Cellular Therapy, IRCCS, C.R.O., Aviano, Pordenone, Italy
› Author Affiliations
Further Information

Publication History

Received: 08 November 2012

Accepted after major revision: 05 February 2013

Publication Date:
22 November 2017 (online)

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Summary

von Willebrand factor (VWF) multimers result from proteolysis by the metalloprotease ADAMTS13. Since C2362F-VWF features abnormally large multimers with their triplet oligomer structure replaced by a diffuse smear, we explored the susceptibility of C2362F-VWF to ADAMTS13. VWF-enriched blood samples, obtained by cryoethanol precipitation of plasma from a patient with von Willebrand disease (VWD) homozygous for the C2362F mutation and a normal subject, were submitted to cleavage by recombinant ADAMTS13 under static conditions in the presence of urea. C2362F-VWF proved completely ADAMTS13-resistant in vitro. At any concentration of recombinant ADAMTS13 (from 0.1 μM to 1 μM), there was no evidence of the abnormally large VWF multimers of C2362F-VWF disappearing, nor any increased representation of triplet multimer bands, unlike the situation seen in normal VWF. This is due partly to a defective ADAMTS13 binding to C2362F-VWF under static conditions, as seen in both the patient’s and recombinant mutated VWF proteins. These findings were associated with a significantly shorter than normal survival of C2362F-VWF after DDAVP, demonstrating that proteolysis and VWF survival may be independent phenomena. Our findings clearly demonstrate that the loss of cysteine 2362 makes VWF resistant to proteolysis by ADAMTS13, at least partly due to an impaired ADAMTS13 binding to VWF. This suggests that the B2 domain of VWF is involved in modulating ADAMTS13 binding to VWF and the consequent proteolytic process. The C2362F-VWF mutation also enables a new abnormality to be identified in the VWF-ADAMTS13 relationship, i.e. an ADAMTS13-resistant VWF.

Keywords

VWF - C2362F VWF mutation - VWD - ADAMTS13 - ADAMTS13 binding to VWF
 

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