Von Willebrand Factor (VWF) Propeptide and Factor VIII (FVIII) Levels Identify the Contribution of Decreased Synthesis and/or Increased Clearance Mechanisms in the Pathogenesis of Type 1 Von Willebrand Disease (VWD) in the Zimmerman Program
Authors: Haberichter Sandra L , Christopherson Pamela A , Flood Veronica H , Gill Joan Cox , Friedman Kenneth D , Montgomery Robert R. 
Published: October 2019

Abstract
Laboratory diagnosis of VWD is challenging, with multiple tests required to obtain an accurate assessment. Plasma VWF level represents a balance between synthesis, secretion, and clearance. Although synthesized together, VWFpp and VWF circulate in plasma independently with differing half-lives. Plasma VWFpp level is used to assess synthesis/secretion, VWFpp/VWF:Ag ratio to indicate clearance of VWF, and FVIII/VWF:Ag ratio to assess VWF synthesis and clearance.
We sought to identify the underlying VWD pathophysiology in subjects enrolled in the Zimmerman Program including 245 healthy controls, 175 "low VWF" (VWF:Ag 30-50), 69 type 1 (VWF:Ag<30), 57 type 1C (VWF:Ag<30 and VWFpp/VWF:Ag>3), and 9 type 1-severe (1S) (VWF:Ag<5). FVIII levels were significantly reduced (p < 0.0001) in VWD subjects compared to controls. The mean FVIII/VWF:Ag ratio in "low VWF" (1.5), types 1 (2.4), 1C (2.9), and 1S (8.7) were significantly different from controls (p < 0.0001). VWFpp levels in all subjects were significantly lower than controls with type 1S subjects demonstrating the most reduced levels. The mean VWFpp/VWF:Ag was significantly increased compared to controls in type 1 (1.9) and 1C (8.4) subjects.
VWFpp level, VWFpp/VWF:Ag and FVIII/VWF:Ag were used to define the underlying pathophysiology in VWD subtypes. A VWFpp/VWF:Ag > 3.0 indicates increased VWF clearance. VWFpp level < 50 IU/dL or FVIII/VWF:Ag > 2.0 indicate reduced VWF synthesis/secretion. A combination of increased clearance/reduced secretion may be identified or neither mechanism. Only 9% of "low VWF" subjects had increased FVIII/VWF:Ag, while 15% had decreased VWFpp suggesting reduced secretion. Other unidentified mechanisms may explain the majority of these "low VWF" cases. In type 1, 38 had increased FVIII/VWF:Ag and 48 had reduced VWFpp indicating reduced secretion in 55-70% of cases. In type 1C, all had increased VWFpp/VWF:Ag, 44 had increased FVIII/VWF:Ag and 19 reduced VWFpp. Reduced secretion may play a role in 33 -77% of these subjects, in addition to increased VWF clearance. All 9 type 1S had reduced VWFpp and 8/9 had increased FVIII/VWF, indicating a reduced secretion mechanism in nearly all cases.
To assess the influence of sequence variant (SV) location on VWF synthesis/secretion, levels were analyzed by presence or absence of SV and SV location by VWF domain. 62% of the VWD cohort had SV identified (23% with >1 SV) while 38% had no SV. Of those with SV, 52% had increased FVIII/VWF:Ag and 48% had decreased VWFpp, suggesting reduced secretion; 31% had increased VWFpp/VWF:Ag indicative of increased clearance; and 32% had neither mechanism. In the group with no SV, only 7% had increased FVIII/VWF:Ag and 17% had decreased VWFpp while 77% had neither mechanism identified.
Of those subjects with reduced secretion as indicated by increased FVIII/VWF:Ag, the majority of SV were found in A1 (29%), D3 (22%) and D1 (13%) which was similar to those with reduced secretion indicated by decreased VWFpp level in A1 (25%), D3 (21%) and D1 (15%). An increased VWFpp/VWF:Ag ratio predicting increased clearance was found in patients with SV in D3, A1 and D4 domains. Patients with both increased clearance and reduced secretion had SV in A1 (53%), D3 (19%) and D4 (11%). Subjects who had neither mechanism identified had SV in A2 (25%), C1-C6 (18%) and D2 (16%), suggesting these SV are associated with other, yet unidentified mechanisms.
Although VWFpp level and FVIII/VWF:Ag are both thought to indicate VWF synthesis/secretion, some discrepancies were observed. VWFpp level may be the more specific marker, as VWFpp and VWF share a common precursor protein. Reduced secretion plays a role in nearly all type 1S, 70% of type 1, and 15% of "low VWF" subjects. Additionally, SV in A1, D3, and D4 may be associated with decreased secretion and/or increased clearance while those in D1 may be associated with decreased secretion alone. No SV were found in 85% of "low VWF" subjects that is consistent with the observation that the majority of these cases (81%) had neither decreased secretion nor increased clearance mechanisms identified. The mechanistic cause of bleeding in these patients remains undefined. Assay of VWFpp and corresponding VWF:Ag ratios may help to define the underlying mechanism in VWD subjects and identify true type 1C VWD patients, which is clinically important for therapeutic treatment.

Disclosures
Flood: CSL Behring: Consultancy; Baxalta: Consultancy. Friedman:Shire: Consultancy; NovoNordisk: Consultancy; CSL Behring: Consultancy; Alexion: Speakers Bureau.