Ruling out the usual suspects: mass spectrometry-based analysis of coagulation protein deficiencies in BDUC
Approximately 70% of patients with mild to moderate bleeding symptoms are diagnosed with bleeding disorders of unknown cause (BDUC). BDUC represents a heterogenous group of individuals where biological mechanisms underlying excessive bleeding remain poorly understood, despite extensive hemostasis testing. This results in difficulty providing adequate treatment and support to patients with bleeding tendencies. Here, we leveraged plasma proteomics aiming to rule out rare protein deficiencies underlying increased bleeding tendency.
Citrated plasma samples, collected at two time points from 105 BDUC patients enrolled in the Vienna Bleeding Biobank, a prospective cohort study on adult patients with mild- to- moderate bleeding tendencies (EC Nr. 206-2009), were analyzed alongside samples from a reference population (n=120). Liquid chromatography-mass spectrometry (LC-MS) was used to quantify plasma proteins using targeted and unbiased acquisition. Bayesian statistics and correlation testing enabled identification of protein differences between the BDUC and reference group, and patient-specific proteomic signatures.
The BDUC cohort is predominantly female (84%) with a high prevalence of blood group O (53.4%).The global plasma proteome of the BDUC cohort did not differ from the reference population and was shaped by demographic characteristics, including correlations of pregnancy zone protein (PZP) levels with sex, von Willebrand factor (VWF) levels with ABO blood group and Alpha-1-acid glycoprotein 2 (ORM2) with body mass index (BMI). Protein quantities were in concordance with diagnostic laboratory measurements, including VWF, fibrinogen and factor IX (r ≥ 0.75). At the group level, no significant deficiencies were detected in proteins listed in the ISTH SSC Tier 1 diagnostic list. Of all detected proteins (n = 450), VWF showed the highest interindividual variation (CV = 97%), and the highest variation between two time-points of the same individual. Patient-specific signatures included low protein Z (PROZ) as well as proteins outside the coagulation cascade.
Individuals with BDUC shared a plasma proteome with healthy controls, apart from some detectable quantitative deficiencies in coagulation factors or the fibrinolytic system, suggesting etiology beyond plasma protein levels. Therefore, we are currently exploring potential hemostatic defects in platelets and cellular systems, and analyzing proteolytic cleavages in plasma upon coagulation to move beyond steady-state protein levels.
