Disease: Von-willebrand Disorder Type 2B, Noncanonical

Patient: 55 YO, Italian male

Note: these heterozygous variants are in cis mutations

Variant1: VWF NM_000552.5 c.2771G>A p.(R924Q), in exon 21, D'D3 domain affected

Variant2: VWF NM_000552.5 c.6532G>T p.(A2178S), in exon 37, D4 domain affected

Phenotypes: Mucosal and cutaneous bleeding, low VWF antigent and VWF activity, mild thrombocytopenia, increased ristocetin-induced platelet aggregation, deficiency in high-molecular-weight multimers. Epistaxis, repeated GI bleeding, easy bruising, ISTH BAT score of 7.

Note: Proband initially diagnosed with VWD type 1 in 2010 but later re-diagnosed as VWD-Type 2.

Family: No reported family history of bleeding, parents unable to be included in family work-up as they were deceased by time of study. Paternity test was performed for family, parental link established. Patient's daughter does not present the same mutation noted in proband but has heterozygous polymorphic variant inherited from mother which is: VWF NM_000552.5 c.3379C>T p.(P1127S), in exon 25. Daughter did not show menorrhagia or hemorrhagic disorder except for large hematoma on the thigh after trauma of moderate intensity.

Molecular workup: Paternity test, Sanger-sequencing to validate genetic variants, multimer analysis with electrophoresis, platelet aggregation analysis with Ristocetin, electron micrographs to observe conformation differences.

Prediction workup: Structural analysis with I-TASSER modeling program to identify perturbed structure. A2178S is predicted as tolerated in all in-silico systems used.

Database Information: Effect of the R924Q mutation is noted in literature database but has contradicting reports of effects altering VWF levels. Some studies do report it as a polymorphic variant

Variant is present in dbSNP database (rs33978901). MAF in European population 0.01< MAF< 0.02

Effect of A2178S variant is present in dbSNP database (rs34230288), MAF = 0.02 in European population.

Disease: Von-willebrand Disorder Type 2M

Patient1: 13 YO male

Patient2: 16 YO female

Note they are siblings

Variant: VWF NM_000552.5: c.5192C>T p.(Ser1731Leu), heterozygous variant, in A3 domain in exon 30

Phenotype patient 1: several bleedings after tosillectomy, recurrent epistaxis, decreased VWF:CB ratio, decreased VWF:CB/VWF:Ag ratio

Phenotype patient 2: menorrhagia, intermittent gum bleeding, easy bruising, decreased VWF:CB ratio, decreased VWF:CB/VWF:Ag ratio

For both patients: VWF:CB ratio corresponds to collagen type 1, multimer analysis was normal, VWF:Ag, VWF:Ac, and factor VIII activity was normal.

Family: Not listed

Present in dbSNP (rs764077750)

Present in gnomAD, rare MAF (ALL: 0.0012%)

Predictions:

SIFT- predicted deleterious (Score 0.01, median 3.34)

MutationTaster- Predicted disease causing (probability score 1)

PolyPhen2- Predicted probably damaging (Score 0.983)

CADD score- 26.5

Authors mention similar AA substitutions at the p.1731 position which leads to reduced binding of VWF to collagen in other patients.

Authors also cite a functional human cell experiment in COS7 cell line which resulted in functional VWF defect.

Authors conclude this novel variant as likely pathogenic.