Identificación por nuestro grupo de una nueva mutación en el gen GP1BA (P.W246L) asociada al fenotipo PT-VWD. Sexta mutación reportada internacionalmente / Identification by our group of a novel mutation in the GP1BA gene (P.W246L) responsible for PT-VWD. The 6th mutation internationally reported

La enfermedad de von Willebrand tipo plaquetario (PT-VWD) y tipo 2B (2B-VWD) son trastornos hemorrágicos raros, caracterizados por agregación plaquetaria a bajas concentraciones de ristocetina (RIPA). El diagnóstico diferencial no es fácil y representa un desafío. Hasta el presente, sólo se habían reportado cinco mutaciones en el gen GP1BA relacionadas con este desorden. Describimos aquí la sexta mutación relacionada con PT-VWD, en un paciente con sintomatología hemorrágica severa, macro-trombocitopenia, leve agregación plaquetaria espontánea, RIPA positivo a 0,3 y 0,4 mg/mL, VWF:RCo/VWF: Ag<0,2 y estudios discriminatorios positivos para PT-VWD. VWFpp/VWF: Ag resultó normal a diferencia del 2B-VWD que en algunas oportunidades resulta afectado. El exón 28 del gen VWF del paciente y su madre no reveló mutaciones. Identificamos una sustitución G>T en el nucleótido 3805 en el gen GP1BA del paciente, resultando en un cambio de Trp a Leu en el residuo 246 (p.W246L), en la región de la GPIBa que une al VWF. Esta mutación no se identificó en su madre ni en 100 controles sanos. Es considerada como dañina por análisis in sílico. Consideramos que esta sustitución es responsable del fenotipo PT-VWD del paciente. Dada la ausencia de la misma en los 100 normales estudiados, no se considera un polimorfismo

Platelet-type von Willebrand disease (PT-VWD) and type 2B von Willebrand disease (2B-VWD) are rare bleeding disorders characterized by increased ristocetin-induced platelet aggregation (RIPA) at low concentrations. Diagnosis of either condition is not easy and the differential diagnosis is especially challenging. Five mutations in the GP1BA gene related to PT-VWD and near 50 patients are currently reported worldwide. We herein describe a patient with severe bleeding symptoms, macro thrombocytopenia, mild spontaneous platelet aggregation, positive RIPA at 0.3 and 0.4 mg/mL, VWF: RCo/VWF: Ag <0.2, normal VWFpp/VWF: Ag ratio, and RIPA mixing tests and cryoprecipitate challenge positive for PT-VWD. GP1BA gene was studied in the patient, his mother, and 100 healthy control subjects. We identified a substitution G>T at nucleotide 3805 in the patient's GP1BA gene, resulting in a Trp to Leu amino acid change at residue 246 (p.W246L), within the VWF binding region. This mutation was absent in his unaffected mother and also in the 100 controls, and was predicted as damaging by in silico analysis. The residue is located in a strongly conserved position in the phylogenetic tree. These findings argue in favor of considering this substitution does not represent a polymorphism, and is therefore responsible for the PT-VWD phenotype of the patient

Identification of p.W246L as a novel mutation in the GP1BA gene responsible for platelet-type von Willebrand disease.

Platelet-type von Willebrand disease (PT-VWD) and type 2B von Willebrand disease (2B-VWD) are rare bleeding disorders characterized by increased ristocetin-induced platelet aggregation (RIPA) at low concentrations of ristocetin. Diagnosis of either condition is not easy and the differential diagnosis between the two entities is especially challenging as evidenced by high levels of misdiagnosis of both conditions, but particularly PT-VWD. Five mutations in the GP1BA gene related to PT-VWD and less than 50 patients are currently reported worldwide. We herein describe a patient with severe bleeding symptoms, macrothrombocytopenia, mild spontaneous platelet aggregation, positive RIPA at 0.3 and 0.4 mg/mL, von Willebrand factor ristocetin cofactor (VWF:RCo) to antigen (VWF:Ag) < 0.2, normal VWF propeptide/VWF:Ag ratio, and RIPA mixing tests and cryoprecipitate challenge positive for PT-VWD. GP1BA gene was studied in the patient, in his mother, and in 100 healthy control subjects. We identified a heterozygous substitution G > T located at nucleotide 3805 in the g.DNA of the patient's GP1BA gene, resulting in a Trp to Leu amino acid change at residue 246 (p.W246L). This mutation was absent in his unaffected mother and also in the 100 controls, and was predicted as damaging by in silico analysis. The residue W246 is located within the VWF-binding region and exists in a strongly conserved position in the phylogenetic tree, which is expected to be unable to tolerate substitutions without changing its functional characteristics. These findings argue strongly in favor of the view that this substitution does not represent a polymorphism and is therefore responsible for the PT-VWD phenotype of the patient.

Endogenous or exogenous coagulation factor level and the response to activated protein C.

BACKGROUND: The abnormal response to activated protein C could be the mechanism to explain the prothrombotic role of elevated coagulation factor levels. OBJECTIVE: We evaluated the effect of factor VIII, II, or X (FVIII, FII, or FX) levels on activated protein C resistance technique and its association with the resistant phenotype. MATERIALS AND METHODS: The correlation between APCR and FVIII was assessed in 36 samples, after Desmopressin infusion and the correlation between FII or FX and APCR in 15 patients with plasma levels between 100-125 U/dl. Also, the effect of the addition of purified human factors (FII, FX) to a normal plasma pool (final concentration: 100, 120, 140, 180, 220 U/dl) was estimated on the APCR technique. RESULTS: APCR values correlated with FVIII increase (r(Spearman) = 0.839; p < 0.001); APCR was abnormal (<2.4) in 9/36 samples, showing higher FVIII values in the abnormal group (VIII(abnormalAPCR) = 176.7 +/- 14.2; VIII(normalAPCR) = 103.5 +/- 8.0). APCR did not correlate with endogenous FII (r(Spearman) = 0.423) or FX (r(Spearman) = -0.169). However, the addition of human FII or FX to the normal plasma pool caused a decrease in APCR (r(SpearmanFII) = -0.843; r(SpearmanFX) = -0.958) without reaching abnormal (<2.4) results. FVIII levels may be associated with a resistant phenotype at values >153.0 U/dl, according to the linear regression analysis. Exogenous FII or FX levels greater than 120 U/dl would affect the APCR, without obtaining abnormal results. CONCLUSIONS: The data do not allow the direct association of the FII or FX increase with a defect in the protein C system in the current conditions.

Anticoagulation in the antiphospholipid syndrome.

Our objectives were to evaluate thrombotic complications in patients with lupus anticoagulant fulfilling Sapporo criteria, anticoagulated with an intended INR 2.0-3.0 due to venous and arterial thrombosis. In our series standard anticoagulation was safe and efficacious in preventing recurrences in patients with systemic lupus erythematosus, with other thrombophilia and with arterial thrombosis.

Antiphospholipid antibodies impact the protein C (PC) pathway behavior.

Antiphospholipid antibodies may interfere with the PC pathway, displaying a resistance to the activated PC (resistant phenotype). This effect was evaluated by the APCR and the ProCG systems in 36 lupus anticoagulant samples, yielding abnormal results in 47% of APCR(original), 17% of APCR(modified), and 22% of ProCG test. ProCG values correlated with APCR(original) but not with APCR(modified). Most of lupus anticoagulants affecting the PC pathway showed abnormal APCR(original) results but not abnormal ProCG values. The different behavior between both systems may be due to the heterogeneity of the antibodies or could be attributed to the fact that, in the ProCG, a PC activator is added, while the APCR employs already activated exogenous PC.