Seidizadeh Omid, Pati Lismaira, Baronciani Luciano, Colpani Paola, Cozzi Giovanna, Pagliari Maria Teresa, Novembrino Cristina, Cairo Andrea, Pappalardo Emanuela, Peyvandi Flora
Department of Pathophysiology and Transplantation, Università degli Studi di Milano, Milan, Italy.
Department of Biosciences, Università degli Studi di Milano, Milan, Italy.
Haemophilia. 2025 Jul;31(4):743-751. doi: 10.1111/hae.70060. Epub 2025 May 10.
The von Willebrand factor (VWF) p.D1472H variant has been shown to artificially lower ristocetin cofactor (VWF:RCo) levels because of impaired VWF binding to ristocetin. Understanding the variant's effect on platelet-dependent VWF activity assays is crucial for avoiding over- or misdiagnosis of von Willebrand disease.
To determine whether p.D1472H affects the VWF:GPIbR latex immunoassay (LIA).
We investigated 150 subjects for VWF:Ag and platelet-dependent VWF activity using four commercially available assays: VWF:GPIbR LIA, VWF:GPIbR chemiluminescence immunoassay (CLIA), VWF:RCo automated, and VWF:GPIbM LIA. p.D1472H was detected directly using Sanger sequencing or as part of other projects using next-generation sequencing (NGS).
A total of 106 subjects were homozygous for aspartic acid [p.D1472H (-)], 41 heterozygous, and 3 homozygous for histidine [p.D1472H (+)]. Using VWF:RCo, p.D1472H (+) subjects (n = 44) showed a significantly lower median VWF activity/VWF:Ag ratio than p.D1472H (-) subjects (p < 0.0001). No significant difference for this ratio was observed between the two groups when assessed using VWF:GPIbR LIA (p = 0.63). Similarly, no significant difference was observed in the presence or absence of the variant (p = 0.31) for the VWF:GPIbR CLIA. Using VWF:GPIbM, a significant difference (p = 0.00052) was observed between the two median ratios.
Our results for the first time show that VWF:GPIbR LIA, despite using ristocetin, is not affected by p.D1472H; hence, it offers reliable results to differentiate VWF quantitative from qualitative deficiencies. We further confirmed that VWF:RCo is highly compromised by p.D1472H, while VWF:GPIbR CLIA is unaffected. VWF:GPIbM LIA results were significantly lower in the p.D1472H (+) subjects than those in p.D1472H (-).
血管性血友病因子（VWF）p.D1472H变异体已被证明会因VWF与瑞斯托霉素结合受损而人为降低瑞斯托霉素辅因子（VWF:RCo）水平。了解该变异体对血小板依赖性VWF活性检测的影响对于避免血管性血友病的过度诊断或误诊至关重要。
确定p.D1472H是否影响VWF:GPIbR乳胶免疫测定（LIA）。
我们使用四种市售检测方法对150名受试者的VWF:Ag和血小板依赖性VWF活性进行了研究：VWF:GPIbR LIA、VWF:GPIbR化学发光免疫测定（CLIA）、VWF:RCo自动化检测和VWF:GPIbM LIA。使用桑格测序直接检测p.D1472H，或作为其他项目的一部分使用下一代测序（NGS）进行检测。
共有106名受试者为天冬氨酸纯合子[p.D1472H（-）]，41名杂合子，3名组氨酸纯合子[p.D1472H（+）]。使用VWF:RCo检测时，p.D1472H（+）受试者（n = 44）的VWF活性/VWF:Ag比值中位数显著低于p.D1472H（-）受试者（p < 0.0001）。使用VWF:GPIbR LIA评估时，两组之间该比值无显著差异（p = 0.63）。同样，对于VWF:GPIbR CLIA，在有或无该变异体的情况下未观察到显著差异（p = 0.31）。使用VWF:GPIbM检测时，两组中位数比值之间存在显著差异（p = 0.00052）。
我们的结果首次表明，VWF:GPIbR LIA尽管使用了瑞斯托霉素，但不受p.D1472H影响；因此，它为区分VWF定量缺陷和定性缺陷提供了可靠的结果。我们进一步证实，VWF:RCo受p.D1472H的影响很大，而VWF:GPIbR CLIA不受影响。p.D1472H（+）受试者的VWF:GPIbM LIA结果显著低于p.D1472H（-）受试者。
