Purpose

Successful diagnosis of Fabry disease is often delayed or missed in patients, especially females, due to clinical heterogeneity and a lack of disease awareness. We present our experience testing for Fabry disease in high risk populations and discuss the relative sensitivities of α-galactosidase A (α-Gal A) enzyme activity in blood, plasma lyso-globotriaosylceramide (lyso-Gb₃) biomarker, and GLA gene sequencing as diagnostic tests for Fabry disease in both males and females.

Methods

Patients with a clinical suspicion of Fabry disease were evaluated with enzyme analysis, biomarker analysis, and GLA sequencing. All three assays were performed from a single tube of EDTA blood. α-Gal A activity was determined in dried blood spots using a fluorometric assay, plasma lyso-Gb₃ by UPLC-MS/MS, and GLA analysis by Sanger sequencing.

Results

Peripheral blood samples were received from 94 males and 200 females, of which 29% of males and 22% of females had a positive family history of Fabry disease. A likely pathogenic or pathogenic variant was identified in 87 (30%) patients (50 males, 37 females), confirming a diagnosis of Fabry disease. Of the remaining patients, 178 (61%) were determined to be unaffected based on normal enzyme activity (males) or normal lyso-Gb₃ and negative sequencing results (females). A VUS was identified in 29 (10%) patients. The positive and negative predictive value of plasma lyso-Gb₃ was 100% and 97% in males and 100% and 99% in females, respectively. This compares with 84% and 100% in males, and 58% and 50% in females for α-Gal A activity testing, respectively.

Conclusions

Plasma lyso-Gb₃ has high sensitivity and specificity for Fabry disease in males and females, and provides supportive diagnostic information when gene sequencing results are negative or inconclusive. α-Gal A activity in dried blood spots (DBS) has high sensitivity, but lower specificity for Fabry disease in males, as not all males with low α-Gal A activities were confirmed to have Fabry disease. Therefore, reflexing to gene sequencing and plasma lyso-Gb₃ is useful for disease confirmation in males. For females, we found that first tier testing consisting of GLA sequencing and plasma lyso-Gb₃ analysis provided the greatest sensitivity and specificity. Enzyme testing has lower sensitivity in females and is therefore less useful as a first-tier test. Enzyme analysis in females may still be helpful as a second-tier test in cases where molecular testing and plasma lyso-Gb₃ analysis are uninformative and in vitro enzyme activity is low.

Summary

Sex-specific testing algorithms that prioritize tests with high specificity and sensitivity offer an effective means of identifying individuals with Fabry disease.

中文翻译：

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男性和女性法布里氏病诊断的综合测试算法。

目的

由于临床异质性和缺乏疾病意识，患者，尤其是女性，法布里疾病的成功诊断常常被延迟或错过。我们介绍了我们在高危人群中对法布里病进行测试的经验，并讨论了血液中血浆α-半乳糖苷酶A（α-GalA）酶活性，血浆溶血球蛋白神经酰胺（lyso-Gb ₃）生物标志物以及GLA基因测序的相对敏感性。法布里氏病男性和女性的诊断测试。

方法

通过酶分析，生物标志物分析和GLA测序对临床怀疑患有Fabry疾病的患者进行评估。所有这三种测定均从单管EDTA血液中进行。使用荧光测定法，通过UPLC-MS / MS的血浆溶酶Gb ₃和通过Sanger测序的GLA分析确定干血斑中的α-GalA活性。

结果

从94名男性和200名女性中采集外周血样本，其中29％的男性和22％的女性具有法布里氏病的阳性家族史。在87名（30％）患者（男50例，女37例）中鉴定出可能的致病或致病变异，证实了法布里病的诊断。在其余患者中，根据正常的酶活性（男性）或正常的lyso-Gb ₃和阴性的测序结果（女性）确定了178名（61％）未受影响。在29名（10％）患者中发现了VUS。男性血浆lyso-Gb ₃的阳性和阴性预测值分别为100％和97％，女性为100％和99％。相比之下，男性进行α-GalA活性测试的男性分别为84％和100％，女性为58％和50％。

结论

血浆溶血Gb ₃对男性和女性的法布里病具有高敏感性和特异性，并且当基因测序结果为阴性或不确定时，可提供支持性诊断信息。干血斑（DBS）中的α-GalA活性具有较高的敏感性，但男性对法布里疾病的特异性较低，因为并非所有具有低α-GalA活性的男性都被证实患有法布里疾病。因此，反思基因测序和血浆溶酶-Gb ₃可用于确认男性的疾病。对于女性，我们发现第一级测试包括GLA测序和血浆溶血Gb ₃分析提供了最大的敏感性和特异性。酶测试对女性的敏感性较低，因此作为一线测试的用处较小。如果分子检测和血浆溶酶-Gb ₃分析的信息不足，而体外酶的活性较低，则女性进行酶分析可能仍可作为第二级检测。

概要

特定性别的测试算法以较高的特异性和敏感性对测试进行优先排序，提供了一种有效的方法来识别患有法布里病的个体。