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A comprehensive testing algorithm for the diagnosis of Fabry disease in males and females
Ashlee R Stiles1, Haoyue Zhang2, Jian Dai2
1Department of Pediatrics, Duke University Medical Center, Durham, NC, USA; Biochemical Genetics Laboratory, Duke University Health System, Durham, NC, USA.
Insights
Diagnosing Fabry disease, especially in females, is challenging. Plasma lyso-globotriaosylceramide (lyso-Gb3) offers high sensitivity and specificity, proving valuable alongside GLA gene sequencing for accurate Fabry disease diagnosis.
Area of Science:
- Genetics
- Biochemistry
- Rare Diseases
Background:
- Fabry disease diagnosis is often delayed, particularly in females, due to varied symptoms and low awareness.
- Effective diagnostic strategies are crucial for timely intervention and management.
Purpose of the Study:
- To evaluate the diagnostic performance of alpha-galactosidase A (α-Gal A) enzyme activity, plasma lyso-globotriaosylceramide (lyso-Gb3) biomarker, and GLA gene sequencing.
- To compare the sensitivities and specificities of these tests in high-risk male and female populations.
Main Methods:
- Assessed patients with suspected Fabry disease using enzyme analysis, biomarker analysis, and GLA sequencing from a single EDTA blood sample.
- α-Gal A activity measured in dried blood spots, plasma lyso-Gb3 by UPLC-MS/MS, and GLA sequencing via Sanger sequencing.
Main Results:
- Plasma lyso-Gb3 demonstrated 100% positive predictive value in males and females.
- GLA sequencing identified pathogenic variants in 30% of patients.
- α-Gal A activity showed high sensitivity but lower specificity in males compared to lyso-Gb3.
Conclusions:
- Plasma lyso-Gb3 is highly sensitive and specific for Fabry disease in both sexes.
- Sex-specific algorithms prioritizing GLA sequencing and plasma lyso-Gb3 are effective for diagnosing Fabry disease.
- Enzyme activity testing has lower utility as a first-tier test in females.
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.
