Hsuan-Chieh Liao

Later Onset Fabry Disease, Cardiac Damage Progress in Silence: Experience With a Highly Prevalent Mutation

BACKGROUND Recently, several studies revealed a much higher prevalence of later onset Fabry disease (FD) than previously expected. It suggested that later onset FD might present as an important hidden health issue in certain ethnic or demographic populations in the world. However, the natural history of its phenotype has not been systemically investigated, especially the cardiac involvement. OBJECTIVES The study analyzed a large-scale newborn screening program for FD to understand the natural course of later onset FD. METHODS To date, 916,383 newborns have been screened for FD in Taiwan, including more than 1,200 individuals with the common, later onset IVS4+919G>A (IVS4) mutation. Echocardiography was performed in 620 adults with the IVS4 mutation to analyze the prevalence of left ventricular hypertrophy (LVH), and gadolinium-enhanced cardiac magnetic resonance imaging was performed in 129 patients with FD, including 100 IVS4 adults. RESULTS LVH was observed in 67% of men and 32% of women older than 40 years. Imaging evidenced significant late gadolinium enhancement in 38.1% of IVS4 men and 16.7% of IVS4 women with the IVS4 mutation but without LVH. Seventeen patients underwent endomyocardial biopsies, which revealed significant globotriaosylceramide substrate accumulation in their cardiomyocytes. CONCLUSIONS Significant cardiomyocyte substrate accumulation in IVS4 patients led to severe and irreversible cardiac fibrosis before development of LVH or other significant cardiac manifestations. Thus, it might be too late to start enzyme replacement therapy after the occurrence of LVH or other significant cardiac manifestations in patients with later onset FD. This study also indicated the importance of newborn screening for early detection of the insidious, ongoing, irreversible cardiac damage in patients with later onset FD.

Very Early Treatment for Infantile-Onset Pompe Disease Contributes to Better Outcomes

OBJECTIVE To evaluate whether very early treatment in our patients would result in better clinical outcomes and to compare these data with other infantile-onset Pompe disease (IOPD) cohort studies. METHODS In this nationwide program, 669,797 newborns were screened for Pompe disease. We diagnosed IOPD in 14 of these newborns, and all were treated and followed in our hospital. RESULTS After 2010, the mean age at first enzyme-replacement therapy (ERT) was 11.92 days. Our patients had better biological, physical, and developmental outcomes and lower anti-rh acid α-glucosidase antibodies after 2 years of treatment, even compared with one group that began ERT just 10 days later than our cohort. No patient had a hearing disorder or abnormal vision. The mean age for independent walking was 11.6 ± 1.3 months, the same age as normal children. CONCLUSIONS ERT for patients with IOPD should be initiated as early as possible before irreversible damage occurs. Our results indicate that early identification of patients with IOPD allows for the very early initiation of ERT. Starting ERT even a few days earlier may lead to better patient outcomes.

Mass Spectrometry but Not Fluorimetry Distinguishes Affected and Pseudodeficiency Patients in Newborn Screening for Pompe Disease

BACKGROUND Deficiency of the lysosomal enzyme acid α-glucosidase (GAA) causes Pompe disease. Newborn screening for Pompe disease is ongoing, and improved methods for distinguishing affected patients from those with pseudodeficiency, especially in the Asian population, would substantially reduce the number of patient referrals for clinical follow-up. METHODS We measured the enzymatic activity of GAA in dried blood spots on newborn screening cards (DBS) using a tandem mass spectrometry (MS/MS) assay. The assay displayed a relatively large analytical range compared to the fluorimetric assay with 4-methylumbelliferyl-α-glucoside. DBS from newborns confirmed to have infantile-onset Pompe disease (IOPD, n = 11) or late-onset Pompe disease (LOPD) (n = 12) and those from patients bearing pseudodeficiency alleles with or without Pompe mutations, or Pompe disease carriers (n = 230) were studied. RESULTS With use of the MS/MS GAA assay in DBS, 96% of the pseudodeficiency newborns and all of the Pompe disease carriers were well separated from the IOPD and LOPD newborns. The fluorimetric assay separated <10% of the pseudodeficiencies from the IOPD/LOPD group. CONCLUSIONS The relatively large analytical range MS/MS GAA assay but not the fluorimetric assay in DBS provides a robust approach to reduce the number of referrals and should dramatically facilitate newborn screening of Pompe disease.

Effects of enzyme replacement therapy for cardiac-type Fabry patients with a Chinese hotspot late-onset Fabry mutation (IVS4+919G>A)

Objective Current studies of newborn screening for Fabry disease in Taiwan have revealed a remarkably high prevalence of cardiac-type Fabry disease with a Chinese hotspot late-onset Fabry mutation (IVS4+919G>A). Design Retrospective cohort study. Setting Tertiary medical centre. Participants 21 patients with cardiac-type Fabry disease (15 men and 6 women) as well as 15 patients with classic Fabry disease (4 men and 11 women) treated with biweekly intravenous infusions of agalsidase β (1 mg/kg) or agalsidase α (0.2 mg/kg) for at least 6 months. Outcome measures These data were collected at the time before enzyme replacement therapy (ERT) began and followed up after ERT for at least 6 months, including patient demographics, medical history, parameter changes of cardiac status and renal functions, plasma globotriaosylsphingosine (lyso-Gb3) and Mainz Severity Score Index. Results After 6–39 months of ERT, plasma lyso-Gb3 was found to be reduced in 89% (17/19) and 93% (14/15) of patients with cardiac-type and classic Fabry disease, respectively, which indicated an improvement of disease severity. For patients with cardiac-type Fabry disease, echocardiography revealed the reduction or stabilisation of left ventricular mass index (LVMI), the thicknesses of intraventricular septum (IVS) and left posterior wall (LPW) in 83% (15/18), 83% (15/18) and 67% (12/18) of patients, respectively, as well as 77% (10/13), 73% (11/15) and 60% (9/15) for those with classic type. Most patients showed stable renal function after ERT. There were statistically significant improvements (p<0.05) between the data at baseline and those after ERT for values of plasma lyso-Gb3, LVMI, IVS, LPW and Mainz Severity Score Index. No severe clinical events were reported during the treatment. Conclusions ERT is beneficial and appears to be safe for Taiwanese patients with cardiac-type Fabry disease, as well as for those with the classic type.

Clinical observations on enzyme replacement therapy in patients with Fabry disease and the switch from agalsidase beta to agalsidase alfa

Background: Fabry disease is an X‐linked inherited lysosomal storage disease that can be treated with the enzymes of agalsidase beta (Fabrazyme) and agalsidase alfa (Replagal). Since June 2009, viral contamination of Genzymes production facility has resulted in a worldwide shortage of agalsidase beta, leading to the switch to agalsidase alfa for patients with Fabry disease in Taiwan. Methods: The medical records were retrospectively reviewed for nine male patients with Fabry disease from the start of agalsidase beta treatment until the switch to agalsidase alfa for at least 1 year. Results: After 12–112 months of enzyme replacement therapy (ERT), decreased plasma globotriaosylsphingosine (lyso‐Gb3) was found in five out of seven patients, indicating improvement in disease severity. Among the six patients with available echocardiographic data at baseline and after ERT, all six experienced reductions of left ventricular mass index. Renal function, including microalbuminuria and estimated glomerular filtration rate, showed stability after ERT. Mainz Severity Score Index scores revealed that all nine patients remained stable at 12 months after switching to agalsidase alfa. ERT improved or stabilized cardiac status and stabilized renal function, while reducing plasma lyso‐Gb3. ERT was well tolerated, even among the three patients who had hypersensitivity reactions. Conclusion: The switch of ERT from agalsidase beta to agalsidase alfa appears to be safe after 1 year of follow‐up for Taiwanese patients with Fabry disease.

Comparison of tandem mass spectrometry to fluorimetry for newborn screening of LSDs

Sir while we have no issues with the data presented in the Letter to the Editor [1], we are compelled to give an alternative conclusion. The majority of newborn labs set their own screen cutoffs, and Illinois uses much higher cutoffs than Missouri. It is only meaningful to compare the number of below-cutoff enzyme activity samples with the use of equivalent cutoffs [2–4]. For example, for mucopolysaccharidosis-I, Missouri steadily lowered their cutoff from 20% to 7% of mean iduronidase activity over the past few years [5], whereas Illinois currently uses a much higher cutoff of 14% [6]. With a 7% cutoff the number of screen positives per 100,000 newborns by digital microfluidics fluorescence in Missouri was 43 [5], whereas tandem mass spectrometry (MS/MS) gave 9 in Washington [4,6], 12 in Illinois [6], and 16 in New York [6]. For Pompe disease with equivalent cutoffs [4,6], Missouri reports 52 screen positives per 100,000 [5], Washington 23 [4,6], Illinois 14 [6], and New York 21 [6]. We hypothesize that the lower number of screen positives with MS/MS is due to the high analytical range (not synonymous with dynamic range) of this method compared to fluorometry [7]. Perhaps the clearest evidence that MS/MS performs better than fluorometry comes from a study in Taiwan using an identical set of DBS showing that MS/MS but not fluorometry separates the pseudodeficiency from the Pompe-affected DBS samples (Fig. 1) [8]. This shows that variation due to DBS sampling is not a dominant effect and disproves the statement that the “analytical/dynamic range is irrelevant” [1]. Additionally, there is no correlation seen in one enzymatic activity compared to another for 111,000 DBS tested for iduronidase, -glucosidase, and -galactosidase [9], showing that variation in white cell count or other DBS quality factors are not dominant effects.

Functional and biological studies of α-galactosidase A variants with uncertain significance from newborn screening in Taiwan

Fabry disease is an X-linked disorder resulted from deficiency of α-galactosidase A (GLA) activity. In Taiwan, a total of 792,247 newborns were screened from 2008 to 2014 in two newborn screening centers, and 13 variants of uncertain significance (VOUS) in the GLA gene were identified. To determine whether these variants were pathogenic or not, functional, biochemical, clinical and pedigree analyses were performed. In vitro functional assay was established through site-directed mutagenesis, and four in silico tools were used to predict pathogenesis. The enzyme activity of dried blood spots and plasma metabolite lyso-Gb3 level from subjects with the variants were measured. Additionally, clinical manifestations were evaluated extensively from the subjects and their relatives. Our results revealed that p.G104V, p.I232T, p.D322H, and p.G360C all exhibited relatively low residual enzyme activities and elevated plasma lyso-Gb3 level. These data strongly suggest that these Fabry mutations may cause classical or later-onset phenotypes. In contrast, neither significantly clinical symptoms nor elevated lyso-Gb3 level was found in cases with p.P60S, p.A108T, p.S304T, p.R356Q, and p.P362T variants, which may be non-pathogenic or milder forms of Fabry variants. More data need to be included for the patients with p.N53D, p.P210S, p.M296L, and p.K391T variants. The established system provides us more information to classify these GLA variants.

Detecting 22q11.2 Deletion Syndrome in Newborns with Low T Cell Receptor Excision Circles from Severe Combined Immunodeficiency Screening

Objective Based on experiences and results from newborn screening for severe combined immunodeficiency (SCID), we evaluated the occurrence of chromosome 22q11.2 deletion syndrome (22q11.2DS) in newborns with different T cell receptor excision circles (TREC) results and established a second tier genetic test for 22q11.2DS. Study design Recalled dried blood spots from 486 newborns with TREC results <90 copies/uL were tested from the SCID newborn screening. Quantitative real‐time polymerase chain reaction assay was used to detect the copy number of TBX1 and HIRA genes by simple DNA extraction method. Multiplex ligation dependent probe amplification was used for further confirmation. Results Four hundred sixty‐eight cases were considered negative because their haploid copy number of TBX1 and HIRA genes was >0.75. Eighteen cases with TBX1 and/or HIRA gene copy number <0.75 were suspected as positive, and 13 cases were further confirmed with 22q11.2DS. Detection rates of 22q11.2DS were 10.7% (6/56) in TREC <30 copies, 6.8% (9/132) in <50 TREC copies, 4.6% (12/260) in <70 TREC copies, and 2.7% (13/486) in <90 TREC copies. Conclusions 22q11.2DS detection can be incorporated into the second‐tier assay in subjects with low TREC copies in SCID screening. The dried blood spot methods were feasible for 22q11.2DS newborn screening.