Abstract
Fabry disease is one of the most common lysosomal storage disorders caused by mutations in the gene encoding lysosomal α-galactosidase A (α-Gal A) and resultant accumulation of glycosphingolipids. The sugar mimetic 1-deoxygalactonojirimycin (DGJ), an orally available pharmacological chaperone, was clinically approved as an alternative to intravenous enzyme replacement therapy. The decision as to whether a patient should be treated with DGJ depends on the genetic variant within the α-galactosidase A encoding gene (GLA). A good laboratory practice (GLP)-validated cell culture-based assay to investigate the biochemical responsiveness of the variants is currently the only source available to obtain pivotal information about susceptibility to treatment. Herein, variants were defined amenable when an absolute increase in enzyme activity of ≥3% of wild type enzyme activity and a relative increase in enzyme activity of ≥1.2-fold was achieved following DGJ treatment. Efficacy testing was carried out for over 1000 identified GLA variants in cell culture. Recent data suggest that about one-third of the variants comply with the amenability criteria. A recent study highlighted the impact of inter-assay variability on DGJ amenability, thereby reducing the power of the assay to predict eligible patients. This prompted us to compare our own α-galactosidase A enzyme activity data in a very similar in-house developed assay with those from the GLP assay. In an essentially retrospective approach, we reviewed 148 GLA gene variants from our former studies for which enzyme data from the GLP study were available and added novel data for 30 variants. We also present data for 18 GLA gene variants for which no data from the GLP assay are currently available. We found that both differences in experimental biochemical data and the criteria for the classification of amenability cause inter-assay discrepancy. We conclude that low baseline activity, borderline biochemical responsiveness, and inter-assay discrepancy are alarm signals for misclassifying a variant that must not be ignored. Furthermore, there is no solid basis for setting a minimum response threshold on which a clinical indication with DGJ can be justified.
Highlights
Fabry disease (FD; MIM# 301500) is a rare X–linked lysosomal storage disorder caused by mutations in the GLA gene encoding for the lysosomal enzyme α-galactosidase A (α-Gal A, E.C. 3.2.1.22)
Before the 178 datasets of the good laboratory practice (GLP)-validated assay were compared with our in-house assay, the following 10 GLA gene variants from previous articles [7,13] were reexamined according to the in-house protocol to evaluate the robustness and reproducibility of the assay: M42V, N139S, G183V, N215S, S247P, L268S, L310F, S345P, R356Q, and G360C
Pharmacological chaperone therapy with the novel chaperone DGJ in Fabry disease depends on the biochemical responsiveness of the GLA gene variant
Summary
Fabry disease (FD; MIM# 301500) is a rare X–linked lysosomal storage disorder caused by mutations in the GLA gene encoding for the lysosomal enzyme α-galactosidase A (α-Gal A, E.C. 3.2.1.22). Pathological changes in the gene and its encoded protein result in a complete cellular absence or insufficiency of α-Gal A enzyme activity. The consequence is a cellular and microvascular dysfunction with multiple organ involvement [1]. The resulting storage of complex sphingolipids in the lysosomes, mainly globotriaosylceramide (Gb3) and its metabolite globotriaosylsphingosine (lyso-Gb3) serve as biomarkers in the diagnosis of FD [2] and are believed to play a major role in disease pathophysiology [3]. Clinical FD manifestation involves acroparesthesia, abdominal pain and fever, angiokeratomas, cornea verticillata, decreased ability to perspire, proteinuria, and progressive renal insufficiency. A broad heterogeneous symptom spectrum can be observed, which is largely associated with the genotype [7]
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