The Changing Landscape of Fabry Disease

Introduction

Fabry disease is a rare X-linked lysosomal storage disorder due to mutations in the GLA gene causing complete or partial deficiency of the enzyme α-galactosidase A (α-Gal A), and subsequent slow accumulation of mainly globotriaocylceramide (Gb3 or GL3) and its deacylated derivative globotriaosylsphingosine (lyso-GL3, also called Lyso-Gb3) in several cell types and body fluids. Early and often asymptomatic cellular damage typically precedes various degrees of organ affection and late organ failure. Clinical symptoms are highly variable and mostly nonspecific (1). Kidney cells, cardiomyocytes, and vascular endothelium are target cells of particular interest in a usually slowly progressive disease. Major complications are secondary to kidney, cardiac, and/or central nervous system affection, usually from the fourth decade onward (2). The disease has had a long journey from the original description of cutaneous angiokeratoma in 1898 (3) to the current recognition of a treatable, highly complex, and heterogenous multisystem disease carrying a high rate of morbidity and mortality (4). The grim natural disease course in the era before enzyme replacement therapy should not be forgotten and was described in 2002 by Branton et al. (5) in 105 hemizygous classic male patients; only 25 of 105 patients survived until the age of 50 and no patients survived past age 60 years. Before the advent of dialysis and kidney transplantation, the most common cause of death was uremia, at a mean age of 41 years (6). Overall, before enzyme replacement was available, a reduced life span of about 25 years in males and 10 years in females was expected compared with the general population (7). Although the pathophysiology is still only partly understood (Figure 1), increasing knowledge of the complexity of mutations and disease manifestations has been fueled by a surge in clinical research and numerous publications after the introduction of enzyme replacement therapy nearly 20 years ago. Agalsidase α and agalsidase β were approved in Europe and the United States (agalsidase β only) in 2001; licensed doses are 0.2 mg/kg per every other week and 1.0 mg/kg per every other week, respectively (8,9). Ten-year follow-up registry data clearly demonstrate a modifying effect of enzyme replacement therapy on serious organ complications and mortality (10,11), and there has been a change in all-cause mortality from predominantly kidney to cardiac deaths (12,13).

Figure 1.

Pathomechanisms of Fabry nephropathy [Eikrem et al. (81)]. GL3, globotriaocylceramide; Lyso-Gb3, globotriaosylsphingosine.

Historically, the disease has been hampered by diagnostic delays and subsequent delays of therapeutic intervention until irreversible organ damage prevails. Hence, the median age at diagnosis was 24 years in males and 31 years in females in a survey of >2200 patients, and the median time between onset of symptoms (usually neurologic pain and gastrointestinal dysfunction) and diagnosis was about 11 years in both sexes (14). The rationale for the increasing focus on early therapy has been clearly demonstrated by several reports highlighting the serious prognostic effect of diagnostic and therapeutic delays (15,16).

Epidemiology, Genetics, and Characterization of Phenotypes

Over the last two decades, general knowledge of Fabry disease and therapeutic challenges have changed dramatically, and prognosis has improved for several reasons. Enormous progress in genetic sequencing technology has led to a paradigm change in the understanding of the complexity of genotype–phenotype interactions. Recently, a general description of clinically relevant categories of variants in Mendelian disorders using the terminology “pathogenic,” “likely pathogenic,” “uncertain significance,” “likely benign,” or “benign” has been suggested (17,18). This is especially relevant for the classification of clinical phenotypes harboring GLA missense mutations accounting for about 60% of >900 known mutations (Human Gene Mutation Database, www.hgmd.org). Importantly, the increasing incidence and prevalence of late-onset, nonclassic mutations, and genetic variants of unclear significance, with milder disease phenotypes have been acknowledged (17,19), with symptoms often confined to a single organ (especially cardiac) and some of these presumably accompanied by no clinical disease at all (17,20). The wide phenotypic spectrum of disease severity, even within the same family, and the fact that not all mutations are causing disease (e.g., polymorphisms) highlight the necessity of careful individual diagnostic and prognostic assessment, to allow correct and timely intervention, as well as avoidance or postponement of unnecessary high-cost treatment in patients with non–disease-causing mutations or very mild disease (19–22). The increasing birth prevalence of GLA mutations, from previous estimates of 1:40,000–170,000 (23) up to 1:1250 in newborn screening studies (19,24,25), reflects the existence of a majority of nonclassic mutations and variants of unknown significance where natural history and effectiveness of enzyme replacement are unknown (19). Currently, research on clinically relevant genotype–phenotype relationships is increasingly prioritized. Clear criteria exist for the diagnosis of classic early-onset Fabry disease (also called Type 1) with absent or low levels of enzyme activity, and typical symptoms (4). However, precise diagnosis may be difficult in late-onset, nonclassic (also called Type 2) male and female patients harboring any variant in the GLA gene with residual enzyme activity, and variable X-chromosome inactivation patterns (females), often presenting with cardiac, minimal, or unclear symptoms (26,27) in early or late adulthood (17,28,29). A widely used algorithm for diagnosing Fabry disease in these latter categories has been published by van der Tol et al. (19,30,31).

New Insight into the Effectiveness of Enzyme Replacement Therapy

The introduction of intravenous enzyme replacement has fueled a tremendous amount of research, opening many new avenues for collaborative work, involving experts from key medical specialties. The way forward has served as a model for learning and organizing clinical research, defining challenges and caveats in the field of rare metabolic diseases (15,32). Although enzyme replacement therapy has undoubtedly turned Fabry disease into a treatable disease, it has become obvious that intervention should be regarded as a disease modifier rather than a cure, and persistent risk of serious complications and increased mortality raises major concerns over current therapeutic strategies (32,33). Furthermore, numerous observational studies and case series published the last decade have disclosed a conundrum of Fabry disease heterogeneity, highlighting the necessity of individual assessment and targeting of therapy, even in patients within the same family (15,34,35). Given a slowly progressive heterogenous disease, new light has been shed on the validity of biases and limitations of few small randomized controlled trials of short duration in comparison with numerous long-term follow-up studies with a large number of patients (36–38). The value of analysis of unpooled data from systematic comprehensive literature searches of observational studies and case series/reports through January 2017 has recently been reported (38). These much larger patient cohorts and attempts to separate data among relevant clinical phenotypes (e.g., children, females, males, classic, and nonclassic), some of them treated for >15 years, have provided new insight that helps the treating physician better define individual patient risk and adequate therapeutic goals. Importantly, this knowledge has strengthened the need for an individual comprehensive multidisciplinary approach, carefully addressing genotype, phenotype, family history, and biomarkers including kidney histology when possible (34,35,39,40).

Individual Therapeutic Goals and Risk Profiles

Organ-specific therapeutic goal recommendations, covering altogether 249 publications (67% male patients including 36 clinical trials), suggest a significant slowing of decline of eGFR and reduction/stabilization of cardiac mass (adult males). The kidney and cardiac therapeutic benefits provide new information expanding the knowledge reported in a previous meta-analysis (33). Although a cardiac benefit was suggested in both sexes, recent data have been generally less robust in females, likely because of a wider disease spectrum ranging from asymptomatic to severely (rare) affected individuals (40). Interestingly, quality of life outcomes were improved in both sexes (34,40). The prognostic importance of younger age and an absence of organ damage when enzyme replacement therapy is initiated have been shown in several studies (10,41). Germain et al. (10) defined “low renal involvement” as urine protein creatinine ratio <0.5 g/g and <50% sclerotic glomeruli in a well-defined observational 10-year follow-up study of 52 classic patients (2 females), mean age 30 years and normal eGFR at start of agalsidase β 1.0 mg/kg every other week. The low renal involvement group (n=32) was younger (mean 25 years at treatment initiation) than the “high renal involvement” group and showed less deterioration of eGFR (mean slope −1.89 versus −6.82 ml/min per 1.73 m² per year). Of note, 94% of the patients were alive at the end of the study and 81% did not experience any events. In patients on enzyme replacement therapy, lower eGFR and higher levels of proteinuria strongly predict faster disease progression (16). Supplemental therapy with renin angiotensin system inhibition to lower proteinuria to ≤0.5 g/d and potentially stabilize GFR should be considered in classic patients with reduced GFR and severe proteinuria (15,42). The effects of enzyme replacement therapy on cerebrovascular events remains unknown, although a recent meta-analysis suggested a potential benefit on stroke prevention (43). Although no clear consensus exists, recommendations for considering withdrawal of enzyme replacement in patients with advanced disease have also been published (44).

Is Higher Agalsidase Dose Beneficial?

Although in vitro milligram per milligram equipotency of agalsidase α and β has been demonstrated (45), the discussion about clinical equipotency of licensed drug regimens remains unsettled. The beneficial effect of higher cumulative agalsidase dose on kidney histology has been reported by Skrunes et al. (41) in serial kidney biopsies in 20 classic patients (median age 21 years, 12 males) with stable microalbuminuria and normal measured GFR followed for 10 years. A clear dose-dependent effect on clearance of podocyte GL3 deposits was found in this cohort, and residual lyso-GL3 correlated with the cumulative enzyme dosage in male patients. The clinical benefits of higher enzyme doses have been corroborated and likely underscored in a larger observational multicenter study with systematic follow-up of a high number of patients of both sexes from three European Fabry centers. The compulsory switch from agalsidase β 1.0 mg/kg every other week to agalsidase α 0.2 mg/kg every other week in many patients (due to the worldwide shortage of agalsidase β supply from June 2009 to January 2012) and subsequent reswitch to agalsidase β 1.0 mg/kg every other week in a number of patients showed conspicuous dose-dependent benefits regarding GFR slopes, lyso-GL3 levels, and gastrointestinal symptoms (46). Moreover, an overview of available evidence indicates that higher doses of agalsidase are beneficial given optimal timing of therapy and selection of patients with classic phenotypes (47,48). Importantly, the majority of literature-based observations of dose-dependent clinical effects so far are confined to classic male patients, and further long-term studies in expanded cohorts of high-risk patients are warranted. Long-term data on therapy outcomes in general are insufficient in female patients and sex-mixed study populations, likely because of variations in X-chromosome inactivation, which are usually not reported in clinical studies (34,40). This may also in part be the reason why no differences were found in clinical events in a recent European multicenter study including a mixture of classic and nonclassic patients [n=387 (192 females), mean age 46±15 years at therapy initiation] comparing licensed doses of agalsidase α and β (49). However, a more robust decrease of lyso-GL3 and better reduction in left ventricular mass were reported in patients receiving a higher enzyme dose (49). There is one study reporting kidney benefit of increasing the dose of agalsidase α to 0.2 mg/kg every week in a limited number of patients (50), but no such evidence is reported for agalsidase β. Although current enzyme substitution regimens fail to normalize elevated lyso-GL3, a clear dose- and age-dependent decrease of lyso-GL3 has been observed after therapy (41,49,51), and the recent therapeutic goal initiative recommends to strive at the lowest possible level of lyso-GL3 (35). More importantly, enzyme replacement therapy has limited effect when started late in the course (15,49,52).

Neutralizing Anti-Agalsidase Antibodies

A major reason for treatment failure is the formation of neutralizing antidrug antibodies (ADAs), which is reported to affect 40% of male patients treated with agalsidase β or α, and leads to subsequent decline of GFR and increase in lyso-GL3 (53). Lenders et al. (53) elegantly demonstrated that agalsidase dose escalation may overcome the detrimental inhibitory effects of these antibodies. A potential therapeutic approach has recently been published, highlighting the need to standardize assays and methods for individual dose escalations to obtain a saturated ADA status (54). Furthermore, future prospective studies are warranted to elucidate the clinical effect of ADAs (54). The role of immunosuppressive therapy is unknown.

Initiation of Pharmacologic Therapy: How Early Is Early Enough?

The strategy of “early treatment” of Fabry disease has been a major focus and is based on the experience in classic patients that progressive disease is more frequent when therapy is delayed until irreversible organ damage is manifest (16,17,52). Our experience in a youngish classic symptomatic patient cohort with normal heart, normal measured GFR, and normo/microalbuminuria suggests that enzyme replacement should be initiated within the teens (before the age of 18 years) (41,51,55), with the goal to prevent or delay the progression to irreversible kidney and heart damage. This “window of opportunity” approach is supported by several authors (15,39,44). Earlier start of therapy in childhood has to be decided on individual basis in patients with severe symptoms and signs. Systematic follow-up to define the individual appropriate window for “early therapy,” often at higher age, is mandatory, especially in slowly progressive nonclassic, late-onset patients (usually females) (34,35,40).

New Therapies

The need for more effective therapies for Fabry disease has stimulated research addressing alternative mechanisms to enhance the efficacy of endogenous or infused enzymes. Substrate reduction therapy (56) and gene therapy trials as listed by ClinicalTrials.gov (57) are currently recruiting patients for phase 1–3 studies.

Migalastat, a small-molecule pharmacological chaperone first approved in Europe in 2016 and the US in 2018, was developed as a stabilizer of specific mutant (amenable) forms of α-Gal to facilitate its normal lysosomal trafficking (58,59). In an 18-month phase 3 trial in predominantly female patients, this agent was well tolerated and was associated with a decrease in left ventricular mass index. Migalastat and enzyme replacement therapy had similar effects on kidney function (59). Another phase 3 study showed modest reduction of GL3 in interstitial capillaries and glomerular cells after 6 and 12 months of therapy (58,60). Thus, migalastat could represent an oral monotherapy alternative to enzyme replacement in such patients (59). Notably, the concept of in vitro and in vivo amenability is under scrutiny, especially in patients with lower range (<10%) enzymatic activity (61). Pegunigalsidase α, a novel polyethylene glycol incubated (PEGylated) enzyme replacement agent, has a prolonged half-life and potential benefits regarding immunogenicity compared with agalsidase. Phase 1 and 2 studies, as well as switch (from agalsidase α) and comparative (agalsidase β) studies, have recently been launched (62).

Plasma and Tissue-Specific Markers

There is no single ideal biomarker in Fabry disease. Elevated plasma lyso-GL3 has been designated a hallmark of Fabry disease (63). Currently, measurement of plasma lyso-GL3 has increasingly replaced plasma and urine GL3 as the most significant, and technically more easily measured, noninvasive diagnostic biomarker (64), supplementing standard measurements of leukocyte GLA enzyme activity and genetic analysis. Lyso-GL3 allows better discrimination between patients with classic and nonclassic disease and subjects without Fabry disease. Furthermore, elevated lyso-GL3 has been linked to clinical events and a higher disease burden (21,29,65). Because skewed X-chromosome inactivation may differ between cells and organs in females, a normal plasma lyso-GL3 value does not rule out the existence of Fabry disease (29,64,65). After initiation of enzyme replacement therapy, a rapid reduction in lyso-GL3 levels is seen in classic males, whereas a slower decline or stabilization typically follows in most nonclassic patients and females (49,66). Beyond being a marker of disease, lyso-GL3 is likely also directly involved in disease pathogenesis via the stimulation of inflammatory and fibrotic mechanisms that are upregulated in Fabry disease (67,68). A novel experimental finding suggesting persistent dysregulated inflammatory signaling in spite of agalsidase-induced podocyte GL3 elimination was recently published (69), shedding new light on potential contributions of glycolipid-stimulated inflammatory markers to vascular remodeling and progressive vasculopathy. Standardization of laboratory assays for lyso-GL3 and limited availability of the analysis in many centers are still a challenge. Further elucidation of the specificity of urinary lyso-GL3 analogs is a matter of ongoing research, especially in late-onset variants with cardiac disease (70). Markers of chronic inflammation in Fabry disease are not yet implemented in clinical practice (67).

In tissue biopsies, abundant lysosomal deposits of glycosphingolipids, mainly GL3, are hallmarks of Fabry disease; these are especially conspicuous in podocytes (71,72) and can easily be diagnosed by bedside stereomicroscopy immediately after a kidney biopsy (73). Recently, measurement of abnormal podocyturia has been tested as a potential early diagnostic and prognostic noninvasive tool in ascertainment of progressive disease and disease burden in patients with classic disease (74). Increased podocyturia has even been reported in very young classically affected children (75). Larger-scale validation of the future role of this parameter is needed. General cardiac biomarkers may also be useful in establishing a full assessment and risk profile for the patient (76).

Kidney Biopsies

Routine clinical laboratory tests (eGFR and albuminuria) are insensitive markers of early progressive Fabry nephropathy. On the other hand, the assessment of specific and nonspecific reversible or irreversible histologic changes provides early information on kidney damage, even in patients with normal GFR and normoalbuminuria. These are crucial diagnostic findings for choosing optimal therapeutic strategies and follow-up of high-risk patients (41,55,77–80). In general, a kidney biopsy is recommended in Fabry patients with atypical symptoms and unclear diagnosis, often with normal kidney function, as well as in patients with unexpected disease course and suspected concomitant diseases (15,30,31). A conspicuous finding in recent case series is the discrepant beneficial effect of enzyme replacement therapy on removal of GL3 inclusion from podocytes, and glomerular capillary endothelial and mesangial cells, contrasting with a worrisome lack of vascular protection. In the study of Tøndel et al. (51) in 12 young classic patients [median age 16.5 (range 7–33) years at treatment initiation, 1 female] treated for 5 years, total clearance of podocyte GL3 was obtained in the youngest patient and microalbuminuria normalized in nearly one-half of the patients. These findings were further expanded by Skrunes et al. (41), who reported paired serial kidney biopsies after 10 years of treatment in an expanded cohort [20 classic patients, 12 males, median age 21 (range 7–61) years], confirming a dose-dependent effect on the elimination of podocyte GL3 deposits (Figure 2) contrasting with a persistent failure to protect smooth muscle cells in media layers of kidney arterioles/arteries (41). This finding suggests that current therapy is insufficient to prevent long-term vascular complications. The underscoring of serious vasculopathy, even in young patients, was further highlighted by the randomized multicenter study of 31 classic pediatric males with minimal disease symptoms and normal measured GFR [median age 12 (range 5–18) years] receiving enzyme replacement (agalsidase beta) for 5 years comparing two treatment regimens [0.5 mg/kg every 2 weeks (n=16) or 1.0 mg/kg every 4 weeks (n=15)] (80). Six patients [mean baseline age 15.5 (range 14–17) years] had repeated paired kidney biopsies; arteriopathy (replacement of arterial/arteriolar muscle cells with hyaline-like material) was evident in all baseline biopsies and, surprisingly, showed progression in all but one patient despite a mean reduction of lyso-GL3 of 71%.

Figure 2.

Histology of Fabry nephropathy. Dose-dependent clearance of podocyte GL3 (red arrows) in two brothers with classic Fabry disease, aged 13 and 15 years at initiation of enzyme replacement therapy. Biopsies are baseline (upper and lower left column, respectively), and after 3, 7, and 13 years of agalsidase therapy. After 6 years of enzyme replacement therapy, the younger brother (upper row) was switched to agalsidase β 1.0 mg/kg every other week. After the switch, the podocytes were virtually completely cleared of GL3, whereas his older brother (lower row), who received a lower cumulative agalsidase dose, continued to have a full podocyte score after a total of 13 years [Skrunes et al. (41)]. Pt, patient.

Systematic kidney biopsies are underused in Fabry disease, and further prospective studies are warranted to help clarify disease mechanisms and potential correlation with clinical phenotypes. Histologic examinations may allow early differentiation between patients with high, low, or no risk of progressive tissue damage, and sometimes unnecessary treatment can be avoided when normal or only minimally affected tissue, or superimposed disease, is identified (20,30,73). Kidney biopsies provide the earliest and most sensitive insights into important cellular and vascular involvement, which are surrogate markers of disease activity. New unbiased stereological histologic methods (79) have shown capacity for assessment of therapeutic response after 1 year of therapy (58,79). To detect and expand potentially relevant pathophysiologic mechanisms, kidney biopsies can be used beyond routine diagnostics by the application of omics-related technologies. A preliminary study from our institution exploited next-generation mRNA sequencing of mRNA from microdissected nephron compartments of serial long-term kidney biopsies with Fabry nephropathy. First analyses pointed toward increased expression of genes, e.g., related to the extracellular matrix, immune response, and inflammation, compared with baseline tissues. Thus, RNA sequencing is feasible in archival Fabry disease kidney biopsies, and may help to delineate potential novel disease markers and therapeutic targets.

Disclosures

Prof. Svarstad reports receiving a grant from Sanofi Genzyme; positions on the advisory boards of Amicus and Sanofi Genzyme; and speaker fees and travel support from Amicus, Sanofi Genzyme, and Shire-Takeda, all outside of the submitted work. Dr. Marti reports receiving grants from Alexion, Amicus, Sanofi Genzyme, and Shire outside of the submitted work.

Funding

This work is supported by the Western Norway Regional Health Authority Helse Vest grant 912233 (to Dr. Marti).