Real World Evidence Study of Danish Fabry Patients: a >20- Year Longitudinal Retrospective Analysis of Prospectively Collected Data.
Conditions: Fabry Disease
Sponsor: Caroline Michaela Kistorp

 
 
 
 
 
 
 
 
Protocol 
 
 
 
PROTOCOL NUMBER: 
EC22-008 
 
 
STUDY TITLE: 
A Real World Evidence Study of Danish Fabry patients: a > 20-year 
Longitudinal Retrospective Analysis of Prospectively Collected Data. 
 
 
 
VERSION: 1.0 (Original) 
DATE: 30 June 2023 
 
 
 
 
 
 
 
 
 
 
 
 
SPONSOR: 
Amicus Therapeutics Ltd. 
One Globeside, Fieldhouse Ln, Marlow SL7 1HZ, UK 

Study information 
Title 
Real World Evidence study of Danish Fabry patients: a 
>20-year longitudinal retrospective analysis of 
prospectively collected data. 
Protocol version identifier 
V1 
Date of last version of protocol 
 
Active substance 
NA 
Medicinal product 
NA 
Product reference 
NA 
Procedure number 
 
Marketing authorisation 
holder(s) 
 
Joint PASS 
 
MAH(s) contact 
 
Research question and 
objectives 
Research Question 
Is the incidence and prevalence of Fabry associated 
clinical 
events 
(FACEs) 
(cardiac, 
renal 
and 
cerebrovascular) associated with sex, genotype, 
phenotype at time of diagnosis, biomarkers, and Fabry 
specific therapy? 
 
Objectives 
To investigate time to first Fabry associated clinical 
events (FACEs) (cardiac, renal, and cerebrovascular) 
with particular focus on Migalastat clinical outcomes 
and treatment outcomes preceding Migalastat 
therapy.  
To investigate the incidence and prevalence of FACEs 
with respect to Fabry specific treatment, Migalastat, 
ERT or no treatment. 
To describe FACEs in accordance with different geno-
and phenotypic groups.  
To investigate the incidence and time to a first fatal or 
non-fatal cardiac, renal and cerebrovascular clinical 
event, separated by each category. 
 

Title 
Real World Evidence study of Danish Fabry patients: a 
>20-year longitudinal retrospective analysis of 
prospectively collected data. 
Primary objectives 
To investigate time to first Fabry associated clinical 
events (FACE) (cardiac, renal, and cerebrovascular) 
with particular focus on Migalastat clinical outcomes 
and treatment outcomes preceding Migalastat 
therapy.  
 
Secondary objectives 
To investigate the incidence and prevalence of FACEs 
with respect to Fabry specific treatment, Migalastat, 
ERT or no treatment. 
To describe FACEs in accordance with different geno-
and phenotypic groups.  
To investigate the incidence and time to a first fatal or 
non-fatal cardiac, renal and cerebrovascular clinical 
event, separated by each category.   
 
. 
 
Country(-ies) of study 
Denmark  

STATEMENT OF COMPLIANCE
The study will be conducted in accordance with the Declaration of Helsinki principles, 
International Conference on Harmonisation (ICH) Good Clinical Practice (GCP) guidelines, 
and applicable Danish regulations and laws. All personnel involved in the conduct of this 
 
 

SIGNATURE PAGE
The signature below constitutes the approval of this protocol and the attachments and 
provides the necessary assurances that this study will be conducted according to all
stipulations of the protocol, including all statements regarding confidentiality, and 
according to local legal and regulatory requirements and applicable regulations and 
guidelines. 
Principal Investigator or Clinical Site Investigator: 
Signed: 
 
Date: 
 
 
Name:  
 
Title:  
 
 
 

Table of contents
Study information .................................................................................19
Primary objectives ............................................................................................... 20
Secondary objectives ........................................................................................... 20
STATEMENT OF COMPLIANCE ................................................................21
SIGNATURE PAGE ..................................................................................22
Table of contents ..................................................................................23
List of abbreviations .............................................................................25
Responsible parties ...............................................................................26
Abstract ................................................................................................28
Primary outcomes ............................................................................................... 28
Secondary outcomes ........................................................................................... 29
Amendments and updates .....................................................................30
Milestones .............................................................................................31
Rationale and background ....................................................................32
Research question and objectives .........................................................32
Research methods ................................................................................33
Study design ...................................................................................................... 33
Endpoints ........................................................................................................... 34
Patient reported outcomes (PRO) .......................................................................... 36
Setting............................................................................................................... 37
Study population ................................................................................................. 37
Treatments ......................................................................................................... 37
Study workflow ................................................................................................... 38
Variables ............................................................................................................ 38
Variables of interest at inclusion ............................................................................ 39
Exploratory outcome variables .............................................................................. 41
Other variables and exploratory outcome variables .................................................. 43
Safety variables .................................................................................................. 43
Safety Endpoints ................................................................................................. 43
Data sources ...................................................................................................... 44
Data sources for primary endpoints ....................................................................... 44
Other data sources .............................................................................................. 44
Study size .......................................................................................................... 44
Data management ............................................................................................... 44
Data collection .................................................................................................... 44
Validation procedures .......................................................................................... 44
Data management ............................................................................................... 45
Medical coding .................................................................................................... 45
Serious Adverse Events/Adverse Events Data Reconciliation ..................................... 45
Database lock ..................................................................................................... 45

Data analysis ...................................................................................................... 45
Descriptive statistics ............................................................................................ 45
Handling of missing data ...................................................................................... 46
Primary endpoints analysis ................................................................................... 46
Secondary endpoints analysis ............................................................................... 46
Quality control .................................................................................................... 46
Monitoring .......................................................................................................... 46
Investigator training ............................................................................................ 46
Audits ................................................................................................................ 46
Inspections ......................................................................................................... 46
Limitations of the research methodology ................................................................ 47
Protection of human subjects ...............................................................47
Ethics and Good Clinical Practice ........................................................................... 47
Regulatory framework of the study ........................................................................ 47
Independent Ethics Committee / Institutional Review Board ..................................... 47
Subject information and consent ........................................................................... 47
Subjects and data confidentiality ........................................................................... 47
Subject Confidentiality ......................................................................................... 47
Data Confidentiality ............................................................................................. 48
Data handling and record keeping ......................................................................... 48
Record keeping and archiving ............................................................................... 48
Transfer of Essential Documents ........................................................................... 48
Financing and insurance ....................................................................................... 48
Financing ........................................................................................................... 48
Management and reporting of adverse events/adverse reactions .........49
Adverse event (AE) ............................................................................................. 49
Plans for disseminating and communicating study results ....................49
References ............................................................................................50
Annex 1. List of stand-alone documents ...............................................52
Annex 2. ENCePP checklist for study protocols .....................................52
Annex 3. Additional information ...........................................................52
List annex 1.1 Description of participating clinical scientists and 
research staff ........................................................................................53
 
 

List of abbreviations
-Gal A
Alpha Galactosidase A
ACE
Angiotensin-Converting Enzyme
ARB
Angiotensin-Receptor Blockers
BPI
Brief Pain Inventory
CRA
Clinical Research Assistant
CRO
Contract Research Organization
eCRF
electronic Case Report Form
EDC
Electronic Data Capture
ERT
Enzyme Replacement therapy
ESRD
 
FACE 
 
End-Stage Renal Disease
 
Fabry Associated Clinical Event 
FPI
First Patient In
GL-3/Gb3
Globotriaosylceramide
Lyso Gb3
Globotriaosylsphingosine
GCP
Good Clinical Practices
GLA
Galactosidase A
IgG
Immunoglobulin G
LPO
Last Patient Out
LV
Left Ventricle
LVH
Left Ventricular Hypertrophy
LVMI 
Left Ventricular Mass Index 
MedDRA 
Medical Dictionary for Regulatory Activities 
MSSI 
Mainz Severity Score Index 
PBI 
Patient Benefit Index 
PBQ 
Patient Benefit Questionnaire  
PCA 
Principal Component Analysis 
PI 
Principal Investigator 
PNQ 
Patient Needs Questionnaire 
PT 
Preferred Term 
Q1 
First Quartile 
Q3 
Third Quartile 
SF-36v2 
The Short Form (36) Health Survey version 2 
SIV 
Site Initiation Visit 
SmPC 
Summary of Product Characteristics 
SOC 
System Organ Class 
TSQM-9 
Treatment Satisfaction Questionnaire for Medication v.9 
QoL 
Quality of Life 
SAE 
Serious Adverse Event 

Responsible parties
SPONSOR CONTACT DETAILS
Name 
Rigshospitalet and Copenhagen University
Department of Medical Endocrinology and Metabolism 
Address 
Rigshospitalet, Copenhagen University Hospital 
Blegdamsvej 9 
DK-2100 Copenhagen 
Denmark 
 
Phone number
+45 3545 2337 
 
COORDINATING INVESTIGATOR
Name / Title 
MD, PhD, Caroline Kistorp, Professor, Senior Consultant at Department of 
Medical Endocrinology and Metabolism, Rigshospitalet and Copenhagen 
University 
 
Address 
Rigshospitalet, Copenhagen University Hospital
Blegdamsvej 9 
DK-2100 Copenhagen 
Denmark 
Phone/Fax
+45 22475699
Email
caroline.michaela.kistorp@regionh.dk
 
 
CONTRACT RESEARCH ORGANIZATIONS INFORMATION
Name
NOT APPLICABLE
Address

Phone/Fax
Email
 
Participating clinical scientists and research staff will be described in Annex 1 
 
 

Abstract
 
Title 
Real World Evidence study of Danish Fabry patients: a > 20-year longitudinal 
retrospective analysis of prospectively collected data. 
Rationale and 
background 
Fabry is a rare X-linked metabolic lysosomal disorder caused by deficiency 
-galactosidase A ( -Gal A) by mutations in the GLA gene, 
encoding the -Gal A enzyme, which catalyses glycosphingolipids, namely 
globotriaosylceramide (Gb3). Reduced or absent -Gal A activity leads to 
accumulation of Gb3 in various organs as well as cellular dysfunction and 
inflammation causing phsyical symptoms and eventual organ failure. 
Migalastat, an oral chaperone therapy was approved for Fabry patients in 
2016 providing an alternative to infusions with enzyme replacement every 2 
weeks. Although the initial trials of Migalastat had some both short and 
extended outcome treatment comparisons, the overall evidence of clinical 
efficacy is based on too small numbers considering the heterogeneity of the 
Fabry patient population as well as the very slow progression of the disease. 
Though the body of real-world evidence is growing, there is a need for more
publications of real-world long-term data on clinical outcomes with a focus 
on treatment with Migalastat.  
Research question 
and objectives 
Research Question 
Is the incidence and prevalence of Fabry associated clinical events (FACEs) 
(cardiac, renal, and cerebrovascular) associated with sex, genotype, 
phenotype at time of diagnosis, biomarkers, and Fabry specific therapy?
 
Objectives 
To investigate time to first Fabry associated clinical events (FACE) (cardiac, 
renal, and cerebrovascular) with particular focus on Migalastat clinical 
outcomes and treatment outcomes preceding Migalastat therapy. 
To investigate the incidence and prevalence of FACEs with respect to Fabry 
specific treatment, Migalastat, ERT or no treatment. 
To describe FACEs in accordance with different geno- and phenotypic 
groups.  
To investigate the incidence and time to a first fatal or non-fatal cardiac, 
renal, and cerebrovascular clinical event, separated by each category.   
Primary outcomes 
Time to first FACE (cardiac, renal, and cerebrovascular) with particular 
focus on Migalastat on clinical outcomes and treatment outcomes 
preceding Migalastat therapy.  

Secondary outcomes 
To investigate the incidence and prevalence of FACEs with respect to Fabry 
specific treatment, Migalastat, ERT or no treatment. 
To describe FACEs in accordance with different geno- and phenotypic 
groups  
To investigate the incidence and time to a first fatal or non-fatal cardiac, 
renal and cerebrovascular clinical event, separated by each category.   
 
Study design 
Retrospective observational, single-center study from the Danish National 
Fabry Center. Describing treatment related outcomes in Fabry Disease over 
time.  
Population 
Male and female patients with a diagnosis of Fabry Disease.  
Registred at the Danish National Fabry center, Rigshospitalet, 
Denmark from 01.01.2001.  
Variables 
The primary endpoint is time to first FACE (cardiac, renal, or 
cerebrovascular events) since each baseline, with particular focus on the 
added value of the recent addition of Migalastat as a treatment option.
Secondary endpoints are prevalence and incidence of FACEs and time to 
first FACE. 
Exploratory endpoints: Organ-specific biomarkers across modalities 
(auditory, lung, heart, kidney, skin, and neurological), and patient-
reported outcomes evaluated by questionnaires. 
Data sources 
Electronic medical records and paper records.  
Study size 
Estimated 115 patients, which constitute all patients with a genetically 
verified diagnosis of Fabry disease in Denmark.  
Data analysis  
Primary endpoint will be addressed using event-driven statistics from a 
baseline based on the date of diagnosis and treatment initiation.
Secondary analyses will address the individual parameters of the FACEs.
Exploratory endpoints will rely on repeated measures statistical models to 
investigate the change in organ-specific function during the 20-year follow-
up. Detailed description is found in statistical analysis plan (SAP).
Milestones 
 
Ethics Committee 
Approval 
Q3 2023 
Start of data collection 
Q3 2023 
End of data collection 
Q1 2024 
Study progress report 
Q1 2024 
Interim report  
(baseline data) 
Q2 2024 

Registration in the EU 
PAS register 
Q3-4 2023 
Final report of study 
results 
Q3 2024 
 
 
Amendments and updates 
None  
 
 

Milestones
 
Milestone 
 
Ethics Committee Approval 
Q3 2023 
Start of data collection 
Q3 2023 
End of data collection 
Q1 2024 
Study progress report 
Q1 2024 
Interim report (baseline data) 
Q2 2024 
Registration in the EU PAS register 
Q3-4 2023 
Final report of study results 
Q3 2024 
 
 
 

Rationale and background 
Fabry disease is a rare X-linked lysosomal disorder in lysosomes, resulting in a multisystemic 
disorder 
-galactosidase A ( -Gal A) due to mutations in the 
GLA gene, encoding the hydrolase -Gal A enzyme, which catalyses glycosphingolipids namely 
globotriaosylceramide (Gb3).  Reduced or absent -Gal A activity leads to accumulation of Gb3 in 
the lysosomes of various organs, as well as cellular dysfunction and inflammation causing physical 
symptoms and eventual organ failure (1,2). Fabry disease as an X-linked disorder with a high 
penetrance, though not 100% in female heterozygotes, although the disease affects both males 
and females. With the X-linked nature, Fabry disease is suitable for an efficient, high diagnostic 
yield cascade screening over at least three generations surrounding an index-patient. The Danish 
National Fabry Centre has now more than 20 years of experience of cascade screening, which has 
provided a patient population of 115 male and female patients. The prevalence of Fabry disease in 
the Danish population is estimated to be 1:58000 (males 1:85000, females 1:44000) (3). 
Primarily male patients, with early onset classical phenotypes have none or very low -Gal A 
activity, and they inevitably develop multiorgan failure without treatment (4). Patients with late 
onset of Fabry disease represent both males and females with higher -Gal A activity and variable 
degrees of organ involvement (5). Fabry disease can cause irreversible damage to multiple organs 
resulting in Fabry associated clinical events (FACEs) such as renal, cardiac, cerebrovascular 
events, and death (5). 
The primary Fabry specific therapy has historically been enzyme replacement therapy (ERT), and 
as of 2016 an oral chaperone therapy, Migalastat, has been available, providing an alternative to 
ERT for patients with amenable mutations by increasing their endogenous -Gal A enzyme activity
(6 14). However, real-world long-term data from well-characterised cohorts with available detailed 
clinical consecutive observations with focus on treatment with Migalastat and Fabry associated 
events has not been fully investigated (12 14).  
 
Research question and objectives 
Research question 
Is the incidence and prevalence of Fabry associated clinical events (FACEs) (cardiac, renal, and 
cerebrovascular) associated with sex, genotype, phenotype at time of diagnosis, biomarkers, and 
Fabry specific therapy? 
 
 
 
 
 
 
 

Objectives 
To investigate time to first Fabry associated clinical events (FACE) (cardiac, renal, and 
cerebrovascular) with particular focus on Migalastat clinical outcomes and treatment outcomes 
preceding Migalastat therapy.  
To investigate the incidence and prevalence of FACEs with respect to Fabry specific treatment, 
Migalastat, ERT or no treatment. 
To describe FACEs in accordance with different geno- and phenotypic groups.  
To investigate the incidence and time to a first fatal or non-fatal cardiac, renal and cerebrovascular 
clinical event, separated by each category.   
Primary objective 
To investigate time to first Fabry associated clinical events (FACEs) (cardiac, renal, and 
cerebrovascular) with particular focus on Migalastat clinical outcomes and treatment outcomes 
preceding Migalastat therapy. 
Secondary objectives 
To investigate the incidence and prevalence of FACEs with respect to Fabry specific treatment, 
Migalastat, ERT or no treatment. 
To describe FACEs in accordance with different geno- and phenotypic groups.  
To investigate the incidence and time to a first fatal or non-fatal cardiac, renal and cerebrovascular 
clinical event, separated by each category. 
Exploratory objectives 
To describe disease progression with focus on organ involvement. 
Research methods 
Study design  
Observational, retrospective, single-center study originating from the Danish National Fabry 
Centre, Centre of Cancer and Organ Disease, Department of Medical Endocrinology, Copenhagen 
University Hospital, Rigshospitalet. Patients are enrolled at the time of sampling leading to a 
genetically verified diagnosis of Fabry disease. Patients are followed from the day of enrolment 
until 31st of December 2022, having received yearly or bi-yearly evaluation during follow-up.
Method 
Retrospective study of prospectively collected patient data with use of secondary patient data from 
both paper and electronic patient files. All patients are treatment naïve at time of diagnosis, which 
is defined as time point for a verified Fabry associated genetic mutation in the GLA gene. A 
baseline time point is further defined by switch of Fabry specific treatment.  Both Migalastat and 
ERT will be studied retrospectively. Also untreated patients will be studied by assessment of 
relevant clinical and laboratory variables. The patients have been followed from 01.01.2001 until 

31.12.2022. All patients with Fabry disease both receiving specific Fabry medication and untreated 
patients have been offered assessment yearly or every other year by thorough workup, including 
targeted examinations of organ function and symptoms known to be associated with Fabry 
Disease.  
Endpoints 
Primary endpoints  
The primary endpoint is time to first FACE (cardiac, renal, or cerebrovascular events) since each baseline, with 
particular focus on the added value of the recent addition of Migalastat as a treatment option.  
FACEs are defined as the following: 
Cardiac clinical events: 
Myocardial infarction 
New symptomatic arrhythmia requiring medication, direct current cardioversion, or 
interventional procedure (e.g., ablation, pacemaker, or defibrillator implantation)
Unstable angina defined by national practice guidelines and accompanied by electrocardiographic changes and non-
ST segment elevation myocardial infarction [UA/NSTEMI]) and accompanied by electrocardiographic changes 
Congestive heart failure requiring hospitalization 
Any major cardiac medical procedure (e.g., valve replacement, stent-implantation, transplant
or persistent atrial fibrillation) 
Cerebrovascular clinical events 
Stroke (documented by a physician) 
Transient ischemic attack (documented by a physician) 
Renal clinical events 
Doubling of serum creatinine level from the start of analysis (2 consecutive values)
End-stage renal disease (ESRD) requiring long-term dialysis or transplantation
Death due to FACEs 
All FACEs will be analyzed together as a composite endpoint and separately in each category (cardiac, 
cerebrovascular, and renal). 
 
 
Secondary endpoints 
The secondary endpoints  
The prevalence of FACEs from each baseline (diagnosis, and start of treatment) to 
compare between Migalastat-treated, ERT-treated patients and non-treated patients
The incidence of FACEs from each baseline (diagnosis, and start of treatment) to 
compare between Migalastat-treated, ERT-treated patients and non-treated patients

Time to the first cardiac, cerebrovascular, and renal clinical event separately by each 
specific category (including death in these categories) from start of treatment or 
baseline 
Incidence of cardiac, cerebrovascular, and renal clinical events separately by each 
specific category (including death in these categories) 
Occurrence of cardiac, cerebrovascular, and renal clinical events separately by each 
specific category (including death in these categories)  
Annualized rate of change in eGFR based on the Chronic Kidney Disease Epidemiology 
Collaboration equation (eGFRCKD-EPI) over time to compare between Migalastat-treated, 
ERT, and untreated patients  
The prevalence and incidence of a clinically significant change in mGFR and/or eGFR 
(according to KDIGO guidelines rapid progression is defined as a sustained decline in 
eGFR of more than 5 ml/min/1.73 m2 /yr) from each baseline (diagnosis, and start of 
treatment) to compare between Migalastat-treated, ERT-treated patients and non-
treated pts 
The prevalence and incidence of micro (> 30 mg/g creatinine) or macroalbuminuria (> 
300) by urinary albumin/creatinine ratio (ACR) or urinary 24 h protein excretion from 
each baseline (diagnosis, and start of treatment) to compare between Migalastat-
treated, ERT-treated patients and non-treated pts 
 
 
Exploratory organ specific Endpoints 
Reflecting the programme of the annual clinical examinations and questionnaires of the Danish 
Fabry patients.   
Lung capacity: clinically significant changes in spirometry measures of obstructive and restrictive 
lung function evaluate by body plethysmography 
Forced expiratory volume after 1 second (FEV1) 
A clinically relevant change is defined as 25% decrease from baseline or as 
incident volume of 20% below predicted value according to an age- and sex-
adjusted reference range.  
Forced vital capacity (FVC) 
A clinically relevant change is defined as 25% decrease from baseline or as 
incident volume of 20% below predicted value according to an age- and sex-
adjusted reference range. 
 
Neurological, audiological assessments  
Standardized sweat test (hypo, normo, hyperhidrosis), standard unit of measure: mg pr. 
30 minutes 
 

Standardized tilt-test (Presence of orthostatism 
yes/no) 
Heart rate variability (Age-dependent  impaired vs not) 
Audiogram (Impaired vs. not) 
Autonomic nerve function (small fibre assessment)) 
Presence of Fabry associated abnormalities by brain MRI 
 
Ophthalmological and dermatological assessments 
Presence of cornea versiculata  
Presence of angiokeratoma  
 
Composite phenotypic score  
Mainz Severity Score Index (Composite score)  
 
Cardiac function (evaluated by echocardiography) 
Left ventricular mass index (LVMi)  
Left ventricular ejection fraction (LVEF)  
Fractional shortening  
Left ventricular end diastolic and systolic volumes  
Mid-wall fractional shortening (MWFS) 
Left ventricular wall thickness  
NT-proBNP and high sensitive Troponin T  will be analysed from the Danish Fabry biobank.
 
Renal function  
Laboratory parameters: 
serum creatinine 
urine creatinine 
urine protein 
urine albumin 
urine ACR 
measured glomerular filtration rate by 99Tc-DTPA or 51Cr-EDTA 
 
Patient reported outcomes (PRO) 
SF-36 Health Questionnaire (Composite score) 

NHP-Health Profile (Daily living questionnaire 
composite score)
Major Depression Inventory (Becks Depression Index II - Composite score)  
 
Pharmacodynamic Endpoints 
Pharmacodynamic endpoints will be assessed by evaluations comparing Migalastat-treated and 
untreated patients and/or Migalastat-treated and ERT-treated patients: 
Plasma lyso-Gb3 
-Gal A enzyme activity  
Urinary and plasma Gb3, lyso-Gb3, -GAL A enzyme activity  
Neutralizing antibodies with ERT 
 
 
Evaluation tools 
Data extraction from electronic and paper medical records. No specific evaluation test or tool 
required. 
Setting  
Study population 
All patients from the Danish National Fabry Centre will be included, estimating the inclusion of 115 
individuals (sex ratio [female:male] 
 2:1). Pre-specified subgroups includes: non-amenable/ 
amenable to Migalastat, males/females. 
Inclusion criteria 
- Established diagnosis of Fabry Disease based on genetic testing and alpha Gal-A activity in males 
and females.  
- Data from baseline visit and at least one follow-up visit.  
Exclusion criteria 
Absence of data from baseline visit.  
Treatments 
Study treatment 
No treatment will be evaluated prospectively. However, patients will be grouped according to 
treatment received during the treatment period (Migalastat, enzyme-replacement therapy or no 
treatment). 
Patients who are switched from one treatment to another will be grouped according to current 
treatment at the assessment time point (if possible, according to a new baseline before switching). 

Details of this procedure will depend on available information and numbers and cannot be described 
until all patients data have been collected. 
Symptomatic therapies 
Data on nonspecific Fabry therapy such as ace enzyme inhibitors (ACE-I), angiotension-II blockers 
(AT-II), 
anticoagulants, 
statin, 
antiarrhythmic 
therapy, 
asthma/bronchodilator 
therapy, 
antidepressive therapy, as well as use of analgesics and other important concomitant medication 
will be recorded.  
Other measures 
If possible, information on comorbidities at each treatment start and switch will be recorded.
 
 
Study workflow  
Fabry patients entered the study at their baseline visit (first visit in the center) from 2001 with last 
follow-up December 2022. Patient data will be grouped for the whole duration of the defined study 
period, with individual datapoints registered at the specific date of event/measurement. Ideally the 
datapoints will be used according to their specific dates but may be aggregated according to 
availability. Details is further described in the statistical analysis plan. 
 
Variables  
Primary outcome variables (Form to be filled at untreated baseline and every change in treatment 
status: From untreated to treated or vice versa, from one treatment to another. 
Registration of treatment 
All patients included in the trial are considered treatment naïve at inclusion in the observational 
period (01.01.2001). Registration of initiation, change or cessation of treatment will follow the 
following structure: 
Date 
(Date) 
Drug 
(Name) 
Action* 
(Categorical) 
Dose 
(Integer) 
Dose interval 
(Comment) 
Comment 
(Comment)
*Action is restricted to four levels: initiation, increase, decrease, cessation. 
 
Cardiac events 
Date of event 
Cardiac event 
Yes/No
 
Myocardial infarction  
Yes  
No 
 
Unstable angina pectoris  
Yes 

No 
 
PCI / CABG  
Yes 
No 
 
Heart Failure  
Yes 
No 
 
Arrhythmia  
(New onset of symptomatic arrhythmia requiring anti-arrhythmic 
 Medication) 
 
Yes 
No 
 
Heart procedure (pacemaker/ICD/valve surgery) 
Yes 
No 
 
Renal events  
Date of event 
Renal event 
Yes/No 
 
Doubling of s-creatinine 
Yes 
No  
 
ESKD  
Yes  
No  
 
Cerebrovascular events  
Date of event 
Cerebrovascular event 
Yes/No 
 
Stroke  
Yes  
No  
 
TIA  
Yes 
No  
Variables of interest at inclusion 
Variables of interest at inclusion consists primarily of time invariant variables. 
Demographics  
Date of Birth
DDMMYYYY 
Sex
Male 
Female 
Ethnicity
White 
Black 
Asian 

Others 
City
Residence City  
Fabry-specific disease background  
Age of first symptoms 
Years
Age of diagnosis 
Years
Fabry family history 
Yes
No 
Unknown 
Index case 
Yes
No  
ALPHA GAL ACTIVITY (LEUCO ASSAY) at 
inclusion 
% of normal activity
MOLECULAR TEST
Amenable mutation found: 
Other mutations  
 
 
 
 

Exploratory outcome variables 
Variables concerning clinical presentation 
 
ANGIOKERATOMA 
Yes 
No 
CORNEA VERTICILLATA
Yes 
No 
ABDOMINAL PAIN
Yes 
No 
REFLUX
Yes 
No 
INDIGESTION
Yes 
No 
DIARRHEA
Yes 
No 
CONSTIPATION
Yes 
No 
DYSPNOEA
Yes 
No 
 
 
NEUROPATHIC PAIN
Yes 
No 
ACROPARAESTHAESIA 
Yes 
No 
DEPRESSION SYMPTOMS
Yes 
No 
 
Variables concerning organ dysfunction 
 
Lung capacity  
 
Clinically significant Changes in spirometry of 
obstructive function  
Yes  
No 
Clinically significant changes in spirometry of 
restrictive function  
Yes  
No  
Neurological / Audiological 
 
Pathological heart rate variability  
Yes  
No  

Impaired hearing audiogram  
Yes  
No  
Vertigo  
Yes  
No  
 
BLOOD PRESSURE
mmHg   
Heart Rate 
BPM 
 
BUN
mmol/l 
Plasma CREATININE
mmol/l 
URINARY CREATININE
mg/dl 
ALBUMIN/CREATININE RATIO
mg/g 
PROTEINURIA
mg 
GFR CKD-EPI
ml/min/1,73m2 
mGFR 
ml/min  
Plasma Gb3
microgram/ml 
Plasma lysoGb3
ng/ml 
Urine GB3
microgram/mmol creatinine 
Alpha GAL-A activity (DBS or 
Leuco assay)  
% 
normal 
activity 
and 
quantitative 
concentration (Method reference) 
 
Variables concerning concomitant medication 
The included concomitant medication to be recorded is restricted to medications believed to be of 
clinical interest. Medication will be recorded as to their use at inclusion and regarding their change 
throughout the study if possible. 
 
Concomitant medication at inclusion
ACE-I/ ARB 
Yes  
No  
Beta-blocker 
Yes  
No  
Ca-blockers 
Yes  
No  

Diuretics 
Yes  
No  
Statins 
Yes 
No  
 
 
Date 
(Date) 
Drug 
(Name) 
Action* 
(Categorical) 
Dose 
(Integer) 
Dose interval 
(Comment) 
Comment 
(Comment)
*Action is restricted to four levels: Initiation, decrease, increase, cessation. 
 
Other variables and exploratory outcome variables  
 
Safety variables 
Safety Endpoints 
Although the study collects retrospective secondary data, adverse event reporting will be 
conducted, if necessary, in line with the requirements of each study supporter. 
 
SAEs  
Vital signs: 
resting systolic blood pressure 
resting diastolic blood pressure 
Body weight 
Overall survival among all patients enrolled, as assessed by recorded patient deaths 
from any cause 
Laboratory parameters: 
serum creatinine 
urine creatinine 
urine protein 
urine albumin 
urine ACR 
ECGs 
ECHOs, including left ventricular mass index (LVMi), if medically indicated and 
requested by a physician 
Occurrence of male infertility. 
Infusion related reactions and other key safety events  

 
Data sources  
Data sources for primary endpoints 
Electronic medical records, paper records and national medical records 
Other data sources 
Not applicable  
Study size  
The study is estimated to comprise of approximately 115 individuals with genetically verified Fabry 
Disease (Ratio of sex [female:male] 2:1).  
Data management  
Data collection  
Investigators or other designated site staff are implemented in the project group will be trained in the 
specific procedural steps of the protocol, data extraction. 
 
Study variables will be recorded in an electronic database/ CRF using validated and secure database 
in RedCap©. Patient data will be handled while preserving confidentiality. Investigators, or other 
designated site staff, will analyze all Fabry patients irrespective of disease-causing mutation and will
extract clinical and laboratory parameters from original medical records (primary data sources).
Validation procedures 
A 10% random sample of records will be reviewed by a second independent observer to assure quality. 
The investigator team will discuss and resolve any noted discrepancies. 
Case report form 
All CRFs will be completed by the Investigators or other designated site staff who have been trained 
to record relevant information.  
 
Data will be acquired retrospectively from medical documentation of enrolled patients. 
Study variables will be recorded in a CRF designed for the specific purpose of this study. CRF design 
will be reviewed. At the end of data acquisition, the extent of missing values will be evaluated, and 
handled according to the description in the statistical analysis plan. 
All patient data will be handled while preserving confidentiality. 

Data management 
All CRFs will be completed by the Investigators or other designated site staff who have been trained 
to record relevant information. Designated staff responsible for data acquisition should have higher 
medical education.  
 
Data will be captured retrospectively from medical record documentation.  
Study variables will be recorded in an electronic CRF by RedCap. CRFs will be reviewed for 
completeness at the end of document completion for data check (of missing data and/or specific 
questions requiring the investigators patient knowledge). All patient data will be handled preserving 
confidentiality. 
 
Medical coding 
Not applicable.  
Serious Adverse Events/Adverse Events Data Reconciliation 
Although the study collects retrospective secondary data, reporting of adverse events will be 
conducted, if necessary, in line with the requirements of each study supporter. In the event of not 
previously reported serious adverse events or suspected, unexpected serious adverse reactions to a 
l be reported to Competent Authorities within 24 hours. 
Database lock 
At the completion of data acquisition, all interested parties will be informed of impending data lock. 
Data lock will transpire if all parties agree, but before the independent validation by an external 
party and before statistical analysis. 
Data analysis  
Descriptive statistics 
Analyses will be performed by descriptive statistical methods. Continuous variables will be described 
with number of patients with valid/missing observations, mean, standard deviation, median, 
minimum and maximum. Non-normally distributed data will be presented as median and interquartile 
range as appropriate.  Categorical variables will be described by frequencies and related percentages.  
In addition to descriptive analysis, many of the endpoints will undergo statistical testing (i.e., p-values, 
confidence limits) to provide inferential summaries of subgroup comparisons (such as gender, age, 
ethnicity). 
Details are going to be described in the Statistical Analysis Plan (SAP). 
Analysis will be stratified by sex, age (> or <40 years), Fabry specific therapy (ERT treated 
(agalsidase alfa and beta separately), Migalastat treated, treatment naïve), baseline organ 
involvement (cardiac, renal, cerebrovascular, peripheral nervous system, and gastrointestinal 
system). Additional analysis may be required and carried out based on the results of the analyses 
mentioned above. 

Handling of missing data 
Missing data will not be imputed.   
Primary endpoints analysis 
Kaplan-Meier plots will be used to assess time to first Fabry associated clinical event (FACEs). Cox 
proportional hazard models will be used to assess the association between baseline characteristics 
and incident Fabry associated clinical event. The following baseline variables will be used in the 
model as covariables: age, time since diagnosis of Fabry disease, previous FACE (renal, cardiac, or 
cerebrovascular).  
Patient benefit index 
Not applicable  
Secondary endpoints analysis 
Incidence rate of Fabry associated clinical events will be evaluated as events per 1000 patient 
years 
If data allows incidence and prevalence will be analyzed in subgroups (i.e., classic males, females, 
late-onset males, and females) 
Renal, cerebrovascular, and cardiac function analyses will be performed in the same subgroups, as 
well as safety analyses per subgroup. 
Quality control  
Monitoring 
Investigator training   
Documents to perform the study will be mailed to physicians who agreed to participate, including 
the investigator agreements (fees agreement and confidentiality agreement).   
Investigators and other designated site staff who have agreed to participate will be trained about 
the protocol, study procedures and CRF completion by teleconference or on-site visits. 
Study variables will be recorded in a CRF, and patient data will be handled preserving 
confidentiality. Investigators, or other designated site staff will analyze all Fabry patients with 
amenable mutations and will extract clinical and laboratory features from original medical records 
(primary source). 
 
Audits 
Not applicable   
Inspections 
Not applicable  

Limitations of the research methodology  
The participating center has been selected as the Danish national Fabry center, with more than 20 
years of experience managing patients with Fabry disease.    
As the clinical data will be collected retrospectively, it is expected that some clinical data will be 
missing. 
Protection of human subjects  
Ethics and Good Clinical Practice 
The study will be conducted in accordance with ICH GCP. All applicable subject privacy requirements, 
and the ethical principles that are outlined in the Declaration of Helsinki 2013, including, but not 
limited to: Danish legal and health authorities and 
approval of study protocol and any subsequent amendments. The study will be reviewed and 
approved by the Danish Data Protection Agency, and the Danish Health Data Authority. With special 
regards to enrolment of patients who is dead, the study will be reviewed and approved by the 
Scientific Ethics Committee, The Capital Region, Denmark according to the specific legal 
requirements. Data will be anonymized (coded) when entered in the database.  
 
Regulatory framework of the study 
No drug will be evaluated prospectively, therefore, regulatory submission to the Danish Medical 
Association is not required in Denmark. Regarding data-collection, approval will be acquired from 
the legal regulatory authorities of the Capital Region of Denmark (see above). 
Independent Ethics Committee / Institutional Review Board 
No other Institute Review Board.  
Subject information and consent 
All patients will have provided written informed consent. The Study Team must maintain the 
original, signed Informed Consent/Assent Form. A copy of the signed Informed Consent/Assent 
Form must be given to the patient and/or their legally authorized representative. 
A dated entry will 
obtained prior to any extraction of data and that the patient or their legally authorized 
representative received a copy of the signed informed consent/assent. 
Subjects and data confidentiality 
Subject Confidentiality 
recorded in the CRF will be documented anonymously, coded with a patient number in such a way 
that only the investigator and site staff may associate data with an identified or identifiable 

individual or his/her medical record. All other parties involved in data management, analysis and 
storage will receive, and subsequently analyze, non-identifiable patient data. 
Data Confidentiality 
information furnished by Amicus to the investigator will be kept in confidence and such information 
will be divulged to any expert committee, affiliated institution, and employees only under an 
appropriate understanding of confidentiality with such committee, affiliated institution, and 
employees. 
Data handling and record keeping 
regulatory restrictions and ethical considerations, Amicus or any Regulatory Agency may consult 
and/or copy study documents to verify the included data. 
Record keeping and archiving 
According to Danish law and to Amicus policies.  
Only authorized persons will have access to identifiable personal details, if required for data 
verification. Data protection and privacy regulations will be observed in capturing, forwarding, 
processing, and storing subject data. If applicable, subjects will be informed accordingly and will 
be requested to give their consent on data handling procedures in accordance with national 
regulations. 
remain anonymous and confidential. Investigators, or investigator partners defined in annex 1 if 
applicable and various government health agencies may inspect the records of the study. Every 
 
During the development and analysis of the study, statistical program codes will be locally stored 
as documents according to best practices. Data protection laws in Denmark will be adhered to at 
all times. Further, the log file produced during the final run of each program will be stored locally 
at the investigator site as accompanying documents along with the results. 
Transfer of Essential Documents 
Not applicable  
Financing and insurance 
Financing 
Senior researchers 2x50% data management 700,000 DKK 
Specialist nurse to extract patient data 390,000 DKK 
2 Medical students to extract data and manage data files 240,000 DKK 
Meetings, IT access, literature review, publication costs 70,000 DKK 

Total 1,400,000 DKK 
Amicus sponsored  
Insurance 
Not applicable  
Management and reporting of adverse events/adverse 
reactions  
Adverse event (AE) 
Collection of adverse events as Individual Case Safety Reports (ICSRs) is not required from such 
retrospective analyses of patient medical records as it is secondary data. 
Plans for disseminating and communicating study results  
The study, and study results, will be disclosed on clinicaltrials.gov and other relevant clinical study 
disclosure websites in accordance with relevant AMICUS policies. The primary publication is 
expected to be in an internationally recognised journal, with the Scientific Committee forming the 
majority of the authorship. Thereafter, any secondary publications will reference the original 
publication and will be authored in accordance with International Committee of Medical Journal 
Editors (ICMJE) requirements. 
Where required by applicable regulatory requirements, an investigator signatory will be identified 
for the approval of the clinical study report. The investigator is encouraged to share the summary 
results with the study subjects, as appropriate.    
 
 

References 
1.  
Germain DP. Fabry disease. Orphanet J Rare Dis. 2010;5(1):30.  
2.  
Sanchez-Nino MD, Sanz AB, Carrasco S, Saleem MA, Mathieson PW, Valdivielso JM, et al. 
Globotriaosylsphingosine actions on human glomerular podocytes: implications for Fabry 
nephropathy. Nephrol Dial Transplant. 2011 Jun 1;26(6):1797 802.  
3.  
Effraimidis G, Rasmussen ÅK, Dunoe M, Hasholt LF, Wibrand F, Sorensen SS, et al. Systematic cascade 
screening in the Danish Fabry Disease Centre: 20 years of a national single-centre experience. 
Branchini A, editor. PLoS One. 2022 Nov 16;17(11):e0277767.  
4.  
Waldek S, Patel MR, Banikazemi M, Lemay R, Lee P. Life expectancy and cause of death in males and 
females with Fabry disease: Findings from the Fabry Registry. Genet Med [Internet]. 2009;11(11):790
6. Available from: https://doi.org/10.1097/GIM.0b013e3181bb05bb 
5.  
Ortiz A, Germain DP, Desnick RJ, Politei J, Mauer M, Burlina A, et al. Fabry disease revisited: 
Management and treatment recommendations for adult patients. Mol Genet Metab. 2018 
Apr;123(4):416 27.  
6.  
Feldt-Rasmussen U, Hughes D, Sunder-Plassmann G, Shankar S, Nedd K, Olivotto I, et al. Long-term 
efficacy and safety of Migalastat treatment in Fabry disease: 30-month results from the open-label 
extension of the randomized, phase 3 ATTRACT study. Mol Genet Metab. 2020 Sep;131(1 2):219 28. 
7.  
Schiffmann R, Bichet DG, Benjamin E, Wu X, Giugliani R. The Migalastat GLP-HEK assay is the gold 
standard for determining amenability in patients with Fabry disease. Mol Genet Metab Reports. 2019 
Sep;20(July):100494.  
8.  
Disease with the Pharmacologic Chaperone Migalastat. N Engl J Med. 2016;375(6):545 55.  
9.  
Hughes DA, Nicholls K, Shankar SP, Sunder-Plassmann G, Koeller D, Nedd K, et al. Oral pharmacological 
chaperone Migalastat compared with enzyme replacement therapy in Fabry disease: 18-month results 
from the randomised phase III ATTRACT study. J Med Genet. 2017 Apr;54(4):288 96.  
10.  
Schiffmann R, Bichet DG, Jovanovic A, Hughes DA, Giugliani R, Feldt-Rasmussen U, et al. Migalastat 
improves diarrhea in patients with Fabry disease: clinical-biomarker correlations from the phase 3 
FACETS trial. Orphanet J Rare Dis. 2018 Dec 27;13(1):68.  
11.  
Germain DP, Nicholls K, Giugliani R, Bichet DG, Hughes DA, Barisoni LM, et al. Efficacy of the 
pharmacologic chaperone Migalastat in a subset of male patients with the classic phenotype of Fabry 
disease and Migalastat-amenable variants: data from the phase 3 randomized, multicenter, double-
blind clinical trial and extension study. Genet Med. 2019 Sep;21(9):1987 97.  
12.  
Nowak A, Huynh Do U, Krayenbuehl P, Beuschlein F, Schiffmann R, Barbey F. Fabry disease genotype, 
phenotype, and Migalastat amenability: Insights from a national cohort. J Inherit Metab Dis. 2020 Mar 
30;43(2):326 33.  
13.  
Riccio E, Zanfardino M, Ferreri L, Santoro C, Cocozza S, Capuano I, et al. Switch from enzyme 
replacement therapy to oral chaperone Migalastat for treating fabry disease: real-life data. Eur J Hum 
Genet. 2020 Dec 9;28(12):1662 8.  
14.  
Müntze J, Gensler D, Maniuc O, Liu D, Cairns T, Oder D, et al. Oral Chaperone Therapy Migalastat for 
Treating Fabry Disease: Enzymatic Response and Serum Biomarker Changes After 1 Year. Clin 

Pharmacol Ther. 2019 May 13;105(5):1224 33.  
 
 
 

Annex 1. List of stand-alone documents 
Documents listed in Annex 1 can be maintained separately from the study protocol. They should 
documents in a table as indicated below. 
Number 
Date 
Section of study 
protocol 
Amendment or 
update 
Reason 
1 
xx.xx.2023 
Text 
Update 
Description of 
participating 
clinical 
scientists and 
research staff 
Annex 2. ENCePP checklist for study protocols  
A copy of the ENCePP Checklist for Study protocols available at 
http://www.encepp.eu/standards_and_guidances/index.html completed and signed by the main 
author of the study protocol should be included in Annex 2.  
The checklist will facilitate the review of the protocol and evaluation of whether investigators have 
considered important methodological aspects.  
In question 9.5 of the Checklist, Revision 1:  
 
 
 
 
Annex 3. Additional information  
Additional annexes may be included if necessary. 
 
 
 
 
 

List annex 1.1 Description of participating clinical scientists and
research staff