Pilot Study to Assess Clinical and Pivotal Biomarkers in the Urine to Predict the Progression of Nephropathy in Fabry Disease
Conditions: Fabry Disease
Sponsor: Lysosomal and Rare Disorders Research and Treatment Center, Inc.

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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CLINICAL STUDY PROTOCOL 
 
 
PROTOCOL TITLE: 
 
 
PROTOCOL NO:     
 
Pilot study to assess clinical and pivotal biomarkers in the urine to 
predict the progression of nephropathy in Fabry disease. 
23-LDRTC-01 
  
 
PRINCIPAL 
INVESTIGATOR: 
 
 
 
 
 
SITE(S):                              
 
 
Ozlem Goker-Alpan MD 
Lysosomal & Rare Disorders Research & Treatment Center, Inc  
3702 Pender Drive, Ste 170 
Fairfax, VA 22030 
Phone:  703-261-6220 
 
Lysosomal & Rare Disorders Research & Treatment Center, Inc  
3702 Pender Drive, Suite 170 
 Fairfax, VA 22030 
 
Lysosomal & Rare Disorders Research & Treatment Center, Inc  
121 Congressional Lane, Suite 320 
Rockville, Maryland 20853 
 
 
 
CO-INVESTIGATOR: 
 
Margarita Ivanova, PhD 
Lysosomal & Rare Disorders Research & Treatment Center, Inc  
3702 Pender Drive, Suite 170 
 Fairfax, VA 22030 
Phone:  571-732-4606 
 
 
 
 
 
 
 
 
Principal Investigator Signature:_________________________________Date: ____________ 
 
                              
                                                                                                               
 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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INTRODUCTION 
 
  Nephropathy is one of the major contributors to morbidity and mortality in Fabry disease (FD). 
The standard non-invasive clinical laboratory tests for kidney disease (creatinine, glomerular 
filtration rate (GFR), and cystatin C) are not sensitive enough and may delay the therapeutic 
decision-making process. Additionally, the disease-specific biomarkers globotriaosylceramide 
(Gb-3) and globotriaosylsphingosine (Lyso-Gb-3) are not considered clinical entities associated 
with kidney damage. Therefore, the precise detection of the presence and the progression of kidney 
involvement is crucial for not only the development of effective therapies but also the prevention 
of late and irreversible complications in FD.  
 
  In FD, the typical clinical signs of kidney involvement may be absent, but biopsies may already 
show chronic glomerular and interstitial damage with fibrosis early in the disease course. The renal 
histology is characterized with intracellular Gb-3 deposits, even with normal GFR and minimal 
proteinuria. A characteristic finding is that the vacuoles correspond to extracted Gb-3 deposits in 
podocytes and epithelial cells. Heterozygote females develop the same kidney involvement as 
hemizygous males with FD (1).  
 
 The progressive accumulation of Gb-3 and its deacylated form lyso-Gb-3 creates an inflammatory 
environment (Figure 1). Furthermore, the accumulated Gb-3 and lyso-Gb-3 have been released 
into the cellular milieu from the ruptured lysosomes, leading to cell damage (2). Naturally 
sequestered inside live cells, these molecules are passively released extracellularly upon cell death 
and loss of membrane integrity. The elevated levels of both molecules in plasma and urine correlate 
with the impact of Fabry disease and may correspond to the short-term response to therapy with 
the reversal of cellular apoptosis.  
 
 Cells can die through several different mechanisms. Two of the significant types of cell death are 
necrosis and apoptosis. Sphingolipids are associated with apoptosis, where cells die while 
maintaining the integrity of their plasma membrane but are ingested by the tissue macrophages, in 
turn releasing pro- or anti-inflammatory signals such as TGF-beta or IL-10 (3). In contrast, necrotic 
cell death occurs in response to an insult such as infection and is associated with rapid loss of cell 
membrane integrity. Pyroptosis is a form of necrotic cell death due to a  pro-inflammatory response 
(4). This pathway is uniquely dependent on Caspase-1, which is not involved in apoptotic cell 
death. Caspase-1 activation in macrophages results in the processing of Il-1 with subsequent 
activation of the transcription factor NF-κB.  
  
 The transcription factor NF-κB regulates multiple aspects of immune functions and serves as a 
pivotal mediator of inflammatory responses (5). NF-κB controls the expression of pro-

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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inflammatory cytokines (e.g., IL-1, IL-6, IL-8, and TNFα), chemokines (e.g., MIP-1, MCP1, 
RANTES, and Eotaxin), and vascular adhesion molecules (e.g., ICAM1, VCAM1, and E-selectin). 
NF-κB is also a central mediator of TGF-β1, the pathway central to the progression of fibrosis in 
multiple organs.  
  The responses induced by cell injury can cause significant damage while trying to heal the injured 
tissue. This is one of the reasons why it's important to elucidate these molecules in the urine not 
only to understand the order of pathogenic events associated with renal injury but also potentially 
block the cellular triggers of inflammation. Thus, this proposed study of pivotal urinary biomarkers 
may offer further insight on renal injury beyond current diagnostic tests.   
 
 
 
Figure 1. Fabry nephropathy. The pathogenesis leads to the activation of inflammatory cascades and 
fibrosis. The model was adopted from publication (6).   
 
   BRIEF RATIONALE AND BACKGROUND 
 
   Progressive nephropathy, glomerulosclerosis, tubular atrophy, and interstitial fibrosis are 
associated with inflammatory cellular infiltration. The Gb-3 accumulation in lysosomes leads to 
early cellular dysfunction followed by cell death, which activates inflammatory pathways and 
stimulates tissue fibrosis in Fabry disease. Progressive nephropathy, including glomerulosclerosis, 
tubular atrophy, and interstitial fibrosis, are associated with inflammatory cellular infiltration in 
the kidney. While Gb-3 accumulation is primarily located in endothelial and epithelial glomerular 
cells, histological studies demonstrate that the buildup of Gb-3 in podocytes plays an essential role 
in glomerular damage (7). Moreover, the study of urine-derived FD kidney epithelial cells and 
podocytes in vitro demonstrates that the cause of this damage lies in deregulated autophagy 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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pathways (7, 8). Deposition of Gb-3 in glomerular cells leads to focal segmental glomerular 
sclerosis. In addition, Gb-3 deposition in podocytes leads to activation of necroptosis (Figure 1). 
Podocyte injury, detachment, and podocyturia might be the initial pathogenic events in FD 
nephropathy (Figure 1). Gb3 accumulation in podocytes has been shown to resist ERT clearance 
partially. Moreover, the Gb3 accumulation in lysosomes continuously activates inflammatory 
pathways and stimulates pathogenetic processes, while the cellular clearance of Gb-3 by ERT 
limits histological damage. 
   It is well known that the NF-kB, TGF-β1, and Nothch1 signaling pathways play an essential role 
in nephropathy (Table 1 and 2). Moreover, Gb-3 deposition is associated with the increased release 
of TGF-β1 and epithelial-to-mesenchymal cell transition, leading to the over-expression of 
profibrotic molecules (Figure 1). TLR4 is involved in the initiation of this process leading to renal 
fibrosis via inflammasome activation in renal epithelial cells and, at the same time, dysregulated 
immune response (Tables 1 and 2).  
 
 
  Common clinical biomarkers related to FD nephropathy (alpha-1-microglobulin, microalbumin, 
creatinine) will be compared with the pivotal biomarkers associated with the inflammatory and 
dysregulated immune pathways.  
Table 2: Suggested pivotal biomarkers and pathways involved in Fabry  nephropathy: 
 
Relationship to pathway or 
involvement 
Biomarkers 
 
 
 
Inflammatory 
biomarkers 
Cytokines 
NF-kB pathway 
IL-1β, IL-6, TNF-α, Il-10 
TGF-β1 pathway 
TGF-β1, Il-4, INF-gamma 
TLR4 pathway 
Il-18, Il-12 
Chemokines 
NF-kB pathway 
MCP1, IL-8, RANTES, BAFF, APRIL 
TLR4 pathway 
PAI-1 
 
Biomarkers of kidney 
function and 
proteinuria 
clinical laboratory urine biomarkers 
alpha-1-microglobulin, albumin, cystatin C, 
GFR 
We will measure in our laboratory 
B2M, bikunin, NGAL, osteopontin, 
clusterin, creatinin. 
 
Acute kidney injury 
Marker of renal tubular injury 
KIM-1 
Inflammation-associated glycoprotein 
YKL-40 
Growth factor, fibrosis 
EGF 
Apoptotic marker 
CK-18 M30 
Pyroptotic markers 
IL-1β, IL-18 (included in the list of 
cytokines) 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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This is a study to assess the markers related to autophagy, apoptosis, pyroptosis, and inflammatory 
markers related to NFkB, TNF-alpha, and TGF-β1 pathways in the urine. Urinary biomarkers will 
then be compared to the standard measures of kidney function and proteinuria: GFR, cystatin-C, 
B2M, bikunin, NGAL. Gb3 and Lyso-Gb3, urine microalbumin, and urine protein-to-creatinine 
(UPCR) ratio. We will also analyze the role of therapy, especially for the inflammatory responses 
in patients on stable enzyme replacement therapy (ERT) with that of patients naïve to therapy.   
 
OBJECTIVE AND SPECIFIC AIMS 
 
 Primary objective:  Identify biomarkers in urine for the detection and progression of kidney 
involvement in naïve male and female patients with FD, and compare with patients on ERT to 
assess the effects of ERT.  
  Specific Aims: 
1) To identify urinary inflammatory biomarkers associated with nephropathy in FD. 
 
2) Analyze the association between urinary inflammatory markers, clinical urine 
biomarkers, circulating lyso-Gb3, and kidney involvement in naïve FD patients. 
 
3) Analyze the association between urinary inflammatory markers, clinical urine 
biomarkers, circulating lyso-Gb3, and kidney involvement in FD patients on stable ERT. 
 
4)  Assess the role of the time of initiation and duration of therapy (ERT), respectively, 
inactivation and reversibility of inflammation cascade, and how these relate to the degree 
of kidney involvement in patients with FD.  
 
SUBJECT SELECTION 
 
Number of Subjects  
 
A total of 50 subjects, male or female, between the ages of 18 to 80 years of age, include 25 
healthy individuals and 25 patients with  Fabry disease.  
 
Study population 
 
FD patients n=25 (5 without clinical evidence of nephropathy and 15 patients with clinical 
evidence of nephropathy),  NAÏVE FD patients (n=5), and healthy controls n=25 (controls, urine 
samples are not actual patients, urine samples will be purchased from a commercial company).       
 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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The study will involve 50 age-and gender-matched subjects and controls divided into the 
following cohorts: 
 
1. FD subjects without clinical evidence of nephropathy (normal eGFR and no 
microalbuminuria) naïve or on ERT (n=5),  
2. FD subjects with clinical evidence of nephropathy (n=15)  
           Subcohorts:  
a. FD subjects on ERT with CKD stage 1 (estimation  n=5) 
b. FD subjects on ERT with CKD stage 2 (estimation n=5) 
c. FD subjects on ERT with CKD type 3 a or b (estimation n=5) 
3. FD patients naïve to therapy (n=5). Can we be included in Cohort group 1 or 2. 
Additional analysis will be stratified NAÏVE vs. ERT.  
4.  Non-FD age-matched controls with no known kidney disease n=25.  
 
 
FD 
Healthy control 
Total FD 
25 
25 
without nephropathy 
5 
 
with nephropathy 
15 
 
NAIVE 
5 
 
 
Table 3. Clinical staging of FD patients:  
 
Clinical 
parameter(s) 
FD  patients and kidney involvement 
Age-matched 
controls 
Stage 1 
Stage 2 
Stage 3a 
Stage 3b 
Kidney involvement 
Kidney damage 
Normal/ 
Mild reduction 
Moderate 
reduction 
Moderate 
reduction  
normal 
GFR ml/m1.73 m2 
>90 
60-89 
45-59 
30-44 
90 min 
Cystatin-C 
0.5-0.9 
1.0-1.2 
1.3-1.5 
1.4-1.9 
0.5-0.9 
Urine B2M 
- 
0.428-0.603 
0.603-0.950 
0.950-1.034 
- 
FD biochemical markers 
α–Gal A 
Deficiency 
Deficiency 
Deficiency 
Deficiency 
Normal 
GLA mutations 
Yes 
yes 
yes 
yes 
No 
Gb-3 
Yes 
yes 
yes 
yes 
No 
Lyso-Gb-3 
Yes 
yes 
yes 
yes 
No 
 
Gender of Subjects 
Subjects of both genders will be included in the research, and it is expected that this inclusion 
will be representative of the FD patients population.  
 
Age of Subjects 
Subjects will be between the age of 18 to 80 years of age. 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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Racial and Ethnic origin 
 
There are no enrollment restrictions based on race or ethnic origin, and it is expected that racial 
and ethnic representations in this study will represent the FD population.  
 
INCLUSION/EXCLUSION CRITERIA 
 
Inclusion criteria: 
 
a. Male and Female subject is greater than 18 but not older than 80 years. 
b. Subject willing to sign the informed consent and/or assent. 
c. Confirmed diagnosis of Fabry disease based on deficient α–Gal A enzymatic activity and 
molecular analysis demonstrating pathogenic variants in the GLA gene.  
Exclusion criteria: 
a. Any other known genetic condition associated with CKD,  
b. Evidence of hepatitis B or C infections or other chronic infectious diseases, 
c. Pregnancy or breastfeeding.  
d. Any other chronic condition, as per PI's discretion, that makes the subject ineligible.  
Description of criteria for withdrawal from the study 
 
A subject may withdraw from the study at any time. The study is entirely voluntary. 
 
RECRUITMENT: 
 
Upon identifying patients with FD disease by the Principal Investigator, the research coordinator 
will recruit the study subjects. The research coordinator will recruit healthy controls in the clinic 
or purchase from a commercial company.   
 
SETTING/LOCATION  
 
 
 
 
 
 
 
       
Research subjects will be recruited at the Lysosomal and Rare Disorders Research and 
Treatment Center, Inc (LDRTC) located at 3702 Pender Drive, Suite 170,  Fairfax, VA 22030, 
and/or 121 Congressional Lane, Ste 320, Rockville, MD, 20852.  
 Subjects recruited at these facilities will be receiving advice from Ozlem Goker-Alpan, MD, and 
be under her clinical care. All experimental/laboratory procedures will be performed by or under 
the supervision of Margarita Ivanova, Ph.D. All informed consent processes will take place at the 
indicated research sites. 
 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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Laboratory tests will be performed at  Lysosomal & Rare Disorders Research & Treatment 
Center, Inc, whereas some assays such as  Lyso-Gb3 will be performed outside collaborating 
laboratories such as Labcorp Diagnostic  Laboratories and/or  Centogene, Rostock, Germany. 
 
STUDY PROCEDURES 
 
Study type 
 
 Descriptive and observational study of molecular components detected in biological specimens 
of urine. 
   
Study design and plan 
 
This prospective study aims to establish novel biomarkers for non-invasive detection of kidney 
involvement in FD. FD patients will be assessed using the definition of Kidney Disease Outcomes 
Quality Initiative (KDOQI) 2002. The patients with CKD stages 3 or less with a progressive 
decline in kidney function, as evidenced by a decreasing GFR and a slope of less than -1.5, will be 
further assessed.  
 
The glomerular filtration rate will be calculated from serum creatinine (CKD-EPI). The UPCR 
will be used to measure proteinuria. Additionally, the patients will be stratified as having low renal 
involvement (LRI) if the UPCR is less than or equal to 0.5 g/g. Anti-drug antibody status and other 
adverse events, such as the occurrence and the progression of cardiomyopathy as evidenced by 
LVMI, LVPWD, and neurological events such as strokes, will be recorded through the study. The 
categorization of kidney involvement via renal biomarkers will be defined (Table 4).  
 
Table 4. Clinical staging of FD patients:  
 
 
FD  patients and kidney involvement 
Age-matched 
controls 
Clinical 
parameter(s) 
Stage 1 
Stage 2 
Stage 3a 
Stage 3b 
Kidney involvement 
Kidney damage 
Normal/ 
Mild reduction 
Moderate 
reduction 
Moderate 
reduction  
Normal 
GFR ml/m1.73 m2 
>90 
60-89 
45-59 
30-44 
90 min 
Cystatin-C 
0.5-0.9 
1.0-1.2 
1.3-1.5 
1.4-1.9 
0.5-0.9 
Urine B2M 
- 
0.428-0.603 
0.603-0.950 
0.950-1.034 
- 
FD biochemical markers 
α–Gal A 
Deficiency 
Deficiency 
Deficiency 
Deficiency 
Normal 
GLA mutations 
Yes 
yes 
yes 
yes 
No 
Gb-3 
Yes 
yes 
yes 
yes 
No 
Lyso-Gb-3 
Yes 
yes 
yes 
yes 
No 
 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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The study will take place over 24 months. The clinical part of the active study to collect clinical 
data and the samples will be over 12 months. The urine samples will be collected three times ( 0, 
6, and 12 months) (Table 5). A physical examination will be performed during the visits, and 
samples will be collected. We will review medical records as a part of the clinical evaluation. After 
a urine specimen collection, samples will be aliquoted and stored at -800 C until biomarkers  are 
screened. We expect to recruit the subjects within 6 months of the study initiation; thus, the last 
visit for the last subject is expected to be 19 months after the study starts. 
 
Table 5. Visit and assessment schedule: 
 
baseline 
 
 
visits 
1 
2 
3 
month 1/ 
enrollment 
month 
6 
month 
12 
Vital signs  
X 
X 
X 
Height and weight  
X 
 
X 
Physical examination 
X 
 
X 
Concomitant medications  
X 
X 
X 
Adverse event reporting 
X 
X 
X 
Informed consent form 
X 
 
 
Eligibility (inclusion/exclusion) 
X 
 
 
Urine pregnancy test (part of eligibility) 
X 
 
 
HBV/HCV testing ( if clinical results are not available ) 
 
 
 
Medical history 
X 
X 
X 
Retrospective chart review 
X 
 
 
Urine sample collection and urinalysis 
X 
X 
X 
 
STUDY METHOD AND PROCEDURES 
 
Informed consent will be obtained prior to the execution of any research procedures. The subjects 
will be followed for a 12 month period during which urine will be collected three times every six 
months. Subsequent samples will be obtained based on the Principal Investigator's order. 
Biological samples are only collected for research purposes and will not be sold or distributed for 
other purposes.  
After the study is completed, biological samples may be retained for possible future exploratory 
analysis by the Principal Investigator and/or by the Co-Investigator.   
   
CONSENT/ASSENT: 
 
Enrollment of a subject or a family member in this study will be completed once all inclusion and 
no exclusion criteria have been met and the subject has signed the informed consent form (ICF). 
The clinical research coordinator will approach subjects identified by the Principal Investigator.  
The Principal Investigator will discuss the research study with the subject explaining research 
procedures, potential risks, and benefits. The Principal Investigator will give the subject an 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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opportunity to ask questions, and ample time will be given to the subject to review the ICF in 
detail. The clinical research coordinator will repeat this process using verbiage found in the consent 
to explain the study. 
The alternative method for consenting:  
The ICF could be emailed or mailed to the patient with a pre-paid addressed return envelope 
provided for convenience. The patient should be contacted at a later time to confirm receipt of the 
ICF and be given an opportunity to ask questions. The potential subject will have ample time to 
review the ICF in detail. Once all questions have been answered to the subject's satisfaction, 
research personnel should provide guidance as to what page requires the subject's printed name, 
signature, date, and witness signature (if applicable). A copy of the original signed ICF will be 
provided to the patient. 
DEMOGRAPHICS  AND MEDICAL HISTORY 
 
A review of demographic parameters, including age, gender, race, and ethnicity, will be performed 
at screening.  
Past and present medical and surgical history will be recorded according to the Schedule of 
Assessments. Any ongoing condition and signs and symptoms observed prior to the initiation of 
the study will be recorded as past medical history.  
Disease history specific to FD will be recorded and includes the date of diagnosis, primary 
symptoms at time of diagnosis, and disease progression history. A separate chart review will be 
conducted as indicated in the schedule of events to assess Fabry disease-related kidney, cardiac, 
and nervous system involvement, history of strokes and presence and nature of pain, and the 
history of acute pain crises, history of the treatment of FD including Enzyme Replacement therapy, 
and other treatments such as ACE and ARBs for proteinuria, presence of anti-drug antibodies and 
history of immune hypersensitivity reactions will be specifically noted. 
CONCOMITANT MEDICATIONS AND PROCEDURES  
Subjects will be asked to report any new or changes in previously reported prescription and non-
prescription medications, including dosage, frequency, and administration dates. Information will 
be entered in the source records. Information regarding any medical or surgical procedures 
performed since the last visit will also be collected. 
URINALYSIS  
In addition to the exploratory laboratory tests, a clinical urinalysis will be performed.    
Urinalysis will include the following: Specific gravity, appearance, color pH, glucose, protein 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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(total), ketones, bilirubin, urobilinogen, hemoglobin, leucocyte esterase, reducing substances, 
and nitrite.  
OTHER LABORATORY TEST  
Virus Serology  
Subjects are required to have negative serology test results for hepatitis B surface antigen and 
hepatitis C antibody at screening to participate in the study. 
Use of Biological Samples for Future Research 
Researchers may use leftover samples from already collected and deidentified biological samples 
obtained from consented subjects during the study for other future investigations  
Vital Signs  
Vital signs will include temperature, blood pressure, respiration rate, and heart rate. Vital signs 
will be measured as specified in the Schedules of Assessments (1) and as needed per PI.  
Physical Exam  
A physical examination will be performed by a study physician or designee, as specified in the 
Schedule of Assessments (Table 2). The physical examination includes skin, head, ears, eyes, nose, 
throat, heart, lungs, abdomen, and neurologic systems. An additional examination may be 
performed by the Investigator. 
Weight and Height  
Weight and height will be measured using proper scales. No bulky clothes or shoes should be 
worn.  
 
BENEFITS: 
There are no immediate benefits to subjects for participation; however, knowledge obtained from 
this study may lead to improved understanding, diagnosis, and therapeutic interventions in the 
future. 
 
SUBJECT COMPENSATION:  
Costs: There are no costs to subjects. Subjects will not be charged for the costs of procedures or 
sample analysis. 
Payment: No subject compensation will be provided.  
HUMAN SUBJECT PROTECTIONS  

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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Risks: 
 
There is some risk to subjects participating in this study: 
There are risks of personal information being inadvertently released. The research team will try its 
best to protect research participants' health and personal information. To minimize this risk, only 
researchers at Lysosomal & Rare Disorders Research & Treatment Center, Inc will have 
access to biological specimens collected in this protocol, and all specimens will be deidentified. In 
the case of any sample being shared with other researchers, they will only receive specimens 
labeled with a generic study identification number that cannot be linked back to the research 
subject. No other identifying information will be shared with them. 
Alternatives: 
 
The subject has the right to decide not to participate in the research study. 
 
Confidentiality: 
 
All patient data will remain confidential in accordance with the Health Insurance and Portability 
and Accountability Act (HIPAA). No patient identifiers will be used to label biological specimens. 
All biological specimens will be deidentified using a generic identification number. 
Adverse Event Reporting: 
 
All adverse events will be reported to the IRB according to the Internal Conference of 
Harmonization (ICH) Good Clinical Practice (GCP) adverse reporting guidelines section 6 and 
Lysosomal & Rare Disorders Research & Treatment Center, Inc standard operating 
procedures section 6 regarding reporting of adverse events and unanticipated problems. 
A
 (AE) 
:
,
•
•
•
•

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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:
•
•
•
•
•
•
•
•
•
•
•
•
•
•
.

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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LDRTC is responsible for reporting serious and non-serious adverse drug reactions suspected of 
being related to to regulatory authorities. The Investigator is responsible for reporting adverse 
drug reactions to the IRB/EC, if required by national law or regulation, within the timelines 
required by such law or regulation. The Investigator shall maintain records of all such 
submissions.  
 
Study-specific committees 
 
 No steering committee, data safety monitoring committee, or clinical endpoint committee will 
be used in this study.  
 
STUDY OVERSIGHT 
 
Premature Termination of Study:  
Ozlem Goker-Alpan, MD, may prematurely terminate the study. Reasons for termination may 
include but is not limited to the following: 
• Series of adverse events or incidences associated with subject participation. 
• Low subject enrollment. 
 
Clinical Research Quality Assurance:  
To ensure that GCP and good clinical research are conducted per regulatory requirements and 
guidelines, the study will be made available for monitoring, auditing, review, and inspection by 
regulatory authorities such as IRB and FDA. This will be accomplished by providing direct access 
to study-related documents.   
Data management 
 
The acquired data will be entered into Excel for further analysis. Data will be analyzed using 
GraphPad Prism software (Graphpad Inc., La Jolla, CA) using ANOVA analysis and Student's t-
test for statistical analysis. p<0.05 is considered significant.  
Samples and data storage 
Once subjects have signed the informed consent form, they will be assigned a personal generic 
research study identification number. This number will be matched with the patient's name and 
demographic information in a file stored on a password-protected computer. Only members of the 
research team at Lysosomal & Rare Disorders Research & Treatment Center, Inc will have 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
15 
 
access to this file. Urine samples shipped for further analysis and viable cells stored long-term in 
liquid nitrogen freezers will be done without any identifiers attached. Research data from urine 
analysis will be stored in a password-protected separate file shared online between researchers at 
Lysosomal & Rare Disorders Research & Treatment Center, Inc. 
IRB REVIEW/ ETHICS/ INFORMED CONSENT 
 
The protocol, informed consent, and all other relevant documents will be submitted to the IRB for 
review, and approval will be obtained before the study is initiated per the required guidelines. This 
study will be conducted with responsible clinical research practices in accordance with the ethical 
principles that have their origins in the Declaration of Helsinki and that are congruent with GCP 
guidelines established at the ICH and other applicable regulatory requirements. The study will be 
conducted in accordance with the regulations of the United States Food and Drug Administration 
(FDA) as described in 21 CFR 50 and 56, applicable laws, and IRB requirements. 
The Principal Investigator will submit any changes to the protocol to the IRB for review and 
approval before implementing such changes. However, a protocol change will be implemented 
immediately if the change is intended to eliminate or minimize harm to subjects. Such change will 
be notified to the reviewing IRB and the FDA within 10 working days.  
The Principal Investigator is responsible for providing each subject with full and adequate verbal 
and written information using the IRB-approved informed consent form as described above. 
Informed consent will be obtained prior to performing any study-related procedures. A copy of the 
signed consent form will be given to the subject.  
Clinical research quality assurance  
To ensure GCP and good clinical research conduct per regulatory requirements and guidelines, the 
study will be made available for monitoring, auditing, review, and inspection to regulatory 
authorities such as IRB and FDA. This will be accomplished by providing direct access to study-
related documents.  
 LDRTC 
will
I
review 
 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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 DATA HANDLING AND RECORDKEEPING  
The 
Site 
will 
collect 
patient 
information 
from 
patient 
medical 
charts. 
The Site will follow its internal procedures for the Development of Study Specifications for the 
Development of the password-protected Excel, where patient data will be extracted. The Site will 
follow their internal procedures for the testing and live production of the study-specific database. 
The Site will document their entry method, QC process, and if entry/readability of file is limited 
to certain staff. The Site will keep Master Study File where they will document data entry 
processes. The Site will follow their internal procedures for the management of user access to the 
system and database. The Site will have data transfer and quality control procedures in place and 
documented to minimize data transcription errors if applicable. All Site staff involved in the 
handling of study data will have documented training on the relevant procedures. 
The Site will ensure that protected health information is handled in compliance with national 
privacy/data protection laws and regulations. The Site will ensure that all personal health 
information has been de-identified. Study data and patient records will be archived per the Site's 
policies and will be made available to Regulatory Agency upon request. AEs, medical history, and 
concurrent medical conditions will be coded using the most recent Medical Dictionary for 
Regulatory Activities. 
CONFIDENTIALITY 
 
All subject data and personal information will remain confidential in accordance to the Health 
Portability and Accountability Act and the Privacy Rule. However, all subject study-related 
records upon request may be disclosed to regulatory agencies such as the United States FDA and 
overseeing IRB for inspection. If shared with them, the Privacy Rule no longer covers subject 
information, but these entities are also committed to keeping subject information confidential. 
Subject information will be kept confidential and will not be made publicly available. If any results 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
17 
 
are published as a result of this research study subject identity will remain confidential, as only 
aggregate data will be reported.  
 
A description of this clinical trial will be available on http://www.ClinicalTrials.gov. 
This Web site will not include information that can identify the subjects. At most, the Web site 
will include a summary of the results. Subjects can search this Web site at any time. 
 
INTENDED USE OF DATA 
 
The data collected and analyzed will be used to understand the renal function in Fabry disease 
better. Data may be used and stored until all research-related activities are completed. 
 
 
REFERENCES 
 
1. 
Fogo AB, Bostad L, Svarstad E, Cook WJ, Moll S, Barbey F, et al. Scoring system for renal 
pathology in Fabry disease: report of the International Study Group of Fabry Nephropathy (ISGFN). 
Nephrol Dial Transplant. 2010;25(7):2168-77. 
2. 
Waldek S, Feriozzi S. Fabry nephropathy: a review - how can we optimize the management of 
Fabry nephropathy? BMC Nephrol. 2014;15:72. 
3. 
Young MM, Kester M, Wang HG. Sphingolipids: regulators of crosstalk between apoptosis and 
autophagy. J Lipid Res. 2013;54(1):5-19. 
4. 
Yu P, Zhang X, Liu N, Tang L, Peng C, Chen X. Pyroptosis: mechanisms and diseases. Signal 
Transduct Target Ther. 2021;6(1):128. 
5. 
Rozenfeld P, Feriozzi S. Contribution of inflammatory pathways to Fabry disease pathogenesis. 
Mol Genet Metab. 2017;122(3):19-27. 
6. 
Feriozzi S, Rozenfeld P. Pathology and pathogenic pathways in fabry nephropathy. Clin Exp 
Nephrol. 2021;25(9):925-34. 
7. 
Liebau MC, Braun F, Hopker K, Weitbrecht C, Bartels V, Muller RU, et al. Dysregulated autophagy 
contributes to podocyte damage in Fabry's disease. PLoS One. 2013;8(5):e63506. 
8. 
Slaats GG, Braun F, Hoehne M, Frech LE, Blomberg L, Benzing T, et al. Urine-derived cells: a 
promising diagnostic tool in Fabry disease patients. Sci Rep. 2018;8(1):11042. 

 
Protocol Number: 23-LDRTC-01 
PI: Ozlem Goker-Alpan  
Protocol version: June 15, 2023  
Lysosomal & Rare Disorders Research & Treatment Center Inc 
 
 
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