Pilot Study to Assess the Effects of Discontinuation of Vitamin K Antagonists on the Rate of Elastin Degradation
Conditions: Emphysema or COPD, Aneurysm, Cystic Fibrosis, Alpha 1-Antitrypsin Deficiency
Sponsor: Canisius-Wilhelmina Hospital

NL60536.091.17 / VKAs and elastolysis
RESEARCH PROTOCOL
Pilot study to assess the effects of 
discontinuation of vitamin K antagonists on the
rate of elastin degradation 
(August 2017)

NL60536.091.17 / VKAs and elastolysis
PROTOCOL TITLE 
The effects of discontinuation of vitamin K antagonists on the rate of elastin 
degradation
Protocol ID
Short title
VKAs and elastolysis
EudraCT number 
NA
Version
4.0
Date
August 21 2017
Coordinating investigator/project 
leader
Rob Janssen, MD, PhD
Department of Pulmonary Diseases
Canisius Wilhelmina Hospital
Weg door Jonkerbos 100
6532 SZ Nijmegen
{ HYPERLINK "mailto:rob.janssen@cwz.nl" }
Principal investigator(s)
Ianthe Piscaer, MD
Department of Respiratory Medicine
Maastricht UMC+
P.Debyelaan 25
6229 HX Maastricht
The Netherlands
Ruben Zaal, MD
Department of Pulmonary Medicine

NL60536.091.17 / VKAs and elastolysis
Canisius-Wilhelmina Hospital 
Weg door Jonkerbos 100
6532 SZ Nijmegen
The Netherlands
Lucie Bloem-de Vries, MD
Head of Comittee Anticoagulation
INR Anticoagulation Clinic – Canisius Wilhelmina 
Hospital
Burgemeester Daleslaan 27 
6532 CL Nijmegen
l.bloem-devries@cwz.nl
Sponsor
Rob Janssen, MD, PhD 
Department of Pulmonary Diseases
Canisius Wilhelmina Hospital
Weg door Jonkerbos 100
6532 SZ Nijmegen
{ HYPERLINK "mailto:rob.janssen@cwz.nl" }
Independent expert (s)
Sami Simons, MD, PhD
Department of Respiratory Medicine
Gelre Ziekenhuizen
Albert Schweizerlaan 31
7334 DZ  Apeldoorn
sami.simons@gelre.nl
Laboratory sites 
Department of Clinical Chemistry 
Canisius Wilhelmina Hospital
Weg door Jonkerbos 100
6532 SZ Nijmegen

NL60536.091.17 / VKAs and elastolysis
R&D Group Vitak
Daughter of the Maastricht University Holding
Oxfordlaan 55
6229 EV Maastricht
Centraal Diagnostisch Laboratorium
Maastricht UMC+
P.Debyelaan 25
6229 HX Maastricht
Pharmacy 
NA

NL60536.091.17 / VKAs and elastolysis
PROTOCOL SIGNATURE SHEET
Name
Signature
Date
Head of Department:
Maarten Wiggers de Vries
Operations Manager Pulmonology
Canisius Wilhelmina Hospital
Principal Investigator:
Rob Janssen, MD, PhD
Pulmonologist 
Canisius Wilhelmina Hospital

NL60536.091.17 / VKAs and elastolysis
TABLE OF CONTENTS
{ TOC \o "1-3" \u }

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LIST OF ABBREVIATIONS AND RELEVANT DEFINITIONS 
AE
Adverse Event
AR
Adverse Reaction
CA
Competent Authority
CCMO
Central Committee on Research Involving Human Subjects; in Dutch: 
Centrale Commissie Mensgebonden Onderzoek
COPD
DOAC
Chronic Obstructive Pulmonary Disease
Direct oral anticoagulant
DSMB
Data Safety Monitoring Board
Dp-uc MGP
Dephosphorylated uncarboxylized Matrix Gla Protein
EudraCT
European drug regulatory affairs Clinical Trials 
IC
Informed Consent
IMP
Investigational Medicinal Product 
IMPD
Investigational Medicinal Product Dossier 
INR
International normalized ratio
METC 
Medical research ethics committee (MREC); in Dutch: medisch ethische 
toetsing commissie (METC)
MGP
Matrix Gla Protein
pDES
Plasma Desmosine
SAE
Serious Adverse Event
SD
Standard deviation
SNP
Single nucleotide polymorphism
Sponsor
The sponsor is the party that commissions the organisation or 
performance of the research, for example a pharmaceutical
company, academic hospital, scientific organisation or investigator. A 
party that provides funding for a study but does not commission it is not 
regarded as the sponsor, but referred to as a subsidising party.
SUSAR
Suspected Unexpected Serious Adverse Reaction
VKA
Vitamin K antagonist
VKORC1
Vitamin K 2,3-epoxide reductase complex 1
WBP
Personal Data Protection Act (in Dutch: Wet Bescherming 
Persoonsgevens)
WMO
Medical Research Involving Human Subjects Act (in Dutch: Wet 
Medisch-wetenschappelijk Onderzoek met Mensen

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SUMMARY
Rationale:
Elastin is a unique protein providing elasticity, resilience and deformability to dynamic 
tissues, such as lungs and vasculature. Elastin fibers are characterized by their high affinity 
for calcium. However, calcified elastin is more prone to the degrading effects of proteases 
and, in turn, partially degraded elastin has an even higher affinity for calcium. A disturbed 
balance between proteases and anti-proteases is a major underlying mechanism in the 
development of chronic obstructive pulmonary disease (COPD). Virtually the only protein that 
can protect elastin from calcification is matrix Gla-protein (MGP), which needs vitamin K for
its activation. In COPD patients, a lower vitamin K status is found when compared to control 
subjects and an inverse association exists between vitamin K status and elastin degradation. 
In addition, vitamin K status is lower and elastin degradation is accelerated in Vitamin K 
antagonist (VKA) users.  
VKAs are widely used. Nowadays, an increasing number of patients uses direct oral 
anticoagulants (DOACs), which do not influence vitamin K status.  We hypothesize that 
discontinuation of VKAs results in an improved vitamin K status and deceleration of elastin 
degradation. In order to test this hypothesis, we want to conduct an observational pilot study 
in which the change in elastin degradation– quantified by plasma desmosine concentrations 
– in patients who discontinue use of VKAs will be used as primary endpoint.
Objective: To evaluate whether discontinuation of VKAs results in a higher vitamin K status 
and deceleration of the rate of mature cross-linked elastin degradation.
Study design: Observational study.
Study population: A total of 30 VKA users who will discontinue the use of VKAs.   
Main study parameters/endpoints: The primary endpoint is the change in the rate of 
elastin degradation quantified by the plasma desmosine assay. Secondary endpoints are the
change in vitamin K status quantified by measuring plasma levels of dephosphorylated 
uncarboxylated, (i.e. dp-uc, inactive) Matrix Gla Protein (MGP), the relation between 
desmosine and dp-ucMGP and differences of desmosine and dp-ucMGP levels among 
subjects with different polymorphisms of the vitamin K 2,3-epoxide reductase complex 1 
(VKORC1) gene. 
Nature and extent of the burden and risks associated with participation, benefit and 
group relatedness: We will draw extra blood collection tubes at two moments. The first time 
is during one of the last regular INR testing at the anticoagulation clinic, therefore no 
additional venipuncture has to be performed. At this moment two additional blood collection 
tubes will be drawn. The second moment is approximately 6 months after discontinuation of 
VKAs. At this point the patient will undergo an additional venipuncture. Patients are asked to 

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fill in a questionnaire concerning age, gender, length, height, indication for VKA use, duration 
of VKA use, smoking status and history, presence of COPD and other pulmonary disease, 
whether patients will switch to DOACs or will completely stop the use of anticoagulants, 
vitamin D supplementation, use of multivitamins and use of medications. Furthermore, we 
will record the INR at baseline. Participants may experience some discomfort during blood 
sampling. Participating in the study has negligible risks. 
1. INTRODUCTION AND RATIONALE
Elastin is a unique protein providing elasticity, resilience and deformability to dynamic 
tissues, such as lungs and vasculature.1 Elastin fibers are almost exclusively developed 
before birth and in the first period of life after which the synthesis of elastin is suppressed at 
a post-transcriptional level.1 Elastin can be degraded by certain proteases, also referred to as 
elastases.1 Elastin has high affinity for calcium,1 and the calcium content of elastin fibers 
therefore increases during aging.2 Calcium changes the conformation of elastin, which gives 
elastases access to elastin fibers leading to degradation.3,4 In turn, partially degraded elastin 
has a higher affinity for calcium due to increased polarity.4 Therefore, elastin calcification and 
elastin degradation are two pathologic processes that stimulate each other. When elastin is 
degraded, desmosine, a component of mature elastin, can be detected in the bloodstream. 
Since desmosine is only found in mature elastin, its plasma levels are a reflection of the rate 
of systemic elastin degradation.5
Chronic obstructive pulmonary disease (COPD) is characterized by a disturbed balance 
between proteases and anti-proteases, resulting in an accelerated degradation of elastin in 
the lungs, but also in blood vessels.6 Besides elastin degradation, calcification of elastin, i.e. 
in coronary arteries, is also accelerated in patients with COPD.7
Matrix Gla protein (MGP) is a potent inhibitor of elastin calcification and needs vitamin K to 
become activated. In an observational epidemiological study, higher vitamin K2 intake was 
associated with less vascular calcification.8 In addition, three years of vitamin K2 
supplementation in postmenopausal women resulted in a decrease of arterial stiffness, while 
an increase was observed in the placebo group.9 The dephophorylated uncarboxylated (i.e. 
inactive) form of MGP, dp-ucMGP, can be measured in plasma and its levels are negatively 
associated with vitamin K status. Therefore, dp-ucMGP levels are used as a measure for 
vitamin K status. 

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Earlier, we found a reduced vitamin K status in patients with COPD compared with patients 
without COPD and an inverse association between vitamin K status (quantified by dp-ucMGP 
levels) and elastin degradation (quantified by desmosine levels) among COPD patients 
(abstract ATS 2016). Recently, in 106 VKA users we found higher desmosine and dp-ucMGP 
levels when compared with patients who do not use VKAs. In VKA users, we also found an 
inverse association between vitamin K status and elastin degradation (quantified by plasma 
dp-ucMGP and desmosine levels, respectively). 
During activation of vitamin K dependent proteins such as MGP, vitamin K is oxidated from 
vitamin K 2,3-epoxide to vitamin K hydroquinone. Subsequently, vitamin K 2,3-epoxide 
needs to be recycled to the quinone form by an enzyme called vitamin K 2,3-epoxide 
reductase complex 1 (VKORC1).  Different single nucleotide polymorphisms of this gene 
exist, each resulting in a different level of expression of the enzyme. Since the VKORC1 
enzyme is the direct pharmacologic target of VKAs, this gene is for an important part 
responsible for the interindividual dose variability of VKAs.  The rs9923231 SNP (g.-
1639G.A) is the main genetic determinant of VKA dose requirement and can alone explain 
between 25% to 30% of the dose variance among patients.10
Nowadays, oral anticoagulants exist which do not influence vitamin K status, the so-called 
direct oral anticoagulants (DOAC). An increasingly number of patients is switching from 
VKAs to DOACs. 
We hypothesize that discontinuation of VKAs results in an improved vitamin K status and a
deceleration of elastin degradation. In order to test this hypothesis, we want to conduct an 
observational pilot study in which the change in elastin degradation– quantified by plasma 
desmosine concentrations – in patients who discontinue use of VKAs will be used as primary 
endpoint.
2. OBJECTIVES
We hypothesize that in persons who use VKAs, discontinuation of vitamin K antagonists
leads to a deceleration of elastin degradation.
Primary Objective:  
To assess the change in the rate of elastin degradation after discontinuation of vitamin K 
antagonists in patients who use VKAs. Quantification of the elastin degradation will be done 

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by measuring plasma desmosine concentrations at baseline and at 6 months after 
discontinuation.
Secondary Objective(s): 
To evaluate the possible effects of discontinuation of VKAs on dp-ucMGP (inversely 
correlated with vitamin K-status).
To evaluate if different polymorphisms of the VKORC1 gene are associated with desmosine 
and dp-ucMGP levels and the change of both parameters after discontinuation of VKAs. 
3. STUDY DESIGN
The study is designed as a non-interventional study. The follow up per patient comprises 6 
months. We will include 30 VKA users who will stop the use of VKAs. At baseline, patients 
are asked to fill in a questionnaire and two additional samples of blood are taken during the 
regular testing of the international normalized ratio (INR). One blood sample is needed for 
determination of the VKORC1 polymorphisms and one for desmosine and dp-ucMGP levels. 
After 6 months, patients are asked to draw one additional blood sample for determination of 
desmosine and dp-ucMGP. At baseline and 6 months blood can be drawn in the Canisius 
Wilhelmina Hospital or one of the outposts of the anticoagulation clinic. Since the blood 
samples have to be stored and frozen in the laboratory in the Canisius Wilhelmina hospital 
within 2 hours after drawing, patients can visit the outposts within one hour before the blood 
samples are transported to the laboratory, i.e. within one hour before closing the outpost. If 
both visiting the hospital or outpost is not possible, the patient can contact the investigator to 
make an appointment for drawing the blood samples at home. 

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An overview of is given in Figure 1.
Figure 1. Overview of study and time schedule
                      
                                                                                        
No VKA use 
0 months
- 2 blood collection tubes during the 
regular visit 
- Questionnaire
6 months
- 1 blood sample during 
additional visit
-Questionnaire
VKA use

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4. STUDY POPULATION
4.1 Population (base)  
We will include 30 VKA users who are going to discontinue the use of VKAs at short time 
4.2 Inclusion criteria 
In order to be eligible to participate in this study, a subject must meet all of the following 
criteria:
-
Use of VKAs for at least 3 months
-
Stop VKAs at short time
-
Written informed consent
-
Age ≥18 years
-
Ability to comply with all study requirements
4.3 Exclusion criteria 
A potential subject who meets any of the following criteria will be excluded from participation 
in this study:
-
Active malignancy or cured malignancy <12 months prior to enrollment
-
Use of maintenance dose oral corticosteroids
-
Serious mental impairment
-
Life expectation of less than 6 months on the basis of concurrent disease
4.4 Sample size calculation 
It concerns a pilot study and therefore sample size calculation was not performed. 
5. METHODS
5.1 Study parameters/endpoints 
5.1.1
Main study parameter/endpoint 
The primary endpoint is the difference in elastin degradation rate, quantified by the change in 
plasma desmosine levels, measured at baseline and 6 months after discontinuation of VKAs.

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5.1.2
Secondary study parameters/endpoints  
Secondary endpoints are:
-
The difference in vitamin K status, quantified by the change in dp-ucMGP, measured 
at baseline and 6 months after discontinuation of VKAs.
-
The association between desmosine and dp-ucMGP in patients who use VKAs and do 
not use VKAs. 
-
Differences in desmosine and dp-ucMGP levels between different VKORC 
polymorphisms. 
5.1.3
Other study parameters  
In addition, the following parameters will be registered, because of their possible confounding 
capacity:
-
Gender
-
Age
-
INR
-
Indication for VKA use
-
Duration of VKA use
-
BMI
-
Smoking status and history
-
Presence of COPD
-
Presence of other pulmonary disease
-
Medication 
-
Vitamin D supplementation
-
Use of multivitamins
-
Switch to DOACs or complete discontinuation of anticoagulant use
5.2 Study procedures 
All participating subjects are asked to fill in a questionnaire at home and will be invited to visit 
the hospital at baseline and approximately 6 months after discontinuation of VKA use for 
taking blood samples. The first blood sample will be drawn during (one of) the latest regular 
control of their INR at the coagulation clinic, so this will not involve an extra visit or 
venipuncture. Patients will be invited to draw blood approximately 6 months after 
discontinuation of VKAs. This concerns an additional visit besides the regular care and 
patients will undergo an extra venipuncture. 
Questionnaire

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Patients are asked to fill in a questionnaire at home, concerning gender, age, indication for 
VKA use, length, height, smoking status and history presence of COPD, presence of other
pulmonary disease, vitamin D supplementation, use of multivitamins and medication use, 
whether subjects switch to DOACs or completely discontinue the use of anticoagulants.
Vena punction:
Two non-fasting blood samples of 15mL (EDTA-plasma) and 10mL (EDTA-plasma) will be 
drawn at baseline. Since patients have regular INR determination at this moment, no extra 
venipunction has to take place. After six months one non-fasting blood sample of 15mL 
(EDTA-plasma) will be drawn. At this moment patients will have an additional vena punction. 
All samples will be collected in the Canisius Wilhelmina Hospital. EDTA-plasma will be 
centrifuged, divided in two portions of ~1 ml and frozen at -80°C within 2 hours after drawing. 
One portion will be used for quantification of vitamin K status and one for quantification of the 
rate of elastin degradation. DNA will be isolated from the samples of 10mL, which will be 
divided in two portions and frozen at -80°C.
Measurement of plasma desmosine levels:
The rate of elastin degradation will be quantified by measuring plasma (p)DES levels. 
Subjects with the highest pDES are assumed to have the highest rates of elastin 
degradation. Isodesmosine and desmosine fractions are measured separately by liquid 
chromatography-tandem mass spectrometry as previously described using deuterium-
labelled 
desmosine 
as 
internal 
standard 
({
HYPERLINK 
"http://www.ncbi.nlm.nih.gov/pubmed/?term=17494786" 
};
{
HYPERLINK 
"http://www.ncbi.nlm.nih.gov/pubmed/?term=23727558" }). 
Measurement of plasma dp-ucMGP levels:
Plasma dp-ucMGP will be measured using a dual-antibody test based on the previously 
described sandwich ELISA developed by VitaK (Maastricht, The Netherlands) ({
HYPERLINK "http://www.ncbi.nlm.nih.gov/pubmed/?term=18401181" }). The intra- and inter-
assay variations of this ELISA are 7.6% and 6.8%, respectively, and its sensitivity is 50 
pmol/L. Dp-ucMGP levels are very stable for up to fifteen years when frozen at -80°C. The 
within subject variability of dp-ucMGP is also very limited; in a cohort study little change in 
dp-ucMGP was observed after 5-6 years.
Determination of VKORC1 polymorphisms
High Pure PCR Template Preparation Kit (Roche Diagnostics, Mannheim, Germany) will be 
used for isolation of DNA, according to the manufacturer’s instructions. For genotyping the 

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C1173T (rs 9934438) and G-1639A (rs 9923231) single nucleotide polymorphisms of the 
VKORC1 gene, real-time PCR Fluorescence Resonance Energy Transfer (FRET) analyses 
will be performed. FRET LightMix® assays (cat.-no 40-0302-16, TIB MOLBIOL, Berlin, 
Germany) on the LightCycler® (Roche Diagnostics) will be used, according tot he 
manufacturer’s protocols. The assay consists of a duplex reaction measuring the melting 
curves of the used specific fluorescent probes in two different channels, each with a distinct 
wavelenght. With each run positive (heterozygote, provided with te kit) and negative 
controles will be determined. 
5.3 Withdrawal of individual subjects 
Subjects can leave the study at any time for any reason if they wish to do so without any 
consequences. The investigator can decide to withdraw a subject from the study for 
urgent medical reasons.
General criteria for withdrawal of individual subjects 
-
Patients should be withdrawn if VKAs are restarted before all blood samples are 
drawn.
-
Patients should also be withdrawn at any time if the investigator concludes that it 
would be in the patient’s best interest for any reason. 
-
Patients may voluntarily withdraw from the trial for any reason at any time. They may 
be considered withdrawn if they state an intention to withdraw, fail to return
visits, become lost to follow up for any other reason, or if any of the following 
occurs: discovery of failure of randomization; discovery of patient ineligibility; errors 
in treatment compliance; missed/unscheduled/off-schedule/incomplete/incorrect 
assessments.
If premature withdrawal occurs for any reason, the investigator must determine the primary 
reason for the patient’s premature withdrawal from the trial and record this information in the 
CRF.
5.4 Replacement of individual subjects after withdrawal 
Up to a maximum of 5 subjects who withdraw from the study will be replaced.

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5.5 Follow-up of subjects withdrawn from treatment 
Patients who prematurely withdraw from the study will be scheduled for an ‘end-of-study visit’ 
as soon as possible. At the end of study visit the exact date and the primary reason for 
withdrawal will be noted in the CRF.
5.6 Premature termination of the study 
If failure to enroll sufficient patients occur, premature termination of the study might be 
warranted. No special actions are needed in case of premature termination of the study. 
Premature termination of the study will not have any negative implications for the study 
participants.
6. SAFETY REPORTING
6.1 Temporary halt for reasons of subject safety 
In accordance to section 10, subsection 4, of the WMO, the sponsor will suspend the study if 
there is sufficient ground that continuation of the study will jeopardise subject health or 
safety. The sponsor will notify the accredited METC without undue delay of a temporary halt 
including the reason for such an action. The study will be suspended pending a further 
positive decision by the accredited METC. The investigator will take care that all subjects are 
kept informed. 
6.2 AEs, SAEs and SUSARs 
6.2.1
Adverse events (AEs) 
Adverse events are defined as any undesirable experience occurring to a subject during the 
study. All adverse events reported spontaneously by the subject or observed by the 
investigator or his staff will be recorded.
Participants may experience some discomfort during blood sampling.
6.2.2
Serious adverse events (SAEs) 
Since this is an observational study, serious adverse events are not expected and will 
therefore not be reported.

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6.3 Follow-up of adverse events 
All AEs will be followed until they have abated, or until a stable situation has been reached. 
Depending on the event, follow up may require additional tests or medical procedures as 
indicated, and/or referral to the general physician or a medical specialist.

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7. STATISTICAL ANALYSIS
7.1 Primary study parameter(s) 
The primary endpoint is the change in plasma desmosine levels. Plasma desmosine levels 
will be determined at baseline and 6 months after discontinuation of VKAs. Univariate 
analysis (ANCOVA) will be used for comparing plasma desmosine levels before and after 
discontinuation of VKAs. Results will be corrected for the following covariates (if statistical 
significant): age, gender, INR, indication for VKA use, duration of VKA use, BMI, smoking 
status and history, presence of COPD or other pulmonary disease, vitamin D 
supplementation, use of multivitamins and use of medication. A P<0.05 will be considered 
statistically significant. Statistics will be performed using SPSS (SPSS Inc., Chicago, Illinois, 
USA).
Both desmosine and dp-ucMGP levels will be log-transformed before performing statistical 
analysis.
7.2 Secondary study parameter(s) 
The change in plasma dephosphorylated uncarboxylated matrix Gla-protein at baseline and 
6 months after discontinuation of VKAs will be a secondary study parameter. Univariate 
analysis (ANCOVA) will be used, corrected for the following covariates (if statistical 
significant):  age, gender, INR, indication for VKA use, duration of VKA use, BMI, smoking 
status and history, presence of COPD or other pulmonary disease, vitamin D 
supplementation, use of multivitamins and use of medication. Univariate analysis will be used 
to assess the relationships between dp-ucMGP and desmosine and Pearson’s rho will be 
calculated. To assess the differences of both pDES and d-ucMGP levels between the various 
VKORC1 polymorphisms, univariate analysis will also be used, corrected for the same 
covariates as listed above. 
Both desmosine and dp-ucMGP levels will be log-transformed before performing statistical 
analysis. 
8. ETHICAL CONSIDERATIONS
8.1 Regulation statement 
This Study will be conducted according to the principles of the Declaration of Helsinki, 64th 
WMA General Assembly, Fortaleza, Brazil, October 2013, and in accordance with the 
‘Medical Research Involving Human Subjects Act’ (WMO).

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8.2 Recruitment and consent 
We will identify users of VKAs who are going to stop the use of VKAs from the anticoagulant 
clinic in the Canisius Wilhelmina Hospital. The anticoagulant clinic has an overview of
patients who will discontinue the use of VKAs. We will not use medical records of patients 
before the patient has signed informed consent. Patients will receive an information letter
and can contact the investigators for more information by email, mail or telephone. 
The patient will get sufficient time to consider participation in this study to ask questions. If 
the patient wants to participate in the study, the patient can sign informed consent (two-fold) 
and thereafter initiate study participation.
8.3 Benefits and risks assessment, group relatedness 
Participating in the study has no/marginal risks.
8.4 Compensation for injury 
We wish to obtain dispensation from the statutory obligation to provide insurance, because 
participating in the study is without risks. We will make a reasoned request to the accredited 
METC. 
 
The sponsor/investigator has a liability insurance which is in accordance with article 7 of the 
WMO. 
The insurance applies to the damage that becomes apparent during the study or within 4 
years after the end of the study.
8.5 Incentives  
Participants will receive a reimbursement for their travelling expenses only for their second 
additional visit (€ 0.19/km).
9.
ADMINISTRATIVE ASPECTS, MONITORING AND PUBLICATION

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9.1 Handling and storage of data and documents 
Patient Information will be handled with care, taking into consideration the required 
confidentiality as stated by the Dutch ‘Wet Bescherming Persoonsgegevens’ (‘Law for the 
Protection of Personal Information’). All patients will be coded by a subject number. A 
separate registry will involve the subject numbers and their corresponding data. Only the 
principal investigator is permitted access to this registry. 
Research documents, from which patient identity can be deduced will only be accessible for 
third parties (for example METC or inspection by competent authorities)
9.2 Amendments  
Amendments are changes made to the research after a favourable opinion by the accredited 
METC has been given. All amendments will be notified to the METC that gave a favourable 
opinion. 
9.3 Annual progress report 
The sponsor/investigator will submit a summary of the progress of the trial to the accredited 
METC once a year. Information will be provided on the date of inclusion of the first subject, 
numbers of subjects included and numbers of subjects that have completed the trial, serious 
adverse events/ serious adverse reactions, other problems, and amendments. 
9.4 Temporary halt and (prematurely) end of study report 
The investigator/sponsor will notify the accredited METC of the end of the study within a 
period of 8 weeks. The end of the study is defined as the last patient’s last visit. 
The sponsor will notify the METC immediately of a temporary halt of the study, including the 
reason of such an action. 
   
In case the study is ended prematurely, the sponsor will notify the accredited METC within 15 
days, including the reasons for the premature termination.
Within one year after the end of the study, the investigator/sponsor will submit a final study 
report with the results of the study, including any publications/abstracts of the study, to the 
accredited METC.

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9.5 Public disclosure and publication policy  
After analysis, data will be used for scientific presentations and for writing manuscripts which 
will be submitted to international peer-reviewed scientific journals for publication. All data will 
be reported anonymously. There are no special arrangements made between the sponsor 
and the investigator concerning the public disclosure and publication of the research data. 

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10. REFERENCES
1. Mithieux SM, Weiss AS. Elastin. Adv Protein Chem 2005:70:437-461.
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