Association of Advanced Glycation End-product Accumulation and Adverse Outcomes in Peritoneal Dialysis and Haemodialysis Patients and the Impact of a Dietetic Intervention on Skin Autofluorescence
Conditions: Chronic Kidney Disease
Sponsor: University of Nottingham

 
 
 
Page 1 of 30 
 
Skin autofluorescence as a risk marker in people receiving dialysis  
Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
 
 
 
Associations of advanced glycation end-product accumulation 
and adverse outcomes in peritoneal dialysis and haemodialysis 
patients and the impact of a dietetic intervention on skin 
autofluorescence 
 
Draft Version 4.5 Final Version 2.0 
4th October 2017 
 
 
 
Short title: Skin autofluorescence as a risk marker in people receiving 
dialysis 
 
 
Acronym:  AGED (Advanced Glycation End-products in Dialysis 
Patients) 
 
 
Trial Registration: 
NCT02878317 
 
 
IRAS Project ID: 
200680 
 
 
Trial Sponsor:  
University of Nottingham 
 
 
Sponsor reference: 
16050 
 
 
Funding Source:  
Internal funds 

 
 
 
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Skin autofluorescence as a risk marker in people receiving dialysis  
Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
TRIAL / STUDY PERSONNEL AND CONTACT DETAILS 
 
Sponsor: 
 
 
University of Nottingham 
Contact name 
 
Ms Angela Shone 
 
 
 
 
Head of Research Governance 
 
 
 
 
Research and Graduate Services 
 
 
 
 
King’s Meadow Campus 
 
 
 
 
Lenton Lane 
 
 
 
 
Nottingham 
 
 
 
 
NG7 2NR 
Email: sponsor@nottingham.ac.uk  
 
Chief investigator:   
Professor Maarten Taal 
Professor of Medicine and Honorary Consultant            
Nephrologist 
Division of Medical Sciences and Graduate Entry Medicine, 
University of Nottingham, Derby Medical School,  
     
Uttoxeter Rd, Derby, DE22 3NE 
Phone: 01332 224618 
Fax: 01332 789352 
Email:m.taal@nottingham.ac.uk; 
maarten.taal1@nhs.net 
 
Co-investigators: 
 
Dr Nick Selby  
Consultant Nephrologist and Honorary Associate             
Professor 
Royal Derby Hospital,  
 
 
 
 
Uttoxeter Rd, Derby, DE22 3NE           
 
 
 
Phone: 01332 788262 
Email: nicholas.selby@nottingham.ac.uk 
 
Dr Huda Mahmoud MBChB 
Research Fellow - studying for PhD 
Renal Research Registrar  
Royal Derby Hospital, Uttoxeter Rd, 
Derby,   DE22 3NE 
Phone: 01332 788262 
Email: huda.mahmoud@nhs.net 
 
Daniela Viramontes Horner,  
MRes Dietetics 
Student, PhD in Medicine and Health 
Royal Derby Hospital,  
 
 
 
 
Uttoxeter Rd, Derby, DE22 3NE 
Phone: 01332 788262 
Email: mzxdv@nottingham.ac.uk 
 
Trial / Study Statistician: 
Not required, all statistical analysis will be performed by the 
investigating team 
 
 
Trial / Study Coordinating Centre:  Renal Unit of the Department of Nephrology Royal Derby 
Hospital, Uttoxeter Rd, Derby, DE22 3NE 

 
 
 
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Skin autofluorescence as a risk marker in people receiving dialysis  
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This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
SYNOPSIS 
 
Title 
Associations of advanced glycation end-product accumulation and 
adverse outcomes in peritoneal dialysis and haemodialysis patients and 
the impact of a dietetic intervention on skin autofluorescence. 
Acronym 
AGED 
Short title 
Skin autofluorescence as a risk marker in people receiving dialysis. 
Chief Investigator 
Professor Maarten Taal 
Objectives 
Study 1: 
• To investigate whether the skin autofluorescence (SAF) levels increase 
over time in prevalent and incident people on peritoneal dialysis (PD) and 
haemodialysis (HD). 
• To investigate one-year survival in relation to increased SAF levels. 
• To determine the most important factors associated with changes in 
SAF over time. 
• To investigate the association of increased SAF levels with morbidity, 
changes in visual function, health-related quality of life and nutritional 
status. 
 
Study 2: 
• To investigate whether the correction of malnutrition by an intensive 
nutritional intervention results in a decrease in SAF levels in people on 
PD and HD. 
Trial Configuration 
Study 1: longitudinal/prospective. 
Study 2: observational non-randomized proof of principle study. 
Setting 
Outpatient secondary care setting. 
Sample size estimate 
Sample size Study 1: 
The primary outcome of the study is one-year survival in relation to 
increased SAF levels in people on HD and PD. With a power of 80%, a 
two-sided alpha of 5% and an expected hazard ratio of 3.5 and 2.0 in 
people on PD and HD, respectively, 100 HD and 40 PD participants will 
be needed. 
 
Sample size Study 2: 
Since this is an observational proof of principle study, it would be 
reasonable to include 40 dialysis participants (either HD or PD). 
Number of participants 
Study 1: 
• 100 participants undergoing maintenance HD treatment (dialysis 
vintage greater than 3 months). 
• 40 PD dialysis participants with a dialysis vintage greater than 3 
months. 
• 30 incident participants starting either HD or PD treatment (dialysis 
vintage <3 months). 
 
Study 2: 
40 malnourished participants on maintenance HD and PD. 
Eligibility criteria 
HD cohort: 
• Age: >18 years (no upper age limit). 
• At least three dialysis sessions per week for 3 or 4 hours. 
• Dialysis with biocompatible membranes. 

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
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• Able to give informed consent. 
 
PD cohort: 
• Age: >18 years (no upper age limit). 
• Dialysis with lactate/bicarbonate-buffered solutions with different 
glucose concentrations as prescribed for routine clinical care. 
• Able to give informed consent. 
Description of 
interventions 
Study 1: no intervention. 
 
Study 2: Dietitian supervised nutritional support aimed at improving 
malnutrition in addition to standard dietetic advice consistent with United 
Kingdom Renal Association guidelines (Wright and Jones, 2010). 
Duration of study 
Study 1:  
Participant recruitment will begin on June 2016 with an anticipated 
recruitment period of 14 months. Therefore, all baseline data and 
measurements will be collected and performed between June 2016 and 
August 2017. 
 
Participant Duration: once recruited, all participants will be followed-up for 
up to five years; consequently, it is expected that the study will be 
completed by August 2022.   
 
Study 2: 
Participant recruitment will begin in December 2017 with an anticipated 
recruitment period of 6 months. Therefore, all baseline data and 
measurements will be collected and performed between December 2017 
and June 2018. 
 
Participant Duration: once recruited, all participants will be followed-up for 
24 months; consequently, it is expected that the study will be completed 
by June 2020.   
Randomization and 
blinding 
Studies 1 and 2: no randomization required. 
Outcome measures 
Study 1: 
• Primary outcomes:  
1. Changes in SAF levels over 1 year. 
2. One-year survival. 
• Secondary outcomes: factors associated with the accumulation of AGE, 
morbidity, biochemistry, visual function, quality of life and nutritional 
status.  
 
Study 2: 
• Primary outcomes: changes in SAF levels. 
• Secondary outcomes: biochemistry, visual function, quality of life and 
nutritional status.  
Statistical methods 
Intragroup comparisons will be performed using Wilcoxon test or paired t-
test for dimensional variables and McNemar test for categorical variables. 
Intergroup comparisons will be performed using Mann Whitney test or 
Student t test for continuous variables and χ2 test or Fisher’s exact test for 
categorical variables. To determine the significance and strength of 
associations, Pearson’s correlation coefficient will be used for continuous 
variables and Spearman rank for nonparametric variables. Linear 

 
 
 
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This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
regression analysis will be used to identify determinants of advanced 
glycation end-product (AGE) accumulation. Cox proportional hazards 
models will be used to investigate the prognostic value of the 
accumulation of AGE for predicting mortality. A p value ≤0.05 will be 
considered to have statistical significance. 

 
 
 
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Skin autofluorescence as a risk marker in people receiving dialysis  
Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
ABBREVIATIONS  
 
AE 
Adverse Event 
 
AGE 
Advanced glycation end-products 
 
CI 
Chief Investigator overall 
 
 
CKD 
Chronic kidney disease 
 
CLF 
Collagen linked fluorescence 
 
CML 
Carboxymethyl lysine 
 
CRF 
Case Report Form 
 
 
EDTRS Early Treatment Diabetic Retinopathy Study 
 
ESKD End stage kidney disease 
 
GCP 
Good Clinical Practice 
 
 
HD 
Haemodialysis  
 
ICF 
Informed Consent Form 
 
IL-6 
Interleukin 6 
 
 
NHS 
National Health Service 
 
PD 
Peritoneal dialysis 
 
 
REC 
Research Ethics Committee 
 
R&D 
Research and Development department 
 
 
SAE 
Serious Adverse Event 
 
SAF 
Skin autofluorescence 
 
SF-36 36-Item Short Form Health Survey 
 
SGA 
Subjective global assessment 
 
TNF-α Tumour necrosis factor-α 

 
 
 
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Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
TABLE OF CONTENTS 
 
 
ABBREVIATIONS ............................................................................................................................... 6 
TRIAL / STUDY BACKGROUND INFORMATION AND RATIONALE .................................... 9 
TRIAL / STUDY OBJECTIVES AND PURPOSE .......................................................................... 10 
PURPOSE 
10 
PRIMARY OBJECTIVES 
10 
SECONDARY OBJECTIVES 
10 
DETAILS OF DEVICE(S) ................................................................................................................. 11 
DESCRIPTION  
11 
MANUFACTURE 
11 
PACKAGING AND LABELLING 
11 
STORAGE, DISPENSING AND RETURN 
11 
PLACEBO 
12 
KNOWN SIDE EFFECTS 
12 
REFERENCE SOURCE:  
12 
TRIAL / STUDY DESIGN ................................................................................................................. 12 
TRIAL / STUDY CONFIGURATION 
12 
Primary endpoints 
12 
Secondary endpoints 
12 
Safety endpoints 
12 
Stopping rules and discontinuation 
13 
RANDOMIZATION AND BLINDING 
13 
Maintenance of randomization codes and procedures for breaking code 
13 
TRIAL/STUDY MANAGEMENT 
13 
DURATION OF THE TRIAL / STUDY AND PARTICIPANT INVOLVEMENT 
14 
End of the Trial 
14 
SELECTION AND WITHDRAWAL OF PARTICIPANTS 
14 
Recruitment 
14 
Eligibility criteria 
14 
Inclusion criteria 
14 
Exclusion criteria 
15 
Expected duration of participant participation 
15 
Removal of participants from therapy or assessments/Participant Withdrawal 
15 
Informed consent 
15 
TRIAL / STUDY TREATMENT AND REGIMEN ......................................................................... 16 
Compliance 
20 
Criteria for terminating trial 
20 
TRANSPORT AND STORAGE OF THE TISSUES 
21 
LABORATORY ANALYSES 
21 
STATISTICS ....................................................................................................................................... 21 
Methods 
21 
Assessment of efficacy 
22 
Primary endpoints 
22 
Secondary endpoints 
22 
Assessment of safety 
22 
Procedures for missing, unused and spurious data 
22 
Definition of populations analysed 
22 

 
 
 
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This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
ADVERSE EVENTS ........................................................................................................................... 23 
ETHICAL AND REGULATORY ASPECTS .................................................................................. 23 
ETHICS COMMITTEE AND REGULATORY APPROVALS 
23 
INFORMED CONSENT AND PARTICIPANT INFORMATION 
23 
RECORDS 
24 
Case Report Forms 
24 
Sample Labelling 
24 
Source documents 
24 
Direct access to source data / documents 
25 
DATA PROTECTION 
25 
QUALITY ASSURANCE & AUDIT ................................................................................................. 25 
INSURANCE AND INDEMNITY 
25 
TRIAL CONDUCT 
25 
TRIAL DATA 
26 
RECORD RETENTION AND ARCHIVING 
26 
DISCONTINUATION OF THE TRIAL BY THE SPONSOR 
26 
STATEMENT OF CONFIDENTIALITY 
26 
PUBLICATION AND DISSEMINATION POLICY ....................................................................... 27 
USER AND PUBLIC INVOLVEMENT ........................................................................................... 27 
STUDY FINANCES ............................................................................................................................ 27 
Funding source 
27 
Participant stipends and payments 
27 
SIGNATURE PAGES ......................................................................................................................... 28 
REFERENCES .................................................................................................................................... 29 
 

 
 
 
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Skin autofluorescence as a risk marker in people receiving dialysis  
Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
TRIAL / STUDY BACKGROUND INFORMATION AND RATIONALE 
 
Chronic kidney disease (CKD) is a global public health problem and is associated with 
multiple adverse outcomes including reduced survival, especially in people requiring renal 
replacement therapy (peritoneal dialysis (PD), haemodialysis [HD] and transplantation) 
(Zhang and Rothenbacher, 2008). Multiple risk factors lead to the development and/or 
progression of CKD, such as obesity, hyperlipidaemia, glomerulonephritis, intercurrent 
infections, smoking, type 2 diabetes and hypertension, the two latter being considered as 
leading causes of CKD worldwide (Zhang and Rothenbacher, 2008; Junaid Nazar et al., 
2014). 
 
People on dialysis develop a variety of complications/abnormalities as a result of loss of 
endocrine or exocrine function of the kidneys, including anaemia, metabolic acidosis, bone 
and mineral disorders, fluid overload, hypertension, electrolyte disturbances and 
dyslipidaemia (KDIGO, 2013). In recent years, inflammation, oxidative stress and endothelial 
dysfunction (other common abnormalities in people on dialysis) have become areas of 
interest because of their strong relationship with higher rates of cardiovascular morbidity and 
mortality in people on dialysis (Oleniuc et al., 2012).  
 
Advanced glycation end-products (AGE) are uremic toxins markedly increased in people on 
dialysis. Formation of AGE starts with a non-enzymatic reaction between proteins and 
glucose molecules called the Maillard reaction; however, AGE are also formed more rapidly 
during oxidative stress with the subsequent formation of reactive carbonyl compounds like 
methyl glyoxal. At this point, AGE synthesis is irreversible and AGE will cross-link with tissue 
proteins; it seems that collagen in the skin and vascular basement membranes are especially 
susceptible to AGE accumulation and subsequent injury. AGE also interact with specific AGE 
receptors that will lead to the activation of systemic inflammation by increasing the release of 
cytokines and, consequently, exacerbate tissue damage (Arsov et al., 2014). Importantly, 
AGE are also formed in food during cooking with dry heat at high temperatures such as in 
frying, grilling or roasting and about 10% of ingested AGE are absorbed. 
 
Skin autofluorescence (SAF) is a relatively new technique that measures the skin 
accumulation of AGE. It is a non-invasive, operator independent, quick (less than 5 minutes) 
and easy to perform technique that utilizes the fluorescent properties of AGE, like the 
extensively used collagen linked fluorescence (CLF) method, and has been validated with 
specific AGE measurements and CLF in skin biopsies (Oleniuc et al., 2012). It has been 
reported that SAF is strongly correlated with overall and cardiovascular mortality in people 
with diabetes and undergoing HD (Hartog et al., 2007; Graff et al, 2014). Several factors 
have been associated with higher SAF values in people on dialysis in cross-sectional 
studies, for instance, chronological age in both dialysis modalities, glucose exposure from 
peritoneal dialysis fluid and dialysis vintage only in people on PD and presence of diabetes in 
people on HD (McIntyre et al., 2010; Smit et al., 2010). 
 
AGE accumulation is postulated as the one of the modulating factors that drives visual 
disorders (Kandarakis et al., 2014); the increased accumulation secondary to 
hyperglycaemia in diabetes is thought to cause vascular basement membrane thickening 
and destruction of pericytes from the retinal capillary bed in diabetic retinopathy (Singh et al., 
2014). The accumulation of AGE has also been reported to be a mechanism in the 
deterioration in visual acuity associated with increasing age (Ishibashi et al., 1998). 
Separately, the accumulation of AGE in other metabolic diseases (e.g. end stage kidney 
disease [ESKD]) has been found to cause nerve and eye dysfunction. Ocular abnormalities 
and therefore visual disturbances are reported in people with ESKD but the mechanism for 
this has not been fully explored or understood. In animal studies, AGE accumulation is seen 

 
 
 
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Skin autofluorescence as a risk marker in people receiving dialysis  
Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
in the lens, cornea and vitreous humour (Darlene et al., 2011). In humans, systemic AGE 
levels and visual acuity scoring have not been previously investigated. 
 
Because of the adverse outcomes strongly associated with higher levels of SAF, several 
options focused on reducing the accumulation of AGE have been proposed. One of these 
promising interventions is the reduction of dietary AGE; it has been suggested that cooking 
techniques that avoid very high temperatures such as poaching, steaming, stewing and 
boiling can significantly reduce the AGE content of food when compared to frying, broiling, 
grilling and roasting (Uribarri et al., 2010); nevertheless, most of the evidence regarding 
dietary modifications to reduce exogenous AGE is of low quality and therefore further studies 
are required (Kellow and Savige, 2013). 
 
Nevertheless, analysis of baseline data from Study 1 has identified strong associations 
between higher SAF and malnutrition whereas no correlations were observed between 
higher SAF and high dietary AGE intake. Correction of malnutrition may therefore represent 
a more important dietary intervention to reduce accumulation of AGE in people receiving 
dialysis. We further reasoned that placing people with malnutrition on a restrictive diet may 
worsen their malnutrition and we have therefore adapted our original research plan to include 
an observational study to assess the impact of correcting malnutrition on SAF by providing a 
dietitian supervised nutritional support intervention, which essentially involves the 
usual/standard dietetic care/advice provided by the NHS with some additional supervision, 
follow-up and approved dietary supplements (also provided by the NHS), rather than a 
randomised trial of dietary AGE restriction. 
 
The results from the present project will benefit people on dialysis because they may 
demonstrate that the correction of malnutrition decreases the SAF levels in this population. 
Published observational studies suggest that reduction of SAF levels will in turn be 
associated with a reduction in the high morbidity and mortality rates associated with chronic 
dialysis and, consequently, healthcare costs. Improved survival and reduced comorbidity 
would also be expected to improve the quality of life of people on dialysis.  
 
TRIAL / STUDY OBJECTIVES AND PURPOSE 
 
PURPOSE 
The purpose of the present project is to investigate the association between AGE 
accumulation and adverse outcomes (mainly mortality) and the impact of a dietary 
intervention on SAF levels in people on HD and PD. 
 
PRIMARY OBJECTIVES 
 
Study 1:  
• 
To investigate whether the SAF levels increase over time in prevalent and incident 
HD and PD participants. 
• 
To investigate one-year survival in relation to increased SAF levels. 
 
Study 2: 
• 
To investigate whether the correction of malnutrition by an intensive nutritional 
intervention results in a decrease in SAF levels in people on PD and HD. 
 
SECONDARY OBJECTIVES 
 
Study 1:  

 
 
 
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Skin autofluorescence as a risk marker in people receiving dialysis  
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This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
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• 
To determine the most important factors associated with change in SAF over time. 
• 
To investigate the association of increased SAF levels with morbidity, health-related 
quality of life and nutritional status. 
• 
To investigate the association between SAF levels and changes in visual function.  
 
Study 2: 
• 
To investigate the effect of an intensive nutritional intervention on the improvement of 
health-related quality of life and nutritional markers. 
 
DETAILS OF DEVICE(S) 
 
No products will be used in the present project. 
 
Description  
The medical device that will be used in the present project is a validated “AGE reader Standard 
Unit (SU)”. The AGE reader is a medical device used in clinical research which can non-
invasively measure tissue accumulation of AGE through a fluorescent technique called SAF.  
 
The AGE reader SU illuminates a skin surface of 1 cm2, guarded against surrounding light, 
with an excitation light source between 300 and 420 nm (peak excitation ≈370 nm). Light from 
the skin is measured with a spectrometer (AVS-USB2000, Avantes Inc., Eerbeek, The 
Netherlands) in the 300-600 nm range, using a 200-µm glass fibre. SAF will be measured as 
the ratio between emission and excitation, calculated in arbitrary units (AU), by dividing the 
intensity of the fluorescent light coming from the skin (measured as area under the curve [AUC] 
of fluorescent wave lengths between 420 and 600 nm) by the intensity of the emitted light 
(measured as AUC of wave lengths between 300 and 420 nm) multiplied by 100. For each 
participant, three measurements will be performed on the volar surface of the lower arm at ≈10 
cm below the elbow, ensuring that the area has normal skin without visible vessels, scars or 
other abnormalities. For HD participants, only the non-fistula arm will be used and evaluations 
will take place prior to dialysis sessions (Siriopol et al., 2015). This technique is non-invasive 
and there is no risk of harm to the participant. Measurements take only a few minutes. 
 
The AGE reader SU will be used by Daniela Viramontes Horner (PhD student) following user 
instructions provided by the manufacturer. 
 
Manufacture 
The name of the manufacturer is DiagnOptics Technologies BV (Aarhusweg 4-9, Groningen, 
The Netherlands) 
 
The AGE reader SU is a distributed CE marked product covered by the "CE Marking of 
Conformity Certificate", reference number: 2085393CE01, delivered by DEKRA Certification 
B.V., Arnhem, The Netherlands, Notified Body Identification Number 0344, and conforms to 
the required technical documentation, in accordance with Annex VII of the "EC-Directive", the 
Council Directive 93/42/EEC of 14 June 1993, concerning medical devices. 
 
The AGE reader SU will be used in the present project in accordance with this CE marking. 
 
Packaging and labelling 
 
Storage, dispensing and return 
The AGE reader SU will be stored securely, in a locked room, or locked cupboard or cabinet. 
Access to the medical device will be limited to the study staff and investigators. 
  

 
 
 
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Skin autofluorescence as a risk marker in people receiving dialysis  
Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
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from the University of Nottingham 
Placebo 
No placebo products will be used in the present project. 
 
Known Side Effects 
The AGE-Reader has been extensively tested in thousands of adults for several years without 
any adverse effects. The AGE Reader has CE approval (2085393CE01) to be used as a 
medical device and thus has passed the rigid safety standard. The amount of ultraviolet light 
exposure is actually far below the safety limits. 
 
Reference source:  
AGE reader. Cardiovascular risk assessment. http://www.diagnoptics.com/age-reader/ 
[Accessed on 12th May, 2016] 
 
TRIAL / STUDY DESIGN 
 
TRIAL / STUDY CONFIGURATION 
Studies 1 and 2 will be single-centre. 
 
Study 1: longitudinal/prospective. 
Study 2: observational non-randomized proof of principle study where malnourished dialysis 
participants will receive a dietitian supervised intensive nutritional support. 
 
Primary endpoints 
Study 1:  
1. Changes in SAF levels after 12 months of follow-up. 
2. One-year survival. 
 
Study 2: changes in SAF levels after 6 months of a dietetic intervention (i.e. intensive 
nutritional support). 
 
Secondary endpoints 
Study 1:  
• 
Factors associated with the accumulation of AGE. 
• 
Association of SAF levels with morbidity, biochemical variables (haemoglobin, 
haemoglobin A1C (HbA1C), urea, creatinine, potassium, phosphate, calcium, sodium, 
albumin, total proteins, cholesterol, triglycerides, and intact parathyroid hormone), 
visual function, circulating AGE levels (carboxymethyl lysine [CML] and pentosidine), 
inflammatory biomarkers (tumour necrosis factor-α [TNF-α], interleukin 6 [IL-6], C 
reactive protein [CRP] and others) health-related quality of life and nutritional status 
(energy, protein and fat intake, dietary AGE intake, anthropometry and subjective 
global assessment [SGA]). 
 
Study 2:  
• 
Association of SAF levels with morbidity, biochemical variables (haemoglobin, HbA1C, 
urea, creatinine, potassium, phosphate, calcium, sodium, albumin, total proteins, 
cholesterol, triglycerides, and intact parathyroid hormone), visual function, circulating 
AGE levels (CML and pentosidine), inflammatory biomarkers (TNF-α, IL-6, CRP and 
others) health-related quality of life and nutritional status (energy,  protein and fat 
intake, dietary AGE intake, anthropometry and SGA). 
 
Safety endpoints 
 
• 
There will not be any safety endpoints for these studies. 

 
 
 
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from the University of Nottingham 
 
Stopping rules and discontinuation 
 
Participant Withdrawal  
Participants will be withdrawn from the study if they withdraw their consent at any time. 
Participants may be withdrawn from the study at the discretion of the Investigator. 
Participants will be made aware that withdrawal will not affect their future care. Participants 
who withdraw will be asked for permission to retain data already collected for use in the final 
analysis. 
 
If participants fail to attend for their study visits, a member of the research team will try to 
contact them by telephone within 24 hours of their appointment and by written 
correspondence within a week of the missed appointment.  Multiple attempts at contact will 
be made by the research team over a period of a month before stating that a participant has 
been lost to follow-up.  
 
Participants will be withdrawn from the study if they: 
• 
Withdraw their consent at any point. 
• 
Become pregnant. 
• 
Decline to comply with the dietetic intervention (Study 2 only). 
 
Criteria for terminating the study 
Study 1 is observational and will therefore only be terminated if it becomes clear that for 
unforeseen reasons the study is not feasible. 
 
The intervention in Study 2 is not expected to be associated with any adverse effects. Study 
2 will therefore only be terminated if it becomes clear that for unforeseen reasons the study is 
not feasible. 
 
RANDOMIZATION AND BLINDING 
 
Maintenance of randomization codes and procedures for breaking code 
 
TRIAL/STUDY MANAGEMENT 
 
The Chief Investigator (CI; Professor Maarten Taal) has overall responsibility for the study 
and shall oversee all study management. The data custodian will be the CI. 
 
The study will be coordinated from the University of Nottingham campus at the Royal Derby 
Hospital. Data will be collected at the Renal Unit of the Department of Nephrology at the 
Royal Derby Hospital (Derby Teaching Hospitals NHS Foundation Trust) and stored at the 
University of Nottingham. 
 
Daniela Viramontes Horner (PhD student), Dr. Nick Selby and Professor Maarten Taal are 
responsible for study planning, recruitment, consenting, data collection, handling and 
interpretation. 
 
 
 
 
 
 

 
 
 
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Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
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from the University of Nottingham 
DURATION OF THE TRIAL / STUDY AND PARTICIPANT INVOLVEMENT 
 
Study Duration: 
 
Study 1:  
Participant recruitment will begin on June 2016 with an anticipated recruitment period of 14 
months. Therefore, all baseline data and measurements will be collected and performed 
between June 2016 and August 2017. 
 
Participant Duration: once recruited, all participants will be followed-up for up to five years; 
consequently, it is expected that the study will be completed by August 2022.   
 
Study 2: 
Participant recruitment will begin in December 2017 with an anticipated recruitment period of 
6 months. Therefore, all baseline data and measurements will be collected and performed 
between December 2017 and June 2018. 
 
Participant Duration: once recruited, all participants will be followed-up for 24 months; 
consequently, it is expected that the study will be completed by June 2020.   
 
End of the Trial 
 
The end of studies 1 and 2 will be when long term follow-up has been completed. 
 
SELECTION AND WITHDRAWAL OF PARTICIPANTS 
 
Recruitment 
 
Potential participants on HD and PD will be recruited from the Renal Unit of the Department 
of Nephrology at the Royal Derby Hospital. The initial details of the study and participant 
information sheet will be provided by the usual care team (which may include the 
researcher). Participants will then be given at least 24 hours to consider whether they wish to 
participate, as well as ask any questions about the study, before being re-contacted by the 
investigators. 
  
If needed, the usual hospital interpreter and translator services will be available to assist with 
discussion of the study, the participant information sheets, and consent forms, but the 
consent forms and information sheets will not be available in other languages. 
 
It will be explained to the potential participant that entry into the study is entirely voluntary 
and that their treatment and care will not be affected by their decision. It will also be 
explained that they can withdraw at any time. In the event of their withdrawal it will be 
explained that their data collected so far cannot be erased and we will seek consent to use 
the data in the final analyses where appropriate. 
 
The participant’s general practitioner will not be informed about the study because there are 
no implications for other aspects of treatment. 
 
Eligibility criteria 
 
Inclusion criteria 
HD cohort: 
• Age: >18 years (no upper age limit). 
• At least three dialysis sessions per week for 3 or 4 hours. 

 
 
 
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from the University of Nottingham 
• Dialysis with biocompatible membranes. 
• Able to give informed consent. 
 
PD cohort: 
• Age: >18 years (no upper age limit). 
• Dialysis with lactate/bicarbonate-buffered solutions with different glucose concentrations as 
prescribed for routine clinical care. 
• Able to give informed consent. 
 
Exclusion criteria 
• Does not wish to participate. 
• Renal transplant.  
• Pregnancy or breast feeding or intending pregnancy. 
• Expected survival less than one year (for Study 1). 
 
Expected duration of participant participation 
Participants will be participating in the Study 1 for five years and in the Study 2 for 24 
months.  
 
Removal of participants from therapy or assessments/Participant Withdrawal 
Participants will be withdrawn from the study if they withdraw their consent at any point, 
receive a renal transplant or decline to comply with the dietetic intervention (Study 2). 
Female participants will be withdrawn from the study if they become pregnant.   
 
Participants may be withdrawn from the study either at their own request or at the discretion 
of the Investigator. The participants will be made aware that this will not affect their future 
care. Participants will be made aware (via the information sheet and consent form) that 
should they withdraw the data collected to date cannot be erased and, with their consent, 
may still be used in the final analysis. 
 
If participants fail to attend for their study visits, a member of the research team will try to 
contact them by telephone within 24 hours of their appointment and by written correspondence 
within a week of the missed appointment.  Multiple attempts at contact will be made by the 
research team over a period of a month before stating that a participant has been lost to follow-
up.  
 
With participant consent, data that is collected prior to withdrawal will be kept and analysed as 
part of the data set. The date, time and reason for participant withdrawal will be documented 
on the case report form (CRF). 
 
To ensure attainment of adequate numbers of participants for the analysis to be valid, the 
research team will attempt to recruit other participants to replace any who withdraw from the 
study. 
 
Informed consent 
The process for obtaining participant informed consent will be in accordance with the 
Research Ethics Committee (REC) guidance, Good Clinical Practice (GCP) and any other 
regulatory requirements that might be introduced. 
 
The research team have both training and experience in assessing participant capacity and 
gaining informed consent in the clinical context. 
 

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
Detailed information will be provided on the study to ensure that participants understand the 
purpose and potential risks involved. The study will not involve participants who are unable to 
represent their own interests or are particularly susceptible to coercion. 
 
All participants will provide written informed consent. The Informed Consent Form will be 
signed and dated by the participant before they enter the study. The Investigator will explain 
the details of the study and provide a Participant Information Sheet, ensuring that the 
participant has sufficient time to consider participating or not. The Investigator will answer 
any questions that the participant has concerning study participation.  
 
The decision regarding participation in the study is entirely voluntary. The Investigator or their 
nominee shall emphasize to them that consent regarding study participation may be 
withdrawn at any time without penalty or affecting the quality or quantity of their future 
medical care. 
 
Informed consent will be obtained from each participant before they undergo any 
interventions (including physical examination and history taking) related to the study. One 
copy of this will be kept by the participant, one will be kept by the Investigator, and a third will 
be retained in the participant’s hospital records. 
 
Should there be any subsequent substantial amendment to the final protocol, which might 
affect a participant’s participation in the study, continuing consent will be obtained using an 
amended Consent Form which will be signed by the participant. 
 
TRIAL / STUDY TREATMENT AND REGIMEN 
 
Study 1 - longitudinal/prospective study 
 
100 participants undergoing maintenance HD treatment (dialysis vintage greater than 3 
months), 40 PD participants with a dialysis vintage greater than 3 months and up to 30 
incident participants starting either HD or PD treatment (dialysis vintage less than 3 months) 
will be recruited from the Renal Unit of the Department of Nephrology at the Royal Derby 
Hospital and will be followed-up for five years.  
 
Data collection and study procedures/assessments 
 
Once participants have given their informed consent, relevant participant characteristics will 
be extracted from electronic data captured in medical records and from a self-report 
questionnaire at baseline and will include demographic characteristics, educational level, 
occupation status, family history, duration of renal replacement therapy, dialysis adequacy, 
aetiology of CKD, medical history (e.g. history of cardiovascular disease, diabetes and 
hypertension), current co-morbidities, alcohol history, smoking status and current medication. 
For the PD participants, information regarding the type of PD and the volume of the different 
PD solutions will also be recorded. 
 
The accumulation of AGE will be assessed by measuring SAF using a validated 
Autofluorescence Reader (AGE Reader, DiagnOptics, Groningen, The Netherlands) at 
baseline, 3, 6, 9 and 12 months of follow-up. The method has been previously described by 
Meerwaldt et al. (2005). As described in the study by Siriopol et al. (2015), “the 
autofluorescence reader illuminates a skin surface of 1 cm2, guarded against surrounding 
light, with an excitation light source between 300 and 420 nm (peak excitation ≈370 nm). 
Light from the skin is measured with a spectrometer (AVS-USB2000, Avantes Inc., Eerbeek, 
The Netherlands) in the 300-600 nm range, using a 200-µm glass fibre”. SAF will be 
measured as the ratio between emission and excitation, calculated in arbitrary units (AU), by 

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
dividing the intensity of the fluorescent light coming from the skin (measured as area under 
the curve [AUC] of fluorescent wave lengths between 420 and 600 nm) by the intensity of the 
emitted light (measured as AUC of wave lengths between 300 and 420 nm) multiplied by 
100. For each participant, three measurements will be performed on the volar surface of the 
lower arm at ≈10 cm below the elbow, ensuring that the area has normal skin without visible 
vessels, scars or other abnormalities. For HD participants, only the non-fistula arm will be 
used and readings will be undertaken within the first hour of HD treatment (Siriopol et al., 
2015).  
 
Routine blood/laboratory tests will be performed at baseline and monthly and will include 
haemoglobin, HbA1C, urea, creatinine, potassium, phosphate, calcium, sodium, albumin, 
total proteins, cholesterol, triglycerides and intact parathyroid hormone. Pre- and post-
dialysis blood samples (10 mL) will be taken at baseline, 3, 6, 9 and 12 months of follow-up 
and centrifuged to obtain at least 2 mL of serum and 2 mL of plasma which will be frozen and 
stored at -80°C for subsequent analysis of circulating AGE levels (CML and pentosidine) and 
markers of inflammation (TNF-α, IL-6, CRP and others). 
 
Nutritional assessments will be performed at baseline and at 6 and 12 months of follow-up 
and will include: three 24-h dietary recalls to assess energy, fat and protein intake, a 
validated food frequency questionnaire to evaluate dietary AGE intake; the 7-point scale 
SGA for the classification of the nutritional status (normal, mild-moderate malnutrition and 
severe malnutrition) and anthropometric measurements and indexes (weight, height, mid-
arm circumference, triceps skinfold thickness, body mass index, hand-grip strength and mid-
arm muscle area). 
 
Participants will have all aspects of their visual acuity assessed at baseline and 12 months. 
Those on HD will be tested before and after a single mid-week dialysis session. Participants 
will initially be asked a screening questionnaire to assess whether or not they note a change 
in their visual acuity or experience visual disturbances peri dialysis. The assessments will 
include near, distance, contrast and colour vision assessed using the Jaeger near vision 
chart, standardized Early Treatment Diabetic Retinopathy Study (EDTRS) testing, Pelli-
Robson Contrast Sensitivity Chart, and Ishihara Color Vision Test, respectively. Participants 
will be asked to bring their glasses or wear their contact lenses on the study day to assess 
best corrected visual acuity.  
 
Health-related quality of life will be evaluated at baseline and at 6 and 12 months of follow-up 
with the 36-Item Short Form Health Survey (SF-36) and EQ5D questionnaire. 
 
After the first year of the study, SAF readings and questionnaires will be repeated once a 
year for up to five years. 
 
Study 2 - Observational non-randomised proof of principle study 
 
Starting December 2017, 40 malnourished (i.e. SGA scores from 1 to 5) participants on 
maintenance HD and PD will be recruited from the Renal Unit of the Department of 
Nephrology at the Royal Derby Hospital and then will be followed-up for 24 months. 
Therefore, Study 2 will be running alongside Study 1. Depending on the results of the 
longitudinal/prospective study, we will decide to include incident HD and PD participants as 
well. 
 
Description of intervention 
 
Once the identified malnourished participants have given their informed consent, they will 
receive intensive dietitian supervised nutritional support with the aim of improving their 

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
malnutrition. In addition, participants will receive standard dietary advice for people on 
dialysis based on the Nutritional Guidelines in CKD published by the Renal Association in 
March 2010 in the UK (Wright and Jones, 2010) and will include the following: energy (35 
kcal/kg/day) and protein intake (1.2 g/kg/day), as well as potassium, phosphate and sodium 
restriction, according with biochemical blood parameters. 
 
Adherence to the intervention will be evaluated by means of three 24-h dietary recalls every 
month (1 dialysis day, 1 non-dialysis day and 1 weekend day for the HD participants and 2 
weekdays and 1 weekend day for the PD participants). 
 
Data collection and study procedures/assessments 
 
Relevant participant characteristics will be extracted from electronic data captured in medical 
records and from a self-report questionnaire at baseline and will include demographic 
characteristics, educational level, occupation status, family history, duration of renal 
replacement therapy, dialysis adequacy, aetiology of CKD, medical history (e.g. history of 
cardiovascular disease, diabetes and hypertension), current co-morbidities, alcohol history, 
smoking status and current medication. For the PD participants, information regarding the 
type of PD and the volume of the different PD solutions will also be recorded. 
 
The accumulation of AGE will be assessed by measuring SAF using a validated 
Autofluorescence Reader (AGE Reader, DiagnOptics, Groningen, The Netherlands) at 
baseline, 3 and 6 months of follow-up according to the method described above. 
 
Routine blood/laboratory tests will be performed at baseline and monthly and will include 
haemoglobin, HbA1C, urea, creatinine, potassium, phosphate, calcium, sodium, albumin, 
total proteins, cholesterol, triglycerides and intact parathyroid hormone. Pre- and post-
dialysis blood samples (10 mL) will be taken at baseline, 3 and 6 months of follow-up and 
centrifuged to obtain at least 2 mL of serum and 2 mL of plasma which will be frozen and 
stored at -80°C for subsequent analysis of circulating AGE levels (CML and pentosidine) and 
markers of inflammation (TNF-α, IL-6, CRP and others). 
 
Nutritional assessments will be performed at baseline and at 6th month of treatment and will 
include: three 24-h dietary recalls to assess energy, fat and protein intake, a validated food 
frequency questionnaire to evaluate dietary AGE intake, the 7-point scale SGA for the 
classification of the nutritional status (normal, mild-moderate malnutrition and severe 
malnutrition) and anthropometric measurements and indexes (weight, height, mid-arm 
circumference, triceps skinfold thickness, body mass index, hand-grip strength and mid-arm 
muscle area).  
 
Participants will have all aspects of their visual acuity assessed at baseline and 6 months. 
Those on HD will be tested before and after a single mid-week dialysis session. Participants 
will initially be asked a screening questionnaire to assess whether or not they note a change 
in their visual acuity or experience visual disturbances peri dialysis. The assessments will 
include near, distance, contrast and colour vision assessed using the Jaeger near vision 
chart, standardized EDTRS testing, Pelli-Robson Contrast Sensitivity Chart, and Ishihara 
Color Vision Test respectively. Participants will be asked to bring their glasses or wear their 
contact lenses on the study day to assess best corrected visual acuity. 
 
Health-related quality of life will be evaluated at baseline and at 6th month of follow-up with 
the SF-36 and EQ5D questionnaire. 
 
After the first 6 months of the study, SAF readings and questionnaires will be repeated once 
every 6 months for up to 2 years. 

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
Schematic diagram of study procedures and stages – Study 1 (timeline showing only 
the first year of study) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Prevalent and incident HD and 
PD participants potentially 
identified 
Participant Information Sheet 
given + Informed consent signed 
NO 
YES 
End of possible participation in 
the study 
Start of the study 
0     
1  
 2     
 3     
 4      
5           6             7            8 
       9           10            11        12      
Months  
     Participant characteristics 
 
     Skin autofluorescence and pre- and post-dialysis bloods 
 
     Biochemistry 
 
     Nutritional and quality of life assessments 
 
     Assessment of visual function 

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
Schematic diagram of study procedures and stages – Study 2 (timeline showing only 
the first 6 months of the study) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Compliance 
 
The Study 1 (prospective/longitudinal) involves SAF measurements and visual function, 
nutritional and quality of life assessments during one year of follow-up. Therefore, there is no 
need for an assessment of participant compliance with the study protocol. Should the 
participant not complete the study day, this will be classified as a withdrawal rather than non-
compliance (see Section Participant Withdrawal). 
 
Compliance with the intervention of Study 2 will be evaluated by means of three 24-h dietary 
recalls every month (1 dialysis day, 1 non-dialysis day and 1 weekend day for the HD 
participants and 2 weekdays and 1 weekend day for the PD participants). 
 
Criteria for terminating trial 
 
Study 1 is observational and will therefore only be terminated if it becomes clear that for 
unforeseen reasons the study is not feasible. 
 
Malnourished HD and PD 
participants potentially identified 
Participant Information Sheet 
given + Informed consent signed 
NO 
YES 
End of possible participation in 
the study 
Start of the study 
     Participant characteristics 
 
     Skin autofluorescence and pre- and post- dialysis bloods 
 
     Biochemistry 
 
     Nutritional and quality of life assessments 
 
     Assessment of visual function 
0     
1  
 2     
 3     
 4      
5           6              
                                               
Months  

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
The intervention in Study 2 is not expected to be associated with any adverse effects. Study 
2 will therefore only be terminated if it becomes clear that for unforeseen reasons the study is 
not feasible. 
 
TRANSPORT AND STORAGE OF THE TISSUES 
 
Blood samples (10 mL) will be collected at intervals as described and centrifuged. At least    
2 mL of serum and 2 mL of plasma will be stored at -80°C for future analysis.  
 
Samples and data will be stored in a linked anonymised format. Samples will be labelled 
using a combination of study reference, unique study identifier and cross referenced with 
location code numbers to permit accurate linkage to study data and the consent form.  
 
The master database will be held by the CI in a password encrypted file.  
 
Where participants do not agree to the future use of the samples, they will be destroyed in 
accordance with the Human Tissue Act, 2004. 
 
LABORATORY ANALYSES 
 
Routine blood/laboratory tests will be analysed in a local NHS laboratory at the Royal Derby 
Hospital, which is serviced and managed according to national standards and governance 
frameworks. No DNA or RNA extraction will occur as part of this study. Assays for CML and 
inflammatory markers will be conducted in research laboratories at the University of 
Nottingham. 
 
STATISTICS 
 
Methods  
 
Statistical analysis will be conducted by Daniela Viramontes Horner (PhD student), Professor 
Maarten Taal and Dr. Nick Selby. Data management and statistical analyses will be 
performed using statistical software SPSS version 22.0.  
 
Data will be presented as mean ± standard deviation, median (percentiles 25%-75%), or 
percentages, as appropriate. 
 
To compare baseline versus final evaluations, Wilcoxon test or paired t-test will be used in 
the case of dimensional variables, and McNemar test in the case of categorical variables. 
Intergroups comparisons will be performed using Mann Whitney test or Student t test for 
continuous variables and χ2 test or Fisher’s exact test for categorical variables. To determine 
the significance and strength of associations, Pearson’s correlation coefficient will be used 
for analyses of associations between continuous variables and Spearman rank for 
nonparametric variables. Linear regression analysis will be used to identify determinants of 
AGE accumulation. Cox proportional hazards models will be used to investigate the 
prognostic value of the accumulation of AGE for predicting mortality. A p-value less than or 
equal to 0.05 will be considered to have statistical significance. 
 
Sample size calculation of Study 1 was performed by using the software nQuery Advisor 
v.6.0. 
 
 
 
 

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
Sample size Study 1: 
 
The primary outcome for sample size determination is one-year survival in relation to 
increased SAF levels in people on HD and PD. With a power of 80%, a two-sided alpha of 
5% and an expected hazard ratio of 3.5 and 2.0 in people on PD and HD, respectively 
(based on similar findings of the studies of Kimura et al. [2014] and Siriopol et al. [2015]), 
100 HD and 40 PD participants will be needed. 
 
Sample size Study 2 
Since this is a proof of principle study, it would be reasonable to include 40 dialysis 
participants (either HD or PD). 
 
Assessment of efficacy 
 
Primary endpoints 
 
Study 1:  
• Mean change in SAF levels (AU) from baseline to 12 months of follow-up. 
• One-year survival. 
  
Study 2:  
Mean change in SAF levels (AU) after 6 months of a dietitian supervised intensive nutritional 
support intervention 
 
Secondary endpoints 
 
Study 1:  
• Regression coefficients for identifying factors associated with the accumulation of AGE. 
• Correlation coefficients for analysing the association of SAF levels with morbidity, 
biochemistry, circulating AGE levels, inflammatory biomarkers, visual function, health-related 
quality of life and nutritional status. 
 
Study 2:  
• Changes in biochemistry, circulating AGE levels, inflammatory biomarkers, visual function, 
health-related quality of life and nutritional status. 
 
Assessment of safety 
 
Tolerability and safety of the nutritional support intervention during the proof of principle 
study (Study 2) will be assessed by direct interview. If any serious adverse event (SAE) is 
reported during the study, it will be recorded on an appropriate SAE report form. 
 
Procedures for missing, unused and spurious data 
 
All SAF measurements, biochemistry, nutritional and quality of life assessments for Study 1 
and 2 will be used in the statistical analysis, including data from participants who did not 
complete the entire study protocol. 
 
Definition of populations analysed 
 
Study 1: 
Full Analysis set: All participants included in the protocol for whom at least one post-baseline 
assessment of the primary endpoint is available.  
 

 
 
 
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Per protocol set: All participants in the Full Analysis set who are deemed to have no major 
protocol violations that could interfere with the objectives of the study. 
 
Study 2: 
Full Analysis set: All randomised participants, who participated in at least one treatment and 
for whom at least one post-baseline assessment of the primary endpoint is available.  
 
Per protocol set: All participants in the Full Analysis set who are deemed to have no major 
protocol violations that could interfere with the objectives of the study. 
 
ADVERSE EVENTS 
 
The occurrence of an adverse event as a result of participation within this study is not 
expected and as such no adverse event data will be collected.  
 
ETHICAL AND REGULATORY ASPECTS 
 
ETHICS COMMITTEE AND REGULATORY APPROVALS 
The study will not be initiated before the protocol, informed consent forms and participant 
information sheets have received approval / favourable opinion from the REC, and the 
respective NHS Research & Development (R&D) department. Should a protocol amendment 
be made that requires REC approval, the changes in the protocol will not be instituted until 
the amendment and revised informed consent forms and participant information sheets (if 
appropriate) have been reviewed and received approval / favourable opinion from the REC 
and R&D departments. A protocol amendment intended to eliminate an apparent immediate 
hazard to participants may be implemented immediately providing that the REC are notified 
as soon as possible and an approval is requested. Minor protocol amendments only for 
logistical or administrative changes may be implemented immediately; and the REC will be 
informed. 
 
The study will be conducted in accordance with the ethical principles that have their origin in 
the Declaration of Helsinki, 1996; the principles of GCP, and the Department of Health 
Research Governance Framework for Health and Social care, 2005. 
 
INFORMED CONSENT AND PARTICIPANT INFORMATION 
 
The process for obtaining participant informed consent will be in accordance with the REC 
guidance, GCP and any other regulatory requirements that might be introduced. The 
investigator or their nominee and the participant shall both sign and date the Informed 
Consent Form before the person can participate in the study. 
 
The participant will receive a copy of the signed and dated forms and the original will be 
retained in the Trial Master File. A second copy will be filed in the participant’s medical notes 
and a signed and dated note made in the notes that informed consent was obtained for the 
study.  
 
The decision regarding participation in the study is entirely voluntary. The investigator or their 
nominee shall emphasize to them that consent regarding study participation may be 
withdrawn at any time without penalty or affecting the quality or quantity of their future 
medical care, or loss of benefits to which the participant is otherwise entitled. No study-
specific interventions will be done before informed consent has been obtained. 
 

 
 
 
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transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
The investigator will inform the participant of any relevant information that becomes available 
during the course of the study, and will discuss with them, whether they wish to continue with 
the study. If applicable they will be asked to sign revised consent forms. 
 
If the Informed Consent Form is amended during the study, the investigator shall follow all 
applicable regulatory requirements pertaining to approval of the amended Informed Consent 
Form by the REC and use of the amended form (including for ongoing participants). 
 
RECORDS  
 
Case Report Forms  
 
Case Report Forms (CRFs) will be the primary data collection instrument. All data requested 
on the CRF will be recorded. Data collected includes inclusion and exclusion criteria, 
demographic characteristics, educational level, occupation status, family history, duration of 
renal replacement therapy, dialysis adequacy, aetiology of CKD, medical history (e.g. history 
of cardiovascular disease, diabetes and hypertension), current co-morbidities, alcohol 
history, smoking status, current medication, type of PD, volume of PD solutions SAF 
measurements, routine blood tests, circulating AGE levels, inflammatory biomarkers, visual 
function, nutritional and quality of life assessments. 
 
Each participant will be assigned a study identity code number for use on CRFs, other study 
documents and the electronic database. The documents and database will also use their 
initials (of first and last names separated by a hyphen or a middle name initial when 
available) and final two digits from their year of birth. 
 
CRFs will be treated as confidential documents and held securely in accordance with 
regulations. The investigator will make a separate confidential record of the participant’s 
name, date of birth, local hospital number or NHS number, and Participant Study Number 
(the Trial Recruitment Log), to permit identification of all participants enrolled in the study, in 
accordance with regulatory requirements and for follow-up as required. 
 
CRFs shall be restricted to those personnel approved by the Chief or local Principal 
Investigator and recorded on the ‘Trial Delegation Log.’ 
 
All paper forms shall be filled in using black ballpoint pen. Errors shall be lined out but not 
obliterated by using correction fluid and the correction inserted, initialled and dated. 
 
The Chief or local Principal Investigator shall sign a declaration ensuring accuracy of data 
recorded in the CRF. 
 
Sample Labelling  
 
Each participant will be assigned a study identity code number for use on the samples, 
consent forms and other study documents and the electronic database. The identifier will 
include subject number, participant initials (of first and last names separated by a hyphen or 
a middle name initial when available) and final two digits from their year of birth. 
 
Source documents  
 
Source documents shall be filed at the investigator’s site and may include but are not limited 
to, consent forms, current medical records, laboratory results and records. A CRF may also 
completely serve as its own source data. Only study staff as listed on the Delegation Log 

 
 
 
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from the University of Nottingham 
shall have access to study documentation other than the regulatory requirements listed 
below. 
 
Direct access to source data / documents 
 
The CRF and all source documents, including progress notes and copies of laboratory and 
medical test results shall made be available at all times for review by the CI, Sponsor’s 
designee and inspection by relevant regulatory authorities (e.g. DH, Human Tissue 
Authority). 
 
DATA PROTECTION  
 
All study staff and investigators will endeavour to protect the rights of the study’s participants 
to privacy and informed consent, and will adhere to the Data Protection Act, 1998. The CRF 
will only collect the minimum required information for the purposes of the study. CRFs will be 
held securely, in a locked room, or locked cupboard or cabinet. Access to the information will 
be limited to the study staff and investigators and relevant regulatory authorities (see above). 
Computer held data including the study database will be held securely and password 
protected. All data will be stored on a secure dedicated web server. Access will be restricted 
by user identifiers and passwords (encrypted using a one way encryption method). 
 
Information about the study in the participant’s medical records/hospital notes will be treated 
confidentially in the same way as all other confidential medical information. 
 
Electronic data will be backed up every 24 hours to both local and remote media in encrypted 
format. 
 
QUALITY ASSURANCE & AUDIT  
 
INSURANCE AND INDEMNITY 
 
Insurance and indemnity for study participants and study staff is covered within the NHS 
Indemnity Arrangements for clinical negligence claims in the NHS, issued under cover of 
HSG (96)48. There are no special compensation arrangements, but study participants may 
have recourse through the NHS complaints procedures. 
 
The University of Nottingham as research Sponsor indemnifies its staff, research participants 
and research protocols with both public liability insurance and clinical trials insurance. These 
policies include provision for indemnity in the event of a successful litigious claim for proven 
non-negligent harm.  
 
TRIAL CONDUCT 
 
Study conduct may be subject to systems audit of the Trial Master File for inclusion of 
essential documents; permissions to conduct the study; Trial Delegation Log; CVs of study 
staff and training received; local document control procedures; consent procedures and 
recruitment logs; adherence to procedures defined in the protocol (e.g. inclusion / exclusion 
criteria, correct randomisation, timeliness of visits); AE recording and reporting; 
accountability of study materials and equipment calibration logs. 
 
The Study Coordinator/Academic Supervisor, or where required, a nominated designee of 
the Sponsor, shall carry out a site systems audit at least yearly and an audit report shall be 
made to the Trial Steering Committee. 
 

 
 
 
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TRIAL DATA  
 
Monitoring of study data shall include confirmation of informed consent; source data 
verification; data storage and data transfer procedures; local quality control checks and 
procedures, back-up and disaster recovery of any local databases and validation of data 
manipulation. The Study Coordinator/Academic Supervisor, or where required, a nominated 
designee of the Sponsor, shall carry out monitoring of study data as an ongoing activity.  
 
Entries on CRFs will be verified by inspection against the source data. A sample of CRFs 
(10% or as per the study risk assessment) will be checked on a regular basis for verification 
of all entries made. In addition, the subsequent capture of the data on the study database will 
be checked. Where corrections are required these will carry a full audit trail and justification. 
 
Study data and evidence of monitoring and systems audits will be made available for 
inspection by REC as required. 
 
RECORD RETENTION AND ARCHIVING 
 
In compliance with the ICH/GCP guidelines, regulations and in accordance with the 
University of Nottingham Research Code of Conduct and Research Ethics, the Chief or local 
Principal Investigator will maintain all records and documents regarding the conduct of the 
study. These will be retained for at least 7 years or for longer if required. If the responsible 
investigator is no longer able to maintain the study records, a second person will be 
nominated to take over this responsibility.  
 
The Trial Master File and study documents held by the CI on behalf of the Sponsor shall be 
finally archived at secure archive facilities at the University of Nottingham. This archive shall 
include all study databases and associated meta-data encryption codes. 
 
DISCONTINUATION OF THE TRIAL BY THE SPONSOR  
 
The Sponsor reserves the right to discontinue this study at any time for failure to meet 
expected enrolment goals, for safety or any other administrative reasons.  The Sponsor shall 
take advice from the Trial Steering Committee and Data Monitoring Committee as 
appropriate in making this decision. 
 
STATEMENT OF CONFIDENTIALITY  
 
Individual participant medical information obtained as a result of this study is considered 
confidential and disclosure to third parties is prohibited with the exceptions noted above. 
Participant confidentiality will be further ensured by utilising identification code numbers to 
correspond to treatment data in the computer files. Such medical information may be given to 
the participant’s medical team and all appropriate medical personnel responsible for the 
participant’s welfare.  
 
If information is disclosed during the study that could pose a risk of harm to the participant or 
others, the researcher will discuss this with the CI and where appropriate report accordingly. 
 
Data generated as a result of this study will be available for inspection on request by the 
participating physicians, the University of Nottingham representatives, the REC, local R&D 
Departments and the regulatory authorities. 
 
 

 
 
 
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This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
PUBLICATION AND DISSEMINATION POLICY 
 
The results of this study will be submitted to peer-reviewed journals for publication as soon 
as data analysis is completed. The results will also be presented at national and international 
scientific conferences. Participants will not be identified in any publications. Participants will 
be contacted either by email or a letter sent to their addresses about the findings of the 
research (unless participants advise the research team that they do not wish to be 
contacted). In addition, participants will be informed of the results of the study via a 
departmental research newsletter, which is made available to all people on dialysis. The 
results of this study will form part of PhD thesis for Daniela Viramontes Horner. The study 
has been registered on www.clinicaltrials.gov (Trial registration number: NCT02878317) 
 
USER AND PUBLIC INVOLVEMENT 
 
Currently a patient focus group including people on HD is being established. 
 
STUDY FINANCES 
 
Funding source  
 
This study is funded by internal funds from the University of Nottingham and Department of 
Renal Medicine, Royal Derby Hospital. 
 
Participant stipends and payments 
 
Participants will not be paid to participate in the study. Travel expenses will be offered for any 
hospital visits in excess of usual care. The majority of study assessments will be timed to 
take place at the time of routine visits for dialysis treatment or assessment. 
 
 
 

 
 
 
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This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
SIGNATURE PAGES 
 
Signatories to Protocol: 
 
Chief Investigator: (name)__________________________________ 
 
 
Signature:__________________________________ 
 
 
Date: ___________ 
 
 
 
 
Co- investigator: 
(name) __________________________________ 
 
 
Signature:__________________________________ 
 
 
Date: ___________ 
 
 
 
 
 
 
 
 
 
 

 
 
 
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Skin autofluorescence as a risk marker in people receiving dialysis  
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This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
REFERENCES 
 
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Advanced glycation end-products and skin autofluorescence in end-stage renal 
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Darlene, A., Dartt Reza, D. and D'Amore, P. (2011) Immunology, inflammation and diseases 
of the eye. Elsevier Press: pp. 287-288. 
 
Graff, R., Arsov, S., Ramsauer, B., Koetsier, M., Sundvall, N., Engels, G.E., Sikole, A., 
Lundberg, L., Rakhorst. G. and Stegmayr, B. (2014) Skin and plasma autofluorescence 
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Hartog, J.W., Voors, A.A., Schalkwijk, C.G., Scheijen, J., Smilde, T.D., Damman, K., Bakker, 
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Ophthalmol 116(12): pp.1629-1632. 
 
Junaid Nazar, C.M., Kindratt, T.B., Ahmad, S.M., Ahmed, M. and Anderson, J. (2014) Barriers 
to the successful practice of chronic kidney diseases at the primary health care level: 
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Kandarakis, S.A., Piperi, C., Topouzis, F. and Papavassiliou, A.G. (2014) Emerging role of 
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Kellow, N.J. and Savige, G.G. (2013) Dietary advanced glycation end product restriction for 
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Kidney Disease: Improving Global Outcomes (KDIGO) CKD Work Group (2013). KDIGO 2012 
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Kimura, H., Tanaka, K., Kanno, M., Watanabe, K., Hayashi, Y., Asahi, K., et al. (2014) Skin 
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Meerwaldt, R., Links, T., Graaff, R., Thorpe, S.R., Baynes, J.W., Hartog, J., Gans, R. and Smit, 
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Skin autofluorescence as a risk marker in people receiving dialysis  
Protocol draft 4.5 Final Version 2.0 04.10.2017 
This protocol is confidential and the property of the University of Nottingham. No part of it may be 
transmitted, reproduced, published, or used by others persons without prior written authorisation 
from the University of Nottingham 
Oleniuc, M., Schiller, A., Secara, I., Onofriescu, M., Hogas, S., Apetrii, M., Siriopol, D. and 
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