Clinical trial NCT04119843
Conditions: Known or Suspected Focal Liver Lesions Severe Renal Impairment
Sponsor: Ascelia Pharma AB
Start date: 2020-02-19; Completion date: 2023-02-17
Enrollment: 87 participants; Allocation: NA; Intervention model: SINGLE_GROUP

Brief summary:
The overall objective of this study is to evaluate the safety and diagnostic efficacy of Mangoral in liver MRI in participants with known or suspected focal liver lesions and severe renal impairment. The diagnostic efficacy of Mangoral will be assessed in terms of visualization of detected focal liver lesions in combined MRI (CMRI: combined Mangoral-enhanced and unenhanced MRI) compared to unenhanced MRI.

Detailed description:
The overall objective of this multicenter, open-label, study is to evaluate the safety and diagnostic efficacy of Mangoral in participants with known or suspected focal liver lesions and severe renal impairment. Study treatment is a single oral dose of Mangoral (800 mg manganese chloride \[II\] tetrahydrate, 500 mg L-alanine, and 800 IU vitamin D3). Adult male and female participants with severe renal impairment or acute kidney injury and who are being evaluated for known or suspected focal liver lesions will be included. Primary diagnostic efficacy in terms of visualization of detected lesions will be evaluated centrally by 3 independent readers. Study MRIs will also be evaluated by the on-site radiologists for the assessment of secondary objectives and for clinical purposes.

Arms: arm 1: All participants will receive a single dose of Mangoral (800 mg Manganese (II) chloride tetrahydrate \[MnCl2 4H2O\]).
Interventions: intervention 1: Mangoral
Intervention descriptions: intervention 1: 800 mg manganese chloride \[II\] tetrahydrate, 500 mg L-alanine, and 800 IU vitamin D3

Protocol documents:
Prot_000.pdf: 
Protocol Number: ASC-Man-P016 
Official Title: A multicentre, open-label study to evaluate the safety and diagnostic efficacy of 
mangoral in patients with known or suspected focal liver lesions and severe renal impairment. 
NCT Number: NCT04119843 
Document Date: 12 October 2021


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Study Protocol 
A Multicentre, Open-label Study to Evaluate the Safety 
and Diagnostic Efficacy of Mangoral in Patients with 
Known or Suspected Focal Liver Lesions and Severe 
Renal Impairment 
EudraCT no.: 2019-001599-12 
IND no.: 102,043 
Trial no.: ASC-Man-P016 
Trial name: SPARKLE 
Version history:  
Version 7.0, 12 October 2021 (including protocol amendment 5) 
Version 6.0, 06 April 2021 (including protocol amendment 4) 
Version 5.0, 06 Jul 2020 (including protocol amendment 3) 
Version 4.0, 23 Apr 2020 (including protocol amendment 2) 
Version 3.0, 26 September 2019 (including protocol amendment 1) 
Version 2.0, 24 June 2019 (original protocol) 
Ascelia Pharma AB 
Hyllie Boulevard 34 
215 32 Malmö 
Sweden 
Confidentiality Statement 
The information in this document is confidential and proprietary to Ascelia Pharma AB. It is understood 
that the information will not be disclosed to any person or institution other than investigators or 
consultants for review, their staff, applicable Ethics Committee(s) / Institutional Review Board(s), 
regulatory authorities and trial subjects without express authorisation by Ascelia Pharma AB. 


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PROTOCOL SYNOPSIS 
Title: 
A multicentre, open-label study to evaluate the safety and 
diagnostic efficacy of mangoral in patients with known or suspected 
focal liver lesions and severe renal impairment. 
Trial no.: 
ASC-Man-P016  
Trial name: 
SPARKLE 
EudraCT no.: 
2019-001599-12 
IND no.: 
102,043 
Clinical phase: 
Phase III 
Investigational medicinal 
product: 
Manganese (II) chloride tetrahydrate (MnCl2•4H2O; working name: 
mangoral) powder for oral administration. 
Study objective(s): 
The overall objective of this study is to evaluate the safety and 
diagnostic efficacy of mangoral (equiv. to 800 mg MnCl2•4H2O) in 
patients with known or suspected focal liver lesions and severe 
renal impairment. 
Primary objective: 
• 
To assess the diagnostic efficacy of mangoral in liver MRI in 
terms of visualisation of detected focal liver lesions in 
combined MRI (CMRI: combined mangoral-enhanced and 
unenhanced MRI) compared to unenhanced MRI 
Lesion visualisation will be determined by qualitative 
assessments of lesion border delineation and lesion 
contrast compared to background liver on 4-point scales for 
up to 15 lesions per patient 
Secondary objectives: 
• 
To assess the diagnostic efficacy of mangoral in liver MRI in 
terms of: 
o number of lesions detected by each MRI method 
(unenhanced MRI, mangoral-enhanced MRI, and 
CMRI) 
o visualisation of detected focal liver lesions in 
mangoral-enhanced MRI compared to unenhanced 
MRI (determination of visualisation will be done in the 
same way as for the primary efficacy variable) 
o confidence in lesion detection and localisation 
separately in unenhanced MRI, mangoral-enhanced 
MRI, and CMRI 
o lesion dimensions (independent off-site readers’ 
assessment): longest diameter of the largest and 
smallest lesion 
o quantitative assessments by measuring percent 
signal intensity enhancement of liver, liver-to-lesion 


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contrast, signal-to-noise ratio, and contrast-to-noise 
ratio of up to 5 lesions per patient 
o number of patients who had at least one new lesion 
identified on CMRI compared to unenhanced MRI 
alone 
• 
To assess a proportion of patients having at least one 
malignant lesion identified on post-mangoral images that 
was not identified on pre-mangoral images.  
• 
To evaluate the safety and tolerability of mangoral 
• 
To evaluate the pharmacokinetics of manganese after a 
single dose of mangoral in a subgroup of patients (including 
a small number of dialysis patients) 
• 
To evaluate the impact of diagnostic performance of CMRI 
and mangoral-enhanced MRI versus unenhanced MRI on 
the patients’ management 
Design: 
Multicentre, open-label, pivotal Phase III study to evaluate the 
safety and diagnostic efficacy of mangoral.  
Treatment: 
Single oral dose of mangoral (800 mg manganese chloride [II] 
tetrahydrate, 500 mg L-alanine, and 800 IU vitamin D3). 
Population: 
Adult male and female patients with severe renal impairment 
(Chronic Kidney Disease [CKD] or Acute Kidney Injury [AKI]) and 
who are being evaluated for known or suspected focal liver lesions. 
Approximately 80% of enrolled patients should have liver 
metastasis or other common focal liver lesions. The proportion 
enrolled HCC patients will be monitored jointly by the CRO/sponsor 
in order to achieve approximately 20% HCC patients in the overall 
study population. The CRO/sponsor may instruct individual sites, 
from time to time, to specifically and exclusively enrol patients with 
either non-HCC or HCC. 
PK subgroup: Pharmacokinetic (PK) evaluations will be performed 
in a subgroup of patients included in the study. This PK subgroup 
will consist of 12 patients, including 4 patients currently on 
maintenance haemodialysis. These patients will be selected from 
few sites that are able to include these patients and follow PK 
requirements. 
Inclusion criteria: 
1. Patients who have been fully informed and have personally 
signed and dated the informed consent 
2. Male and female patients 18 years and older 
3. Patients who have known or suspected focal liver lesions 
based on medical history and previous laboratory and/or 
imaging examinations 
4. Patients with severe renal impairment; 
a. Chronic 
kidney 
disease 
(CKD) 
(estimated 
glomerular 
filtration 
rate 
[eGFR] 


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< 30 mL/min/1.73 m2) based on medical history and 
previous laboratory examinations,  at least once 
within the last 3 months prior to the Baseline Visit 
OR 
b. Acute kidney injury (AKI) with an increase in serum 
creatinine ≥ 0.3 mg/dL within 48 hours or ≥ 50% 
within 7 days,  prior to the Baseline Visit 
 
5. Female patients who are not of childbearing potential, or 
female patients who are of childbearing potential but are 
using highly effective contraception (including hormonal 
contraceptives, such as combined oral contraceptives, 
patch, vaginal ring, injectables, and implants, intrauterine 
device, 
intrauterine 
hormone-releasing 
system, 
vasectomised partner, and tubal ligation) or practising 
sexual abstinence (only accepted if this is the usual and 
preferred lifestyle of the patient), and have a negative urine 
or serum pregnancy test within 24 hours prior to the 
administration of contrast agent 
6. Sexually active male patients (who are not vasectomised) 
must be willing to use condoms after administration of 
mangoral until the last follow-up visit 
7. Patients who agree to comply with all study procedures as 
outlined within the study protocol and informed consent 
8. Not applicable 
Exclusion criteria: 
1. Patients with simple liver cysts only 
2. Patients who have received any investigational drug or were 
treated with an investigational device within 6 weeks prior to 
the Baseline Visit 
3. Patients who have received any MRI contrast media within 
6 weeks prior to the Baseline Visit or are scheduled to 
receive any other contrast medium other than mangoral 
before the last study visit 
4. Patients who have previously been enrolled in the study and 
underwent mangoral-enhanced MRI 
5. Patients who are clinically unstable or have an acute illness, 
as judged by the investigator; i.e. the underlying clinical 
condition is such that clinically relevant changes in condition 
can be reasonably anticipated during the study period 
6. Patients with severe hepatic impairment (according to Child-
Pugh score C) 
7. Not applicable 
8. Patients scheduled for surgery before the last study visit 
9. Pregnant or lactating women 
10. Patients with a history of severe allergies (e.g. anaphylaxis), 
as judged by the investigator 


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11. Patients with a known hypersensitivity to the IMP or any 
excipients in the IMP 
12. Patients who are, in the investigator’s opinion, unable to 
comply with the study requirements including follow-up 
procedures 
13. Patients with known portosystemic shunts 
14. Patients with encephalopathy, as judged by the investigator 
15. Patients with neurodegenerative disorders 
16. Patients with acute neurological disorders, as judged by the 
investigator 
17. Patients with haemochromatosis 
18. Patients receiving blood transfusion within 14 days prior to 
the Baseline Visit 
19. Patients who cannot fast for 8 hours 
20. Patients who cannot undergo MRI due to MRI incompatible 
pacemaker, metal implants, or other MRI contraindications 
21. Patients with known inherited disorders of manganese 
metabolism 
22. Patients with known pheochromocytoma 
23. Patients with history of surgical/other interventional 
procedures or conditions which may interfere with 
absorption or excretion of IMP (e.g. small bowel obstruction, 
gastric bypass, gastric resection, biliary obstruction), as 
judged by the investigator 
24. Patients who have been institutionalised by official or court 
order and therefore are not completely free in their decision-
making process 
25. Patients dependent on the sponsor, investigators, personnel 
of the trial site or any other individual, whose willingness to 
volunteer may be unduly influenced by the expectation, 
whether justified or not, of benefits associated with 
participation. 
Methodology: 
The study consists of the following periods/visits: 
• 
Screening period (day -28 to -1; assessments can be 
performed anytime during the 4-week period): informed 
consent, inclusion/exclusion criteria, patient characteristics, 
serum pregnancy test (female patients of childbearing 
potential) 
• 
Baseline period (day -1 to day 0, i.e. within 24 hours prior to 
the administration of mangoral): safety assessments, 
neurological assessments, unenhanced MRI examination of 
the liver (baseline MRI), pre-dose blood samples for 
manganese 
measurements, 
pregnancy 
test 
(female 
patients of childbearing potential) 


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• 
Day of MRI (day 0): intake of mangoral after a fast of at least 
4 hours and after unenhanced MRI (note: fasting must be 
continued until after the mangoral-enhanced liver MRI 
examination has been performed (ie 4 [± 1] hours after 
mangoral administration) 
• 
Follow-up visits following contrast administration (24 [± 4] 
hours, 48 [± 4] hours, and 5 [±2] days post-dose): safety 
assessments, neurological assessment, blood samples for 
manganese concentrations.  
Note: In case a patient is not able to return for the follow-up visits to 
the study site, these follow-up assessments may be performed at an 
off-site location (e.g. at the patient’s home or at an associated off-site 
clinic by a trained examiner): Patients with long travelling distances 
may stay at a hotel near the study site at the expense of the sponsor. 
If further adjustments are required, Visit 4 (at 24 [± 4] hours) and Visit 
5 (48 [± 4] hours) can be performed as a remote visits, at the 
discretion of the investigator. 
• 
A subgroup of patients who underwent brain MRI due to any 
clinical reason within the last 6 months prior to 
mangoral-enhanced MRI may also undergo an optional 
brain MRI 7 (+ 2) days after mangoral-enhanced MRI of the 
liver 
PK subgroup: additional blood samples (6 mL per time point) will be 
drawn for the determination of PK parameters on the day of the liver 
MRI examination (day 0) at 0.25, 0.5, 1, 2, 3, 4, 6, 8, and 12 hours 
post-dose. 
Primary diagnostic efficacy in terms of visualisation of detected 
lesions will be evaluated centrally at an imaging core laboratory by 
three independent readers who are experienced in liver imaging. A 
fourth reader who is not involved in efficacy reads will track and 
match the detected lesions on unenhanced, mangoral-enhanced 
and CMRI to confirm lesion numbering and lesion location across 
the modalities. Lesion tracking will also confirm lesions that are 
identified on pre-mangoral, post-mangoral and CMRI images. Study 
MRIs will also be evaluated by the on-site radiologists for the 
assessment of secondary objectives and for clinical purposes. Both 
on-site and independent radiologists will be trained prospectively in 
a standardised way. 
Efficacy variables: 
Primary efficacy variable: 
The primary efficacy endpoint will be the visualisation of detected 
focal liver lesions in combined MRI (CMRI, mangoral-enhanced 
MRI plus unenhanced MRI) as compared to unenhanced MRI.  
Visualisation will be measured by two co-primary variables: 
• lesion border delineation 


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• lesion contrast compared to liver background 
Both parameters will be determined for each lesion (up to 15 lesions 
per patient) and by each of the three independent readers by 
qualitative assessment on 4-point scales with categories ‘poor’ (1), 
‘partial/moderate’ (2), ‘good’ (3), and ‘excellent’ (4) lesion border 
delineation/lesion contrast. 
Based on the scores on the 4-point scales, two sum scores will be 
calculated for each patient and separately for each MRI method: a 
lesion border delineation sum score and a lesion contrast sum 
score. Only lesions detected at baseline (unenhanced MRI) will be 
included in the assessment of diagnostic efficacy.  
Secondary efficacy variables: 
• 
Number of lesions detected by each MRI method: 
unenhanced MRI, mangoral-enhanced MRI, and CMRI (on-
site and independent off-site readers’ assessments) 
• 
Visualisation of focal liver lesions in mangoral-enhanced 
MRI as compared to unenhanced MRI (independent off-site 
readers’ assessment; visualisation will be assessed as for 
the primary efficacy endpoint) 
• 
Confidence in lesion detection separately in unenhanced 
MRI, mangoral-enhanced MRI, and CMRI (3-point scale, 
on-site and independent off-site readers’ assessments of up 
to 15 lesions per patient) 
• 
Confidence in lesion localisation separately in unenhanced 
MRI, mangoral-enhanced MRI, and CMRI (3-point scale, 
on-site and independent off-site readers’ assessments of up 
to 15 lesions per patient) 
• 
Lesion 
dimensions 
(independent 
off-site 
readers’ 
assessment): longest diameter of the largest and the 
smallest lesion 
• 
Quantitative assessments will be performed for up to 5 
detected 
lesions 
(independent 
off-site 
readers’ 
assessment). These lesions will be the same for pre- and 
post-contrast assessments: 
o liver signal intensity (SI) enhancement (%) 
o liver-to-lesion contrast (LLC) 
o signal-to-noise ratio (SNR) 
o contrast-to-noise ratio (CNR) 
• 
Change(s) in patients’ management based on the diagnostic 
performance of CMRI or mangoral-enhanced MRI versus 
unenhanced MRI: 
o Any changes in patient management based on MRI 
findings (yes/no) 
o Next steps in patient management based on MRI 
findings (i.e. chemotherapy, surgery, local ablation 
procedure, combination therapy, or other [specify]) 


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Subgroup analyses: 
The primary efficacy endpoint will be evaluated in appropriate 
subgroups which will be defined in the statistical analysis plan (e.g. 
subgroups by MRI scanner’s magnetic field strength, lesion type, 
lesion diameter, age, and eGFR). 
Safety variables: 
Vital signs, physical examination, neurological assessment, 
electrocardiogram (ECG), clinical safety laboratory testing 
(haematology, 
biochemistry, 
urinalysis), 
blood 
manganese 
concentrations, monitoring of adverse events, and evaluation of 
brain MRI in a subgroup of patients. 
Pharmacokinetic 
variables 
The following PK variables will be determined: AUC (area under the 
curve) for the interval 0-120 hours, Cmax (maximum or peak 
concentration), tmax (time to achieve maximum concentration), t1/2 
(terminal half-life) and Clapp (total apparent clearance). 
Statistical methods: 
The two co-primary variables (patients’ lesion border delineation 
and patients’ lesion contrast) will be analysed for the full analysis 
population (i.e. all patients who received mangoral and for whom all 
unenhanced/enhanced MRI are assessable). The superiority of 
CMRI over unenhanced MRI regarding both co-primary variables 
(using the mean score differences between the paired sum scores 
weighted by the number of lesions determined per patient) will be 
evaluated for each independent reader.  
The study will be considered successful if the reading results of at 
least two of three independent readers demonstrate the superiority 
of CMRI versus unenhanced MRI for both lesion border delineation 
and lesion contrast. Hypotheses will be tested using a one-sided 
paired t-test. 
A comparison of unenhanced MRI and mangoral-enhanced MRI 
alone will be evaluated in the same way as the primary analysis. 
The secondary efficacy variables will be analysed in an exploratory 
manner using descriptive statistics (including confidence intervals). 
Safety and PK data will be analysed descriptively. 
Number of patients: 
Approximately 197 patients will be enrolled to ensure a sample size 
of 167 patients for the primary analysis at an overall study power of 
80%, assuming 85% of the image sets being completely evaluable. 
To adjust for testing the simultaneous success of two co-primary 
variables, the power of each individual test will be set to 90%. The 
simultaneousness does not influence the type I error. Thus, the 
tests will be performed at a global 1-sided level of significance of 
0.025. 
Number of sites: 
Approximately 60 sites. 
 
 




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SERIOUS ADVERSE EVENT REPORTING CONTACT DETAILS 
All SAEs must be reported immediately (within 24 hours from the investigator’s awareness) to:  
E-mail: 
 
 
Please refer to Section 6.3.7.4 for more details. 
 
Sponsor’s Safety Responsible Person: 
 
Email: 
 
Phone: 
 
 
 
 


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ABBREVIATIONS 
ADR 
Adverse drug reaction 
AE 
Adverse event 
AKI 
Acute Kidney Injury 
ALAT 
Alanine aminotransferase 
ASAT 
Aspartate aminotransferase 
BD 
Border delineation 
BUN 
(serum) Blood Urea Nitrogen 
CKD 
Chronic Kidney Disease 
CMRI 
Combined mangoral-enhanced and unenhanced MRI 
CNR 
Contrast-to-noise ratio 
CRO 
Contract research organisation 
CT 
Computed tomography 
CTCAE 
Common Terminology Criteria for Adverse Events 
DICOM 
Digital Imaging and Communications in Medicine 
DSMB 
Data Safety Monitoring Board 
DWI 
Diffusion-weighted imaging 
ECG 
Electrocardiogram 
eCRF 
Electronic case report form 
eGFR 
Estimated glomerular filtration rate 
EudraCT 
European Union Drug Regulating Authorities Clinical Trials 
FDA 
Food and Drug Administration 
 γ-GT 
Gamma-glutamyltransferase 
GBCA 
Gadolinium-based contrast agent 
GCP 
Good Clinical Practice 
GMP 
Good Manufacturing Practice 
GRAS 
‘Generally Recognised As Safe’ 
HCC 
Hepatocellular carcinoma 
ICH 
International Council for Harmonisation of Technical Requirements for 
Pharmaceuticals for Human Use 
ICP-MS 
Inductively coupled plasma mass spectrometry 
IDSMB 
Independent Data Safety Monitoring Board 
IEC 
Independent ethics committee 
IMP 
Investigational medicinal product 


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INR 
International normalised ratio 
IRB 
Institutional review board 
IRC 
Independent review charter 
LC 
Lesion contrast compared to liver background (qualitative) 
LLC 
Liver-to-lesion contrast (quantitative) 
MDRD 
Modification of Diet in Renal Disease 
MedDRA 
Medical Dictionary for Regulatory Activities 
Mn 
Manganese 
MnCl2•4H2O 
Manganese (II) chloride tetrahydrate 
MRI 
Magnetic resonance imaging 
N 
Number of patients 
NSF 
Nephrogenic systemic fibrosis 
PK 
Pharmacokinetics 
ROI 
Region of interest 
SAE 
Serious adverse event 
SD 
Standard deviation 
SI 
Signal intensity 
SNR 
Signal-to-noise ratio 
SOP 
Standard operating procedure 
SUN 
Serum urea nitrogen 
SUSAR 
Suspected Unexpected Serious Adverse Reaction 


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TABLE OF CONTENTS 
Protocol synopsis ................................................................................................................ 2 
Flow-chart ............................................................................................................................. 9 
Serious adverse event reporting contact details ............................................................. 10 
Abbreviations ..................................................................................................................... 11 
Table of contents ................................................................................................................ 13 
1 
Introduction ...................................................................................................... 18 
1.1 
Background ........................................................................................................ 18 
1.1.1 
Focal liver lesions .......................................................................................... 18 
1.1.2 
Imaging of focal liver lesions .......................................................................... 18 
1.1.3 
Manganese-based contrast agents for MRI ................................................... 19 
1.1.4 
Mangoral ....................................................................................................... 19 
1.2 
Rationale of the study......................................................................................... 21 
1.3 
Risk-benefit assessment .................................................................................... 21 
2 
Study objectives and purpose ........................................................................ 23 
2.1.1 
Primary objective ........................................................................................... 23 
2.1.2 
Secondary objectives ..................................................................................... 23 
3 
Overall study design and plan description..................................................... 24 
3.1 
Discussion of study design ................................................................................. 25 
4 
Selection of study population ......................................................................... 26 
4.1 
Inclusion criteria ................................................................................................. 26 
4.2 
Exclusion criteria ................................................................................................ 27 
4.3 
Withdrawal criteria .............................................................................................. 28 
5 
Trial treatments ................................................................................................ 29 
5.1 
Investigational medicinal product(s) ................................................................... 29 
5.1.1 
Administration ................................................................................................ 29 
5.1.2 
Identity of the investigational medicinal product ............................................. 29 
5.1.3 
Method of assigning patients to treatment groups .......................................... 30 
5.1.4 
Selection of doses in the study ...................................................................... 30 
5.1.5 
Selection and timing of dose for each patient ................................................ 31 
5.1.6 
Supply, manufacturing, packaging and labelling ............................................ 31 
5.1.7 
Storage .......................................................................................................... 31 
5.1.8 
Shipment ....................................................................................................... 31 
5.1.9 
Blinding and randomisation............................................................................ 31 
5.1.10 
Investigational medicinal product compliance and accountability ................... 31 
5.1.11 
Management of investigational medicinal product overdose .......................... 31 


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5.2 
Prior and concomitant therapy ............................................................................ 32 
5.2.1 
Prohibited or prohibited with restrictions ........................................................ 32 
5.2.2 
Permitted oral intake ...................................................................................... 32 
6 
Variables and methods .................................................................................... 32 
6.1 
Population Characteristics .................................................................................. 32 
6.1.1 
Demographics ............................................................................................... 32 
6.1.2 
Medical history ............................................................................................... 33 
6.1.2.1 
General medical history ...................................................................................... 33 
6.1.2.2 
Referral diagnosis .............................................................................................. 33 
6.1.2.3 
Child-Pugh score ................................................................................................ 33 
6.1.2.4 
Estimated glomerular filtration rate (eGFR) ........................................................ 34 
6.2 
Diagnostic efficacy ............................................................................................. 35 
6.2.1 
Definition of liver segments ............................................................................ 35 
6.2.2 
Primary and secondary efficacy variables ...................................................... 36 
6.2.2.1 
Primary efficacy variable .................................................................................... 36 
6.2.2.2 
Secondary efficacy variables .............................................................................. 36 
6.2.3 
Derivation of efficacy variables ...................................................................... 38 
6.2.3.1 
Visualisation of focal liver lesions ....................................................................... 38 
6.2.3.2 
Confidence in lesion detection ............................................................................ 39 
6.2.3.3 
Confidence in lesion localisation ........................................................................ 40 
6.2.3.4 
Lesion dimensions ............................................................................................. 40 
6.2.3.5 
Quantitative assessments .................................................................................. 40 
6.2.4 
Imaging procedures ....................................................................................... 41 
6.2.4.1 
Image acquisition ............................................................................................... 41 
6.2.4.2 
Image handling and processing .......................................................................... 43 
6.2.5 
Efficacy assessments .................................................................................... 43 
6.2.5.1 
Off-site (central) assessments ............................................................................ 43 
6.2.5.2 
On-site assessment ........................................................................................... 43 
6.3 
Safety ................................................................................................................. 43 
6.3.1 
Vital signs ...................................................................................................... 44 
6.3.2 
Physical examination ..................................................................................... 44 
6.3.3 
Neurological assessments ............................................................................. 44 
6.3.4 
Electrocardiogram (ECG) .............................................................................. 44 
6.3.5 
Laboratory variables ...................................................................................... 45 
6.3.5.1 
Screening laboratory tests .................................................................................. 45 
6.3.5.2 
Clinical safety laboratory tests ............................................................................ 45 
6.3.5.3 
Blood manganese concentration ........................................................................ 46 
6.3.5.4 
Pregnancy test ................................................................................................... 47 
6.3.6 
Additional safety assessments ....................................................................... 47 
6.3.6.1 
MRI of the brain – Optional................................................................................. 47 


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6.3.7 
Adverse events .............................................................................................. 47 
6.3.7.1 
Definition of adverse events ............................................................................... 48 
6.3.7.2 
Assessment of adverse events ........................................................................... 48 
6.3.7.3 
Follow-up of adverse events ............................................................................... 50 
6.3.7.4 
Documentation and reporting of adverse events ................................................ 50 
6.4 
Pharmacokinetics ............................................................................................... 51 
6.4.1 
Pharmacokinetic variables ............................................................................. 51 
6.4.2 
Blood sampling time points, handling and labelling of blood samples ............ 51 
7 
Study conduct .................................................................................................. 53 
7.1 
Study schedule................................................................................................... 53 
7.2 
Observations by visit .......................................................................................... 56 
7.2.1 
Screening visit: V1 (day -28 to -1) .................................................................. 56 
7.2.1.1 
Screening ........................................................................................................... 56 
7.2.1.2 
Re-screening and re-consenting ........................................................................ 56 
7.2.2 
Baseline period: V2 (day -1) to V3 (day 0), within 24 hours pre-dose ............. 56 
7.2.3 
Day of Dosing: V3 (day 0), dosing, mangoral-enhanced MRI and post-dose 
assessments ................................................................................................. 57 
7.2.4 
Follow-up visit V4 (day 1) .............................................................................. 57 
7.2.5 
Follow-up visit V5 (day 2) .............................................................................. 58 
7.2.6 
Follow-up visit V6 (day 5) .............................................................................. 58 
7.2.7 
Follow-up visit V7 (day 7) in optional brain MRI subgroup ............................. 58 
7.2.8 
Withdrawal Visit ............................................................................................. 59 
7.3 
Independent Data Safety Monitoring Board (IDSMB) ......................................... 59 
7.4 
Duration of the study .......................................................................................... 59 
7.4.1 
Planned duration for the individual patient ..................................................... 59 
7.4.2 
Premature termination ................................................................................... 60 
8 
Statistics ........................................................................................................... 61 
8.1 
Statistical and analytical plans ............................................................................ 61 
8.1.1 
Analysis populations ...................................................................................... 61 
8.1.2 
Adaptive study design .................................................................................... 62 
8.1.3 
Primary efficacy analysis ............................................................................... 62 
8.1.4 
Secondary efficacy analyses ......................................................................... 62 
8.1.5 
Subgroup analyses ........................................................................................ 63 
8.1.6 
Safety analyses ............................................................................................. 63 
8.1.7 
Pharmacokinetic analyses ............................................................................. 64 
8.1.8 
Missing data .................................................................................................. 64 
8.1.9 
Multicentre trial .............................................................................................. 64 
8.1.10 
Patient data listings ....................................................................................... 64 
8.1.11 
Deviations from the planned statistical analysis ............................................. 64 
8.1.12 
Interim analysis .............................................................................................. 64 


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8.1.13 
Software used for statistical analysis ............................................................. 64 
8.1.14 
Determination of sample size ......................................................................... 64 
9 
Data handling and data quality assurance ..................................................... 67 
9.1 
Data management .............................................................................................. 67 
9.2 
Documentation ................................................................................................... 67 
9.2.1 
Screening log and patient identification list .................................................... 67 
9.2.2 
Source data and patient records .................................................................... 67 
9.2.3 
Electronic case report form (eCRF) ............................................................... 68 
9.3 
Direct access to source data/documents ............................................................ 68 
9.4 
Monitoring .......................................................................................................... 68 
9.5 
Quality assurance .............................................................................................. 69 
10 
Ethical, legal and administrative aspects ....................................................... 70 
10.1 
Approval procedures .......................................................................................... 70 
10.2 
Protocol amendments ........................................................................................ 70 
10.3 
Informed consent ............................................................................................... 70 
10.4 
Confidentiality and data protection ..................................................................... 71 
10.5 
Liability, insurance and finances ......................................................................... 71 
10.6 
Clinical study report, publication and use of study results ................................... 72 
10.7 
Archiving of study documents ............................................................................. 72 
11 
List of references ............................................................................................. 73 
12 
Clinical Study Protocol - Signature Pages ..................................................... 76 
13 
Appendices ....................................................................................................... 78 
I 
Declaration by the Investigator  
 
List of Tables 
Table 1 
Overview on previous Phase I and II studies on mangoral .................................20 
Table 2 
Properties of the investigational medicinal product .............................................30 
Table 3 
Physio-chemical properties of manganese (II) chloride tetrahydrate ...................30 
Table 4 
Child-Pugh scoring system .................................................................................34 
Table 5 
Overview of efficacy variables ............................................................................38 
Table 6 
MRI parameters .................................................................................................42 
Table 7 
Screening laboratory parameters .......................................................................45 
Table 8 
Clinical safety laboratory parameters (baseline and post-dose) ..........................46 
Table 9 
Study schedule...................................................................................................54 
Table 10 
Definition of analysis populations .......................................................................61 
Table 11 
Sample sizes ......................................................................................................65 
 


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List of Figures 
Figure 1 
Schematic representation of liver segments according to Couinaud ...................35 
Figure 2 
Sample sizes depending on correlation and mean differences ...........................66 
 


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1 
INTRODUCTION 
1.1 
Background 
1.1.1 
Focal liver lesions 
The widespread use of imaging modalities has increased the detection rates of focal liver 
lesions, either incidentally in asymptomatic patients or in high-risk patients, such as patients 
with liver cirrhosis. These lesions can be either benign, including focal nodular hyperplasia, 
hepatocellular adenoma and hepatic cysts, or malignant.  
Liver metastases are the most frequent type of malignant focal liver lesion. Although cancer 
can be hard to control when it has spread, some types of metastatic cancer can be cured, or 
the growth and burden of the metastases reduced or eliminated, leading to improved life 
expectancy and relieved symptoms if detected at an early stage and handled appropriately. 
This is especially relevant for liver metastases. The liver is the second most common organ 
for metastasis after the lymph nodes [1]. Many solid cancers originating e.g. from the lungs, 
breasts, colon, stomach or pancreas metastasise to the liver, and very often the liver is the first 
site of metastatic disease. If liver metastases from colorectal cancer are detected early and 
deemed eligible for surgical resection, the survival rate can be significantly improved, and 
sometimes full recovery is possible. The 5-year overall survival rate for patients undergoing 
resection for colorectal liver metastases has been reported to be 46% compared to only 6% 
for patients who were not subjected to surgical treatment of their liver metastases [2]. Surgical 
resection of liver metastases from non-colorectal primary tumours such as breast cancer has 
also been reported to lead to improved survival outcome [3, 4].  
Hepatocellular carcinoma (HCC) is the most common primary malignant hepatic tumour and 
occurs mainly in patients with underlying chronic liver disease and cirrhosis. Time to recurrence 
is longer and 5-year survival rate is higher in patients with early HCC than in those with 
advanced stages [5]. Thus, early detection of HCC is crucial for successful treatment and 
reduction of mortality. 
Factors affecting patient management include number, size and location of focal liver lesions. 
Cross-sectional imaging plays an essential role in both initial staging (i.e. determining the stage 
of the cancer), preoperative planning, monitoring of treatment effect and surveillance for 
recurrence of disease [6]. Contrast-enhanced computed tomography (CT) and contrast -
enhanced magnetic resonance imaging (MRI) have shown higher sensitivity and specificity as 
compared to unenhanced CT / MRI for detecting and localizing focal liver lesions. However, in 
patients with severe renal impairment and focal liver lesions utilisation of both iodinated and 
gadolinium-based contrast agents is either restricted or contra-indicated. Oral mangoral is 
being developed to provide manganese-based contrast enhancement that has the potential to 
provide higher accuracy in lesion detection and visualisation as compared to unenhanced MRI.  
1.1.2 
Imaging of focal liver lesions 
Several imaging modalities are available for the screening and evaluation of focal liver lesions, 
including liver metastases and HCC, such as CT, positron emission tomography combined 
with unenhanced CT, MRI, and ultrasound. Although the choice of imaging techniques may be 
based on the preference and expertise of the institution, contrast-enhanced CT and contrast-
enhanced MRI are preferred techniques. Magnetic resonance is rapidly emerging as a 
preferred imaging option, especially for detection and characterisation of small liver lesions, 
due to its high accuracy resulting from optimal lesion-to-liver contrast and absent radiation 
exposure [7].  


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Contrast enhancement is an essential component of a liver MRI examination as it improves 
visualisation and delineation of focal liver lesions against normal liver parenchyma. Liver-
specific imaging contrast agents offer greater lesion-to-liver contrast than conventional 
extracellular fluid space agents [8]. 
1.1.3 
Manganese-based contrast agents for MRI 
Manganese (Mn) is an essential trace element in humans, animals and plants and is found in 
nature in a wide variety of minerals, often in combination with iron. The manganese cation 
(Mn2+) has a great potential for use in MRI contrast agents as it has powerful paramagnetic 
properties due to its 5 unpaired electrons. Similarly to gadolinium- and copper-containing 
contrast agents, manganese-based contrast agents increase the signal intensity of 
T1-weighted images [9, 10, 11]. Manganese is present in many intracellular organelles, 
especially in mitochondria, where it plays a role in protein synthesis. Thus, organs such as the 
liver consisting of cells with high mitochondrial content are predestinated targets for 
manganese-based contrast agents. Currently marketed liver MRI contrast agents are 
intravenous injections based on the lanthanide heavy metal gadolinium; however, there has 
been increasing safety concern with the use of gadolinium-based contrast agents (GBCAs), 
especially in patients with impaired renal function. Manganese based contrast agents offer an 
alternative to gadolinium [12]. A chelated form of manganese was used as the liver MRI 
contrast agent mangafodipir trisodium, which has been the only US Food and Drug 
Administration (FDA)-approved and marketed manganese-based contrast agent so far, but the 
agent was withdrawn from the US market in 2003 and from the European market in 2012. 
Mangafodipir trisodium was approved for intravenous injection. Following injection, the 
chelated manganese dissociates into the chelate and the manganese ion. The latter binds to 
serum proteins and is accumulated in several organs, including the liver, kidney and heart 
muscle. Adverse events (AEs) that were frequently reported in clinical studies of mangafodipir 
trisodium-included nausea, headache, pruritus, and sensations of heat and flushing; 
cardiovascular effects were also observed [9]. 
1.1.4 Mangoral 
Mangoral is a manganese-based contrast agent for liver MRI developed for oral administration. 
It contains manganese (II) chloride tetrahydrate (MnCl2•4H2O) as its active pharmaceutical 
ingredient. As manganese is poorly absorbed from the intestine, L-alanine and vitamin D3 are 
included in the formulation and function as absorption promoters to increase the uptake of 
manganese into hepatocytes in the liver [13]. As manganese accumulates in the normal 
hepatocytes but not in liver metastases, the hypointense lesions are clearly visible against the 
hyperintense liver parenchyma on MRI images.  
The advantage of the oral route of administration of manganese is its portal venous transport 
to the liver and its fast uptake into hepatocytes resulting in a very low systemic availability of 
manganese. As mainly the enterohepatic circulation is involved, the risk for toxicities in the 
central nervous system, heart, or kidney is drastically reduced.  
To date, mangoral has been studied in 6 completed clinical Phase I and II trials in healthy 
volunteers (N = 52, in which 2 received placebo) and patients with known liver metastases or 
suspected liver lesions (N = 75). An overview of these studies is given in Table 1. In addition, 
77 patients received mangoral in a compassionate use program based on referral diagnoses 
that involved different types of focal liver lesions and diseases of the intestines (e.g. different 
types of primary cancers with liver metastases, hepatocellular carcinoma, hemangiomas, 


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jaundice, cholangitis, infections, cysts and other diseases) regardless of kidney function. All 
patients in the compassionate use program received 1600 mg mangoral. 
Taken together, the diagnostic efficacy analyses of the conducted clinical studies showed 
improved diagnostic quality scores after administration of mangoral. Compared to unenhanced 
liver MRI, administration of mangoral resulted in increased relative signal intensity in the liver, 
and improved delineation and visualisation of liver lesions. Dose-dependency was observed 
for some of the efficacy variables assessed, with the 800 mg dose giving the best balance 
between safety and efficacy. In comparison to an intravenous gadobenate dimeglumine 
protocol, mangoral was comparable in the sensitivity to detect colorectal cancer liver 
metastases. 
No safety concerns have been identified based on monitoring of AEs, clinical safety laboratory 
assessments, vital signs, electrocardiograms, and pharmacokinetic (PK) analyses in healthy 
volunteers and / or patients with liver metastases included in the clinical development program 
so far. Adverse drug reactions were mainly mild and transient gastrointestinal tract symptoms. 
In addition, blood concentrations of manganese were within the normal range at all four 
investigated doses. Thus, mangoral is expected to have a favourable safety profile compared 
to current intravenous and gadolinium-based MRI contrast media. 
 
Table 1 
Overview on previous Phase I and II studies on mangoral 
Study 
No. subjects 
Study design 
Mangoral 
(MnCl2•4H2O 
equiv.) dose 
Key results 
CMC-P001 
[14] 
Healthy subjects 
N=20 (2 received 
placebo) 
Phase I, 
open-label, 
dose-rising 
800 mg / 
1600 mg 
Data suggested that mangoral 
may be an effective MRI 
contrast agent 
CMC-P002 
[15] 
Patients with liver 
metastases 
N=18 
Phase II, 
open-label 
1600 mg 
Diagnostic quality scores 
improved after mangoral 
CMC-P003 
[16] 
Patients with liver 
metastases 
N=20 
Phase II, 
randomised, 
parallel group, 
open-label 
800 mg / 
1600 mg 
Robust liver signal intensity 
enhancement; optimal 
diagnostic time window 
between 2 and 6 hours 
CMC-P004 
[17] 
Patients with liver 
metastases 
N=20 
Phase II, 
randomised, 
cross-over 
1600 mg 
Sensitivity to detect liver 
metastases was comparable 
between mangoral and 
gadobenate dimeglumine 
CMC-P005 
[not 
published] 
Patients with liver 
metastases 
N=17 
Phase II, 
randomised, 
parallel group, 
open-label 
800 mg / 
1600 mg 
Improved delineation of focal 
liver lesions after mangoral  
CMC-P010 
[18] 
Healthy subjects 
N=32 
Phase II, 
randomised, 
double-blind, 
cross-over, 
dose-response 
200 mg / 
400 mg / 
800 mg 
Increase in liver-to-muscle 
signal intensity ratio and image 
quality superior after the 
800 mg dose 
 


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Note: for a more detailed description of individual clinical study results and an overview on pre-
clinical studies, refer to the current version of the Mangoral investigator’s brochure [19], which 
will be provided to all investigators. Pharmacological and pharmaceutical aspects are also 
reviewed by Jørgensen et al [20]. 
1.2 
Rationale of the study 
Mangoral obtained Orphan Drug Designation by the FDA for use in diagnostic liver MRI in 
patients where the use of GBCAs may be medically inadvisable or where GBCAs cannot be 
administered. The currently used MRI contrast agents based on gadolinium have been 
associated with nephrogenic systemic fibrosis (NSF) in patients with severely impaired kidney 
function. Consequently, regulatory authorities have contraindicated or warned against the use 
of these contrast agents in patients with an estimated glomerular filtration rate (eGFR) 
< 30 ml/min/1.73 m2 [21]. Currently, there is no safe and effective contrast agent available for 
these patients to improve the detection of liver lesions in MRI examinations. 
Due to the high first-pass of manganese in the liver upon oral administration, and the small 
fraction of manganese reaching the circulatory system, mangoral is expected to be a safe 
alternative to gadolinium-based MRI contrast agents or iodinated contrast agents used for 
computed tomography. Thus, mangoral may fill the unmet medical need for improved lesion 
detection in these patients. 
This study is a Phase III study to investigate the diagnostic efficacy and safety of mangoral in 
patients with known or suspected focal liver lesions who also have severe renal insufficiency.  
1.3 
Risk-benefit assessment 
Mangoral consists of manganese (II) chloride tetrahydrate, vitamin D3, and L-alanine. The 
components were evaluated by the FDA as GRAS (‘Generally Recognised As Safe’) 
substances (manganese: 21 CFR 184, 1446; vitamin D3: 21 CFR 184, 1950) or Direct Food 
Additive (L-alanine: 21 CFR 172, 320). 
Reports of manganese intoxication have been associated with occupational exposure to 
airborne manganese, e.g. in miners, smelters, or welders, or environmental exposure through 
manganese-containing pesticides. Manganese toxicity mainly affects the central nervous 
system, resulting in symptoms similar to Parkinson’s disease, but also cardiac, lung (upon 
inhalation of manganese), liver, reproductive and foetal toxicity have been observed [22].  
A prerequisite for the potential exertion of toxic effects is the distribution of manganese via the 
systemic circulation and accumulation in tissue and organs. Serum manganese has been 
suggested as a reasonable biomarker for recent exposure to manganese [23].  
Through the oral route of intake, the passage of manganese into the blood system is largely 
prevented, because manganese is very efficiently filtered out by the liver and subsequently 
excreted into the bile and finally the intestine [24]. This first-pass effect has been shown to be 
approximately 95%, leaving only trace amounts of manganese in the liver veins. Thus, 
approximately 5 % of the administered dose is absorbed by the small intestine. No correlations 
between manganese blood concentration and clinical chemistry including liver enzymes, after 
mangoral administration were observed [9]. 
The terminal plasma elimination of all manganese compounds in man is reported to be 5 to 
11 hours [25]. The half-life for the elimination of injected Mn2+ ion is reported to be 10.1 ± 
20.3 hours in healthy humans but 26.7 ± 19.0 hours in patients with hepatic impairment [26]. 
The biological half-life of manganese in the body is 37 days [27].  


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Pharmacokinetic results of previous clinical studies with mangoral showed that at the used oral 
doses of 800 mg and 1600 mg MnCl2•4H2O effects on blood manganese concentrations were 
relatively small (see investigator’s brochure [19] for PK results of previous studies). No clinically 
relevant changes between pre- and post-contrast manganese blood concentrations or out-of-
range values were observed [16, 18]. 
In these studies, both dose levels were found to be safe and not associated with serious 
adverse reactions. Overall, the frequency of AEs was higher with the higher dose level, but all 
events were of mild or moderate intensity. AEs that were assessed as being related to the 
administration of mangoral most frequently affected the gastrointestinal tract: the most 
common adverse reactions were diarrhoea (particularly at the higher dose level), nausea, and 
flatulence (refer to the investigator’s brochure [19] for a summary of current safety data). 
Together with the evaluation of clinical safety laboratory parameters, vital signs, 
electrocardiogram (ECG) results, and PK analyses, these safety data support the conclusion 
that the administration of mangoral raises no safety concerns, especially at the dose level of 
800 mg that will be used in this study. So far, clinical experience does not indicate potential 
hepatic, neurological or cardiovascular risks following the administration of the investigational 
medicinal product. 
A risk for a hepatotoxic effect of mangoral is not foreseen, due to the fast elimination half-life, 
available preclinical data in dogs (section 4.2 of the IB [19]) and the absence of any clinical or 
laboratory data in humans suggesting such a risk. For further protection of the patients 
participating in this study, patients with severe hepatic impairment, i.e. those with a Child-Pugh 
score C (exclusion criterion 6) are excluded. 
Efficacy results of the completed clinical studies demonstrate improved diagnostic quality of 
liver MRI after Mangoral enhancement and improved visualisation of liver and bowel. Individual 
patients eligible for inclusion into this study may benefit from their participation in this study in 
such a way that a more accurate diagnosis of their condition may have a positive impact on 
further treatment decisions regarding their cancer therapy. This research project may help to 
offer patients with severe renal impairment, who are in need for contrast-enhanced liver MRI, 
but cannot undergo gadolinium-enhanced MRI, a safe, efficacious, and convenient alternative 
to current contrast agents. 


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2 
STUDY OBJECTIVES AND PURPOSE 
The overall objective of this study is to evaluate the safety and diagnostic efficacy of mangoral 
in patients with known or suspected focal liver lesions and severe renal impairment. 
2.1.1 Primary objective 
• 
To assess the diagnostic efficacy of mangoral in liver MRI in terms of visualisation of 
detected focal liver lesions in combined MRI (CMRI: combined mangoral-enhanced 
and unenhanced MRI) compared to unenhanced MRI. 
Lesion visualisation will be determined by qualitative assessments of lesion border 
delineation and lesion contrast compared to background liver on 4-point scales for up 
to 15 lesions per patient. 
2.1.2 Secondary objectives 
• 
To assess the diagnostic efficacy of mangoral in liver MRI in terms of: 
o number of lesions detected by each MRI method (unenhanced MRI, 
mangoral-enhanced MRI, and CMRI) 
o visualisation of detected focal liver lesions in mangoral-enhanced MRI 
compared to unenhanced MRI (determination of visualisation will be done in the 
same way as for the primary efficacy variable) 
o confidence in lesion detection and localisation separately in unenhanced MRI, 
mangoral-enhanced MRI, and CMRI 
o lesion dimensions (independent off-site readers’ assessment): longest diameter 
of the  largest and the smallest lesion 
o quantitative assessments by measuring percent signal intensity enhancement 
of liver, liver-to-lesion contrast, signal-to-noise ratio, and contrast-to-noise ratio 
of up to 5 lesions per patient 
o number of patients who had at least one new lesion identified on CMRI 
compared to unenhanced MRI alone 
• 
To assess a proportion of patients having at least one malignant lesion identified on 
post-mangoral images that was not identified on pre-mangoral images. 
• 
To evaluate the safety and tolerability of mangoral 
• 
To evaluate the pharmacokinetics of manganese after a single dose of mangoral in a 
subgroup of patients (including a small number of dialysis patients) 
• 
To evaluate the impact of diagnostic performance of CMRI and mangoral-enhanced 
MRI versus unenhanced MRI on the patients’ management. 
 


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3 
OVERALL STUDY DESIGN AND PLAN DESCRIPTION 
This will be a multicentre, open-label, pivotal Phase III study to evaluate the safety and 
diagnostic efficacy of mangoral in patients with known or suspected focal liver lesions and 
concurrent severe renal impairment. 
Approximately 197 patients will be enrolled at about 60 investigational sites in Europe, Asia, 
USA and South America. Patients must fulfil all of the inclusion criteria and none of the 
exclusion criteria (Section 4 for inclusion and exclusion criteria). 
An overview on the schedule of assessments is provided in Section 7.1. After signing the 
informed consent, patients will undergo a screening examination within 4 weeks prior to the 
investigational medicinal product administration to verify the patients’ eligibility, to document 
patient characteristics, including medical and surgical history, and to determine the Child-Pugh 
score to assess the patients’ hepatic impairment status. A serum pregnancy test will be 
performed in women of childbearing potential at the local laboratory at screening (Section 
6.3.5.4). 
Within 24 hours prior to the administration of mangoral, patients will undergo safety 
assessments, including clinical safety laboratory tests, electrocardiogram (ECG), vital signs, 
and physical examination, and neurological assessments. A urine or serum pregnancy test will 
be performed in women of childbearing potential at the local laboratory (Section 6.3.5.4). Prior 
and concomitant therapies (taken within 14 days pre-dose) will be recorded in the electronic 
case report form (eCRF) (Section 5.2). 
On the day of contrast administration (day 0), patients will undergo an unenhanced MRI 
examination of the liver prior to the intake of mangoral (baseline MRI; unenhanced MRI can 
also be done on day -1, i.e. within 24 hours prior to contrast administration). Blood samples for 
measurement of blood manganese concentrations will be drawn immediately prior to dosing. 
Each enrolled patient will receive a single dose of orally administered mangoral after a fast of 
at least 4 hours (Section 5.1). A mangoral-enhanced liver MRI examination will be performed 
4 (± 1) hours after the mangoral administration. Patients must continue to fast until after the 
mangoral-enhanced liver MRI examination has been performed. 
Follow-up evaluations (FU 1, FU 2, FU 3) will be performed on 3 visits following contrast 
administration, at 24 (± 4) hours, 48 (± 4) hours, and 5 (± 2) days post-dose, for safety 
assessments including clinical safety laboratory tests, measurement of blood manganese, 
ECG, vital signs, physical examination, including neurological assessments. Changes in 
concomitant therapies will be recorded. Follow-up evaluations at 24 (± 4) hours and 48 (± 4) 
hours can at the discretion of the investigator be performed as remote visits when required to 
ease the burden for the patient. Participants in the PK subgroup are required to have all 
assessments including blood samples as scheduled.  
An additional follow-up visit (FU 4) for the optional brain MRI may be performed 7 (+ 2) days 
after mangoral-enhanced MRI of the liver only in those patients who underwent brain MRI due 
to any clinical reason within the last 6 months prior to mangoral-enhanced MRI and for whom 
previous brain MRI images are available for comparison (Section 6.2.4.1.2).  
In a subgroup of patients, additional blood samples (6 mL per time point) will be drawn for the 
determination of pharmacokinetic parameters on the day of the liver MRI examination (day 0) 
at 0.25, 0.5, 1, 2, 3, 4, 6, 8, and 12 hours post-dose (Section 6.4). This PK subgroup will consist 
of 12 patients, including 4 patients currently on maintenance haemodialysis. 


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Each unenhanced and each mangoral-enhanced liver MRI examination will consist of axial T1- 
and T2-weighted image sequences and a diffusion-weighted imaging (DWI) sequence. Details 
of MRI acquisition parameters are described in Section 6.2.4.1.1. 
Primary diagnostic efficacy in terms of visualisation of detected focal liver lesions will be 
evaluated centrally at an imaging core laboratory by 3 independent readers (Section 6.2.5). 
Reading will be performed in 3 parts (Part I: unenhanced MRI alone; Part II: combined MRI, 
i.e. paired reading of both unenhanced and mangoral-enhanced images; Part III: mangoral-
enhanced MRI alone). There will be a minimum 2-weeks gap between the reads to reduce 
recall bias. All independent central readers will be blinded to clinical data, site, and country 
information. 
The site investigators will also evaluate unenhanced and mangoral-enhanced MRI images (on-
site read). Their assessments will be used for the analysis of secondary variables. 
Both on-site and independent central (off-site) readers will be prospectively trained utilizing 
standardised methodology. The details of site training, image collection, anonymisation, read 
methodology and read platform will be described in an imaging manual and/or independent 
review charter (IRC) developed by a centralised imaging core laboratory. 
3.1 
Discussion of study design 
Mangoral is currently being developed for visualisation of focal liver lesions, including liver 
metastases, in patients where the use of GBCAs may be medically inadvisable or cannot be 
administered. Patients with severe renal impairment are at risk of experiencing serious adverse 
effects if they are given any of the intravenously administered contrast agents currently 
available on the market for liver MRI enhancement. NSF is a rare, but potentially fatal, 
complication of GBCAs that has been reported in patients with renal failure [21]. 
Previous clinical Phase I and II studies included healthy volunteers and patients with liver 
metastases. This Phase III study will be performed to assess the safety and diagnostic efficacy 
of mangoral in a patient cohort with severe renal impairment (eGFR < 30 mL/min/1.73 m2, or 
increase in serum creatinine ≥ 0.3 mg/dL within 48 hours or ≥ 50% within 7 days) and who are 
being evaluated for known or suspected focal liver lesions. In addition, as patients with severe 
renal impairment are likely to need dialysis, patients on haemodialysis or peritoneal dialysis 
will be included in this study to obtain information on safety, pharmacokinetics, and diagnostic 
efficacy of mangoral in such patients. 
A multicentre and multinational design is used to increase representativeness and to reduce 
bias. 
Results of a previous clinical Phase II study demonstrated that signal intensity enhancement 
increased in liver parenchyma until 3 hours post-dose and there was no significant difference 
in the liver-to-lesion contrast between 3 and 6 hours after mangoral administration [16]. Thus, 
the primary time point for mangoral-enhanced liver MRI chosen in this study is 4 hours after 
intake with a time window of ± 1 hour.  


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4 
SELECTION OF STUDY POPULATION 
Approximately 197 patients with severe renal impairment (Chronic Kidney Disease [CKD] or 
Acute Kidney Injury [AKI]) who are being evaluated for known or suspected focal liver lesions 
will be enrolled. Patients will be recruited among in- and outpatients at approximately 60 
radiological sites. 
The proportion enrolled HCC patients will be monitored jointly by the CRO/sponsor in order to 
achieve approximately 20% HCC patients in the overall study population. The CRO/sponsor 
may instruct individual sites, from time to time, to specifically and exclusively enrol patients 
with either non-HCC or HCC. 
It is intended that potential patients for this study will be identified from a group of patients 
currently attending, or being referred to the study site for the diagnosis and treatment of known 
or suspected focal liver lesions. Potentially eligible patients will be approached by the 
investigator to ascertain whether they would be interested in participating in the study. 
Competitive patient recruitment between countries and sites will be used during the entire 
recruitment period to ensure that the required number of patients are enrolled within the 
planned recruitment period. 
4.1 
Inclusion criteria 
For inclusion in this study, the patients must fulfil all of the following criteria: 
1. Patients who have been fully informed and have personally signed and dated the 
informed consent 
2. Male and female patients 18 years and older 
3. Patients who have known or suspected focal liver lesions based on medical history and 
previous laboratory and/or imaging examinations 
4. Patients with severe renal impairment presented as: 
a) Chronic kidney disease [CKD] (estimated glomerular filtration rate [eGFR] 
< 30 mL/min/1.73 m2) based on medical history and previous laboratory 
examinations, at least once, within the last 3 months prior to the Baseline Visit 
OR  
b) Acute kidney injury [AKI]) with an increase in serum creatinine ≥ 0.3 mg/dL within 
48 hours or ≥ 50% within 7 days, prior to the Baseline Visit 
5. Female patients who are not of childbearing potential, or female patients who are of 
childbearing potential but are using highly effective contraception (including hormonal 
contraceptives, such as combined oral contraceptives, patch, vaginal ring, injectables, 
and 
implants, 
intrauterine 
device, 
intrauterine 
hormone-releasing 
system, 
vasectomised partner, and tubal ligation) or practising sexual abstinence (only 
accepted if this is the usual and preferred lifestyle of the patient), and have a negative 
urine or serum pregnancy test within 24 hours prior to administration of contrast agent 
6. Sexually active male patients (who are not vasectomised) must be willing to use 
condoms after administration of mangoral until the last follow-up visit 
7. Patients who agree to comply with all study procedures as outlined within the study 
protocol and informed consent 


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8. Not applicable 
4.2 
Exclusion criteria 
Patients will not be included in the study if any of the following criteria are met: 
1. Patients with simple liver cysts only 
2. Patients who have received any investigational drug or were treated with an 
investigational device within 6 weeks prior to the Baseline Visit 
3. Patients who have received any MRI contrast media within 6 weeks prior to the 
Baseline Visit or are scheduled to receive any other contrast medium other than 
mangoral before last study visit 
4. Patients who have previously been enrolled in the study and underwent 
mangoral-enhanced MRI 
5. Patients who are clinically unstable or have an acute illness, as judged by the 
investigator; i.e. the underlying clinical condition is such that clinically relevant changes 
in condition can be reasonably anticipated during the study period 
6. Patients with severe hepatic impairment (according to Child-Pugh score C) 
7. Not applicable 
8. Patients scheduled for surgery before the last study visit 
9. Pregnant or lactating women 
10. Patients with a history of severe allergies (e.g. anaphylaxis), as judged by the 
investigator. 
11. Patients with a known hypersensitivity to the IMP or any excipients in the IMP 
12. Patients who are, in the investigator’s opinion, unable to comply with the study 
requirements including follow-up procedures 
13. Patients with known portosystemic shunts 
14. Patients with encephalopathy, as judged by the investigator 
15. Patients with neurodegenerative disorders 
16. Patients with acute neurological disorders, as judged by the investigator 
17. Patients with haemochromatosis  
18. Patients receiving blood transfusion within 14 days prior to the Baseline Visit 
19. Patients who cannot fast for 8 hours 
20. Patients who cannot undergo MRI due to MRI incompatible pacemaker, metal implants, 
or other MRI contraindications 
21. Patients with known inherited disorders of manganese metabolism 
22. Patients with known pheochromocytoma 
23. Patients with a history of surgical/other interventional procedures or conditions which 
may interfere with the absorption or excretion of the IMP (e.g. small bowel obstruction, 
gastric bypass, gastric resection, biliary obstruction) as judged by the investigator 


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24. Patients who have been institutionalised by official or court order and therefore are not 
completely free in their decision-making process 
25. Patients dependent on the sponsor, investigators, personnel of the trial site or any other 
individual, whose willingness to volunteer may be unduly influenced by the expectation, 
whether justified or not, of benefits associated with participation 
4.3 
Withdrawal criteria 
Premature study termination for an individual patient should be considered under the following 
circumstances: 
• 
The patient has an AE that may jeopardise the patient’s safety (e.g. allergic reaction to 
IMP) 
• 
Non-compliance (e.g. break of required fast before the examination) 
• 
Lost to follow-up (the patient did not show up for the examination and study personnel 
were unable to contact the patient) 
Decisions concerning the premature study termination of individual patients will be made in 
agreement with the sponsor. 
The trial must be terminated for an individual patient if the patient withdraws his / her consent. 
Patients may decide to withdraw their consent to participate at any time and for any reason 
without prejudice to their further medical care. The investigator should make a reasonable 
effort to ascertain the reason(s) for the patient’s withdrawal while fully respecting the patient’s 
rights.  
The reason(s) for premature termination are to be recorded on the eCRF. For all patients who 
are prematurely withdrawn, all observations and test results available at the time of withdrawal 
are to be recorded in the eCRF. 
Patients who terminate the study prematurely will not be replaced. 
For criteria concerning the termination of the whole study please refer to Section 7.4.2. 


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5 
TRIAL TREATMENTS 
5.1 
Investigational medicinal product(s) 
5.1.1 Administration 
The Investigational medicinal product is mangoral combined with L-alanine and vitamin D3. 
Mangoral is intended for diagnostic use and single-dose administration only. 
All patients will receive a single dose of IMP (equivalent to 800 mg MnCl2•4H2O) orally after a 
fast of at least 4 hours. Patients must continue to fast until after the mangoral-enhanced liver 
MRI examination has been performed (i.e. 4 [± 1] hours after IMP administration). Patients are 
allowed to drink water during the fasting period. 
Note: please inform and consult with the sponsor or designee in case administered IMP has 
not been fully absorbed by the patient (i.e. if the patient vomited after IMP intake and prior to 
the MRI examination). 
The IMP is dispensed in two stick packs, one containing the active pharmaceutical ingredient 
and the other containing the two absorption promoters. The contents in both stick packs 
(Section 5.1.2) should be dissolved together in 200 mL of water, preferably cold, and ingested 
as an oral solution within 2 hours after dissolution. The patient should drink the solution slowly 
within 10 minutes. 
5.1.2 Identity of the investigational medicinal product 
  
 
 
 
 
 
 physio-chemical properties of the active ingredient MnCl2•4H2O are 
summarised in 
 Table 3
. 
 
 


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Table 3 
Physio-chemical properties of manganese (II) chloride tetrahydrate 
Solubility: 
Soluble in water (1980 g/L [20°C]) and alcohol 
pH: 
A 5% solution has a pH between 3.5 and 6.0 
Melting point: 
+58°C 
Density: 
2.01 g/cm3 
Hygroscopicity: 
Hygroscopic 
 
5.1.3 Method of assigning patients to treatment groups 
Not applicable; all patients will receive the same dose the IMP. 
5.1.4 Selection of doses in the study 
Results from a previous Phase I study have shown that relative signal intensity (SI) of the liver 
on T1-weighted images was significantly increased at both dose levels of 800 mg and 1600 mg 
MnCl2•4H2O [14]. Similar relative SI enhancement after administration of the 800 mg or 
1600 mg dose was seen in a Phase II study in patients with liver metastases [16]. Another 
Phase II study showed that a dose of 800 mg MnCl2•4H2O resulted in a significantly higher 
increase in liver-to-muscle SI and a superior overall image quality compared to lower doses of 
400 mg and 200 mg MnCl2•4H2O, respectively [18].  
Safety experiences in previous clinical Phase I and Phase II studies showed that the number 
of AEs, especially of gastrointestinal tract reactions, tended to be higher in the 1600 mg dose 
group than in the 800 mg dose group. Thus, based on the results of the dose comparisons and 
the observed better tolerability of 800 mg versus 1600 mg MnCl2•4H2O, the 800 mg dose was 
chosen for this pivotal Phase III study. 


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5.1.5 Selection and timing of dose for each patient  
The IMP will be administered 4 (± 1) hours prior to mangoral-enhanced MRI of the liver. The 
timing of dose was chosen based on the results of a previous Phase II study showing that the 
optimal diagnostic time window with mangoral is between 2 and 6 hours [16]. 
Patients must have been fasting for at least 4 hours prior to the administration of IMP. 
5.1.6 Supply, manufacturing, packaging and labelling 
The IMP stick packs for oral administration will be provided under the responsibility of the 
sponsor. 
Each study site will be supplied with two stick packs for each patient (one stick pack 1 and one 
stick pack 2) for use within this study to match the expected number of patients to be enrolled 
at a given site. 
The IMP will be manufactured, packed, labelled, and released (by a Qualified Person) and 
distributed under the responsibility of the sponsor in accordance with the principle of Good 
Manufacturing Practice (GMP) and the applicable national and/or local regulatory 
requirements. The labels will be available in local languages. 
5.1.7 Storage 
The IMP (stick pack 1 and stick pack 2) is to be kept at the study site at room temperature 
(15 – 25°C) in a place with authorised access only. 
5.1.8 Shipment 
The IMP will be shipped according to local regulations. The responsible investigator / 
responsible study personnel at the study site will confirm correct receipt of the IMP in writing 
and ensure it is stored safely and correctly. The investigator will document the distribution date 
and the amount used on the forms provided for this purpose. 
5.1.9 Blinding and randomisation 
Not applicable as this is an open-label study with regard to the administration of IMP.  
5.1.10 Investigational medicinal product compliance and accountability 
The oral solution of the IMP, containing the content of both stick packs, will be prepared by the 
investigator or by study personnel assigned by the investigator at the study site and handed to 
the patient. Date and time of IMP administration will be recorded in the eCRF.  
The investigator will ensure that the IMP provided will be used only within the framework of 
this study and as directed in the study protocol.  
IMP accountability must be recorded at each site and will be checked by a study monitor. Upon 
the completion of the study, all IMPs will be reconciled and destroyed locally or returned to 
sponsor or designee according to local regulations and site standard operating procedures. 
Receipt, distribution and destruction of the IMP must be recorded according to the study-
specific instructions provided separately. 
5.1.11 Management of investigational medicinal product overdose 
Accidental overdosing of IMP is unlikely as the oral solution will be prepared by qualified 
personnel at the study sites. In case of accidental overdosing, the patient should be closely 


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monitored and treated, if necessary, at the discretion of the investigator. A dose of 1600 mg 
MnCl2•4H2O, which is double the dose used in this trial, has been well-tolerated in previous 
clinical Phase I and II studies. Cases of overdose must be reported as adverse events (Section 
6.3.7). If an overdose is associated with a serious adverse reaction, this reaction is subject to 
expedited reporting, i.e. within 24 hours (Section 6.3.7.4). 
5.2 
Prior and concomitant therapy 
Any medication taken within 14 days prior to the administration of IMP will be recorded in the 
eCRF, regardless of whether the treatment will be continued during the study. Any changes in 
concomitant medication or the addition of a new treatment during the study will be recorded. 
This also includes any dietary supplements and hormonal contraception. The following details 
must be given: nature of the disease (indication), name of the medicine (including active 
ingredients) or specification of measures, dosage, unit, route of administration, and the start 
and end dates of treatment. 
5.2.1 Prohibited or prohibited with restrictions 
Several dietary components, such as iron [28], magnesium, calcium, vitamin C, and vitamin 
D3 [20], and medications, such as tetracycline or tetracycline derivatives [20], may negatively 
or positively influence the absorption of manganese from the intestine. Therefore, the use of 
the following substances is prohibited during this study from 6 hours prior to IMP administration 
until after the mangoral-enhanced MRI: 
• 
Dietary iron supplementation, including multivitamin that contains iron, may interfere 
with manganese absorption through competition for the same transporters 
• 
Supplemental magnesium and/or supplemental calcium may reduce manganese 
absorption 
• 
Supplemental vitamin C or vitamin D3, which promote manganese absorption 
• 
Magnesium-containing antacids and laxatives 
• 
Quinolone and tetracycline antibiotics 
5.2.2 Permitted oral intake 
The use of following is allowed: 
• 
Glucose or juice intake (e.g. orange juice) for the purpose to prevent or treat 
hypoglycaemia in patients with diabetes mellitus type I or II 
6 
VARIABLES AND METHODS 
6.1 
Population Characteristics 
6.1.1 Demographics 
The following demographic characteristics will be recorded in the eCRF: 
• 
Year of birth 
• 
Sex 


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• 
Ethnicity and race (only if the collection of such data is permitted by local regulations) 
• 
Weight and height 
6.1.2 Medical history 
6.1.2.1 
General medical history 
General medical history findings considered relevant to the study by the investigator will be 
recorded in the eCRF together with the respective dates and periods (whether the condition is 
a past disease/illness or still ongoing at study entry). Relevant findings include, but may not be 
limited to, major surgeries, heart diseases, respiratory diseases, central nervous system and 
neurological diseases, psychiatric disorders, blood disorders, hepatorenal disorders, 
genitourinary disorders, and known allergies.  
In addition, it will be recorded whether or not the patient underwent recent brain MRI. If yes, 
the date and the clinical reason for the examination will be recorded. In case that at the day of 
the mangoral-enhanced liver MRI the date of the brain MRI lays within the last 6 months and 
previous brain MRI images are available, the patient may consent to undergo a brain MRI 7 
(+ 2) days after the mangoral-enhanced liver MRI. 
6.1.2.2 
Referral diagnosis 
Details of the underlying condition related to the evaluation of known or suspected focal liver 
lesions will be recorded in the eCRF, including: 
• 
Diagnosis 
• 
Diagnosis of underlying primary malignancy, if applicable 
• 
Date of first diagnosis 
• 
Presence of metastases (body region) 
• 
Clinical signs and symptoms 
6.1.2.3 
Child-Pugh score 
The Child-Pugh score [29, 30, 31, 32] will be determined during the screening period to assess 
hepatic impairment of the patient.  
The score comprises 5 measures of liver disease, which will each be scored 1, 2, or 3, 
depending on the severity as shown in Table 4. The sum of the scores provides the Child-
Pugh score (class A, B, or C). 
 




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6.2 
Diagnostic efficacy 
6.2.1 Definition of liver segments 
Independent readers and on-site radiologists will assess focal liver lesions on images from 
unenhanced and mangoral-enhanced MRI in 8 anatomical segments of the liver according to 
the Couinaud classification [34, 35], which divides the liver into the following 8 segments 
(Figure 1):  
Segment I: 
caudate lobe, situated posteriorly around the inferior vena cava 
Segment II: 
left lateral superior segment, situated to the left of the left hepatic vein and 
falciform ligament superior to the portal plane 
Segment III: 
left lateral inferior segment, situated to the left of the left hepatic vein and 
falciform ligament inferior to the portal plane 
Segment IV: 
left medial segment, situated between the left and middle hepatic veins, 
subdivided into IVa (superior) and IVb (inferior) subsegments 
Segment V: 
right anterior inferior segment, situated below the portal plane between 
the middle and right hepatic veins 
Segment VI: 
right posterior inferior segment, situated below the portal plane to the right 
of the right hepatic vein 
Segment VII: 
right posterior superior segment, situated above the portal plane to the 
right of the right hepatic vein 
Segment VIII: 
right anterior superior segment, situated above the portal plane between 
the middle and right hepatic veins 
 
Figure 1 
Schematic representation of liver segments according to Couinaud 
IVC = inferior vena cava (image taken from: https://clinicalgate.com/liver-3/) 
 
 


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6.2.2 Primary and secondary efficacy variables 
6.2.2.1 
Primary efficacy variable 
The primary efficacy endpoint will be the visualisation of detected focal liver lesions in 
combined MRI (CMRI, mangoral-enhanced MRI plus unenhanced MRI) as compared to 
unenhanced MRI.  
Visualisation will be measured by two co-primary variables: 
• 
lesion border delineation 
• 
lesion contrast compared to liver background 
Both parameters will be determined for each lesion (up to 15 lesions per patient) and by each 
of the three independent readers by qualitative assessment on 4-point scales with categories 
‘poor’ (1), ‘partial/moderate’ (2), ‘good’ (3), and ‘excellent’ (4) lesion border delineation/lesion 
contrast (Section 6.2.3.1). Based on the scores on the 4-point scales, two sum scores will be 
calculated for each patient and separately for each MRI method: a lesion border delineation 
sum score and a lesion contrast sum score. Both scores will be weighted by the number of 
lesions per patient. Only lesions detected at baseline (unenhanced MRI) will be included in the 
assessment of diagnostic efficacy. 
Superiority of CMRI over unenhanced MRI regarding visualisation of focal liver lesions will be 
analysed by testing the superiority of each of the two co-primary variables separately for each 
of the three independent readers. Reader success will be achieved if the reading results of a 
reader demonstrate superiority of CMRI versus unenhanced MRI for both lesion border 
delineation and lesion contrast. For the success of the trial, two of three readers must achieve 
reader success. 
6.2.2.2 
Secondary efficacy variables 
The secondary efficacy variables will be based on the evaluations by independent readers 
(off-site readers’ assessment) and by the on-site radiologist (on-site readers’ assessment): 
• 
Number 
of 
lesions 
detected 
by 
each 
MRI 
method: 
unenhanced 
MRI, 
mangoral-enhanced MRI, and CMRI (on-site and independent off-site readers’ 
assessments) 
• 
Visualisation of focal liver lesions in mangoral-enhanced MRI as compared to 
unenhanced MRI (independent off-site readers’ assessment; visualisation will be 
assessed as for the primary efficacy endpoint) 
• 
Confidence in lesion detection separately in unenhanced MRI, mangoral-enhanced 
MRI, and CMRI (3-point scale, on-site and independent off-site readers’ assessments 
of up to 15 lesions per patient) 
• 
Confidence in lesion localisation separately in unenhanced MRI, mangoral-enhanced 
MRI, and CMRI (3-point scale, independent on-site and off-site readers’ assessments 
of up to 15 lesions per patient) 
• 
Lesion dimensions (independent off-site readers’ assessment): longest diameter of the 
largest and the smallest lesion 
• 
Quantitative assessments will be performed for up to 5 detected lesions (independent 
off-site readers’ assessment). These lesions will be the same for pre- and post-contrast 
assessments: 
o liver signal intensity (SI) enhancement (%) 


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o liver-to-lesion contrast (LLC) 
o signal-to-noise ratio (SNR) 
o contrast-to-noise ratio (CNR) 
• 
Change(s) in patients’ management based on the diagnostic performance of CMRI or 
mangoral-enhanced MRI versus unenhanced MRI (on-site and independent off-site 
readers’ assessments): 
o Any changes in patient management based on MRI findings (yes/no); 
o Next steps in patient management based on MRI findings (i.e. chemotherapy, 
surgery, local ablation procedure, combination therapy, or other [specify]). 
An overview of the primary and secondary efficacy variables by off- and/or on-site assessment 
of MRI images) is provided in Table 5. For details on deriving the efficacy variables 
Section 6.2.3. 
 




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Lesion border delineation:  
1 = 
Poor: 
lesion border is poorly distinct, the lesion cannot be separated from 
surrounding tissues or structures 
2 = 
Partial: 
delineation of the lesion border is fair, but not complete; the lesion cannot 
be clearly separated from surrounding tissues or structures 
3 = 
Good: 
delineation of the lesion border is complete; the lesion is adequately 
separated from surrounding tissues or structures 
4 = 
Excellent: 
lesion border is sharply and clearly distinct, the lesion is sharply separated 
from surrounding tissues or structures 
Lesion contrast:  
1 = 
Poor: 
the difference in signal intensity between the lesion and the surrounding 
normal liver tissue is poor; the lesion can barely be identified and it is not 
possible to evaluate and measure the size (maximum diameter) of the 
lesion 
2 = 
Partial: 
the difference in signal intensity between the lesion and the surrounding 
normal liver tissue is fair; the lesion can be identified, but it is not possible 
to evaluate and measure the size (maximum diameter) of the lesion 
3 = 
Good: 
the difference in signal intensity between the lesion and the surrounding 
normal liver tissue is adequate; the lesion can be identified and its size 
(maximum diameter) can be evaluated and measured 
4 = 
Excellent: 
the difference in signal intensity between the lesion and the surrounding 
liver is marked; the lesion can be optimally identified and its size 
(maximum diameter) can be easily measured 
 
Assessments will be done by the three independent readers during central reading sessions 
(Section 6.2.5): part I (unenhanced MRI alone) and part II (paired reading of both unenhanced 
and mangoral-enhanced images) and part III (mangoral-enhanced images alone). 
For this study, a specific MRI manual will be prepared for training the investigators on MRI 
acquisition parameters, and local reading procedures and rules. An Independent Review 
Charter will be prepared for the independent reading process. The independent readers will 
be trained on MRI assessments and the training will be documented.  
The handling of MRI scans with more than 15 lesions will be described in these documents. 
Briefly, the readers will identify the smallest lesion and the largest lesion. If there are more than 
15 lesions, the reader will start counting starting from the smallest lesion to the largest lesion 
and will stop at 15 lesions. 
6.2.3.2 
Confidence in lesion detection 
Confidence in lesion detection will be evaluated for each lesion during the central reading 
sessions (Section 6.2.5), i.e. part I (unenhanced MRI alone), part II (CMRI), and part III 
(mangoral-enhanced 
MRI 
alone), 
by 
the 
three 
independent 
readers, 
and 
on 
mangoral-enhanced MRI and unenhanced MRI images by the on-site radiologists. Up to 15 
lesions per patient will be evaluated on the following 3-point scale: 
1 = 
The lesion is detected with low confidence 
2 = 
The lesion is detected with moderate confidence 
3 = 
The lesion is detected with high confidence 


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6.2.3.3 
Confidence in lesion localisation 
Confidence in lesion localisation will be assessed for each lesion during the central reading 
sessions (Section 6.2.5), i.e. part I (unenhanced MRI alone), part II (CMRI), and part III 
(mangoral-enhanced 
MRI 
alone), 
by 
the 
three 
independent 
readers, 
and 
on 
mangoral-enhanced MRI and unenhanced MRI images by the on-site radiologists using the 
Couinaud liver segment classification system [34]. Up to 15 lesions per patient will be 
evaluated on the following 3-point scale: 
1 = 
The lesion is localised to a liver segment with low confidence 
2 = 
The lesion is localised to a liver segment with moderate confidence 
3 = 
The lesion is localised to a liver segment with high confidence 
Note: If a lesion is large enough to be present in more than one segment it shall be 
considered to be in the segment in which its site lies. 
6.2.3.4 
Lesion dimensions 
Lesion dimension measurements will be done by each of the three independent readers on 
mangoral-enhanced MRI and unenhanced MRI images. The longest diameter of the largest 
and of the smallest visualised lesions, respectively, will be measured.  
6.2.3.5 
Quantitative assessments 
For the quantitative analysis, the signal intensities (SI) of liver parenchyma and liver lesion, as 
well as the standard deviation (SD) of the background noise will be determined by each of the 
three independent readers during central reading session part II (paired reading of both 
unenhanced and mangoral-enhanced images; Section 6.2.5). The assessment will be done 
only during reading session part II as this allows the readers to assess the same lesions on 
unenhanced and enhanced images in the absence of formal lesion matching. Up to 5 lesions 
per patient of ≥ 2 cm in diameter will be evaluated and these lesions will be the same on 
pre-and post-contrast images.  
Quantitative SI will be measured by positioning circular regions of interest (ROIs) in a 
homogenous area in the liver and the assessed liver lesion on the same image. The ROI 
placed in the lesion should at least encompass half of the lesion. ROIs of a constant size will 
be placed in the same locations on all pre- and post-contrast images. SD of the background 
noise will be measured using the largest possible rectangular ROI vertical to the patient's 
abdomen in the direction of the phase-encoding gradient. 
The following quantitative measures will be determined: 
Signal intensity (SI) enhancement: 
• 
Liver SI enhancement (%) =  
([SIliv post contrast – SIliv pre contrast]/[SIliv pre contrast]) × 100 
Liver-to-lesion contrast (LLC): 
• 
LLC = (SIliv – SIles)/(SIliv + SIles) 
Signal-to-noise ratio (SNR): 
• 
SNR = SIliv/SDnoise 


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Contrast-to-noise ratio (CNR): 
• 
CNR = (SIliv – SIles)/SDnoise 
with SIliv = signal intensity of the liver, SIles = signal intensity of the lesion, SDnoise = standard 
deviation of the background noise. 
6.2.4 Imaging procedures  
6.2.4.1 
Image acquisition 
Details of imaging procedures and acquisition will be provided to each study site in a 
study-specific imaging manual (imaging site manual).  
6.2.4.1.1 
MRI examination of the liver 
Unenhanced MRI of the liver will be performed during the baseline period, i.e. either on the 
day prior to the mangoral-enhanced MRI or pre-dose on the same day as the 
mangoral-enhanced MRI. Mangoral-enhanced MRI of the liver will be performed 4 (± 1) hours 
after mangoral administration. 
MRI examinations will be done using a 1.5 Tesla or 3.0 Tesla MRI scanner. An overview of 
image sequences utilised for both pre- and post-contrast examinations and MRI parameters is 
given in Table 6. All relevant details of MRI methodology will also be given in the imaging site 
manual. 
6.2.4.1.2 
MRI examination of the brain 
Unenhanced MRI of the brain will be performed 7 (+ 2) days after mangoral-enhanced MRI of 
the liver in a subgroup of patients who underwent brain MRI due to any clinical reason within 
the last 6 months prior to mangoral-enhanced MRI and for whom previous brain MRI images 
are available for comparison.  
MRI examination should be done with the same parameters used in the brain MRI images 
performed before participating in this study. If parameters from previous brain MRI are 
unknown, standard settings can be used. 
MRI scans will be evaluated by the off-site imaging core laboratory. 
 


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Table 6 
MRI parameters 
General Instructions 
Scanner type 
1.5 Tesla or 3.0 Tesla 
Patient Orientation 
Supine 
Breathing instructions 
Breath-hold technique or ultra-fast acquisition 
Sequences 
(both pre- and post-contrast) 
• 
T1-gradient echo (T1-GRE),  
• 
T1-gradient echo with fat suppression 
(T1-GRE-Fat Sat); 
• 
T2-fast spin echo without fat suppression 
(T2-SS-FSE, HASTE),  
• 
Optional: T2- fast spin echo with fat suppression 
(T2-RTR-FSE) 
• 
Diffusion-weighted imaging (DWI). 
Sequence parameters 
 
T1-GRE 
T1-GRE Fat Sat 
T2-SS-FSE, 
HASTE 
T2-RTR-FSE 
(optional) 
DWI 
Scanner 
(Tesla) 
1.5 
3.0 
1.5 
3.0 
1.5 
3.0 
1.5 
3.0 
1.5 
3.0 
Plain 
Axial 
Axial 
Axial 
Axial 
Axial 
Axial 
Axial 
Axial 
Axial 
Axial 
Dimension 
2D or 3D 
2D or 3D 
2D or 
3D 
2D or 3D 
2D 
2D 
2D 
2D 
2D 
2D 
TR (msec) 
2D: 100-
200 
3D: 3-10 
2D: 100-
200 
3D:3 -10 
2D: 100-
200 
3D: 3-10 
2D: 100-
200 
3D: 3-10 
800-
2500 
800-
2500 
R-R 
inter-val 
R-R 
inter-val 
1000-
5400 
(min.) 
1000-
5400 
(min.) 
TE (msec) 
1.5-3.0 
& 4.0-
6.0 
1.0-1.5 
& 2.0-
3.0 
1.5-3.0 
& 4.0-
6.0 
1.0-1.5 
& 2.0-
3.0 
80-200 
80-200 
70-200 
70-200 
Minimum 
Minimum 
Flip 
angle 
(°) 
2D: 
60-80 
3D: 
10-15 
2D: 
60-80 
3D: 
10-15 
2D: 
60-80 
3D: 
10-15 
2D: 
60-80 
3D: 
10-15 
90 
90 
90 
90 
90 
90 
NEX 
0.6-1 
0.6-1 
1 
1 
0.5-1 
0.5-1 
2-4 
2-4 
2-6 
2-6 
Matrix 
(frequency) 
250-350 
250-350 
250-350 
250-350 
250-350 
250-350 
250-350 
250-350 
80-192 
80-192 
Matrix 
(phase) 
180-256 
180-256 
180-256 
180-256 
180-256 
180-256 
180-256 
180-256 
80-192 
80-192 
Phase 
direction 
AP 
AP 
AP 
AP 
AP 
AP 
AP 
AP 
AP 
AP 
Slice 
thickness 
(mm) 
2D: 
4-6 
(true) 
3D: 3-5 
(true) 
2D: 
4-6 
(true) 
3D: 3-5 
(true) 
2D: 
4-6 
(true) 
3D: 3-5 
(true) 
2D: 
4-6 
(true) 
3D: 3-5 
(true) 
4-8 
4-8 
4-8 
4-8 
4-8 
4-8 
Slice 
gap 
(mm) 
2D: 
0-2 
3D: 
0 
2D: 
0-2 
3D: 
0 
2D: 
0-2 
3D: 
0 
2D: 
0-2 
3D: 
0 
0-3 
0-3 
0-3 
0-3 
0-3 
0-3 
B-values 
- 
- 
- 
- 
- 
- 
- 
- 
0-100, 
500-800 
0-100, 
500-800 
AP = anterior to posterior, min. = minimum, NEX = number of excitations, TE = echo time, TR = repetition time 


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6.2.4.2 
Image handling and processing 
Independent off-site assessments will be performed at a centralised imaging core laboratory 
(lab) in a digital fashion using a validated software. The independent readers will be blinded 
with regard to patient identity and all clinical information of a patient.  
Each site will transfer all image data acquired to the imaging core laboratory. All study-related 
image data have to be submitted in DICOM (Digital Imaging and Communications in Medicine) 
format. In accordance with Good Clinical Practice (GCP) guidelines, investigational sites are 
required to maintain the original images as source documentation of the study. 
The imaging core laboratory will check and document image quality and completeness of 
image sets prior to the independent reading sessions. All independent read operational details 
will be described in the IRC developed by the core lab. 
6.2.5 Efficacy assessments 
6.2.5.1 
Off-site (central) assessments 
All unenhanced and mangoral-enhanced MRI image sets will be evaluated centrally at the core 
imaging laboratory by 3 independent readers.  
The central evaluation of all MRI images will consist of 3 parts: 
• 
Part I: 
unenhanced MRI alone 
• 
Part II: combined 
MRI, 
i.e. 
paired 
reading 
of 
both 
unenhanced 
and 
mangoral-enhanced images 
• 
Part III: mangoral-enhanced MRI alone 
Each of the three independent readers will assess all images of all patients during each reading 
part. There will be a 2-week gap between reading parts to reduce recall bias. 
The analysis of the primary efficacy variable (Section 6.2.2.1) will be based on the qualitative 
assessments performed centrally. 
Lesion tracking and lesion matching  
At the time of efficacy reads, the three independent readers will identify the location of the 
detected lesions in liver segments. A fourth reader who is not involved in efficacy reads will 
track and match the detected lesions on unenhanced, mangoral-enhanced and combined 
(unenhanced plus mangoral-enhanced MRI) to confirm lesion numbering and lesion location 
across the modalities. Lesion tracking will also confirm lesions that are identified on 
pre-mangoral, post-mangoral and combined (unenhanced plus mangoral-enhanced) MRI 
images. 
6.2.5.2 
On-site assessment 
On-site evaluation will be done by each site investigator (or on-site appointed radiologist) within 
5 working days after the MRI examination (Table 5).  
6.3 
Safety 
Section 7.1, study schedule provides details on assessments per visit, if a visit is conducted 
as a remote visit (see Sections 7.2.4 and 7.2.5 as applicable).  


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6.3.1 Vital signs 
Vital signs will be measured at baseline within 3 hours prior to contrast administration, and 
after contrast administration at 6 (± 1) hours, 24 (± 4) hours, 48 (± 4), and 5 (± 2) days post-
dose: 
• 
Systolic blood pressure (mmHg) 
• 
Diastolic blood pressure (mmHg) 
• 
Pulse rate (bpm) 
• 
Respiratory rate (breaths per minute) 
• 
Body temperature (°C) 
Patients must be sitting, and measurements must be started after at least 4 minutes of rest. 
The investigator will evaluate any worsening in vital signs. Any new or worsening events 
(irrespective of clinical significance) observed after mangoral administration compared to 
previous pre-dose assessments must be reported as an AE. 
6.3.2 Physical examination 
A physical examination will be performed at the following times: baseline (within 3 hours prior 
to contrast administration), and at 6 (± 1) hours, 24 (± 4) hours, 48 (± 4) hours, and 5 (± 2) 
days post-dose. Any new or worsening observed changes (irrespective of clinical significance) 
after administration of mangoral compared to previous pre-dose assessments must be 
reported as an AE (Section 6.3.7). 
6.3.3 Neurological assessments 
Neurological assessments to monitor for any potential neurotoxic effects will be performed pre-
dose during the baseline period and 24 (± 4) hours, 48 (± 4) hours, and 5 (± 2) days hours after 
the administration of mangoral. At each site, a trained examiner (e.g. nurse practitioner, 
physician) will be appointed for the neurological assessments after receiving appropriate 
training (as applicable). 
Any new or worsening observed changes (irrespective of clinical significance) after mangoral 
administration compared to previous pre-dose assessments must be reported as an AE 
(Section 6.3.7). 
6.3.4 Electrocardiogram (ECG) 
A 12-lead ECG will be performed after 5 minutes supine rest at baseline (within 3 hours prior 
to contrast administration) and after administration of mangoral at 6 (± 1) and 24 (± 4) hours 
post-dose.  
Standard ECG parameters will be recorded and assessed (including corrected QT interval, RR 
interval, PR interval, and QRS interval). 
The investigator will record on the eCRF whether the results are normal, abnormal (not 
clinically significant or clinically significant). If recorded as abnormal and clinically significant, 
the abnormality must be reported in the eCRF as an AE (Section 6.3.7).  


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6.3.5 Laboratory variables 
6.3.5.1 
Screening laboratory tests 
Table 7 provides an overview of the laboratory parameters to be measured at the local 
laboratory during the screening period for the determination of the Child-Pugh score. 
Table 7 
Screening laboratory parameters 
Child-Pugh: 
(local lab) 
Serum bilirubin 
Serum albumin 
International normalised ratio (INR) 
eGFR: 
(local lab) 
Serum creatinine 
Serum albumin 
Serum urea nitrogen 
(SUN/BUN) 
Note: eGFR measurement/assessment is only required if not available within the last 3 months 
prior to the Baseline Visit. 
6.3.5.2 
Clinical safety laboratory tests  
Blood samples for haematology and biochemistry tests as well as a urine sample for urinalysis 
for safety monitoring will be collected at baseline (within 24 hours prior to contrast 
administration), and 2 (± 0.5) hours, 6 (± 1) hours, 24 (± 4) hours, 48 (± 4) hours, and 5 (± 2) 
days after administration of mangoral. A blood volume of approximately 12 mL will be drawn 
per time point. An overview of measured parameters is given in Table 8. 
 


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Table 8 
Clinical safety laboratory parameters (baseline and post-dose) 
Haematology: 
(central lab) 
Haemoglobin 
Haematocrit 
Erythrocytes 
Leucocytes 
Thrombocytes 
Neutrophils 
Eosinophils, 
Basophils 
Lymphocytes 
Monocytes 
INR  
 
Biochemistry: 
(central lab) 
Alkaline phosphatase 
Aspartate aminotransferase (ASAT) 
Alanine aminotransferase (ALAT) 
Gamma-glutamyl transferase (γ-GT) 
Creatinine 
Glucose 
Urea 
Total Bilirubin 
Albumin 
Sodium 
Potassium 
Calcium 
Magnesium 
Chloride 
Urinalysis: 
(local lab) 
 
pH 
Leucocytes 
Blood 
Protein 
Glucose 
Urobilinogen 
Ketones 
Bilirubin 
Nitrite 
Specific density 
 
All blood tests (haematology, biochemistry) will be analysed at a central laboratory. Special 
tubes, labels, packaging, and sample handling instructions will be provided by the central 
laboratory. Blood sampling and handling procedures are described in the study-specific 
laboratory manual. 
Urinalysis will be performed by using sites’ standard of care dipsticks or using dipsticks 
provided by the central laboratory. 
The investigator must document the clinical significance of abnormal laboratory values in the 
eCRF. If the clinically significant abnormal value is observed after administration of mangoral, 
the abnormality must also be recorded as an AE (Section 6.3.7). All clinically significant 
abnormal laboratory values observed at 5 (± 2) days post-dose will be followed upon until they 
have normalised, or until there is no medical necessity for further blood tests in the 
investigator’s opinion (Section 6.3.7.3). 
After completion of all analyses, any leftover blood or blood samples will be destroyed 
immediately after the termination of the study. 
6.3.5.3 
Blood manganese concentration 
Blood samples for manganese measurements will be taken on the day of mangoral 
administration immediately (within 1 hour) prior to dosing, and at 24 (± 4), 48 (± 4), and 5 (± 2) 
days post-dose in all patients. A blood volume of 6 mL will be drawn per time point. Additional 


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blood samples for PK assessments will be taken on the day of mangoral administration only in 
the PK subgroup (Section 6.4.2). 
Blood samples for determination of manganese concentrations will be handled as described 
in Section 6.4.2. 
6.3.5.4 
Pregnancy test 
A serum pregnancy test will be performed in women of childbearing potential at the local 
laboratory at the screening. A urine or serum pregnancy test will be performed in women of 
childbearing potential at the local laboratory at baseline, i.e. within 24 hours prior to the 
administration of mangoral. Pregnancy test dipsticks will be provided by the central laboratory 
or sites will be using their standard of care dipsticks. 
If local regulations require serum pregnancy tests, a blood sample will be taken and a serum 
beta-hCG test will be performed at the local laboratory. In case of pregnancy, the patient is not 
allowed to enter the study or will be withdrawn from further study participation. 
All pregnancies diagnosed after mangoral intake will be reported to the sponsor/ designee 
(page 10 for contact details) on a paper pregnancy reporting form and will be monitored by 
the investigator up to the final outcome. In addition, the event should be captured on 
dedicated CRF pages as timelines detailed in the eCRF- Completion Guidelines. The 
outcome, including premature termination, must be reported to the sponsor as a follow-up to 
the initial report.  
6.3.6 Additional safety assessments 
Manganese is required for normal brain function as it plays a role as co-factor or regulator of 
brain enzymes, in particular of glutamine synthetase. However, it has been shown that 
overexposure to manganese, when absorbed in large amounts exceeding the homeostatic 
range, may have neurotoxic effects resulting in neuropsychological alterations, psychiatric 
symptoms and Parkinsonian symptoms with a significant decrement of performance in motor 
functions related to coordination and tremor [22, 36].  
Manganese accumulation in the brain occurs mainly in the globus pallidus and the striatum of 
the basal ganglia which can be detected by MRI imaging as high signal intensity in these 
regions on T1-weighted images [37, 38] 
6.3.6.1 
MRI of the brain – Optional 
An unenhanced MRI examination of the brain (Section 6.2.4.1.2) will be done 7 (+ 2) days after 
the administration of mangoral only in those patients who underwent brain MRI due to any 
clinical reason within the last 6 months prior to mangoral-enhanced MRI and for whom previous 
brain MRI images are available for comparison. Note that the brain MRI procedure is optional 
and requires a consent from the patient if to be performed. Post-dose images will be evaluated 
for any signs of manganese accumulation in the brain and will be compared by the independent 
off-site radiologist to the earlier brain MRI results. 
6.3.7 Adverse events 
Any untoward medical occurrence that is experienced by the patient from the time of signing 
the informed consent to the time immediately prior to the administration of IMP will be recorded 
in the eCRF as a pre-dose event. A pre-dose event cannot be causally related to the IMP; 
however, it may be causally related to study-related procedures (e.g. blood sampling during 


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the screening period). Thus, for each pre-dose event, relationship to study-related procedures 
and seriousness will be assessed and recorded. 
All untoward medical occurrences which are noticed after the administration of IMP up until the 
end of the follow-up period (end of day 7 [+2] in the brain MRI subgroup, end of day 5 [±2] in 
all other patients) will be reported as adverse events (AEs). The relationship to the IMP and 
also to study procedures will be recorded for all AEs that occur after administration of IMP.  
6.3.7.1 
Definition of adverse events 
An AE is any untoward medical occurrence in a patient or clinical investigation subject 
administered a pharmaceutical product and does not necessarily have a causal relationship 
with this treatment. An AE can therefore be any unfavourable and unintended sign (including 
abnormal laboratory finding), symptom, or disease temporally associated with the use of an 
IMP, whether or not related to the IMP. 
Symptoms or clinically significant laboratory or instrumental (e.g. electrocardiographic) 
abnormalities of a pre-existing condition, such as cancer or other diseases, should not be 
considered an AE. However, the occurrence of new symptoms, or laboratory, or instrumental 
abnormalities, and the worsening of pre-existing symptoms, are considered AEs. 
Laboratory values outside the normal laboratory-specific reference ranges that make 
intervention necessary and / or are considered clinically significant by the investigator must be 
reported as AEs. If the abnormal laboratory value is a sign of a disease, only this diagnosis 
should be reported as an AE. 
An AE is considered an adverse drug reaction (ADR) if a causal relationship between a 
medicinal product and an AE is at least a reasonable possibility.  
6.3.7.2 
Assessment of adverse events 
The investigator will identify the occurrence of pre-dose events and / or AEs through non-
leading questioning and examination of the patient. 
For each AE, the investigator will document the following: 
• 
Signs and symptoms of the event (if a specific disease can be diagnosed, this disease 
should be the reported AE; if only signs and symptoms can be evaluated, each sign or 
symptom should be reported as a separate AE) 
• 
Onset date and time (if a change from pre-dose in a laboratory test is reported as an AE, 
the start date is the date of collection of the first laboratory sample that shows the change) 
• 
End date and time (if a change from pre-dose in a laboratory test is reported as an AE, 
the end date is the date of collection of the first laboratory sample that shows a return to 
pre-dose level) 
• 
Measure taken (none, drug treatment required, hospitalisation or prolonged 
hospitalisation, study discontinuation, other measures [specification]) 
• 
Outcome (recovered/resolved, recovering/resolving, recovered/resolved with sequelae, 
not recovered/not resolved, fatal, unknown) 
In addition, each AE will be rated by the investigator according to the following categories (the 
sponsor must carry out a separate assessment for causal relationship, seriousness, and 
expectedness): 


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Relationship to the investigational medicinal product 
The causal relationship between an AE and the IMP will be evaluated and classified as follows: 
Related: 
 
After careful medical consideration, there is enough evidence (e.g. reasonable 
time sequence, known response pattern, not attributable to concomitant 
medications or concurrent diseases) to assume a causal relationship between 
the AE and the investigational medicinal product or a causal relationship the 
AE and the investigational medicinal product cannot be ruled out with certainty 
Not related: 
After careful medical consideration, the AE is clearly and incontrovertibly due 
to causes other than the investigational medicinal product (e.g. documented 
pre-existing condition) 
Intensity 
Regardless of the classification of an AE as serious or non-serious (see below), its intensity 
must be assessed according to medical criteria alone using the Common Terminology Criteria 
for Adverse Events (CTCAE) classification (v5.0): 
Grade 1 (mild): 
asymptomatic or mild symptoms; clinical or diagnostic 
observations only; intervention not indicated 
Grade 2 (moderate): 
minimal, local or non-invasive intervention indicated; limiting age-
appropriate instrumental activities of daily living (e.g. preparing 
meals, shopping, managing money 
Grade 3 (severe): 
severe or medically significant but not immediately life-
threatening; hospitalisation or prolongation of hospitalisation 
indicated; disabling; limiting self-care activities of daily living (e.g. 
bathing, dressing, undressing, feeding self, using the toilet, 
taking medications 
Grade 4 (life-threatening): 
life-threatening consequences; urgent intervention indicated 
Grade 5 (death:) 
death related to AE 
It should be noted that a severe AE does not necessarily have to be serious in nature and that 
a serious AE does not need to be of severe intensity. 
Seriousness 
A serious adverse event (SAE) is an AE that at any dose: 
• 
Results in death 
• 
Is life-threatening (i.e. the patient was at risk of death at the time of the event; it does not 
refer to an event that hypothetically might have caused death if it were more severe) 
• 
Requires in-patient hospitalisation or prolongation of existing hospitalisation 
• 
Results in persistent or significant disability/incapacity (defined as a permanent or 
substantial disruption of the patient’s ability to carry out normal life activities) 
• 
Is a congenital anomaly or birth defect 
or 
• 
Is an important medical event that does not have to be immediately life-threatening or result 
in death or hospitalisation, but may, based upon appropriate medical judgment, jeopardise 


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the patient or may require medical or surgical intervention to prevent any of the outcomes 
listed above. Examples of such events include allergic bronchospasm requiring intensive 
treatment in an emergency room or at home, and blood dyscrasias or convulsions that do 
not result in hospitalisation, or development of drug dependency or drug abuse. Note: 
events of seizure, stroke, cerebral venous thrombosis, QTcF or QTcB greater than 480 
msec or QTcF or QTcF increase of 60 msec over baseline must be reported as serious. 
All AEs that do not fall into any of the above categories are defined as non-serious. 
Note: planned hospitalisation or prolongation of hospitalisation for a pre-existing condition or a 
procedure required by this study protocol, without a serious deterioration in health, is not 
considered an SAE. 
Expectedness (only evaluated by the sponsor): 
Expected: 
An ADR is considered expected if its description agrees in nature, severity, 
frequency and specificity with the description of the up-to-date reference safety 
information included in the investigator's brochure 
Unexpected: An unexpected ADR is an adverse reaction for which the nature or severity 
is not consistent with the applicable product information (e.g., investigator’s 
brochure) 
A Suspected Unexpected Serious Adverse Drug Reaction (SUSAR) is defined as an untoward 
and unintended response to an IMP that is not listed in the applicable product information and 
meets one of the above mentioned serious criteria. The sponsor will ensure that all SUSARs 
associated with the IMP will be reported to the applicable regulatory authority/competent 
authorities and the relevant independent ethics committees (IECs)/institutional review boards 
(IRBs) in accordance with the applicable national/local regulations (usually within 7 calendar 
days for life-threatening and fatal SUSARs or within 15 calendar days for all other SUSARs). 
The sponsor/CRO will also promptly inform all investigators about any SUSAR. 
6.3.7.3 
Follow-up of adverse events 
Each unresolved AE at the patients’ individual study termination must be followed up until it 
subsides or until the cause is known or an adequate final assessment can be given. The 
maximum follow-up period will be 4 weeks after the patient’s study termination. If after 4 weeks 
no final outcome can be acquired, the outcome of the AE is ‘unresolved’. 
For all patients with clinically significant abnormal laboratory findings at 5 (± 2) days post-dose 
compared to the baseline evaluation (within 24 hours prior to the administration of IMP), the 
laboratory test will be repeated to ensure the validity of the abnormal result. Confirmed 
abnormal values will be followed up until normalisation to baseline level or until there is no 
medical necessity for further blood tests in the investigator’s opinion. Therefore, the patient 
might have to come to the site for one or more unscheduled follow-up visits. 
6.3.7.4 
Documentation and reporting of adverse events 
Any AE must be recorded in the eCRF with the specifications detailed in Section 6.3.7.2. 
All SAEs must be reported by the investigator immediately (within 24 hours) to the sponsor/ 
designee (minimum information required: patient number, site identifier, investigational 
medicinal product, event term, and causality assessment) on a dedicated paper SAE form. 


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A study-specific SAE form for immediate reporting will be provided to the investigators as part 
of investigator site file. In case of an SAE, this form should be completed and submitted 
immediately (no later than 24 hours after the investigator’s awareness of the event) to the 
sponsor/ designee (page 10 for contact details). Secure email of the SAE form is the preferred 
method to transmit this information. In addition, the event should be captured on the dedicated 
(Serious) Adverse Event page of the eCRF within 3 days. 
In order to perform an independent medical assessment of the reported case, the Sponsor or 
a designee may follow-up by telephone, electronic mail, and/or a monitoring visit to obtain 
additional case details and outcome information (e.g. hospital discharge summaries, 
consultant reports, autopsy reports). 
Follow-up reports will be provided by the investigator each time new  information on the SAE 
becomes available (until the case is resolved or the patient has recovered). Every effort should 
be made to follow all SAEs considered to be related to the IMP or trial-related procedures until 
a final outcome can be reported. For information, a similar procedure as for reporting of initial 
SAEs will apply.  
Investigators are not obliged to actively seek AE or SAE after the end of the AE reporting 
period (defined as 5 [± 2] days after IMP intake). However, if the investigator learns of any 
SAE, at any time after the end of AE reporting period, and he/she considers the event to be 
reasonably related to the IMP or study participation, the investigator must promptly notify the 
sponsor.  
Notification of the IECs/IRBs and competent authorities about all relevant events will be 
performed by the sponsor (or designated CRO) and / or by the investigator (as applicable) 
according to all applicable national/local regulations. 
6.4 
Pharmacokinetics 
6.4.1 Pharmacokinetic variables 
Pharmacokinetics will be evaluated only in the PK subgroup (N = 12) at selected study sites. 
Blood concentrations of manganese will be corrected for baseline (pre-dose) concentration of 
manganese. 
The following PK variables will be determined: 
• 
AUC (area under the curve): for the interval 0-120 hours 
• 
Cmax (maximum or peak concentration): determined from tabulated observed blood level 
data 
• 
tmax (time to achieve maximum concentration) 
• 
t1/2 (terminal half-life): determined as 0.693/Kel, where Kel is a first-order elimination rate 
constant determined from the slope of the terminal phase of semilog plot of concentration 
vs. time data 
• 
Clapp (total apparent clearance): calculated as dose/AUC 
6.4.2 Blood sampling time points, handling and labelling of blood samples 
Patients, who are part of the PK subgroup shall have V4 and V5 at the study site.  
Blood samples for PK evaluation will be collected by venipuncture (or by using an indwelling 
catheter) during the screening period, immediately prior to the administration of the IMP (within 


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1 hour pre-dose), and 0.25, 0.5, 1, 2, 3, 4, 6, 8, 12, 24, 48, and 120 hours post-dose. A blood 
volume of 6 mL will be drawn per time point. 
Handling and labelling of blood samples for PK analysis will be performed according to the 
instructions provided by the central laboratory. Special tubes, labels, packaging, and 
instructions for storage and shipment will also be provided by the central laboratory. Blood 
samples should be shipped for analysis on the day of collection. 
Blood manganese concentrations will be determined from all collected samples using a 
validated sensitive analytical procedure (e.g. inductively coupled plasma mass spectrometry) 
in accordance with the EMA Guideline on Bioanalytical Method Validation and the FDA 
Guidance for Industry at a specialised central analytical laboratory. 
After completion of all analyses, any leftover blood or blood samples will be destroyed 
immediately after the termination of the study. 


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7 
STUDY CONDUCT 
7.1 
Study schedule 
The schedule of visits and study-related procedures at each visit are presented in Table 9.  
Patients who are interested in participating in the study will sign an informed consent form after 
having been thoroughly informed about the purpose and procedures of the study and before 
any screening examination is performed. Screening will be performed within 4 weeks (between 
days -28 and -1) prior to the day of the mangoral-enhanced liver MRI.  
Depending on the clinical condition of the patient and the routine at the study site, 
hospitalisations for up to 2 days for the baseline evaluations and the performance of 
unenhanced/enhanced MRI, as well as other study-related procedures on the day of the MRI, 
may be necessary. Patients in the PK subgroup will be hospitalised at baseline for at least 2 
days because these patients will undergo additional blood sampling for PK assessments within 
1 hour pre-dose and between 0.25 and 12 hours post mangoral administration. 
Scheduled follow-up visits (FU1, FU2, FU3, and FU4, if required) may be performed as 
outpatient visits. In case a patient is not able to return to the study site for the follow-up visits, 
the follow-up assessments may be performed by a trained examiner at an off-site location, e.g. 
at the patient’s home or at an associated off-site clinic. For patients with long travelling 
distances, it may be possible that a hotel room will be booked for the patient and an 
accompanying person (caregiver) by the study site coordinator in agreement with the sponsor. 
Hotel costs will be at the expense of the sponsor. Costs for travelling to the study sites may 
also be reimbursed upon request and submission of tickets and/or receipts.  
In the event, a patient is not part of the PK subgroup, FU1 (V4 at 24 [± 4] hours) and FU 2 (V5 
at 48 [± 4] hours) can be performed as remote visits at the discretion of the investigator, if 
further adjustments are required to ease the burden for the patient. If an on-site visit is replaced 
by a remote visit, information on possible adverse events and changes in concomitant 
therapies shall be collected timely. 
Only a subgroup of patients, i.e. patients who underwent brain MRI due to any clinical reason 
within the last 6 months prior to mangoral-enhanced MRI and for whom previous brain MRI 
images are available, will return to the site for the FU4 visit. At this visit, which will take place 
7 (+ 2) days after the mangoral-enhanced liver MRI, an MRI of the brain will be performed.  
 




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n  Clinical safety laboratory tests (Section 6.3.5.2) to be performed within 24 hours prior to mangoral administration (either on the 
day before or on the day of the MRI examination) and after 2 (± 0.5) and 6 (± 1) hours post-dose on the day of the MRI 
examination. All clinically significant abnormal laboratory values observed at 5 (± 2) days post-dose will be followed up until 
they have normalised, or until there is no medical necessity for further blood tests in the investigator’s opinion. 
o  Blood samples for manganese measurements will be taken on the day of mangoral administration immediately prior to dosing 
(within 1 hour pre-dose; Section 6.3.5.3) in all patients; additional blood samples will be taken at 0.25, 0.5, 1, 2, 3, 4, 6, 8, and 
12 hours post-dose only in the PK subgroup (Section 6.4.2). 
p  AE reporting starts from the time of signing the informed consent; an event that occurs prior to the administration of mangoral 
will be recorded as ‘pre-dose event’ in the eCRF. Any unresolved AE at the patients’ individual study termination must be 
followed up until it subsides or until the cause is known or an adequate final assessment can be given for a maximum follow-
up period of 4 weeks, Section 6.3.7.3). 


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7.2 
Observations by visit 
7.2.1 Screening visit: V1 (day -28 to -1) 
A Screening Visit will be performed within 4 weeks and up to 1 day prior to the planned 
mangoral-enhanced MRI examination. Patients who are interested in participating in the 
study will be informed verbally and in writing about the purpose, nature, risks, and benefits 
of the study.  
The following will be carried out and / or recorded at the Screening Visit: 
• 
Patient signs informed consent form as a part of the written patient information including 
the agreement concerning protection of personal data and direct access to source 
documents 
• 
Demographic data (incl. year of birth, age, sex, height, weight) 
• 
Relevant general medical history (Section 6.1.2.1) 
• 
Clinical signs and symptoms (including diagnosis) of the underlying primary malignancy 
(Section 6.1.2.2) 
• 
Pregnancy test (serum test for female patients of childbearing potential) (Section 6.3.5.4) 
• 
Review of inclusion and exclusion criteria (Sections 4.1 and 4.2) 
• 
Child-Pugh evaluations/determination of Child-Pugh score (Sections 6.1.2.3 and 6.3.5.1) 
• 
Concomitant and prior medications taken within 14 days pre-dose (Section 5.2) 
7.2.1.1 
Screening 
Potential participants will be screened based on inclusion and exclusion criteria. 
7.2.1.2 
Re-screening and re-consenting 
Re-screening is allowed if the patient has not undergone the IMP-enhanced MRI, earliest 5 
days post dosing attempt. 
A new screening number will be assigned in the eCRF. 
7.2.2 Baseline period: V2 (day -1) to V3 (day 0), within 24 hours pre-dose 
The following will be performed and/or recorded during the baseline period, i.e. within 24 hours 
prior to mangoral administration (note: some of the baseline assessments may be 
done/recorded on the day/evening before the MRI, or all baseline assessments may be 
done/recorded on the day of the MRI): 
• 
Concomitant and prior medications taken within 14 days pre-dose (Section 5.2) 
• 
Vital signs within 3 hours prior to mangoral administration (Section 6.3.1) 
• 
Physical examination within 3 hours prior to mangoral administration (Section 6.3.2) 
• 
ECG within 3 hours prior to mangoral administration (Section 6.3.4) 
• 
Neurological assessments within 24 hours prior to mangoral administration (Section 6.3.3) 
• 
Verification of inclusion/exclusion criteria (eligibility check), as applicable 
• 
Pregnancy test (urine or serum test for female patients of childbearing potential) within 
24 hours prior to the administration of mangoral (Section 6.3.5.4) 
• 
Blood sampling for clinical safety laboratory tests (haematology, biochemistry) within 
24 hours prior to mangoral administration (Section 6.3.5.2) 


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• 
Urine collection for urinalysis within 24 hours prior to mangoral administration 
(Section 6.3.5.2) 
• 
Unenhanced MRI of the liver either on day -1 or on day 0 prior to mangoral administration 
(Section 6.2.4.1.1) 
• 
Blood sampling for determination of manganese concentration immediately (within 1 hour) 
before administration of mangoral (Section 6.3.5.3) 
• 
Pre-dose events (for definition Section 6.3.7) 
7.2.3 Day of Dosing: V3 (day 0), dosing, mangoral-enhanced MRI and post-dose 
assessments  
After the performance/documentation of all baseline assessments (Section 7.2.2), the following 
examinations will be carried out and/or recorded on the day of MRI: 
• 
Administration of IMP (to be taken after fasting for at least 4 hours; Section 5.1) 
• 
Mangoral-enhanced MRI of the liver at 4 (± 1) hours after mangoral administration (Section 
6.2.4.1.1) 
• 
Any changes in concomitant therapies (including medications taken only once and any 
non-prescription medicine) 
• 
Vital signs at 6 (± 1) hours post-dose (Section 6.3.1) 
• 
Physical examination at 6 (± 1) hours post-dose (Section 6.3.2) 
• 
ECG at 6 (± 1) hours post-dose (Section 6.3.4) 
• 
Blood sampling for clinical safety laboratory tests (haematology, biochemistry) at 2 (± 0.5) 
and 6 (± 1) hours post-dose (Section 6.3.5.2) 
• 
Urine collection for urinalysis at 2 (± 0.5) and 6 (± 1) hours post-dose (Section 6.3.5.2) 
• 
Only in the PK subgroup: blood sampling for determination of manganese concentration at 
0.25, 0.5, 1, 2, 3, 4, 6, 8, and 12 hours post-dose (PK; Section 6.4.2) 
• 
Adverse events (Section 6.3.7) 
7.2.4 Follow-up visit V4 (day 1) 
The following examinations will be carried out and / or recorded on the first day following the 
day of mangoral dosing and the mangoral-enhanced MRI (24 [± 4] hours post-dose): 
• 
Any changes in concomitant therapies (including medications taken only once and any 
non-prescription medicine) 
• 
Vital signs at 24 hours post-dose (Section 6.3.1) 
• 
Physical examination at 24 hours post-dose (Section 6.3.2) 
• 
Neurological assessments at 24 hours post-dose (Section 6.3.3) 
• 
ECG at 24 hours post-dose (Section 6.3.4) 
• 
Blood sampling for clinical safety laboratory tests (haematology, biochemistry) at 24 hours 
post-dose (Section 6.3.5.2) 
• 
Urine collection for urinalysis at 24 hours post-dose (Section 6.3.5.2) 
• 
Blood sampling for determination of manganese concentration at 24 hours post-dose 
(Section 6.3.5.3) 
• 
Adverse events (Section 6.3.7) 


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If Visit 4 is conducted as a remote visit, the reason for the change of the visit type shall be 
recorded in the medical records. As a minimum, information on possible adverse events and 
changes in concomitant medication must be collected.  
7.2.5 Follow-up visit V5 (day 2) 
The following examinations will be carried out and / or recorded on the second day following 
the day of mangoral dosing the mangoral-enhanced MRI examination (48 [± 4] hours 
post-dose): 
• 
Any changes in concomitant therapies (including medications taken only once and any 
non-prescription medicine) 
• 
Vital signs at 48 hours post-dose (Section 6.3.1) 
• 
Physical examination at 48 hours post-dose (Section 6.3.2) 
• 
Neurological assessments at 48 hours post-dose (Section 6.3.3) 
• 
Blood sampling for clinical safety laboratory tests (haematology, biochemistry) at 48 hours 
post-dose (Section 6.3.5.2) 
• 
Urine collection for urinalysis at 48 hours post-dose (Section 6.3.5.2) 
• 
Blood sampling for determination of manganese concentration at 48 hours post-dose 
(Section 6.3.5.3) 
• 
Adverse events (Section 6.3.7) 
If Visit 5 is conducted as a remote visit, the reason for the change of the visit type shall be 
recorded in the medical records. As a minimum, information on possible adverse events and 
changes in concomitant medication must be collected.  
7.2.6 Follow-up visit V6 (day 5) 
The following examinations will be carried out and/or recorded on the third day following the 
day of the day of IMP dosing and mangoral-enhanced MRI examination (5 [± 2 ] days 
post-dose): 
• 
Any changes in concomitant therapies (including medications taken only once and any 
non-prescription medicine) 
• 
Vital signs at 5 (± 2) days post-dose (Section 6.3.1) 
• 
Physical examination at 5 (± 2) days post-dose (Section 6.3.2) 
• 
Neurological assessments at 5 (± 2) days post-dose (Section 6.3.3) 
• 
Blood sampling for clinical safety laboratory tests (haematology, biochemistry) at 5 (± 2) 
days post-dose (Section 6.3.5.2) 
• 
Urine collection for urinalysis at 5 (± 2) days post-dose (Section 6.3.5.2) 
• 
Blood sampling for determination of manganese concentration at 5 (± 2) days post-dose 
(Section 6.3.5.3) 
• 
Adverse events (Section 6.3.7) 
7.2.7 Follow-up visit V7 (day 7) in optional brain MRI subgroup 
The following examinations will be carried out and/or recorded on the seventh day following 
the day of the MRI: 
• 
Any changes in concomitant therapies (including medications taken only once and any 
non-prescription medicine) 


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• 
Unenhanced MRI of the brain (Section 6.2.4.1.2) 
• 
Adverse events (Section 6.3.7) 
7.2.8 Withdrawal Visit 
Patients who withdraw from treatment or the study prior to the last Follow-Up Visit will be asked 
to attend a Withdrawal Visit, if at all possible. The visit must be scheduled as soon as possible 
after withdrawal. 
No new information will be collected from patients who withdraw from the study, except 
information collected in relation to the scheduled Follow-up Visits (Section 7.2.4, 7.2.5, 7.2.6) 
and information needed for the follow-up of AEs (Section 6.3.7). The reason for withdrawal 
must be recorded in the eCRF. 
7.3 
Independent Data Safety Monitoring Board (IDSMB) 
An Independent Data Safety Monitoring Board (IDSMB) will review all safety data available 
once 30 patients have completed the third follow-up visit (5 days after the administration of 
mangoral), and make recommendations to the Sponsor to either continue, modify or 
discontinue the study.  
After consideration, the Sponsor will inform the IDSMB of any action that will be taken in 
response to the IDSMB’s recommendations.  
The role and responsibilities of the IDSMB, their operational procedures, and methods of 
communication with the Sponsor, are described in a separate IDSMB Charter. The IDSMB will 
consist of three expert members who are independent of the Sponsor. The IDSMB will be 
composed by: 
• 
A radiologist, who participated in the previous Phase II study or Phase I studies 
• 
A neurologist and 
• 
A biostatistician experienced in DSMB evaluation 
All IDSMB members will have experience in the conduct of clinical studies. Members will not 
be investigators in the trial, nor will they have any conflict of interest with the Sponsor. Members 
will not have a bias with regard to the use of mangoral for imaging diagnostic. Sponsor 
representatives are not eligible for membership on the IDSMB 
7.4 
Duration of the study 
7.4.1 Planned duration for the individual patient 
After a screening visit within 4 weeks prior to the administration of mangoral, the core study 
phase will last 4-5 days for each individual participant and comprises a baseline period of up 
to 24 hours prior to mangoral administration, the day of mangoral administration and 3 follow-
up visits on 24 (± 4) hours, 48 (± 4) hours and 5 (± 2) days following the day of mangoral 
administration. In a subgroup of patients (i.e. patients who underwent brain MRI for any clinical 
reasons within 6 months prior to the mangoral-enhanced liver MRI and for whom previous 
brain MRI images are available), a fourth follow-up visit will be performed 7 (+ 2) days after 
contrast administration for an unenhanced brain MRI examination. 
In case of clinically significant abnormal laboratory findings, the patient will be asked to return 
to the study site for follow-up blood tests until normalisation of the value(s) to baseline level is 
observed or until further blood sampling is not medically necessary in the investigator’s opinion. 


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7.4.2 Premature termination 
Study 
The study as a whole may be terminated prematurely by the sponsor at any time if: 
• 
New toxicological or pharmacological findings or SAEs invalidate the earlier positive 
benefit–risk assessment 
• 
The development of the IMP is discontinued, a marketing authorisation is no longer 
intended or the study does not meet the expected goal; or 
• 
Other important reasons, not named above 
Site 
A participating study site can be excluded from participation by the sponsor if: 
• 
The site fails to comply with the requirements of the protocol 
• 
The site fails to comply with GCP standards; or 
• 
The first patient is not recruited within a reasonable period after initiation of the site 
Patient 
Individual patients are to be withdrawn from the trial according to the criteria specified in 
Section 4.3. 


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8 
STATISTICS 
8.1 
Statistical and analytical plans 
All statistical analyses will be outlined in detail in a statistical analysis plan which will be 
prepared and signed prior to any study-specific analyses. In general, all continuous measures 
will be summarised descriptively, including the number of available values, minimum, 1st 
quartile, median, mean, standard deviation, 3rd quartile, maximum, if appropriate. Categorical 
data will be presented by frequency and percentage. Ordinal ratings may be handled as 
continuous data, if appropriate. 
Inferential methods will be based on a significance level of 0.05 for two-sided tests and 0.025 
for one-sided tests. All statistical significance tests besides the primary efficacy analysis should 
be interpreted in an exploratory manner. 
8.1.1 Analysis populations 
All patients who have taken a single dose of the IMP will be included in the safety analysis 
(safety population). The efficacy analysis will be performed in the full analysis set of patients 
and on a per-protocol basis, see definitions below (Table 10). Patients with major protocol 
violations will be excluded from the latter. For this purpose, protocol violations that could 
interfere with the objectives of the study, e.g. incorrect concomitant medications or violation of 
the inclusion/exclusion criteria will be assessed as minor or major in collaboration with the 
sponsor. The criteria for this assessment will be defined before analysis. Listings will be 
prepared to show the eligibility of all patients. 
The primary analysis will be done according to the intention-to-treat principle for the full 
analysis set. In addition, a per-protocol analysis will be prepared as an assessment of 
sensitivity. 
 
Table 10 
Definition of analysis populations 
Population 
Description 
Safety population 
All patients  enrolled in the study (fulfil all inclusion criteria, but none 
of the exclusion criteria and have been included in the clinical trial 
at Visit 2). 
Full analysis set (FAS) 
All patients of the safety population who received the IMP and for 
whom the primary efficacy variable is assessable, i.e. all 
unenhanced/enhanced liver MRI images are assessable. 
Per-protocol set (PPS) 
The per-protocol set defines the subset of the patients in the FAS 
without major protocol violations, including violations of 
inclusion/exclusion criteria. 
PK population 
All patients in the PK subgroup with PK blood samples taken after 
mangoral administration.  
PK dialysis subgroup population 
(DSP) 
All patients in the PK subgroup, currently on maintenance 
haemodialysis. 
Brain MRI subgroup population 
(BMSP) 
All patients with brain MRI performed within 6 months prior to V3 who 
underwent the optional brain MRI 7 (+ 2) days after mangoral-
enhanced MRI of the liver. 
 


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8.1.2 Adaptive study design 
The study design is not adaptive since no interim analysis is planned. 
8.1.3 Primary efficacy analysis 
The primary population for this analysis will be the full analysis set (FAS). Efficacy will be 
assessed by testing the superiority of each of the 2 co-primary variables:  
• 
lesion border delineation (BD), and 
• 
lesion contrast compared to liver background (LC). 
The testing will be done separately for each of the three independent readers and will be based 
on the mean scores of BD and LC. As described in Section 6.2.5, each of the 
three independent readers will assess all unenhanced and enhanced liver MRI images of all 
patients distributed over 3 reading parts (part I: unenhanced alone, part II: paired reading, part 
III: enhanced alone). Each reading part will yield a qualitative assessment of BD and LC on 
4-point scales (score 1 to 4; Section 6.2.2.1) per lesion.  
The two sum scores (sBD and sLC) are determined for each patient by summarising the 
individual scores over all lesions detected at baseline for part I (unenhanced MRI) and part II 
(CMRI). Then, the mean score differences between the paired sum scores weighted by the 
number of lesions are determined per patient: 
Diff(BD) =
 sBDcombined − sBDunenhanced
Number of lesions
 . 
Diff(LC) =
 sLCcombined − sLCunenhanced
Number of lesions
 . 
For each independent reader, the hypotheses for the superiority of CMRI versus unenhanced 
MRI will be tested using a 1-sided paired t-test using the means of the paired score differences: 
 
H0;BD: µDiff(BD) ≤ 0 versus  H1;BD: µDiff(BD) > 0 
and 
 
H0;LC: µDiff(LC) ≤ 0 versus  H1;LC: µDiff(LC) > 0 
If both tests indicate the superiority of CMRI at the one-sided significance level of 0.025, reader 
success is achieved. The corresponding two-sided 95% confidence intervals will be calculated 
as well. These results will be presented in a table together with a summary of the mean scores 
per reader. For the success of the trial, two of three readers must achieve reader success. 
As a sensitivity analysis, the analogous 1-sided Wilcoxon signed-rank test will be performed 
additionally. 
The primary variable will be analysed for the full analysis population and additionally in an 
exploratory manner for the per-protocol population using the above-described methodology.  
8.1.4 Secondary efficacy analyses 
A comparison of part I (unenhanced MRI alone) and part III (mangoral-enhanced MRI alone) 
will be analysed in the same way as the primary endpoint.  
The other secondary efficacy variables (Section 6.2.2.2) will be analysed in an exploratory 
manner using descriptive statistics (including two-sided 95% confidence intervals) for the full 
analysis population and the per-protocol population. Exploratory statistical tests may be 


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defined if deemed beneficial in addition to the confidence intervals. Details will be defined in a 
statistical analysis plan, which will be signed prior to any study-specific analyses. 
Inter-reader and intra-reader agreement:  
Additionally, assuming repeated assessments per subject by three independent readers, an 
intra-reader and inter-reader reproducibility will be evaluated separately for MRI readings in 
Parts I, II and III using linear mixed-effects models fitted to individual mean scores of border 
delineation (BD) and lesion contrast (LC), considering patients as a random effect. Details of 
the statistical model will be specified in the SAP. 
8.1.5 Subgroup analyses 
The primary efficacy endpoint will be evaluated in appropriate subgroups which will be defined 
in the statistical analysis plan. These subgroups may include but are not limited to: 
• 
Subgroups by MRI scanner’s magnetic field strength (1.5 Tesla or 3 Tesla) 
• 
Subgroup by lesion type (e.g. liver metastasis, primary liver tumour) 
• 
Subgroups by lesion diameter (average of the measurements by the three independent 
readers):  
o small lesion (diameter < 1 cm) 
o medium lesion (diameter ≥ 1 to ≤ 3 cm) 
o large lesion (diameter > 3 cm) 
• 
Subgroups by age (≤ 65 years, > 65 years) 
• 
Subgroups by eGFR (eGFR < 15, ≥ 15 to ≤ 30, > 30 mL/min/1.73m2) 
• 
Subgroups by status of liver disease (no liver disease, Child-Pugh A [e.g. mild fibrosis])  
• 
Subgroup of patients with PK data 
• 
Subgroup of patients with brain MRI 
8.1.6 Safety analyses 
AEs will be encoded using the MedDRA thesaurus in the most recent version available at the 
beginning of the study. This version will be used throughout the entire study. Updates of the 
MedDRA thesaurus will not be applied afterwards. The same will apply to the Anatomic 
Therapeutic Chemical Classification System developed by the World Health Organisation, 
which will be used for the coding of previous and concomitant medications. Frequency tables 
for coded previous and concomitant medication will be compiled 
Frequency tables by system organ class and preferred term will be prepared based on patients 
experiencing an AE and based on the number of AEs. These frequency tables will also be 
prepared for AEs assessed as related to the investigational medicinal product. A frequency 
table will be created to give an overview of numbers of AE and patients with an AE in relation 
to investigational product, severity and seriousness (if an AE is recurring with the same term, 
the highest reported severity will be used for categorisation). 
Results of vital sign measurements, physical examinations, neurological assessments, brain 
MRI assessment and ECG will be presented using descriptive statistics. 


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Laboratory data will be presented using descriptive statistics as well as using shift-tables with 
respect to normal ranges. 
Concomitant diseases and medical history will be presented using frequency tables. 
8.1.7 Pharmacokinetic analyses 
The analysis of PK variables (Section 6.4) will be based on the PK population (for definition 
see Table 10). PK variables will be summarized descriptively. A plot will show the manganese 
concentration over time for each patient (spaghetti plots). 
8.1.8 Missing data 
Adverse events with unknown onset times will be counted as post mangoral administration 
AEs. AEs with missing relationship to mangoral will be counted as possibly related AEs. 
Missing efficacy data will not be replaced. Technical errors, e.g. laboratory values that have to 
be regarded as technical errors from a medical point of view, will not be included in the 
statistical analysis and will only be reported in the patient data listings. Exclusion of medically 
implausible data will be decided during data cleaning or a data review meeting before database 
lock. 
8.1.9 Multicentre trial 
This trial will be performed at multiple sites. The assessments used for the primary analysis 
will be performed by three independent readers for the pool of all MRI images of all sites. 
Therefore, the efficacy analysis will be performed for each reader separately and not stratified 
by site. Each of the study sites is expected to include only a small number of patients; therefore, 
also the analysis of secondary efficacy parameters based on on-site reads will not be stratified 
by site.  
8.1.10 Patient data listings 
All recorded data will be presented in the patient data listings. 
8.1.11 Deviations from the planned statistical analysis 
Any deviations from the planned statistical analysis have to be discussed in the final study 
report and should be defined in the statistical analysis plan. If the deviation has any impact on 
the confirmatory analysis, the deviation must be specified in an amendment to the study 
protocol. 
8.1.12 Interim analysis 
No interim analysis is planned. 
8.1.13 Software used for statistical analysis 
The SAS software version 9.4 or higher will be used for the statistical analysis and for the 
reporting of this trial. 
8.1.14 Determination of sample size 
The primary endpoint analysis consists of two co-primary one-sided t-tests at the conventional 
one-sided significance level of 0.025. Following the FDA’s draft guidance on multiple endpoints 
in clinical trials, adjustment of the type-I-error is not necessary since both primary variables 


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need to show superiority simultaneously. Nonetheless, this affects the type II error. To 
counteract the inflation of the type II error, each test’s power is set to 0.9 to achieve a study 
power of at least 0.9 x 0.9 = 0.81. 
Effect size 
The primary efficacy is based on qualitative parameters, but no exact estimates are available 
from previous studies due to the use of DWI in this study. A study investigating a 
manganese-based contrast agent with T1 / T2 and DWI comparing also unenhanced versus 
CMRI [39] reported qualitative score results on a scale from 1 to 5. These results are used to 
derive estimates for mean and standard deviation (SD) and scaled down for the 1 to 4 scale 
used in this study. Calculating an average score per patient and averaging readers after 
rescaling, leads to averages of 2.3 for unenhanced versus 2.6 for CMRI. This is a difference 
of 0.3 with each sample’s standard deviation of ~1.9, or ~1.5 after rescaling. For estimation of 
the SD of the samples’ differences, a high correlation (0.8) is assumed, which results in 0.95. 
Conservatively assuming no effect of the subpopulation of the HCC patients (20% of the study 
population), the effect is 0.3 x (1-0.2) = 0.24. 
Sample size calculation was performed using SAS® version 9.4 resulting in 167 patients 
needed for analysis (Table 11, Figure 2). Assuming, that only 85% of the image sets will be 
completely evaluable (all enhanced and unenhanced liver MRI images) and that no patients 
will drop out (since the efficacy procedure is completed within a few hours for each patient), 
197 patients need to be included in the study. 
 
Table 11 
Sample sizes 
Computed N Pairs 
SD paired 
Mean Difference 
Correlation 
Actual Power 
N Pairs 
1.5, 1.5 
0.2 
0.75 
0.9 
298 
1.5, 1.5 
0.2 
0.8 
0.9 
239 
1.5, 1.5 
0.2 
0.85 
0.9 
180 
1.5, 1.5 
0.22 
0.75 
0.9 
247 
1.5, 1.5 
0.22 
0.8 
0.9 
198 
1.5, 1.5 
0.22 
0.85 
0.9 
149 
1.5, 1.5 
0.24 
0.75 
0.9 
208 
1.5, 1.5 
0.24 
0.8 
0.9 
167 
1.5, 1.5 
0.24 
0.85 
0.9 
126 
1.5, 1.5 
0.26 
0.75 
0.9 
177 
1.5, 1.5 
0.26 
0.8 
0.9 
142 
1.5, 1.5 
0.26 
0.85 
0.9 
107 
1.5, 1.5 
0.3 
0.75 
0.9 
134 
1.5, 1.5 
0.3 
0.8 
0.9 
108 
1.5, 1.5 
0.3 
0.85 
0.9 
81 
 


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Figure 2 
Sample sizes depending on correlation and mean differences 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 


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9 
DATA HANDLING AND DATA QUALITY ASSURANCE 
9.1 
Data management 
Data will be captured using the eCRF provided by the designated CRO. The eCRF is 
specifically designed to meet the data recording requirements of the clinical study protocol. 
Only the investigator and his/her authorised staff (as listed on the form ‘Task Delegation and 
Signature Form’ provided by the CRO) who are trained appropriately on the eCRF system are 
allowed to fill-in the eCRFs or to make corrections, and will have access to the data or may 
change data (however without the possibility to delete the original entries). After completion, 
each eCRF will be electronically signed and dated by the investigator. Central laboratory test 
results will be imported automatically into the eCRF. For all laboratory and central imaging 
data, the units or any transformation of units must be clearly defined (if not otherwise agreed). 
The overall procedures for quality assurance of clinical trial data are described in the 
respective standard operating procedures (SOPs) of the CRO. A document describing the 
functional specification will be prepared by the CRO in cooperation with the eCRF provider 
and be approved by the sponsor prior to the start of the trial. This functional specification 
will describe in detail the structure and functionality of the eCRF. In addition, the CRO will 
prepare a data management plan to describe the procedures and processes of data 
collection and data coding. 
Electronic validation of the data entered in the eCRFs will take place (e.g. check of ranges, 
consistency, and plausibility). All plausibility checks will be defined in the data validation 
plan to be approved by the sponsor. Additionally, the eCRFs will be reviewed by qualified 
data management personnel and medically qualified personnel for completeness, 
consistency, and plausibility. Any data anomalies will be communicated to the site(s) for 
clarification/resolution (data query). Procedures for data entry and correction will be 
tracked and appropriately recorded in the audit trail of the eCRF. 
9.2 
Documentation 
9.2.1 Screening log and patient identification list 
A screening log will be kept by the investigator, listing all patients screened for this study. 
Patients will be identified by initials and date of birth. The status of the patient, i.e. whether 
enrolled in this study or not, will be entered on the log.  
All patients who have given informed consent to study participation – regardless whether 
the patient has received any IMP or not – have to be entered on the patient identification 
list by the investigator, giving initials, date of birth, and patient identification code.  
The screening log and patient identification list will be kept in the investigator's file. 
9.2.2 Source data and patient records 
In this study, all collected data in the patient file are considered source data, including MRI 
images and interpretations. Prior to including patients in the study, the investigator must sign 
a source data agreement that identifies the source documents (original documents, data, and 
records) at the site. The document will also list which data may be recorded directly on the 
eCRFs or any electronic device (as applicable). 


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Definition of source data: All information in original records and certified copies of original 
records of clinical findings, including observations or other activities in a clinical trial necessary 
for the reconstruction and evaluation of the trial.  
9.2.3 Electronic case report form (eCRF) 
An appropriate eCRF in English will be developed by the CRO. An eCRF completion manual 
for the use of the electronic data capture system will be provided prior to study start and 
investigators will be trained on how to enter all study-specific data into the eCRF. Investigators 
must enter the information required by the protocol into the eCRF and should complete the 
eCRF as soon as possible, but latest on the third working day after a patient visit (or in case 
of on-site assessment results: within 3 working days after the on-site assessment). No 
questions must be left unanswered. Note: all SAEs must be reported by the investigator 
immediately (within 24 hours) via the SAE paper form and additionally in the eCRF in timelines 
specified in CRF completion manual (Section 6.3.7.4). 
For all countries, the eCRFs have to be completed in English. Monitors will review the eCRFs 
for completeness and accuracy, and instruct site personnel to make any required corrections 
or additions. The eCRFs are saved immediately after entering the data by the investigator. All 
eCRF data are stored in a central database at the eCRF provider while the trial is ongoing and 
at the responsible CRO for final analysis after trial completion.  
After trial completion, the trial site will receive a digital media with a compilation of the eCRF 
trial data per site (electronic PDF files) for filing with its trial documents. The sponsor will 
receive electronic, bookmarked PDF files of individual eCRFs (including audit trail). The 
sponsor will also receive a list of queries and SAE forms after database closure. 
9.3 
Direct access to source data/documents 
In accordance with ICH GCP E6(R2), the investigator/study site will permit direct access to 
source data/documents for trial-related monitoring, audits, IECs/IRBs review, and regulatory 
inspection(s). Direct access includes permission to examine and verify any records that are 
important for the evaluation of the study. Patients will consent in writing to the direct access to 
their original medical record for trial-related monitoring (source data verification), audits and 
regulatory inspections. 
9.4 
Monitoring 
Monitoring will be done in accordance with the stipulations of Section 5.18 of the ICH GCP 
E6(R2) guideline and sponsor’s / CRO’s SOPs by designated representatives of the sponsor 
or CRO (study monitor). The sponsor / CRO will develop a separate clinical monitoring plan 
tailored to the specific patient protection and data integrity risks of the study. 
The study monitor will visit the study site prior to starting the first study-related procedure at 
this site (initiation visit) to ensure the eligibility of the study site, qualification of staff, and 
appropriateness of technical equipment, to discuss the protocol, study-related procedures, and 
responsibilities, and to ensure that the investigator receives all documents (including an up-to-
date version of the mangoral investigator’s brochure [19]) needed to conduct the study 
properly. The investigator will allow the study monitor to make regular visits during the course 
of the study to: 


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• 
Verify that written informed consent was obtained prior to each patient’s participation 
in the study 
• 
Discuss any emergent problems 
• 
Check the eCRFs for completeness and plausibility 
• 
Review accountability, correct storage, and handling of the investigational medicinal 
product 
• 
Ensure that the study is being conducted according to pertinent regulatory 
requirements, ICH GCP E6(R2), and the protocol 
eCRF entries will be verified against source documentation in the patient’s medical record or 
in the location stated in the source data agreement. Study monitors will maintain confidentiality 
of the patients’ personal data. 
A close-out visit will be performed upon conclusion of the study site’s participation in the study. 
9.5 
Quality assurance 
Risk-based quality management will be implemented in this study based on the requirements 
of the Integrated Addendum of the Good Clinical Practice guideline ICH E6(R2). Aspects of 
risk-based quality management will be detailed in a risk management plan. 
A member of the sponsor’s (or designated CRO’s) quality assurance unit may conduct an audit 
at the study site to ensure compliance with the protocol, GCP and applicable regulatory 
requirements. 
In addition, inspections by regulatory health authority representatives and IEC(s) / IRB(s) are 
possible. The investigator or his / her designee should notify the sponsor immediately of any 
such inspection. 
The investigator is expected to cooperate with such inspections and to discuss any findings 
with the auditor. The observations and findings of the auditor will be recorded and the 
investigator will be informed of the audit outcome. 


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10 ETHICAL, LEGAL AND ADMINISTRATIVE ASPECTS 
The study will be carried out in conformity with the ethical principles enunciated in the 
Declaration of Helsinki [40]. The currently valid version according to applicable national 
regulatory and legal requirements will be considered. 
The study will also be carried out in accordance with the Integrated Addendum of the ICH 
guideline on Good Clinical Practice E6(R2), the Regulation EU No. 536/2014, which will 
replace the EU Clinical Trial Directive 2001/20/EC, the Commission Directive 2005/28/EC 
(April 8, 2005), and all local laws, regulatory requirements, and guidelines applicable in the 
participating countries.  
By signing the Investigator’s Declaration (Appendix I), the investigator agrees to conduct the 
study as set out in this protocol and in accordance with the moral, ethical and scientific 
principles governing clinical research. 
The study personnel and vendors (i.e. key study personnel) is provided in Appendix II. 
The signatures of sponsor and coordinating investigator can be found in Section 12. 
10.1 Approval procedures 
Before the start of the study, the study protocol, patient information / informed consent form, 
and other relevant documents will be submitted by the investigator or designated CRO to the 
relevant IEC/IRB for all participating sites/countries in accordance with national laws, 
guidelines, and GCP provisions. Written IEC approval must be obtained prior to patient 
enrolment.  
The relevant local authorities will be notified/asked for approval of the intended study in 
accordance with local legal requirements. 
10.2 Protocol amendments 
After initiation of the study, any change in this protocol will require a formal amendment. The 
amendment must be signed by all of the signatories to the original protocol. Once the study 
has started, amendments will be made only in exceptional cases. 
If ethically relevant aspects are concerned, the IEC/IRB will be informed of amendments and 
approval will be sought. All protocol amendments will be submitted to the regulatory authorities 
as regulated by national law. 
Changes to the protocol may only be implemented after all appropriate requirements listed 
above (ethics, regulatory approval of all responsible personnel) have been fulfilled. 
10.3 Informed consent 
The procedure of informed consent will be carried out in accordance with all applicable laws 
and regulations. The purpose and nature of the study, possible risks and benefits of IMP 
administration, planned procedures and potential discomforts, and usage, transfer, and 
disclosure of personal data and study-related health data will be explained in writing in a patient 
information sheet, which will be handed to the patient by the investigator, or an authorised 
designee. In addition, the investigator, or an authorised designee, will verbally provide all 
relevant information to the patient. Sufficient time will be allowed to discuss any questions 
raised. The patient will be made aware that participation in this study is voluntary and that 
he/she can withdraw their consent to participate at any time without giving a reason and without 
having to fear any disadvantageous effects on his/her current or future medical care. The 


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investigator will inform the patient that in providing informed consent, he / she is giving 
permission for representatives of the sponsor, ethic committees, or competent authorities to 
inspect their medical records for data verification. Patients will be informed that all personal 
information made available for inspection will be handled in the strictest confidence and in 
accordance with all applicable data protection laws and regulations. 
Only after written and verbal information has been provided, can consent for participation be 
given by the patient by personally signing and dating the approved informed consent form. The 
signed documents must be retained by the investigator as part of the study records. A copy of 
the signed informed consent form will be given to the patient. 
The investigator will not undertake any procedures required for this study until informed 
consent has been obtained. The date of obtaining informed consent will be entered in the 
eCRF.  
After releasing an amendment to the protocol, that might influence the patient's decision for 
participation, the patient information sheet and the informed consent form must be amended 
accordingly. Revised informed consent sheets must be submitted to the relevant IECs/IRBs 
and the relevant authority as required by local law. In addition, any relevant new information 
about the IMP that becomes available during the study will be passed on to the investigators 
and the patients. Depending on the nature of the amendment, it might be necessary for patients 
who are already enrolled to confirm their informed consent on the basis of the new information. 
10.4 Confidentiality and data protection 
All relevant provisions of the European General Data Protection Regulation (GDPR [EU] 
2016/679, 27 April 2016) and all local legal requirements regarding the protection of personal 
data will be adhered to. 
The anonymity of trial patients will be maintained. Patients’ names will not be supplied to the 
sponsor or designated CRO. Patients will be identified on all eCRFs, image media and other 
documents by a specific patient identification (screening) number. Documents that identify the 
patient (e.g. the signed informed consent) must be maintained in confidence by the 
investigator. Sponsor, CRO, central laboratory, and off-site readers will only be given 
pseudonymised data, laboratory samples, or images. 
The patients will be informed about all media used in this study for documentation, storing, and 
transferring the patients’ study findings and will be assured that all data will be handled in the 
strictest confidence. 
This study protocol, any other unpublished documentation (e.g. eCRF), any information regarding 
the IMP (e.g. investigator’s brochure), and any results derived from the study will be regarded as 
confidential. The investigators and study site members will not be allowed to disclose such 
information without prior written approval from the sponsor. 
10.5 Liability, insurance and finances 
The sponsor has taken out appropriate third-party liability insurance coverage in accordance 
with all local legal requirements in the respective country in which the study is performed. It 
covers the eventuality that personal injury may be caused by using the IMP or by any trial-
specific procedure carried out according to this protocol.  
The general insurance conditions will be kept in the investigator's file and will be made 
available for patients at any time. 


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The financial agreements for each site (e.g. Clinical Trial Agreement) are addressed in one or 
more documents. The parties must sign the agreement before each site is initiated. All 
investigators and other relevant site staff participating in this study must complete a Financial 
Disclosure Form. 
10.6 Clinical study report, publication and use of study results 
An integrated study report according to the standards of the ICH E3 guideline, covering clinical 
and biometrical aspects, will be prepared by the sponsor or its designee within 1 year of the 
end of the study (last protocol-defined contact with any enrolled patient). The report will be 
reviewed and approved by the coordinating investigator. 
A publication policy will be prospectively defined before the start of the study. Any results 
derived from this study may be published in a scientific journal or presented at a scientific 
meeting with the sponsor's consent. The sponsor will be provided with a copy of the manuscript 
for review and approval prior to any such submission. 
The sponsor will make the protocol and results of this study, publicly available on the internet 
at www.clinicaltrials.gov and in the European Union Drug Regulating Authorities Clinical Trials 
(EudraCT) database. Upon completion of the study and when the study results are available, 
the patient has the right to be informed by the investigator about the overall study results. 
10.7 Archiving of study documents 
Essential documents as listed in ICH GCP E6(R2), Section 8, shall be archived safely and 
securely in such a way that they are readily available upon authorities’ request.  
Copies of the protocol, patient information sheet / informed consent form, patient identification 
list, a printout of eCRF (or e.g. digital media with the eCRF), the patient record with all original 
data and original images, and all other documents pertaining to this study will be retained at 
the study site for a minimum of 25 years after the completion and approval of the integrated 
study report or for a period that is in accordance with national regulatory requirements, 
whichever is longest. . The investigator site file is not to be destroyed without the sponsor’s 
approval. Ascelia Pharma or a designee will remind the investigator in writing of this obligation 
when the clinical study report synopsis is distributed to the site. 
The final integrated study report must be retained by the sponsor for 5 years beyond the 
lifetime of the IMP. 
 


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11 LIST OF REFERENCES 
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study. Arch Pathol Lab med. 2008;132(6):931-9. 
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2016;15(4):e183-92. 
3. Yoo TG, Cranshaw I, Broom R, Pandanaboyana S, Bartlett A. Systematic review of 
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4. Uggeri F, Ronchi PA, Goffredo P, et al. Metastatic liver disease from non-colorectal, 
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5. Singal AG, Pillai A, Tiro J. Early detection, curative treatment, and survival rates for 
hepatocellular carcinoma surveillance in patients with cirrhosis: a meta-analysis. PLoS 
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7. Namasivayam S, Martin DR, Saini S. Imaging of liver metastases: MRI. Cancer 
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8. Imam K, Bluemke DA. MR imaging in the evaluation of hepatic metastases. Magn 
Reson Imaging Clin N Am. 2000;8(4):741-56.  
9. Chabanova E, Logager VB, Moller JM, Thomsen HS. Manganese based MR contrast 
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10. Balci NC, Semelka RC. Contrast agents for MR imaging of the liver. Radiol Clin North 
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11. Reimer P, Schneider G, Schima W. Hepatobiliary contrast agents for contrast-
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13. Thomsen HS, Svendsen O, Klastrup S. Increased manganese concentration in the liver 
after oral intake. Acad Radiol. 2004;11(1):38-44. 
14. Chabanova E, Thomsen HS, Løgager V, et al. Effect of new manganese contrast agent 
on tissue intensities in human volunteers: comparison of 0.23, 0.6 and 1.5 T MRI, a 
part of a phase I trial. MAGMA. 2004;17(1):28-35.  
15. Dekker H, Barentsz J, Hoogeveen Y. Oral manganese as contrast medium for MR 
imaging Detection of liver metastases: a phase II clinical study. Abstract SSG08-03. 
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16. Rief M, Huppertz A, Asbach P, et al. Manganese-based oral contrast agent for liver 
magnetic resonance imaging: evaluation of the time course and dose response of liver 
signal intensity enhancement. Invest Radiol. 2010;45(9):565-71. 


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17. Brismar TB, Kartalis N, Kylander C, Albiin N. MRI of colorectal cancer liver metastases: 
comparison of orally administered manganese with intravenously administered 
gadobenate dimeglumine. Eur Radiol. 2012;22(3):633-41. 
18. Albiin N, Kartalis N, Bergquist A, Sadigh B, Brismar TB. Manganese chloride 
tetrahydrate (CMC-001) enhanced liver MRI: evaluation of efficacy and safety in 
healthy volunteers. MAGMA. 2012;25(5):361-8. 
19. Ascelia Pharma AB. Investigator’s Brochure: Mangoral. Edition 11. 
20. Jørgensen JT, Rief M, Brismar TB, Wagner M, Albiin N. A new manganese-based oral 
contrast agent (CMC-001) for liver MRI: pharmacological and pharmaceutical aspects. 
Acta Radiol. 2012;53(7):707-13. 
21. Schlaudecker JD, Bernheisel CR. Gadolinium-associated nephrogenic systemic 
fibrosis. Am Fam Physician. 2009;80:711-14. 
22. Crossover J, Zheng W. Manganese toxicity upon overexposure. NMR Biomed. 2004; 
17(8):544-53. 
23. Lu L, Zhang LL, Li GJ, Guo W, Liang W, Zheng W. Alteration of serum concentrations 
of manganese, iron, ferritin, and transferrin receptor following exposure to welding 
fumes among career welders. Neurotoxicology. 2005;26(2):257-65. 
24. Hauser RA, Zesiewicz TA, Rosemurgy AS, Martinez C, Olanow CW. Manganese 
intoxication and chronic liver failure. Ann Neurol. 1994;36(6):871-5. 
25. Crossgrove J., Zheng W. (2004). Manganese toxicity upon overexposure. NMR in 
Biomedicine, Vol. 17, 544-553. 
26. Kirchin, M.A., Runge, V.M. (2003). Contrast agents for magnetic resonance imaging: 
safety update. Top Magn Reson Imaging. Vol. 14, 426-35. 
27. Casarett and Doull’s Toxicology: The basic science of poisons. (1995). Edited by 
Klaassen, C.D., Amdur, M.O and Doull, J. 5th Edition Published by McGraw-Hill. 
28. Fitsanakis VA, Zhang N, Garcia S, Aschner M. Manganese (Mn) and iron (Fe): 
interdependency of transport and regulation. Neurotox Res. 2010;18(2):124-31. 
29. Pugh RN, Murray-Lyon IM, Dawson JL, et. al. Transection of the oesophagus for 
bleeding oesophageal varices. Br J Surg. 1973; 60:646. 
30. Child CG, Turcotte JG. The Liver and Portal Hypertension. Philadelphia, WB Saunders 
Co. 1964. 
31. UK Medicines Information (UKMi). What is the Child-Pugh score? Available from: 
https://www.sps.nhs.uk/wp-content/uploads/2014/05/UKMI_QA_What_is_the_Child-
Pugh_score_update_May_2017.docx. 
32. FDA. Guidance for Industry – Pharmacokinetics in patients with impaired hepatic 
function: study design, data analysis, and impact on dosing and labeling. May 2003. 
33. Levey AS, Bosch JP, Lewis JB, et al. A more accurate method to estimate glomerular 
filtration rate from serum creatinine: a new prediction equation. Modification of Diet in 
Renal Disease Study Group. Ann Intern Med. 1999;130:461-70. 
34. Couinaud C. Le foie, etudes anatomiques et chirurgicales. Masson, 1957. 


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35. St-Amant M, Jones J. Couinaud classification of hepatic segments. Available from: 
https://radiopaedia.org/articles/couinaud-classification. 
36. Zoni S, Albini E, Lucchini R. Neuropsychological testing for the assessment of 
manganese neurotoxicity: A review and a proposal. Am J Ind Med. 2007;50(11):812–
30. 
37. Da Silva CJ, da Rocha AJ, Jeronymo S, et al. A Preliminary Study Revealing a New 
Association in Patients Undergoing Maintenance Hemodialysis: Manganism 
Symptoms and T1 Hyperintense Changes in the Basal Ganglia. Am J of Neuroradiol. 
2007;28(8):1474-9. 
38. Uchino A, Noguchi T, Nomiyama K, et al. Manganese accumulation in the brain: MR 
imaging. Neuroradiology. 2007;49:715-20. 
39. Koh DM, Brown G, Riddell AM, et al. Detection of colorectal hepatic metastases using 
MnDPDP MR imaging and diffusion-weighted imaging (DWI) alone and in combination. 
EurRadiol. 2008;18(5):903-10. 
40. World Medical Association (WMA). Ethical Principles for Medical Research Involving 
Human Subjects. Declaration of Helsinki, adopted in 1964 by the 18th Assembly, 
amended by the 29th, 35th, 41st, 48th, and 52nd WMA General Assemblies and the 
Note of Clarification on Paragraph 29 added by the 53th WMA General Assembly, 
Washington 2002, the Note of Clarification on Paragraph 30 added by the 55th WMA 
General Assembly, Tokyo 2004, the 59th WMA General Assembly, Seoul 2008, and 
the 64th WMA General Assembly, Fortaleza, Brazil, October 2013. 
 
 






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13 APPENDICES 
I 
Declaration by the Investigator  
Kept separate 
II 
Study personnel and vendors (i.e. key study personnel) 
Kept separate 

SAP_001.pdf: 
Protocol Number: ASC-Man-P016 
 
Official Title: A multicentre, open-label study to evaluate the safety and diagnostic efficacy of 
mangoral in patients with known or suspected focal liver lesions and severe renal impairment. 
 
NCT Number: NCT04119843 
 
Document Date: 02 June 2023 






Statistical Analysis Plan (SAP) 
Version Date: 02-Jun-2023 
 
Sponsor: Ascelia Pharma AB 
 
Protocol No: ASC-Man-P016 version 7.0 
PRA-QMS-02745 9.0 
Page 3 of 34 
Strictly Confidential.  © 2024, PRA Health Sciences.  All rights reserved. 
 Table of Contents 
 Approvals ................................................................................................................................................ 1 
 Change History ....................................................................................................................................... 2 
 Table of Contents .................................................................................................................................... 3 
 Abbreviations .......................................................................................................................................... 5 
 Purpose ................................................................................................................................................... 7 
 Scope ...................................................................................................................................................... 7 
 Introduction ............................................................................................................................................. 7 
7.1 Changes from Protocol .................................................................................................................... 7 
 Study Objectives ..................................................................................................................................... 8 
8.1 Primary Objective ............................................................................................................................. 8 
8.2 Secondary Objectives ...................................................................................................................... 8 
 Study Design ........................................................................................................................................... 9 
9.1 Overview .......................................................................................................................................... 9 
9.2 Parameters .................................................................................................................................... 10 
9.2.1 Visualization of lesions ........................................................................................................ 10 
9.2.2 Primary Efficacy Parameter ................................................................................................. 11 
9.2.3 Secondary Efficacy Parameters .......................................................................................... 11 
9.2.4 Safety Parameters ............................................................................................................... 12 
9.2.5 Pharmacokinetic parameters ............................................................................................... 16 
9.3 Sample Size Considerations .......................................................................................................... 16 
9.4 Randomization ............................................................................................................................... 18 
 General Statistical Considerations ...................................................................................................... 18 
10.1 Descriptive Statistics .................................................................................................................... 18 
10.2 Inferential Statistics ...................................................................................................................... 19 
10.3 Analysis Populations .................................................................................................................... 19 
10.4 Protocol Deviation ........................................................................................................................ 20 
 Conventions and Derivations .............................................................................................................. 21 
11.1 Data Handling .............................................................................................................................. 21 
11.1.1 Imputation of missing data ................................................................................................. 21 
11.1.2 Handling of dates and times .............................................................................................. 21 
11.1.3 Representation of Missing Data ........................................................................................ 21 


Statistical Analysis Plan (SAP) 
Version Date: 02-Jun-2023 
 
Sponsor: Ascelia Pharma AB 
 
Protocol No: ASC-Man-P016 version 7.0 
PRA-QMS-02745 9.0 
Page 4 of 34 
Strictly Confidential.  © 2024, PRA Health Sciences.  All rights reserved. 
11.2 Definitions and Derived Variables ................................................................................................ 21 
11.2.1 Definitions .......................................................................................................................... 21 
11.2.2 Visualization of focal liver lesions ...................................................................................... 23 
11.2.3 Confidence in lesion detection ........................................................................................... 23 
11.2.4 Confidence in lesion localization ....................................................................................... 23 
11.2.5 Quantitative assessments ................................................................................................. 24 
11.3 Statistical Software ...................................................................................................................... 24 
 Interim Analyses .................................................................................................................................. 24 
 Statistical Methods .............................................................................................................................. 25 
13.1 Patient Disposition ....................................................................................................................... 25 
13.2 Protocol Deviations ...................................................................................................................... 25 
13.3 Demographic Data and Baseline Characteristics ........................................................................ 25 
13.4 Medical History and Signs and Symptoms .................................................................................. 25 
13.5 Prior and Concomitant Medications ............................................................................................. 26 
13.6 Investigational Medicinal Product Compliance and Accountability .............................................. 26 
13.7 Efficacy Analysis .......................................................................................................................... 26 
13.7.1 Primary Efficacy Analysis .................................................................................................. 26 
13.7.2 Secondary Efficacy Analysis ............................................................................................. 27 
13.7.3 Subgroup Analyses............................................................................................................ 27 
13.7.4 Pharmacokinetic Analysis .................................................................................................. 28 
13.8 Safety Analysis ............................................................................................................................ 28 
13.8.1 Adverse Events .................................................................................................................. 28 
13.8.2 Vital Signs .......................................................................................................................... 29 
13.8.3 Physical Examination and Neurological Assessments ...................................................... 29 
13.8.4 12-Lead ECG ..................................................................................................................... 29 
13.8.5 Laboratory Parameters ...................................................................................................... 30 
 References .......................................................................................................................................... 30 
 APPENDIX .......................................................................................................................................... 31 
15.1 Data derivation and analysis rules ............................................................................................... 31 
15.1.1 General specifications ....................................................................................................... 31 
15.1.2 Disposition ......................................................................................................................... 31 
15.1.3 Demographics and other baseline characteristics ............................................................. 31 


Statistical Analysis Plan (SAP) 
Version Date: 02-Jun-2023 
 
Sponsor: Ascelia Pharma AB 
 
Protocol No: ASC-Man-P016 version 7.0 
PRA-QMS-02745 9.0 
Page 5 of 34 
Strictly Confidential.  © 2024, PRA Health Sciences.  All rights reserved. 
15.1.4 Safety analysis ................................................................................................................... 32 
15.2 Statistical output documentation .................................................................................................. 34 
 
  Abbreviations 
AE 
Adverse event 
ATC 
Anatomic therapeutic chemical classification 
BD 
Border delineation 
CI 
Confidence interval 
CMRI 
Combined mangoral-enhanced and unenhanced MRI 
CNR 
Contrast-to-noise ratio 
CRF 
Case report form 
CRO 
Clinical research organization 
DWI 
Diffusion-weighted imaging 
GCP 
Good clinical practice 
FAS 
Full analysis set 
HCC  
Hepatocellular carcinoma  
ICH 
International conference on harmonization 
IMP 
Investigational medicinal product 
LC 
Lesion contrast compared to liver background (qualitative) 
LLC 
Liver-to-lesion contrast (quantitative) 
LLOQ 
Lower limit of quantification 
Max 
Maximum 
MedDRA 
Medical dictionary for regulatory activities 
MeMRI 
Mangoral-enhanced MRI 
Min 
Minimum 
MRI 
Magnetic resonance imaging 
N 
Number of non-missing observations 
Nmiss 
Number of missing observations 
PK 
Pharmacokinetics 
PPS 
Per protocol set 
PT 
Preferred term 


Statistical Analysis Plan (SAP) 
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SAE 
Serious adverse event 
SAF 
Safety analysis set 
SAP 
Statistical analysis plan 
SAS 
Statistical analysis system 
sBD 
Sum of scores of border delineation 
SD 
Standard deviation 
SI 
Signal intensity 
sLC 
Sum of scores of lesion contrast compared to liver 
background (qualitative) 
SOC 
System organ class 
SOP 
Standard operating procedure 
TFLs 
Tables, Figures, Listings 
UnMRI 
Unenhanced MRI 
V 
Visit 
WHO 
World health organization (United Nations) 
WHO-DD 
World health organization drug dictionary 


Statistical Analysis Plan (SAP) 
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 Purpose 
The Statistical Analysis Plan (SAP) describes the statistical methods to be used during the reporting and 
analyses of data collected under Ascelia Pharma AB Protocol ASC-Man-P016 version 7.0 dated 12-Oct-
2021. 
 Scope 
The Statistical Analysis Plan outlines the following: 
 
Study Objectives 
 
Study Design 
 
Study Endpoints 
 
Applicable Study Definitions 
 
Statistical Methods. 
 Introduction 
This Statistical Analysis Plan (SAP) is based on the relevant sections of study protocol. Tables, Figures 
and Listings (TFLs) Specifications are described in a separate document (TFL Shells). The purpose of the 
SAP is to describe in more detail how the analyses are to be performed and presented. It quotes the relevant 
statements directly from the protocol, but it does not give a full description of study design etc. 
The TFL Shells specify the output to support the analysis. The TFL Shells are considered as a guideline 
for producing the output.  
The SAP will be finalized prior to database lock or any unblinding of study team members. 
The investigational medicinal product (IMP) is mangoral (previous candidate name: CMC-001) containing 
manganese (II) chloride tetrahydrate (MnCl2 4H2O) combined with L-alanine and vitamin D3. It will be 
referred to hereafter as mangoral in this document. Mangoral is for diagnostic use and for single dose 
administration only. 
The aim of this study is to evaluate the safety and diagnostic efficacy of mangoral in patients with known or 
suspected focal liver lesions and severe renal impairment. 
7.1 Changes from Protocol 
The planned analysis will be performed according to the study protocol, its amendments and this statistical 
analysis plan. If there are contradictions between the study protocol or its amendments and this statistical 
analysis plan, the analysis will be performed according to this analysis plan. Any deviation from the planned 
analysis according to the study protocol has to be described in the integrated report. 
The protocol states that the per protocol set is all patients in the full analysis set without any important 
protocol deviations. The SAP has been revised to state that only a subset of important deviations will result 
in patients being excluded from the per protocol set. All important protocol deviations will be manually 
reviewed by ICON and Ascelia to agree which could have impact on the per protocol set. 
The protocol mentions the requirement for adverse drug reactions (defined per protocol as a causal 
relationship between a medicinal product and an AE to be at least a reasonable possibility) to be collected 


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and summarized. However, in this study adverse events are only assessed as related or not related so this 
will be used for the outputs. 
The wording and definition of the PK dialysis subgroup population is amended. 
The protocol Section 8.1.5 mentions subgroup analysis to be performed on the co-primary variables to be 
done by status of liver disease, on the subgroup of patients with PK data and also subgroup of patients with 
brain MRI, this is not included in the SAP as the patient groups are considered to be too small for a 
meaningful comparison. Subgroup analysis on the co-primary variables by sex will be added. Additionally 
subgroup analyses is mentioned in the protocol for by lesion diameter on the primary endpoint, this is not 
planned but different subgroup analyses is planned as described in Section 13.6.3.  
The derivations for liver-to-lesion contrast and contrast to noise ratio are amended slightly as described in 
Section 11.2.5. 
QTcB interval (msec) and QTcF interval (msec) were derived in the database but issues were found with 
these calculations, for this reason these parameters were re-derived in the datasets per the derivations in 
Section 11.2.1. 
 Study Objectives 
8.1 Primary Objective 
To assess the diagnostic efficacy of mangoral in liver MRI in terms of visualization of detected focal liver 
lesions in combined MRI (CMRI: combined mangoral-enhanced and unenhanced MRI) compared to 
unenhanced MRI. 
8.2 Secondary Objectives 
 
To assess the diagnostic efficacy of mangoral in liver MRI in terms of: 
- 
number of lesions detected by each MRI method (unenhanced MRI, mangoral-enhanced MRI 
and CMRI); 
- 
visualization of detected focal liver lesions in mangoral-enhanced MRI compared to unenhanced 
MRI (determination of visualization will be done in the same way as for the primary efficacy 
variable); 
- 
confidence in lesion detection and localization separately in unenhanced MRI, mangoral-
enhanced MRI and CMRI; 
- 
lesion dimensions (independent off-site readers’ assessment): longest diameter of largest and 
smallest lesion;  
- 
quantitative assessments by measuring percent signal intensity enhancement of liver, liver-to-
lesion contrast, signal-to-noise ratio and contrast-to-noise ratio of up to 5 lesions per patient; 
- 
number of patients who had at least one new lesion identified on CMRI compared to 
unenhanced MRI alone. 
 
To assess a proportion of patients having at least one malignant lesion identified on post-mangoral 
images that was not identified on pre-mangoral images.  
 
To evaluate the safety and tolerability of mangoral; 
 
To evaluate the pharmacokinetics of manganese after a single dose of mangoral in a subgroup of 
patients (including a small number of dialysis patients); 
 
To evaluate the impact of diagnostic performance of CMRI and mangoral-enhanced MRI versus 
unenhanced MRI on the patients’ management. 






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to confirm lesion numbering and lesion location across the modalities. Lesion tracking will also confirm 
lesions that are identified on pre-mangoral, post-mangoral and combined MRI images for inclusion in the 
analysis of the co-primary variables.  
9.2.2 Primary Efficacy Parameter 
The primary efficacy endpoint is the visualization of detected focal liver lesions in combined MRI (CMRI, 
mangoral-enhanced MRI plus unenhanced MRI) as compared to unenhanced MRI.  
The visualization will be assessed using two co-primary variables: 
 
lesion border delineation (BD) 
 
lesion contrast compared to the liver background (LC).  
Both parameters will be determined for each lesion (up to 15 lesions per patient) and by each of the 
3 independent readers by qualitative assessment on 4-point scales with categories ‘poor’ (1), ‘partial / 
moderate’ (2), ‘good’ (3) and ‘excellent’ (4) lesion border delineation/lesion contrast (defined in the Section 
6.2.3.1 of the study protocol). Based on the scores on the 4-point scales, two sum scores will be calculated 
for each patient and separately for each MRI method: a lesion border delineation sum score and a lesion 
contrast sum score. Both scores will be weighted by the number of lesions per patient. 
For primary efficacy endpoint analysis, only lesions that are seen on both pre- and post-contrast (Part I and 
Part II) will be considered and results of individual readers will be reported separately. The matcher will 
match lesions for unenhanced, enhanced and combined MRI. It is possible, that lesions are not seen on an 
unenhanced MRI scan, but become visible on the combined MRI and/or enhanced MRI scan. Since the 
primary endpoint refers to the visualization of the lesion, in this event, these lesions will not be considered 
for the primary efficacy analysis. 
9.2.3 Secondary Efficacy Parameters 
The secondary efficacy variables will be based on the evaluations by independent readers (off-site readers’ 
assessment) and selected endpoints (number of lesions per MRI method, change in patients’ management 
and confidence in lesion detection and localization and change in patients’ management) also by the on-
site radiologist (on-site readers’ assessment) as specified in Table 1. The secondary efficacy endpoints 
assessed off-site will be evaluated using the same number of lesions as the primary endpoint, if not 
specified otherwise. 
9.2.3.1 Number of lesions detected by each MRI method and proportions of patients 
Number of lesions detected by each MRI method (unenhanced MRI, mangoral-enhanced MRI and CMRI) 
per patient will be presented using descriptive statistics.  
In addition, the following proportions of patients will be calculated for assessments performed by 
independent off-site readers: 
- 
The number of patients having at least one lesion identified on post-mangoral images that was not 
identified on pre-mangoral images. 
- 
The number of patients having at least one malignant lesion identified on post-mangoral images that 
was not identified on pre-mangoral images based on patients with at least one malignant lesion at 
baseline. For patients with malignant lesions at baseline any new lesions identified are also assumed 
to be malignant. 
- 
The number of patients who had improvement in at least one co-primary variable for at least one lesion 
on post-mangoral images compared to pre-mangoral images. 


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9.2.3.2 Visualization in mangoral-enhanced MRI vs. unenhanced MRI 
Visualization of focal liver lesions in mangoral-enhanced MRI as compared to unenhanced MRI will be 
assessed in the same way as for the primary efficacy endpoint (by the off-site readers only). 
9.2.3.3 Confidence in lesion detection and localization 
Confidence in lesion detection is defined in Section 11.2.3 and will be assessed separately in unenhanced 
MRI, mangoral-enhanced MRI and CMRI (3-point scale, on-site and independent off-site readers’ 
assessments of up to 15 lesions per patient).  
Confidence in lesion localization is defined in Section 11.2.4 and will be assessed separately in unenhanced 
MRI, mangoral-enhanced MRI and CMRI (3-point scale, independent on-site and off-site readers’ 
assessments of up to 15 lesions per patient).  
The lesions will be listed in ordinal sequence from the smallest to the largest with specification of the liver 
segment in which they have been identified. For assessments provided by on-site readers, all lesions 
detected per MRI method (regardless if they are matched compared to unenhanced MRI) will be included 
in descriptive statistics.  
9.2.3.4 Lesion dimensions  
Lesion dimension measurements (diameter measured in mm) will be done by each of the 3 independent 
readers on mangoral-enhanced MRI and unenhanced MRI images for all lesions. The longest diameter of 
the largest and of the smallest visualized lesions, respectively, will be summarized.  
9.2.3.5 Quantitative assessments  
Quantitative assessments are specified in Section 11.2.5 and will be performed for up to 5 detected lesions 
(independent off-site readers’ assessment). These lesions will be the same for pre- and post-contrast 
assessments: 
- 
liver signal intensity (SI) enhancement (%); 
- 
liver-to-lesion contrast (LLC); 
- 
signal-to-noise ratio (SNR); 
- 
contrast-to-noise ratio (CNR). 
9.2.3.6 Change in patients’ management  
Change(s) in patients’ management based on diagnostic performance of CMRI or mangoral-enhanced MRI 
versus unenhanced MRI (on-site and independent off-site readers’ assessments): 
- 
Any changes in patient management based on MRI findings (yes / no); 
- 
Next steps in patient management based on MRI findings (i.e. chemotherapy, surgery, local 
ablation procedure, combination therapy). 
A patient can have more than one recommended management per MRI method, for this reason all results 
will be included in the listings and the tables. 
9.2.4 Safety Parameters 
This section defines the general principles for the analysis of categorical and continuous parameters, unless 
specified otherwise. In general, the analysis of all parameters will be done by visit/time point and treatment 
group and overall.  


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Categorical parameters will be descriptively summarized per visit/time point and overall post-baseline. For 
an overall post-baseline analysis, the worst value during the on-treatment period, including unscheduled 
visits, will be derived. If applicable, the number of not exact values will be displayed in the respective 
table. Continuous parameters will be descriptively summarized. Changes from baseline will be presented 
for all post-baseline visits/time points. Baseline values are defined as described in Section 11.2.1. 
Where applicable, continuous laboratory parameters will be classified as low, normal, or high based on 
reference (normal) ranges as provided by the central laboratories.  
A listing of patients with values outside the reference ranges will be provided. All parameter values will be 
presented in a patient data listing including respective flagging with respect to reference ranges. 
Unscheduled measurements of parameters will be presented in the patient data listing. In general, 
unscheduled measurements will not be included in the analysis. 
9.2.4.1 Vital signs 
Vital signs will be measured at baseline (within 3 hours prior to contrast administration) and after contrast 
administration at 6 (± 1) hours, 24 (± 4) hours, 48 (± 4) and 5 (± 2) days post-dose: 
• 
Systolic blood pressure [mmHg]; 
• 
Diastolic blood pressure [mmHg]; 
• 
Pulse rate [bpm]; 
• 
Respiratory rate [breaths per minute]; 
• 
Body temperature [°C]. 
The investigator will report any worsening from baseline in vital signs as AE (see Section 6.3.7 of the 
protocol). 
9.2.4.2 Physical examination 
A physical examination will be performed at the following times: baseline (within 3 hours prior to contrast 
administration) and at 6 (± 1) hours, 24 (± 4) hours, 48 (± 4) hours and 5 (± 2) days post-dose. Any 
abnormalities must be specified in the eCRF and recorded as an AE (see Section 6.3.7 of the protocol). 
9.2.4.3 Neurological assessments 
Neurological assessments to monitor for any potential neurotoxic effects will be performed pre-dose 
during the baseline period and 24 (± 4) hours, 48 (± 4) hours and 5 (± 2) days hours after the 
administration of mangoral.  
Results of the assessments will be recorded in the eCRF. All new (i.e. no pre-existing condition) abnormal 
findings observed after mangoral administration must be recorded and reported as AEs (see Section 
6.3.7 of the protocol). 
9.2.4.4 Electrocardiogram (ECG) 
A 12-lead ECG will be performed after 5 minutes supine rest at baseline (within 3 hours prior to contrast 
administration) and after administration of mangoral at 6 (± 1) and 24 (± 4) hours post-dose.  
ECG parameters will be recorded and assessed: overall interpretation, heart rate (beats/min), QT interval 
(msec), RR interval (msec), PR interval (msec), QRS duration (msec), QTcB interval (msec) and QTcF 
interval (msec).  


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The investigator will record on the eCRF whether the results are normal, abnormal (not clinically 
significant or clinically significant). If recorded as abnormal and clinically significant, the abnormality must 
be specified in the eCRF and if observed after administration of mangoral, the abnormality must also be 
recorded as an AE. 
Note: As specified in Section 6.3.7 of the protocol, if electrocardiographic abnormalities observed in a pre-
existing condition are preserved also after the administration of mangoral, these should not be considered 
as AEs. Only if abnormalities are worsening of pre-existing ones, they are considered as AEs. 
9.2.4.5 Laboratory variables 
Details of laboratory parameters are specified in Section 6.3.5 of the protocol. In summary, Table 2 
provides an overview of the screening laboratory tests and Table 3 provides an overview of other 
laboratory parameters measured at baseline and post-dose visits. 
The investigator has to document the clinical significance of abnormal laboratory values in the eCRF. If 
the clinically significant abnormal value is observed after administration of mangoral, the abnormality 
must also be recorded as an AE. 
Table 2 
Screening laboratory parameters 
Child-Pugh: 
(local lab) 
Serum bilirubin 
Serum albumin 
International normalized ratio (INR) 
eGFR: 
(local lab) 
Estimated Glomerular Filtration 
Rate (eGFR) 
Serum creatinine 
Serum albumin 
Serum urea nitrogen (SUN/BUN) 
 


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Table 3 
Clinical safety laboratory parameters (baseline and post-dose) 
Hematology: 
(central lab) 
Hemoglobin 
Hematocrit 
Erythrocytes 
Leukocytes 
Platelets 
Neutrophils 
Eosinophils, 
Basophils 
Lymphocytes 
Monocytes 
INR  
 
Biochemistry: 
(central lab) 
Alkaline phosphatase 
Aspartate aminotransferase (AST) 
Alanine aminotransferase (ALT) 
Gamma-glutamyl transferase (GGT) 
Creatinine 
Glucose 
Urea 
Total Bilirubin 
Albumin 
Sodium 
Potassium 
Calcium 
Magnesium 
Chloride 
Urinalysis: 
(local lab) 
 
pH 
Leucocytes 
Erythrocytes 
Protein 
Glucose 
Urobilinogen 
Ketones 
Bilirubin 
Nitrite 
Specific gravity 
All clinically significant abnormal laboratory values observed at 5 (± 2) days post-dose are to be followed 
up until they have normalized, or until in the investigator’s opinion there is no medical necessity for further 
blood tests.  
9.2.4.6 Blood manganese concentration 
Blood samples for manganese measurements will be taken on the day of mangoral administration 
immediately (within 1 hour) prior to dosing and at 24 (± 4), 48 (± 4) and 5 (± 2) days post-dose in all 
patients. A blood volume of 6 mL will be drawn per time point.  
For patients in the PK subgroup (selected from a few sites) additional blood samples for PK assessments 
will be taken on the day of mangoral administration. Blood samples will be taken on the day of mangoral 
administration immediately (1 hour pre-dose) and 0.25, 0.5, 1, 2, 3, 4, 6, 8, 12, 24, 48 and 120 hours 
post-dose.  
9.2.4.7 Pregnancy test 
A serum pregnancy test will be performed in women of childbearing potential at the local laboratory at 
screening. A urine or serum pregnancy test will be performed in women of childbearing potential at the 
local laboratory at baseline, i.e. within 24 hours prior to the administration of mangoral. In the case of 
pregnancy, the patient is not allowed to enter the study or will be withdrawn from further study 
participation. The event is to be captured on dedicated CRF page.  


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9.2.5 Pharmacokinetic parameters 
The aim is to evaluate the pharmacokinetics of manganese in a subgroup of patients for whom blood 
samples were taken after mangoral administration according to a specific sampling schedule. These 
patients will be selected from a few sites that are able to include these patients and follow PK requirements. 
The pharmacokinetic evaluation of manganese will be performed using non-compartmental pharmaco-
kinetic analysis by or under the supervision of the sponsor. Based on the individual blood concentration-
time profiles (using the actual sampling times), the pharmacokinetic parameters given in Table 4 will be 
derived. 
Table 4 
Pharmacokinetic parameters 
Parameter 
Description 
AUC 
Area under the concentration-time curve from zero up to infinity with 
extrapolation of the terminal phase. 
AUC0-120 
Area under the concentration-time curve from zero until time (120 hours).  
This parameter may contain an extrapolated portion if t is replaced by a 
definite time. 
AUC0-24 
Area under the concentration-time curve from zero until time (24 hours).  
This parameter may contain an extrapolated portion if t is replaced by a 
definite time. 
Cmax 
Maximum observed concentration after administration. 
λz 
First order rate constant associated with the terminal elimination phase 
estimated by linear regression of the logarithmic transformed concentration-
time plot. 
R2adj 
Adjusted coefficient of determination of the terminal regression line. 
t1/2,z 
The half-life associated with the terminal elimination phase. 
tmax  
Time to attain Cmax. 
Clapp 
Total apparent clearance calculated as dose/AUC 
LLOQ = lower limit of quantification. 
This analysis will also be repeated considering minimum concentration adjusted results (by patient) as 
below, the new pharmacokinetic parameters will be referred as “adjusted by Cmin“: 
Minimum concentration adjusted = Concentration at timepoint - Minimum concentration  
Further the percentages of:  
(AUC0-24 h adjusted by Cmin / AUC0-24 h unadjusted)*100 
(Cmax adjusted by Cmin / Cmax  unadjusted)*100 
9.3 Sample Size Considerations 
The primary endpoint analysis consists of two co-primary one-sided t-tests at the conventional one-sided 
significance level of 0.025. Following the FDA’s draft guidance on multiple endpoints in clinical studies, 
adjustment of the type-I-error is not necessary since both primary variables need to show superiority 
simultaneously. Nonetheless, this affects the type-II-error. To counteract the inflation of the type-II-error, 
each test’s power is set to 0.9 to achieve a study power of at least 0.9 x 0.9 = 0.81. 


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Effect size 
The primary efficacy is based on qualitative parameters. When designing the ASC-Man-P016 study, a 
conservative approach was applied when calculating the sample size due to uncertainties regarding the 
assumed treatment effect overall and in patients with HCC lesions in particular, and the impact of using 
diffusion-weighted imaging (DWI). 
The original sample size determination was based on a study investigating a manganese-based contrast 
agent with T1 / T2-weighted imaging and DWI. Efficacy was evaluated by comparison of unenhanced 
versus CMRI qualitative score results for “likelihood that a lesion represented a metastasis” on a scale 
from 1 to 5 [1]. These results were used to derive estimates for mean and standard deviation (SD) and 
scaled down for the 1 to 4 scale used in this study. Calculating a mean score per patient and averaging 
readers after rescaling, lead to averages of 2.3 for unenhanced versus 2.6 for CMRI. This is a difference 
of 0.3 with each sample’s standard deviation of ~1.9, or ~1.5 after rescaling. For estimation of the SD of 
the samples’ differences, a high correlation (0.8) was assumed, which resulted in SD of differences of 
0.95. 
Conservatively assuming no effect of the subpopulation of the HCC patients (20% of the study population), 
the difference in the full population was estimated to 0.3 x (1-0.2) = 0.24. 
This resulted in a need of 167 patients for the analysis to reach 90% power for each test. With an 
assumption that only 85% of the image sets would be evaluable (all mangoral-enhanced and unenhanced 
MRI images) and that no patients would drop out (since the MRI procedures are completed within a few 
hours for each patient), it was concluded that 197 enrolled patients were required as per the original sample 
size estimation. 
However, since the original sample size estimation, new information relevant to the underlaying 
assumptions of the effect size has become available. This includes a recent re-analysis of the early phase 
CMC-P004 study [2]. In Study CMC-P004, the MRI images were assessed by 2 investigating radiologists 
in consensus, who were not blinded to the treatment or the clinical information. A re-read of images from 
Study CMC-P004 has been performed by utilizing the 3-reader paradigm that is recommended in the 
guidance for developing medical imaging drugs [3] and is to be used in the present study (Study ASC-Man-
P016). The primary objective of the re-read (P004A) was to estimate the diagnostic efficacy of mangoral 
MRI (combined mangoral-enhanced and unenhanced MRI) and MultiHance® MRI (combined MultiHance®-
enhanced and unenhanced MRI) for visualization of focal liver lesions assessed by 3 independent 
radiologists. The MRI images from Study CMC-P004 were re-assessed according to the procedures 
described in Section 9.2.2. The preliminary analysis showed that the median effect size (CMRI vs. 
unenhanced) was 0.74 (data on file). 
Study CMC-P004 used a dose of 1600 mg manganese chloride tetrahydrate, which is conservatively 
assumed to give a 20% higher effect than the 800 mg dose that is used in the ASC-Man-P016 study 
resulting in an effect size of 0.74 / 1.2 = 0.62. Further, the effect in the subpopulation with HCC lesions is 
assumed to be half the effect seen in the population with metastases and potentially constituting up to 30% 
of the total patient population resulting in the further reduction of effect size to 0.52. The impact of DWI has 
conservatively been assumed to reduce the contrast effect by further 25%. Taken together, an effect size 
of 0.52/1.25=0.42 is considered relevant and has been used for a new estimation of the sample size. 
Sample size 
With the effect size of 0.42, 59 evaluable patients are required to achieve 81% power in a single reader 
test. With the success rule of 2 out of 3, this corresponds to 90% power to meet the co-primary efficacy 
endpoint. With this smaller sample size, a somewhat larger margin to account for non-evaluable patients 


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has been applied. In the original calculation 85% evaluable patients of the total population was assumed, 
but now ∼75% of the total population is assumed to be evaluable. With this assumption, at least 80 patients 
with both unenhanced and mangoral-enhanced MRI will be needed for the primary analysis. 
9.4 Randomization 
No randomization procedure is used, this study is open-label and only has 1 treatment for all patients. 
However, central read data of the liver from the imaging vendor will be masked to ICON 
statistics/programming and Ascelia prior to database lock (ICON data management will be unblinded for 
reconciliation purposes).  
 General Statistical Considerations 
In general, all continuous measures will be summarized descriptively, including number of available values, 
minimum, 1st quartile, median, mean, standard deviation, 3rd quartile and maximum. Categorical data will 
be presented by frequency and percentage. Ordinal ratings may be handled as continuous data. 
10.1 Descriptive Statistics  
The following descriptive statistics will be calculated for continuous data and for ordered categorical data 
(ordinal data): 
 
 


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Summary statistics are displayed with the following digits:  
Description 
Characteristic 
Number of 
decimal places 
Count 
n 
0 
Count corresponding to the number of patient for a treatment 
group 
N 
0 
Mean 
Mean 
As in source + 1 
Standard Deviation 
SD 
As in source + 2 
Minimum, Maximum 
Min , Max 
As in source 
Median 
Median 
As in source + 1 
1st Quartile / 3rd Quartile 
Q1/Q3 
As in source + 1 
Percentage relative to N 
% 
1 
Coefficient of Variation (%) 
CV (%) 
1 
+ Number of decimal places will not be displayed where the count is zero, also if percentage is 100% no decimal 
places will be shown, 
 
For ordered categorical data and nominal data, absolute and relative frequencies (in %) will be calculated: 
• 
Display percentage information only for number of events bigger than zero. 
• 
Percentages will be displayed to 1 decimal place except if it is 100%. 
All data will be presented in the patient data listings.  
If an output does not include any observations, then the following placeholder will be used: “NO DATA 
CONTRIBUTED TO THIS TABLE / LISTING / FIGURE”. 
10.2 Inferential Statistics 
Inferential methods will be based on a significance level of 0.05 for 2-sided tests and 0.025 for 1-sided 
tests. All statistical tests besides the primary efficacy analysis will be interpreted in an exploratory manner. 
The testing of efficacy will be done separately for each of the 3 independent readers and will be based on 
mean scores of BD and LC. For each independent reader, the hypotheses for the superiority of CMRI 
versus unenhanced MRI will be tested using a 1-sided paired t-test comparing mean score differences 
between treatment groups, assuming that the differences are normally distributed. As a sensitivity analysis, 
the non-parametric Wilcoxon signed-rank test will be also performed. 
In all statistical tables, p-values will be reported as specified by the statistical program used, at least up to 
three decimal places. The p-values less than 0.001 will be reported as provided by SAS (e.g. '<0.001').  
10.3 Analysis Populations 
All patients enrolled in the study (fulfil all inclusion criteria, but none of the exclusion criteria and have 
been included in the clinical study at Visit 2) will be included in the safety analysis (safety population). The 
efficacy analysis will be performed in the full analysis set of patients (i.e. who have taken a single dose of 
the investigational medicinal product) and on a ‘per protocol’ basis, see definitions below (Table 5).  


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Table 5 
Definition of analysis populations 
Population 
Description 
Screened Patients 
All patients with date of informed consent not missing. 
Safety population (SAF) and 
enrolled patients 
All patients enrolled in the study (fulfil all inclusion criteria, but none of the 
exclusion criteria and have been included in the clinical study at Visit 2). 
Dosed Patients 
All patients of the safety population who received the IMP. 
Full analysis set (FAS) 
All patients of the safety population who received the IMP and for whom the 
primary efficacy variable is assessable, i.e. all unenhanced / enhanced liver 
MRI images are assessable. 
Per protocol set (PPS) 
The ‘per protocol’ set defines the subset of the patients in the FAS without 
important protocol deviations that could affect the study objectives. All 
important protocol deviations will be manually reviewed by ICON and 
Ascelia to agree which to exclude from PPS. 
PK population (PKS) 
Defines the subset of patients in the FAS who have a blood sample taken 
at a PK timepoint (0.5, 1, 2, 3, 4, 6, 8 or 12 hours post-dose). These 
patients will be selected from a few sites that are able to include these 
patients and follow PK requirements. 
Dialysis subgroup population (DSP) 
All in the FAS currently on maintenance hemodialysis. 
Brain MRI subgroup population 
(BMSP) 
All patients in the FAS with brain MRI performed within 6 months prior to V3 
who underwent the optional brain MRI 7 (+2) days after mangoral-
enhanced MRI of the liver. 
 
Demographic data and baseline characteristics will be displayed for the SAF, FAS and PPS populations. 
Efficacy data will be displayed for the FAS and the PPS populations. Safety data will be displayed for the 
SAF population. Manganese blood concentration data will be displayed for the FAS population, but PK 
parameters data will only displayed for the PKS population. 
10.4 Protocol Deviation 
The relevant protocol deviations have to be defined by a systematic data review prior to database closure 
and unblinding. For this purpose, protocol deviations that occurred during the study such as deviations of 
inclusion/exclusion criteria or forbidden concomitant medications or patient non-compliance will be 
assessed as ‘important’ depending on their potential to interfere with the primary endpoint and the safety 
endpoints of the study. Listings will be prepared to show the eligibility of all patients. Comprehensive 
justification for the classification of a protocol deviation as “important” will be given in the integrated 
clinical study report. 
Important protocol deviations and the assessment of analysis sets will be defined during last data review 
before database closure. All definitions given in the Minutes of the Final Data Review will be taken into 
account in the analysis.  
The list of protocol deviations will be reviewed by the Medical Director, discussed with and approved by 
sponsor and finalized before locking the database. The sponsor will identify important protocol deviations 
which will lead to the exclusion of patients from the per protocol set (PPS). Important protocol deviations 
related to study objectives for patients excluded from PPS will be flagged in the protocol deviation listing. 


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 Conventions and Derivations 
11.1 Data Handling  
11.1.1 Imputation of missing data 
There will be no imputation of missing data. All data will be analyzed as they appear in the database. 
Missing data will be displayed in patient data listings and will be declared in tables as appropriate. 
11.1.2 Handling of dates and times 
• 
Adverse events with unknown onset date/time will be counted as treatment emergent AEs. 
• 
Adverse events with unknown end date/time will be counted as an ongoing AEs, in addition to 
those marked as ongoing in the CRF. 
• 
Adverse events with unknown relationship to IMP will be counted as related in AE summary 
tables. 
• 
Any adverse events with missing seriousness will be counted as serious in AE summary tables. 
• 
Adverse events with partial onset/end date/time will be treated as specified in Section 15.1.4.1. 
• 
For medications with partial end dates see Section 15.1.3.2. 
11.1.3 Representation of Missing Data 
Times should be printed in the format "HR:MI". "HR" represents the 2--digit hour portion of the time of 24h-
format. "MI" represents the 2-digit minute portion of the time. Both hour and minute portions of time are 
zero padded integer values. Missing time portions should be represented on patient listings as dashes 
("10:--" and/or "--:--").  
11.2 Definitions and Derived Variables 
11.2.1 Definitions 
For the statistical analysis, the different MRI methods will be denoted according to the following table. 
Table 6 
Labeling of MRI methods 
MRI Method 
Short label (for tables etc.) 
unenhanced MRI 
UnMRI 
mangoral-enhanced MRI 
MeMRI 
combined MRI (mangoral-enhanced MRI plus unenhanced 
MRI) 
CMRI 
 
In the following table, the definitions and calculation of derived variables are summarized. 
Table 7 
Further definitions 
Variable / Term 
Definition / Way of calculation 
Baseline 
The last assessment made before the administration of IMP will be used as 
baseline.  


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Variable / Term 
Definition / Way of calculation 
Change from baseline  
at visit  
The difference between a value at time point and the value at baseline 
Study day 
The day of IMP is defined as study Day 1. 
Calculate the study day according to the following rules: 
If date < date of IMP, then study day = date of IMP - Date. 
If date ≥ date of IMP, then study day = Date – date of IMP + 1. 
Age [years] 
Year of informed consent – Year of birth  
Age categories 
>= 18 - < 65 years 
>= 65 - < 75 years  
>= 75 years 
Body Mass Index (BMI) 
BMI [kg/m²] = Weight [kg]
Height [𝑚𝑚]² 
Study duration [days] 
Date of completion/discontinuation – date of informed consent + 1  
Prior medication 
Any medication taken and stopped before the date of administration of 
mangoral. 
If the end date of prior medication is missing or partial see Section 15.1.3.2. 
Concomitant medication  
Any medication taken prior to the administration of mangoral and continuing after 
treatment or started after the date of administration of mangoral.  
If the end date of concomitant medication is missing or partial see Section 
15.1.3.2. 
Adverse events 
All untoward medical occurrences after signing the informed consent. These can 
be adverse events either prior to treatment or after treatment as collected in the 
CRF. 
Time to onset of adverse events 
 
Time to onset of adverse events will be calculated based on the administration 
start date time of IMP based on the imputed value for adverse event start 
date/time.   
Duration of adverse events 
 
Duration of adverse events will be calculated based on the imputed values for 
adverse event start date/time and stop date/time.  
If the duration of the adverse event could not be calculated due to unknown date 
information, the following assessment to categories will be used: 
• 
If the adverse event is marked as “ongoing” in the CRF or the end date 
is missing, the duration will be categorized as “ongoing”. 
• 
Otherwise, the duration category will be set to “missing”. 
Flagging of laboratory 
parameters 
Continuous parameters will be classified as low, normal, or high based on 
reference (normal) ranges. In addition, laboratory parameters are flagged also 
according to CTCAE grading (G) as G1, G2, G3, or G4 
Reproducibility 
A consistency of results obtained when the same imaging is performed at short 
intervals on the same patients using different methods and by different readers. 
QTcB interval (msec) 
QTcB interval (msec) =
QT interval (msec)
(RR interval (sec))1/2 


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Variable / Term 
Definition / Way of calculation 
QTcF interval (msec) 
QTcF interval (msec) =
QT interval (msec)
(RR interval (sec))1/3 
The raw datasets will be in the Study Data Tabulation Model (SDTM) and the analysis datasets will be in 
the Analysis Data Model (ADaM), where ADaM datasets are derived from SDTM datasets. Throughout the 
entire study only one CDISC version will be used, there will be no change even if a higher version is 
available. 
11.2.2 Visualization of focal liver lesions 
Visualization of focal liver lesions will be measured by the two variables ‘lesion border delineation’ (BD) 
and ‘lesion contrast compared to liver background’ (LC). Both variables will be determined by qualitative 
assessment on the following 4-point scales for up to 15 lesions per patient as defined in Section 6.2.3.1 of 
the protocol: 1=‘poor’, 2=‘partial / moderate’, 3=‘good’, 4=‘excellent’.  
For each reader, the two sum scores (sBD and sLC) are determined for each patient by summarizing the 
individual scores over all lesions detected by treatment groups UnMRI (unenhanced MRI), MeMRI 
(mangoral-enhanced MRI) and CMRI (combined mangoral-enhanced MRI plus unenhanced MRI). Both 
scores are weighted by the number of lesions per patient. Mean difference (sum score weighted by 
number of matched lesions) will also be derived for matched lesions: Unenhanced MRI vs. Combined 
MRI and Unenhanced MRI vs. Enhanced MRI. 
The handling of MRI scans with more than 15 lesions is described in specific documents (MRI manual for 
training the investigators on MRI reading procedures and rules and Independent Review Charter) 
prepared for the independent reading process. If there are more than 15 lesions, the reader will start 
counting starting from the smallest lesion to the largest lesion and will stop at 15 lesions. 
11.2.3 Confidence in lesion detection 
Confidence in lesion detection will be evaluated for each lesion during the central reading sessions, i.e. 
part I (unenhanced MRI alone), part II (CMRI) and part III (mangoral-enhanced MRI alone), by 
3 independent readers and by on-site radiologists. Up to 15 lesions per patient will be evaluated on 3-point 
scale: 
1 = 
The lesion is detected with low confidence 
2 = 
The lesion is detected with moderate confidence 
3 = 
The lesion is detected with high confidence. 
11.2.4 Confidence in lesion localization 
Confidence in lesion localization will be assessed for each lesion during the central reading sessions (part I, 
part II and part III) by 3 independent readers and by on-site radiologists. Up to 15 lesions per patient will be 
evaluated on 3-point scale: 
1 = 
The lesion is localized to a liver segment with low confidence 
2 = 
The lesion is localized to a liver segment with moderate confidence 


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3 = 
The lesion is localized to a liver segment with high confidence. 
Note: If a lesion is large enough to be present in more than 1 segment it shall be considered to be in the 
segment in which its center lies. 
11.2.5 Quantitative assessments 
For the quantitative analysis, the signal intensities (SI) of liver parenchyma and liver lesion as well as the 
standard deviation (SD) of the background noise will be determined by each of the three independent 
readers during central reading session part II (paired reading of both unenhanced and mangoral-enhanced 
images). The assessment will be done only during reading session part II as this allows the off-site readers 
to assess the same lesions on unenhanced and enhanced images. Up to 5 lesions per patient of ≥ 2 cm in 
diameter will be evaluated and these lesions will be the same on pre- and post-contrast images. If in a 
patient, all lesions are < 2 cm in diameter, only normal liver SI and background noise will be measured.  
Quantitative SI will be measured by positioning circular regions of interest (ROIs) in a homogenous area in 
the liver and in the assessed liver lesion on the same image. The ROI placed in the lesion should at least 
encompass half of the lesion. ROIs of a constant size will be placed in the same locations on all pre- and 
post-contrast images. SD of the background noise will be measured using the largest possible rectangular 
ROI vertical to the patient's abdomen in the direction of the phase-encoding gradient. 
The following quantitative measures will be determined: 
Signal intensity (SI) enhancement: 
• 
Liver SI enhancement (%) =  
([SIliv post contrast – SIliv pre contrast]/[SIliv pre contrast]) × 100 
Liver-to-lesion contrast (LLC): 
• 
LLC = (SIliv – Mean of SIles)/(SIliv + Mean of SIles) 
Signal-to-noise ratio (SNR): 
• 
SNR = SIliv/SDnoise 
Contrast-to-noise ratio (CNR): 
• 
CNR = (SIliv – Mean of SIles)/SDnoise 
 
with SIliv = signal intensity of the liver, SIles = signal intensity of the lesion, SDnoise = standard deviation of 
the background noise, Mean of SIles is the mean of all lesion signal intensities for a patient. 
11.3 Statistical Software 
All statistical analyses will be performed with SAS®, Version 9.4 or later. Working instructions are printed 
in the highlighted italic text below the shells. 
 Interim Analyses 
No interim analysis is planned. 


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 Statistical Methods 
13.1 Patient Disposition 
Patient disposition will be tabulated with number and percentages of patients enrolled, dosed, completed 
according to the protocol (completion of Visit 7 if part of the brain MRI subgroup population, otherwise 
completion of Visit 6) or discontinued prematurely. The patients who prematurely discontinued the study 
and the reasons for their discontinuation will be presented. The number and percentage of patients 
screened, not enrolled and reasons for non-enrollment will also be presented. The number and percentage 
of patients in each analysis population will also be presented as part of the disposition output. 
13.2 Protocol Deviations 
Protocol deviations will be summarized based on the assessment of major (important) or minor (non-
important) by type for the SAF, FAS and PPS populations. In addition, for the subset of patients with major 
(important) or minor (non-important) deviations we will present the number and percentage who completed 
the study and those who discontinued the study. 
All protocol deviations will be listed. 
13.3 Demographic Data and Baseline Characteristics 
The demographic data and baseline characteristics will be analyzed using descriptive statistics in 
summary or frequency tables for the following parameters for the SAF, FAS and PPS populations: 
• 
Sex 
• 
Age 
• 
Age group (>= 18 - < 65 years, >= 65 - < 75 years and >= 75 years) 
• 
Ethnicity 
• 
Race 
• 
Height (cm) 
• 
Weight (kg) 
• 
BMI (kg/m^2) 
The screening laboratory results will also be summarized for the SAF, FAS and PPS populations showing 
the below: 
• 
Laboratory results related to GFR (eGFR categorization, eGFR value). Serum creatinine, serum 
albumin and serum urea nitrogen values are recorded in the source documents at site for assessing 
GFR but will not be summarized. 
• 
Laboratory results related to Child-Pugh scoring (Child-Pugh score, encephalopathy grade, ascites, 
serum bilirubin, serum albumin and INR) by liver disease based on medical history records. 
Disease history of focal liver lesions will be summarized and also listed. 
13.4 Medical History and Signs and Symptoms 
All general medical history conditions, surgical history and signs and symptoms will be coded in MedDRA 
version 25.0 or higher. General medical history conditions and signs and symptoms will be summarized by 
system organ class and preferred term for the SAF, FAS and PPS populations. The version of the utilized 
dictionary will be presented as part of the provided tables and listings.  


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All medical history, surgical history and signs and symptoms will be listed. Any medical history marked as 
ongoing on the CRF or with a missing end date will be considered as “ongoing”. 
13.5 Prior and Concomitant Medications 
Prior and concomitant medications will be coded according to the World Health Organization (WHO) Drug 
Dictionary (2022MAR GLOBAL B3) or later and summarized frequencies and percentages according to 
ATC level 2 and preferred term for the SAF, FAS and PPS populations. Patients taking the same medication 
multiple times will be counted once per medication. The version of the utilized dictionary will be presented 
as part of the provided tables and listings. 
13.6 Investigational Medicinal Product Compliance and Accountability 
IMP administration will be summarized for the SAF, FAS, PPS and PKS. Frequencies and percentages 
and/or descriptive statistics will be provided for each category. In addition, the number and percentage of 
patients receiving 200 mL of IMP as planned per protocol will be provided. 
13.7 Efficacy Analysis 
13.7.1 Primary Efficacy Analysis 
The primary population for this analysis will be the full analysis set (FAS). The two sum scores (sBD and 
sLC) are determined for each patient (both combined and unenhanced) by summarizing the individual 
scores over matched lesions. Then, the mean score differences between the paired sum scores weighted 
by the number of matched lesions are determined per patient: 
Diff(BD) =
 sBDcombined − sBDunenhanced
Number of lesions
  
Diff(LC) =
 sLCcombined − sLCunenhanced
Number of lesions
  
For each independent reader, the hypotheses for the superiority of CMRI versus unenhanced MRI will be 
tested using a 1-sided paired t-test using the means of paired score differences: 
 
H0;BD: µDiff(BD) ≤ 0 
versus  
H1;BD: µDiff(BD) > 0 
and 
 
H0;LC: µDiff(LC) ≤ 0 
versus  
H1;LC: µDiff(LC) > 0 
. 
If both tests indicate superiority of CMRI at the 1-sided significance level of 0.025, reader success is 
achieved. The corresponding 2-sided 95% confidence intervals will be also calculated. These results will 
be presented in a table together with a summary of the mean scores per reader.  
Additionally, as a sensitivity analysis, the analogous 1-sided Wilcoxon signed rank test will be performed. 
The primary analysis will be also repeated in an exploratory manner for the per protocol set using the 
above-described methodology.  
Superiority of CMRI over unenhanced MRI regarding visualization of focal liver lesions will be analyzed by 
testing the superiority of each of the two co-primary variables separately for each of the three independent 
readers. Reader success will be achieved if the reading results of a reader demonstrate superiority of CMRI 
versus unenhanced MRI for both lesion border delineation and lesion contrast. The acceptance by two out 
of three readers will be considered to be a successful demonstration of efficacy in the study. 


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13.7.2 Secondary Efficacy Analysis 
A comparison of part I (unenhanced MRI alone) and part III (mangoral-enhanced MRI alone) will be 
analyzed in the same way as the primary endpoint.  
All other efficacy variables specified in Section 9.2.3 (and listed in Table 1) will be analyzed using descriptive 
statistics (including 2-sided 95% confidence intervals) for the full analysis population (FAS) and the per 
protocol set (PPS). 
For each reader, the percentage of patients who had improvement in at least one co-primary variable for 
at least one lesion on part II (CMRI) and part III (mangoral-enhanced MRI alone) compared to part I 
(unenhanced MRI) will be calculated. In addition, the percentage of patients who had improvement in both 
BD and LC for at least one lesion on part II (CMRI) and part III (mangoral-enhanced MRI alone) compared 
to part I (unenhanced MRI) will be calculated. Similarly, there will be also calculated percentage of patients 
who had at least one new lesion identified on part II (CMRI) and part III (mangoral-enhanced MRI alone) 
compared to part I (unenhanced MRI). 
Additionally, assuming repeated assessments per patient (for approximately 20 patients) by three 
independent readers, an intra-reader reproducibility will be evaluated separately for readings of MRIs in 
parts I, II and III. Based on a patient-level approach, the intra-reader and inter-reader variability will be 
analyzed using linear mixed-effects models fitted to individual mean scores of border delineation (BD) and 
lesion contrast (LC). Basically, the model will include reader, patient and replicate as a random effects and 
divide the total variability into an intra- and inter-reader component, in case this model does not converge 
we will have patient as a fixed effect instead.  
Considering reader as a random effect, this model is used to express the observed value as the true 
value plus the intra-reader error with inter-reader variance 𝜎𝜎𝐵𝐵
2 = 𝑣𝑣𝑣𝑣𝑣𝑣(𝜇𝜇𝑖𝑖) and intra-reader variance 𝜎𝜎𝑊𝑊
2 =
𝑣𝑣𝑣𝑣𝑣𝑣(𝑒𝑒𝑖𝑖𝑖𝑖). As a relative measure of reproducibility, the intra-reader coefficient of variation (wCV) defined as 
wCV = σW / μ where μ = E(μi) is taken as the mean of true value. The Intraclass Correlation Coefficient 
(ICC) will also be derived as the inter-reader variability divided by the total variability. 
Bland–Altman plots [4] (showing differences versus mean, with mean difference and 95% limits of 
agreement) will be used for graphical presentations of intra- and inter-reader agreements for part I 
(unenhanced MRI) versus part II (CMRI) and part I (unenhanced MRI) versus part III (mangoral-enhanced 
MRI alone) as well. 
For inter-reader plots, separate plots will be shown for BD and LC. For each MRI type, plots will be done 
for each pairwise comparison: reader 2 vs. reader 1, reader 3 vs. reader 1 and reader 3 vs. reader 2. 
For a number of patients BD and LC will be assessed twice by each reader and MRI type. For intra-
reader plots, separate plots will be shown for BL and LC, for each reader and MRI type plots will be done 
against first vs. second read (for those matched lesions). 
13.7.3 Subgroup Analyses 
The primary efficacy endpoint and the secondary efficacy endpoint visualization in mangoral-enhanced MRI 
vs. unenhanced MRI will be also evaluated in the following subgroups of patients: 
‒ 
Subgroups of patients by the MRI scanner’s magnetic field strength 1.5 Tesla and 3 Tesla. 
‒ 
Subgroups of patients by the lesion type (benign, malignant, unknown, malignant: HCC, malignant: 
metastasis) 
‒ 
Subgroups of patients with age < 65 years and ≥ 65 years. 


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‒ 
Subgroups of patients by eGFR: < 15, ≥ 15 to ≤ 30 and > 30 mL/min/1.73 m2. 
‒ 
Subgroups of patients by sex 
These subgroup analyses will be done using the FAS population. Additional subgroup analysis may be 
done by race and/or ethnicity depending on the availability of data. A forest plot will be produced to show 
the comparison of paired differences (mean difference and 95% confidence interval) for the primary analysis 
and subgroup analysis in both BD and LC for combined vs. unenhanced and enhanced vs. unenhanced. 
For lesions diameter (small lesion (diameter < 1 cm); medium lesion (diameter ≥ 1 to ≤ 3 cm); large lesion 
(diameter > 3 cm)), subgroup analyses will not be done for the primary endpoint but for the below outputs 
on FAS: 
- 
Number of lesions detected by MRI method: This will summarize number of lesions by lesion 
diameter category and methods. 
- 
Number of patients with at least one new lesion identified compared to unenhanced MRI: This 
will summarize for patients with new lesions if they are categorized as small, medium or large. 
The denominator for percentages will be number of patients in the FAS. 
13.7.4 Pharmacokinetic Analysis 
The analysis of PK parameters will be performed using the PK Set (PKS). Descriptive and graphical 
methods will be used to summarize the study results. 
The CV and the geometric mean will be additionally included in the descriptive analyses of PK 
parameters, except for tmax. The 2-sided 95% confidence intervals for the geometric means of Cmax, 
AUC0-t and AUC will be constructed. 
Missing data will be treated as such and will not be imputed or replaced in any way if not specified otherwise. 
Individual, mean and overlay blood concentration-time profiles will be plotted on both linear and semi-
logarithmic scales. Individual values and descriptive statistics for blood concentration data will be 
presented by the nominal time point using the Safety Population. 
13.8 Safety Analysis 
The analysis of safety data will be performed using the SAF. Safety data will be summarized using 
descriptive statistics. 
13.8.1 Adverse Events 
Safety analysis will include the investigation of adverse events (AEs), serious adverse events (SAEs), AEs 
related to IMP and SAEs related to IMP.  
The following events shall be reported always as SAEs: 
• 
seizure 
• 
stroke 
• 
cerebral venous thrombosis 
• 
QTcF or QTcB greater than 480 msec 
• 
OTcF or QTcB increase of 60 msec over baseline 
 
For all AEs, a summary containing the following counts of patients, percentages of patients and number of 
events will be presented by pre and post dose (per CRF): 


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 
Number of AEs; 
 
Number of related AEs; 
 
Number of AEs leading to study discontinuation (AE caused the patient to be discontinued from the 
study); 
 
Number of related AEs leading to study discontinuation; 
 
Number of SAEs; 
 
Number of related SAEs; 
 
Number of grade 3 or greater AEs; 
 
Number of grade 3 or greater related AEs; 
 
Number of AEs recovered; 
 
Number of AEs recovering; 
 
Number of AEs with outcome of death. 
Further, all AEs, SAEs, AEs related to IMP and non-serious AEs will be summarized in a frequency table 
by MedDRA system organ class (SOC) and preferred term (PT), presented by pre and post dose (per CRF). 
The version of the utilized dictionary will be presented as part of the provided tables and listings. Tables 
will be sorted by decreasing frequency for SOC then PT within SOC using the overall column. 
A summary of AEs by SOC, PT and maximum CTCAE grade will be presented. 
Additionally, a summary of frequent post dose AEs (experienced by ≥ 5% of dosed patients) by PT will be 
displayed summarizing time to event (hours) and duration of event (hours) with summary statistics. For 
handling partial date/times refer to Section 15.1.4.1. If an AE has no end date (ongoing at end of study) this 
will not be included in the analysis of duration of event.  
All AEs, SAEs (other than death) and AEs leading to study discontinuation will be listed. In addition, a listing 
for AEs leading to death will be presented. Listings of AEs leading to death and SAEs (other than death) 
will be presented on screened patients and other AE listings will be presented on enrolled patients.  
13.8.2 Vital Signs 
Summary statistics for vital signs specified in Section 9.2.4.1 and their changes from baseline will be 
presented. 
All vital signs data will be listed. 
13.8.3 Physical Examination and Neurological Assessments 
Parameters of physical examination (specified in Section 9.2.4.2) and neurological assessments (specified 
in Section 9.2.4.3) there will be summarized using descriptive statistics.  
All physical and neurological examination data will be listed. 
13.8.4 12-Lead ECG 
Summary statistics for 12-lead ECG parameters specified in Section 9.2.4.4 and their changes from 
baseline will be presented.  
Additionally, the number and percentage of patients with maximum QT value in certain ranges per ICH E14 
will be summarized (≤ 450 msec, 450 to 480 msec, 480 to 500 msec and > 500 msec). The number and 
percentage of patients with maximum change from baseline in QT value in certain ranges will be 
summarized (≤ 30 msec, 30 to 60 msec and >60 msec). All ECG data will be listed. 


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13.8.5 Laboratory Parameters 
Summary statistics for laboratory parameters specified in Section 9.2.4.5 will be presented by 
International System of Units overall. Shift-tables for hematology and biochemistry with respect to CTCAE 
v5 will be presented as well. 
For urinalysis parameters, categories such as “positive” and “negative” will be used. The positive category 
could be further classified as positive “+” or borderline positive “(+)”.  
For presentation of laboratory values as out of normal range, the flags are created using the normal 
ranges as provided by the central laboratory. 
Box plots will be provided for hematology and biochemistry parameters showing the reference ranges for 
the parameters. 
All laboratory data will be listed. In addition, all out of range laboratory data will be listed. 
 References 
[1] Koh DM, Brown G, Riddell AM, et al. Detection of colorectal hepatic metastases using MnDPDP MR 
imaging and diffusion-weighted imaging (DWI) alone and in combination. Eur Radiol. 2008;18(5):903-
10. 
[2] Brismar TB, Kartalis N, Kylander C, Albiin N. MRI of colorectal cancer liver metastases: comparison of 
orally administered manganese with intravenously administered gadobenate dimeglumine. Eur Radiol. 
2012;22(3):633-41. 
[3] FDA Guidance for Industry. Developing medicinal imaging drug and biological products, Part 3: Design, 
analysis, and interpretation of clinical studies. June 2004. 
[4] Bland JM, Altman DG: Statistical methods for assessing agreement between two methods of clinical 
measurement. Lancet. 1986; 1(8476):307–10. 
Further references are given in the study protocol. 
 


Statistical Analysis Plan (SAP) 
Version Date: 02-Jun-2023 
 
Sponsor: Ascelia Pharma AB 
 
Protocol No: ASC-Man-P016 version 7.0 
PRA-QMS-02745 9.0 
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 APPENDIX 
15.1 Data derivation and analysis rules 
The purpose of this section is to give technical details for the implementation of the SAP.  
15.1.1 General specifications 
15.1.1.1 Visit windows 
The exact times at which IMP administration and mangoral-enhanced MRI are performed are defined in 
Section 5.1.5 of the protocol. 
15.1.2 Disposition 
All presentations for patient disposition will be done overall. 
15.1.2.1 Patient discontinuation 
Reasons for patient discontinuation as specified in the End-of- study page of the CRF will be used.  
15.1.3 Demographics and other baseline characteristics 
15.1.3.1 Patient demographics 
Age as collected in CRF demographics form will be used.  
15.1.3.2 Prior and concomitant medication 
Prior and concomitant medication is collected as of enrollment in the CRF and described like that in the 
study protocol. For the analysis, the definition as described in the following is used. 
The following rules are used to define the categories “prior” and “concomitant” medication. 
Pre-requisite is a complete date/time of IMP administration (entered or imputed). 
Stop of medication  
Date/time  
Condition 
Category 
Complete date/time is 
available 
Stop date/time is earlier than date/time of first dose of IMP. 
Prior 
Missing month 
Year of stop date is earlier than year of first dose of IMP. 
Prior 
Missing day 
Month/year of stop date are earlier than month/year of first dose of 
IMP. 
Prior 
Missing hours 
Day/month/year of stop date are earlier than day/month/year of first 
dose of IMP. 
Prior 
Missing minutes 
Day/month/year/hours of stop date/time are earlier than 
day/month/year/hours of first dose of IMP. 
Prior 
Otherwise 
Concomitant 
Medication ticked in the CRF as “continuing” will be classified as concomitant. 


Statistical Analysis Plan (SAP) 
Version Date: 02-Jun-2023 
 
Sponsor: Ascelia Pharma AB 
 
Protocol No: ASC-Man-P016 version 7.0 
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15.1.4 Safety analysis 
15.1.4.1 Adverse events 
The result of all imputation strategies (e.g., incomplete start dates of adverse events) and new derived 
information must be stored in the corresponding analysis dataset. 
Handling of missing date information 
The term missing date/time refers to a completely missing date/time or to an incomplete date/time where 
parts are not available, e.g., missing hours.  
Missing start and end date/times will be imputed conservatively, i.e., missing values will be imputed in 
such a way that the duration of the adverse event is considered with the longest possible duration. 
Imputation 
I1: Impute AE start date with: 
A replacement of missing year for AE start information is not foreseen. If needed, this will be considered 
on a case-by-case decision which must be documented. 
a) If year of AE is the same as year of treatment then set start date to the date of treatment, otherwise 
use first calendar day and/or first calendar month. 
Imputation will be done based on the available partial information starting with month and then day: 
Missing date 
Date of treatment 
Imputed date 
2014-Mar 
2014-Mar-XX 
2014-Mar-XX 
2014-Mar 
Not 2014-Mar 
2014-Mar-01 
2014 
2014-XXX-XX 
2014-XXX-XX 
2014 
Not 2014  
2014-Jan-01 
b) First hour and/or first minute. 
Imputation will be done based on the available partial information starting with hour and then minutes:  
Missing time 
Date time of treatment 
Imputed time 
11:-- 
11:XX 
11:XX 
11:-- 
Not 11 
11:00 
--:-- 
Known  
Time of treatment 
--:-- 
Unknown 
00:00 
I2: Impute AE stop date with: 
a) Last calendar day and/or calendar last month. 
Imputation will be done based on the available partial information starting with month and then day. The 
respective last month and day will be chosen for imputation: 
Missing date 
Imputed data 
2014-Mar 
2014-Mar-31 
2014 
2014-Dec-31 
For February, leap years must to be taken into account when calculating the last day in February. 


Statistical Analysis Plan (SAP) 
Version Date: 02-Jun-2023 
 
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Protocol No: ASC-Man-P016 version 7.0 
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b) Last hour and/or last minute. 
Imputation will be done based on the available partial information starting with hour and then minutes. 
The respective last hour of a day and last minute will be chosen for imputation. “23:59” will be considered 
as the last hour/last minute per day. 
Missing time 
Imputed time 
11:-- 
11:59 
--:-- 
23:59 
 
List of deaths 
Death will be identified by the outcome of adverse event if the outcome equals “fatal” or reason for end of 
study is “dead”. 
Patient experiencing a non-serious adverse event 
All patients who had at least 1 non-serious AE will be taken into account regardless of the experience of a 
serious AE.  
15.1.4.2 Laboratory parameters, vital signs and further safety parameters 
The tables will display the number of patients still in the study (n) at the time point/visit. For the overall 
presentation, all post-baseline values on treatment are taken into account.  
For all ordinary levels of the categorical parameter including missing values, both the number and the 
corresponding percentage is displayed.  
For descriptive statistics of continuous parameters, n is the number of patients with recorded values at 
the respective time point/visit. The analysis of changes from baseline (e.g., baseline versus End of 
treatment) is based on patients with non-missing values at both visits; for all other patients, the change is 
missing. 
Unscheduled visits 
Unscheduled visits are time points not planned in the protocol. 
In listings, unscheduled visits will be listed as recorded. All visits will be ordered chronologically including 
the dates of unscheduled visits. 
Unscheduled visits will be incorporated in the overall post-baseline summary in tables. If the date is 
incomplete, but it can be determined whether values were measured in the on-treatment period, they will 
be incorporated in overall post-baseline summaries. Unscheduled visits will be excluded from the per time 
point/visit presentation. 
Not exact values  
Not exact laboratory values such as < x, > x will not be included in the analysis of continuous parameters. 
The frequency of occurrence of not exact values will be displayed in the respective table where 
applicable.  
Handling of not exact values in a categorical analysis is described in the section below (“Values out of 
range”). 


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Version Date: 02-Jun-2023 
 
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Protocol No: ASC-Man-P016 version 7.0 
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Ordering of parameters 
Laboratory parameters, vital signs and further safety parameters will be ordered alphabetically within their 
parameter group (e.g., hematology, clinical chemistry and urinalysis). Time points/visits will be sorted 
chronologically. If changes from baseline are displayed by time point/visit, all visits will be displayed first 
followed by all the changes from baseline. 
15.2 Statistical output documentation 
The outputs of statistical procedures for primary analysis will be included in the statistical output 
documentation.