An Open Label, Phase III, Randomized Controlled, Multicentre Non-Inferiority Trial to Compare Efficacy and Safety of Miltefosine and Paromomycin With SSG and PM Combination for Treatment of Primary Visceral Leishmaniasis (VL) Patients in Eastern Africa
Phase: PHASE3
Conditions: Visceral Leishmaniasis
Sponsor: Drugs for Neglected Diseases

Confidential  
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CLINICAL TRIAL PROTOCOL 
 
An Open Label, Phase III, Randomized Controlled, 
Multicentre Non-Inferiority Trial to Compare Efficacy 
and Safety of Miltefosine and Paromomycin with 
Sodium Stibogluconate and Paromomycin 
Combination for Treatment of Primary Visceral 
Leishmaniasis (VL) Patients in Eastern Africa 
 
 
Short title 
MF/PM Phase III 
Name of product(s) 
Miltefosine (Impavido®) (MF) 
Paromomycin (PM) 
Sodium Stibogluconate (SSG) 
Drug Class 
Alkylphosphocholine for MF 
Aminoglycoside for PM 
Pentavalent antimonial for SSG 
Phase 
Phase III 
Indication 
Primary Visceral Leishmaniasis 
Clinical Trial Protocol 
Number 
DNDi-MILT/PM-01-VL 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential 
Page 2 of 68 
Sponsor 
DNDi (Drugs for Neglected Diseases initiative), Chemin 
Louis Dunant, 15, 1202 Geneva, Switzerland 
Phone: +41 22 906 9230 
Acting through its DNDi Africa office, Tetezi Towers, 3rd 
Floor, George Padmore Road, Kilimani, P.O. Box 21936 - 
00505, Nairobi, Kenya 
Phone: +254 20 3995 000 
Sponsor’s representative: 
Dr Monique Wasunna, Director, DNDi Africa Regional 
Office, Nairobi, Kenya 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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Investigators and Study 
Team 
Investigators: 
Dr Jane Mbui, Centre for Clinical Research, KEMRI, 
Kenya 
Dr Simon Njenga, Centre for Clinical Research, KEMRI, 
Kenya 
Prof Joseph Olobo, College of Health Sciences, 
Makerere University, Uganda 
Dr Patrick Sagaki, Amudat Hospital, Uganda 
Prof Dr AM Musa, Institute of Endemic Diseases (IED), 
Sudan 
Prof Eltahir Awad Gasim Khalil, IED, Sudan 
 
Dr Brima Younis Musa, IED, Professor Elhassan Centre 
for Tropical Diseases, Dooka, Sudan 
 
Dr Rezika Mohammed & Prof Asrat Hailu, University of 
Gondar, Ethiopia 
Dr Koert Ritmeijer, MSF Operational Centre Amsterdam 
(MSF Holland) 
Dr Gabriel Alcoba, MSF Operational Centre Geneva 
(MSF Switzerland) 
Head of Visceral Leishmaniasis Program: 
Dr Fabiana Alves, DNDi, Geneva, Switzerland 
Statisticians: 
Dr Bruno Scherrer, Consultant and 
Dr Thaddaeus Egondi, DNDi Africa Regional Office, 
Nairobi, Kenya 
Clinical Pharmacologist: 
Dr Thomas Dorlo, Antoni van Leeuwenhoek Hospital – 
Netherlands Cancer Institute, The Netherlands 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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Senior Medical Manager: 
Dr Gina Muthoni Ouattara, DNDi Africa Regional Office, 
Nairobi, Kenya 
 
Clinical Project Manager: 
Alexandra Solomos, DNDi, Geneva, Switzerland 
Current Clinical Trial 
Protocol Version / Date 
Draft version 5.0, 13 Mar 2020 
Previous Clinical Trial 
Protocol Version / Date 
Final version 1.0, 20 Feb 2017 
Final version 2.0, 5 Sep 2017 
Final version 3.0, 4 May 2018 
Final version 4.0, dated 22 Jul 2019 
The information contained in this document is confidential. It is to be used by investigators, 
potential investigators, consultants, or applicable independent ethics committees. It is understood 
that this information will not be disclosed to others without written authorisation from DNDi, except 
where required by applicable local laws. 
 
 

 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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TABLE OF CONTENTS 
TABLE OF CONTENTS ................................................................................................. 9 
LIST OF FIGURES & TABLES .................................................................................... 11 
ABBREVIATIONS – GLOSSARY OF TERMS ............................................................ 12 
PROTOCOL SYNOPSIS .............................................................................................. 14 
1. 
BACKGROUND AND STUDY RATIONALE ..................................................... 17 
2. 
STUDY OBJECTIVES AND ENDPOINTS ........................................................ 22 
2.1 Objectives ...................................................................................................................................... 22 
2.1.1. 
Primary Objective ...................................................................................................... 22 
2.1.2. 
Secondary Objectives ............................................................................................... 22 
2.2. 
Study Endpoints ..................................................................................................................... 22 
2.2.1. 
Primary Endpoint ....................................................................................................... 22 
2.2.2. 
Secondary Endpoint(s) .............................................................................................. 23 
3. 
STUDY DESIGN AND STUDY DESIGN RATIONALE ...................................... 23 
3.1. 
Study design............................................................................................................................ 23 
3.2. 
Study duration and duration of subject participation ......................................................... 25 
3.3. 
Rationale of study design ...................................................................................................... 25 
4. 
SELECTION OF SUBJECTS ............................................................................ 26 
4.1. 
Inclusion criteria ..................................................................................................................... 27 
4.2. 
Exclusion criteria .................................................................................................................... 27 
5. 
SCHEDULE OF EVENTS .................................................................................. 29 
5.1 Schedule of PK/PD events - Intensive cohort ............................................................................ 31 
6. 
ENROLMENT PROCEDURES .......................................................................... 32 
7. 
STUDY TREATMENTS ..................................................................................... 32 
7.1. 
DNDi study treatment ............................................................................................................. 32 
7.2. 
Doses and treatment regimens ............................................................................................. 33 
7.2.1 
Test arms ........................................................................................................................ 33 
7.2.2 
Reference arm ................................................................................................................ 37 
7.3. 
Study treatments labelling, packaging ................................................................................. 37 
7.4. 
Accountability ......................................................................................................................... 37 
7.5. 
Storage ..................................................................................................................................... 37 
7.6. 
Blinding and procedures for unblinding .............................................................................. 38 
7.7. 
Concomitant treatments ......................................................................................................... 38 
7.8. 
Rescue treatment .................................................................................................................... 38 
8. 
STUDY ASSESSMENTS ................................................................................... 39 
8.1. 
Timing of Assessments .......................................................................................................... 39 
8.2. 
Baseline Assessments ........................................................................................................... 39 
8.3. 
Assessment of Efficacy .......................................................................................................... 40 
8.3.1 
Clinical assessment of VL ............................................................................................. 40 
8.3.2. 
Parasitological assessment ...................................................................................... 41 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
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8.3.3. 
Clinical Lab assessments related to VL evolution ................................................. 42 
8.3.4. 
Post-kala-azar dermal leishmaniasis (PKDL) .......................................................... 43 
8.4. 
Pharmacokinetics and Pharmacodynamics Assessments ................................................ 43 
8.4.1. 
Pharmacokinetic Assessments ................................................................................ 43 
8.4.1.1. 
Paromomycin pharmacokinetic assessments ........................................................ 44 
8.4.1.2. 
Miltefosine pharmacokinetic assessments ............................................................. 44 
8.4.2. 
Pharmacodynamic Assessments ............................................................................. 45 
8.5. 
Assessment of Safety ............................................................................................................. 45 
8.5.1. 
Laboratory examinations .......................................................................................... 46 
8.6. 
Adverse event definitions and reporting .............................................................................. 46 
8.6.1. 
Adverse Event definition ........................................................................................... 46 
8.6.2. 
Assessment of laboratory abnormalities ................................................................ 47 
8.6.3. 
Serious Adverse Event .............................................................................................. 48 
8.6.4. 
Eliciting Adverse Event information ........................................................................ 49 
8.6.5. 
Management of Adverse Events .............................................................................. 49 
8.6.6. 
Adverse Event reporting period ............................................................................... 49 
8.6.7. 
Adverse Event reporting requirements ................................................................... 49 
8.6.8. 
Grading of Adverse Event severity .......................................................................... 50 
8.6.9. 
Adverse Event causality assessment ...................................................................... 51 
8.6.10. 
Exposure in utero ...................................................................................................... 51 
8.6.11. 
Adverse event follow up............................................................................................ 52 
9. 
WITHDRAWAL CRITERIA ................................................................................ 52 
9.1. 
Rules in case of treatment suspension or interruption ...................................................... 53 
9.2. 
Rules for permanently interrupting study treatment .......................................................... 53 
9.3. 
Subject withdrawal from the study and subject replacement ............................................ 53 
10. 
DATA ANALYSIS AND STATISTICAL METHODS .......................................... 53 
10.1. 
Sample size determination ..................................................................................................... 53 
10.2. 
Definition of study populations included in the analysis ................................................... 54 
10.3. 
Subject Disposition ................................................................................................................ 55 
10.4. 
Baseline ................................................................................................................................... 55 
10.5. 
Treatment Compliance ........................................................................................................... 55 
10.6. 
Efficacy Analysis .................................................................................................................... 55 
10.7. 
Safety Analysis ....................................................................................................................... 58 
10.8. 
PK Analysis ............................................................................................................................. 58 
10.9. 
PD Analysis ............................................................................................................................. 59 
11. 
DATA SAFETY MONITORING BOARD ........................................................... 59 
12. 
QUALITY ASSURANCE AND QUALITY CONTROL PROCEDURES ............. 59 
12.1. 
Investigator’s file .................................................................................................................... 59 
12.2. 
Case report forms (CRFs) ...................................................................................................... 59 
12.3. 
Source documents .................................................................................................................. 60 
12.4. 
Record Retention .................................................................................................................... 60 
12.5. 
Monitoring, audits and inspections ...................................................................................... 60 
12.6. 
Audits and inspections .......................................................................................................... 61 
12.7. 
Data Management ................................................................................................................... 61 
12.8. 
Confidentiality of trial documents and subjects records ................................................... 61 
13. 
PROTOCOL AMENDMENTS ............................................................................ 61 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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14. 
EARLY TERMINATION OF THE STUDY .......................................................... 62 
15. 
ETHICS .............................................................................................................. 63 
15.1. 
Informed consent process ..................................................................................................... 63 
15.2. 
Ethical aspects of subject inclusion and study procedures .............................................. 64 
15.3. 
Ethical aspects of study treatments ..................................................................................... 65 
15.4. 
HIV status and VCT ................................................................................................................. 66 
15.5. 
Patient costs ............................................................................................................................ 66 
16. 
INSURANCE AND LIABILITY ........................................................................... 66 
17. 
REPORTING AND PUBLICATION ................................................................... 66 
18. 
REFERENCES .................................................................................................. 66 
 
 
LIST OF FIGURES & TABLES 
 
Figure 1- Overall study design ................................................................................................................ 26 
 
Table 1 - Daily allometric miltefosine dose for female children based on fat-free mass .................. 34 
Table 2 - Daily allometric miltefosine dose for male children based on fat-free mass ..................... 35 
Table 3 - Standard logarithmic criteria for Visceral Leishmaniasis parasitological diagnosis ....... 42 
 
 
 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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ABBREVIATIONS – GLOSSARY OF TERMS 
AE 
Adverse event 
ALB 
Albumin 
ALT 
Alanine aminotransferase (SGPT) 
AR 
Adverse Reaction 
AST 
Aspartate aminotransferase 
AUC 
Area under the concentration-time curve 
BID (b.i.d.) 
Twice a day 
BMI 
Body Mass Index 
CI 
Confidence Interval 
CIOMS 
Council for International Organizations of Medical Sciences 
Cmax 
Maximum concentration 
CRE 
Creatinine 
CRF 
Case report form 
CTCAE 
Common Terminology Criteria for Adverse Events 
DNDi 
Drugs for neglected diseases initiative 
DSMB 
Data Safety Monitoring Board 
ECG 
Electrocardiogram 
EMA 
European Medicines Agency 
EOT 
End of treatment 
FAS 
Full Analysis Set 
FDA 
Food and Drug Administration 
GCP 
Good clinical practice 
GMP 
Good manufacturing practice 
HED 
Human Equivalent Dose 
HIV 
Human immunodeficiency virus 
IA 
Interim Analysis 
ICH 
International Conferences on Harmonization 
IEC 
Independent ethics committee 
IM 
Intramuscularly 
IMP 
Investigational Medicinal Product 
ITT 
Intention to treat 
IV 
Intravenous 
LC-MS/MS 
Liquid chromatography mass spectrometry 
LEAP 
Leishmaniasis East Africa Platform 
MedDRA 
Medical Dictionary for Regulatory Activities 
MF 
Miltefosine 
MoH 
Ministry of Health 
MUAC 
Mid Upper Arm Circumference 
MSF 
Medecins Sans Frontières (Doctors without Borders) 
NCE(s) 
New Chemical Entity(ies) 
PCR 
Polymerase chain reaction 
PD 
Pharmacodynamic 
PI 
Principal investigator 
PK 
Pharmacokinetics 
PKDL 
Post Kala azar Dermal Leishmaniasis 
PM 
Paromomycin 
PP 
Per Protocol 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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PT 
Prothrombin Time 
QD (q.d.) 
Daily 
RDT 
Rapid Diagnostic Test 
SAE 
Serious adverse event 
SOP 
Standard Operating Procedure 
SSG 
Sodium Stibogluconate 
TBIL 
Total Bilirubin 
TEAE 
Treatment emergent adverse event 
ULN 
Upper limit of normal 
VCT 
Voluntary counselling and testing programme 
VL 
Visceral Leishmaniasis 
WBC 
White blood cell 
WHO 
World Health Organization 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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PROTOCOL SYNOPSIS 
 
Protocol title 
An Open Label, Phase III, Randomized Controlled, Multicentre Non-
Inferiority Trial to Compare Efficacy and Safety of Miltefosine and 
Paromomycin with Sodium Stibogluconate and Paromomycin 
Combination for Treatment of Primary Visceral Leishmaniasis (VL) 
Patients in Eastern Africa 
Short title 
MF/PM Phase III 
Clinical Trial 
Protocol Number 
DNDi-MILT/PM-01-VL 
Sponsor 
DNDi, Chemin Louis Dunant, 15, 1202 Geneva, Switzerland. 
Phone: +41 22 906 9230 
PROTOCOL SUMMARY 
 
Current WHO-recommended treatment for Visceral Leishmaniasis (VL) in Eastern Africa 
is a combination of Sodium Stibogluconate (SSG) and paromomycin (PM) administered 
for 17 days, with an efficacy of 91%. This treatment is far from optimal as it requires 17 
days of two separate painful injections, necessitating patients to be hospitalized during 
the whole treatment period. In addition, the antimonial SSG exhibits life-threatening 
toxicities such as cardiotoxicity, hepatotoxicity and pancreatitis. Therefore, there is an 
urgent need to explore alternatives that are efficacious, safe, ideally of short duration, 
affordable and suitable to be used in remote areas where VL occurs.  
Until new chemical entities are developped, opportunities with currently available 
compounds should be assessed to improve on current treatment options, with the main 
aim to replace the toxic and patient-unfriendly SSG treatment-component.  
Miltefosine (MF) is the only oral drug available for VL treatment. It has been extensively 
used in Asia for VL treatment as monotherapy for 28 days, with satisfactory cure rates 
(>90%). However, MF as monotherapy showed lower efficacy in Eastern Africa (72%, 
95%CI: 60–85%) than in Asia. PK data indicated under-exposure and higher relapse 
rates in children compared to adults with the conventional linear 2.5 mg/kg/day MF dose. 
To overcome this under-dosage in children, an allometrically scaled dosing regimen has 
been developed. This allometric dosage was assessed for paediatric VL in the LEAP 
0714 trial in Kenya and Uganda. The 28 days regimen of MF allometric dose showed a 
cure rate of 90.0% (95% CI: 73.5-97.9%) at 6 months follow up in a population of 30 
patients aged 4 to 12 years. These results showed that the efficacy level could be 
increased in children treated with the allometric dose, reaching similar efficacy observed 
in adults (i.e. 86.2%).   
PM has been well studied in Eastern Africa during the development of the combination 
SSG-PM. PM monotherapy of 20mg/kg/d IM for 21 days showed an overall efficacy at 6 
months of 84.3%. Intramuscular PM can be administered at primary health care level, 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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requires minimal training of health personnel and the drug can be stored at room 
temperature.  
Replacing the toxic SSG with oral MF can bring better safety and a more field-adapted, 
patient-friendly treatment.  
Originally, the study was planned to assess the safety and efficacy of two combination 
regimen of PM and MF (arm 1: PM/MF for 14 days and arm 2: PM for 14 days and MF 
for 28 days) as compared to the standard of care SSG-PM for the treatment of primary 
VL patients in Eastern Africa. Due to unforeseen challenges in recruitment, the study 
design has been amended to prematurely discontinue recruitment in one investigational 
arm in order to complete the trial in an acceptable timeframe and ensure the 
achievement of the objectives.  
Following DSMB review of the available data (meeting held on 7 May 2019), including 
143 study participants enrolled, among which 135 had completed treatment and Day 28 
follow up assessment, and 66 had completed their day 210 follow-up visit, two main 
changes were approved: first, the revision of the exclusion criteria in order to include a 
more VL representative population, and second, to discontinue recruitment in the 
investigational arm 2.  
These modifications in agreement with the DSMB will ensure retention of the 
investigational arm 1, on the basis of 1) shorter treatment duration than current standard 
of care, 2) it can be administered within the hospitalization period, 3) it is a cheaper 
option for the control programs, and therefore from a public health perspective will be a 
potentially more favorable option for patients.  
In this revised protocol version, the analysis remains as planned, but comparison will be 
performed only between one investigational arm (arm 1 MF/PM 14 days) and the 
standard of care (SSG-PM). Data from patients randomized in arm 2 (PM 14 days/MF 
28 days), which recruitment is now stopped, will be summarized in a descriptive analysis, 
but not included in the formal comparative efficacy analysis. 
The current study aims to determine the safety and efficacy of one combination regimen 
of PM and MF as compared to SSG-PM for the treatment of primary VL patients in 
Eastern Africa. This is an open label, Phase III, randomized controlled, parallel arm 
multicentre non-inferiority clinical trial. The investigational arms are: 
• Arm 1: Paromomycin 20 mg/kg/d IM for 14 days combined with oral miltefosine 
allometric dosing BID for 14 days 
• Arm 2: Paromomycin 20 mg/kg/d IM for 14 days combined with oral miltefosine 
allometric dosing BID for 28 days (recruitment is now discontinued in this arm) 
The control arm is the current standard treatment for VL in East Africa: 
• Arm 3: Sodium Stibogluconate 20 mg/kg/day IM/IV combined with Paromomycin 
15 mg/kg/day IM for 17 days 
MF allometric dosing will be calculated according to patient’s weight, height and sex. For 
patients weighing < 30 kg, an easy-to-use table with allometric dosing scheme by weight, 
height and sex will be provided to the investigators to define the exact daily dose to be 
administered. For patients weighing ≥ 30 kg, the allometric dose will correspond to the 
conventional dose in mg/kg. Therefore, patients weighing ≥ 30 to < 45 kg will receive 

Clinical Trial Protocol number (DNDi-MILT/PM-01-VL) 
 
 Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
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100 mg/day and patients ≥ 45 kg will receive 150 mg/day. 
Subjects are hospitalized for 14 days of PM and MF treatment in both arm 1 and arm 2. 
MF treatment starts at the same time as PM treatment. At discharge (on day 15), patients 
allocated to arm 2 used to be instructed to continue MF treatment on an out-patient basis 
until completion of the 28 days treatment. Subjects were receiving clear instructions as 
well as a daily diary to guide them in their treatment schedule at home.  
SSG-PM combination therapy will be administered for 17 days according to routine VL 
treatment guidelines and patients will remain hospitalized for the entire duration of the 
treatment. Patients in all arms will come back for the end-of-treatment assessments visit 
at day 28. 
The target population will be primary VL patients from 4 to 50 years. It is important to 
include a paediatric population which is particularly vulnerable and is a major factor in 
the East African disease burden. The trial will be run in the VL endemic countries at 5 
LEAP sites: Kacheliba in Kenya, Amudat in Uganda, Doka and Um El Kher in Sudan and 
Gondar in Ethiopia, as well as 2 MSF sites: Tabarakallah in Sudan and Abdurafi in 
Ethiopia. 
Revised enrollment target in protocol version 4.0 dated 22 July 2019 is 170 in each study 
arm 1 and 3. The total number of patients included in the trial will be approximately 420, 
taking into account the number of patients randomized to treatment arm 2 before 
discontinuation of this arm (approximately 80 patients). Each patient’s participation in the 
study will be for approximately 7 months. This will consist of baseline assessments, 
treatment period (14 or 17 days) and 6 months follow-up. Recruitment for the entire trial 
is expected to take approximately 32 months assuming that 15 to 30 % of all VL patients 
will meet the eligibility criteria. Therefore, study duration (first patient in to last patient, 
last visit) is expected to take approximately 39 months. Taking into account the analysis 
and reporting period, the study shall last at most 45 months. 
Primary efficacy endpoint will be the cure at day 210, based on clinical examination 
(absence of clinical signs and symptoms of VL and no requirement for rescue treatment 
during the trial). 
Safety assessments will be done through routine monitoring of adverse events. A 
characterization of the nature and frequency of SAEs, AEs that lead to treatment 
discontinuation and overall frequency and severity of AEs from start of the treatment until 
6 months follow-up will be made. 
Pharmacokinetics (PK) profile of PM and MF will be described and parasite clearance in 
each arm as indicated by direct microscopy and quantitative polymerase chain reaction 
(qPCR) will be evaluated as pharmacodynamics (PD) markers of cure. The relationship 
between PK and PD measurements will be further assessed through drug exposure-
response modelling.  
Finally, compliance to MF treatment in an outpatient setting will also be described. If 
proven non-inferior to SSG-PM, the combination of PM and MF is expected to improve 
significantly treatment safety and to reduce pain, while ensuring low cost and suitability 
to be used in remote areas. It will provide the East African region with the first non-
antimony-based effective treatment for VL. 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
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1. Background and Study Rationale 
Visceral leishmaniasis (VL) is a parasitic disease caused by Leishmania. It is fatal if 
not treated. The disease is characterized by fever, weight loss, hepatosplenomegaly, 
lymphadenopathy, anaemia, leucopenia and thrombocytopenia. Globally there are 
200-400,000 estimated new cases of VL occurring annually and 90% of these occur in 
six countries: Brazil, Ethiopia, India, Sudan, Somalia and South Sudan (Alvar et al., 
2012). In Eastern Africa, VL is caused by Leishmania donovani, affecting mainly 
children (Harhay et al., 2011).   
Eastern Africa is expected to become the VL region with the highest burden worldwide 
for global VL control, following significant VL elimination efforts in Southeast Asia. 
WHO revised the recommended VL treatment for Eastern Africa in 2010 from sodium 
stibogluconate (SSG) monotherapy (30 days treatment) to a combination of SSG and 
PM administered for 17 days, following a phase III trial conducted in the region (WHO, 
2010). The efficacy of SSG-PM combination was 91% in the intention to treat 
population using a complete case analysis, which was non-inferior to a 30-day SSG 
monotherapy (Musa et al., 2012; Hailu et al., 2010). This new treatment is an 
improvement over the 30-day SSG monotherapy, but a few important drawbacks and 
limitations still preclude its general use. Although the treatment regimen is shorter, it 
still requires 17 days of two separate painful injections, necessitating patients to be 
hospitalized during the whole treatment period. In addition, there are life-threatening 
toxicities associated with the use of antimony-based treatments such as SSG. These 
include cardiotoxicity, hepatotoxicity and pancreatitis.  
AmBisome® (liposomal amphotericin B) is used as 2nd line drug for rescue treatment 
and for specific target populations such as pregnant women, severe disease or HIV 
co-infection. The need for cold chain, high cost and the administration by well trained 
personnel limit the widespread use of AmBisome® in Eastern Africa. 
Therefore, there is a need to explore alternatives that are efficacious, safe, ideally of 
short duration, affordable and suitable to be used in remote areas where VL occurs. In 
this context, an oral efficacious and safe treatment would be more adapted to field 
conditions and would allow shorter hospitalization time. 
In order to respond to this need, upstream pre-clinical research at DNDi has been 
focused on identification of orally available new chemical entities (NCEs) that meet 
these requirements. Promising NCEs are expected to transition from pre-clinical to 
clinical phase in 2017-2018, bringing new opportunities for innovation in VL therapy. 
However, the full development of a new oral treatment is not expected before 2022-
23. As we wait for the NCEs, new opportunities with currently available compounds 
should be assessed to improve on current treatment options, with the main aim to 
replace the toxic and patient-unfriendly SSG treatment-component.  
In Asia, a 10-days combination of PM with MF has been very successful with a 
definitive cure rate at 6 months of 98.7% (95% CI: 95.1-99.8%) (Sundar et al., 2011). 
These results were confirmed in an implementation study in India where the MF/PM 
combination showed 96.9% (95%CI: 95.0-98.2%) efficacy at the end of 6 months 
follow-up, which was similar for all age groups. The regimen needs to be adapted to 
the African disease context which is expected to require longer duration and higher 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
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doses to achieve satisfactory efficacy, as described below based on the experience of 
miltefosine and paromomycin treatment in East Africa. 
Currently, miltefosine (MF) is still the only oral drug available for VL treatment. As a 
phosphorylcholine ester of hexadecanol, it is a membrane-active alkylphospholipid, 
which interferes with the membrane lipid metabolism (e.g. de novo synthesis of 
phosphatidylcholine). It acts through numerous interactions with cell membrane 
components and cell signaling pathways. It also induces apoptosis through the PI3K-
Akt pathway and possibly through mitochondrial dysfunction. Numerous immune-
modulatory actions of MF have been described, potentially contributing indirectly to its 
mechanism of action in vivo. 
MF has been registered in India for VL indication in 2002, and it has been extensively 
used in Asia for VL treatment as monotherapy for 28 days. Treatment of up to 12 weeks 
is recommended for Post Kala azar Dermal Leishmaniasis (PKDL) in Asia. It is well 
tolerated, the main side effects are vomiting and nausea, and transient increases in 
liver enzymes and creatinine. The majority of the events are mild and do not require 
treatment discontinuation. The main limitation of MF is its teratogenicity, which requires 
contraception in female patients of childbearing age during treatment (28 days) and 
after treatment for at least 5 months. Furthermore, in the US Product Information 
(Paladin Therapeutics, 2014) there is a warning on potential male fertility toxicity, which 
has been observed in rats. The rationale for this warning was based on findings 
observed at the Human Equivalent Dose (HED) calculations. However, when 
alternative analysis on risk of fertility impairment was done based on exposure data 
(PK) in animals rather than HED, a better safety margin could be derived of 2-3 fold in 
male rats and at least 3.5 fold in female rats, taking into consideration the lowest dose 
where reversible findings were observed in animals. This is in line with retrospective 
data from VL patients treated with MF in India showing 69% of proven fertility, vs 52% 
in the Amphotericin B control arm (assessments were done between 11 and 57 months 
after start of MF treatment) (FDA Advisory Committee & Book, 2013). In another study 
in Columbia, sperm tests were performed in CL patients, resulting in no clinically 
relevant effect on sperm viability or spermatogenesis.  However, these studies had 
methodological issues and were not considered appropriate to rule out the risk on male 
fertility. 
MF was evaluated as monotherapy and in combination with Ambisome in a phase II 
clinical trial in Kenya and Sudan (LEAP0208, Omollo et al., 2011; Wasunna et al., 
2016). None of the treatment regimens reached the predefined acceptable satisfactory 
efficacy level of > 90% (point estimate) to be taken forward in a Phase III trial to 
compare with SSG+PM: Ambisome + MF (10 days regimen) showed a cure rate of 
77% (95% CI: 64-90%), and MF as monotherapy (28 days regimen) had 72% (95% 
CI: 60-85%) efficacy at 6 months follow up. Although not statistically significant (study 
was not powered for this comparison), there was a clear trend of a poorer efficacy in 
children (7-12y) as compared to adolescents/adults (13-60y) in the trial, especially in 
the MF monotherapy arm (59.1% vs 86.2%, p= 0.061). The conventional MF dose 
linearly based on weight (mg/kg), calculated as 2.5 mg/kg/day, did not provide similar 
drug exposure in children as compared to adults. This is in line with previous published 
MF PK data from India and Nepal (Dorlo, Huitema, Beijnen, & De Vries, 2012; Ostyn 
et al., 2014). MF has a long half-life (7 days), and keeps accumulating over the 
treatment duration to eventually reach steady-state at the 4th week of treatment. In the 
LEAP0208 study, for 28 days treatment, the mean end-of-treatment MF concentration 

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was 22.1 μg/mL (IQR 16.8-29.1 μg/mL) vs 30.2 μg/mL (IQR 24.7-36.3 μg/mL) 
(difference 37%, p<0.001), respectively for patients weighing <30 kg vs patients >30 
kg.  
To overcome this under-dosage of children, an allometric dosage for paediatric VL has 
been assessed in the LEAP 0714 trial in Kenya and Uganda. This MF dosing is based 
on an allometric algorithm by fat free mass. In practice, individual weight, height and 
sex are used to determine the dose, which entails administering a relatively higher 
mg/kg/day dose in patients with lower body weight compared to patients with a higher 
body weight (Dorlo et al., 2012). The 28 days regimen of MF allometric dose showed 
a cure rate of 96.7% (95% CI: 82.8-99.9%) at Day 28 and a cure rate of 90.0% (95% 
CI: 73.5-97.9%) at 6 months follow up in a population of 30 patients aged 4 to 12 years. 
These results showed that efficacy level was increased in children treated with the 
allometric dose as compared to the conventional dose, reaching similar efficacy 
observed in adults in the LEAP0208 trial (i.e. 86.2%). In general, MF was well tolerated 
and compliance in the hospital setting was 100%. There was no treatment 
discontinuation due to drug related adverse events. Two SAEs were reported (anaemia 
and transfusion reaction), none of them related to MF, which is consistent with the 
good safety profile of this drug. PK analysis was performed to further characterize the 
drug exposure and PK properties of MF comparing allometric versus historic data on 
the conventional regimen. The overall MF exposure (area under the concentration-time 
curve from zero to infinity) was 12% higher compared to conventional dosing in the 
same pediatric age group. MF seemed to accumulate faster in LEAP0714 with the 
allometric dosing in the first week of treatment, resulting in a higher drug exposure 
during the first half of the treatment: e.g. day 7 MF concentrations were generally much 
higher for LEAP0714 (median 5,880 ng/mL) than for LEAP0208 (median 2,670 ng/mL), 
although this difference was not significant due to high variability. While total MF 
exposure was thus increased, children treated with MF allometric dosing did not 
achieve the expected exposure i.e. similar to adults treated with conventional therapy 
of 2.5 mg/kg/d for 28 days. A potential reason for this was an apparent stagnation of 
the MF accumulation between day 14 and day 21 for 40% of the LEAP0714 patients, 
after which MF concentrations rose again between day 21 and day 28. Nevertheless, 
the variability in MF concentrations and overall exposure was higher for the linear 
dosing regimen than for the allometric regimen. This had an effect on the proportion of 
patients reaching a PK target threshold, with less patients being low exposed. It was 
previously shown that the duration someone was above 10x EC50 (17.9 µg/mL) as 
measure of MF PK was associated with a lower probability of disease relapse and 
subsequent rescue treatment (T. P. C. Dorlo et al., 2014). 15% of paediatric patients 
enrolled in LEAP0714 did not reach the 17.9 µg/mL threshold (10xEC50), compared 
to 29% for LEAP0208 in the paediatric subset. This in combination with the 12% overall 
higher total exposure to miltefosine could possibly explain the improved clinical efficacy 
observed in children treated with the allometric regimen.  
Based on the results of this trial, MF is an oral drug that remains an attractive option 
for combination with other drugs, and for children (< 30Kg), the allometric dosing is a 
better regimen, both in terms of drug exposure and therapeutic outcome, while 
maintaining a good safety profile.  
Paromomycin (PM) is a broad spectrum aminoglycoside antibiotic effective against a 
wide range of bacteria and protozoa. It has been extensively studied in Asia, where 
the 10-day combination of PM with MF is currently one of the treatment options used 

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for VL, with a very safe profile and high efficacy of 98% in the phase III trial and also 
in the effectiveness study in field conditions. The most frequently reported AEs related 
to PM are injection site pain, transient mild or moderate increases in alanine 
aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase, 
creatinine and bilirubin enzymes, and reversible ototoxicity (Paromomycin Package 
Insert, 2016).  
PM has been well studied in Eastern Africa during the development of the combination 
SSG-PM. The initial PM monotherapy arm in the LEAP 0104 trial was 15mg/kg/day IM 
for 21 days, based on satisfactory efficacy data from Asia. The overall efficacy at 6 
months was 63.8% (81/127 cured) with high variability across treatment sites (14.3 to 
96.6%) (Hailu et al., 2010). The study arm was stopped and a dose-finding study was 
implemented in Sudan to assess the efficacy of PM monotherapy given as 
15mg/Kg/day for 28 days (extended duration) or 20mg/Kg/d for 21 days (increased 
daily dose); the efficacy at the 6-months follow-up was 81% (17/21, 95%CI 58-95%) 
and 80% (16/20, 95%CI 56-94%) respectively (Musa et al., 2010). The gain in efficacy 
was similar either by extending duration or increasing PM daily dose, suggesting that 
it is driven by the overall drug exposure. The protocol was amended, to replace the PM 
monotherapy arm with the regimen of 20mg/kg/d IM for 21 days. The overall efficacy 
of the 20mg/kg/d regimen at 6 months was 84.3% (167/198 cured), with more 
homogeneous profile across sites (80% to 93.3%) (Musa et al., 2012). Doses of 
15mg/kg/d and 20mg/kg/d presented very similar safety profiles. Increasing the PM 
dose to 20mg/kg/d did not increase the incidence of liver or kidney toxicity, ototoxicity 
or serious adverse events. Therefore, both regimens could be considered for 
combination with MF. Considering that the PM efficacy is driven by overall exposure, 
and both regimens are safe, the 20 mg/kg/d regimen is a better option as it allows for 
shorter treatment duration. Intramuscular PM can be administered at primary health 
care level, requires minimal training of health personnel and the drug can be stored at 
room temperature. Moreover, it is the cheapest of currently available anti-leishmanial 
drugs. 
The current study aims to determine if combined MF and PM treatment regimen is non-
inferior to the SSG-PM combination currently used as first line treatment in Eastern 
African VL patients. If proven efficacious (non-inferior) and safe, this therapy can be 
an alternative over the use of SSG as a component of the current treatment. It would 
minimize the number of injections, remove the direct antimonial toxicity, could be an 
attractive option for children that represents a high proportion of the population at risk 
and the elderly who are most at risk of SSG-induced toxicity, therefore potentially 
improving the overall benefit/risk profile of VL treatment. Another important advantage 
of this combination is the reduced cost and increased suitability to be used in remote 
areas.  
Originally, the study protocol had a trial design including three arms: two investigational 
arms with MF and PM and the reference treatment arm SSG-PM for 17-day course 
treatment. The strategy was to combine one parenteral drug for a short period (PM 20 
mg/kg/d for 14 days) with an oral drug for the same or longer duration (MF allometric 
dosing for 14 or 28 days) aiming to reduce as much as possible the hospitalization 
period, while achieving appropriate exposure and satisfactory efficacy levels.  
The trial started with both test arms: MF/PM for 14 days and MF for 28 days/PM for 14 
days. During the first year of recruitment in the trial, the number of VL cases in the 

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region dropped by 20% as compared to the preceding 4 years before commencing the 
trial. In addition, only 4 out of 7 sites have been actively recruiting patients for various 
reasons, including political instability in the countries, affecting local operations. Finally, 
the enrolment rate (% of ‘VL patients recruited in the trial / VL confirmed cases’) has 
been lower than anticipated and variable across sites, with an overall enrolment rate 
of only 16% as compared to the 30% planned. Therefore, the study with its original 
design could not be conducted in a reasonable timeframe and important changes have 
been considered in order reach the study objectives. 
In order to mitigate the low enrollment, the protocol was amended to v3.0 dated 4 May 
2018 to include 2 MSF sites (Tabarakallah in Sudan and Abdurafi in Ethiopia). The 
sites were expected to compensate for the low patient numbers overall. In addition, 
because of the high proportion of patients in Ethiopia excluded due to severe 
malnutrition (30-40%), an Ethiopia-specific amendment was implemented to modify 
the indicator for severe malnutrition from body mass index (BMI) to mid-upper arm 
circumference (MUAC) to allow for a better representativeness of the Ethiopian 
population. Finally, active case search and referral networks were initiated at some of 
the sites. Nevertheless, these measures taken to improve recruitment have not 
resulted in a significant boost of the enrolment and further actions needed to be taken. 
New oral promising NCEs are expected to be at the clinical stage of development for 
VL patients by 2021 and the plan is to complete this trial before then so that results 
can be of benefit to the VL patients. Moreover, if new trial(s) with NCEs start before 
the end of this study, the recruitment of patients will compete with these NCEs studies 
and will become even more challenging to develop new treatments for visceral 
leishmaniasis in Eastern Africa region. 
Following DSMB review of the available data (meeting held on 7 May 2019), two main 
changes to the protocol were agreed to achieve completion of the trial in an acceptable 
timeframe and ensure the achievement of the objectives of the trial. First, to revise the 
exclusion criteria in order to include a more VL representative population, and second, 
to discontinue recruitment in the investigational arm 2.  
Revision of the exclusion criteria. The reasons for exclusions leading to a low 
enrollment rate were analyzed. It was noted that across the sites, 15 to 20% of patients 
were excluded because of laboratory abnormalities. In most cases, the baseline 
laboratory abnormality was not considered clinically significant by the investigator to 
indicate AmBisome therapy and a majority of these patients were therefore treated with 
SSG-PM with a good therapeutic response. For this reason, individual laboratory 
parameters in the exclusion criteria were replaced by investigator judgement on the 
need of AmBisome, based on the severity of the clinical manifestations of the disease 
(such as jaundice, bleeding and oedema) and laboratory parameters.  
Discontinue recrutiment in investigational arm 2 (PM 14d / MF 28 days). Following 
review of the preliminary available data by the DSMB, including 143 study participants 
enrolled, among which 135 had completed treatment and Day 28 assessment, and 66 
of them had completed their day 210 follow-up visit, it was decided in agreement with 
the DSMB to ensure retention of the investigational arm 1, on the basis of 1) shorter 
treatment duration than current standard of care, 2) it can be administered within the 
hospitalisation period, 3) it is a cheaper option for the control programs, and therefore 
from a public health perspective will be a potentially more favorable option for patients. 

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In this revised protocol version, the analysis remains as planned, but comparison will 
be performed only between one investigational arm (arm 1 MF/PM 14 days) and the 
standard of care (SSG-PM). Data from patients randomized in arm 2 (PM 14 days/MF 
28 days), which recruitment is now stopped, will be summarized in a descriptive 
analysis, but will not be included in the comparative hypothesis testing.   
The total dose of PM will be 280 mg, similar to the total dose given in the SSG-PM 
combination therapy (255 mg). For MF, PK data indicated that with 14 days allometric 
MF treatment, 91% of patients will at least reach EC90 in plasma, which would 
represent appropriate exposure.  
If demonstrated non-inferior to SSG-PM, the 14-day MF-PM regimen will be preferable 
to be taken forward for future standard of care or alternative treatment to SSG-PM. 
Data will be presented to Ministry of Health (MoH) and regulatory authorities in the 
region as evidence to support policy change for the national control programs. 
2. Study Objectives and Endpoints 
2.1 Objectives 
2.1.1. Primary Objective 
• To compare the efficacy of a 14-day combination regimen of MF and PM with the 
standard 17-day course of SSG-PM for the treatment of primary VL patients in Eastern 
Africa 
2.1.2. Secondary Objectives 
• To assess the safety of two combination regimens of PM (14 days) and MF (14 or 28 
days) with the standard 17-day course of SSG-PM 
• To describe the efficacy of the PM (14 days) and MF (28 days) investigational arm 
• To describe the pharmacokinetic (PK) profiles of PM and MF (14 days and 28 days 
regimen) in primary VL patients 
• To evaluate parasite clearance in each arm as indicated by direct microscopy and 
quantitative polymerase chain reaction (qPCR) 
• To assess the relationship between PK and PD measurements (parasitological and 
clinical outcome) 
• To describe compliance to MF treatment in an outpatient setting 
2.2. Study Endpoints 
2.2.1. Primary Endpoint 
Definitive cure - cure at 6 months follow up defined as absence of clinical signs and 
symptoms of VL at D210 and no requirement for rescue treatment during the trial (i.e. 
no relapse or initial treatment failure). 
 

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2.2.2. Secondary Endpoint(s) 
• Safety: 
Frequency of SAEs and AEs requiring treatment discontinuation. 
Frequency and severity of adverse events from the start of treatment through the last 
visit, at D210. 
• Efficacy: 
Initial cure - cure at the end of treatment (Day 28), defined as recovery of clinical signs 
and symptoms; absence of parasites (microscopy) and no rescue treatment 
administered up to and including Day 28.  
Probable cure - absence of clinical signs and symptoms of VL at D56 and no prior 
requirement for rescue treatment. 
• Pharmacokinetics: 
Total and partial blood plasma exposure to PM and MF (bioanalysis performed by a 
validated LC-MS/MS assay), defined as the area under the concentration-time curve, 
for MF, during treatment and until the last point of follow-up, and for PM based on full 
curves both on the first day of treatment (day 1) and the last day of treatment (day 14). 
• Pharmacodynamics: 
Blood parasite clearance over time (qualitative and quantitative), as measured by 
qPCR from blood samples, from baseline until day 210, and at any suspicion of relapse 
during the trial. 
Tissue parasite loads, as semi-quantified by microscopy and qPCR from tissue 
samples collected at baseline, at the end of treatment (D28) and at any suspicion of 
relapse during the trial. 
• Compliance to MF treatment in an outpatient setting will be described through patients’ 
hospital records history, drug accountability and PK measurements. 
3. Study design and study design rationale 
3.1. Study design 
This is an open label, Phase III, randomized, controlled, parallel arm multicentre non-
inferiority clinical trial to compare the efficacy of a combination regimen of MF and PM 
with SSG-PM for the treatment of primary adult and children VL patients in Eastern 
Africa. 
In addition, safety of two combination regimens of PM (14 days) and MF (14 or 28 
days) will be assessed, and the efficacy of the PM (14 days) and MF (28 days) 
investigational arm 2 will be described. 
The study will be conducted at 5 LEAP sites and 2 MSF sites: Kacheliba in Kenya, 

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Amudat in Uganda, Doka, Um El Kher and Tabarakallah (MSF) in Sudan and Gondar 
and Abdurafi (MSF) in Ethiopia. 
The 2 treatment regimens to be tested were as below but with patient’s recruitment in 
arm 2 now discontinued: 
• Arm 1: Paromomycin 20 mg/kg/d IM for 14 days combined with oral miltefosine 
allometric dosing for 14 days 
• Arm 2: Paromomycin 20 mg/kg/d IM for 14 days combined with oral miltefosine 
allometric dosing for 28 days (recruitment is now discontinued in this arm but 
patients’ data available will be analyzed and results described) 
The reference arm is the current standard treatment for VL: 
• Arm 3: Sodium Stibogluconate 20 mg/kg/day IM/IV combined with Paromomycin 15 
mg/kg/day IM for 17 days 
For patients weighing < 30 kg, an easy-to-use table with allometric dosing scheme by 
weight, height and sex will be provided to the investigators to define the exact daily 
dose to be administered. For patients weighing ≥ 30 kg, the allometric dose will 
correspond to the conventional dose in mg/kg. Therefore, in order to simplify the dose 
calculation, patients weighing ≥ 30 to < 45 kg will receive 100 mg/day and patients ≥ 
45 kg will receive 150 mg/day. 
The target population will be VL patients from 4 to 50 years old in order to cover both 
paediatric and adult population. The limit of 50 years for inclusion is due to higher 
mortality rate and lower efficacy observed in patients > 50y when treated with SSG-
PM in a recent pharmacovigilance program (Kimutai et al., 2017). MF and PM will be 
administered together, both starting at D1. PM dosage will be 20 mg/kg/d for 14 days 
administered once-a-day intramuscularly, whereas MF allometric dose will be 
administered orally BID for 14 days in arm 1 and for 28 days in arm 2.  
Patients are hospitalized for 14 days of PM and MF treatment for both arm 1 and arm 
2. MF treatment starts at the same time as PM treatment and for arm 2 it used to 
continue on an out-patient basis until completion of the 28 days treatment.  
During hospitalization, compliance to treatment is assured. During this period, patients 
or parent/guardian will be instructed on MF treatment, the daily dose (morning and 
afternoon), and the administration with food to avoid or minimize any vomiting. Patients 
in arm 2 were discharged after 14 days in hospital with clear instructions on how to 
continue MF treatment, and to return for the Day 28 visit. Compliance during the 14 
days treatment outside the hospital was assessed by collecting the empty blisters pack 
at the day 28 visit and checking for any unused drug. A daily diary was provided to 
these patients in order to guide them in their treatment schedule at home. Any episode 
of vomiting reported by the patient was captured by the clinician at day 28 visit. In case 
of multiple episodes of vomiting or any other adverse events, patients were instructed 
to come to the hospital for an assessment and management in an unscheduled visit. 
Compliance will be further assessed through cross-check with PK data. 
SSG-PM combination therapy will be administered for 17 days according to routine VL 

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treatment guidelines. Patients will remain hospitalized for the entire duration of SSG-
PM treatment. Patients in all arms will come back for the end-of-treatment 
assessments visit at day 28. 
All patients will have pharmacokinetics measurement of MF at D28 and D56 in order 
to measure end of treatment MF concentration and verify adherence to the treatment. 
In addition, a subset of patients in Kenya and Sudan will be asked to participate to 
intensive sampling for both PM and MF PK, defined as the intensive cohort: originally, 
40 in each MF/PM combination arms, with selection of 20 pediatric (≥7 to ≤12 yrs) 
patients and 20 adolescent/adult patients (>12 to ≤50 years). Because of premature 
discontinuation of arm 2, the number of patients enrolled in this arm in the PK/PD 
intensive cohort will probably not reach 40, however samples collected will be 
analysed. Patients who agree to be part of the intensive cohort will be asked to provide 
a separate written informed consent. The intensive cohort will also have additional 
qPCR blood samples collected (see section 5.1 Schedule of PK/PD events – Intensive 
cohort, as well as section 8.4 Pharmacokinetics and Pharmacodynamics 
Assessments). 
3.2. Study duration and duration of subject participation 
Each patient’s participation in the study will be for approximately 7 months. This will 
consist of baseline assessments, treatment period (14, 17 or 28 days) and 6 months 
follow-up. Recruitment for the entire trial is expected to take approximately 32 months 
assuming that 15 to 30% of all VL patients will meet the eligibility criteria. Therefore, 
study duration (first patient in to last patient, last visit) is expected to take approximately 
39 months. Taking into account the analysis and reporting period, the study shall last 
at most 45 months. 
3.3. Rationale of study design  
The strategy to combine one parenteral drug for a short period (PM 20 mg/Kg/d IM for 
14 days) with an oral drug for the same or longer duration (MF allometric dosing for 14 
or 28 days) was to reduce as much as possible the hospitalization period and to allow 
the treatment to achieve appropriate exposure and satisfactory efficacy levels for the 
treatment of primary VL patients. In addition, replacing the toxic SSG with oral MF can 
bring better safety and a more field-adapted, patient-friendly treatment. NCE 
candidates won’t be fully developed for VL treatment before 2027. Therefore, currently 
available compounds should be assessed to improve on current recommended 
treatment and avoid adverse drug reactions observed with SSG. 
The primary endpoint of this protocol is cure at 6 months follow up defined as absence 
of signs and symptoms of VL (and no rescue treatment), aiming to assess efficacy of 
the regimen of MF combined with PM as compared to the routine VL treatment SSG-
PM. The long term follow-up of 6 months is necessary in VL trials due to risk of relapse 
after a negative test of cure at the end of treatment.  
This phase III trial should provide the evidence to determine if the investigational study 
arm is non-inferior to SSG-PM. Patient tolerability will improve with short course 
combination regimens that are better tolerated and more patient-friendly.  
Safety assessments will include clinical, haematological and biochemistry evaluations 
at regular time points (day 0, 3, 7, 14, 28, 56 and 210) to ensure safety is adequately 

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assessed and treatment failures can be caught. 
  
Samples size estimation is based on the power and precision to estimate the efficacy 
of definitive cure at 6 months (D210) which is the primary endpoint and main parameter 
of interest. Taking into consideration the data available on PM monotherapy in East 
Africa, the results of LEAP 0208 on miltefosine efficacy (Wasunna et al., 2016) and 
latest efficacy results of MF allometric dose in children in Eastern Africa, the expected 
efficacy for the combination of PM with MF at the end of 6 months follow-up will be 
defined as 93%. The margin defined as acceptable for concluding the non-inferiority of 
the tested treatments compared to the reference arm (SSG-PM, 91% expected 
efficacy) will be 7%, considering the advantages of MF/PM combination in terms of 
safety, shorter hospitalization and more patient-friendly. 
An interim analysis (IA) will not be performed in this trial. Unfortunately, there is no 
early marker of cure for VL that would allow an interim analysis using data from end of 
treatment to predict the outcome at the 6 months follow-up.  
Pharmacodynamic parameters of repeated qPCR measurements in blood and in tissue 
(baseline and EOT), and clinical outcome up to D210 will be assessed to allow for a 
simultaneous population PK-PD analysis to investigate in detail the drug exposure-
treatment response relationship.  
Figure 1- Overall study design   
 
4. Selection of Subjects 
170 patients will be enrolled in each study arm 1 and 3. The total number of patients 
included in the trial will be approximately 420, taking into account the number of 
patients randomized to treatment arm 2 before discontinuation of this arm 
(approximately 80 patients). Patients will be primary VL cases with age range between 
4 and 50 years of age. 

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In order to meet recruitment target, 5 LEAP sites and 2 MSF sites are included in the 
trial:  Kacheliba in Kenya; Amudat in Uganda; Doka, Um El Kher and Tabarakallah 
(MSF) in Sudan; and Gondar and Abdurafi (MSF) in Ethiopia.  
Sites will be initiated as soon as the required EC/regulatory approvals have been 
obtained in their respective country. 
Approximately 150 VL patients are treated routinely per month in the 7 sites. Based on 
historical data from previous trials, whereby it is estimated that 15% of patients will not 
be in the age range of 4-50 years, 20% will be excluded based on lab and/or nutrition 
criteria, 15% will be female patients not willing or not able to comply with contraception 
and an additional 30% may not be enrolled for other criterion, the enrollment rate for 
this trial will thus be approximately 20% of the total number of patients diagnosed with 
VL. Therefore, the recruitment capacity expected is 20 patients per month when all trial 
sites are active. Considering a mean approval process of 6-12 months in the countries, 
the delay in the construction of Umelkher site and the fact that MSF sites have joined 
the trial at a later stage, the target enrollment of 170 patients per arm is planned to 
take approximately 32 months. 
There will not be a minimum requirement of number of patients enrolled per site; sites 
are expected to recruit competitively until the total sample size is achieved.  
Enrollment will be boosted by increasing patient outreach through local strategies such 
as active case search. 
The following eligibility criteria were designed to select subjects for whom the protocol 
treatment is considered appropriate. All relevant medical and non-medical conditions 
should be taken into consideration when deciding whether this protocol is suitable for 
a particular subject. Eligibility criteria may not be waived by the investigator. Any 
questions regarding a subject’s eligibility should be discussed with the DNDi Medical 
Coordinator prior to subject’s enrollment. 
4.1. Inclusion criteria 
Subjects must meet all of the following inclusion criteria to be eligible for enrolment 
into the study:  
• Patients with clinical signs and symptoms of VL and confirmatory parasitological 
microscopic diagnosis  
• Patients aged 4 to ≤ 50 years who are able to comply with the study protocol.  
• Patients for whom written informed consent has been obtained (if aged 18 years and 
over) or signed by parents(s) or legal guardian for patients under 18 years of age. In 
the case of minors, assent from the children also needs to be obtained as per each 
country regulatory requirements 
4.2. Exclusion criteria 
The presence of any of the following will exclude a subject from study enrolment: 
• Patients who are relapse cases 

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• Patients with Para-Kala azar dermal leishmaniasis grade 3 
• Patients who have received any anti-leishmanial drugs in the last 6 months 
• Patients with severe malnutrition (for children aged <5 years: weight-for-height WHO 
reference curves by sex, z score <-3; for patients 5-18 years: BMI-for-age WHO 
reference curves by sex, z score < -3; for adults >18 years: BMI < 16)*  
• Patients with positive HIV diagnosis 
• Patients with previous history of hypersensitivity reaction or known drug class allergy 
to any of the study treatments 
• Patients with previous history of cardiac arrhythmia or with a clinically significant 
abnormal ECG 
• Patients suffering from a concomitant severe infection such as TB, schistosomiasis or 
any other serious underlying disease (e.g. cardiac, renal, hepatic) or chronic condition 
which would preclude evaluation of the patient’s response to study medication  
• Pregnant or lactating women 
• Female patients of child bearing age who do not accept to have a pregnancy test done 
at screening and/or who do not agree to use contraception from treatment period until 
5 months after the end of treatment (see section 15.2) 
• Patients with haemoglobin < 5g/dl 
• Patient with signs of severe VL according to Investigator’s judgement, requiring an 
indication for AmBisome therapy based on the clinical manifestations (such as 
jaundice, bleeding, edema) and clinically significant abnormalities in the following 
laboratory parameters: haemoglobin, WBC, platelets, liver enzymes (ALT and AST), 
total bilirubin and creatinine 
• Patients with pre-existing hearing loss based on audiometry at baseline 
• Patients who cannot comply with the planned scheduled visits and procedures of the 
study protocol 
 
* Note: for Ethiopia only: Patients with severe malnutrition (for patients 4-18 years: 
MUAC cut-off based on MUAC-for-height reference table; for patients > 18 years: 
MUAC < 170 mm)

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5. Schedule of events 
 
Protocol Activities and Forms to Be 
Completed 
Screening 
Treatment period (day) 
+/- 1d except for D28 
  
Follow up visits 
D-7 to D0 
D1 
D3 
D7 
D14 
D28 
(EOT) 
+1d 
  
D56 
+/- 7d 
D210 
+/- 14d 
Consent form & consent for HIV test 
X 
  
  
  
  
  
  
  
  
Demographic data and medical history 
X 
 
 
 
 
 
 
 
 
Clinical assessment 
X 
 X 
X 
X 
X 
X 
  
X 
X 
Nutritional status 
X 
 
 
 
 
 
 
 
 
Audiometric test 
X 
 
 
 
X 
X 
  
 
X 
HIV test 
X 
  
  
  
  
  
  
  
X1 
Pregnancy test2 
X 
  
  
  
  
X  
  
X  
X  
Spleen/bone marrow/lymph node aspiration, 
parasitology assessment (microscopy) 
X 
  
  
  
  
X 
  
X5 
X5 
Hematology (hemoglobin, WBC with 
differential, platelets) 
X 
  
X 
X 
X 
X 
  
X 
X 
Biochemistry analysis (AST/ALT, bilirubin, 
creatinine) 
X 
  
X 
X 
X 
X 
  
  
 
Biochemistry analysis (Albumin) 
X 
 
 
 
 
X 
 
 
X 
ECG3 
X 
 
 
X 
X 
X 
 
 
 
Assessment for PKDL 
X 
  
  
  
  
X 
  
X 
X 
Blood sample for miltefosine PK4 
 
 
  
 
X 
X 
  
X 
 
Tissue (bone marrow/spleen/lymph node) and 
blood sample for PD (qPCR) 
X 
 
 
 
 
X 
  
X5 
X5 
Treatment arm 1  
 
PM QD/MF allometric dose BID for 14 days 
 
 
 
 
Treatment arm 2 – Discontinued 
 
PM QD for 14 days /MF allometric dose BID for 28 days 
 
 
 
Treatment arm 3  
 
SSG/PM QD for 17 days 
 
 
 
 
Safety assessment 
SAEs/non-
serious study 
related AEs 
-------------------------- SAEs and AEs monitoring during the study -------------------------- 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 30 of 68 
 
 
1 HIV test at D210 will be done only for patients in Gondar and Abdurafi. 
 
2 Only for woman of child-bearing potential. 
 
3 ECG will be performed at baseline and at D14 in all arms, and in addition at D7 and D28 in the reference arm SSG/PM. 
 
4 MF PK assessments at D14 will apply to all patients enrolled in arm 1, while D28 and D56 samples will apply to all patients enrolled in 
arm 1 and 2. MF PK sample will be collected prior to the morning dose at D14 and D28 (arm 2). D28 (arm 1) and D56 samples will be 
collected in the morning.  
 
5 Parasitology and collection of tissue and blood samples for qPCR will be done during the follow-up period after D28 (at D56 and D210) 
only if clinically indicated (i.e. anytime if suspect of relapse, with reappearance of symptoms and signs of VL). 
 
Total volume of blood to be collected for patient not included in the intensive PK/PD cohort: 
Assumptions: Ht= 1mL EDTA, BQ= 2mL dried tube, MF PK= 2mL EDTA, PD= 1mL EDTA. Volume of blood to be collected will be 
minimized as much as possible, according to the volume of tubes available in the market. The maximum volume to be collected at each 
study visit will be 6mL. 
 
General note: PK and PD assessments in this schedule of events do not apply to patients included in the intensive cohort 
(see section 5.1 Schedule of PK/PD events – Intensive cohort). 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 31 of 68 
 
 
5.1 Schedule of PK/PD events - Intensive cohort 
 
40 patients from Kenya and Sudan in each arm 1 and 2 (20 paediatric ≥7 to ≤12 yrs patients and 20 adolescent/adult patients >12 to 
≤50 yrs) will be included in the intensive cohort and will have a separate schedule of PK and PD events as described below. Because 
of discontinuation of arm 2 in protocol v4.0 dated 22 July 2019, the number of patients in arm 2 enrolled in the PK/PD intensive cohort 
is expected to be probably less than 40.  
Protocol Activities and Forms to Be 
Completed 
Screening 
Treatment period (day) 
+/- 1d except for D28 
  
Follow up visits 
D-7 to D0 
D1 
D3 
D7 
D14 
 
D211 
 
D28 
(EOT) 
+1d 
  
D56 
+/- 7d 
D210 
+/- 14d 
Sampling Time-points: 
 
 
 
 
 
 
 
 
 
 
Blood sample for paromomycin PK2 
X 
X 
 
 
X 
 
 
 
 
 
Blood sample for miltefosine PK3 
X 
X 
  
X 
X 
X 
X 
  
X 
 
Blood sample for PD (qPCR) 
X 
 
X 
X 
X 
 
X 
  
X4 
X4 
Sampling Volumes: 
 
 
 
 
 
 
 
 
 
 
Total EDTA tube blood volume (mL) 
4 
12 
2 
4 
12 
2 
4 
 
4 
2 
   Blood volume for hematology (mL) 
1 
 
1 
1 
1 
 
1 
 
1 
1 
   Blood volume for PK plasma (mL) 
2 
12 
 
2 
10 
2 
2 
 
2 
 
   Blood volume for PD qRT-PCR (mL) 
1 
 
1 
1 
1 
 
1 
 
1 
1 
Total dried tube blood volume (for 
biochemistry) 
2 
 
2 
2 
2 
 
2 
 
 
2 
TOTAL BLOOD VOLUME (mL) 5 
6 
12 
4 
6 
14 
2 
6 
 
4 
4 
 
1 D21 visit will only be applicable to the subset of patients allocated to intensive PK sampling in arm 2 (collection of this sample is not applicable anymore in this 
protocol version). 
2 PK PM sample at D1 and D14 (arm 1 and 2): 
In order to minimize blood volume collected, a subset of patients will be randomly allocated to one sparse PK sampling schedule:  
- 
D1: either 1, 2, 4, 24h or 1, 2, 8, 24h 
- 
D14: either 0, 1, 2, 4, 24h or 0, 1, 2, 8, 24h 
3 PK Miltefosine sample at D1 and D14 will be collected at the same time points as PK PM (according to the sparse sampling scheme the patient is allocated to). 
8h sample will be collected always before the 2nd daily dose. PK samples will be collected prior to the morning dose at D7 and D28 (arm 2). D28 (arm 1) and D56 
samples will be collected in the morning. 
4 Blood samples for qPCR will be collected during the follow-up period (at D56 and D210) only if clinically indicated (i.e. anytime if suspect of relapse, with 
reappearance of symptoms and signs of VL). 
5Volume of blood to be collected will be minimized as much as possible, according to the volume of tubes available in the market. 
The maximum volume to be collected at a study visit will be 14 mL on day 14. 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 32 of 68 
 
6. Enrolment procedures  
The study will be conducted in the LEAP and MSF sites. These sites are located in 
endemic areas of VL and are reference treatment centers in the regions, where 
routinely VL patients are diagnosed and treated. 
Patients may come to the treatment center passively or through referral from health 
professionals. Referral can be done on the basis of clinical signs and symptoms and/or 
rapid diagnostic test. In some of the sites, active case detection is performed by mobile 
teams visiting regularly highly endemic communities. This procedure will be boosted 
during the inclusion period of the clinical trial. 
Subjects between the ages of 4 and 50 years who fulfill the inclusion / exclusion criteria, 
and from whom informed consent has been obtained (see 15.1 Informed Consent 
process) will be enrolled in the study. Patients who have entered screening 
assessments for this trial will be documented in a Patient Screening/Enrolment Log. 
Each eligible patient will be assigned a patient identification number by the investigator 
in the site, sequentially, which will be recorded in the Patient Screening/Enrolment Log. 
A note describing the ICF signature and enrolment in the trial will also be made in the 
patient's file.  
Patients were originally randomized centrally to treatment in a 1:1:1 allocation ratio 
using varying block sizes of 6, 9 or 12 subjects in random order. Following 
discontinuation of arm 2, a 1:1 allocation ratio is now used with varying block sizes of 
4, 6 or 8 subjects. Randomization codes will be prepared by the DNDi Data Center in 
Nairobi and will be accessible through an online system. If the principal investigator 
has no internet connection available, randomization codes will also be provided by 
phone.  
Treatment should be started within 7 days after the screening assessment. Otherwise 
compliance with eligibility criteria has to be reassessed (blood tests). 
Enrolment status of subjects at each site will be informed weekly by the sites to the 
study monitor and updated by data centre. Recruitment will stop once 170 subjects 
have been enrolled in each trial’s arm 1 and 3. 
7. Study treatments 
7.1. DNDi study treatment  
Name: Miltefosine (Impavido®) 
Class: Phosphocholine analogue. 
Mechanism of action: interferes with the synthesis and metabolism of phospholipids. 
It may also interfere with the parasite’s membrane signal transduction, and 
glycosylphosphatidylinositol anchor biosynthesis. 
Commercial source: Paladin labs and Knight Therapeutics Inc. 
Product appearance: comes as 10mg and 50mg capsules in pack of 56 capsules 
sealed in 8 aluminum blister stripes, each containing 7 capsules. 
Administration: oral treatment  
For MF, there are no known interactions with other commonly used medications, 
though this cannot be excluded. 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 33 of 68 
 
Name: Paromomycin 
Class: Aminoglycoside 
Mechanism of action: inhibits protein synthesis by binding to 16S ribosomal RNA.  
Commercial source: Gland Pharma, India; distributed by IDA Foundation. 
Product appearance: Each box of paromomycin contains a pack of 10 amber-colored 
glass ampoules containing 750 mg of paromomycin base in 2 ml (375 mg/ml) 
equivalent to 1 g of paromomycin sulphate in 2 ml (500 mg/ml).  
The 20 mg/kg sulfate is equivalent to 15 mg/kg of base.  
Administration: intramuscular injection. 
Name: Sodium Stibogluconate 
Class: Pentavalent antimonial 
Mechanism of action: decrease in ATP and GTP synthesis contributes to decreased 
macromolecular synthesis and to decreased Leishmania viability. 
Commercial source: Albert David Ltd., India; distributed by IDA Foundation. 
Product appearance: A faintly straw coloured fluid in a multi-dose brown opaque vial 
of 30ml containing SSG BP equivalent to 100mg pentavalent antimony in each ml (total 
3g). 
Administration: Given IV (slow over 5 minutes) or IM at a daily dose of 20mg/kg body 
weight. 
7.2. Doses and treatment regimens 
7.2.1 Test arms 
Arm 1: Paromomycin 20 mg/kg/d IM q.d. for 14 days combined with oral miltefosine 
allometric dosing b.i.d. for 14 days 
Arm 2: Paromomycin 20 mg/kg/d IM q.d. for 14 days combined with oral miltefosine 
allometric dosing b.i.d. for 28 days (recruitment is now discontinued in this arm) 
MF dosing will be defined as follows: 
- 
Patients <30 kg: the allometric MF daily dose will be calculated according to subject’s 
weight, height and sex 
- 
Patients ≥ 30 to <45 kg: 100 mg/day  
- 
Patients ≥ 45 kg: 150 mg/day 
PM and MF will be administered together, both starting at D1. PM dosage will be 20 
mg/kg/d for 14 days administered intramuscularly, whereas MF allometric dose will be 
administered orally for 14 days in arm 1 (it used to be administered for 28 days in arm 
2).  
Patients are hospitalized for 14 days of PM and MF treatment in both arm 1 and arm 
2. The MF treatment will start at the same time as the PM treatment and for arm 2 it 
used to continue on an out-patient basis until completion of the 28 days treatment.  
Miltefosine administration 
MF will be administered orally for 14 days in arm 1 and used to be administered for 28 
days in arm 2. 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 34 of 68 
 
For patients less than 30 kg, the allometric dose will be defined according to weight, 
height and sex as indicated in Table 1, divided in 2 administrations per day for 14 days 
in arm 1 or 28 days in arm 2 (not applicable now recruitment has been discontinued in 
arm 2). 
Table 1 - Daily allometric miltefosine dose for female children based on fat-free mass 
 
 
 
FEMALE
HT (cm)
80-89
90-99
100-109
110-119
120-129
130-139
140-149
150-159
160
WT (kg)
7
8
30
9
30
RISK OF SEVERE MALNUTRITION
10
30
40
REFER TO THE WHO REFERENCE CURVES
11
40
40
40
12
40
40
40
13
40
40
40
14
40
40
40
50
15
40
40
50
50
16
40
50
50
50
17
40
50
50
50
50
18
50
50
50
50
60
19
50
50
50
60
60
20
50
50
50
60
60
60
21
50
50
60
60
60
60
22
50
50
60
60
60
60
23
50
50
60
60
60
70
70
24
50
50
60
60
60
70
70
25
50
60
60
60
70
70
70
26
50
60
60
60
70
70
70
70
27
50
60
60
70
70
70
70
80
28
50
60
60
70
70
70
80
80
29
50
60
60
70
70
70
80
80
30
50
60
60
70
70
80
80
80
80

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 35 of 68 
 
Table 2 - Daily allometric miltefosine dose for male children based on fat-free mass 
 
MF 10 mg and 50 mg strengths will be supplied through MSF Supply, Belgium, and 
Knight Therapeutics, US respectively in alu/alu blister packs of 56 capsules. 
Administration of the daily dosage must be divided in 2 administrations, during or after 
a meal, to avoid direct gastrointestinal adverse effects of MF, as follows: 
 
MALE
HT (cm)
80-89
90-99
100-109
110-119
120-129
130-139
140-149
150-159
160
WT (kg)
7
8
40
9
40
RISK OF SEVERE MALNUTRITION
10
40
40
REFER TO THE WHO REFERENCE CURVES
11
40
40
12
40
50
13
50
50
50
14
50
50
50
15
50
50
60
60
16
50
50
60
60
17
50
60
60
60
18
50
60
60
60
70
19
50
60
60
70
70
20
50
60
60
70
70
21
60
60
60
70
70
70
22
60
60
70
70
70
80
23
60
60
70
70
80
80
24
60
60
70
70
80
80
80
25
60
60
70
70
80
80
80
26
60
70
70
80
80
80
100
27
60
70
70
80
80
80
100
28
60
70
70
80
80
100
100
100
29
60
70
70
80
80
100
100
100
30
60
70
80
80
100
100
100
100
Total daily 
dose, 
divided: 
30mg 
40mg 
50mg 
60mg 
70mg 
80mg 
100mg 150mg 
Morning 
(breakfast) 
2x 
10mg 
2x 
10mg 
1x 
50mg 
1x 
50mg 
1x 
50mg 
1x 
50mg 
1x 
50mg 
2x 
50mg 
 
 
 
 
 
 
 
 
 
Afternoon 
(dinner) 
1x 
10mg 
2x 
10mg 
----- 
1x 
10mg 
2x 
10mg 
3x 
10mg 
1x 
50mg 
1x  
50mg 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 36 of 68 
 
During the 14 days of hospitalization, administrations for all patient’s arms will be 
directly observed by the study nurse to assure compliance. The number of capsules 
administered (morning and afternoon), the time of administration and any vomiting 
episode will be documented in a patient dispensing log. After each drug intake the 
patients will be observed for vomiting for 30 minutes. All vomiting episodes will be 
recorded. In case of vomiting within 30 minutes after intake, the dose will be re-
administered. If a repeated vomiting occurs (vomiting after re-dose) the treatment will 
not be re-administered.    
Compliance will be assessed by total dose taken by the patient. Full compliance will 
be considered if 90-110% of the prescribed dose is administered.  
If the patient vomits a dose within 30min of drug intake and is re-dosed (without 
vomiting this 2nd re-dose), the re-dose will be considered for compliance. However, if 
the patient is not re-dosed within 30min or if s/he vomits the re-dose, this dose will not 
be accounted for compliance. In the case of a patient vomiting after 30min of MF intake, 
the dose will be accounted for compliance. 
Patients in arm 2 used to continue MF intake after discharge at day 14 on an out-
patient basis. Compliance during the 14 days treatment outside the hospital was 
assessed by collecting the empty blisters at the day 28 visit and checking for any 
unused drug. A diary including illustrations that is made understandable for illiterate 
subjects was provided to these patients in order to guide them in their treatment 
schedule at home. Any episode of vomiting reported by the patient was captured by 
the clinician at day 28 visit. In case of multiple episodes of vomiting, patients were 
instructed to come to the hospital for an assessment in an unscheduled visit. 
Compliance will be further assessed through cross-check with PK data. 
Paromomycin administration 
PM will be given for 14 days in both arm 1 and arm 2. Exact daily dose of 20 mg/kg will 
be calculated using patient’s body weight and will be administered once-a-day with 
intramuscular injection into the buttock muscle.  
PM sulfate will be supplied through IDA Foundation, Holland, in boxes of 10 amber-
colored glass ampoules each, containing an equivalent 750 mg of PM base in 2 ml 
(375 mg/ml). 
All administrations will be directly observed by the study nurse or clinician to assure 
compliance.  
Treatment interruption 
MF treatment in arm 1 (14 days) will last at least 13 days in order to be considered 
90% compliant. In case of treatment interruption longer than 24 hours, missing doses 
will be compensated after day 14. 
MF treatment in arm 2 (28 days) could be temporarily interrupted (due to AEs or at 
discretion of investigator) and resumed if this interval was no longer than 72hs. In this 
case, the total administered dose over a total of 25 days was still considered within 
90% range of full compliance, therefore it was not considered a compliance deviation. 
The missing doses was not compensated after the final dose on day 28, and the 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 37 of 68 
 
schedule of assessments would remain the same. 
If treatment needs to be interrupted for > 72hs due to an AE related to MF, or if 
treatment needs to be interrupted due to an AE related to PM or SSG, rescue treatment 
will be considered, at the discretion of the investigator. 
7.2.2 Reference arm 
Arm 3: Sodium Stibogluconate 20 mg/kg/day Intravenous/Intramuscular (IV/IM) q.d. 
and Paromomycin 15 mg/kg/day IM q.d. for 17 days 
SSG will be given IM/IV at a dose of 20 mg/kg once daily for 17 days. PM base will be 
administered IM at a dose of 15 mg/kg once daily for 17 days. The dosing regimen will 
be the combination of both SSG and PM as described above. 
7.3. Study treatments labelling, packaging 
Commercially available MF, PM and SSG will be used. Trial specific labels will be 
applied on the boxes prior to use with the following statement “For Clinical Trial use 
Only” and will include protocol code, Sponsor and PI contact details as well as direction 
for use. 
7.4. Accountability 
Study medications must be kept in a locked room that can be accessed only by the 
pharmacist or designated personnel by the investigator. The study medications must 
not be used for other purposes other than this protocol. Under no circumstances the 
investigator or site staff may supply study medications to other investigators or sites, 
or allow the medications to be used other than as directed by this protocol without prior 
authorization from DNDi. Under no circumstances can the study specific products 
(appropriately labelled) be replaced by other commercially available product (not 
specifically trial labelled).  
Specific study forms for drug accountability will be designed to allow for adequate 
records on receipt, use, return, loss, or other disposition of medications. These 
accountability forms must be maintained by the study pharmacist or designated 
personnel and stored in the locked room with the study medication.  
7.5. Storage 
Study drugs must be kept in a locked cabinet and/or in a room with restricted access, 
under the control of the study pharmacist.  
Temperature of the storage location will be monitored daily and recorded in a 
temperature log. In the event of temperature excursions below or above the allowable 
range, site staff will inform the study monitor. 
Miltefosine: MF must be stored at 20-25°C; excursions permitted between 15-30°C. 
The product should be stored in the original package and protected from moisture. The 
shelf life is 5 years. Packaging should be undamaged prior to use. 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 38 of 68 
 
Paromomycin: The product should be stored below 30°C and protected from light. 
Partially used ampoules must not be stored for future patient use. The shelf life is 
approximately 3 years. Packaging should be undamaged prior to use. 
Sodium Stibogluconate: The product should be stored below 30°C and protected 
from light. The shelf life is 3 years. The contents should not be used for more than one 
month after opening. 
7.6. Blinding and procedures for unblinding 
This is an un-blinded open label study. 
7.7. Concomitant treatments 
All medications required for concomitant conditions should be postponed until after the 
end of treatment for VL, unless warranted by immediate medical need. Common 
conditions such as malaria and respiratory tract infections should be treated prior to 
starting VL treatment. 
Details of all concomitant medications used during the study will be recorded in the 
patients’ charts and in the CRF, with reason for use and dates of administration. 
Concomitant medications will be coded using the WHO Drug Dictionary Enhanced. 
Drugs that prolong the QT interval (e.g. quinine) should be avoided while SSG is being 
administered. If it is urgently required to give such drugs, SSG should be stopped; 
otherwise alternatives should be sought (e.g. artemisinin derivatives). For MF, there 
are no known interactions with other commonly used medications, though this cannot 
be excluded. While giving PM, the use of other aminoglycosides must be avoided (e.g. 
gentamicin). 
7.8. Rescue treatment 
In case of treatment failure or intolerability (treatment must be permanently 
interrupted), rescue treatment will be provided to the patient, at the discretion of the 
study physician and based on the national VL treatment guidelines. For patients in arm 
1 and 2, rescue treatment is expected to be SSG/PM (SSG 20 mg/kg/day, PM 15 
mg/kg/day) or AmBisome multiple dose (30 mg/kg total dose). In arm 3, rescue 
treatment is expected to be AmBisome multiple dose (30 mg/kg total dose).  
Patients will be given rescue treatment if they have: 
• failed to respond to MF/PM treatment or to SSG/PM reference treatment within the 
treatment period (lack of improvement or worsening of the clinical signs and symptoms 
of VL). 
• failed to respond to MF/PM treatment or to SSG/PM reference treatment at the EOT 
assessments (D28). This will be reflected by positive parasitological assessment 
and/or lack of improvement or worsening of the clinical signs and symptoms of VL. 
• failed to tolerate trial medication / occurrence of adverse event(s) during receipt of trial 
drugs that requires permanent treatment interruption, and rescue treatment is indicated 
by the investigator (see section 9). 
• relapsed, i.e. recurrence of symptoms, signs and presence of parasites after EOT 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 39 of 68 
 
(D28) during the follow up period 
• developed severe post kala azar dermal leishmaniasis (PKDL) that necessitates 
rescue treatment (see section 8.3.4). 
8. Study Assessments 
8.1. Timing of Assessments 
Assessments will be carried out at Screening period (day -7 to 0) and on days 1, 3, 7, 
14, 21, 28, 56 and 210. It will include clinical, parasitological, haematological, 
biochemistry, safety, pharmacokinetic and pharmacodynamics assessments. 
A table with the schedule of events is listed under section 5. 
All patients will be hospitalized for screening/baseline procedures until completion of 
day 14 (arm 1 and 2) and day 17 (arm 3) procedures (approximately 21-24 days).  Day 
28, 56 and 210 assessments will be conducted as outpatient visits. 
Assessments performed during study visit at D28 will have allowable window of +1day, 
due to critical PK assessments at this EOT visit. Therefore, it is acceptable to have 
patients readmitted at the hospital the day before (D27). In the case of patients coming 
from very remotes areas, mobilization will be strengthened and it will be allowed to 
readmit those patients 2 to 3 days before D28 assessments. 
All other assessments performed during treatment period (D1 until < D28) will have an 
allowable window of +/-1 day. In the follow-up period, visit windows will be: day 56 (±7 
days) and day 210 (±14 days). 
8.2. Baseline Assessments 
The following baseline assessments will be done during screening (and after obtaining 
patient consent) in order to confirm VL diagnosis and verify inclusion and exclusion 
criteria. 
• Demographic data including age and sex 
• Medical history including relevant past history, previous VL episode, concomitant 
medications and medication taken in previous 14 days  
• Clinical examination: Physical examination including clinical signs and symptoms 
of VL, PKDL assessment, body weight, height, vital signs (Temperature, Heart rate 
and Blood pressure), spleen and liver size 
• Nutritional status: severe malnutrition will be defined as follows*: 
- 
for children aged <5 years with weight-for-height WHO reference curves by sex, z 
score <-3;  
- 
for patients 5-18 years with BMI-for-age WHO reference curves for sex, z score < 
-3;  
- 
for adults ≥19 years as BMI < 16 
• ECG 
• Audiometric test 
• Clinical safety laboratory evaluations: 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 40 of 68 
 
- 
Haematology: Haemoglobin, total WBC with differential, platelets  
- 
Biochemistry: Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), 
total bilirubin (TBIL), Creatinine (CRE), Albumin (ALB) 
- 
Urine pregnancy test 
- 
HIV test – routine counselling for HIV test will be given for patients’ parents, as part 
of the study informed consent procedure interview or a separate consent will be 
requested, according to the local requirements. Testing will be done according to 
national guidelines and undergo a single rapid diagnostic test for the purposes of 
the clinical trial.  If the RDT for HIV is positive, the patient must be excluded from 
participating in the clinical trial and receive current standard treatment for VL and 
further assessment and treatment for HIV according to the national HIV treatment 
Guidelines. 
- 
Spleen, bone marrow or lymph node aspirate for parasitology (microscopy and 
culture, see additional information in section 8.3.2. Parasitological assessment) 
and pharmacodynamics assessment (qPCR)**  
- 
Blood sample for pharmacokinetics and pharmacodynamics assessment 
* Note: for Ethiopia, severe malnutrition will be defined as follows: 
- 
For patients 4-18 years, MUAC cut-off based on the following MUAC-for-height 
reference table: 
Height 
MUAC cut-off for 
severe malnutrition 
65-<110 cm height 
< 115 mm 
110- <120 cm 
< 120 mm 
120-<130 cm 
< 125 mm 
130-<140 cm 
< 135 mm 
140-<150 cm 
< 145 mm 
150-<160 cm 
< 150 mm 
160-<170 cm 
< 155 mm 
≥170 cm 
< 160 mm 
 
- 
For patients >18 years, MUAC < 170 mm 
MUAC is the circumference of the left upper arm and is measured at the mid-point 
between the tips of the shoulder and elbow. 
** Note: microscopy is mandatory. Culture and qPCR material to be collected if there 
is remaining material after preparing the slides. 
8.3. Assessment of Efficacy 
8.3.1 Clinical assessment of VL 
The clinical evaluation* will involve measuring temperature (axillary), spleen size, liver 
size, body weight at screening, on day 1, 3, 7, 14, 21**, 28 and at all follow-up 
assessments on day 56 and 210. In addition, the following VL symptoms will be 

Protocol number (DNDi-MILT/PM-01-VL)  
 
Miltefosine Paromomycin Phase III Trial 
Protocol Final v5.0 / 13 Mar 2020 
Confidential  
Page 41 of 68 
 
systematically checked at each study visit: fever, abdominal swelling, loss of appetite, 
diarrhea, coughing, epistaxis or other bleeding signs, jaundice. 
The size of the liver will be measured in the midclavicular line for its total span; the 
spleen size will be measured from the left costal margin on the anterior axillary line to 
the tip of the spleen medially. 
* Note: for Ethiopia, clinical evaluation will include measurement of MUAC at screening 
and on day 28, 56 and 210. 
** Note: D21 visit was only applicable to the subset of patients allocated to PK sampling 
in arm 2 (now discontinued). 
8.3.2. Parasitological assessment 
Parasitological assessment will be done at baseline and at day 28 through spleen or 
bone marrow aspirate and microscopic examination.  
Spleen aspirate should not be performed if platelets count is < 40,000/mm3, Hb ≤ 
5g/dL, if the patient has signs of bleeding, jaundice, or if the spleen is not palpable. In 
these cases, bone marrow aspiration is recommended. 
In addition, sites performing splenic aspiration for parasitology must also perform 
prothrombin time as an additional compulsory screening test to determine whether it is 
safe to proceed with splenic aspiration. If there is more than a 5 second difference 
between the normal control value and prothrombin time result obtained, the patient is 
not eligible for splenic aspiration at that point in time. In this case, the investigator 
should, either,   
a. Administer Vitamin K for 3 days and repeat the prothrombin time test on day 
4. If prothrombin time result is within acceptable range (not more than 5 
seconds difference from normal control), and all other parameters for splenic 
aspiration are within the required range, then a splenic aspirate can be done 
OR 
b. Perform bone marrow aspiration instead 
If the site does not have the capability to perform the prothrombin time test, splenic 
aspiration should not be performed, and bone marrow aspiration should be performed 
instead.  
Due to the risk of hemorrhage post spleen aspiration, the patient must remain under 
observation. Record pulse and blood pressure every half hour for 4 h, then every hour 
for 6 h. Patient must remain in bed for 12 hours.  
Patient monitoring post-splenic aspiration should strictly be observed as per the study 
protocol and in-country national VL treatment guidelines. Any suspicion of bleeding 
such as a drop in blood pressure, rising heart rate, abdominal discomfort or pain, or 
any other sign as per investigator’s judgement should be followed by immediate 
emergency response measures as per national VL treatment guidelines. 

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During the follow-up visits at day 56, day 210 or unscheduled visit, parasitology will be 
performed when clinically indicated, i.e. anytime if suspect of relapse (reappearance 
of symptoms and signs of VL).  
Aspirates will be smeared on slides, stained and read under microscope.  
For spleen aspirate, slide reading will be graded as positive or negative according to 
the standard logarithmic criteria as defined in Table 3. 
Table 3 - Standard logarithmic criteria for Visceral Leishmaniasis parasitological 
diagnosis 
 
Count Oil Immersion x 100 
6+ 
> 100 parasites per field 
5+ 
10-100 parasites per field 
4+ 
1-10 parasites per field 
3+ 
1-10 parasites per 10 fields 
2+ 
1-10 parasites per 100 fields 
1+ 
1-10 parasites per 1000 fields 
0 
0 parasite per 1000 fields 
 
Optionally, if there is sufficient tissue aspirate sample remaining after the slide has 
been prepared, this will be processed and used for pharmacodynamics analysis (see 
section 8.4.2.) and culture.  
8.3.3. Clinical Lab assessments related to VL evolution 
Hematology parameters are commonly affected in a patient with VL. At baseline, the 
patient is likely to present anemia, low WBC and possibly low platelets. 
Exclusion criteria are defined as not to include in the clinical trial patients with severe 
anemia (Hb< 5g/dL). In addition, Hb level, platelets and WBC counts will be assessed 
by the investigator together with the clinical presentation and other laboratory 
parameters to judge if patient has signs of severe VL disease, requiring AmBisome 
therapy.    
Hematology parameters must be recorded at screening, D3, D7, D14, D28, D56 and 
D210. It is expected that the hematological parameters recover with time after 
treatment. 
Albumin will also be assessed before and after treatment, at screening, Day 28 and 
Day 210. 

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8.3.4. Post-kala-azar dermal leishmaniasis (PKDL) 
Patients will be monitored closely for post-kala-azar dermal leishmaniasis (PKDL) 
through the course of the study. Diagnosis will be made clinically based on the typical 
appearance and distribution of the rash. Presence of PKDL will be assessed at 
screening, on day 28 and during follow-up assessments on day 56 and day 210. 
Grading will be noted during the assessment times as follows:  
• Mild (Grade 1): lesions mainly on the face and head, with others scattered on arms, 
chest and back  
• Moderate (Grade 2): the upper part of the chest and arms are also affected  
• Severe (Grade 3): lesions may be found on the whole body.  
Patients who develop severe PKDL (grade 3) or have mucosal and/or eye involvement 
(any grade) will require treatment with rescue medication at the discretion of the 
attending physician.  
8.4.  Pharmacokinetics and Pharmacodynamics Assessments  
8.4.1. Pharmacokinetic Assessments 
Patients in arm 1 will have pharmacokinetics measurement of MF at D14, whereas all 
patients in arm 1 and 2 will have PK measurement of MF at D28 and D56 in order to 
verify adherence to the treatment.   
In addition, intensive sampling for both PM and MF PK will be performed in a subset 
of patients defined as the intensive cohort: originally 40 in each MF/PM combination 
arms, with selection of 20 pediatric (≤12 yrs) patients and 20 adolescent/adult patients 
(>12 to ≤50 yrs). Because arm 2 is discontinued, the number of patients enrolled in this 
arm in the PK/PD intensive cohort will most probably not reach 40, however samples 
collected will still be analysed. Enrollment in this intensive cohort will be limited to 
specific sites located in Sudan and Kenya and as far as possible should be balanced 
between the two countries. MSF sites will not participate to the enrollment in the 
intensive PK/PD cohort. These sites will be selected based on their experience, 
equipment and resources available. Since previous trials (LEAP0714 and LEAP0208) 
have raised questions on both the (reduced) absorption rate, but also extent of 
absorption (bioavailability), of MF in East African VL patients, denser sampling is 
required in this trial than previously performed for MF pharmacokinetics. Due to the PK 
nonlinearities observed in pediatric patients receiving the allometric MF dosing 
(LEAP0714), the inclusion of children in this PK sub-study is important and therefore 
the aim is to have equal numbers of patients below 12 years and above 12 years 
included in the PK sub-study, to allow for comparison. For PM, we have included a 
sparse sampling scheme to enable assessment of pharmacokinetic profiles, with 
subsequent use of population PK modelling to analyze the data. PM PK will be 
assessed on both day 1 and day 14 of treatment to assess the influence of changing 
patient characteristics during the treatment period (e.g. increasing body weight, 
reduced liver damage, etc.) on the PK of PM and to assess potential accumulation of 
the drug during the treatment period.  

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In order to avoid multiple blood sampling in young children, only children from 7 years 
of age and weighing more than 20 kg will be included in the PK intensive cohort. 
To reduce the number of total samples/blood volume per patient in the intensive cohort, 
a mixed sampling approach will be performed. Patients will be randomly allocated to 
sparse sampling schemes to allow for a pooled PK analysis. 
As much as possible, a single blood draw should be performed per study visit to avoid 
multiple vein punctures. Sample for both MF and PM PK will be in general a 2mL EDTA 
whole blood sample/time point. For time-points where only MF PK is to be assessed, 
also 2 ml EDTA whole blood sample/time point will be collected to accommodate with 
the lowest volumes of tubes currently available on the market.  
The pharmacokinetics of both PM and MF will be determined using validated liquid 
chromatography–tandem mass spectrometry (LC-MS/MS) bioanalytical assays (PM to 
be validated). The pharmacokinetics of both drugs will be determined in plasma, unless 
a dried-blood-spot method is developed for PM prior to initiation of this trial.   
Samples will be stored on site at minimally -20°C and regularly shipped under frozen 
conditions to the respective capitals for shipment to Amsterdam, the Netherlands. 
Samples will be analysed for their MF/PM content at the Bioanalytical Laboratory at 
the Netherlands Cancer Institute – Antoni van Leeuwenhoek Hospital, following a 
validated procedure using liquid chromatography coupled to tandem mass 
spectrometry (LC-MS/MS). The development and validation of the assay for MF has 
previously been published (Dorlo et al., 2008). A bioanalytical assay for paromomycin 
in human plasma has been described in literature previously and will be validated 
during the progressing review period and implementation of this protocol.    
 
8.4.1.1. 
Paromomycin pharmacokinetic assessments 
PM PK assessments will be performed only in the intensive cohort (subset of 40 
patients in each MF/PM combination arms) at Screening, D1 and D14. The screening 
sample can be collected at any day during screening period. PM PK sample at D1 and 
D14 will be collected at 0, 1, 2, 4, 8, 24 hrs post-dose.  
In order to minimize blood volume collected, each patient will be randomly allocated to 
one of the following sparse PK sampling schedules:  
- 
D1: either 1, 2, 4, 24h or 1, 2, 8, 24h. 
- 
D14 (arm 1 and 2): either 0, 1, 2, 4, 24h or 0, 1, 2, 8, 24h. 
Time of drug administration and time of PK samples collection have to be recorded 
systematically.  
8.4.1.2. 
Miltefosine pharmacokinetic assessments 
MF PK assessments will be performed at D14 for subjects in arm 1 and at D28 and 
D56 for all subjects in arm 1 and 2 to confirm adherence to treatment. Subjects who 
consent to participate to the intensive cohort will have MF PK assessments performed 
at Screening, D1, D7, D14, D28 and D56 in both arms and an additional assessment 

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used to be done at D21 in arm 2 only (not applicable anymore in this protocol version). 
The screening sample can be collected at any day during screening period or before 
the administration of the 1st dose at D1. 
In the intensive cohort, MF PK sample at D1 and D14 will be collected at the same 
time points as PM PK at 0, 1, 2, 4, 8 and 24h (according to the sparse sampling scheme 
the patient is allocated to). The 8h MF PK sample will be collected always before the 
2nd daily dose. D7 and D21 (arm 2 only, not applicable in this protocol version) PK 
samples will be collected prior to the morning dose. D28 PK sample used to be 
collected prior to the morning dose in patients allocated to arm 2 and will be collected 
in the morning in patients allocated to arm 1.  In the follow-up period (D56), PK sample 
shall be collected in the morning. Time of drug administration and time of PK samples 
collection have to be recorded systematically.  
8.4.2. Pharmacodynamic Assessments 
Parasite load in blood and tissue aspirate (bone marrow or spleen): PD assessment 
will be based on real time quantitative polymerase chain reaction (qPCR) for L. 
donovani in EDTA blood or tissue aspirate, based on the amplification of kinetoplastid 
DNA (kDNA) (other Leishmania DNA targets may also be considered), to assess the 
parasite clearance over time during the treatment with two combination regimen of PM 
and MF or treatment with SSG/PM. 
All patients will have qPCR in tissue and blood at screening and at Day 28 assessment; 
and during follow-up if there is suspicious of relapse. 
In addition, the patients (40 in arm 1 and probably fewer patients in arm 2 now this arm 
is discontinued) in the intensive cohort who consent to have multiple sampling for PK 
will also have qPCR blood samples collected at D0, 3, 7, 14, 28, 56 and 210. 
Approximately 1 ml EDTA whole blood will be immediately separated from blood EDTA 
samples taken (e.g. for Complete Blood Count or PK analysis) for the qPCR analysis. 
The blood samples and also the remainder of bone marrow/spleen aspirate sample 
(optional) will be immediately diluted with DNA stabilizing buffer solution (L6 buffer) 
and can then be stored up to 2 years at frozen temperature.  
Furthermore, additional relevant PD markers will be considered for analysis in order 
to monitor treatment effect. 
8.5.  Assessment of Safety  
Safety of the different treatment regimens will be assessed through routine monitoring 
of adverse events. At each study visit, the patients will be enquired about current 
adverse events or any events observed during the period previous to the visit. In 
addition, evaluation of hematology and blood chemistry parameters, regular 
measurement of vital signs and physical examinations will be made at scheduled 
follow-up visit. 
Audiometric test will be performed at baseline and on days 14, 28 and 210 for all 
patients. 
ECG will be performed at baseline and on day 14 in all arms, and in addition on days 

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7 and 28 in the reference arm SSG/PM. SOPs will be developed to guide investigators 
on audiometry and ECG assessments. 
Urine pregnancy tests will be performed in woman of child-bearing potential at baseline 
to exclude any pregnancy, and on D28, D56 and D210 visits to monitor eventual 
unexpected pregnancy. 
HIV test will be repeated at the D210 visit only for patients in Gondar and Abdurafi 
sites, due to high prevalence of HIV/VL co-infected patients (15%) in this specific 
population. 
The frequency and severity of AEs will be described, as well as frequency of SAEs or 
AEs that lead to treatment discontinuation. See section 8.6 for AE definition and 
reporting. 
8.5.1. Laboratory examinations  
Hematology parameters hemoglobin, WBC with differential and platelets will be 
analyzed at screening, on days 3, 7, 14, 28, 56 and 210.  
Biochemistry parameters ALT, AST, TBIL and CRE will be analyzed at screening, on 
days 3, 7, 14 and 28. 
Samples will be analyzed at the local laboratory using standardized equipment. 
Approximately 3 ml of blood will be collected at each visit: 1 ml in EDTA for the 
hematology and 2 ml in dried tube for the biochemistry. 
Lab parameters abnormal values will be assessed for clinical significance and will be 
graded according to CTCAE v4.03 (see section 8.6.7). 
 
8.6. Adverse event definitions and reporting 
8.6.1. Adverse Event definition 
Any untoward medical occurrence in a clinical trial subject administered a medicinal 
product and which does not necessarily have a causal relationship with this treatment.  
It can therefore be any unfavourable and unintended sign (e.g. an abnormal laboratory 
finding), symptom, or disease temporally associated with the use of a medicinal 
product, whether or not considered related to the medicinal product. 
Definition of an AE includes worsening (in severity and frequency) of pre-existing 
conditions (“Medical history”) before first Investigational Medicinal Product (IMP) 
administration and abnormalities of procedures (i.e. ECG, X-ray…) or laboratory 
results which are assessed as “clinically significant” (see details in section 8.6.2). 
What is not an AE? 

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• Medical conditions present at the initial study visit that do not worsen in severity or 
frequency during the study are NOT considered as AE. 
• Symptoms, exacerbation or worsening of the studied disease will NOT to be 
considered as AE nor captured on the AE page of the CRF if consistent with the 
anticipated natural progression of the disease (overall and for this given subject). 
• Lack of efficacy of the IMP is NOT considered as AE. 
 
8.6.2. Assessment of laboratory abnormalities 
For every laboratory assessment, the investigator will evaluate if the lab test is normal 
or abnormal. If abnormal, the investigator will assess if this finding is clinically 
significant or not. If a lab parameter is abnormal and clinically significant, it should be 
reported as an adverse event (AE).  
An abnormal lab test must be compared with the previous value.  
An AE is a new event after the administration of the 1st dose of the study drug or a 
worsening in the condition (in the case of lab tests, increase in severity by CTCAE 
v4.03) which is judged clinically significant by the investigator. 
Laboratory abnormalities should be assessed as “clinically significant” (and therefore 
have to be reported as an AE) only if they meet AT LEAST ONE of the following 
conditions:  
• The abnormality suggests a disease and/or organ toxicity AND this abnormality was 
not present at the screening visit or is assessed as having evolved since the 
screening visit 
• The abnormality results in discontinuation of the study drug 
• The abnormality requires medical intervention or concomitant therapy 
Lab parameters which are classified with regards of severity as CTCAE v4.03 Grade 
3 or Grade 4 must be carefully assessed, as they are likely to require medical 
intervention and/or be life-threatening in terms of severity.   
For the purpose of this protocol, all clinically significant Grade 3 and Grade 4 lab 
abnormalities must be reported as an AE. 
When reporting an abnormal lab, a clinical diagnosis should be recorded rather than 
the abnormal value itself, if available (for example, “hypokalemia” rather than 
“decreased potassium levels”). Furthermore, lab abnormalities which present with 
clinical signs and symptoms will also be considered clinically significant. In these 
cases, again the adverse event should be recorded as the syndromic clinical diagnosis 
rather than clinical signs and symptoms (ex. acute pancreatitis instead of each finding 
separately: high levels of amylase, high levels of lipase, abdominal pain and vomiting). 
For any abnormal clinically significant lab value, the investigator will carefully assess if 
the event is due to the natural disease progression (e.g. anaemia during the first days 
of treatment). If the abnormality is associated with the anticipated VL disease 
symptoms, it does not need to be reported as an AE, unless if it meets the criteria of a 

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clinically significant event.  
8.6.3. Serious Adverse Event 
An adverse event will be defined as serious if it  
− results in death, 
i.e. causes or contributes to the death. 
− is life-threatening, 
in this context refers to an AE/Adverse Reaction (AR) in which the patient was at 
risk of death at the time of the AE/AR; it does not refer to an AE/AR that 
hypothetically might have caused death if more severe. 
− requires in-patient hospitalisation or prolongation of existing hospitalisation, 
i.e. the AE requires at least an overnight admission or prolongs a hospitalization 
beyond the expected length of stay.  
In this protocol, the patient is expected to be hospitalized for 14 days, with 
discharge expected on D15. If the patient presents an AE that requires 
hospitalization for more than 24hs after D15, this event will be considered an SAE. 
However, if the patient remains hospitalized for social reasons, it will NOT be 
considered as an SAE.    
Hospital admissions or prolongation of hospitalisation for surgery planned before 
study entry, for social reasons, for any elective surgery (i.e. plastic surgery) or for 
normal disease management (including treatment adjustment) are NOT to be 
considered as SAE according to this criterion (i.e. if the protocol or the standard 
management of the disease under study requires planned hospitalization). 
− results in persistent or significant disability or incapacity, 
i.e. the AE resulted in a substantial disruption of the subject’s ability to conduct 
normal activities. 
− is a congenital anomaly / birth defect, 
i.e. an adverse event outcome in a child or foetus of a subject exposed to the 
Investigational Medicinal Product (or marketed medicinal product) before 
conception or during pregnancy. 
− is an important medical event, i.e. is medically significant. 
Medical and scientific judgment should be exercised in deciding whether other 
situations should be considered serious events/reactions, such as important 
medical events that might not be immediately life-threatening or result in death or 
hospitalization but might jeopardize the patient or might require intervention to 
prevent one of the other outcomes listed above.  

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In addition, any suspected transmission via a medicinal product of an infectious 
agent is also considered a serious adverse event/reaction. 
Note: SAE onset date is the SAE start date or date a pre-existing AE becomes serious. 
8.6.4. Eliciting Adverse Event information 
The investigator is required to report all directly observed adverse events and all 
adverse events spontaneously reported by the trial subject using concise medical 
terminology. In addition, each trial subject will be questioned about the occurrence of 
adverse events at each study visit (scheduled and non-scheduled) since the time of 
the last study visit, as well as follow-up from previously recorded adverse event. 
8.6.5. Management of Adverse Events 
All adverse events will be managed by the investigators according to local treatment 
guidelines. The sites are equipped with appropriate resources to treat adequately the 
adverse events. However, in case the patient cannot be managed on site, he/she will 
be referred for specialized care in the nearest referral hospital. 
 
8.6.6. Adverse Event reporting period 
The adverse events reporting period begins upon subject enrolment in the trial (after 
signature of informed consent) and ends at the end of the subject participation in the 
trial (D210 follow-up visit).  
In this protocol, the reporting period is different for AEs and for SAEs: 
• Non-serious adverse events: All non-serious AEs will be reported upon 
administration of the first dose of trial medication until the end of subject 
participation in the trial (including post-treatment follow-up period up to D210). 
Furthermore, non-serious AEs that occur in the screening period (from signature 
of the informed consent) AND are judged as study-related (e.g. hematoma due to 
venipuncture) will be reported in the AE CRF.   
• Serious adverse events: All SAEs must be reported upon subject enrolment in the 
trial (after signature of informed consent during screening period) until the end of 
the subject participation in the trial (including post-treatment follow-up period up to 
D210). 
• Screening failure: beyond the date of screening failure (to be recorded), only 
serious study-related events will be followed-up. 
All adverse events that occur during the adverse event reporting period specified 
in the protocol must be reported to DNDi, whether or not the event is considered 
medication related. In addition, any adverse event that occurs subsequent to the 
adverse event reporting period that the investigator assesses as possibly related 
to the investigational medication should also be reported as an adverse event. 
8.6.7. Adverse Event reporting requirements 

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Information on adverse events must be evaluated by a physician. Each adverse event 
is to be classified by the investigator as serious or non-serious. This classification will 
determine the reporting procedure for the event to DNDi and Health Authorities/Ethics 
Committees (as per local regulatory requirements). 
All serious adverse events (SAE) are to be reported immediately (within 24 hours of 
awareness of SAE by the investigator) to DNDi (SAEMFPMstudy@dndi.org), using the 
SAE report form.  This includes a description of the event, onset date and seriousness 
criteria, duration, severity, relationship to study drugs, outcome, measures taken and 
all other relevant clinical and laboratory data. The initial report is to be followed by 
submission of additional information (follow-up SAE form) as it becomes available. Any 
follow-up reports should be submitted as soon as possible, and if possible within 5 
working days.  
In addition to immediately reporting SAEs to DNDi, Investigators are responsible for 
reporting SAEs occurring at their site to the Ethics Committee (IEC), and any periodic 
safety reporting if applicable, in compliance with the local regulatory requirements. 
Serious adverse events should also be reported on the clinical trial adverse event case 
report form (CRF). It should be noted that the form for reporting of SAE (SAE form) is 
not the same as the adverse event section of the CRF. Where the same data are 
collected, the two forms must be completed in a consistent manner, and the same 
medical terminology should be used. 
Non-serious adverse events are to be reported on the CRF, including description of 
the event, onset date, duration, severity, seriousness, relationship to all study drugs, 
actions taken and outcome. In the CRF, a given adverse event will be recorded only 
one time per patient, and the severity recorded will be the maximum level reached. If 
several distinct episodes of the same condition occur, their number will be recorded in 
the CRF. 
 
8.6.8. Grading of Adverse Event severity 
Severity is a clinical determination of the intensity of an AE. The severity for an AE 
should be graded using the National Cancer Institute’s Common Terminology Criteria 
for Adverse Events (CTCAE, version 4.03).  
In case of AEs that are not described in the CTCAE v4.03, the investigator will use the 
terminology MILD, MODERATE, SEVERE, LIFE-THREATENING or DEATH to 
describe the maximum severity of the adverse event as follows:  
MILD (grade 1) 
 
The subject is aware of the event or symptom, but the event 
or symptom is easily tolerated (e.g. no reduction in daily activities is required) 
MODERATE (grade 2) 
The subject experiences sufficient discomfort to interfere 
with or reduces his or her usual level of activity. 
SEVERE (grade 3)  
Significant impairment of functioning: the subject is unable 
to carry out usual activities and/or the subject’s life is at risk from the event 

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LIFE-THREATENING (grade 4) The subject is at risk of death at the time of the adverse 
event. It does not refer to an AE that hypothetically might have caused death if more 
severe (Life-threatening consequences, urgent intervention required) 
DEATH (grade 5) 
 
Death related to AE 
This information on AE grading will be entered in the adverse event CRF. 
It is to be noted the distinction between severity (intensity) and seriousness (regulatory 
definition) of adverse events. A severe adverse event is not necessarily a serious 
adverse event. 
8.6.9. Adverse Event causality assessment 
For both serious and non-serious adverse events, the investigator is required to assess 
if there is a causal relationship between the adverse event and all the study drugs, i.e. 
to determine whether there is at least a reasonable possibility that one of both study 
drugs caused or contributed to the adverse event. This means that there are facts and 
arguments to suggest a causal relationship.  
To help investigators with the decision binary tree in the evaluation of causality, the 
CIOMS VI group recommends that investigators be asked to consider the following 
before reaching a decision: 
• Medical history 
• Lack of efficacy/worsening of existing condition 
• Study medications 
• Other medications (concomitant or previous) 
• Withdrawal of study medication, especially following trial discontinuation / end of 
study medication 
• Erroneous treatment with study medication (or concomitant) 
• Protocol related procedure 
Using this information the investigator will classify each adverse event as: 
Not related:   There is no reasonable possibility of causal relationship  
between an AE and study drug 
Related:  
There is at least a reasonable possibility of a causal relationship 
between an AE and a study drug. This means that there are facts 
(evidence) or arguments to suggest a causal relationship. 
The decision to suspend, and resume treatment or to permanently interrupt treatment 
due to an adverse event will be left to the clinician in charge. 
8.6.10. 
Exposure in utero 

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All female patients of child bearing age will be excluded unless they agree to undergo 
a pregnancy test and explicitly consent to take an acceptable method of contraception 
(i.e. Depo-Provera). Urine pregnancy tests will be performed in woman of child-bearing 
potential at baseline to exclude any pregnancy, and on D28, D56 and D210 visits to 
monitor eventual unexpected pregnancy. All pregnant women will be excluded from 
the study. This criterion was set because of the teratogenic potential of MF 
(demonstrated in rats). The effective contraception should be used for a minimum 
period of 5 months after end of treatment with MF. Therefore, Depo-Provera injection 
will be required before treatment onset and around day 90 to cover for the entire 
exposure period. 
However, in the unlikely event that any trial subject becomes or is found to be pregnant 
while receiving any of the study treatment drugs or during follow-up period, the 
investigator must submit the event on a Pregnancy report form. The information 
submitted should include the anticipated date of delivery. 
The investigator will follow the subject until completion of the pregnancy or until 
pregnancy termination (i.e., induced / spontaneous abortion). The investigator will 
provide pregnancy outcome information on a Pregnancy outcome form.  
A pregnancy is not an SAE. 
In the case of a live birth, a medically qualified person should assess the infant at the 
time of birth and submit a Child report. An SAE should be declared in the case of 
unfavorable pregnancy outcome (abortion, still birth) or congenital abnormality (in 
addition to the Child report). 
8.6.11. Adverse event follow up 
All adverse events should be followed until they are resolved or the investigator 
assesses them as chronic or stable or the subject participation in the trial ends (i.e., 
until a final report is completed for that subject).  
In addition, all serious adverse events and those non-serious events assessed by the 
investigator as possibly related to the investigational drug must continue to be followed 
even after the subject participation in the trial is over. Such events should be followed 
until they resolve or until the investigator assesses them as “chronic” or “stable.” 
Resolution of such events is to be documented on the CRF and SAE form. 
9. Withdrawal criteria 
A subject should be withdrawn from the study treatment if, in the opinion of the 
investigator, it is medically necessary, or if it is the wish of the subject.  
If a subject does not return for a scheduled visit, every effort should be made to contact 
the subject.  
In any circumstance, every effort should be made to document subject outcome, if 
possible. 
If the subject withdraws consent, no further evaluations should be performed and no 
attempts should be made to collect additional data, with the exception of safety data, 

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which should be collected if possible and in accordance with patient consent. 
If a subject withdraws from the study, the reason must be noted on the CRF.  
If a subject is withdrawn from the study because of a treatment limiting adverse event, 
thorough efforts should be made to clearly document the outcome of AE. 
9.1. Rules in case of treatment suspension or interruption 
In the event that treatment is interrupted (e.g. due to an adverse event), the decision 
to resume treatment will be taken by the site Principal Investigator.  
If treatment interruption lasts > 72hs due to an AE related to MF, or if treatment needs 
to be interrupted due to an AE related to PM or SSG, rescue treatment will be 
considered, at the discretion of the investigator. 
Once rescue treatment is given, the patient will be considered a treatment failure, and 
will be withdrawn from the study. 
If rescue treatment is not indicated, the patient may continue to participate in the study, 
and perform the scheduled visits and procedures.   
9.2. Rules for permanently interrupting study treatment 
If a subject is withdrawn from the study before the full course of the treatment is 
completed, the physician must make all necessary arrangements to ensure that the 
subject receives the appropriate treatment for the relevant medical condition (e.g. with 
drug/s currently recommended by the national policy). 
9.3. Subject withdrawal from the study and subject replacement 
Subjects withdrawn from the study will not be replaced. 
10. Data Analysis and Statistical Methods 
10.1. Sample size determination 
Samples size calculation is based on the efficacy (definitive cure) at 6 months (D210) 
which is the primary endpoint and main assessment criterion of interest. 
The initial sample size assumptions were based on an expected efficacy at the end of 
6 months follow-up of 93% for the tested combination and 91% for the reference arm 
SSG-PM, a non-inferiority margin of 7%, and 1.25% alpha (one-sided adjusted for 
multiplicity), the sample size required to reach a power of at least 80% per comparison 
of interest (each investigational arm vs SSG-PM) was 173 patients per arm rounded to 
175. With the provision for lost to follow-up (LTFU), the original sample size for each 
arm was 192 (total of 576 patients). 
In this revised protocol, the analysis will remain as planned, but comparison will be 
performed only between one investigational arm (arm 1 MF/PM 14 days) and the 
standard of care SSG-PM. Because the number of comparisons is now limited to one, 
there is no multiplicity of testing issue and therefore, no need for adjustment of type 

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one error if the change in the protocol is not linked to an interim analysis or a shift to 
an adaptive design such as “drop the looser”. Reasons for stopping a dose regimen is 
not based on significant data but rather related to the paucity of patients fulfilling 
inclusion criteria. Moreover, the number of patients who reached the primary time point 
was around 10% of the planned sample size and considering the expected success 
rate (93%), the number of observed failures cannot be informative unless it would have 
been a major discrepancy with expectation. Even if collected data are considered 
informative, the use of O’Brien Fleming curve in the alpha spending function when 10% 
of the sample size was available indicates that the fraction of alpha spent at the 
potential interim analysis becomes quite negligible at the final analysis. Consequently, 
no adjustment of alpha was considered necessary in the current situation. Using same 
assumptions for efficacy and non-inferiority margin, a power of at least 80%, and 
considering an alpha of 0.025 (one-sided, no adjustment for multiplicity) the required 
sample size is 153 patients per arm. A 10% provision for potential LTFU to ensure a 
power of at least 80% brings the sample size to 170 patients per arm (1 and 3). 
The total number of patients included in the trial will be approximately 420, taking into 
account the number of patients randomized to treatment arm 2 before discontinuation 
of this arm (approximately 80 patients).    
10.2.   Definition of study populations included in the analysis 
The following patient populations will be defined for analysis: 
• Intention-To-Treat (ITT): all randomized patients in the arms to which they were 
randomly assigned regardless of their adherence with the inclusion-exclusion 
criteria, the treatment they received, and subsequent withdrawal from treatment or 
deviation from the protocol. 
• Modified Intention-To-Treat (mITT) or Full Analysis Set (FAS): All randomized 
patients receiving at least one dose of treatment. In case of error of treatment 
allocation, the actual treatment received will be used in the analysis. Modified ITT 
set of patients will be used in the primary efficacy analysis and in safety analysis. 
• Set of completers: subset of patients belonging to the FAS, who attend the 6-
months follow-up visit. This set will be used in a sensitivity analysis of the primary 
efficacy analysis. This analysis will be based on the definitive cures.  
• Per protocol (PP) set of patients: subset of patients belonging to the FAS and who 
are free from major protocol violations. Major protocol violations are any violation 
which might bias the result of the non-inferiority test. Major violations will be defined 
a priori in the statistical analysis plan. They will be identified and documented 
during the blind review of data. This set of patients will be used in the primary 
sensitivity analysis.           
The primary population for efficacy analysis at D210 will be the modified intention to 
treat population (mITT or FAS). Furthermore, per-protocol set of patients and the set 
of completers sensitivity analyses will be performed. 
All patients enrolled in the study and who have been administered the first dose of 
study medication will be included in the safety analysis. 

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For the secondary endpoint on PK, all subjects exposed to at least one dose of the 
medication and who have a PK sample collected will be included in the analysis. 
Patients’ data randomized in arm 2 (PM 14 days/MF 28 days), which recruitment is 
now discontinued, will be summarized in a descriptive analysis, but not included in the 
comparative efficacy analysis. 
10.3. Subject Disposition  
The number of patients who were screened, failed screening, randomized, completed 
the treatment period, completed the follow-up, early terminations and reasons for early 
termination of study will be summarized by number of patients (n) and 
percentages (%). 
10.4. Baseline  
Routine baseline characteristics including age, sex, height, weight, BMI for age*, and 
symptoms (fever, weight loss, epistaxis, etc.) will be documented and summarised as 
appropriate according to data variable type (e.g. mean for normally distributed 
continuous data). 
* Note: for Ethiopia, MUAC will be documented in addition to BMI for age 
 
10.5. Treatment Compliance  
All test drugs will be supervised on administration by study medical staff. The dose 
given will be recorded in the patient dispensing log and in the CRF. Full compliance 
will be considered if 90-110% of the prescribed dose is administered. 
10.6. Efficacy Analysis 
Primary endpoint:   
- Definitive cure or treatment success:  
o absence of signs and symptoms of VL at D210,  
o no rescue treatment at any time up to D210 (included). 
- Treatment failure:  
o Patient who requires rescue treatment due to any of the following: 
- 
Initial failure;  
- 
occurrence of adverse events that leads to treatment discontinuation 
- 
relapse at any time; 
o death during treatment period or death during follow-up period that is related to VL 
o Lost to follow-up at D56 or before.  
- Unconfirmed status at D210:  

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o Patient who did not fail at day 28 (initial cure) and at day 56 (probable cure), did 
not relapse before D210 (no rescue treatment, no signs and symptoms of VL 
before day 210) and did not attend day 210 visit. 
Definitions: 
Initial failure: patient who did not respond to the treatment and required rescue 
therapy within the 28 days treatment period; or patient who had a positive parasitology 
(by microscopy) at the D28 assessment. 
Relapse: patient who had responded to treatment and had a negative parasitology at 
the D28 assessment, but who presents signs and/or symptoms of VL during follow-up, 
with VL confirmed by presence of parasites in a parasitological investigation.  
Secondary efficacy endpoints: 
Initial cure - cure at the end of treatment (Day 28), defined as recovery of clinical signs 
and symptoms; absence of parasites (microscopy) and no rescue treatment 
administered at Day 28 or before. 
Probable cure - absence of clinical signs and symptoms of VL at D56 and no prior 
requirement for rescue treatment. 
Primary Efficacy Analysis 
The primary efficacy analysis will be based on  
• the modified intention to treat population (mITT or FAS); 
• the primary endpoint (cure at D210); 
• the primary imputation approach (see below) and the primary test of non-inferiority, 
which is the Blackwelder test. 
• primary comparison: MF-PM 14 days vs SSG-PM.  
The significance limit will be 0.025 (one-sided). If the non-inferiority test is significant 
the test of superiority will be performed.  
Handling of unconfirmed status 
The handling of unconfirmed status can be based on imputing a failure to all 
unconfirmed status and in that case the success rate is probably underestimated (ITT 
worst-case scenario). Another approach is to consider all cases of probable cure at 
D56 as definitive cure at D210.  This is equivalent to the use of last observation carried 
forward approach or even considering an absence of proven failure as a success. This 
approach is probably overestimating the success rate. A third intermediary approach 
consists in imputing a probability of success which is neither a failure (Imputed 
probability of success = IPS = 0) nor a success (IPS = 1) but an imputed probability of 
success (0 < IPS < 1). This is the basic idea leading to the primary imputation method. 

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The estimated probability of success (PS) is equal to 1 minus the probability of relapse 
(PR) between day 56 and day 210. The PR is estimated from the sample of all patients 
irrespective of treatment group who were a probable cure at D56. PR is equal to the 
number of relapses between D56 and D210 divided by the number of probable cure at 
D56 who attend D210 visit.  
The imputed probability of success (IPS) will be set at 1 − 2PR and the number of 
imputed success in a given treatment group will be equal to the number of unconfirmed 
status multiplied by 1 − 2PR. Because the Blackwelder test is based on binary data, 
the value (1 – 2Pr) will be rounded to the lower whole number (number of successes 
in patients with unconfirmed status) and added to the number of successes in other 
patients. 
The relapse rate (PR) is multiplied by two to impose a penalty when the status at D210 
is unknown.  Considering that study arms 1 and 3 in this non-inferiority trial are 
expected to have similar true relapse rate after day 56 efficacy at D210 (similar 
proportion of relapses), the imputation method will consider pooled PR as the best 
estimation of failure rate. If the MF/PM combination has a smaller relapse rate, then 
the measure of the effect size will be in disfavor of this combination.    
In addition, the following sensitivity analyses will be performed: 
1) Replacement of the primary imputation approach by the conservative approach 
(LTFU = failure). 
2) Replacement of the primary imputation approach by the optimistic approach (LTFU 
= success). 
3) Replacement of the primary imputation approach by the realistic approach (LTFU 
have the same rate of relapse as completers: PS = 1 − PR). 
4) Replacement of the mITT by the per-protocol set of patients.     
5) Replacement of the mITT by the set of completers. This analysis avoids any 
imputation but is based on a subset of patients. The result should be close to the third 
sensitivity analysis if there are not many patients with unconfirmed status. 
Justification of the non-inferiority margin 
 
The non-inferiority margin of 7% has been defined after consultation with experts. It is 
justified by the following advantages of the tested combination arm over the reference 
arm: replacing SSG with oral MF will improve treatment safety by removing the toxic 
antimonial component of the current SSG-PM treatment; it will be a more patient-
friendly treatment by replacing one of the two daily injections with an oral 
administration; and hospitalization time will be reduced by at least 3 days. Finally, the 
new combination will ensure suitability in remote areas where VL occurs. 
Shift from non-inferiority to superiority 
If the tested combination is significantly non-inferior to the reference arm, the test of 
superiority will be performed (Likelihood ratio test) without adjustment of type error. 

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The result of this test as well as the 95% confidence interval of the excess success 
rate will be used to assess the benefit risk of the tested combination with regards to 
the reference arm but not to claim the superiority. The analysis will be the same as the 
primary analysis after replacing the non-inferiority test by the superiority.  
Interim analysis 
No interim analysis (IA) will be performed in this trial.   
10.7.  Safety Analysis 
All patients enrolled and who received at least one dose of study medication (mITT set 
of patients) will be included in the safety analysis (treatment-emergent AEs). The 
incidence rate of the most pertinent and frequent ADR (> 5%) of each treatment group 
will be compared and tested. No adjustment for multiplicity will be performed. These 
results will be used for assessing the benefit risk ratio. 
The proportion of patients with SAE and/or AEs leading to treatment discontinuation 
will be described per System Organ Class (using preferred terms defined by MedDRA), 
and summarised according to relationship to treatment and severity (CTCAE, version 
4.03).  In addition, a narrative for each of the SAE and AE leading to treatment 
discontinuation will be developed describing all aspects of the medical event. 
The proportion of patients presenting at least one treatment emergent AE (TEAE) will 
be presented. All TEAEs will be described per study arm using the same classification 
as presented above. AEs will be tabulated by severity (CTCAE V4.03), and causality.  
Abnormalities of laboratory/procedures results will be tabulated with respect to clinical 
significance. In addition, shift tables and graphs will be generated to describe the 
laboratory parameter values over time, per treatment arm. 
10.8.  PK Analysis 
Initially, a standard two-stage non-compartmental analysis will be performed using the 
ncappc script in R to derive the following basic descriptive PK parameters for MF and 
PM in the intensive PK cohort: Cmax, tmax, AUC0-last time-point, AUC0-end of treatment (MF). 
Groups of children ≤ 12 years and patients >12 years will be compared with each other. 
All results will be descriptively compared to historic data of published MF PK and 
unpublished PM PK data from the region. Subsequently, in a later phase, a population 
PK approach will be employed to derive better estimates of drug exposure and also to 
obtain standard primary PK parameters using a structural compartmental model. Using 
this model, a more sophisticated covariate analysis will be performed to assess the 
influence of combining both drugs on each other’s PK parameters and additionally 
other time-constant and time-varying patient characteristics. Also sparse MF PK 
samples from all patients (in contrast to only the intensive cohort) will be included as 
well in such a population PK analysis. Exact details and procedures of this population 
PK analysis will be described in a population PK analysis plan. Such a structural 
population PK model will also allow for better assessment of exposure-response 
relationships, which will focus on linking individual exposure parameters to initial cure, 
final cure/relapse and data on parasite levels (both microscopy and qPCR) and 
potentially other measured markers of clinical improvement in patients.  

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10.9.  PD Analysis 
In this trial the blood parasite counts will be evaluated as a pharmacodynamic indicator 
to derive the systemic parasite clearance rate and thus as an experimental measure 
of response to treatment. Individual or pooled average (depending on the individual 
quality of data) parasite clearance rates will be derived using log-linear regression on 
positive parasite counts. This rate will be correlated to treatment outcome to evaluate 
the predictive value of this marker for the initial and final treatment outcome. 
Subsequently, using the developed population PK models, these will be linked in a 
sequential manner to the pharmacodynamic parasite counts. By following parasite 
clearance using qRT-PCR and evaluating PK of MF and PM, population PK-PD 
modeling will enable us to further explore the relationship between drug exposure and 
parasitological outcome in a more mechanistic way, which can be seen as an essential 
component in the development of new treatment regimens. Clinical outcome and 
measures (spleen and liver size), appropriate biochemistry values and other selected 
biomarkers will also be integrated as possible influential covariates in the PK-PD 
model, as appropriate. Eventually, the population PK-PD model will be used to identify 
which PK parameter and/or measure of exposure is critically relating to therapeutic 
response for MF and PM, which will be of pivotal value in defining the optimal treatment 
duration and intensity of MF in future combination regimens. 
 
11. Data Safety Monitoring Board 
A Data Safety Monitoring Board (DSMB), composed of a minimum of 3 members 
including a statistician and a physician all of whom must be independent of the 
investigator and sponsors, will be set up prior to study initiation. The DSMB will monitor 
the study in order to ensure that harm is minimized and benefits maximized for the 
study subjects. All safety and available outcome data will be presented to the DSMB 
for review. The data and intervals will be agreed prior to the study initiation and 
documented in the DSMB Charter. 
 
12. Quality Assurance and Quality Control Procedures 
12.1. Investigator’s file 
The investigator must maintain adequate and accurate records to enable the conduct 
of the study to be fully documented and the study data to be subsequently verified. 
These documents include Investigator’s Site File, subject clinical source documents 
and screening / enrolment logs. The Investigator’s Site File will contain the 
protocol/protocol amendments, CRF/SAE and query forms, IEC and regulatory 
approval with correspondence, sample informed consent, drug accountability records, 
staff 
curriculum 
vitae 
and 
authorization 
forms 
and 
other 
appropriate 
documents/correspondence etc.  
12.2. Case report forms (CRFs) 
Data will be collected by laboratory technicians, medical doctors, clinical officers and 
nurses authorized by the investigator. It will be supervised by the Investigator and 
signed by the investigator or by an authorised staff member. Study-specific information 
will be entered into the Case Report Form (CRF) and SAE form (if applicable). Data 

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that are derived should be consistent with the source documents or the discrepancies 
should be explained. All CRF data should be anonymised, i.e. identified by study 
patient number only. 
The investigator at each trial site should ensure the accuracy, completeness, legibility, 
and timelines of all data reported to the sponsor in the CRFs/SAE forms and any other 
additional information that is required. The investigator is responsible for keeping all 
consent forms, screening forms, CRF/SAE form and the completed subject 
identification code list in a secure location. 
12.3.  Source documents 
The verification of the CRF/SAE form data must be by direct inspection of source 
documents. Source documents include subject hospital/clinic records, physician's and 
nurse's notes, appointment book, original laboratory reports, ECG, X-ray, pathology 
and special assessment reports, signed informed consent forms, consultant letters, 
and subject screening and enrolment logs. 
The investigator must maintain source documents such as laboratory and consultation 
reports, history and physical examination reports, etc., for possible review and/or audit 
by DNDi and/or Regulatory Authorities. The Investigator / designee will record the date 
of each subject’s visit in patient file together with a summary of their status and 
progress in the study. 
12.4.  Record Retention 
The investigator must keep all study documents on file for at least 25 years after 
completion or discontinuation of the study. After that period of time the documents may 
be destroyed with prior permission from DNDi, subject to local regulations. 
Should the investigator wish to assign the study records to another party or move them 
to another location, DNDi must be notified in advance. 
12.5.  Monitoring, audits and inspections 
Monitoring visits to the trial site will be made periodically by DNDi representatives or 
designated clinical monitors to ensure that GCPs and all aspects of the protocol are 
followed.  Source documents and SAE forms/Data clarification forms (DCFs; sent to 
DNDi pharmacovigilance) will be reviewed for verification of consistency with data on 
CRFs. The investigator will ensure direct access to source documents by DNDi or 
designated representatives. It is important that the investigators and their relevant 
personnel are available during the monitoring visits. 
The investigators will permit representatives of DNDi and/or designated clinical 
monitors to inspect all CRFs/SAE forms/DCFs, medical records, laboratory work 
sheets and to assess the status of drug storage, dispensing and retrieval at any time 
during the study. The corresponding source documents for each subject will be made 
available provided that subject confidentiality is maintained in accord with local 
regulations. The inspections are for the purpose of verifying the adherence to the 
protocol and to ensure the study is conducted according to GCP. It is important that 
the investigators and other trial site staff are available at these visits. 

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The monitoring visits provide DNDi with the opportunity to evaluate the progress of the 
study, verify the accuracy and completeness of CRFs/SAE forms/DCFs, resolve any 
inconsistencies in the study records, as well as to ensure that all protocol requirements, 
applicable regulations, and investigator's obligations are being fulfilled. Four visit types 
are planned: pre-study, study start, during the study, and study end. Visits may also 
be performed by regulatory authorities. 
It will be the clinical monitor’s responsibility to inspect the CRFs/SAE forms/DCFs at 
regular intervals throughout the study, to verify the adherence to the protocol and the 
completeness, consistency and accuracy of the data being entered on them. The 
investigator agrees to cooperate with the clinical monitor to ensure that any problems 
detected in the course of these monitoring visits are resolved. 
12.6.  Audits and inspections 
The trial site may also be subject to quality assurance audits by DNDi or designated 
representatives and/or to inspection by regulatory authorities or Independent Ethics 
Committees (IEC). 
It is important that the investigators and their relevant personnel are available for 
possible audits or inspections. 
12.7.  Data Management  
In order to ensure data quality, a uniform hard copy CRF will be designed for use at all 
the sites. After the CRFs have been completed by the site investigators and monitored 
by the clinical monitor, they will be collected and sent on an ongoing basis to DNDi 
data centre, based in Nairobi for data entry (double independent data entry), cleaning 
and analysis.  
CRFs and source documents will be monitored by the clinical monitor. Discrepancies 
noted either by the monitor or data centre will be queried to the site PI. The trial data 
will be stored in a computer database maintaining confidentiality in accordance with 
national data legislation and Good Clinical Practice. 
12.8.  Confidentiality of trial documents and subjects records 
The investigator must assure that subjects’ anonymity will be maintained and that their 
identities are protected from unauthorized parties. On CRFs or other documents 
submitted to the sponsor, subjects should not be identified by their names, but 
exclusively by an identification code. The investigator should keep a subject enrolment 
list showing codes, names, and addresses. The investigator should maintain 
documents for submission to sponsor authorized representative, and subject’s signed 
written consent forms, in strict confidence.  
13. Protocol Amendments 
The Principal investigator will ensure that the study protocol is strictly adhered to 
throughout, and that all data are collected and recorded correctly on the CRF. The 
Principal investigator may contact the medical coordinator for a protocol waiver for 
minor deviations from the protocol, e.g. patient unable to attend during visit window. 

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All protocol modifications must be documented in writing. Any protocol amendment 
must be approved and signed by the sponsor and the Principal investigator and is to 
be submitted to the appropriate IEC for information and approval in accordance with 
local requirements, and to regulatory agencies if required. Approval by IEC (and 
Regulatory Authority, if applicable) must be awaited before any changes can be 
implemented, except for changes necessary to eliminate an immediate hazard to trial 
subjects, or when the change involves only logistical or administrative aspects of the 
trial (e.g. change in clinical monitor(s), change of telephone number(s)). 
The protocol amendment can be initiated by either sponsor or by any Principal 
investigator.  
The investigator will provide in writing the reasons for the proposed amendment and 
will discuss with the medical coordinator and sponsor.  
14. Early Termination of the Study 
Both the sponsor and the investigator reserve the right to terminate the study at any 
time prior to inclusion of the intended number of subjects, but they intend to exercise 
this right only for valid scientific or administrative reasons. Should this be necessary, 
both parties will arrange the procedures on an individual study basis after review and 
consultation. In terminating the study, the sponsor and the investigator will assure that 
adequate consideration is given to the protection of the subject’s interest. 
Reasons for early termination by the sponsor(s) may include but not be limited to:  
• Too low enrolment rate. 
• Protocol violations. 
• Inaccurate or incomplete data. 
• Unsafe or unethical practices. 
• Questionable safety of the test article. 
• Suspected lack of efficacy of the test article. 
• Following the recommendation of the DSMB or IEC 
• Administrative decision. 
Reasons for early termination by the investigator may be: 
• Insufficient time or resource to conduct the study 
• Lack of eligible patients 
In the event that a study is early terminated either by the sponsor or by the 
investigator, the investigator has to: 
• Complete all CRFs to the greater extent possible 

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• Return all test articles, CRF, and related study materials to the sponsor who 
provided them 
• Answer all questions of the sponsors or their representatives related to data of 
subjects enrolled at the site prior to study termination 
• Ensure that subjects enrolled in the study who had not yet reached a follow up time 
point are followed up with the necessary medical care. 
• Provide in writing the reasons for his decision to the national health authority and 
the sponsor. 
15. Ethics 
The experimental protocol for this study has been designed in accordance with the 
general ethical principles outlined in the Declaration of Helsinki and ICH guidelines for 
Good Clinical Practice (International Committee for Harmonization). DNDi assures that 
it will comply with all applicable state, local and foreign laws for protecting the rights 
and welfare of human subjects.  This protocol and any protocol amendments will be 
reviewed / approved by an IEC before its implementation.  
It is the responsibility of the Global or National Coordinating Investigator/Investigator 
to apply for review to the IEC of the country where the study takes place regarding 
local rules and regulations. Written approval from all involved IECs must be obtained 
before implementation of any protocol-specified intervention /investigation provided to 
the subject (such as subject information sheets or descriptions of the study). 
Any modifications made to the protocol after receipt of the IEC approval must also be 
submitted by the investigator in writing to the IEC in accordance with local procedures 
and regulatory requirements. 
15.1.  Informed consent process 
Inclusion in the study will occur only if the subject (for adults) or the parent/guardian 
(for children) gives written informed consent. It is the responsibility of the investigator / 
designee to obtain written informed consent from each individual participating in this 
study, after adequate presentation of aims, methods, anticipated benefits, and 
potential hazards of the study. The written informed consent document will be 
translated into the local language or a language understood by the subject or the 
parent/guardian. If needed, the person will be given time to discuss the information 
received with members of the community or family before deciding to consent. The 
subject or parent/guardian will be asked to provide written and signed consent. 
If the subject is illiterate or unable to write, a literate witness must sign (this person 
should have no connection to the research team and the sponsor, and, if possible, 
should be selected by the participant). The investigator should also obtain the assent 
of children (if appropriate and according to country regulatory requirements), but their 
assent must be completed by the permission of a parent or guardian.  
Patients who agree to participate to the intensive PK and PD sampling (intensive 
cohort) will be asked to sign a separate informed consent document describing the 

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additional procedures to be performed.  
In addition, depending on local country regulations, a separate consent form for sample 
storage will also be used as applicable. 
If new safety information results in significant changes in the risk/benefit assessment, 
the consent form should be reviewed and updated if necessary. All subjects (including 
those already being treated) should be informed of the new information, given a copy 
of the revised form and give their consent to continue in the study.  
15.2.  Ethical aspects of subject inclusion and study procedures 
Potential study subjects will be selected from the routine VL diagnosis and treatment 
activities at the study sites. Children are a majority of patients affected by VL in Eastern 
Africa. Following the routine procedures for diagnosis of Kala-azar, patients with a 
confirmed VL diagnosis, aged 4-50y will be invited to participate in the study. The limit 
of 50 years of age for inclusion is due to higher mortality rate and lower efficacy 
observed in patients >50 years when treated with SSG-PM in a recent 
pharmacovigilance program in Eastern Africa (Kimutai et al., 2017). This study showed 
a higher risk of death for patients aged ≥50 years (9.1%) versus the overall population 
(0.9%). The end of treatment effectiveness for this population was significantly lower 
(81.8%) as compared to the overall EOT effectiveness (95.1%). 
The ICF process will be done with the subjects or parent/guardian of subjects in case 
of children. 
Inclusion/exclusion criteria will be systematically assessed during screening phase and 
only those meeting the study criteria will be eligible for enrolment. 
Parasitological confirmation of VL requires an invasive procedure, as tissue needs to 
be collected either from bone marrow or spleen. Tissue aspiration will only be 
performed by authorized medical personnel following the standard operation 
procedure. Patients with platelet count < 40,000/mm3, Hb < 5g/dL or with signs of 
bleeding should be diagnosed by bone marrow rather than spleen aspirate. 
HIV co-infected patients are excluded from the study, as they present lower efficacy 
rates, higher toxicity and higher lethality as compared to the non-HIV patients. 
Therefore, these patients require different case management and care.   
Female patients of child bearing age who do not accept to have a pregnancy test done 
at screening and/or who do not agree to use contraception from treatment period until 
5 months after the end of treatment will be excluded from the trial. Depending on 
countries and communities, it is very sensitive to introduce family planning to the young 
or unmarried female population who is not so familiar with contraception and is often 
illiterate. Community leaders’ opinion will be requested to guide the trial staff in the 
best approach to open the discussion on this topic, taking into consideration the local 
cultural context from the different communities. 
Study procedures are designed to ensure accurate assessment (i.e. clinical, PK 
sampling, laboratory assessments) while ensuring close follow up for safety. Patients 
in test arms 1 and 2 will be treated as in-patients up to day 14 to closely monitor the 
treatment safety and response to treatment. Patients in arm 2 used to continue on an 
out-patient basis until completion of the 28 days treatment. Since MF safety profile has 

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been studied in the past, no major safety concerns were expected during this period. 
In case of any safety concern, patients were instructed to either contact the investigator 
on phone or to come to the hospital for an unscheduled visit. Both patients in arm 1 
and 2 are assessed at the end of treatment visit on day 28. 
Furthermore, in order to record long-term clinical outcome, patients will be requested 
to return for the 6 months follow-up visit.    
Biological samples will be drawn only for the purpose of clinical assessments, 
pharmacokinetic (PK) and pharmacodynamics (PD) analysis. The volume of blood 
collected will be minimised as much as possible, and only one venepuncture will be 
performed per planned scheduled visits.  
Blood will be collected for PK analysis only in a subset of patients. Provided the PK 
nonlinearities observed in pediatric patients receiving the allometric MF dosing 
(LEAP0714), the inclusion of children ≤12 years old in the PK sub-study in addition to 
adolescents/adults >12 to ≤50 years is important to better study MF exposure in this 
population. In order to avoid multiple blood sampling in young children, only children 
from 7 years of age and weighing more than 20 kg will be included in the PK sampling 
schedule. 
A mixed sampling approach will be performed in order to reduce as much as possible 
the number of total samples and volume of blood to be collected (see section 8.4).  
It is expected that the maximum total volume of blood to be collected in the intensive 
PK/PD cohort (subset of patients, 40 in arm 1 and probably less than 40 in arm 2 that 
is now discontinued) will be approximately 58mL over the 6 months participation in the 
trial, assuming:  
6mL will be collected at study visits: screening, D7 and D28 
12mL will be collected at D1  
4mL will be collected at D3, D56 and D210 
14mL will be collected at D14 
2mL will be collected at D21 (in arm 2 only, not applicable in this protocol version). 
Patients not included in the intensive PK/PD cohort will have a total of 32 ml of whole 
blood collected over the entire study, for blood safety assessments, MF PK and for 
qPCR. 
There may be slight variations in the blood volume due to tubes available in the market. 
In any case, the maximum volume of blood to be collected in a study visit will be 14mL. 
Biological samples may be kept after the completion of the study in order to evaluate 
how other biomarkers or immunological markers respond following VL treatment. 
15.3.  Ethical aspects of study treatments 
MF is a drug marketed for Visceral Leishmaniasis indication and it has been 

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extensively used as primary treatment for VL in Asia, with very good safety profile. PM 
has been well studied in Eastern Africa during the development of the combination 
SSG-PM, which is now the current WHO-recommended treatment for VL. Safety 
profiles of these drug are well-known and potential side effects will be monitored 
throughout study duration to guarantee patient’s safety. 
All screened but ineligible VL patients will be treated with current standard treatment 
according to national guidelines. 
15.4.  HIV status and VCT 
All patients will be offered counselling and screening for HIV (voluntary counselling and 
testing programme (VCT). This may either be done at the same time as consent is 
obtained for inclusion in the trial or at a later date according to hospital practice.  
Patients who are found to be HIV positive will not be eligible to participate in this trial 
but will receive appropriate treatment, according to national treatment guidelines. 
Additionally, they will be referred onwards for treatment, surveillance and follow up 
according to the national protocol for HIV positive patients.  
15.5.  Patient costs 
Patients will be reimbursed for travel to and from the study site but will not receive any 
payment for trial participation. Any medication that is required during the trial period 
will be provided free of charge to the patient. Food during the in-patient treatment 
phase will also be provided free of charge to the patient. This is seen as an essential 
part of the patient care plan bearing in mind the high prevalence of malnutrition and 
the poverty of these patients. 
16. Insurance and Liability 
DNDi is insured to indemnify the investigator against any claim for damages brought 
by a research subject who suffers from a research related injury during the 
performance of the trial according to the protocol. The insurance in each country will 
also cover for the costs of medical care required as a result of the subject participation 
in the trial (ex. AE related to study drug or study procedure). 
17. Reporting and publication 
All clinical trials will be registered with a recognised clinical trial registry such as 
www.clinicaltrials.gov. Once the trial is completed and published, open access policy 
for data sharing will be followed. 
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