Clinical trial NCT03315039
Conditions: Leukemia, Myeloid, Acute
Sponsor: Moleculin Biotech, Inc.
Start date: 2018-03-28; Completion date: 2020-06-20
Enrollment: 7 participants; Allocation: NA; Intervention model: SEQUENTIAL

Brief summary:
This is a multi-center, open-label, dose escalation study that will determine the maximum tolerated dose (MTD) and recommended Phase 2 dose (RP2D) of liposomal annamycin as a single agent for the treatment of subjects with AML that is refractory to or relapsed after standard induction therapy

Detailed description:

Arms: arm 1: 2-hour intravenous infusion liposomal annamycin daily for 3 consecutive days followed by 18 days off study drug (i.e., one treatment cycle = 21 days).
Interventions: intervention 1: Liposomal Annamycin
Intervention descriptions: intervention 1: 2-hour intravenous infusion liposomal annamycin daily for 3 consecutive days followed by 18 days off study drug (i.e., one treatment cycle = 21 days).

Protocol documents:
Prot_SAP_001.pdf: 
Protocol No. MB-104 
Version 5.0 March 13, 2019 
Page 1 
CONFIDENTIAL 
TITLE: 
Phase 1/2 Study of Liposomal Annamycin for the Treatment of 
Subjects with Acute Myeloid Leukemia (AML) that is Refractory 
to or Relapsed after Standard Induction Therapy  
PROTOCOL NO. 
MB-104 
IND Number 
134 860 
EudraCT number: 
2017-002337-27 
INVESTIGATIONAL 
DRUG: 
Liposomal Annamycin 
SPONSOR: 
Moleculin Biotech, Inc. 
5300 Memorial Drive  
Suite 950 
Houston, TX, 77007 
Telephone: 862-273-1432 
Email: dpicker@moleculin.com 
MONITOR: 
Richard S. Ungerleider, MD 
Senior Vice President, Clinical Research Affairs 
Theradex  
4365 Route 1, South Suite 101 
Princeton, New Jersey 08540 
Telephone: 609-799-7580 extension 3067 
Mobile: 609-356-4072 
Facsimile: 609-799-4148 
Email: rungerleider@theradex.com 
DATE OF VERSION: 
March 13, 2019 
VERSION: 
5.0 


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CONFIDENTIAL 
TABLE OF CONTENTS 
 
PROTOCOL SIGNATURE SHEET ...............................................................................................5 
INVESTIGATOR SIGNATURE SHEET .......................................................................................6 
LIST OF ABBREVIATIONS ..........................................................................................................7 
SUMMARY OF CHANGES ...........................................................................................................9 
PROTOCOL SYNOPSIS ..............................................................................................................14 
1.1  Acute Myeloid Leukemia .................................................................................................20 
1.  BACKGROUND ......................................................................................................................20 
1.2  Liposomal Annamycin ......................................................................................................20 
1.3  Nonclinical Studies ...........................................................................................................21 
1.4  Clinical Studies .................................................................................................................22 
1.5  Rationale for the Current Study ........................................................................................23 
1.6  Consideration of Cardiotoxicity in the Current Study ......................................................23 
2.  STUDY OBJECTIVES .............................................................................................................25 
3.  STUDY DESIGN......................................................................................................................25 
3.1  Design Summary ...............................................................................................................25 
3.2  Dosing ...............................................................................................................................26 
3.3  Safety Considerations .......................................................................................................27 
3.3.1  Cardiotoxicity ..........................................................................................................28 
3.3.2  Myalgia – Adverse Event Workup ..........................................................................28 
3.4  Criteria for Dose-Limiting Toxicity..................................................................................28 
3.5  Evaluation of Efficacy ......................................................................................................29 
3.5.1  Bone Marrow Evaluation .........................................................................................29 
3.6  Premedications and Prophylactic Treatments ...................................................................29 
3.6.1  Tumor Lysis Syndrome Prophylaxis .......................................................................29 
3.6.2  Allergic Reactions ....................................................................................................29 
3.6.3  Oral Prophylaxis for Mucositis ................................................................................30 
4.  ELIGIBILITY CRITERIA........................................................................................................30 
4.1  Inclusion Criteria ..............................................................................................................30 
4.2  Exclusion Criteria .............................................................................................................32 
4.3  Subject Withdrawal ...........................................................................................................32 
4.3.1  Withdrawal Procedures ............................................................................................33 
4.3.2  Subject Replacement ................................................................................................33 
5.  STUDY TREATMENT ............................................................................................................34 
5.1  Liposomal Annamycin (Study Drug)................................................................................34 


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5.1.1  Description of Liposomal Annamycin .....................................................................34 
5.1.2  Preparation of Liposomal Annamycin .....................................................................34 
5.1.3  Liposomal Annamycin Administration....................................................................34 
5.1.4  Storage .....................................................................................................................34 
5.2  Concomitant Medications .................................................................................................35 
5.3  Measuring Subject Compliance ........................................................................................35 
5.4  Drug Accountability..........................................................................................................35 
6.  STUDY PROCEDURES ..........................................................................................................36 
6.1  Screening...........................................................................................................................36 
6.2  Baseline (Day 1) ...............................................................................................................37 
6.3  On-Study Evaluations .......................................................................................................38 
6.3.1  Study Drug Administration ......................................................................................38 
6.3.2  Induction Cycle ........................................................................................................39 
6.4  End of Study Evaluations ..................................................................................................40 
6.5  Follow-up Evaluations ......................................................................................................40 
6.6  Survival Follow-up ...........................................................................................................40 
7.  ASSESSMENT OF EFFICACY...............................................................................................41 
7.1  Efficacy Parameters ..........................................................................................................41 
7.1.1  Response Criteria .....................................................................................................41 
7.2  Methods for Measuring Efficacy ......................................................................................42 
8.  ASSESSMENT OF SAFETY ...................................................................................................42 
8.1  Safety Parameters..............................................................................................................42 
8.2  Methods for Measuring Safety ..........................................................................................42 
8.3  Procedures for Recording Adverse Events .......................................................................43 
8.3.1  Routine Reporting of Adverse Events .....................................................................43 
8.3.2  Reporting of Serious Adverse Drug Events, Including Deaths ...............................44 
8.3.3  Reporting of Suspected Unexpected Serious Adverse Reactions ............................45 
8.3.4  Severity of Adverse Drug Events ............................................................................45 
8.3.5  Relationship of Adverse Events to the Study Drug .................................................45 
8.4  Withdrawal From Study Treatment Due to Adverse Events ............................................46 
8.5 Establishment of a Safety Review Committee/Consultant ................................................46 
8.6 Monitoring of Cardiovascular Toxicity .............................................................................46 
9.  PHARMACOKINETIC ASSESSMENT .................................................................................46 
10.  STATISTICS ..........................................................................................................................47 
11.  ACCESS TO SOURCE DOCUMENTS ................................................................................47 
12.  QUALITY CONTROL AND QUALITY ASSURANCE ......................................................48 
13.  ETHICS...................................................................................................................................48 
13.1  Declaration of Helsinki ...................................................................................................48 
13.2  Institutional Review Board .............................................................................................48 


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CONFIDENTIAL 
13.3  Informed Consent............................................................................................................49 
14.  DATA HANDLING AND RECORDING .............................................................................49 
15.  PUBLICATION POLICY ......................................................................................................49 
16.  PROTOCOL AMENDMENTS AND MODIFICATIONS ....................................................49 
17.  REFERENCES .......................................................................................................................51 
APPENDIX A: SCHEDULE OF EVENTS ..................................................................................54 
APPENDIX B: EASTERN COOPERATIVE ONCOLOGY GROUP PERFORMANCE 
STATUS SCALE ............................................................................................................57 
APPENDIX C: COMMONLY USED CHEMOTHERAPY REGIMENS AND THEIR 
ANTHRACYCLINE CONTENT ...................................................................................58 
APPENDIX D: DEFINITION OF HY’S LAW.............................................................................59 
APPENDIX E: CALCULATIONS OF ANTHRACYCLINE EXPOSURE .................................60 
 
 
 


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CONFIDENTIAL 
PROTOCOL SIGNATURE SHEET 
 
The undersigned have reviewed the format and content of this protocol and have approved 
Protocol No. MB-104 for issuance. 
 
 
 
March 13, 2019 
Sponsor Signature 
 
Date 
 
 
 


Protocol No. MB-104 
Version 5.0 March 13, 2019 
Page 6 
 
 
CONFIDENTIAL 
INVESTIGATOR SIGNATURE SHEET 
 
I have read the attached protocol and agree that it contains all the necessary details for 
performing the study. 
 
I will provide copies of the protocol and of the preclinical and clinical information on the test 
article, which was furnished to me by the Sponsor, to all members of the study team responsible 
to me who participate in the study.  I will discuss this material with them to assure that they are 
fully informed regarding the test article and the conduct of the study. 
 
Once the protocol has been approved by the Institutional Review Board (IRB)/Independent 
Ethics Committee (IEC), I will not modify this protocol without obtaining the prior approval of 
the Sponsor and of the IRB/IEC.  I will submit the protocol modifications and/or any informed 
consent modifications to the Sponsor and the IRB/IEC, and approval will be obtained before any 
modifications are implemented. 
 
I understand the protocol and will work according to the US FDA Code of Federal Regulations, 
the principles of Good Clinical Practice (current International Conference of Harmonisation 
[ICH] guidelines), European Clinical Trial Directive 2001/20/EC,  European Union GCP 
Directive 2005/28/EC and the Declaration of Helsinki (1964) including all amendments up to 
and including the Fortaleza, Brazil, revision (2013). 
 
 
 
 
 
 
 
Investigator's Signature 
 
Date 
 
 


Protocol No. MB-104 
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Page 7 
CONFIDENTIAL 
LIST OF ABBREVIATIONS 
AE 
adverse event 
ALL 
acute lymphocytic leukemia 
AML 
acute myeloid leukemia 
BM 
bone marrow 
BUN 
blood urea nitrogen 
CABG 
coronary artery bypass graft 
CAD 
coronary artery disease,  
CBC 
complete blood count 
CNS 
central nervous system 
CR 
complete response 
CRi 
CR with incomplete blood count recovery meets all criteria for CR except for 
neutropenia (<1,000; CRi) but must include transfusion independence 
CRp 
CR with incomplete blood count recovery meets all criteria for CR except for 
thrombocytopenia (<100,000; CRp) but must include transfusion independence 
eCRF/EDC 
Electronic Case Report Form/Electronic Data Capture 
CTCAE 
Common Terminology Criteria for Adverse Events 
CV 
cardiovascular 
CYP 
Cytochrome P450 family of enzymes 
DLT 
dose-limiting toxicity 
DMSO  
dimethyl sulfoxide 
ECG 
electrocardiogram 
ECHO 
echocardiogram 
ECOG 
Eastern Cooperative Oncology Group 
EFS 
event-free survival 
EOS 
end of study 
EudraCT 
European Union Drug Regulating Authorities Clinical Trials 
FDA 
Food and Drug Administration 
GCP 
Good Clinical Practice 
HSCT 
hematopoietic stem cell transplantation 
IEC 
Independent Ethics Committee 
ICH 
International Conference of Harmonisation 
IRB 
Institutional Review Board 
IV 
intravenous(ly) 
LDH 
lactate dehydrogenase 
LV 
left ventricular  
LVEF 
left ventricular ejection fraction 
LVH 
left ventricular hyperthropy 
MDR 
multidrug-resistant 
MI 
myocardial infarction 
MTD 
maximum-tolerated dose 
MUGA 
multiple-gated acquisition 
NCI 
National Cancer Institute 
OS 
overall survival 
PCI 
percutaneous coronary intervention 


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CONFIDENTIAL 
PK 
 
pharmacokinetic 
PR 
 
partial response 
RP2D  
recommended Phase 2 dose 
SAE 
 
serious adverse event 
SGOT  
serum glutamic-oxaloacetic transaminase 
SGPT  
serum glutamic-pyruvic transaminase 
SRC 
 
Safety Review Committee/Consultant 
ULN  
upper limit of normal 
VHD  
valvular heart disease 
 
 


Protocol No. MB-104 
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CONFIDENTIAL 
Summary of Changes 
Page 
Section 
Description of Text 
All 
Headers 
Versioning updated 
Global 
Global 
Minor editorial changes were made throughout the document, including 
correction of grammatical and typographical errors, use of numeric forms, and 
addition of abbreviations. 
Global 
Global 
NCI CTCAE v4.03 5 
14, 16, 
17, and 
19 
Protocol Synopsis 
Methodology 
Inclusion Criteria 
Exclusion Criteria 
Statistical 
Analyses 
For each dose level, groups of 3 subjects will receive a treatment cycle of 
liposomal annamycin daily as a 2-hour intravenous (IV) infusion for 
3 consecutive days followed by 18 days off study drug (i.e., the induction 
treatment cycle = 21 days).  If no subject experiences a dose-limiting toxicity 
(DLT), based on National Cancer Institute (NCI) Common Terminology 
Criteria for Adverse Events (CTCAE) v4.03 5, which constitutes a study drug-
related irreversible or not medically manageable Grade 3 or higher toxicity, 
further defined in Section 3.4, during the induction treatment through Day 42 
Day 28, the subsequent group of 3 subjects will receive the next higher 
liposomal annamycin dose. 
8 
Prior anthracycline cumulative dose (see Table 2 and Appendix E) below 
550 <551 mg/m2 or the daunorubicin equivalent, which is the 
recommended noncardiotoxic level.  
b.
Table 2 shows the maximal doses for other anthracyclines and the 
conversion factor for a daunorubicin equivalent is provided in
Appendix E Potential cardioprotective measures for anthracycline 
drugs and analogues by limitation of cumulative doses are 
demonstrated by the following conversion factors:  
Daunorubicin = 1 
Doxorubicin = 2.2 
Epirubicin = 1.1 
Mitoxantrone = 5 
Idarubicin = 5.3 
c.
Appendix E shows examples of how the maximal allowance based on
daunorubicin equivalence is calculated The maximal allowance will 
be calculated as a daunorubicin equivalent added to the 
anticipated exposure to liposomal annamycin with the maximal 
exposure of anthracyclines capped at 550.9 mg/m2 (Appendix E) 
d.
Exposure to annamycin projected for subjects enrolled in a given
cohort in this study will be assessed on the basis of prior total
exposure added to the anticipated designated cohort dose level (up to
120 mg/m2, which, after 3 doses, representing a cumulative exposure
of <550 551-mg/m2 daunorubicin equivalents).
5 
Prior anthracycline cumulative dose more than 50% above recommended 
noncardiotoxic levels, Left ventricular ejection fraction (LVEF) <50%, 
valvular heart disease, or severe hypertension (see Table 1) 
Study enrollment in the expansion phase portion of the study willbe halted if >2 
of 6 or >3 of 10 subjects experience a DLT at the recommended Phase 2 dose.  
The Medical Monitor and the SRC will continually review all data and decide 
if the study should continue and if any changes to the protocol before restarting 
enrollment must be made. 
24 
1.6 Consideration 
A value of 550  <551 mg/m2 will be used as the correlative value for 


Protocol No. MB-104 
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CONFIDENTIAL 
of Cardiotoxicity 
in the Current 
Study  
liposome-encapsulated annamycin for purposes of assessment of maximal 
exposure, with a conversion factor of 1.   
25-29 
3.1  Design 
Summary 
3.2  Dosing 
3.3.1 
Cardiotoxicity 
3.4  Criteria for 
Dose-Limiting 
Toxicity  
For each dose level, groups of 3 subjects will receive a treatment cycle of 
liposomal annamycin daily as a 2-hour intravenous (IV) infusion for 
3 consecutive days followed by 18 days off study drug (i.e., the induction 
treatment cycle = 21 days). If no subject experiences a dose-limiting toxicity 
(DLT), based on National Cancer Institute (NCI) Common Terminology 
Criteria for Adverse Events (CTCAE) v4.03 5, which constitutes a study drug-
related irreversible or not medically manageable Grade 3 or higher toxicity, 
further defined in Section 3.4, during the induction treatment through Day 42 
Day 28, the subsequent group of 3 subjects will receive the next higher 
liposomal annamycin dose. 
Study enrollment in the expansion phase portion of the study willbe halted if >2 
of 6 or >3 of 10 subjects experience a DLT at the recommended Phase 2 dose.  
The Medical Monitor and the SRC will continually review all data and decide 
if the study should continue and if any changes to the protocol before restarting 
enrollment must be made. 
The initial group of 3 subjects will be treated with a 2-hour intravenous infusion 
of 100 mg/m2/day (Dose Level 1) liposomal annamycin daily for 3 consecutive 
days followed by 18 days off study drug (i.e., 1 treatment cycle = 21 days).  
Provided that no subject experiences a DLT (defined in Section 3.4) during the 
induction treatment cycle through Day 42 Day 28, the ensuing group of 
3 subjects will receive 120 mg/m2/day (Dose Level 2) liposomal annamycin- 
Subjects will also have clinical and electrocardiographic examinations 
evaluated at the anticipated maximal plasma concentrations, and again weekly 
during the induction cycle of treatment with  multiple-gated acquisition 
(MUGA) or echocardiogram (ECHO [2-dimensional accepted, 3-dimensional 
preferred]) scans, or ECHO  strain evaluations performed every 3 weeks by 
using the same modality and instrument.  Any subject demonstrating any of the 
above manifestations of cardiotoxicity, or >10% decrease in ejection fraction 
based on scans, will be discontinued from the study.  Between the Day 18 and 
Day 28 visits or at the End of Treatment visit (if the subject is prematurely 
terminated), a final MUGA or ECHO scan, or an ECHO strain evaluation , 
using the same modality and instrument, will be conducted by using the same 
modality and instrument one week after the treatment cycle or after the last 
study drug administration, if the treatment period is prematurely terminated 
The DLT evaluation period is defined as the first day of treatment (Day 1) until 
Day 42 Day 28.   
The following hematologic toxicities that occur after the cycle of treatment that 
do not resolve by Day 42 will be considered a DLT: 
• 
•Peripheral absolute neutrophil count 
•
(ANC) <500/mm3 (Grade ≥4);
• 
•Non-transfusion-dependent platelet count <20,000/mm3 due to 
documented bone marrow 
•
aplasia/hypoplasia (vs. malignant infiltration or other cause).  Bone
marrow aplasia/hypoplasia is defined as overall marrow cellularity 
<20%; 
•
Platelet count <50,000/mm3 (Grade ≥3) that is associated with


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3.5.1  Bone 
Marrow 
Evaluation 
bleeding; 
Platelet count <25,000/mm3 (Grade 4) that requires platelet transfusion. 
Bone marrow aspirate (a biopsy if there are no spicules present) 14-21 15 to 35 
days after the start of therapy will be performed. to document hypoplasia.  If 
hypoplasia is not documented or indeterminate, a repeat biopsy should be 
obtained in another 7-14 days to clarify the persistence of leukemia.  If 
hypoplasia is present, then a repeat biopsy at the time of hematologic recovery 
based on peripheral blood counts should be obtained to document remission. 
31, 32, 
and 33 
4.1 Inclusion 
Ctiteria 
4.2 Exclusion 
Criteria 
4.3.2  Subject 
Replacement 
8 
Prior anthracycline cumulative dose (see Table 2 and Appendix E) below 
550 <551 mg/m2 or the daunorubicin equivalent, which is the 
recommended noncardiotoxic level.  
c.
Potential cardioprotective measures for anthracycline drugs and 
analogues by limitation of cumulative doses are demonstrated by 
the conversion factors in Table 2 shows the maximal doses for other 
anthracyclines and the conversion factor for a daunorubicin equivalent
is provided in Appendix E 
d.
Appendix E shows examples of how the maximal allowance based on
daunorubicin equivalence is calculated The maximal allowance will 
be calculated as a daunorubicin equivalent added to the 
anticipated exposure to liposomal annamycin with the maximal 
exposure of anthracyclines capped at 550.9 mg/m2 (Appendix E) 
e.
Exposure to annamycin projected for subjects enrolled in a given
cohort in this study will be assessed on the basis of prior total
exposure added to the anticipated designated cohort dose level (up to
120 mg/m2, which, after 3 doses, representing a cumulative exposure
of <550 551-mg/m2 daunorubicin equivalents).
5 
Prior anthracycline cumulative dose more than 50% above recommended 
noncardiotoxic levels, LVEF <50%, valvular heart disease, or severe 
hypertension (see Table 1) 
Subjects withdrawing in the cohort dose-escalation period for reasons other than 
toxicity who have not taken all 3 doses during the induction cycle and/or fail to 
complete restaging after completion of the treatment cycle (~21 days) such that 
safety or efficacy is not evaluable will be replaced. However, subjects 
experiencing a DLT in the dose-finding cycle will not be replaced. 
34 
5.1.2  Preparation 
of Liposomal 
Annamycin 
The study drug must be reconstituted prior to use.  The diluted product can be 
held for up to 4 hours 24 hours when stored between 25 to 37ºC 34 to 42ºC, 
after which time unused drug should not be administered to subjects. 
39-40 
6.3.2  Induction 
Cycle 
6.4  End of Study 
In addition to the weekly evaluations during the induction cycle, the following 
evaluations will be conducted at the end of the 3-week treatment cycle (between 
Days 14-21): 
1.
Physical examination including weight, vital signs, and oral exam
1.
MUGA or ECHO scan or ECHO strain evaluation between Days 18
and 28.  Cardiology consultation should be requested if any question
arises about cardiac function. The MUGA scan should also be
conducted at the Early Termination visit for subjects who are
withdrawn prematurely.
2.
Bone marrow aspirate and peripheral blood specimens (complete blood
count, differential, and platelet count; flow cytometry is optional)
between Days 15 and 35 to document response


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Evaluations 
6.5  Follow-up 
Evaluations 
6.6.  Survival 
Follow-up 
1.
Physical examination including weight, vital signs, and oral exam
2.
CBC with differential and platelet count, if subject had a hematological
toxicity which could be classified as a DLT (as defined in Section 3.4) 
Subjects will be followed every 3 months from study drug discontinuation to 
monitor resolution or stabilization of any treatment-related medical events and 
obtain survival data.  If possible, a MUGA scan, ECHO scan, or ECHO 
strain evaluation will be collected. 
42 
7.2  Methods for 
Measuring 
Efficacy 
Bone marrow aspirate/biopsy and peripheral blood specimens (complete blood 
count, differential, and platelet count) to document response will be collected at 
14-21 days 15 to 35 days after the start of therapy to document hypoplasia.  If 
hypoplasia is not documented or indeterminate, a repeat biopsy should be 
obtained in 7-14 days to clarify the persistence of leukemia.  If hypoplasia is 
present, then a repeat biopsy at time of hematologic recovery should be 
obtained to document remission.  
47 
10. Statistics
Study enrollment in the expansion phase portion of the study will be halted if 
>2 of 6 or >3 of 10 subjects experience a DLT at the recommended Phase 2 
dose.  The Medical Monitor and the SRC will continually review all data and 
decide if the study should continue and if any changes to the protocol before 
restarting enrollment must be made. 
54-56 
Appendix A: 
Schedule of 
Events 
Day 21 3 (At the End of the Induction Cycle) 
CBC, differential, platelet countc,d 
Insertion of columns for Day 15, Day 18, and Day 35 
Day 42 CBC, differential, platelet count: Xo 
Day 18 to 28 MUGA, ECHO, or ECHO strain: X 
Survival Follow-up: MUGA, ECHO, or ECHO strain: Xq 
Day 15 to 35 Bone marrow aspirate and peripheral bloodh for measuring 
disease: X 
g A final MUGA or ECHO scan or an ECHO strain ealuation will be 
conducted between the Day 18 and Day 28 visits or at the the End of 
Treatment visit if the subject is prematurely terminated. 
Footnotes d, e, f, g, h, i, j, k, l, m, n, and o were renumbered to e, f, h, i, j, k, l, 
m, n, o, p, and d, respectively. 
f h Bone marrow aspirate and peripheral blood (complete blood count, 
differential, and platelet count; flow cytometry is optional) within 15 days prior 
to the first dose of liposomal annamycin to document disease status.  Bone 
marrow aspirate/biopsy 14-21 15 to 35 days after the start of therapy will be 
performed. to document hypoplasia.  If hypoplasia is not documented or 
indeterminate, a repeat biopsy should be obtained in another 7-14 days to clarify 
the persistence of leukemia.  If hypoplasia is present, then a repeat biopsy at the 
time of hematologic recovery based on peripheral blood counts should be 
obtained to document remission. If cytogenetics were initially abnormal, 
include cytogenetics as part of the remission documentation. 
q  A MUGA scan, ECHO scan, or ECHO strain will be collected at the Q3-


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month survival timepoint if possible. 
59 
Appendix E 
Calculations of 
Anthracycline 
Exposure 
All daunorubicin equivalents are based on 550 <551 mg/m2 as being the 
maximal allowable dose with the maximal exposure of anthracyclines 
capped at 550.9 mg/m2. 
Example calculations: 
A subject with prior FLA(G)-IDA Induction (Idarubicin 24 mg/m2) for 1 
course: 
24 mg/m2 per course 
24 × 5.3 (conversion factor of daunorubicin  = 127. 2 mg/m2: daunorubicin 
equivalents 
With maximal exposure of anthracyclines based on daunorubicin equivalent 
capped at 550 mg/m2: 
550 mg/m2– 127.2 mg/m2 = 422.8 mg/m2: 
Therefore, additional liposomal annamycin of no more than 422.8 
422.8-mg/m2 exposure will be permitted (subject allowed to receive 
3 doses at 120 mg/m2) 
A subject with prior D(60)A 3+7 Induction (Daunorubicin 180 mg/m2) 
With maximal exposure of anthracyclines based on daunorubicin equivalent 
capped at 550 mg/m2: 
Daunorubicin 550 mg/m2– 180 mg/m2 = 370 mg/m2 
Therefore, additional liposomal anthramycin of no more than 370-mg/m2 
exposure is permitted (subject allowed to receive 3 doses at 120 mg/m2) 


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CONFIDENTIAL 
PROTOCOL SYNOPSIS 
 
Protocol No.: 
MB-104 
Protocol Title: 
Phase 1/2 Study of Liposomal Annamycin for the Treatment of Subjects with Acute 
Myeloid Leukemia (AML) that is Refractory to or Relapsed after Standard 
Induction Therapy   
Clinical Phase: 
Phase 1/2  
Investigator(s): 
TBD 
Study Center(s): 
Approximately 7  
Name of Sponsor/ 
Company: 
Moleculin Biotech, Inc. 
Objectives: 
The primary objective of this study is to evaluate the safety and identify the 
recommended Phase 2 dose (RP2D) of liposomal annamycin for the treatment of 
subjects with AML that is refractory to or relapsed after standard induction therapy.   
 
Secondary objectives: 
1. Pharmacokinetics of annamycin and its metabolite, annamycinol  
2. Preliminary assessment of the antileukemic activity of liposomal 
annamycin as second line therapy for subjects with refractory or relapsed 
AML based on established response criteria, including complete response 
(CR), partial response (PR), event-free survival (EFS), overall survival 
(OS; Kaplan-Meier), and time to and duration of remission/response  
Patient Population: 
Subjects diagnosed with AML that is refractory to or relapsed after standard 
induction therapy. 
Methodology: 
This is a multicenter, open-label, dose-escalation study that will determine the 
maximum-tolerated dose (MTD) and RP2D of liposomal annamycin as a single 
agent for the treatment of subjects with AML that is refractory to or relapsed after 
standard induction therapy.   
 
During the dose-escalation phase of the study, eligible subjects could have received 
any number of previous therapies for their relapsed AML as long as the prior 
anthracycline cumulative dose is below the recommended noncardiotoxic level).  
During the expansion phase of the study, after the MTD or RP2D has been 
established, study treatment will be second-line therapy, thus subjects could not 
receive prior therapy for their relapsed AML. 
 
Enrollment will occur in cohorts of 3 subjects in a conventional 3+3 escalating dose 
design, starting at a dose level of 100 mg/m2/day administered for 3 days.  Dose 
escalation will take place on the basis of safety assessments in this first cohort with 
the next dose of liposomal annamycin escalated to 120 mg/m2 in the absence of 
safety concerns.   
 
For each dose level, groups of 3 subjects will receive a treatment cycle of liposomal 
annamycin daily as a 2-hour intravenous infusion for 3 consecutive days followed 
by 18 days off study drug (i.e., the induction treatment cycle = 21 days).. If no 
subject experiences a dose-limiting toxicity (DLT), based on National Cancer 
Institute (NCI) Common Terminology Criteria for Adverse Events (CTCAE) v5, 
which constitutes a study drug-related irreversible or not medically manageable 
Grade 3 or higher toxicity during the induction treatment through Day 28, the 
subsequent group of 3 subjects will receive the next higher liposomal annamycin 
dose.  However, if 1 of the 3 initial subjects experiences a DLT, the cohort of 
subjects at that dose level will be expanded to 6 subjects.  If at least 2 of the 6 
subjects experience a DLT, this will be considered a nontolerated dose and then 3 
subjects will be treated at a lower dose.   


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CONFIDENTIAL 
The MTD is defined as the highest dose of liposomal annamycin at which fewer 
than 2 (of a cohort of up to 6) subjects experience a DLT.   
 
The RP2D is defined as the optimal dose to be determined by the Sponsor in 
conjunction with an independent Safety Review Committee/Consultant (SRC), 
consisting of at least 1 independent consultant, on the basis of review of available 
clinical and laboratory safety and efficacy data.  Up to 21 additional subjects will be 
enrolled at either the MTD or RP2D, to better define toxicity and evaluate efficacy 
at this dose.  
 
If a subject discontinues treatment for reasons other than study drug-related adverse 
events such that safety and efficacy of the drug cannot be fully evaluated during the 
induction cycle, a replacement subject may be enrolled; these circumstances will be 
reviewed on a case-by-case basis by the SRC in conjunction with the Sponsor.  
Number of Subjects: 
Three subjects minimum (6 maximum) for each dosing cohort.  Evaluation of a 
RP2D based on the aggregate assessment of data will be determined by the SRC in 
conjunction with the Sponsor, with consideration of additional dose exploration 
based on safety assessments.  Up to 21 additional subjects will be enrolled at the 
MTD or RP2D to better define toxicity and evaluate efficacy at this dose.  
Therefore up to 33 subjects will be enrolled in this study: up to 12 subjects in the 
dose-escalation phase and up to 21 additional subjects at the MTD or RP2D.  
Study Drug: 
Annamycin is a lipophilic anthracycline antibiotic that incorporates 4 structural 
modifications from doxorubicin: 2’-iodo, 3’-hydroxy, 4’-epi, and 4-demethoxy 
doxorubicin. 
 
Liposomal annamycin is supplied in 50-mL vials containing annamycin as a 
lyophilized powder.   
 
Subjects will be treated daily for 3 consecutive days with a 2-hour intravenous 
infusion of liposomal annamycin followed by 18 days off liposomal annamycin 
(i.e., 1 treatment cycle = 21 days).  The starting dose of liposomal annamycin (dose 
for subjects in Cohort 1) is 100 mg/m2/day.  The liposomal annamycin doses will be 
escalated up to 120 mg/m2/day in the next cohort in the absence of safety concerns.   


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Inclusion Criteria: 
Inclusion Criteria 
1. 
A pathologically confirmed diagnosis of AML by World Health 
Organization classification 
2. 
AML that is refractory to or relapsed after standard induction therapy  
3. 
Age ≥18 years at the time of signing informed consent 
4. 
No chemotherapy, radiation, or major surgery within 2 weeks prior to 
first dose of study drug and/or recovered from the toxic side effects of 
that therapy, unless treatment is indicated due to progressive disease 
5. 
No investigational therapy within 4 weeks of the first dose of study drug 
6. 
Eastern Cooperative Oncology Group (ECOG) performance status 0 to 2 
7. 
Adequate laboratory results including the following: 
a. 
Bilirubin ≤1.5 times the upper limit of normal unless due to Gilbert 
Syndrome 
b. Serum glutamic-oxaloacetic transaminase , serum glutamic-pyruvic 
transaminase, and alkaline phosphatase <3 times the upper limit of 
normal) unless due to organ involvement 
c. 
Adequate renal function (The Cockroft-Gault equation will be used 
to estimate creatinine clearance.  This equation is as follows:  
Creatinine clearance in milliliters per minute = [140 – age] × body 
weight [kg]/72 × plasma creatinine [mg/dL], multiplied by 0.85 for 
women.  By using this equation, adequate renal function will be 
deemed to be a creatinine clearance of greater than 60 mL/minute.) 
8. 
Prior anthracycline cumulative dose <551 mg/m2 or the daunorubicin 
equivalent, which is the recommended noncardiotoxic level.  This will be 
calculated as follows: 
a. 
Determination of prior exposure will be based on total milligrams 
per meter squared of prior anthracycline therapy). 
b. Potential cardioprotective measures for anthracycline drugs and 
analogues by limitation of cumulative doses are demonstrated by the 
following conversion factors:  
Daunorubicin = 1 
Doxorubicin = 2.2 
Epirubicin = 1.1 
Mitoxantrone = 5 
Idarubicin = 5.3 
c. 
The maximal allowance will be calculated as a daunorubicin 
equivalent added to the anticipated exposure to liposomal annamycin 
with the maximal exposure of anthracyclines capped at 550.9 mg/m2. 
d. Exposure to annamycin projected for subjects enrolled in a given 
cohort in this study will be assessed on the basis of prior total 
exposure added to the anticipated designated cohort dose level (up to 
120 mg/m2, which, after 3 doses, representing a cumulative exposure 
of <551-mg/m2 daunorubicin equivalents). 
9. 
Subject can understand and sign the informed consent document, can 
communicate with the Investigator, and can understand and comply with 
the requirements of the protocol. 
10. Women of childbearing potential must have a negative serum or urine 
pregnancy test. 
11. All men and women must agree to practice effective contraception during 
the entire study period and after discontinuing study drug, unless 
documentation of infertility exists.   
a. 
Sexually active, fertile women must use 2 effective forms of 
contraception (abstinence, intrauterine device, oral contraceptive, or 
double barrier device) from the time of informed consent and until at 
least 6 months after discontinuing study drug 


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b.
Sexually active men and their sexual partners must use effective
contraceptive methods from the time of subject informed consent and
until at least 3 months after discontinuing study drug
Exclusion Criteria: 
Exclusion Criteria 
1.
Subjects diagnosed with acute promyelocytic leukemia
2.
Concomitant therapy that includes other chemotherapy that is or may be
active against AML except for prophylaxis and/or treatment of
opportunistic or other infection with antibiotics, antifungals, and/or
antiviral agents
3.
Prior mediastinal radiotherapy
4.
Any condition that, in the opinion of the Investigator, places the subject at
unacceptable risk if he/she were to participate in the study.
5.
Left ventricular ejection fraction (LVEF) <50%, valvular heart disease, or
severe hypertension. Cardiac subjects with a New York Heart Association
classification of 3 or 4 will be excluded. (Cardiology consultation should
be requested if any question arises about cardiac function.) This also
includes subjects with baseline QT/QTc interval >480 msec, a history of
additional risk factors for torsade des pointes (e.g., heart failure,
hypokalemia, family history of Long QT Syndrome), and use of
concomitant medications that significantly prolong the QT/QTc interval.
6.
Clinically relevant serious comorbid medical conditions including, but
not limited to, active infection, recent (less than or equal to 6 months)
myocardial infarction, unstable angina, symptomatic congestive heart
failure, uncontrolled hypertension, uncontrolled cardiac arrhythmias,
chronic obstructive or chronic restrictive pulmonary disease, active
central nervous system disease uncontrolled by standard of care, known
positive status for human immunodeficiency virus and/or active hepatitis
B or C, cirrhosis, or psychiatric illness/social situations that would limit
compliance with study requirements
7.
Pregnant, lactating, or not using adequate contraception
8.
Known allergy to anthracyclines
9.
Any evidence of mucositis/stomatitis or previous history of severe
(≥Grade 3) mucositis from prior therapy
10. Required use of strong inhibitors and inducers of Cytochrome P450
family of enzymes (CYP) and transporters that cannot be held during
treatment days
Criteria for Evaluation: 
(Safety): 
1.
Clinical adverse events (AEs), routine hematological and biochemical
parameters, physical examination and vital signs, 12-lead
electrocardiograms (ECGs), ECOG performance status, pregnancy testing,
and monitoring of concomitant medications will be assessed according to
the Schedule of Events.
2.
All AEs and laboratory abnormalities will be graded for severity by using
the NCI CTCAE v5.  The relatedness of all AEs and laboratory
abnormalities and their seriousness will be determined by the Investigator
and adjudicated by the SRC in conjunction with the Sponsor.  The SRC
will consist of an external, independent (not participating in the clinical
trial) committee/consultant providing guidance with regard to safety
issues, including approval of dose escalation and designation of DLTs.
3.
AEs will be documented from the time of first dose of study drug until
39 days after the last dose administered on study.
4.
Strategies for detection of cardiotoxicity that include cardiac imaging
(echocardiography, nuclear imaging, cardiac magnetic resonance) and
biomarkers (troponin, natriuretic peptides) will be based on clinical
assessments at screening, as well as current relevant guidelines


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Criteria for Evaluation 
(Efficacy): 
All subjects evaluable for efficacy will be assessed for response to treatment by 
using the recommendations of the International Working Group for standardization 
of response criteria, treatment outcomes, and reporting for therapeutic trials.  The 
primary efficacy variable is leukemia response rate, evaluated by the Investigator at 
the end of the induction cycle and at the end of study, based on bone marrow 
aspirate and peripheral blood collected at the end of the induction cycle.  A bone 
marrow (BM) aspirate/biopsy should be repeated in 1 week if there is a question of 
residual leukemia in assessing efficacy based on an initial BM specimen.  
 
Primary Efficacy Endpoints: 
1. Morphologic leukemia-free state 
a. 
BM <5% blasts in an aspirate with spicules (a BM biopsy should be 
performed if spicules are absent) 
b. No blasts with Auer rods or persistence of extramedullary disease  
2. Complete remission: 
a. 
Morphologic leukemia-free state in which 
i. Subject is independent of transfusions 
ii. Absolute neutrophil count of >1,000  
iii. Platelets of ≥100,000 
3. Cytogenetic complete response – cytogenetics normal in those with 
previously abnormal cytogenetics 
4. CRi and CRp: CR with incomplete blood count recovery meets all criteria 
for CR except for either neutropenia (<1,000; CRi) or thrombocytopenia 
(<100,000; CRp) but must include transfusion independence 
 
Secondary Endpoints: 
 
1. EFS: Time from enrollment until disease progression or death from any 
cause 
2. OS: Time from enrollment until death from any cause 
3. PR:  All of the hematologic values for a CR, with normalization of blood 
counts, but with a decrease of at least 50% in the percentage of blasts to 
5% to 25% in the bone marrow aspirate  
4. Time to and duration of remission/response 
Pharmacokinetic 
Analyses: 
Pharmacokinetics of annamycin and its metabolite, annamycinol, will be measured 
after the first dose of liposomal annamycin on Day 1 and Day 3 of the induction 
cycle.   
 
Blood samples for pharmacokinetic analysis will be collected on these days at 
predose and at 0.25, 0.5, 1, 2, 4, 8, and 24 hours after the start of liposomal 
annamycin infusion for 3 subjects at each dose and 6 subjects at the RP2D. 
 
ECGs and documentation of concomitant medications potentially affecting CYP 
enzymes will be obtained to ensure that these are a consideration in the evaluation 
of the safety and pharmacokinetics of this drug.  
Statistical Analyses 
This is a multicenter, open-label, dose-escalation, MTD/RP2D safety study of 
liposomal annamycin given to patients with refractory or relapsed AML.  Therefore, 
only descriptive statistics will be utilized.  A maximum of 33 patients will be 
recruited for the study depending on the level at which toxicity is observed. 
 
Data from all subjects who receive 1 or more doses of liposomal annamycin will be 
incorporated into the final safety analysis.  Safety and tolerability will be assessed 
by adverse events, vital signs, physical examination, laboratory parameters, and an 
ECHO or multiple-gated acquisition (MUGA) scan or an ECHO strain evaluation, 
as well as cardiotoxicity biomarkers.  Adverse events will be listed by subject and 


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will be tabulated and summarized as the number and percentage of subjects who 
reported each adverse event.  Descriptive statistics will be generated as appropriate; 
otherwise, no formal statistical analyses are planned. 
 
The Medical Monitor and the SRC will continually review all data and decide if the 
study should continue and if any changes to the protocol before restarting 
enrollment must be made. 


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1.  BACKGROUND 
 
1.1  Acute Myeloid Leukemia 
 
Adult acute myeloid leukemia (AML) is the most common type of leukemia in adults, 
accounting for approximately 25% of leukemia diagnoses, with an estimated 19,950 new cases 
and an estimated 10,430 patients will die from AML this year in the US (NCI 2016; Deschler et 
al. 2006).  The 5-year survival rate in patients with AML is approximately 25%.  AML evolves 
rapidly and develops resistance to therapy because of its polyclonal and heterogeneous features 
(Ferrara et al. 2013).  Cytogenetic and molecular mutation testing may be important for risk 
assessment and prognosis, but they are not predictive.  So far, ascertaining a specific mutation 
does not predict what therapy is likely to work (Patel et al. 2012). 
 
AML patients initially respond well to the decades’ old and still standard 3 + 7 induction 
regimen if they are candidates for this intensive therapy.  Response and survival are associated 
with multiple factors, including patient age, performance status, antecedent hematologic 
conditions (de novo vs. secondary AML), and cytogenetic and molecular features.  Responses in 
the second line of therapy rely on these prognostic factors but also include the achievement of 
minimal residual disease status and the duration and type of response, e.g., complete response 
(CR) or CRi (complete response with incomplete bone marrow recovery) in the first line (Chen 
et al. 2015).  Patients in first relapse with a first CR duration of 12 months or longer are offered 
high-dose cytarabine-based regimens.  These include fludarabine with cytarabine and idarubicin, 
clofarabine with idarubicin and cytarabine, cladribine with idarubicin and cytarabine, twice daily 
fludarabine, cytarabine, or high-dose cytarabine-based investigational approaches.  For patients 
in first salvage with a first CR duration <12 months and all patients in second salvage or beyond, 
Phase 1 to 2 investigational approaches are reasonable (Kantarjian 2016). 
 
 
1.2  Liposomal Annamycin 
 
Doxorubicin is one of the most effective antitumor agents against hematological malignancies, 
including AML, and certain human solid tumors such as breast carcinoma and osteosarcoma.  Its 
use is limited by myelosuppression, chronic cardiotoxicity, and natural or acquired drug 
resistance.  Important advances in understanding the mechanisms of acquired resistance to 
doxorubicin and other structurally unrelated antitumor agents have occurred recently.  
Overexpression of a membrane glycoprotein, P-glycoprotein, which acts as a drug efflux pump, 
mediates acquired resistance to doxorubicin in some in vitro systems and in vivo animal tumor 
models (DeVita et al. 1993).  Overexpression of P-glycoprotein occurs in a significant number of 
patients when their tumors progress or relapse after treatment with doxorubicin.  No standard 
regimen exists for the treatment of patients with relapsed AML, particularly in patients with first 
remission duration of less than 1 year. 
 


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Extensive efforts have been devoted to the synthesis of anthracycline analogs with improved 
properties.  Initial efforts were directed towards analogs with reduced cardiotoxicity (Dalton et 
al. 1993; Moscow et al. 1988; Consoli et al. 1996).  More recently, triggered by the discovery of 
multidrug resistance and identification of P-glycoprotein, the efforts have focused on analogs 
with non-cross-resistant properties (Acton et al. 1984; Ganapathi et al. 1989; Barbieri et al. 1987; 
Priebe et al. 1993).  Several non-cross-resistant analogs have been identified.  They all have in 
common a markedly increased lipophilicity.  Some of them have a similar mechanism of 
cytotoxicity to that of doxorubicin, i.e., topoisomerase II inhibition.  Others have a different 
mechanism of action, i.e., DNA alkylation (Acton et al. 1984). 
 
Liposomes have been used extensively as carriers of doxorubicin and daunorubicin (Gabizon et 
al. 1982; Rahman et al. 1986).  Liposomal doxorubicin was found to be less cardiotoxic and 
more active than doxorubicin in models of liver metastases in mice (Mayhew et al. 1983; 
Herman et al. 1983).  Clinical studies have been conducted with different liposomal formulations 
of doxorubicin.  These trials have shown a maximum-tolerated dose (MTD) similar to that of 
free doxorubicin, a significant reduction of certain toxicities, such as gastrointestinal and 
vesicant effects, and an unchanged dose-limiting toxicity, i.e., myelosuppression.  These studies 
have also suggested a reduced cardiotoxic potential. 
 
Annamycin is an anthracycline antibiotic with a mechanism of action that appears to be 
inhibition of topoisomerase-II.  This drug was selected for its lack of cross-resistance and a high 
affinity for lipid membranes (Perez-Soler et al. 1990).  Because of this latter property, 
annamycin is an ideal compound for liposome entrapment, thus as this drug is completely 
insoluble in water solutions, liposomes can be used as a carrier for its intravenous administration.  
Liposomal annamycin was developed to combine the intrinsic favorable properties of the 
compound (lack of cross-resistance) with the potential advantages associated with liposome 
delivery (reduced cardiotoxicity and preferential distribution to certain organs), as well as the 
potential for resisting multidrug-resistant (MDR) activity (Perez-Soler et al. 1997; Kolonias et al. 
1999).  A liposomal formulation of annamycin was developed at MD Anderson Cancer Center 
that has high entrapment efficiency, physical stability, and chemical stability.   
 
1.3  Nonclinical Studies 
 
The in vitro cytotoxicity of liposomal annamycin was tested on a panel of 4 different parental 
cell lines and their respective MDR-1-expressing cell lines (Zou et al. 1994).  In all MDR cell 
lines tested (KB-V1, P388/Dox, CEM/Vbl, and 8226/R), there was minimal resistance to 
annamycin; however, as expected, resistance to doxorubicin was high.  Studies with MDR 
modulators and metabolic inhibitors indicated significant differences in the cellular 
transmembrane transport systems between doxorubicin and annamycin and suggest that 
annamycin efflux is not mediated by P-glycoprotein MDR (Perez-Soler et al. 1994).   
 


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In vivo, liposomal annamycin has shown lack of cross-resistance in KB-VI human xenografts (an 
MDR-resistant cell line) and enhanced antitumor activity compared with doxorubicin in several 
mouse tumor models such as leukemia (L-1210), melanoma (B16), reticulosarcoma (M5076), 
and Lewis lung carcinoma cells (Zou et al. 1994; Perez-Soler et al. 1990).  Results in KB and 
KB-V1 human xenografts demonstrate that liposomal annamycin was at least as effective as 
doxorubicin (Zou et al. 1994).  Liposomal annamycin was more active than doxorubicin in a 
leukemia L--1210 mouse tumor model (Perez-Soler et al. 1990).   
 
In mice, at a single intravenous dose of 15.7-mg/kg liposomal annamycin, myelosuppression was 
noted.  With weekly intravenous doses of 5.2-mg/kg liposomal annamycin for 6 weeks or 
3.1- and 4.2-mg/kg liposomal annamycin for 10 weeks, the cardiotoxicity of liposomal 
annamycin was less than equitoxic doses of doxorubicin (Zou et al. 1995).  In dogs, a single 
15-minute intravenous infusion of up to 1.42-mg/kg liposomal annamycin was well tolerated, 
with no clinically significant adverse effects, hematological and chemical changes, and 
pathological changes (Zou et al. 1995). 
 
Additional information about the nonclinical data with liposomal annamycin can be found in the 
Investigator's Brochure. 
 
1.4  Clinical Studies 
 
A Phase 1, open-label, dose-escalating clinical study was conducted in subjects with refractory 
or relapsed AML, myelodysplastic syndrome, or acute lymphocytic leukemia (ALL) (Study No. 
AR522-21) (Andreeff et al. 2001).  The study was initiated in March 1999 and completed in 
November 2000.  Cohorts of 3 subjects were to be treated, with a starting dose for the first cohort 
of 190 mg/m2/day intravenously (IV) for 3 consecutive days.  Dosing for additional cohorts was 
on an escalating basis (230-, 280-, and 350-mg/m2/day IV liposomal annamycin for 
3 consecutive days).  Therapy was to be repeated every 4 to 6 weeks. 
 
Twenty subjects were treated: 3 at 190 mg/m2/day, 2 at 230 mg/m2/day, 2 at 280 mg/m2/day, and 
13 at 350 mg/m2/day.  Subjects received up to 3 cycles of therapy.  An encouraging response was 
obtained in a subject with refractory ALL.  Specifically, 1 out of 3 ALL subjects treated had a 
complete remission.  This subject had 3 relapses prior to treatment with liposomal annamycin, 
and was treated with 350-mg/m2/day liposomal annamycin for 1 cycle and 280 mg/m2/day 
liposomal annamycin for 1 cycle.  The duration of response was 5 weeks.  An encouraging 
response was also obtained in a subject with refractory AML.  Specifically, 1 out of the 16 AML 
subjects treated had a complete remission.  This subject had failed induction therapy with CAT, 
and was treated with 280-mg/m2/day liposomal annamycin for 1 cycle and 210 mg/m2/day for 
2 cycles.  The duration of response was 13 weeks. 
 
A Phase 1/2, open-label, dose-escalating clinical study was conducted in subjects with refractory 
or relapsed ALL (Protocol No. CP-103; NCT00271063) (Wetzler et al. 2013).  The study was 
initiated in December 2005 and was completed in November 2008.  The starting annamycin dose 
was 190 mg/m2/day as a 2-hour intravenous infusion for 3 consecutive days followed by 18 days 
without study drug (treatment cycle equaled 21 days).   
 


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A total of 31 subjects were treated.  The MTD was determined to be 150 mg/m2/day for 3 days.  
Other than tumor lysis syndrome, which developed in 2 subjects, 3 severe adverse events (AEs) 
considered deﬁnitely related to the study drug consisted of Grade 3 mucositis, which defined the 
MTD.  There was also 1 case each of Grade 3 diarrhea, typhlitis, and nausea.  Left ventricular 
ejection fractions (LVEFs) were unchanged after 1 course of treatment.  Four subjects received 
more than 1 course of treatment; LVEF decreased in 2 of them (from 50%-55% to 38% in 1 and 
from 50%-55% to 35% in the other) after the second course of treatment and stayed the same in 
the other 2 subjects.  Two additional subjects who received only 1 course of annamycin had late 
LVEF determinations that were unchanged. 
After the MTD was determined, 10 subjects were enrolled in the Phase 2 section of the trial.  
Eight of the subjects completed 1 cycle of the 3 days of treatment at the MTD.  Of these, 5 (62%) 
had an efﬁcacy signal with complete clearing of circulating peripheral blasts.  Three of these 
subjects also cleared bone marrow blasts with 1 subsequently proceeding on to successful stem 
cell transplantation.  The other 2 developed tumor lysis syndrome and died before response 
assessment. 
In addition to the 2 clinical trials in leukemia subjects, liposomal annamycin has been studied in 
2 clinical trials to determine the safety and efficacy in subjects with relapsed solid tumors, 
doxorubicin-resistant breast cancer, and metastatic breast cancer (NCT00012129) (Booser et al. 
2002; Booser et al. 2000).  Doses in these 2 studies ranged from 3- to 240-mg/m2 IV liposomal 
annamycin given every 21 days. 
Additional information about the previous human experience with liposomal annamycin can be 
found in the Investigator's Brochure. 
1.5  Rationale for the Current Study 
Based on the positive clinical results in subjects with acute leukemia in 2 previous clinical 
studies of single-agent liposomal annamycin (Study No. AR522-21 and Study No. CP-103; see 
Section 1.4), the nonclinical data showing both lack of cross-resistance with anthracyclines 
(which is the standard of care for first-line treatment of AML), the increased activity in certain 
leukemia cells lines compared with doxorubicin, and the potential for decreased cardiotoxicity, 
the current study (Study No. MB-104) was designed to explore liposomal annamycin in a 
Phase 1/2 investigation for the treatment of relapsed/refractory AML.  Safety, pharmacokinetics, 
and clinical response will be evaluated.  The objective of the study is to determine the safety and 
tolerability of annamycin, identify dose-limiting toxicities, and define the MTD, as well as the 
recommended Phase 2 dose (RP2D).  The design of this study incorporates a 3+3 dose-escalation 
design and expansion at the RP2D for evaluation of clinical efficacy in up to 21 additional 
subjects. 
1.6  Consideration of Cardiotoxicity in the Current Study 
On the basis of concerns about the possible risk of cardiotoxicity in subjects who have likely 
received prior anthracycline therapy (Zamorano et al. 2016; Gianni et al. 2008), Table 1 and 
Table 2 establish some important considerations, including baseline risk factors and maximal 


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prior anthracycline doses, to be calculated on the basis of equivalent anthracycline exposure 
anticipated.  This protocol incorporates clear criteria for risk factors, as well as prior 
anthracycline exposure, to exclude subjects at cardiac risk from this study.   
 
 
Table 1: Baseline Risk Factors for Cardiac Disease (Zamorano et al. 2016) 
 
Current Myocardial Disease: 
Heart failure (with either preserved or reduced ejection fraction) 
Asymptomatic LV dysfunction (LVEF <50% or high natriuretic peptide) 
Evidence of CAD (previous MI, angina, PCI or CABG, myocardial ischemia) 
Moderate and severe VHD with LVH or LV impairment 
Hypertensive heart disease with LVH 
Cardiomyopathy (hypertrophic, dilated, or restrictive) 
Cardiac sarcoidosis with myocardial involvement 
Significant cardiac arrhythmias 
Demographic and Other CV Risk Factors: 
Age (>65 years) 
Family history of premature CV disease 
Arterial hypertension 
Diabetes mellitus 
Hypercholesterolemia 
Lifestyle Risk Factors: 
Smoking 
High alcohol intake 
Obesity 
Sedentary 
Abbreviations: CABG = coronary artery bypass graft; CAD = coronary artery disease; CV = cardiovascular; 
LV = left ventricular; LVEF = left ventricular ejection fraction; LVH = left ventricular hyperthropy; 
MI = myocardial infarction; PCI = percutaneous coronary intervention; VHD = valvular heart disease. 
 
Table 2:  Cardioprotective Measures for Anthracycline Drugs (adapted from Zamorano et 
al. 2016) 
 
Potential Cardioprotective Measures for Anthracycline 
Drugs and Analogues by Limitation of Cumulative Doses 
Drug 
Conversion Factor 
(Daunorubicin 
Equivalents) 
Daunorubicin 
1 
Doxorubicin 
2.2 
Epirubicin 
1.1 
Mitoxantrone 
5 
Idarubicin 
5.3 
 
A value of <551 mg/m2 will be used as the correlative value for liposome-encapsulated 
annamycin for purposes of assessment of maximal exposure, with a conversion factor of 1.  


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Consideration of the equivalence with conversion factor and sample calculations of these 
anthracyclines to calculate prior exposure are provided in Appendix E. 
 
2.  STUDY OBJECTIVES 
 
The primary objective of this study is to evaluate the safety and identify the RP2D of liposomal 
annamycin for the treatment of subjects with AML that is refractory to or relapsed after standard 
induction therapy.   
 
Secondary objectives: 
1. Pharmacokinetics of annamycin and its metabolite, annamycinol 
2. Preliminary assessment of the antileukemic activity of liposomal annamycin as second-
line therapy for subjects with refractory or relapsed AML based on established response 
criteria, including complete response (CR), partial response (PR), event-free survival 
(EFS), overall survival (OS; Kaplan-Meier), and time to and duration of 
remission/response  
 
3.  STUDY DESIGN 
 
3.1  Design Summary 
 
This is a multicenter, open-label, dose-escalation study that will determine the MTD and RP2D 
of liposomal annamycin as a single agent for the treatment of subjects with AML that is 
refractory to or relapsed after standard induction therapy.   
 
During the dose-escalation phase of the study, eligible subjects could have received any number 
of previous therapies for their relapsed AML as long as the prior anthracycline cumulative dose 
is below the recommended noncardiotoxic level (as defined in Table 2 and Appendix E).  During 
the expansion phase of the study, after the MTD or RP2D has been established, study treatment 
will be second-line therapy, thus subjects could not have received prior therapy for their relapsed 
AML. 
 
Enrollment will occur in cohorts of 3 subjects in a conventional 3+3 escalating dose design, 
starting at a dose level of 100 mg/m2/day administered for 3 days.  Dose escalation will take 
place on the basis of safety assessments in this first cohort with the next dose of liposomal 
annamycin escalated to 120 mg/m2 in the absence of safety concerns.   
 
For each dose level, groups of 3 subjects will receive a treatment cycle of liposomal annamycin 
daily as a 2-hour IV infusion for 3 consecutive days followed by 18 days off study drug (i.e., the 
induction treatment cycle = 21 days).  If no subject experiences a dose-limiting toxicity (DLT), 
based on National Cancer Institute (NCI) Common Terminology Criteria for Adverse Events 
(CTCAE) v5, which constitutes a definite study drug-related irreversible or not medically 
manageable Grade 3 or higher toxicity, further defined in Section 3.4, during the induction 
treatment cycle through Day 28, the subsequent group of 3 subjects will receive the next higher 
liposomal annamycin dose.  However, if 1 of the 3 initial subjects experiences a DLT, the cohort 
of subjects at that dose level will be expanded to 6 subjects.  If at least 2 of the 6 subjects 


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experience a DLT, this will be considered a nontolerated dose and then 3 subjects will be treated 
at a lower dose.   
 
The MTD is defined as the highest dose of liposomal annamycin at which fewer than 2 (of a 
cohort of up to 6) subjects experience a DLT.   
 
The RP2D is defined as the optimal dose to be determined by the Sponsor in conjunction with 
the Safety Review Committee/Consultant (SRC), consisting of at least 1 to up to 3 independent 
consultants, on the basis of review of available clinical and laboratory safety and efficacy data.  
Up to 21 additional subjects will be enrolled at either the MTD or RP2D to better define toxicity 
and evaluate efficacy at this dose.  Therefore, up to 33 subjects will be enrolled in this study: up 
to 12 subjects in the dose-escalation phase and up to 21 additional subjects at the MTD or RP2D.  
 
The Medical Monitor and the SRC will continually review all data and decide if the study should 
continue and if any changes to the protocol before restarting enrollment must be made. 
 
If a subject discontinues treatment for reasons other than study drug-related adverse events such 
that safety or efficacy of the drug cannot be fully evaluated during the induction cycle, a 
replacement subject may be enrolled; these circumstances will be reviewed on a case-by-case 
basis by the SRC in conjunction with the Sponsor.  
 
3.2  Dosing  
 
The initial group of 3 subjects will be treated with a 2-hour intravenous infusion of 
100-mg/m2/day (Dose Level 1) liposomal annamycin daily for 3 consecutive days followed by 
18 days off study drug (i.e., 1 treatment cycle = 21 days).  Provided that no subject experiences a 
DLT (defined in Section 3.4) during the induction treatment cycle through Day 28, the ensuing 
group of 3 subjects will receive 120-mg/m2/day (Dose Level 2) liposomal annamycin.   
 
The liposomal annamycin dose levels planned for investigation are outlined in Table 3.  The 
starting dose is Dose Level 1. 
 
 
 
Table 3: Liposomal Annamycin Dose Levels 
 
Dose Level Designation 
Liposomal Annamycin Dose 
(mg/m2/day) × 3 days 
-1 
90 
1 
100 
2 
120 
 
For each dosing cohort, if 1 of the 3 initial subjects experiences a DLT (Section 3.4), the cohort 
of subjects at that dose level will be expanded to 6 subjects (Figure 1).  If at least 2 of the 
6 subjects experience a DLT, then 3 subjects will be treated at a lower dose.  If a DLT occurs in 
at least 2 out of 6 subjects at Dose Level 1, the next 3 subjects will be enrolled into 
Dose Level -1.  The MTD is defined as the highest dose of liposomal annamycin at which fewer 
than 2 (of a cohort of up to 6) subjects experience a DLT.  The RP2D is defined as the dose to be 


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explored in the expansion phase portion of the study, based on the aggregate data of safety and 
efficacy. 
 
Figure 1:  Schematic of Dosing Plan 
 
Liposomal annamycin will be administered for a single induction cycle. 
 
 
Abbreviations: DLT = dose-limiting toxicity; HSCT = hematopoietic stem cell transplantation. 
 
3.3  Safety Considerations 
 
Subjects will be hospitalized for the full 3 days of liposomal annamycin administration.  If a 
subject experiences a DLT as defined in Section 3.4, or any severe or life-threatening event 
occurs at any time over the first 3 days, dosing should be stopped and the subject should be 
permanently discontinued from the study.  In addition to the DLT criteria in Section 3.4, the 
severe or life-threatening events include (but are not limited to) development of an anaphylactic 
reaction during or after the infusion, clinical or radiological evidence of left ventricular 
dysfunction with dyspnea and pulmonary rales, cardiomegaly or signs of pulmonary 
hypertension on chest x-ray, or acute cardiotoxicity (Section 3.3.1).   
 
Additional criteria for safety evaluation after the initial dose during the induction cycle includes 
creatinine levels increasing to 1.5 times those of baseline or liver function tests increasing to 
2 times those of baseline, in which case the next dose of the study drug will be decreased by 
25%.  If there is further deterioration of organ function after the next dose, the subject will be 
permanently discontinued from the trial.  If these changes are acutely 2 or 3 times those of 
baseline, respectively, the study drug will be held, and subjects will be monitored for resolution 
of these changes.  If there is prompt resolution (within 2 to 5 days), a rechallenge at a 25% lower 
dose will be attempted, and if any additional toxicity is observed, the subject will be permanently 


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discontinued from the trial.  If resolution takes longer than 5 days, the subject will not be 
rechallenged and will be permanently discontinued from the trial. 
Subjects successfully completing the first 3 days of dosing will then be evaluated weekly 
thereafter during the induction cycle of treatment (the induction cycle consists of 21 days 
[3 weeks total], with the first 3 consecutive days of daily liposomal annamycin treatment 
followed by 18 days off of liposomal annamycin).  
3.3.1  Cardiotoxicity 
Subjects will be evaluated before the start of liposomal annamycin treatment and during the first 
3 days of liposomal annamycin treatment, with special consideration for potential cardiotoxicity 
based on documented literature (Section 1.6) that provides currently relevant recommendations.  
Since acute cardiotoxicity may occur immediately to weeks following treatment and can present 
as arrhythmias, ST and T wave abnormalities, pericarditis-myocarditis syndrome, and acute heart 
failure, the subject will be observed in the hospital with daily, diligent attention to evaluation of 
cardiac status, inasmuch as prompt therapy can provide substantial recovery (Cardinale et al. 
2015).  Guidelines for limiting the total cumulative dose of anthracyclines will be followed, as 
per an inclusion criterion that limits any subject with significant cumulative dosing due to prior 
regimens in addition to anticipated exposure to annamycin (See Section 4 and Appendix E).  
Subjects will also have clinical and electrocardiographic examinations evaluated at the 
anticipated maximal plasma concentrations, and again weekly during the induction cycle of 
treatment with multiple-gated acquisition (MUGA) or echocardiogram (ECHO) scans or 
ECHO strain evaluations performed every 3 weeks by using the same modality and instrument.  
Any subject demonstrating any of the above manifestations of cardiotoxicity or >10% decrease 
in ejection fraction based on scans will be discontinued from the study.  Between the Day 18 and 
Day 28 visits or at the the End of Treatment visit (if the subject is prematurely terminated), a 
final MUGA or ECHO scan or an ECHO strain evaluation will be conducted by using the same 
modality and instrument.   
3.3.2  Myalgia – Adverse Event Workup 
If a subject develops study drug-related ≥Grade 3 myalgia on study, total creatine phosphokinase 
and creatine phosphokinase isoenzyme measurements should be performed.  Appropriate 
treatment with analgesics should also be initiated. 
3.4  Criteria for Dose-Limiting Toxicity 
A DLT is defined as any of the following clinically significant AEs or laboratory abnormalities 
that occur during the DLT evaluation period except those that are clearly and incontrovertibly 
due to extraneous causes, including disease progression, underlying disease, concurrent illness or 
concomitant medications.  The DLT evaluation period is defined as the first day of treatment 
(Day 1) until Day 28.  DLT observation periods for safety monitoring in expansion cohorts will 
be as above. The severity of AEs will be graded according to the NCI CTCAE v5.  


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Any Grade ≥ 3 nonhematologic or extramedullary toxicity with the following exceptions: 
•
Anorexia or fatigue;
•
Grade 3 nausea and/or vomiting if not requiring tube feeding or total parenteral nutrition
or diarrhea if not requiring or prolonging hospitalization that can be managed to Grade ≤2
with standard antiemetic or antidiarrheal medications used at prescribed dose within
7 days of onset;
•
Grade 3 mucositis that resolves to Grade ≤2 within 7 days of onset;
•
Grade 3 fever with neutropenia, with or without infection; or
•
Grade 3 infection.
Any subject demonstrating the defining criteria for Hy’s Law (Appendix D) will be discontinued 
from the study, and this will be considered a DLT. 
Subjects who do not achieve a remission of leukemia will not be evaluable for hematological 
toxicity. 
3.5  Evaluation of Efficacy 
3.5.1  Bone Marrow Evaluation 
Bone marrow aspirate (a biopsy if there are no spicules present) 15 to 35 days after the start of 
therapy will be performed.  If cytogenetics were initially abnormal, include cytogenetics as part 
of the remission documentation. 
3.6  Premedications and Prophylactic Treatments 
3.6.1  Tumor Lysis Syndrome Prophylaxis 
All subjects will receive tumor lysis syndrome prophylaxis as per institutional guidelines or as 
per published information (Jones et al. 2015; Coiffier et al. 2010). 
3.6.2  Allergic Reactions 
Antiallergic premedication with an antihistamine (e.g., diphenhydramine at a dose of 50 mg 
administered intravenously) must be administered before each dose of liposomal annamycin. 
Subjects who develop allergic reactions must have their infusions stopped and be treated with 
corticosteroids and/or diphenhydramine and other supportive measures as deemed appropriate by 
the Investigator on the basis of institutional standards of care.  After the reaction has completely 
resolved, the infusion can be restarted at 25% of the original rate and slowly increased to 50% of 
the original rate.  For subjects who experience an allergic reaction, prophylaxis with 
corticosteroids, as well as an antihistamine administered intravenously, will be required for all 
subsequent infusions. If any subjects experience an anaphylactic reaction to therapy, they will be 
permanently discontinued from the study. 


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3.6.3  Oral Prophylaxis for Mucositis 
3.6.3.1  Mouthwash 
Subjects will receive mouthwash as per the institution’s guidelines and practices. 
3.6.3.2  Palifermin (Kepivance) 
The use of Kepivance should be considered if institutional guidelines and the Investigator 
recommend it, but is not required.  Kepivance is indicated as supportive care for preparative 
regimens predicted to result in ≥Grade 3 mucositis in the majority of subjects.  The 
recommended dose of Kepivance is 60 µg/kg/day administered as an intravenous bolus injection 
for 3 consecutive days before and 3 consecutive days after myelotoxic therapy for a total of 
6 doses. 
3.6.3.3  Oral Cryotherapy 
Subjects must receive oral cryotherapy (ice chips) during liposomal annamycin administration, 
utilize the recommended administration at 5 minutes prior to treatment, and continue for 2 to 
6 hours thereafter (Lilleby et al. 2006). 
4. ELIGIBILITY CRITERIA
Subjects were diagnosed with AML refractory to or relapsed after standard induction therapy.  
All subjects must meet the inclusion and exclusion criteria listed below to be eligible for 
enrollment.  Subjects with central nervous system (CNS) involvement will be eligible for this 
study.  They will receive CNS therapy as per standard of care. 
4.1  Inclusion Criteria 
A subject may be included in this study if he/she meets all of the following criteria: 
1. A pathologically confirmed diagnosis of AML by World Health Organization
classification
2. AML that is refractory to or relapsed after standard induction therapy
3. Age ≥18 years at the time of signing informed consent
4. No chemotherapy, radiation, or major surgery within 2 weeks prior to first dose of study
drug and/or recovered from the toxic side effects of that therapy unless treatment is
indicated due to progressive disease
5. No investigational therapy within 4 weeks of the first dose of study drug
6. Eastern Cooperative Oncology Group (ECOG) performance status 0 to 2


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7. Adequate laboratory results including the following: 
a. Bilirubin ≤1.5 times the upper limit of normal unless due to Gilbert Syndrome. 
b. Serum glutamic-oxaloacetic transaminase (SGOT), serum glutamic-pyruvic 
transaminase (SGPT), and alkaline phosphatase <3 times the upper limit of normal 
unless due to organ involvement. 
c. Adequate renal function (The Cockroft-Gault equation will be used to estimate 
creatinine clearance.  This equation is as follows:  Creatinine clearance in milliliters 
per minute = [140 – age] × body weight [kg]/72 × plasma creatinine [mg/dL], 
multiplied by 0.85 for women.  By using this equation, adequate renal function will 
be deemed to be a creatinine clearance of greater than 60 mL/minute.) 
 
8. Prior anthracycline cumulative dose (see Table 2 and Appendix E) <551 mg/m2 or the 
daunorubicin equivalent, which is the recommended noncardiotoxic level.  This will be 
calculated as follows: 
a. Determination of prior exposure will be based on total milligrams per meter squared 
of prior anthracycline therapy (see Appendix C for commonly used chemotherapy 
regimens incorporating anthracyclines). 
b. Potential cardioprotective measures for anthracycline drugs and analogues by 
limitation of cumulative doses are demonstrated by the conversion factors in Table 2. 
c. The maximal allowance will be calculated as a daunorubicin equivalent added to the 
anticipated exposure to liposomal annamycin with the maximal exposure of 
anthracyclines capped at 550.9 mg/m2 (Appendix E). 
d. Exposure to annamycin projected for subjects enrolled in a given cohort in this study 
will be assessed on the basis of prior total exposure added to the anticipated 
designated cohort dose level (up to 120 mg/m2, which, after 3 doses, represents a 
cumulative exposure of <551-mg/m2 daunorubicin equivalents). 
 
9. Subject can understand and sign the informed consent document, can communicate with 
the Investigator, and can understand and comply with the requirements of the protocol. 
 
10. Women of childbearing potential must have a negative serum or urine pregnancy test. 
 
11. All men and women must agree to practice effective contraception during the entire study 
period and after discontinuing study drug, unless documentation of infertility exists.   
a. Sexually active, fertile women must use 2 effective forms of contraception 
(abstinence, intrauterine device, oral contraceptive, or double barrier device) from 
the time of informed consent and until at least 6 months after discontinuing study 
drug. 
b. Sexually active men and their sexual partners must use effective contraceptive 
methods from the time of subject informed consent and until at least 3 months after 
discontinuing study drug. 
 


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4.2  Exclusion Criteria 
 
A subject will be excluded from this study if he/she meets any of the following criteria: 
 
1. Subjects diagnosed with acute promyelocytic leukemia 
 
2. Concomitant therapy that includes other chemotherapy that is or may be active against 
AML, except for prophylaxis and/or treatment of opportunistic or other infection with 
antibiotics, antifungals, and/or antiviral agents 
 
3. Prior mediastinal radiotherapy 
 
4. Any condition that, in the opinion of the Investigator, places the subject at unacceptable 
risk if he/she were to participate in the study 
 
5. LVEF <50%, valvular heart disease, or severe hypertension (see Table 1).  Cardiac 
subjects with a New York Heart Association classification of 3 or 4 will be excluded. 
(Cardiology consultation should be requested if any question arises about cardiac 
function.)  This also includes subjects with baseline QT/QTc interval >480 msec, a 
history of additional risk factors for torsade des pointes (e.g., heart failure, hypokalemia, 
family history of Long QT Syndrome), and use of concomitant medications that 
significantly prolong the QT/QTc interval. 
 
6. Clinically relevant serious comorbid medical conditions including, but not limited to, 
active infection, recent (less than or equal to 6 months) myocardial infarction, unstable 
angina, symptomatic congestive heart failure, uncontrolled hypertension, uncontrolled 
cardiac arrhythmias, chronic obstructive or chronic restrictive pulmonary disease, active 
CNS disease uncontrolled by standard of care, known positive status for human 
immunodeficiency virus and/or active hepatitis B or C, cirrhosis, or psychiatric 
illness/social situations that would limit compliance with study requirements  
 
7. Pregnant, lactating, or not using adequate contraception 
 
8. Known allergy to anthracyclines 
 
9. Any evidence of mucositis/stomatitis or previous history of severe (≥Grade 3) mucositis 
from prior therapy 
 
10. Required use of strong inhibitors and inducers of the Cytochrome P450 family of 
enzymes (CYP) and transporters that cannot be held during treatment days 
 
4.3  Subject Withdrawal 
 
Subjects may voluntarily withdraw from the study at any time for any reason and without 
prejudice to further treatment or the Investigator may withdraw subjects if necessary. 


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Subjects who are removed from the study will undergo all study evaluations described for the 
End of Study visit (Section 6.4).  The reasons for withdrawal will be recorded.   
 
The reasons for which a subject may be prematurely discontinued include the following: 
 
1. Subject has unacceptable toxicity, subject demonstrates manifestations of cardiotoxicity, 
or subject has >10% decrease in ejection fraction based on scans.  All subjects who are 
discontinued as a result of an adverse event should be followed for as long as necessary 
to document the resolution or stabilization of the event. 
 
2. Subjects with disease progression 
 
3. Subject withdrawal of consent 
 
4. Investigator's discretion 
 
5. General or specific changes in the subject's condition that renders the subject ineligible 
for further treatment 
 
6. If pregnancy occurs or is suspected during study drug treatment, annamycin must be 
discontinued immediately.  The subject should be referred to an obstetrician/gynecologist 
experienced in reproductive toxicity for further evaluation and counseling.  If none is 
immediately available, contact Moleculin Biotech for a referral. 
 
7. If the subject's continued participation poses unacceptable risk for any other reason or if 
they are not compliant with the study  
 
4.3.1  Withdrawal Procedures 
 
Subjects withdrawn from the study will complete the end of study (EOS) procedures described in 
Section 6.4 and be followed until resolution of any AEs or until the unresolved AEs are judged 
by the Investigator to have stabilized. 
 
The Investigator will promptly notify the Medical Monitor of a subject’s withdrawal for reasons 
related to toxicity. 
 
4.3.2  Subject Replacement 
 
Subjects withdrawing in the cohort dose-escalation period for reasons other than toxicity who 
have not taken all 3 doses during the induction cycle and/or fail to complete restaging after 
completion of the treatment cycle such that safety is not evaluable will be replaced. However, 
subjects experiencing a DLT in the dose-finding cycle will not be replaced. 
 


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5.  STUDY TREATMENT 
 
5.1  Liposomal Annamycin (Study Drug) 
 
5.1.1  Description of Liposomal Annamycin 
 
Annamycin is a lipophilic anthracycline antibiotic that incorporates 4 structural modifications 
from doxorubicin: 2’-iodo, 3’-hydroxy, 4’-epi, 4-demethoxy doxorubicin.  Annamycin is 
completely insoluble in water but soluble in chloroform and dimethyl sulfoxide. 
 
Liposomal annamycin will be supplied by Moleculin Biotech, Inc., in 50-mL vials containing 
annamycin as a lyophilized powder.  The study drug also contains phospholipids 
(dimyristoylphosphatidyl choline and dimyristoylphosphatidyl glycerol) and polysorbate 20.  
 
Study drug will be shipped to the pharmacy at the study site in individually labeled vials.  Each 
vial is labeled with the following information: drug name, strength, lot number, date of 
manufacture, storage conditions, sponsor’s address, manufacturer’s name, and the statement 
“Caution:  New Drug - Limited to Investigational Use.” 
 
This study is an open-label study, and study drug will not be blinded, nor will placebo product be 
administered. 
 
5.1.2  Preparation of Liposomal Annamycin 
 
The study drug must be reconstituted prior to use.  The diluted product can be held for up to 
24 hours when stored between 34 to 42ºC, after which time unused drug should not be 
administered to subjects. 
 
Caution should be exercised in the handling and preparation of liposomal annamycin.  The use of 
gloves is required.  If liposomal annamycin comes into contact with skin or mucosa, immediately 
wash thoroughly with soap and water.  Liposomal annamycin should be handled in a manner 
consistent with other anticancer drugs.   
 
5.1.3  Liposomal Annamycin Administration 
 
Subjects will be treated daily for 3 consecutive days with a 2-hour intravenous infusion of 
liposomal annamycin followed by 18 days off liposomal annamycin (i.e., 1 treatment 
cycle = 21 days).  A peripheral vein may be used.  The starting dose of liposomal annamycin 
(dose for subjects in Cohort 1) is 100 mg/m2/day.  The liposomal annamycin doses will be 
escalated in sequential cohorts.  Discussions of dose escalation and modifications are provided in 
Section 3.2 and Table 3. 
 
5.1.4  Storage 
 
At the study site, all investigational study drugs will be stored in a locked, safe area to prevent 
unauthorized access.  The lyophilized liposomal annamycin powder should be stored in the 


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freezer at -10 to -25°C (-13 to 14°F).  The reconstituted liposomal annamycin suspension is 
prepared as described in the Pharmacy Manual.  
 
5.2  Concomitant Medications 
 
All medications, whether prescription or nonprescription and including pharmacologic doses of 
vitamins, must be recorded on the Electronic Case Report Form/Electronic Data Capture 
(eCRF/EDC).  
 
Subjects must not be receiving chemotherapeutics that are or may be active against AML, 
including investigational agents, while taking study drug, except for prophylaxis and/or 
treatment of opportunistic or other infection with antibiotics, antifungals, and/or antiviral agents.  
Subjects should not be undergoing concurrent radiation therapy.  No new investigational 
treatments are to be initiated during the subject’s participation in the study.  
 
On the basis of the known metabolism of anthracyclines by CYP3A and CYP2B, enzymes of the 
Cytochrome P450 family, strong inducers and inhibitors of these enzymes will be prohibited 
during the days of treatment.  Information about relevant drugs can be accessed through this 
website: http://bioinformatics.charite.de/transformer/, which easily provides drug interaction 
information; however, any concerns should be discussed with the Sponsor. 
 
5.3  Measuring Subject Compliance 
 
Study drug will be administered only at the investigational site and to subjects enrolled in this 
clinical study under the supervision of the Investigator in accordance with the protocol and 
institutional practice.  Administration of drug will be recorded on the eCRF/EDC. Under no 
circumstances is the Investigator allowed to release clinical supplies for use by another physician 
not named on Form Food and Drug Administration (FDA) 1572, or to administer assigned study 
drugs to a subject who is not enrolled in this study. Study drug will be dispensed only at the 
institution specified on Form FDA 1572 for US sites. For EU sites, FDA form will only be 
collected if permitted by the management of the participating institutions or local/national 
guidelines and regulations. 
 
5.4  Drug Accountability 
 
In accordance with current Good Clinical Practice (GCP), the investigational site will account for 
all supplies of annamycin.  Details of receipt, storage, administration, and return or destruction 
will be recorded in the study drug accountability record according to the standard operating 
procedures of the investigational site.  Copies of the study drug accountability record will be 
provided to the Sponsor. 
 
At the end of the study, Moleculin Biotech, Inc., will instruct the Investigator on the return or 
destruction of unused medication.  If any medication is lost or damaged, its disposition should be 
documented.  Investigational supplies will be retained at the clinical site pending instructions for 
disposition by Moleculin Biotech, Inc., at the end of the study. 
 


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6.  STUDY PROCEDURES 
 
See Appendix A and Appendix B. 
 
6.1  Screening  
 
The Investigator is responsible for keeping a record of all subjects screened for entry into the 
study and subsequently excluded.  The reason(s) for exclusion must also be recorded.  Written 
informed consent must be given before any study related diagnostic or screening procedures, 
unless performed as part of standard of care.  The following screening procedures must be 
performed within 35 days prior to study entry: 
 
1. Informed consent 
 
2. Refractory or relapsed AML diagnosis confirmed  
 
3. ECOG performance status (Appendix B) 
 
4. Complete medical history including documentation of all treatments given for AML, 
cytogenetic and molecular abnormalities obtained at diagnosis, as well as a full cardiac 
history including assessment of prior anthracycline exposure and calculation of potential 
additional exposure based on assignment to dosing cohort (Section 4) 
 
5. Physical examination including height, weight, vital signs, and oral examination 
 
6. Electrocardiogram (ECG) 
 
7. Blood chemistry (sodium, potassium, chloride, bicarbonate, calcium, magnesium, 
phosphorus, blood urea nitrogen [BUN], creatinine, glucose, albumin, total protein, 
alkaline phosphatase, total bilirubin, SGOT, SGPT], lactate dehydrogenase [LDH], and 
uric acid) with creatinine clearance to be calculated from these results 
 
8. Complete blood count (CBC) with differential and platelet count 
 
9. Urinalysis 
 
10. Concomitant medications used within 30 days prior to the first dose of study drug 
 
11. Pregnancy test (with sensitivity of at least 50 mIU/mL) for women of childbearing 
potential within 72 hours prior to first dose of study drug 
 
12. MUGA or ECHO scan or ECHO strain evaluation within 2 weeks prior to first dose of 
study drug 
 


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6.2  Baseline (Day 1) 
 
The subject will be seen at the scheduled baseline visit (Day 1).  The Investigator will certify that 
the subject continues to satisfy all inclusion and exclusion criteria.  If eligible, the subject will be 
enrolled into the study and will be assigned an identification number in sequential order of 
enrollment.  The following baseline evaluations will be made just prior to dosing on Day 1 
(unless otherwise indicated):  
 
1. Updated medical history 
 
2. Physical examination including weight, vital signs, and oral examination 
 
3. ECOG performance status (Appendix B) 
 
4. Blood samples for pharmacokinetic analysis and cardiac biomarkers (troponin-I and 
troponin-T) collected just prior to the start of the annamycin infusion on Day 1 
 
5. Bone marrow aspirate and peripheral blood (complete blood count, differential, and 
platelet count; flow cytometry is optional) within 15 days prior to the first dose of 
liposomal annamycin to document disease 
 
6. Updated concomitant medications 
 
If screening was ≥14 days prior to first dose of study drug, the following should be repeated: 
 
1. Blood chemistry (sodium, potassium, chloride, bicarbonate, calcium, magnesium, 
phosphorus, BUN, creatinine, glucose, albumin, total protein, alkaline phosphatase, total 
bilirubin, SGOT, SGPT, LDH, and uric acid) with creatinine clearance to be calculated 
from these results 
 
2. CBC with differential and platelet count 
 
3. Urinalysis 
 
4. ECG 
 


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6.3  On-Study Evaluations  
 
6.3.1  Study Drug Administration  
 
Dosing will be in the morning at approximately the same time each day.  Subjects will be treated 
with a daily 2-hour intravenous infusion of liposomal annamycin for 3 consecutive days. 
Prophylaxis as described in Section 3.6 will be administered.  Antiallergic premedication with 
diphenhydramine at a dose of 50 mg administered intravenously must be administered before 
each dose of liposomal annamycin.  During each period of study drug administration subjects 
must be observed for at least 4 to 6 hours following daily liposomal annamycin administration.  
The following evaluations will be conducted during the 3 days of liposomal annamycin 
administration (Days 1 to 3): 
 
1. Physical examination including weight, vital signs, and oral examination on Days 2 and 3 
 
2. Blood samples for pharmacokinetic analysis collected at predose and at 0.25, 0.5, 1, 2, 4, 
8, and 24 hours after the start of liposomal annamycin infusion on Day 1 and Day 3 for 
3 subjects at each dose and 6 subjects at the RP2D who complete the 3 days of dosing 
 
3. Blood samples for cardiac biomarkers (troponin-I and troponin-T) collected at predose, at 
the end of infusion on Day 3, and on Day 21 
 
4. ECG monitoring at predose and at 4 hours and 24 hours after the start of liposomal 
annamycin infusion on Day 1, Day 2, and Day 3 for all subjects at each dose and 
6 subjects on Day 1 and Day 3 only at the RP2D who complete the 3 days of dosing, 
coinciding with pharmacokinetic (PK) evaluations  
 
5. Monitoring of adverse events and review of concomitant medications and concurrent 
illnesses (treatment-emergent or worsening illness must be recorded as an adverse event).  
Evaluation of blood chemistries and blood counts daily.  Over the 3 days of initial dosing, 
if any toxicity occurs, including blood count abnormalities, dosing should be interrupted 
until resolution, with rechallenge at a 25% lower dose, based on guidelines in 
Sections 3.3 and 3.4.  If any DLT is encountered (as defined in Sections 3.3 and 3.4) 
including any severe or life-threatening event, the subject should be discontinued from 
the study.  These specific events include (but are not limited to) development of an 
anaphylactic reaction during or after the infusion or clinical or radiological evidence of 
left ventricular dysfunction with dyspnea and pulmonary rales, cardiomegaly, or signs of 
pulmonary hypertension on chest x-ray. 
 
6. Concomitant medications  
 


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6.3.2  Induction Cycle 
Subjects will return to the clinic weekly on Days 7, 14, and 21 during the induction cycle.  
The evaluations listed below will be conducted at each weekly evaluation during the induction 
cycle (Appendix A):   
1. Monitoring of adverse events and review of concurrent illnesses (treatment-emergent or
worsening illness must be recorded as an adverse event)
2. Concomitant medications
3. Physical examination including weight, vital signs, and oral examination
4. CBC with differential and platelet count
5. Blood chemistry (sodium, potassium, chloride, bicarbonate, calcium, magnesium,
phosphorus, BUN, creatinine, glucose, albumin, total protein, alkaline phosphatase, total
bilirubin, SGOT, SGPT, LDH, and uric acid)
6. Pregnancy test (with sensitivity of at least 50 mIU/mL) for women of childbearing
potential on Day 7
7. ECG
In addition to the weekly evaluations during the induction cycle, the following evaluations will 
be conducted at the end of the 3-week treatment cycle: 
1. MUGA or ECHO scan or ECHO strain evaluation between Days 18 and 28.  Cardiology
consultation should be requested if any question arises about cardiac function. The
MUGA scan should also be conducted at the Early Termination visit for subjects who are
withdrawn prematurely.
2. Bone marrow aspirate and peripheral blood specimens (complete blood count, 
differential, and platelet count; flow cytometry is optional) between Days 15 and 35 to 
document response
3. Pregnancy test (with sensitivity of at least 50 mIU/mL) for women of childbearing
potential on Day 21
4. Blood sample for cardiac biomarkers on Day 21


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6.4  End of Study Evaluations 
 
The subject will return to the clinic for an EOS evaluation within 1 week of their treatment cycle 
or last study drug administration if the treatment period was prematurely terminated or subject 
was withdrawn.  In addition, any subject with a treatment-related medical event that requires 
additional follow-up must continue to be monitored as medically appropriate until resolution or 
stabilization (in the event that resolution is not possible).  The procedures listed below are to be 
undertaken at the EOS visit: 
 
1. ECOG performance status (Appendix B) 
 
2. Blood chemistry (sodium, potassium, chloride, bicarbonate, calcium, magnesium, 
phosphorus, BUN, creatinine, glucose, albumin, total protein, alkaline phosphatase, total 
bilirubin, SGOT, SGPT, LDH, and uric acid)  
 
3. CBC with differential and platelet count  
 
4. Monitoring of adverse events and review of concurrent illnesses (treatment-emergent or 
worsening illness must be recorded as an adverse event) 
 
5. Concomitant medications 
 
6.5  Follow-up Evaluations 
 
Follow-up evaluations will occur 2 weeks after the EOS visit.  The procedures listed below are to 
be undertaken at the Follow-up visit: 
 
1. Physical examination including weight, vital signs, and oral examination 
 
2. Monitoring of adverse events and review of concurrent illnesses (treatment-emergent or 
worsening illnesses must be recorded as an adverse event).  Any subject with a 
treatment related medical event must continue to be monitored as medically appropriate 
until resolution or stabilization (in the event that resolution is not possible).   
 
3. Concomitant medications 
 
6.6  Survival Follow-up 
 
Subjects will be followed every 3 months from study drug discontinuation to monitor resolution 
or stabilization of any treatment-related medical events and obtain survival data.  If possible, a 
MUGA scan, ECHO scan, or ECHO strain evaluation will be collected. 
 


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7.  ASSESSMENT OF EFFICACY 
 
7.1  Efficacy Parameters 
 
7.1.1  Response Criteria 
 
All subjects evaluable for efficacy will be assessed for response to treatment by using the 
recommendations of the International Working Group for standardization of response criteria, 
treatment outcomes, and reporting for therapeutic trials (Cheson et al. 2003). The primary 
efficacy variable is leukemia response rate, evaluated by the Investigator at the end of the cycle 
and at the EOS visit, based on bone marrow aspirate and peripheral blood collected at the end of 
the treatment cycle.  A bone marrow (BM) aspirate (biopsy if there are no spicules present) 
should be repeated in 1 week if there is a question of residual leukemia in assessing efficacy 
based on an initial BM specimen.  
 
An evaluation of response to therapy is one of the objectives of this study.  Response criteria are 
described below: 
 
Primary Endpoints: 
 
1. Morphologic leukemia-free state 
a. BM <5% blasts in an aspirate with spicules (a BM biopsy should be performed if 
spicules are absent) 
b. No blasts with Auer rods or persistence of extramedullary disease  
2. Complete remission:  
a. Morphologic leukemia-free state in which 
i. Subject is independent of transfusions  
ii. Absolute neutrophil count of >1,000  
iii. Platelets of ≥100,000 
3. Cytogenetic complete response – cytogenetics normal in those with previously abnormal 
cytogenetics 
4. Cri and CRp: CR with incomplete blood count recovery meets all criteria for CR except 
for either neutropenia (<1,000; CRi) or thrombocytopenia (<100,000; CRp) but must 
include transfusion independence 
 
Secondary Endpoints: 
 
1. EFS: Time from enrollment until disease progression or death from any cause 
2. OS: Time from enrollment until death from any cause 
3. PR: All of the hematologic values for a CR, with normalization of blood counts, but with 
a decrease of at least 50% in the percentage of blasts to 5% to 25% in the bone marrow 
aspirate  
4. Time to and duration of remission/response  
 


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7.2  Methods for Measuring Efficacy 
 
Bone marrow aspirate/biopsy and peripheral blood specimens (complete blood count, 
differential, and platelet count) to document response will be collected at 15 to 35 days after the 
start of therapy. If cytogenetics were initially abnormal, include cytogenetics as part of the 
remission documentation. After documentation of remission, these procedures should be 
performed every cycle thereafter.  In addition, flow cytometry analysis of peripheral blood may 
be utilized for response assessment.   
 
8.  ASSESSMENT OF SAFETY 
 
8.1  Safety Parameters 
 
A primary objective of this study is to evaluate the safety and identify the MTD of liposomal 
annamycin when given in doses starting at 100 mg/m2/day and ranging to as high as 
120 mg/m2/day, or the MTD, whichever is lower, in subjects with refractory or relapsed AML.  
The Investigators, meeting on a regular basis, will discuss any safety issues arising in the study 
in conjunction with the Sponsor and the medical monitor.  The designation of an SRC (Section 
8.5) will be available to adjudicate any issues concerning safety, attribution, or subject 
disposition. 
 
8.2  Methods for Measuring Safety 
 
Clinical AEs, routine hematological and biochemical parameters, physical examination and vital 
signs, 12-lead ECG, ECOG performance status, pregnancy testing, and monitoring of 
concomitant medications will be assessed according to the Schedule of Events (Appendix A).   
 
All AEs and laboratory abnormalities will be graded for severity by using the NCI CTCAE v5. 
The relatedness of all AEs and laboratory abnormalities and their seriousness will be determined 
by the Investigator and adjudicated by the SRC in conjunction with the Sponsor. The SRC will 
consist of at least 1 and up to 3 external, independent (not participating in the clinical trial) 
consultants, providing guidance with regard to safety issues, including approval of dose 
escalation and designation of DLTs. 
 
AEs will be documented from the time of first dose of study drug until 39 days after the last dose 
administered on study.   
 
Strategies for detection of cardiotoxicity that include cardiac imaging (echocardiography, nuclear 
imaging, cardiac magnetic resonance) and biomarkers (troponin, natriuretic peptides) will be 
based on clinical assessments at screening, as well as current relevant guidelines (Zamorano et 
al. 2016; Cardinale et al. 2015). 
 


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8.3  Procedures for Recording Adverse Events 
 
All adverse drug events encountered during the clinical study will be recorded on the 
eCRF/EDC.  Adverse drug events are defined in the International Conference on Harmonisation 
(ICH) Guidance for Industry E6: Good Clinical Practice as follows: 
 
An AE is any untoward medical occurrence in a subject or clinical investigation 
subject administered a pharmaceutical product and that does not necessarily have 
a causal relationship with this treatment.  An AE can therefore be any unfavorable 
and unintended sign (including an abnormal laboratory finding), symptom, or 
disease temporally associated with the use of a medicinal (investigational) 
product, whether or not related to the medicinal (investigational) product 
(ICH April 1996). 
 
An adverse event is any adverse change from the subject’s baseline condition, including any 
clinical or clinically significant laboratory test value abnormality that occurs during the course of 
the clinical study after the experimental drug has been utilized, whether the adverse event is 
considered related to the treatment or not. 
 
All adverse events will be followed until resolution.  This may require obtaining clinical blood 
samples for appropriate laboratory tests until their values return to baseline levels or performing 
end of study or follow-up physical examinations until resolution of identified abnormalities. 
 
8.3.1  Routine Reporting of Adverse Events 
 
Adverse events, whether or not associated with study drug administration, will be recorded on 
the Adverse Event Report form of the eCRF/EDC and will be submitted to the Sponsor at 
regularly scheduled intervals. 
 
The information to be entered in the eCRF/EDC will include: 
 
1. The time of onset of any new adverse event or the worsening of a previously observed 
adverse event 
 
2. The specific type of reaction in standard medical terminology 
 
3. The duration of the adverse event (start and stop dates) 
 
4. The severity/grade of the adverse event.  The severity should be rated according to the 
NCI CTCAE v5. 
 
5. An assessment should be made of the relationship of the adverse event to the study drug, 
i.e., according to the definitions in Section 8.3.4. 
 
6. Description of action taken in treating the adverse event and/or change in study drug 
administration or dose 


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7. Outcome 
 
Follow-up assessments should be repeated to document return of any abnormalities to normal or 
to document other outcome of the adverse event. 
 
8.3.2  Reporting of Serious Adverse Drug Events, Including Deaths 
 
A serious adverse event (SAE), as defined in 21 Code of Federal Regulations 312.32 is: 
 
Any adverse drug event occurring at any dose that results in any of the following outcomes:  
 
1. Death 
 
2. A life-threatening adverse drug event (i.e., immediate risk of death from the event as it 
occurred; this does not include an adverse event that, had it occurred in a more serious 
form, might have caused death)  
 
3. Inpatient hospitalization or prolongation of existing hospitalization 
 
4. A persistent or significant disability/incapacity 
 
5. Congenital anomaly/birth defect 
 
Important medical events that may not result in death, be life-threatening, or require 
hospitalization may be considered a serious adverse drug event when, based upon appropriate 
medical judgment, they may jeopardize the subject or subject and may require medical or 
surgical intervention to prevent 1 of the outcomes listed in this definition.  Examples of such 
medical events include allergic bronchospasm requiring intensive treatment in an emergency 
room or at home, blood dyscrasias or convulsions that do not result in inpatient hospitalization, 
or the development of drug dependency or drug abuse. 
 
Serious adverse events, including death due to any cause, which occur from the time subject has 
taken the first dose of study drug or within 30 days following the last dose of study medication, 
regardless of study drug relation to the SAE, or any SAE occurring later than 30 days following 
the last dose of study medication with Investigator opinion as related to the study medication, 
must be reported immediately to Theradex Safety Desk (within 24 hours of the site becoming 
aware of the event) by email, fax, or telephone.  Notification by email is preferred. The fax and 
telephone numbers and the email address listed below may be used during both business and 
nonbusiness hours.  During nonbusiness hours a recorded message will provide the telephone 
caller with the contact information for the on-call monitor. 
 
All SAEs require that a serious adverse event report form be completed and forwarded either via 
facsimile or as a PDF via email to Theradex® at the fax number or email listed below within 
24 hours of the site becoming aware of the event. 
 


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All SAEs will be reported to: 
Theradex Safety Desk 
Email: 
SafetyDeskUS@theradex.com 
Fax: 
1-609-799-1567 
Telephone: 
1-609-799-7580 
 
In addition to the reporting of SAEs to the Theradex Safety Desk, the SAE must be reported to 
the Independent Ethics Committee/Institutional Review Board (IEC/IRB) according to their 
requirements, as well as to regulatory agencies based upon local reporting requirements. 
 
If the SAE is not previously documented in the Investigator’s Brochure and is thought to be 
related to the investigational treatment the Sponsor may urgently require futher information from 
the Investigator for Health Authority reporting.  The Sponsor may need to issue an Investigator 
Notification to inform all Investigators involved in any study with the same investigational 
treatment that this SAE has been reported. 
 
Hospitalizations due to the underlying disease will not be reported as an SAE unless there is 
reason to suspect a causal relationship with the study drug. 
 
8.3.3  Reporting of Suspected Unexpected Serious Adverse Reactions 
 
Suspected Unexpected Serious Adverse Reactions will be collected and reported to the 
competent authorities and relevant ethics committees in accordance with EU Guidance 
2001/C172/01 or as per national regulatory requirements in participating countries. 
 
8.3.4  Severity of Adverse Drug Events 
 
Adverse drug events will be graded according to the criteria in the NCI CTCAE v5.  The toxicity 
criteria can be accessed through the NCI website 
(https://ctep.cancer.gov/protocolDevelopment/electronic_applications/ctc.htm). 
 
8.3.5  Relationship of Adverse Events to the Study Drug 
 
The Investigator should record on the eCRF/EDC whether the relationship of the adverse event is 
best described as unrelated, possibly related, probably related, or related to the study medication, 
according to the following definitions. 
 
Unrelated: There is evidence that the adverse event definitely has an etiology other than 
the assigned study drug. 
 
Possibly Related: The adverse event has a temporal relationship to study drug 
administration.  However, an alternative etiology may be responsible for the adverse 
event.  Information on drug/product withdrawal may be lacking or unclear. 
 
Probably Related: The adverse event has a temporal relationship to study drug 
administration.  The event is unlikely to be related to an alternative etiology.  There is a 


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reasonable response on withdrawal (dechallenge).  Rechallenge information is not 
required. 
 
Related: The adverse event has a temporal relationship to study drug administration and 
resolves when the drug is discontinued.  An alternative etiology is not apparent.  If the 
subject is rechallenged with the assigned study drug, the adverse event recurs.  
Rechallenge is not necessarily required. 
 
8.4  Withdrawal From Study Treatment Due to Adverse Events 
 
Subjects withdrawn from receiving additional study drug as a result of an adverse event will be 
followed by the Investigator until the outcome is determined.  Additional reports will be 
provided to the Sponsor or regulatory authorities when requested.  Every effort will be made to 
follow the subject for the full study period as per the Schedule of Events (Appendix A). 
 
8.5 Establishment of a Safety Review Committee/Consultant  
 
At least 1 and up to 3 clinicians with the relevant expertise in AML, as well as experience in 
clinical trials, who are not participating in the current clinical study, will be assigned as the SRC 
to review safety considerations that affect dosing and drug administration.  Regular as well as 
well as ad hoc meetings will be convened to ensure subject disposition is based on an 
independent review of data and assessment of risk/benefit for subjects.  Designation of the 
appropriate individual(s) will be based on review and approval by the responsible IRBs. 
 
8.6 Monitoring of Cardiovascular Toxicity 
 
Blood samples will be drawn to assess cardiac biomarkers (troponin-I and troponin-T) at 
predose, at the end of the infusion on Day 3, and on Day 21.  
 
9.  PHARMACOKINETIC ASSESSMENT 
 
The pharmacokinetics of annamycin and its metabolite, annamycinol will be determined.  Blood 
samples for pharmacokinetic analysis collected at predose and at 0.25, 0.5, 1, 2, 4, 8, and 
24 hours after the start of liposomal annamycin infusion on Day 1 and Day 3 for 3 subjects at 
each dose and 6 subjects at the RP2D who complete the 3 days of dosing. 
 
ECGs and documentation of concomitant medications affecting CYP enzymes will be obtained 
to ensure that these are a consideration in the evaluation of the safety and pharmacokinetics of 
this drug.  ECG determinations for subjects at the RP2D will only be required on Days 1 and 3, 
coinciding with PK assessments. 
 
Instructions for the collection, processing, and shipment of samples will be provided in the 
Pharmacokinetic Manual. 
 
Plasma levels of annamycin and annamycinol will be determined by validated liquid 
chromatography with tandem mass spectroscopy.  Concentration-time profiles of annamycin and 


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annamycinol will be analyzed by noncompartmental methods and/or nonlinear least squares 
regression by using WinNonlin (Pharsight Corporation).  Pharmacokinetic parameters and 
variables will be calculated according to standard equations.   
 
10.  STATISTICS 
 
This is a multicenter, open-label, dose-escalation, MTD/RP2D safety study of liposomal 
annamycin given to subjects with refractory or relapsed AML. Therefore only descriptive 
statistics will be utilized.  A maximum of 33 subjects will be recruited for the study depending 
on the level at which toxicity is observed. 
 
Data from all subjects who receive 1 or more doses of liposomal annamycin will be incorporated 
into the final safety analysis.  Safety and tolerability will be assessed by adverse events, vital 
signs, physical examinations, laboratory parameters, and MUGA or ECHO scans or ECHO strain 
evaluations, as well as cardiotoxicity biomarkers.  Adverse events will be listed by subject, and 
will be tabulated and summarized as the number and percentage of subjects who reported each 
adverse event.  Descriptive statistics will be generated as appropriate; otherwise, no formal 
statistical analyses are planned on the safety data. 
 
Pharmacokinetic parameters (maximum concentration, area under the time-concentration curve , 
elimination half-life, volume of distribution, and clearance) will be calculated for annamycin and 
its metabolite, annamycinol, on the basis of plasma concentrations.  Pharmacokinetic data will be 
analyzed by descriptive statistics as appropriate. 
 
ECGs and documentation of concomitant medications affecting CYP enzymes will be obtained 
to ensure that these are a consideration in the evaluation of the safety and pharmacokinetics of 
this drug. 
 
The Medical Monitor and the SRC will continually review all data and decide if the study should 
continue and if any changes to the protocol before restarting enrollment must be made. 
 
11.  ACCESS TO SOURCE DOCUMENTS 
 
The Investigator will make the source documents for this trial available to the sponsor or its 
representatives, or to the regulatory authority or health authority inspectors. 
 
Subject medical information obtained as a result of this study is considered confidential and 
disclosure to third parties other than those noted below is prohibited.  All reports and 
communications relating to subjects in this study will identify each subject only by their initials 
and number.  Medical information resulting from a subject’s participation in this study may be 
given to the subject’s personal physician or to the appropriate medical personnel responsible for 
the subject’s welfare.  Data generated as a result of this study are to be available for inspection 
on request by FDA or other government regulatory agency auditors, the Sponsor Clinical 
Monitor (or designee), and the IRB/IEC. 
 


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The information developed in this clinical study will be used by the Sponsor in the clinical 
development of the study medication and therefore may be disclosed by the Sponsor as required 
for disclosure as a public company to other clinical investigators, to other pharmaceutical 
companies, to the FDA, and to other government agencies. 
 
Any information, inventions, or discoveries (whether patentable or not), innovations, 
suggestions, ideas, and reports, made or developed by the Investigator(s) as a result of 
conducting this study shall be promptly disclosed to the Sponsor and shall be the sole property of 
the Sponsor.  The Investigator agrees upon the Sponsor’s request and at the Sponsor's expense to 
execute such documents and to take such other actions as the Sponsor deems necessary or 
appropriate to obtain patents in the Sponsor’s name covering any of the foregoing. 
 
The Investigator shall retain the clinical study records until such time as directed by Moleculin.  
At a minimum, records are to be retained for at least 15 years after completion of the study. 
The Investigator will notify the sponsor prior to destroying any study records.  Should the 
investigator wish to assign the study records to another party or move them to another location, 
the sponsor must be notified in writing in advance. 
 
If the Investigator cannot guarantee this archiving requirement at the study site for any or all of 
the documents, special arrangements will be made between the Investigator and the sponsor for 
storage.  If source documents are required for continued care of the subject, appropriate copies 
for storage off site will be made. 
 
12.  QUALITY CONTROL AND QUALITY ASSURANCE 
 
Study monitors will periodically audit, at mutually convenient times during and after the study, 
all eCRF/EDCs and corresponding office and clinical laboratory records for each subject.  The 
monitoring visits provide the sponsor with the opportunity to evaluate the progress of the study, 
to verify the accuracy and completeness of eCRF/EDCs, to resolve any inconsistencies in the 
study records, and to assure that all protocol requirements, applicable Health Authority 
regulations, other requirements, and Investigator’s obligations are being fulfilled. 
 
13.  ETHICS   
 
13.1  Declaration of Helsinki 
 
The study will be conducted in accordance with the Declaration of Helsinki (1964) including all 
amendments up to and including the Fortaleza, Brazil revision (2013) as described in “Ethical 
Principles for Medical Research Involving Human Subjects,” which can be accessed at 
http://www.wma.net/en/30publications/10policies/b3/. 
 
13.2  Institutional Review Board 
 
The protocol, informed consent form, and any materials (such as advertisements, subject 
information sheets, or descriptions of the study used to obtain informed consent) for this study 
will be reviewed and approved by a duly constituted IRB/IEC.  


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The Investigator will ensure that all aspects of the IRB/IEC review are conducted in accordance 
with current institutional, local, and national regulations.  A letter documenting the IRB/IEC 
approval and, in Europe, also the national Competent Authority approval will be provided to the 
sponsor prior to initiation of the study.  Amendments to the protocol will be subject to the same 
requirements as the original protocol.  The Investigator or, in Europe, the Sponsor’s 
representative will submit all periodic reports and updates that the IRB/IEC may require, 
including any final close out reports.  The Investigator or, in Europe, the Sponsor’s 
representative will inform the IRB/IEC of any reportable adverse events. 
 
13.3  Informed Consent 
 
Each subject will be provided with oral and written information describing the nature and 
duration of the study in a language they can understand and must consent in writing to participate 
before undergoing screening.  The date of the consent shall be entered by the subject.  The 
original signed consent form will be retained with the study center’s records.  Each subject will 
also be given a copy of his/her signed consent form.   
 
14.  DATA HANDLING AND RECORDING 
 
eCRF/EDCs must be completed for every subject who signs an informed consent and has 
received study drug. 
 
The Investigator is responsible for the completeness and accuracy of information collected on the 
eCRF/EDCs for each individual enrolled.  The eCRF/EDCs will be signed and dated by the 
Investigator.   
 
The sponsor or designee will conduct data processing.  Completed eCRF/EDCs will be reviewed 
carefully for accuracy and completeness.  If necessary, the study site will be contacted for 
corrections and/or clarifications.  All data will be entered into a study database for analysis and 
reporting.  Any data captured electronically (such as laboratory data) will be transferred 
electronically to the database.  Upon completion of data entry, the database will receive a quality 
assurance check to ensure acceptable accuracy and completeness. 
 
15.  PUBLICATION POLICY 
 
The result of this study will be published under the direction of the Sponsor.  Results will not be 
published without prior review and approval by the Sponsor. 
 
16.  PROTOCOL AMENDMENTS AND MODIFICATIONS 
 
The Investigator will ensure that the study is conducted in accordance with the procedures and 
evaluations described in this protocol.  The Sponsor is responsible for all protocol amendments 
and modifications, except those intended to reduce immediate risk to subjects.  The Sponsor is 
responsible for submitting protocol amendments to the appropriate government regulatory 
authorities.  In the USA, the Investigator is responsible for submitting protocol amendments to 


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the appropriate IRB/IEC.  Approval by the IRB will be obtained before changes are implemented 
unless the Investigator concludes that an emergency exists.  In Europe, the Sponsor 
representative will obtain Competent Authority and IEC approval on behalf of the participating 
sites.  Other local/national approvals may be required depending on the country. 
 
When an emergency occurs that requires a departure from the protocol for a subject, a departure 
will be only for that subject.  The Investigator or other physician in attendance in such an 
emergency will, if circumstances and time permit, contact Moleculin Biotech, Inc., immediately 
by telephone.  Such contacts will be made as soon as possible to permit a decision as to whether 
or not the subject is to continue in the study.  The departure from the protocol will be described 
completely, including the reasons for the departure, in the subject's eCRF/EDC.  In addition, the 
IRB will be notified in writing of such departure from protocol. 
 


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and doxorubicin entrapped in cardiolipin liposomes. Cancer Res. 1986;46(5):2295-9. 
 
Wetzler M, Thomas DA, Wang ES, Shepard R, Ford LA, Heffner TL, et al. Phase I/II trial of 
nanomolecular liposomal annamycin in adult patients with relapsed/refractory acute 
lymphoblastic leukemia. Clin Lymphoma Myeloma Leuk. 2013;13:430-4. 
 
Zamorano JL, Lancellotti P, Rodriguez Muñoz D, Aboyans V, Asteggiano R, Galderisi M, et al; 
ESC Scientific Document Group. 2016 ESC Position Paper on cancer treatments and 
cardiovascular toxicity developed under the auspices of the ESC Committee for Practice 
Guidelines. The Task Force for cancer treatments and cardiovascular toxicity of the European 
Society of Cardiology (ESC). Eur Heart J. 2016;37(36):2768-2801. Erratum in: Eur Heart J. 
2018;39(10):839. 
 
Zou Y, Ling YH, Van NT, PriebeW, Perez-Soler R. Antitumor activity of free and liposome-
entrapped annamycin, a lipophilic anthracycline antibiotic with non-cross-resistance properties. 
Cancer Res. 1994;54(6):1479-84. 
 
Zou Y, Priebe W, Stephens LC, Perez-Soler R. Preclinical toxicity of liposome-incorporated 
annamycin: selective bone marrow toxicity with lack of cardiotoxicity. Clin Cancer Res. 
1995;1(11):1369-74. 
 


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CONFIDENTIAL 
APPENDIX A: SCHEDULE OF EVENTS 
Event 
Screening 
Baselinea 
On Treatment (Induction) 
End of 
Study 
Follow
-up 
Survival 
Follow-
up 
 
 
First 3 Dosing 
Days 
Weekly 
 
 
 
 
≥Day -35 
Day 1 
(Prior to 
Dosing) 
Day 
1 
Day 
2 
Day 
3 
Day 
7 
Day 
14 
Day 
15 
Day 
18 
Day 21  
(At the 
End of the 
Induction 
Cycle) 
Day 
28 
Day 
35 
Day 42 
Q3 
months 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
Relapsed/refractory 
AML diagnosis 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
ECOG performance 
status 
X 
X 
 
 
 
 
 
 
 
 
X 
 
 
 
Medical history 
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
Physical 
examinationb 
X 
X 
 
X 
X 
X 
X 
 
 
X 
 
 
X 
 
Electrocardiogram 
X 
Xi 
 
Xk 
Xk 
Xk 
X 
 
 
X 
 
 
 
 
Blood chemistryc 
X 
Xi 
 
X 
X 
X 
X 
 
 
X 
X 
 
 
 
CBC, differential, 
platelet countc,d 
X 
Xi 
 
X 
X 
X 
X 
 
 
X 
X 
 
 
 
Urinalysis 
X 
Xi 
 
 
 
 
 
 
 
 
 
 
 
 
Pregnancy test 
Xe 
 
 
 
 
Xe 
 
 
 
Xe 
 
 
 
 
MUGA, ECHO, or 
ECHO strain 
Xf 
 
 
 
 
 
 
 
Xg___________ ________________X 
 
 
Xq 
Bone marrow 
aspirate and 
peripheral bloodh 
for measuring 
disease 
 
Xj 
 
 
 
 
 
Xj_______________________________________________X 
 
 


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Abbreviations: AML = acute myeloid leukemia; BUN = blood urea nitrogen; CBC = complete blood count; CYP = Cytochrome P450 family of enzymes; 
DLT = dose-limiting toxicity; ECOG = Eastern Cooperative Oncology Group; ECG = electrocardiogram; ECHO = echocardiogram; LDH = lactate 
dehydrogenase; PK = pharmacokinetic; RP2D = recommended Phase 2 dose; SGOT = serum glutamic-oxaloacetic transaminase; SGPT = serum 
glutamic-pyruvic transaminase. 
a Baseline (Day 1) assessments are to be completed prior to dosing on Day 1. 
b Including height (screening only), weight, vital signs, and oral examination; this will occur daily for the first 3 days of drug administration. 
c Sodium, potassium, chloride, bicarbonate, calcium, magnesium, phosphorus, BUN, creatinine, glucose, albumin, total protein, alkaline phosphatase, total 
bilirubin, SGOT, SGPT, LDH, and uric acid with creatinine clearance to be calculated from these results at screening; CBC with differential and platelet count; 
these evaluations will occur daily for the first 3 days of drug administration. 
d CBC with differential and platelet count will be repeated if subject experiences a hematological toxicity that could be classified as a DLT after the first dose of 
the study drug. 
e Pregnancy test for women of childbearing potential within 72 hours prior to first dose of study drug and on Day 7 and Day 21 
f Within 2 weeks prior to the first dose of study drug 
g A final MUGA or ECHO scan or an ECHO strain evaluation will be conducted between the Day 18 and Day 28 visits or at the the End of Treatment visit if the 
subject is prematurely terminated. 
h For peripheral blood, include CBC, differential, and platelet count.  In addition, flow cytometry analysis of peripheral blood may be utilized for measuring 
disease (optional).  Bone marrow aspirate is sufficient if there are spicules present; if not, bone marrow biopsy is required (designated aspirate/biopsy below) 
i CBC, blood chemistry, urinalysis, and ECGs should be repeated if screening was ≥14 days prior to first dose of study drug. 
j Bone marrow aspirate and peripheral blood (complete blood count, differential, and platelet count; flow cytometry is optional) within 15 days prior to the first 
dose of liposomal annamycin to document disease status.  Bone marrow aspirate/biopsy 15 to 35 days after the start of therapy will be performed. If cytogenetics 
were initially abnormal, include cytogenetics as part of the remission documentation. 
k Blood samples for PK analysis collected at predose and at 0.25, 0.5, 1, 2, 4, 8, and 24 hours if clinically feasible and within a reasonable timeframe of the 
indicated times after the start of liposomal annamycin infusion on Day 1 and Day 3 for 3 subjects at each dose and 6 subjects at the RP2D who complete the 
Blood samples for 
pharmacokinetic 
analysis 
Xk
Xk
Xl 
Xk,l
X
Blood samples for 
cardiac biomarkers 
Xm 
Xm 
X 
Study drug 
administration 
Xn
Xn
Xn
Prophylactic 
mouthwash 
Xo
Xo
Xo
Antihistamine 
administration p 
X 
X 
X 
Adverse event 
monitoring 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Concomitant 
medication 
X 
X 
X 
X 
X 
X 
X 
X 
X 


Protocol No. MB-104 
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3 days of dosing. ECGs will also be collected for all subjects involved in pharmacokinetic analyses at predose and at 4 and 24 hours after dosing on Days 1 to 3, 
except for the 6 subjects at the RP2D who will have ECGs on Days 1 and 3 only.  Documentation of concomitant medications affecting CYP enzymes will be 
obtained to ensure that these are a consideration in the evaluation of the pharmacokinetics of this drug. 
l PK/ECG collection timepoint of 24 hours after the start of the infusion on Days 1 and 3, per above, will occur on Day 2 (predose) and Day 4.   
m Blood sample for cardiac biomarkers (troponin-I and troponin-T) will be collected at predose and at the end of the infusion on Day 3. 
n Liposomal annamycin is administered as a daily 2-hour intravenous infusion for 3 consecutive days followed by 18 days off liposomal annamycin (i.e., 
1 treatment cycle = 21 days) every 3 weeks.  Subjects will be hospitalized for the full 3 days of liposomal annamycin administration. 
o Mouthwash per institution’s guidelines and practices
p Antiallergic premedication with diphenhydramine at a dose of 50 mg administered intravenously must be administered before each dose of liposomal 
annamycin. 
q A MUGA scan, ECHO scan, or ECHO strain will be collected at the Q3-month survival timepoint if possible. 


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APPENDIX B: EASTERN COOPERATIVE ONCOLOGY GROUP PERFORMANCE 
STATUS SCALE 
 
Grade 
Details 
0 
Fully active, able to carry on all predisease performance without restriction  
1 
Restricted in physically strenuous activity but ambulatory and able to carry out work of a 
light of sedentary nature, e.g., light house work, office work 
2 
Ambulatory and capable of all self care but unable to carry out any work activities.  Up 
and about more than 50% of waking hours 
3 
Capable of only limited selfcare, confined to bed or chair more than 50% of waking 
hours 
4 
Completely disabled.  Cannot carry on any self care.  Totally confined to bed or chair. 
5 
Dead 
 


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CONFIDENTIAL 
APPENDIX C: COMMONLY USED CHEMOTHERAPY REGIMENS AND THEIR 
ANTHRACYCLINE CONTENT  
 
Regimen 
Anthracycline 
Dose per course 
(mg/m2) 
ABVD 
Doxorubicin 
50 
AC 
Doxorubicin 
60 
Baseline BEACOPP 
Doxorubicin 
25 
CHOP (R_CHOP) 
Doxorubicin 
50 
Cisplatin/Eirubicin 
Epirubicin 
90 
CODOX-M 
Doxorubicin 
40 
D(90)A 3+7 
Daunorubicin 
270 
D(60)A 3+7 
Daunorubicin 
180 
DA 3+ 
Daunorubicin 
150 
DA 2+5 
Daunorubicin 
100 
DAE 
Daunorubicin 
150 
Dclo 
Daunorubicin 
150 
Esc BEACOPP 
Doxorubicin 
35 
EC90 
Epirubicin 
90 
Epi-CEM 
Epirubicin 
150 
FEC75 
Epirubicin 
75 
FEC100 
Epirubicin 
100 
FLA(G)-Ida 
Idarubicin (IV) 
24 
FMD 
Mitoxantrone 
12 
Hyper-CVAD 
Doxorubicin 
50 
Ida-Ara-c 
Idarubicin (IV) 
30 
IDARAM 
Idarubicin (IV) 
20 
IVE 
Epirubicin 
50 
MAXI-CHOP 
Doxorubicin 
75 
MCD 
Mitoxantrone 
12 
MiDAC 
Mitoxantrone 
50 
PAD 
Doxorubicin 
36 
Stanford V 
Doxorubicin 
25 
VAD 
Doxorubicin 
36 
Z-DEX 
Idarubicin (PO) 
40 
Vyxeos 
Daunorubicin 
132 
Abbreviations: IV = intravenous; PO = per os (orally). 
 
 
 


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CONFIDENTIAL 
 
APPENDIX D: DEFINITION OF HY’S LAW  
 
• The drug causes hepatocellular injury, generally defined as an elevated alanine 
aminotransferase or aspartate aminotransferase by 3-fold or greater above the upper 
limit of normal. 
• Among subjects showing such aminotransferase elevations, they also have elevation of 
their serum total bilirubin of greater than 2 × the upper limit of normal, without 
findings of cholestasis (defined as serum alkaline phosphatase activity less than 
2 × the upper limit of normal).  
• No other reason can be found to explain the combination of increased 
aminotransferase and serum total bilirubin, such as viral hepatitis, alcohol abuse, 
ischemia, preexisting liver disease, or another drug capable of causing the observed 
injury.   
 
 
 
Reference: Guidance for Industry Drug-Induced Liver Injury:  Premarketing Clinical Evaluation  
[Internet]. Rockville: U.S. Department of Health and Human Services Food and Drug 
Administration Center for Drug Evaluation and Research (CDER) Center for Biologics 
Evaluation and Research (CBER); c2009 [cited 2019 Feb 17] Available from: 
https://www.fda.gov/downloads/Drugs/.../Guidances/UCM174090.pdf 
 
 
 


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CONFIDENTIAL 
APPENDIX E: CALCULATIONS OF ANTHRACYCLINE EXPOSURE  
 
Cardioprotective Measures for Anthracycline Drugs (adapted from Zamorano et al. 2016) 
 
Potential Cardioprotective Measures for Anthracycline Drugs and Analogues by 
Limitation of Cumulative Doses 
Drug 
Conversion Factor (Daunorubicin Equivalents) 
Daunorubicin 
1 
Doxorubicin 
2.2 
Epirubicin 
1.1 
Mitoxantrone 
5.0 
Idarubicin 
5.3 
Liposomal Annamycin 
1 
 
All daunorubicin equivalents are based on <551 mg/m2 as being the maximal allowable dose 
with the maximal exposure of anthracyclines capped at 550.9 mg/m2. This will be calculated as a 
daunorubicin equivalent added to the anticipated exposure to liposomal annamycin.   The 
calculations and results will be entered into the appropriate eCRF page. 
 
Additional References: 
 
Alderton PM, Gross J, Green MD. Comparative study of doxorubicin, mitoxantrone, and 
epirubicin in combination with ICRF-187 (ADR-529) in a chronic cardiotoxicity animal model. 
Cancer Res. 1992;52(1):194-201.  
 
Anderlini P, Benjamin RS, Wong FC, Kantarjian HM, Andreeff M, Kornblau SM, et al. 
Idarubicin cardiotoxicity: a retrospective study in acute myeloid leukemia and myelodysplasia. J 
Clin Oncol. 1995;13(11):2827-34. 
 
Avilés A, Neri N, Nambo JM, Huerta-Guzman J, Talavera A, Cleto S. Late cardiac toxicity 
secondary to treatment in Hodgkin's disease. A study comparing doxorubicin, epirubicin and 
mitoxantrone in combined therapy. Leuk Lymphoma. 2005;46(7):1023-8.  
 
Feijen EA Leisenring WM, Stratton KL, Ness KK, van der Pal HJ, Caron HN, et al. Equivalence 
ratio for daunorubicin to doxorubicin in relation to late heart failure in survivors of childhood 
cancer. J Clin Oncol. 2015;33(32):3774-80. 
 
Lefrak EA, Pitha J, Rosenheim S, Gottlieb JA. A clinicopathologic analysis of adriamycin 
cardiotoxicity. Cancer. 1973;32(2):302-14. 
 
McGowan JV, Chung R, Maulik A, Piotrowska I, Malcolm Walker J, Yellon DM. Anthracycline 
chemotherapy and cardiotoxicity. Cardiovasc Drugs Ther. 2017;31:63-75. 
 
Ryberg M, Nielsen D, Skovsgaard T, Hansen J, Jensen BV, Dombernowsky P. Epirubicin 
cardiotoxicity: an analysis of 469 patients with metastatic breast cancer. J Clin Oncol. 
1998;16(11): 3502-8. 
 


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Takemura G, Fujiwara H. Doxorubicin-induced cardiomyopathy from the cardiotoxic 
mechanisms to management. Prog Cardiovasc Dis. 2007;49:330-52. 
 
Von Hoff DD, Rozencweig M, Layard M, Slavik M, Muggia FM. Daunomycin-induced 
cardiotoxicity in children and adults a review of 110 cases. Amer J Med. 1977;62:200-8.