2015 05-21 TCRabHaplo2010 PRT v7.0 FPU

Miltenyi Biotec GmbH Clinical Study Protocol
M2011-238; EudraCT No. 2011-005562-38 TCRalpha/beta-Haplo2010
CLINICAL TRIAL PROTOCOL

A multi-center phase I/II safety and feasibility study using
CliniMACS TCRα/β and CD19 depleted stem cell grafts from
haploidentical donors for haematopoietic progenitor cell
transplantation in children and adults
TCRalpha/beta-Haplo2010
EUDRACT No.: 2011-005562-38
Sponsor Study No.: M2011-238
Study drug: Stem cell graft from haploidentical donors depleted of
+ +
TCRα/β and CD19 cells using the CliniMACS TCRα/β
and CD19 Systems
Development phase: I/II
Sponsor: Miltenyi Biotec GmbH
Friedrich-Ebert-Straße 68
51429 Bergisch Gladbach
Germany
Medical Director of the Sponsor Liane Preußner, MD
Miltenyi Biotec GmbH (address see above)
Phone: +49 (0)2204 8306-3950
Fax: +49 (0)2204 8306-6699
E-mail: lianep@miltenyibiotec.de
Trial Management of the Sponsor Sandra Karitzky, PhD
Miltenyi Biotec GmbH (address see above)
Phone: +49 (0)2204 8306-6560
Fax: +49 (0)2204 8306-6699
E-mail: sandrak@miltenyibiotec.de
Coordinating Investigator Prof. Rupert Handgretinger, MD
University Children’s Hospital University Clinic Tübingen
Hoppe-Seyler-Str. 1
72076 Tübingen, Germany
Phone: +49 (0)7071 298-4744
Fax: +49 (0)7071 29-3966
E-mail: rupert.handgretinger@med.uni-tuebingen.de
National Coordinating Investigator (‘Leiter Prof. Rupert Handgretinger, MD
der Klinischen Prüfung’, LKP)
Leading Investigator (adult patients) Prof. Wolfgang Bethge, MD, PhD
University Clinic Tübingen II (Department Hematology,
Oncology, Immunology and Rheumatology)
Leading Investigator (children) Prof. Peter Lang, MD, PhD
University Children’s Hospital University Clinic Tübingen
(Department 1)
Document status: Final Version 7.0
Elaboration date: 21. May 2015
Amendments: 1.0-Final Version 1.0, 03. December 2013
No. 2 Final Version 1.0, 10 March 2014
No. 3 Final Version 1.0, 12. September 2014
No. 4 Final Version 1.0, 21. May 2015
The study will be conducted according to the protocol and in compliance with Good Clinical Practice (GCP), with
the Declaration of Helsinki, and with other applicable regulatory requirements.
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Confidential
This document contains confidential information of Miltenyi Biotec GmbH.
Do not copy or distribute without written permission from the Sponsor.
A multi-center phase I/II safety and feasibility study using CliniMACS TCRα/β
and CD19 depleted stem cell grafts from haploidentical donors for
haematopoietic progenitor cell transplantation in children and adults
Signatures
Declaration of Sponsor
This study protocol was subjected to critical review. The information it contains is
consistent with current knowledge of the risks and benefits of the investigational
product, as well as with the moral, ethical and scientific principles governing clinical
research as set out in the actual Declaration of Helsinki and the guidelines on Good
Clinical Practice.
Liane Preußner, MD (Signature) (Date)

(Medical Director Sponsor)
Declaration of the Coordinating Investigator (incl. National Coordinating

Investigator in Germany (‘LKP’))
This study protocol was subjected to critical review and has been approved by the
Sponsor. The information it contains is consistent with the current risk/benefit
evaluation of the investigational product as well as with the moral, ethical and
scientific principles governing clinical research as set out in the actual Declaration of
Helsinki and the guidelines on Good Clinical Practice.
Prof. R. Handgretinger, MD (Signature) (Date)

(Coordinating Investigator, incl.
National Coordinating Investigator in Germany (‘LKP’))
Leading Investigator:
Prof. P. Lang, MD (Signature) (Date)

(Leading Investigator Paediatric Patients)
Leading Investigator:
Prof. W. Bethge, MD (Signature) (Date)

(Leading Investigator Adult Patients)
Biometrician:
(Date)
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A multi-center phase I/II safety and feasibility study using

CliniMACS TCRα/β and CD19 depleted stem cell grafts from
haploidentical donors for haematopoietic progenitor cell
transplantation in children and adults
Signatures
Declaration of Study Site Principal Investigator and Deputy
All documentation for this study that is supplied to me and that has not been
previously published will be kept in the strictest confidence. This documentation
includes this study protocol, Investigator’s Brochure, equipment for electronic data
documentation, and other scientific data.
I have read this study protocol and agree that it contains all the information required
to conduct the study. I agree to conduct the study as set out in this protocol. I will not
enrol any patients without the prior written approval of a properly constituted
Independent Ethics Committee (IEC). l agree to obtain, in the manner described in
the study protocol written informed consent to participate from all patients enrolled in
this study and to keep the consent forms for 15 years. I will provide a curriculum vitae
before the study starts. No changes will be made to the study protocol without the
prior written approval of the Sponsor and the IEC, except where necessary to
eliminate an immediate hazard to the patients.
Name (Investigator) (Signature) (Place, Date)

Institution
Name (Deputy) (Signature) (Place, Date)

Institution
Name (Deputy) (Signature) (Place, Date)

Institution
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TABLE OF CONTENTS
Signatures................................................................................................................................. 2
Contact Addresses and Responsibilities ..................................................................................... 8
List of Abbreviations and Definitions of Terms ........................................................................... 9
1. PROTOCOL SYNOPSIS ............................................................................................. 11
2. BACKGROUND AND RATIONALE............................................................................. 19
2.1. Study Rationale............................................................................................................. 23
2.2. Risk-Benefit Assessment ............................................................................................... 24
2.2.1. Potential study specific benefits for recipients ............................................................................. 24
2.2.1.1. Low GVHD rates ........................................................................................................................ 24
2.2.1.2. Expedited immune reconstitution ............................................................................................ 25
2.2.1.3. Reduced rate of infections ........................................................................................................ 25
2.2.1.4. Lower numbers of CD34 positive stem cells ............................................................................. 25
2.2.1.5. Prevention of PTLD .................................................................................................................... 25
2.2.1.6. Reduction of immunosuppression post transplantation........................................................... 25
2.2.2. Potential study specific risks for recipients ................................................................................... 25
2.2.2.1. Graft-versus-host Disease ......................................................................................................... 25
2.2.2.2. Lymphoproliferative Syndrome (LPS, PTLD) .............................................................................. 26
2.2.2.3. Potential Sensitization to Murine Proteins ................................................................................ 26
2.2.3. Discussion of the Risk/Benefit Assessment and Conclusion ......................................................... 26
2.2.4. Major risks of allogeneic hematopoetic stem cell transplantation independent of the IMP ....... 27
2.2.4.1. PBSC Infusion............................................................................................................................. 27
2.2.4.2. Infections ................................................................................................................................... 27
2.2.4.3. Graft Failure / Poor Marrow Function....................................................................................... 27
2.2.4.4. Graft-versus-host Disease ......................................................................................................... 28
2.2.4.5. Veno-occlusive Disease (VOD) of the Liver ............................................................................... 28
2.2.4.6. Death ......................................................................................................................................... 28
2.2.4.7. Therapy Toxicities...................................................................................................................... 28
3. STUDY OBJECTIVES AND ENDPOINTS...................................................................... 29

3.1. Objectives..................................................................................................................... 29
3.2. Study hypothesis .......................................................................................................... 31
4. STUDY DESIGN ....................................................................................................... 32
4.1. Study Overview ............................................................................................................ 32
4.2. Rationale for Study Design ............................................................................................ 32
4.3. Study Population .......................................................................................................... 33
4.3.1. Selection Criteria ........................................................................................................................... 34
4.3.2. Withdrawal and Replacement....................................................................................................... 37
4.3.2.1. Patient Withdrawal and Replacement ...................................................................................... 37
4.3.3. Patient Identification and Randomization..................................................................................... 38
4.3.4. Protocol Violations ........................................................................................................................ 38
4.3.5. Premature Termination of the Study ............................................................................................. 38
4.4. Study Treatment ........................................................................................................... 39
4.4.1. Preparation of the Hematopoietic Stem Cell Graft (IMP) .............................................................. 39
4.4.2. Packaging, Labeling and Storage .................................................................................................. 40
4.4.3. Transport of Investigational Product............................................................................................. 41
4.4.4. Administration of Investigational Product .................................................................................... 41
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4.5. Compliance ................................................................................................................... 41

4.6. Graft and Device Accountability .................................................................................... 41
5. STUDY PROCEDURES .............................................................................................. 42
5.1. Study Treatment Plan ................................................................................................... 42
5.1.1. Mobilization and Collection of Donor PBSC................................................................. 42
5.1.2. Conditioning Regimen for Patients ............................................................................. 43
5.1.3. PBSC Transplantation ................................................................................................ 44
5.1.4. Prophylaxis, Supportive Care and Concomitant Treatments ........................................ 45
5.2. Study Assessments........................................................................................................ 50
5.2.1. Evaluation of Grafts ...................................................................................................................... 50
5.2.2. Evaluation of Donors ..................................................................................................................... 50
5.2.3. Evaluation of Patients ................................................................................................................... 52
5.2.4. Adverse Events .............................................................................................................................. 57
5.2.5. Concomitant medication ............................................................................................................... 58
5.2.6. Outcome Parameters .................................................................................................................... 58
5.3. Study Visits Schedule .................................................................................................... 60
5.3.1. Donors’ visit schedule .................................................................................................................... 61
5.3.2. Patients’ visits schedule................................................................................................................. 61
5.3.3. Donors’ Assessments ..................................................................................................................... 62
5.3.4. Patients’ Assessments by Visit ....................................................................................................... 62
5.4. Unscheduled visits ........................................................................................................ 72
5.5. Early Termination Visit (ETV) ......................................................................................... 72
6. ADVERSE EVENTS................................................................................................... 73
6.1. Definitions .................................................................................................................... 73
6.1.1. Adverse event (AE) ........................................................................................................................ 73
6.1.2. Adverse Reaction (AR) ................................................................................................................... 74
6.1.3. Serious Adverse Events (SAEs) ....................................................................................................... 74
6.1.4. Serious Adverse Reactions (SARs) and Suspected Unexpected Serious Adverse Reactions (SUSARs)
75
6.2. Assessment of Adverse Events/Therapy-Related Toxicities............................................. 75
6.2.1. Severity .......................................................................................................................................... 75
6.2.2. Causality ........................................................................................................................................ 75
6.3. Monitoring, Recording and Reporting of Adverse Events ................................................ 76
6.3.1. General Requirements ................................................................................................................... 76
6.3.2. Reporting of Serious Adverse Events ............................................................................................. 77
6.3.3. Other Procedures for Recording and Reporting Adverse Events ................................................... 78
6.4. Adverse Events of Specific Interest ................................................................................ 78
6.5. Follow-Up of Adverse Events ......................................................................................... 78
6.6. Therapy Toxicities of Conditioning ................................................................................. 79
6.7. Unblinding of treatment / Emergency identification ...................................................... 79
7. STATISTICS............................................................................................................. 79
7.1. General Considerations ................................................................................................. 79
7.2. Determination of Sample Size ....................................................................................... 80
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7.3. Analysis Sets ................................................................................................................. 80

7.3.1. Definitions ..................................................................................................................................... 80
7.3.2. Eligibility, Protocol Deviations ....................................................................................................... 80
7.4. Graft Evaluation ............................................................................................................ 80
7.5. Demographics and Baseline Characteristics ................................................................... 81
7.6. Treatment and Study Compliance .................................................................................. 81
7.7. Analyses of the Primary Endpoint .................................................................................. 81
7.8. Analyses of the Secondary Endpoints ............................................................................ 81
7.9. Safety Analyses ............................................................................................................. 82
7.9.1. Adverse Events .............................................................................................................................. 82
7.9.2. Laboratory ..................................................................................................................................... 82
7.9.3. Physical Examination..................................................................................................................... 83
7.9.4. Vital Signs ...................................................................................................................................... 83
7.10. Safety Monitoring of Key Safety Endpoints ................................................................. 83
7.10.1. Statistical Stopping Guidelines ...................................................................................................... 83
7.11. Interim Analysis......................................................................................................... 85
7.12. Data Handling............................................................................................................ 85
8. ETHICAL ASPECTS................................................................................................... 85
8.1. Independent Ethics Committee Approval ....................................................................... 85
8.2. Informed Consent ......................................................................................................... 86
8.3. Data confidentiality ...................................................................................................... 87
8.4. Liability and Insurance .................................................................................................. 87
9. ADMINISTRATIVE PROCEDURES ............................................................................. 87
9.1. Regulatory aspects ........................................................................................................ 87
9.2. Protocol Approval and Amendment............................................................................... 87
9.3. Duration of the Study .................................................................................................... 88
9.4. Data Safety Monitoring Board (DSMB)........................................................................... 88
9.5. Other Ethical and Regulatory Issues ............................................................................... 89
9.6. Data Quality Assurance ................................................................................................. 89
9.7. Case Report Forms and Source Documentation.............................................................. 89
9.8. Access to Source Data ................................................................................................... 90
9.9. Data Processing ............................................................................................................ 90
9.10. Archiving Study Records ............................................................................................ 90
9.11. Publication Policy ...................................................................................................... 90
10. Reference List ........................................................................................................ 92
A. Appendices ............................................................................................................ 97
Appendix 1: Glucksberg Criteria for grading of acute GVHD ...................................................... 98
Appendix 2: Grading of Chronic GvHD...................................................................................... 99
Appendix 3: Karnofsky / Lansky Performance Indeces .............................................................103
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Appendix 4: Sorror Comorbidity Index [72] .............................................................................104

Appendix 5: List of Study Personnel ........................................................................................105
Appendix 6: List of Clinical Study Sites and Principal Investigators ...........................................107
Appendix 7: List of Manufacturing Sites of the IMP / GMP Laboratories ..................................109
Appendix 8: List of Central Laboratories..................................................................................110
Appendix 9: Independent Physician (NL) .................................................................................111
LIST OF TABLES
Table 1: Incidences of oncological and non-malignant diseases considered in this study ................. 33
Table 2: Conditioning regimen with the use of ATG Fresenius............................................................... 44
Table 3: Conditioning regimen with the use of TNI ................................................................................... 44
Table 4: Blood and bone marrow sampling for standard of care routine monitoring, performed at the
local laboratories of the hospital....................................................................................................................... 56
Table 5: Blood and bone marrow sampling for additional assessments: see section 5.2.3.2, X. ...... 57
Table 6: Evaluations and assessments of patients: flow-chart ............................................................... 70
Table 7: Interim safety monitoring stopping boundaries for aGVHD grade III–IV at Day 100 post-
transplantation .................................................................................................................................................... 84
Table 8: Interim safety monitoring stopping boundaries for NRM at Day 100 post-transplantation .. 84
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Contact Addresses and Responsibilities
List of Sponsors Study Personnel: see Appendix 5
Coordinating Investigator: see Appendix 5
National Coordinating Investigator: see Appendix 5
Safety Monitoring Board: see Appendix 5
Statistician: see Appendix 5
Project Management: see Appendix 5
Monitoring: see Appendix 5
Data Management: see Appendix 5
List of Clinical Study Sites and Principal see Appendix 6

Investigators:
List of Manufacturing Sites of IMP: see Appendix 7
List of Laboratories: see Appendix 8
Independent Physician (NL): see Appendix 9
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List of Abbreviations and Definitions of Terms

ADV Adenovirus
AE Adverse event
aGVHD Acute graft-versus-host disease
ALAT Alanine aminotransaminase (=GPT, glutamic pyruvic transaminase)
ALL Acute lymphoblastic leukemia
AML Acute myeloic leukemia
AMG Arzneimittelgesetz (German Medicinal Products Law)
ANC Absolute neutrophil count
AP Alkalic phosphatase
AR Adverse reaction
ASAT Aspartate aminotransferase (=GOT, glutamic oxaloacetic transaminase)
ATG Anti-thymocyte globulin
ATIII Antithrombin III
BID Bis in die (twice daily)
BM Bone marrow
BfArM Federal Institute for Drugs and Medical Devices
(Bundesinstitut für Arzneimittel und Medizinprodukte)
BS Body surface
BW Body weight
CBC Complete blood count
cGVHD Chronic graft-versus-host disease
CK Creatine phosphokinase
CML Chronic myeloid leukemia
CMV Cytomegalovirus
CNS Central nervous system
CR Complete remission
CRF Case report form
CRP C-reactive protein
CSF Colony stimulating factor
CTC Common toxicity criteria (for adverse events)
DFS Disease-free survival
DLCO Diffusion capacity of carbon dioxide
DLI Donor lymphocyte infusion
DNA Deoxyribonucleic acid
DSMB Data Safety Monitoring Board
EBMT European Group for Blood and Marrow Transplantation
EBV Epstein-Barr virus
ECHO Echocardiography
EDTA Ethylenediaminetetraacetic acid
EFS Event-free survival
ESR Erythrocyte sedimentation rate
FACS Fluorescence activated cell sorting
G-CSF Granulocyte colony stimulating factor
GGT Gamma-glutamyl transpeptidase
GVT Graft-versus-tumor
GVHD Graft-versus-host disease
HbsAG Hepatitis B surface antigen
HCG Human chorionic gonadotropin
HCT Hematopoietic cell transplantation
HCV Hepatitis C virus
HHCT Haploidentical allogeneic hematopoietic cell transplantation
HHV-6 Human herpes virus 6
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HIV Human immunodeficiency virus

HLA Human leucocyte antigen
HLH Haemaphagocytic lymphohistiocytosis
HSV Herpes simplex virus
HSCT Hematopoietic stem cell transplantation
HTLV I Human T-cell leukemia virus
ICH-GCP Guideline for Good Clinical Practice of the International Conference on
Harmonization
ICSR Individual Case Safety Report
IEC Independent ethics committee
IMP Investigational medicinal product
IMPD IMP dossier
INR International normalized ratio
IRB Institutional review board
KIR Killer cell immunoglobulin-like receptor
LDH Lactate dehydrogenase
LKP Leiter der klinischen Prüfung (‚National Coordinating Investigator, according
to German drug law [AMG])
MDS Myelodysplastic syndrome
MM Multiple myeloma
MMF Mycophenolate mofetil
MPS Myeloproliferative syndrome
MRD Minimal residual disease
NK Natural killer (cell)
NL The Netherlands
NHL Non-Hodgkin lymphoma
NRM Non-relapse mortality
OS Overall survival
PB Peripheral blood
PBMC Peripheral blood mononuclear cells
PBSC Peripheral blood stem cell
PBSCT Peripheral blood stem cell transplantation
PCR Polymerase chain reaction
PEI Paul-Ehrlich Institute
PTLD Post-transplant lymphoproliferative disease
PTT Partial prothrombin time
RIC Reduced intensity conditioning
RNA Ribonucleic acid
SAE Serious adverse event
SAR Serious Adverse Reaction
SF Shortening fraction
SUSAR Suspected Unexpected Serious Adverse Reaction
TBI Total body irradiation
T cells Thymus-derived withe blood cells
TCR T-cell receptor
TNI Total nodal irradiation
TNF Tumor necrosis factor
TREC T-cell receptor excision circle: circular, stable extrachromosomal DNA
fragment generated during T-cell receptor diversification
Treg T regulatory cell
TSH Thyreoidea stimulating hormone
VZV Varicella zoster virus
WBMV Wet op bijzondere medische verrichtingen
WMO Wet medisch-wetenschappelijk onderzoek met mensen (Medical Research
Involving Human Subjects)
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1. PROTOCOL SYNOPSIS
Title: A multi-center phase I/II safety and feasibility study using CliniMACS TCRα/β
and CD19 depleted stem cell grafts from haploidentical donors for
haematopoietic progenitor cell transplantation in children and adults
Short title: TCRalpha/beta-Haplo2010
EudraCT No.: 2011-005562-38
Sponsor study no.: M2011-238
Phase: I/II
Study design: Multi-center, open-label, single-arm clinical trial
Study centers: 13 in total, 7 centers for adult patients, 6 pediatric centers
Number of subjects: 60 in total, 30 adult, 30 pediatric
Coordinating Investigator: Prof. Rupert Handgretinger, MD, Tübingen
Lead investigator paediatric patients: Prof. Peter Lang, MD, Tübingen
Lead investigator adult patients: Prof. Wolfgang Bethge, MD, Tübingen
Data Safety Monitoring Board: Consisting of at least three experts in the field
Planned duration of study:
• First visit of first patient: Q4 2012
• First visit of last pediatric patient: Q4 2015
• First visit of last adult patient: Q3 2016
• Last visit of last pediatric patient: Q4 2017
• Last visit of last adult patient: Q3 2018
• Final study report: Q1 2019
The planned duration of the study for each patient will be approximately 2 years with
primary endpoint reached 3 months after transplantation (visit XIV) and two follow-up
phases: phase I until 1 year after transplantation (visits XV – XVII) and phase II until
2 years after transplantation (visit XVIII).
The total duration of the study will be about 5.75 years.
Drug Substance:
Mobilized peripheral blood stem cells from allogeneic donors depleted of
TCRα/β+ and CD19+ cells using the CliniMACS TCRα/β-Biotin and CD19
Systems
Drug Products (IMP):
• TCRabCD19PBSC
• TCRabCD19PBSC_cryo
Targeted Cellular Composition:
• Viable CD34+CD45+ cells
Target cell number ≥4 × 106/kg BW, percentage of viable cells ≥95%
• TCRαβ+ cells
Target cell number ≤25 × 103/kg BW
Note: It is not permitted to exceed the indicated target cell number of
25 × 103 TCRαβ+ cells/kg BW unless necessary to reach the target cell
number of ≥4 × 106 CD34+CD45+ cells/kg BW. A maximum cell number of
1 × 105 TCRαβ+ cells/kg BW must not be exceeded.
• CD20+ cells
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Targeted Cell number ≤1 × 105/kg BW

Additional IMP specifications concern TCRγ/δ+, CD3+ and CD45+/WBC counts,
CD45+ vitality, haematocrit and result of visual control. Sterility will be tested but is
not relevant for release.
Route of administration: IV
Comparator: Not applicable
Patient population:
Patients suffering from hematological and non-hematological malignancies, and non-
malignant diseases, requiring and eligible for allogeneic stem cell transplantation.
Inclusion criteria—indications:
• Male or female without childbearing potential (e.g. prepubertal child, postmeno-
pausal, surgically sterile) or using medically adequate contraception
• Adult and paediatric patients with hematological malignancies in complete
remission (CR), partial remission (PR) or with stable disease
- Acute myeloid leukemia (AML):
Patients with high-risk AML in CR1
Patients with relapsed or primary therapy-refractory AML
- Acute lymphoid leukemia (ALL):
Patients with high-risk ALL in CR1
Patients with relapsed or primary refractory ALL
- Hodgkin’s disease: Patients with relapsed or primary refractory Hodgkin’s
disease
- Non-Hodgkin’s lymphoma: Patients with relapsed or primary refractory Non-
Hodgkin’s lymphoma
- Myelodysplastic Syndrome (MDS)/ Myeloproliferative Syndrome (MPS):
Patients with refractory MDS/MPS
- Multiple myeloma (MM): Patients with relapsed or refractory multiple myeloma
• Adult and paediatric patients with non-hematological malignancies without curative
treatment option, mainly
- Neuroblastoma: Patients with relapsed metastatic nmyc-negative or -positive or
local nmyc-positive neuroblastoma
- Soft tissue sarcoma:
relapsed metastatic soft tissue sarcoma (rhabdomyosarcoma, Ewing
sarcoma, peripheral neuroectodermal tumor)
soft tissue sarcoma with primary bone metastases or bone involvement in
patients older than 10 years
• Adult and paediatric patients with the following non-malignant diseases with HSCT
as curative treatment option
Hematologic diseases, acquired and congenital
Severe aplastic anemia (patients not responding to immune suppression)
Paroxysmal nocturnal haemoglobinuria (PNH)
Haemaphagocytic lymphohistiocytosis (HLH)
Congenital immunodeficiencies
Severe combined immune deficiency (SCID) and related diseases
Chediak Higashi syndrome
Congenital metabolic disorders
Malignant osteopetrosis
Lysosomal storage disorders (mucopolysacharidoses, leukodystrophies,
glycoprotein disorders)
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Additional patient inclusion criteria:

• No HLA-identical stem cell donor available, as determined by high-resolution
typing (maximum of 1 antigen or allelic mismatch are acceptable [9/10 match]), but
eligible haploidentical donor with >1 antigenic or allelic mismatch (9/10-match)
identified and at call; Exception: Haplo-identical HSCT is medically indicated even
if an HLA-identical donor is available and decision for haplo-identical HSCT has
been made according to hospital routine prior to inclusion of the patient into this
study.
• Patients aged ≥8 weeks to ≤65 years
• Karnofsky (patients >16 years)/Lansky (patients ≤16 years) index >60%
• Patient in good clinical condition without concomitant diseases significantly
increasing the risk of transplantation, see exclusion criteria
• Adult patients without active infections at the time of transplantation
• Pediatric patients without uncontrollable, progressive infections at the time of
transplantation
• Informed consent given (patient or legal representative).
Main exclusion criteria for patients:
• Age >65 years or <8 weeks
• Patients with progressive disease prior HCT
• <3 months after preceding hematopoietic cell transplantation (HCT)
• History of neurological impairment (active seizures, severe peripheral neuropathy,
signs of leukencephalopathy, active CNS infection)
Note: For patients with HLH or Malignant Osteopetrosis or other patients with
heavy pretreatment with irradiation or intrathecal chemotherapy pre-transplant
CNS MRI and neurological consultation are mandatory.
• Fungal infections with radiological and clinical progression
• Liver function abnormalities with bilirubin >2 mg/dL and elevation of transaminases
higher than 400 U/L
• Chronic active viral hepatitis
• Adult patients: ejection fraction <40% on echocardiography; pediatric patients:
ejection fraction <40% or shortening fraction <25% on echocardiography
• Patients with > grade II hypertension by Common Toxicity Criteria (CTC)
• Creatinine clearance below threshold defined for stem cell transplantation
according to local clinical standard
• Respiratory failure necessitating supplemental oxygen
• HIV infection
• Female patients who are pregnant or breast feeding, or adults of reproductive
potential not willing to use an effective method of birth control during study
treatment and for at least 12 months thereafter
Note: Women of childbearing potential must have a negative serum pregnancy
test at study entry.
• Concurrent severe or uncontrolled medical disease (e.g. uncontrolled diabetes,
congestive heart failure, myocardial infarction within 6 months prior to the study,
unstable and uncontrolled hypertension, chronic renal disease, or active
uncontrolled infection) which by assessment of the treating physician could
compromise participation in the study
• Patients with a history of psychiatric illness or a condition which could interfere
with their ability to understand the requirements of the study (this includes
alcoholism/drug addiction)
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• Patients unwilling or unable to comply with the protocol or unable to give informed
consent
• Treatment with any investigational product within 4 weeks prior to study treatment
(transfusion of the IMP)
Donor selection:
• Haploidentical family member previously identified as eligible donor by
donor/recipient cross-matching including HLA-typing
Note: In case of positive cross-match results for donor-reactive anti HLA
antibodies an alternative donor with negative cross-match results should be
preferred, if available. If no alternative donor with negative cross-match results is
available, removal of anti HLA antibodies is highly recommended to prevent graft
rejection (see 5.1.4.3)
• Donor age ≥16 years
Note: Positive evaluation for allogeneic hematopoietic cell donation has to be
performed at the collection centre according to local standard practice. Informed
consent for mobilization and collection of peripheral blood stem cells according to
local institutional guidelines has to be given in this context independently of the
present clinical study. Stem cell mobilization and collection procedures are not part of
this study and will be performed at the collection centre according to local standard
procedures.
• Study specific informed consent given.
Objectives and Outcome Parameters:
An overview of study objectives and corresponding outcome parameters is shown in
the following table:
Study Objectives Outcome Parameters
Primary
Evaluation of safety/tolerability and Acute graft-versus-host disease grade II–IV defined as GVHD
feasibility of haploidentical PBSC grafts occurring within 100 days after SCT
+ +
depleted of TCRα/β and CD19 cells Severity graded according to Seattle (Glucksberg) criteria
using the CliniMACS TCRα/β/CD19 (Appendix 1):
System in adult and paediatric patients Incidence of GVHD grades II–IV
with hematological and non-hematological Time until occurrence of GVHD grades II–IV
malignancies and specific non-malignant
diseases: Incidence of grade II–IV acute
graft-versus-host disease (GVHD) until
Day 100 post-transplantation
Secondary
Safety outcome parameters
Incidence of grade I acute GVHD until GVHD grade I occurring within 100 days after SCT.
Day 100 post-transplantation Severity graded according to Seattle (Glucksberg) criteria
(Appendix 1)
Incidence of GVHD grade I
Time until occurrence of GVHD grade
Incidence and severity of chronic GVHD Chronic graft-versus-host disease: Incidence/severity
after 1 year and 2 years graded according to standard criteria (Appendix 2)
Incidence of NRM at all visits throughout NRM (non-relapse- mortality): defined as death between
the study day of transplantation (Day 0) and day of assessment, not
due to disease relapse/recurrence.
Incidence and severity of acute infusional Infusional toxicity: maximum toxicity on the days of
toxicities transfusion evaluated by measuring vital signs prior to and
at different times after transfusion
Graft failure from Day 0 to Day 28 Primary graft failure: failure to achieve an absolute ANC
>500/µl at Day +28
Secondary graft failure: initial neutrophil engraftment
followed by a decline in absolute neutrophil count (ANC)
<500/µl and unresponsiveness to growth factor therapy
Feasibility outcome parameters
Final 7.0 14 of 111 21. May 2015

Neutrophil and platelet engraftment from Neutrophil engraftment: cell counts determined by flow
Day 0 to Day 28 cytometry, measurements of ANC ≥500/µL on three
consecutive days and
Time to neutrophil engraftment as time from last stem cell
transplantation to engraftment
Platelet engraftment: cell counts determined by flow
cytometry, measurements of platelet count ≥20,000/µL on
three consecutive days and without platelet transfusion
support for seven days
Time to platelet engraftment as time from last stem cell
transplantation to engraftment
Secondary (continued)
Feasibility outcome parameters (continued)
Overall survival at Day 100 and after 1 Overall survival rate (OS): time from transplantation to
and 2 years death or last follow-up
Disease-free survival at Day 100 and Disease-free survival (DFS): minimum time to relapse/
after 1 and 2 years recurrence, to death or to the last follow-up
Transfusion requirement from Day 0 to Number of thrombocyte infusions after transplantation
Day 100 Time to last thrombocyte infusion from Day 0.
Number of erythrocyte infusions after transplantation
Time to last erythrocyte infusion from Day 0.
Number of infusions of other blood products after
transplantation
Time to last infusion (other blood products) from Day 0.
Incidence of relapse at Day 100 and after Relapse rate: number of patients with relapse at the times
1 and 2 years of assessment
Time to relapse calculated as time of transplantation to
time of relapse
Days of (re)hospitalization at Day 28 and Number of days hospitalized after transplantation
Day 100 assessed at Day 28
Number of days re-hospitalized after primary discharge
assessed at Day 100
Quality of life at baseline, Day 100 and EQ-5D for adults (age ≥18 years), PedsQL for pediatric
after 1 and 2 years patients (age <18 years) and FACT-BMT (age ≥18 years)
at baseline, Day 100 and after 1 and 2 years
Laboratory outcome parameters (routine local assessments)
Donor chimerism Assessed by PCR-analysis of peripheral blood samples
collected on Days 7, 14, 21, 28, 35, 42, 49, 56, 63, 70,
100 and Month 6, 9 and 12 post-transplantation
compared to samples from donor and recipient collected
prior to transplantation.
Assessed by PCR-analysis of bone marrow samples of
patient with haematolocial malignancies collected on
Days 28 and 100 post-transplantation
Laboratory outcome parameters (additional central assessments)
Reconstitution of T, B, NK and T regulatory Immune cell phenotyping of T, B, NK and Treg cell subsets:
+ + + + +
(Treg) cell subsets by immune cell Cell counts of CD3 , CD4 , CD8 , CD3 CD56 ,
CD3 TCRα/β , CD3 TCRγ/δ T cells, naive CD4 TCRα/β ,
+ + + + + +
phenotyping
+ + + +
memory CD4 TCRα/β , naive CD8 TCRα/β , memory
CD8 TCRα/β , DN TCRα/β , TCR Vd2 TCRγ/δ , TCR Vd2
+ + + + + –
TCRγ/δ , naive Tregs, ‘memory’ Tregs, B-cells, monocytes,

+
NK cells , neutrophils, eosinophils;

samples collected on Days 7, 14, 21, 28, 63, 100 and
Month 6 and 12 post-transplantation
Reconstitution of T Vbeta repertoire TCR Vbeta spectratyping (PCR analysis) of T cell
diversification; samples collected on Days 28, 63 and 100
post-transplantation
Reconstitution of T gamma/delta repertoire TCR Vgamma/delta spectratyping (PCR analysis); samples
collected on Days 28, 63 and 100 post-transplantation
Thymic function by TREC Analysis TREC (PCR analysis); samples collected on Days 28, 63
and 100 post-transplantation
KIR genotyping and reconstitution of NK- KIR genotyping of donor and recipient at baseline
cell KIR repertoire KIR repertoire (FACS); samples collected on Days 28, 63
T-cell activity Analysis for CMV-, EBV- and ADV-reactive T cells;
samples collected on Days 28, 63 and 100 post-
Final 7.0 15 of 111 21. May 2015

transplantation
Analysis of T cell stimulation/proliferation; samples
collected on Days 28, 63 and 100 post-transplantation
NK cell activity NK cell activity; samples collected on Days 28, 63 and 100
post-transplantation
Performance outcome parameters GMP-Lab (routine local assessments)
+ +
Performance of the CliniMACS Percentage of viable CD34 CD45 cells recovered after
TCRα/β/CD19 System TCRα/β and CD19 depletion procedure: target value
≥95%
Secondary (continued)
Performance outcome parameters GMP-Lab (routine local assessments)
Performance of the CliniMACS Log Depletion of TCRα/β+ cells
TCRα/β/CD19 System Log Depletion of CD19
+ + +
Cell counts: CD34 CD45 blood stem cells, CD20 B
+ +
cells, CD56 CD16 NK cells, TCRα/β and TCRγ/δ cells,
+ +
CD3 cells and CD45 /WBC cells analysed by flow
cytometry after processing prior to transplantation
+
The percentage of recovered viable CD45 cells after
TCRα/β and CD19 depletion procedure: target value
≥90%
Haematocrit value in graft in mL/mL erythrocytes
Number of grafts with ≥4 × 10 CD34 CD45
6 + +
cells/kg BW
Number of grafts with ≤25 × 10 TCRα/β cells/kg BW
3
Number of grafts with ≤1 × 10 CD20 cells/kg BW

5 +
Sterility of IMP
Safety parameters: adverse events, concomitant medication
Infections: Incidence of CMV, ADV, EBV Recurrence or newly occurring infectious diseases:
and aspergillus, as well as other viral, CMV, ADV, EBV, aspergillus and other at Day 100 and
bacterial and fungal infections 1 year post-transplantation
Infections: Number of reactivations of Number of virus reactivations of CMV, ADV, EBV by
CMV, ADV, EBV by PCR and aspergillus PCR and aspergillus by PCR or serology once to twice
by PCR or serology a week until Day 28, weekly until Day 70 and on
Day 100
Incidence, severity and type of adverse Documentation of serious adverse events throughout
events/serious adverse events the study
Documentation of ‘therapy-related toxicity, known or
unknown’ from Day –12 to Day 0 (during conditioning
and prior to stem cell transplantation)
Documentation of adverse events from Day 0 to
Vital signs and physical examination Vital signs and physical examination including
Karnofsky/Lansky index throughout the study
Safety laboratory (blood count, blood Laboratory values for clinical chemistry and complete
chemistry) blood counts at baseline and from Day 4 to Day 100
Concomitant medication Documentation of concomitant medication from baseline
until Day 100
Documentation of new treatment with cellular products
(erythrocytes, thrombocytes, donor lymphocyte
infusions [DLIs] or antigen-specific T cells) after
Day 100
Enrollment in any other clinical study after Day 100
Study procedures:
Patients will undergo an intensity-reduced conditioning regimen prior to the
intravenous infusion of the IMP. The exact regimen used will depend on the clinical
state of the patient. For all patients GVHD prophylaxis with mycophenolate mofetil
(2 × 20 mg/kg/d) from Day –1 until Day 30 post-transplantation is planned.
Treatment of GVHD will be performed according to the respective institutional
practice guidelines. Patients will have to attend 18 visits in total, starting with the
baseline visit up to the last follow-up visit two years post-transplantation. Donors will
have to attend one visit at the transplantation center.
Final 7.0 16 of 111 21. May 2015

Donors:
• Since stem cell transplantation is the only potentially curative therapeutic option for
the critically ill patients in this study it is planned independently of this trial. All
procedures related to donors (stem cell mobilization and stem cell apheresis) will
therefore be performed according to clinical routine and independent of this study.
The only procedure for donors is the collection of a reference blood sample.
Accordingly the donor has to consent to data transfer and collection and genetic
analysis of one blood sample prior to start of conditioning (Visit II) of the patient.
For information about these requirements and collection of the blood sample the
donor has to visit a study center once.
Preparation of IMP:
• Stem cell apheresis of the donor will be depleted of TCRα/β and CD19 positive
cells using the CliniMACS TCRα/β/CD19 System (consisting of CliniMACS Plus
device and CliniMACS TCRα/β and CliniMACS CD19 reagents). Stem cell
apheresis and subsequent depletion will continue until a post selection target of
≥4 × 106 CD34+CD45+ cells per kg BW of the recipient and ≤25 × 103 TCRα/β+
cells per kg BW of the recipient is reached following at least two but not more than
three stem cell apheresis procedures. No upper limit of CD34+CD45+ progenitor
cells has been defined. It is not permitted to exceed the target cell number of
25 × 103 TCRαβ+ cells/kg BW unless necessary to reach the target cell number of
≥4 × 106 CD34+CD45+ cells/kg BW. However, a maximum cell number of 1 × 105
TCRαβ+/kg BW must not be exceeded.
Active treatment—patients:
• Conditioning for patients with immunodeficiencies and for patients with
hematological and non-hematological malignancies in complete remission
(Minimal Residual Disease [MRD] burden ≤104 or <5% blasts): Fludarabine
(1 × 40 mg/m2/d, Days –8 to –5), Thiotepa (2 × 5 mg/kg BW/d, Day –4), Melphalan
(1 × 70 mg/m2/d, Days –3 to –2) and ATG Fresenius S (1 × 1 mg/kg/d on Day –12,
1 × 9 mg/kg/d on Day –11, 1 × 10 mg/kg/d on Day –10 and 1 × 10 mg/kg/d on
Day -9).
• Conditioning for patients with therapy-refractory hematological malignant disease
(incomplete remission, MRD burden >104 or >5% blasts) and for patients with an
increased risk of graft failure (independent of their state of remission) defined as
patients with non-malignant diseases and chemotherapy naïve patients except of
patients with immunodeficiencies: Either the same dose-reduced conditioning
regimen of Fludarabine, Thiotepa, Melphalan and ATG Fresenius S as patients in
CR or as an alternative option, no ATG Fresenius S, but instead TNI (7 Gy on
Day –1) according to hospital routine. The responsible investigator has to decide
case by case, about the most appropriate approach for each patient.
*Note: Graft failure will be analysed after treatment of the first 10 patients with
incomplete remission and with TNI instead of ATG. If graft failure occurred in <3 of
the 10 patients with this treatment regimen, all patients independent of their state of
remission—complete and incomplete—enrolled afterwards can be transplanted using
ATG Fresenius S or TNI in the conditioning regimen. It is the responsibility of the
investigator to decide case by case, about the most appropriate approach for each
patient.
• Haploidentical stem cell transplantation: 1–3 subsequent intravenous infusions
of haploidentical TCRα/β+ and CD19+ depleted PBSC graft
• GVHD prophylaxis: mycophenolate-mofetil (2 × 20 mg/kg BW/d, Days -1 to +30,
then tapering off)
Final 7.0 17 of 111 21. May 2015

Follow-up for patients:

Assessment of safety/tolerability and feasibility variables at follow-up visits until
Day 365 (±2 weeks) post-transplantation. An additional safety follow-up visit is
planned for Month 24 (±4 weeks), that is, one year after the visit at Day 365.
Reference treatment: Not applicable.
Statistical methods:
The statistical analyses in this study will be exploratory since the study is not
powered to address any pre-defined statements but to generate valid hypotheses on
safety and feasibility issues. A formal sample size calculation as for confirmatory
trials was not done. Therefore, all resulting p-values and confidence intervals are to
be interpreted in the exploratory sense, only.
Data will be appropriately summarized and analysed using tabulation and graphs for
demographic/baseline characteristics and safety/tolerability and feasibility
observations and measurements.
The primary endpoint will be the incidence of grade II–IV acute graft-versus-host
disease (GVHD) on Day 100 post-transplantation. Frequency tabulations of the
number and percentage of patients with acute GVHD by severity (i.e. the ‘crude
incidence rates’) will be presented and displayed graphically along with the two-sided
95% confidence interval (Clopper-Pearson). In addition, time to acute GVHD will be
evaluated to assess the incidence and severity of acute GVHD from the first day of
transplantation (Day 0). The first day of acute GVHD onset at a certain grade will be
used to calculate a cumulative incidence curve for that acute GVHD grade. An overall
cumulative incidence curve will be computed along with a 95% confidence interval at
100 days post-transplantation with death considered as a competing risk.
Survival distributions of secondary endpoints will be estimated using the Kaplan-
Meier method. Binomial proportions will be estimated using the observed proportion
and Clopper-Pearson interval estimator. Incidence rates will also be estimated using
the cumulative incidence function.
Further safety results based on adverse events, physical examinations, vital signs
and clinical laboratory tests will be listed by patient and will be analysed by
descriptive statistics as appropriate.
The main analysis will be presented after completion of the 100 days post-
transplantation visit, i.e. when all patients have either completed the 100 days period
after transplantation, are lost to follow-up or have died within this period. Additional
analyses will be done on the 1-year post-transplantation and on the 2-year follow-up
data, i.e. when all patients have completed the 1-year or 2-year period after
transplantation, are lost to follow-up or have died within these periods.
Determination of Sample Size
No formal estimation of the sample size was performed. It is planned to enroll a total
of n = 60 patients, overall, in 13 centers, with 30 adult patients in 7 centers and 30
paediatric patients in 6 centers.
Safety monitoring and statistical stopping guidelines:
Patient safety during the study will be assessed continuously throughout the study by
monitoring incidence and severity of acute GVHD and incidence of NRM until
Day 100 post-transplantation and type of adverse events. In addition, changes in
findings of physical examination, vital signs and clinical laboratory results (complete
blood count, differential and platelet count and blood chemistry) will be evaluated for
the times defined as appropriate. Each case of acute GVHD grade III–IV and each
Final 7.0 18 of 111 21. May 2015
case of NRM have to be reported immediately and will be announced to the DSMB.
Pre-defined statistical stopping guidelines for GVHD grade III – IV and NRM have
been implemented and will be used as trigger for consultation with the DSMB to
decide about further study continuation. In case of premature study termination
patients included will be observed until their individual end of treatment as scheduled.
Graft failure will also be analysed and assessed by the DSMB for decision on the
conditioning regimen after treatment of the first 10 patients who have not received
ATG Fresenius. If graft failure occurred in less than 3 of these patients, all patients
enrolled after this analysis—both, patients in CR and patients with therapy-refractory
disease—can be transplanted with TNI instead of ATG Fresenius in the conditioning
regimen.
The responsible investigator has to decide case by case, about the most appropriate
approach for each patient.
Study Protocol Version Final 7.0, 21. May 2015
2. BACKGROUND AND RATIONALE

Hematopoietic stem cell transplantation (HSCT) has been established as treatment
option for a growing number of diverse congenital and acquired disorders, both
malignant and non-malignant. The latest activity survey of the European Group for
Blood and Marrow Transplantation (EBMT) reports the performance of 31,322
HSCTs in 2009 in Europe [8]. Conditions treated comprise hematological
malignancies (AML [32%], ALL [17%], MDS/MPS [14%], Non-Hodgkin lymphoma
[9%], and CML, CLL and Hodgkin’s Lymphoma [3% each]) but also solid tumors (1%)
and different non-malignant conditions (plasma cell disorders and bone marrow
failures [5% each], haemoglobinopathies and primary immune deficiencies [3%
each], inherited disorders of metabolism [1%] and autoimmune disease [<1%]).
Generally, HSCT is still associated with very serious risks for the patients and a high
incidence of complications and it remains therefore restricted to patients with life-
threatening diseases. However, in a number of hematological and non-hematological
indications it alone offers a curative option. This is self-evident for all hematological
malignant disorders (leukemias, lymphomas, myelomas) but it is also significant for a
growing number of patients with solid tumors, and non-malignant conditions as for
example congenital or acquired stem cell defects and immunodeficiencies (e.g. SCID
and congenital neutropenia), hematologic diseases (e.g. paroxysmal nocturnal
haemoglobinuria, aplastic anemia, and haemophagocytic lymphohistiocytosis) and
lysosomal storage disorders [see for example 26, 59, 56, 74, 66, 71, 2, 61, 1, 44, 25].
Rationales of the use of HSCT in the non-malignant indications are mostly the
restoration of a functional immune or hematopoietic system and the supplementation
of defective enzyme activities by the transplanted blood cells (e.g. lysosomal enzyme
activity of monocytes in Hurler’s syndrome [2]).
Stem cell transplantation may also be indicated for adult and paediatric patients
suffering from various solid tumors whose bone marrow has been destroyed by
previous myeloablative therapeutic strategies in the course of treating the underlying
oncological disease, e.g. by a previously explored autologous transplantation. In
these patients, HSCT offers the therapeutic option of overcoming dose-limiting
toxicities of intensive chemo-/radiotherapeutic approaches and e.g. neuroblastoma or
Ewing’s sarcoma are meanwhile established indications for HSCT [34, 46, 53, 65,
10]. Additionally, possible graft-versus-tumor effects of the transplant are of
considerable importance.
Final 7.0 19 of 111 21. May 2015
A fast onset of therapy is mandatory for the majority of patients treated in this study.
Rapid disease progression is for example observed in many hematological
malignancies as high-risk ALL or AML even in patients who have already achieved
clinical remission after first-line treatment. Children with inherited SCID need to be
transplanted as soon as possible. Therefore, it is often not feasible to wait for the
identification of an HLA-identical donor. Furthermore, evidence has been
accumulating that after autologous stem cell transplantation the restoration of the
patient’s immune system is not sufficient to eradicate the underlying disease. In this
setting, too, non-autologous transplantation may offer important advantages in
providing an additional graft-versus-tumor activity which may act in the sense of a
graft-mediated immune therapy [11, 18, 48].
However, to avoid serious graft-versus-host disease (GVHD) and graft failure in all
allogeneic stem cell transplantation settings it has long been inevitable to find donors
with the highest possible match to the respective recipients with respect to essential
cellular markers, especially HLA-characteristics. The therapeutic use of
hematopoietic stem cell transplantation thus is limited by the availability of a suitable
HLA-matched donor. A matched related donor can be found for only 30% of the
patients, 70% of the patients thus have to rely on finding a matched unrelated donor
[55]. Though donors can be identified for most of these patients, the search takes at
least several weeks if not months. If the aggressive course of the disease requires
the fast identification of a suitable donor, this is too long for a significant number of
patients.
In order to find a solution, haploidentical stem cell transplantation using closely
related, but only partially matched family donors has been exploited in several
therapeutic settings, recently. In theory, virtually every patient has a potentially
suitable haploidentical related donor—parent, sibling or child—and thus a successful
strategy for haploidentical allogeneic hematopoietic cell transplantation (HHCT) may
clearly be the solution for the ‘lacking donor’ problem. Yet, once again in only partially
matched donors and recipients the difficulties of resulting graft-versus-host disease
and graft failure arise, and initially trials of HHCT were complicated by a high
incidence of GVHD, engraftment failure, and infectious complications resulting in an
unacceptably high treatment related morbidity and mortality [3]. Recent efforts have
therefore been directed on developing therapeutic strategies to minimize these
complications. Graft rejection and GVHD are primarily mediated by T cells of host
and donor. Therefore attempts to overcome the HLA-barrier have focused on
strategies for effective host and graft T cell depletion.
CD34+ Enriched Stem Cell Grafts

An approach pioneered by the groups of Bachar-Lustig et al. and Aversa et al. was to
overcome the rejection of T cell depleted bone marrow cells by using a ‘megadose’ of
CD34+ cells (i.e. >10 × 106 CD34+ cells/kg) for transplantation. The harvesting of
such megadoses was achieved by treating donors with hematopoietic growth factors
prior to graft recovery. Using this setting, Aversa et al. treated patients with advanced
acute leukemia in different trials [5]. To avoid graft failure, intensive and highly
straining patient conditioning regimens with total-body irradiation, thiotepa,
fludarabine, and antithymocyte globulin had to be used. For patients transplanted in
complete remission, fast engraftment rates of neutrophils and platelets were
observed together with a low rate of GVHD (<10%) and a promising disease-free
survival of 47% at two years. For patients transplanted in relapse, however, EFS was
only 4% [6].
Final 7.0 20 of 111 21. May 2015

NRM, however, was very high in these settings due to the intensive myeloablative
conditioning regimens needed and a delayed immune reconstitution which resulted in
a high number of serious infections or regimen related toxicities (e.g. 36.5% in adult
patients in a study by Aversa and colleagues [6] and 66% in advanced AML [22).
Furthermore, a high incidence of relapses in patients not in CR at the time of HCT
was observed [6, 50, 36] and it had to be learned that CD34+ enriched stem cell
grafts offer no curative option for these patients. Another major obstacle to this
approach is the ‘megadose’ of CD34+ stem cells necessary for overcoming major
HLA-barriers. However, at doses below 10 × 106 CD34+ cells/kg BW of the recipient
rejection rates increased and engraftment and immune reconstitution were delayed
(most pronounced in patients receiving less than 8 × 106 CD34+ cells/kg BW [49]).
CD3+ and CD19+ Depleted Stem Cell Grafts

The aim of another therapeutic approach therefore was to develop a strategy to
improve engraftment independent from the infused stem cell doses. A reduced-
intensity conditioning regimen (RIC) was developed to reduce NRM [13, 14]. First
promising experiences were published for a paediatric population [9, 12, 39, 21].
Here, the Miltenyi CliniMACS cell sorting system was used for CD3+/CD19+ depletion
of the grafts using selective immunomagnetic beads. Still, profound T cell depletion is
a fundamental prerequisite for successful HHCT to avoid severe GVHD, while B cell
depletion is obligatory to avoid EBV-related post-transplant lymphoproliferative
disease (PTLD).
On the other hand, earlier investigations had demonstrated the important therapeutic
potential of alloreactive NK-cells in the graft [63]). (increased disease-free survival
[64, 50], engraftment facilitating and graft-versus-tumor effects [62] and a smaller
number of relapses and better survival [63]). NK-cells are involved in the immune
response against bacterial, viral and fungal infections and in transplantation settings
with KIR-mismatched donors patients had lower incidences e.g. of CMV-reactivation
[24, 57]. It was therefore expected that grafts selectively depleted of T and B cells
which still contain not only CD34+ stem cells but also a significant number of graft
facilitating cells such as NK-cells, monocytes and granulocytes would lead to
improved engraftment and better immune reconstitution.
Two phase I/II trials were initiated (adult patients with high risk disease of AML, ALL,
NHL, MM and CML and paediatric patients suffering from acute lymphatic and
myeloic leukemias, MDS, solid tumors and non-malignant diseases). No G-CSF was
administered post-transplantation and mycophenolate mofetil (MMF, 15 mg/kg bid)
was used only if the T-cell content in the graft exceeded 5 × 104 CD3+ cells/kg. The
regimen was well tolerated and engraftment was rapid (median time to >500
granulocytes/µL 13 days and to >20,000 platelets/µL 11 days). Furthermore, all but
one patient engrafted with full donor chimerism by day 14–28 post-transplantation. In
the trial with paediatric patients graft rejection occurred in 13%. Engraftment kinetics
thus were similar to those reported by Aversa et al. after HHCT with CD34+-
megadoses [6] although patients received a clearly lower dose of CD34+ cells/kg.
After TNI based reconditioning and second haploidentical stem cell donation, final
engraftment in paediatric patients was achieved in 100% [12]. However, in adult
patients immune reconstitution was delayed due to the profoundly T-cell depleted
grafts. NK-cell reconstitution was fast, probably due to the high NK-cell content of the
CD3+/CD19+ depleted grafts.
Final 7.0 21 of 111 21. May 2015

In this setting 9/36 (25%) adult patients died of NRM within the first 100 days and
incidence of grade II–IV GVHD was 36% overall (grade II=9, III=2 and IV=2 patients;
11% for aGVHD grade III–IV). One patient developed lethal aGVHD grade IV. Thus,
incidence and degree of GVHD in adult patients after HHCT with CD3+/CD19+
depleted grafts was higher than that reported for patients receiving HHCT with CD34+
selected grafts [6]. Bethge et al. also reported a high incidence rate for acute GVHD
of 48% in adult patients (10/29 patients with GVHD grade II, 2/29 each GVHD grade
III and grade IV with one patient with GVHD grade IV dying) and an NRM in the first
100 days of 20% [14]. In paediatric patients, 30% and 7% had aGVHD grades II and
III. No NRM was seen at day 100. In paediatric patients with acute leukemias and
MDS in remission at time of transplantation, a favorable 2-year EFS of 60% and a
low relapse rate of 20% after 2 years were observed. However, patients with active
disease (>5% blasts) still had extremely poor prognoses (2-year EFS 10% [21, 39]).
Especially in those patients, additional therapeutic strategies are urgently needed.
In conclusion, this regimen allowed haploidentical HCT in an older or heavily
pretreated patient population even without megadoses of CD34+ stem cells.
However, several factors still needed to be improved: the reconstitution of T cells,
especially the recovery of CD4 cells was poor, and the high numbers of graft failures
and high relapse rates, in particular in patients who were not in complete remission
prior to transplantation have to be addressed.
TCRα/β+ and CD19+ Depleted Stem Cell Grafts

The significant impact of graft composition and conditioning regimen on engraftment
has already been demonstrated by the fast engraftment kinetics observed in patients
receiving CD3+/CD19+ depleted grafts compared to merely CD34+ enriched grafts.
Cell species endangering transplantation outcome have meanwhile been identified in
even more detail. Mainly the subsets of CD3+ cells with TCRα/β receptors mediate
graft-versus-host activity while, contrarily, CD3+ cells with TCRγ/δ receptors show the
highly interesting graft-versus-tumor activity [11, 15, 18, 27, 31, 48, 75]. The
beneficial effects of NK cells in the graft have already been discussed (see above).
Furthermore, recent studies have revealed the existence of CD34-negative stem
cells, which are probably precursors of CD34+ stem cells with a high repopulating
capacity [77]. Additional graft facilitating cells have also been defined, such as CD8-
positive T-cells, monocytes and antigen-presenting cells (APCs) [19, 62, 45, 36, 73,
30]. The immunoregulatory properties of CD34+ stem cells have been demonstrated
previously [37, 38].
An efficient depletion of GVHD-mediating T cells presenting CD3 and TCRα/β as
surface markers is imperative to prevent acute graft-versus-host disease. On the
other hand, the remaining transfusional product has to have a high number of CD34+
hematopoietic stem cells. Additional benefits are expected from retaining cell
populations like NK cells, monocytes and TCRγ/δ cells that provide graft-versus-
tumor and anti-infectious activity. Further improvement of the cell product is achieved
by the depletion of CD19+ B cells because this reduces the risk for EBV-related
lymphoproliferative disorder (PTLD) post-transplantation. PTLD has previously been
a major risk in transplantation settings [33]. In consequence a new cell sorting
strategy for processing of the cell grafts has been developed using again the Miltenyi
CliniMACS cell sorting system. The stem cell grafts are selectively depleted of
TCRα/β+ and CD19+ cells by using paramagnetic microbeads in a single processing
step and the resulting cell grafts are rich in a variety of blood cells with diverse
immunological properties [20].
Final 7.0 22 of 111 21. May 2015
First clinical experiences in haploidentical HCT using TCRα/β- and CD19-depleted

PBSC grafts have been obtained with single paediatric patients in Tübingen [51, 70].
All were at extremely high risk and had poor prognosis with all currently available
transplantation methods, including CD3+/CD19+ depletion. They were pretreated with
a RIC conditioning regimen and received no post-transplantation immuno-
suppression. In these pilot patients engraftment and immune reconstitution were
rapid. No acute side-effects were noted and only one patient had an aGVHD grade III
of the skin requiring topical treatment, only. By now, treatment of 23 paediatric
patients has been reported in Tübingen and Rome [43]. Of these, ten patients
suffered from advanced and refractory leukemia (Tübingen) and 12/13 patients
suffered from relapsed/refractory ALL, AML and NHL (Rome). In Tübingen patients
received an RIC conditioning regimen with melphalan, thiotepa, fludarabine or
clofarabine and OKT-3 or ATG. In Rome conditioning was myeloablative
(fractionated TBI, thiotepa, fludarabine and ATG). In both settings, no post-
transplantation GVHD prophylactic immunosuppression was given.
In all cases engraftment was rapid (Tübingen: PMN recovery after median 9 days
[range 8–12] and platelet recovery after median 15 days [range 6–28]; Rome: PMN
recovery after median 11 days [range 7–13] and platelet recovery after median 12
days [range 10–16]). All patients treated in Tübingen using RIC conditioning showed
rapid immune reconstitution (already until day 28). Thus, immune recovery was
markedly improved compared to patients after CD3+/CD19+ depletion. In both
cohorts, of Tübingen and of Rome, TCRγ/δ cells expanded faster than TCRα/β cells
early after transplantation, though at day 100 TCRα/β cells were predominant.
Furthermore, V beta spectratyping showed a broad spectrum of T-cell receptors early
after transplantation. In Tübingen 5/10 patients showed aGVHD grade I, 2/10
developed aGVHD of the skin grade II and 1/10 developed aGVHD of the skin grade
III, which was, however, transient and required only topical treatment. In Rome only
2/13 patients developed aGVHD of the skin grade I. Up to now, 7/10 patients treated
in Tübingen and 10/13 patients treated in Rome are alive and disease-free (follow-up
3–12 months in Tübingen and 1–9 months in Rome). In Tübingen three patients
relapsed and no NRM was observed. In Rome two patients relapsed of whom one
died. Furthermore, one patient of the cohort treated in Rome experienced fatal lung
aspergillosis.
In summary, in all pilot patients treated so far (compare also [51, 70]) rapid and
sustained engraftment, rapid immune reconstitution, and a low incidence of GVHD
were observed. Cell sorting using the CliniMACS device proved to be efficient since a
high TCRα/β log depletion was achieved and the TCRα/β/CD19 depletion was
demonstrated to be effective and feasible while good recovery rates for stem cells
and innate effector cells were observed with a high viability of the resulting cells in
the transplant.
2.1. Study Rationale

As shown above, preliminary results of studies in pilot patients have been very
promising and suggest a clear benefit for patients suffering from hematological and
certain non-hematological malignancies requiring and eligible for haploidentical stem
cell transplantation. This also holds true for the severe non-malignant indications
included in this study. We expect that the use of TCRα/β+ and CD19+ depleted
haploidentical cell grafts in combination with the less aggressive conditioning
regimen needed for patient preparation will be associated with a low risk of acute
Final 7.0 23 of 111 21. May 2015

severe GVHD and graft failure and will increase speed, spectrum and functionality of
immune system reconstitution. This is supposed to reduce the incidence of severe
infections leading to lower rates of Non-relapse mortality (NRM). Preliminary data
have shown that the incidence of acute GVHD III–IV after transplantation of
TCRα/β+/CD19+ depleted haploidentical grafts is comparable to that seen after
transplantation of CD3+/CD19+ depleted and of CD34+ enriched grafts with a low
incidence of NRM [28, 29].
Furthermore, it is expected that engraftment rates will be higher and relapse
probability will be lower due to the presence of various effector cells with potential
anti-tumor activity, which are retained in the graft during the highly selective depletion
of TCRα/β+ and CD19+ cells. These retained beneficial cells might develop cytotoxic
GVT activity immediately after infusion. Since these effector cells with potential anti-
tumor activity are contained in the transplant at Day 0 they will be present already
during the first week after transplantation. This latter feature is of additional
importance in patients suffering from therapy-refractory solid tumors eligible in this
study. Therefore, it appears to be reasonable to offer TCRα/β+ and CD19+ depleted
stem cell grafts to a broad spectrum of patients suffering from high risk hematological
malignancies or therapy-refractory solid tumors with a curative treatment option
requiring and eligible for haploidentical stem cell transplantation, which have the
indication for an allogeneic HCT but are lacking a suitable HLA-matched donor and
have an eligible haploidentical donor at call. The study will comprise both, patients in
complete remission and patients with therapy-refractory malignancies and a
considerable disease burden. Patients with the life-threatening non-malignant
disorders eligible for stem cell transplantation as curative option included in this trial
will also profit from the presence of the manifold effector cells in the graft and the
resulting fast immune reconstitution.
The main study objectives are to demonstrate safety and feasibility of the proposed
treatment regimen in a mixed population of adult and paediatric patients with a great
variety of malignant hematological, other oncological and non-malignant conditions.
To increase the accessible patient population and to avoid center-specific effects
seven clinical sites will be involved.
2.2. Risk-Benefit Assessment

Stem cell mobilization and apheresis are not part of the study-specific procedures.
Donors will be informed separately and according to institutional guidelines of the
respective collection center regarding potential risks and side effects. The only study-
specific procedure for donors is the additional collection of one blood sample which is
not associated with any serious risk.
2.2.1. Potential study specific benefits for recipients
The transplantation of TCRα/β and CD19 depleted haploidentical stem cell grafts
offers potential benefits to participants in the study:
2.2.1.1. Low GVHD rates
TCRα/β cell depletion is expected to result in similarly low GVHD rates compared to
CD34 positive selection and CD3/19 depletion, thereby enabling haploidentical
transplantation.
Final 7.0 24 of 111 21. May 2015

2.2.1.2. Expedited immune reconstitution

The selective removal of TCRα/β-cells enables a stem cell graft that approximates a
regular T cell replete graft as much as possible, retaining potentially beneficial
effector cells like CD3 positive NK-cells and CD3 positive TCRγ/δ-cells. These
retained cells could support engraftment, exert graft-versus-malignancy effects,
reduce the risk of infections, and expedite immune reconstitution, which is currently
one of the major challenges after allogeneic and especially haploidentical stem cell
transplantation.
2.2.1.3. Reduced rate of infections
A high rate of functional immune cells such as NK cells and TCRγ/δ cells will be
transfused with the graft. Thus, a reduced rate of infections is expected when
compared to historical data from studies, in which CD34+-enriched and total CD3+ T-
cell depleted grafts were used.
2.2.1.4. Lower numbers of CD34 positive stem cells
The graft composition might enable the use of lower numbers of CD34+ stem cells
contained in the graft. This could be especially advantageous for adult patients due
to the higher body weight.
2.2.1.5. Prevention of PTLD
The concurrent depletion of B cells provides a preventive measure against the
development of PTLD.
2.2.1.6. Reduction of immunosuppression post transplantation
The combination of a selective TCRα/β/CD19 cell depleted stem cell graft and a
reduced dose intensity conditioning regimen allows the reduction of post-transplant
pharmaceutical GVHD prophylaxis. This treatment scheme is expected to result in
markedly reduced treatment-related toxicity and post-transplant morbidity. This
might also enable to increase eligibility for patients with advanced age, other
comorbidities and with donors unable to mobilize high numbers of CD34+ cells, which
might otherwise not be able to undergo transplantation.
2.2.2. Potential study specific risks for recipients
Recipients of allogeneic transplants are subject to risks from transplant related
procedures and medication utilized. These major risks are independent of the
TCRα/β/CD19 cell depletion using the CliniMACS TCRα/β/CD19 System and are laid
out in the later part of this chapter (2.2.4). It is expected that administration of
TCRα/β/CD19 depleted grafts might have positive effects on these risks.
Additional, study specific risks due to the depletion of TCRα/β/CD19 cells are as
follows:
2.2.2.1. Graft-versus-host Disease
Incomplete or insufficient removal of TCRα/β alloreactive cells might result in
potentially life threatening GVHD, acute or chronic. Measuring of residual TCRα/β
positive T cells is therefore of great importance and has been implemented in the
graft release criteria.
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Additionally incidence and severity of acute GVHD will be monitored continuously

throughout the study to assess patients’ safety. Each case of aGVHD grade III–IV
has to be reported immediately and will be announced to the DSMB (see sections
7.10 and 9.4).
2.2.2.2. Lymphoproliferative Syndrome (LPS, PTLD)
Incomplete or insufficient removal of CD19 positive B cells might result in PTLD,
which could also be life threatening. This risk is not different from T replete or un-
manipulated transplants, where no routine B cell removal is carried out.
Nevertheless, PCR analysis for EBV infection/reactivation will be performed (see
section 5.2.3.1).
2.2.2.3. Potential Sensitization to Murine Proteins
Patients could become sensitized against murine proteins, which are used during the
CliniMACS selection process. Although the procedure is based on the depletion of
the cells that are marked with the respective mouse-anti-CD19 and mouse-anti-biotin
antibodies it cannot be excluded that a few marked cells will be infused into a patient.
If the recipient has a pre-existing allergy, he or she may be at risk of allergic
reactions during infusion of the cell graft. The residual amount of murine protein in
the final product, however, is very low due to the depletion procedure (estimated
maximum dose of 30 µg for a patient with 50 kg body weight). To date, no allergic
reactions have been reported in patients receiving enriched cells processed with the
CliniMACS System. Thus, this risk is considered to be low due to the longstanding
experience with the reagents in different selection techniques (i.e. CD34 selection: no
clinically relevant sensitization has been reported since entering the market in 1997).
The most severe potential risk, graft versus host disease, is addressed in this trial
through the release specifications, the institution of statistical stopping guidelines and
continuous monitoring of the patient with the possibility to intervene if GVHD ensues.
Neither PTLD nor murine protein sensitization are considered a risk that would
prohibit this clinical trial.
2.2.3. Discussion of the Risk/Benefit Assessment and Conclusion

Allogeneic hematopoetic stem cell transplantation and especially the transplantation
of a haploidentical stem cell graft, carries substantial, well known risks that have to
be weighed against the risk of the malignancy or underlying non-malignant disease
as well as the consideration of other treatment options. For patients entering this
study, an allogeneic HSCT has been deemed necessary by the treating physician
and due to unavailability of a matched related or unrelated donor, haploidentical
transplantation is considered.
Disease indications allowed to enter this trial are severe and have a poor long term
outcome if transplantation is not performed. The transplantation of TCRα/β and CD19
depleted haploidentical stem cell grafts offers considerable potential benefits. Due to
the efficient TCRα/β cell depletion similar GVHD rates are expected as observed with
CD34+-selected or CD3/19-depleted grafts. However, GVHD as the most severe
potential risk is addressed in this trial through the release specifications, the
institution of statistical stopping guidelines and continuous monitoring of the patient
with the possibility to intervene if GVHD ensues. Neither PTLD nor murine protein
sensitization are considered a risk that would prohibit this clinical trial.
Benefits of this regimen are expected to be mediated by beneficial effector cells
which are retained in the graft during the depletion procedure. These cells are
Final 7.0 26 of 111 21. May 2015

expected to facilitate engraftment, exert GVT effects, reduce the risk of infections,
and improve immune reconstitution. Especially the latter is currently one of the major
challenges after stem cell transplantation. Additionaly, these beneficial cells offer the
advantage of using dose- and toxicity-reduced conditioning regimens and the
efficient and selective depletion of TCRα/β+ cells allows the reduction of
immunosuppressants post-transplantation for GVHD prophylaxis. In summary, this is
expected to translate into a reduced treatment-related toxicity. Preliminary data from
transplantation of single patients with TCRα/β- and CD19-depleted haploidentical
stem cell grafts show a low rate of acute GVHD as well as high engraftment rates
and a fast recovery of the immune system compared to data published for patients
transplanted with CD34-selected or CD3/CD19-depleted hematopoietic stem cell
grafts. Therefore, overall, the available information suggests that the present study
has a favourable risk-benefit ratio.
2.2.4. Major risks of allogeneic hematopoetic stem cell transplantation independent

of the IMP
Major risks specific to allogeneic hematopoetic stem cell transplantation for the
recipients, which are independent of the CliniMACS TCRα/β/CD19 cell depletion
using the CliniMACS system are described in the following sections. It is expected
that administration of the IMPs (TCRα/β/CD19 depleted grafts) might have positive
effects on these risks.
2.2.4.1. PBSC Infusion
Symptoms may include changes in heart rate, arrhythmia, changes in blood
pressure, fever, chills, sweats, nausea, vomiting, diarrhoea, abdominal cramping,
haemoglobinuria, acute renal failure, allergic reactions, respiratory dysfunction, or
headache.
2.2.4.2. Infections
Due to the underlying disease and to risks associated with immune suppression
during conditioning, transplantation generally puts the patient at higher risk for
potentially life-threatening bacterial, viral, or fungal infections. These risks are
potentially higher with T-cell depleted cell grafts, which are mandatory in
haploidentical transplantation. However, in this study a high rate of functional
immune cells such as NK cells and TCRγ/δ cells will be transfused with the graft.
Thus, a reduced rate of infections is expected when compared to historical data from
studies, in which CD34+-enriched and total CD3+ T-cell depleted grafts were used.
Nevertheless, as prophylactic measures patients will be closely monitored for signs
of infections (see section 5.1.4.1) and will receive preemptive treatment as per
institutional guidelines.
2.2.4.3. Graft Failure / Poor Marrow Function
Generally, T cell depletion of donor cells is associated with an increased incidence of
graft failure in allogeneic transplant recipients. After allogeneic transplantation, the
recipient’s marrow function may be poor and leucopenia, anemia, or
thrombocytopenia may result. Graft failure may result in death if not reversed.
In the present study, however, selectively depleted cell grafts will be used, which still
contain a high number of functional immune cells. Preliminary results in single
patients have shown a remarkably fast functional reconstitution and good
engraftment rates after transplantation of the TCRα/β and CD19 depleted grafts [43].
Final 7.0 27 of 111 21. May 2015
Furthermore, the selective depletion of TCRα/β+ cells allows for a dose-reduced

conditioning regimen with reduced immune toxicity. Therefore, a lower rate of graft
failures and severely impaired marrow function after transplantation is expected
compared to previous studies of haploidentical PBSC transplantation using differently
processed cell grafts.
In case of suspected graft failure the coordinating investigator or the respective
leading investigator will have to be consulted for advice regarding treatment.
2.2.4.4. Graft-versus-host Disease
Potentially disabling acute or chronic GVHD may develop after allogeneic
transplantation and can lead to death. GVHD is thought to be mainly initiated by
TCRα/β+ cells contained in the PBSC graft. Selective depletion of the graft of
TCRα/β+ cells thus is expected to reduce the incidence of severe GVHD. This
assumption is supported by preliminary experiences in single patients [43]. Still,
GVHD is possible in the setting of the present study. See section 5.1.4.3 for
treatment according to this protocol. In this study incidence and severity of acute
GVHD will be monitored continuously throughout the study to assess patients’ safety.
Each case of aGVHD grade III–IV has to be reported immediately and will be
announced to the DSMB (see section 9.4).
2.2.4.5. Veno-occlusive Disease (VOD) of the Liver
VOD is a manifestation of damage of the liver which can be caused by the
conditioning regimens needed in haploidentical stem cell transplantation. It usually
develops within two weeks after allogeneic transplantation and is characterized by at
least two of the following:
• Hyperbilirubinemia (total bilirubin >2 mg/dL)
• Hepatomegaly or right upper quadrant pain
• Sudden weight gain (>5% above baseline)
Recipients developing VOD will have to be monitored closely and will receive
appropriate supportive care and therapy (e.g. Defibrotide) including careful fluid
management.
2.2.4.6. Death
After haploidentical stem cell transplantation patients have a considerable risk of
treatment related mortality within the first year after transplantation (depending on the
study, values between 20 and 50% have been published). This results from severe
regimen related toxicity, and risks of haemorrhage, opportunistic infections, or other
complications. In this study, incidence and severity of NRM will be monitored
continuously throughout the study to assess patient’s safety. Each case of NRM until
Day 100 post SCT has to be reported immediately and will be announced to the
DSMB (see section 9.4).
2.2.4.7. Therapy Toxicities
The conditioning regimens used for patient preparation prior to PBSC transplantation
are associated with a considerable number of toxicities. Treatment-related side
effects of the drugs and interventions used are described in more detail in
section 6.6.
Final 7.0 28 of 111 21. May 2015

3. STUDY OBJECTIVES AND ENDPOINTS

3.1. Objectives
Primary objective:
Evaluation of the safety/tolerability and feasibility of haploidentical PBSC grafts
depleted of TCRα/β+ and CD19+ cells using the CliniMACS TCRα/β/CD19 System in
adult and paediatric patients with hematological and non-hematological malignancies
and specific non-malignant diseases, defined as the incidence of grade II–IV acute
graft-versus-host disease (GVHD) on Day 100 post-transplantation.
Secondary objectives:
Secondary objectives are
Safety outcome variables
• Incidence of grade I acute GVHD until Day 100 post-transplantation
• Incidence and severity of chronic GVHD after 1 year and 2 years
• Incidence of NRM at all visits throughout the study
• Incidence and severity of acute infusional toxicity on the days of stem cell
transfusion
• Graft failure, where primary graft failure is defined as the failure to achieve an
absolute neutrophil count (ANC) >500/µl at Day +28 and where secondary graft
failure is defined as initial neutrophil engraftment followed by a decline in ANC
<500/µl that is unresponsive to growth factor therapy
Feasibility outcome variables
• Neutrophil and platelet engraftment from Day 0 to Day 28 defined as
- Neutrophil engraftment: ANC cell counts and time to reach >500 neutrophils/µl
for three consecutive days
- Platelet engraftment: platelet counts and time to reach ≥20.000 platelets/µl for 3
consecutive days with independence from platelet transfusions for 7 days
• Overall survival at Day 100 and after 1 and 2 years
• Disease free survival at Day 100 and after 1 and 2 years
• Transfusion requirement from Day 0 to Day 100 defined as
- Number of transfusions (thrombocytes, erythrocytes and other blood products)
- Time to last transfusion of thrombocytes, erythrocytes and other blood products
• Incidence of relapse at Day 100 and after 1 and 2 years: rate and time to relapse
• Days of (re)hospitalization assessed at Day 28 and Day 100
• Quality of Life Assessment using EQ-5D (adult patients ≥18 years), PedsQL
(paediatric patients, age <18 years) and FACT-BMT (adult patients ≥18 years) at
baseline, Day 100 and after 1 and 2 years
Laboratory outcome parameters (routine local assessments)
• Donor chimerism by
- PCR-analysis of peripheral blood samples collected on Days 7, 14, 21, 28, 35,
42, 49, 56, 63, 70, 100, 180 and 270 post-transplantation compared to samples
from donor and recipient collected prior to transplantation
- PCR-analysis of bone marrow samples of haematological malignancies
collected on Days 28 and 100 post-transplantation
Laboratory outcome parameters (additional central assessments)
• Reconstitution of T, B, NK and T regulatory (Treg) cell subsets assessed by
immune cell phenotyping: Cell counts of CD3+, CD4+, CD8+, CD3+CD56+,
CD3+TCRα/β+, CD3+TCRγ/δ+ T cells, naive CD4+TCRα/β+, memory
Final 7.0 29 of 111 21. May 2015
CD4+TCRα/β+, naive CD8+TCRα/β+, memory CD8+TCRα/β+, DN TCRα/β+,

TCR Vd2+TCRγ/δ+, TCR Vd2–TCRγ/δ+, naive Tregs, ‘memory’ Tregs, B cells,
monocytes, NK cells, neutrophils, eosinophils;
samples collected on Days 7, 14, 21, 28, 63, 100 and 6 and 12 months post-
transplantation
• Reconstitution of T Vbeta repertoire: TCR V beta spectratyping (PCR analysis);
samples collected on Days 28, 63 and 100 post-transplantation
• Reconstitution of T gamma/delta repertoire: TCR V gamma/delta spectratyping
(PCR analysis); samples collected on Days 28, 63 and 100 post-transplantation
• Thymic function by TREC analysis (PCR analysis), samples collected on Days 28,
63 and 100 post-transplantation
• KIR genotyping of donor and recipient (baseline reference sample) and
reconstitution of NK-cell KIR repertoire by FACS analysis; samples collected on
Days 28, 63 and 100 post-transplantation
• T cell activity by
- Analysis for CMV-, EBV- and ADV-reactive T cells; samples collected on
Days 28, 63 and 100 post-transplantation
- Analysis of T cell stimulation/proliferation; samples collected on Days 28, 63
• NK cell activity; samples collected on Days 28, 63 and 100 post-transplantation
• Performance of the CliniMACS TCRα/β/CD19 System utilizing the CliniMACS
TCRα/β-Biotin and CliniMACS CD19 reagents to produce a graft with defined cell
content as assessed by:
- Percentage of CD34+CD45+ cells recovered after TCRα/β and CD19 depletion
procedure: target value ≥95%
- Log depletion of CD19+ cells
- Log depletion of TCRα/β+ cells
- Cell counts of viable CD34+CD45+ blood stem cells, CD20+ B cells,
CD56+CD16+ TCRα/β and TCRγ/δ cells, CD3+ cells and CD45+/WBC cells
analysed by flow cytometry after processing prior to transplantation
- The percentage of recovered viable CD45+ cells after TCRα/β and CD19
depletion procedure: target value ≥90%
- Haematocrit value in graft in mL/mL erythrocytes
- Number of grafts with ≥4 × 106 CD34+CD45+ stem cells/kg BW of the recipient
- Number of grafts with ≤25 × 103 TCRα/β+ cells/kg BW of the recipient
- Number of grafts with ≤1 × 105 CD20+ cells/kg BW of the recipient
- Sterility of IMP.
Safety Objectives:
• Incidence and type of infections
- Incidence of CMV, ADV (if done according to hospital routine), EBV and
aspergillus infections as well as other viral, bacterial and fungal infections at
Day 100 and 1 year post-transplantation
- Number of virus reactivations of CMV, ADV, EBV by PCR once to twice a week
until Day 28, weekly until Day 70 and on Day 100
• Incidence, severity and type of adverse events and serious adverse events,
clinically relevant vital signs and safety laboratory parameters
- Serious adverse events throughout the study
- ‘Unknown therapy-related toxicity’ as AE and ‘known therapy related toxicitiy’
from Day –12 to Day 0 (during conditioning and prior to stem cell
transplantation)
Final 7.0 30 of 111 21. May 2015
- All adverse events from Day 0 to Day 100 in detail

- Vital signs throughout the study
• Safety laboratory parameters (clinical chemistry and complete blood counts) at
baseline and from Day –4 to Day 100
• Concomitant medication
- Concomitant medication from baseline to Day 100
- Documentation of new treatment with cellular products (erythrocytes,
thrombocytes, donor lymphocyte infusions [DLIs] or antigen-specific T cells)
after Day 100
- Enrollment in any other clinical study after Day 100.
3.2. Study hypothesis

Due to the diverse patient collective and the exploratory nature of the study no exact
statistical hypothesis will be stated. It is expected, however, that the incidence rate of
aGVHD Grade II–IV up to Day 100 will be comparable to rates after transplantation
with CD3/CD19 depleted grafts (i.e. approximately 27% for paediatric patients [40]
and 54% in adult patients [29]). These rates are slightly higher than aGVHD rates
achieved with transplantation of CD34+ enriched grafts (10–15%; [40]). Apart from
this, improvement of quality of life and clinical state are expected for all patients,
although outcomes will differ according to the different oncological and non-malignant
conditions.
There are no comparator and no control groups in this study. Therefore, comparisons
are possible only versus data from other clinical trials in the same indications.
Generally, it is supposed, that compared to historical data patients will benefit from
• improved engraftment rates,
• improved immune reconstitution (increased speed, spectrum, functionality of
immune system reconstitution) leading to a lower rate of severe infections,
• reduced non-relapse- mortality, due to the lower toxicity of the conditioning
regimen and a lower rate of severe infections,
• reduced relapse frequency and longer times of disease-free survival and,
• overall, an improved quality of life.
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4. STUDY DESIGN
4.1. Study Overview
This multi-center, open-label, single-arm, phase I/II clinical trial will assess
safety/tolerability and feasibility of transplantation with haploidentical peripheral blood
stem cell grafts depleted of TCRα/β+ and CD19+ cells using the Miltenyi CliniMACS
TCRα/β/CD19 System in adult and paediatric patients suffering from various
hematological and non-hematological malignancies and non-malignant diseases.
Patients will be prepared for transplantation by a moderate T cell depletion regimen
prior to HSCT. For all patients GVHD prophylaxis with MMF from Day –1 until Day
+30 is planned. Safety will be primarily assessed by determining occurrence and time
to acute GVHD grade II–IV at Day 100 post-transplantation. Secondary endpoints are
the occurrence of acute infusional toxicity, the rate of NRM, engraftment and graft
failure, chronic GVHD, reconstitution and functionality of relevant cells of the immune
system (T, B and NK cell subsets as well as T regulatory cells), T cell chimerism after
transplantation, activation of and newly occurring infectious diseases, relapse rate,
overall and disease-free survival and quality of life up to 2 years after transplantation.
To assess the efficiency of the CliniMACS TCRα/β/CD19 System cell counts of the
stem cell grafts will be determined pre-transplantation and the proportion of grafts
sufficiently depleted or with more residual TCRα/β and CD20+ cells as well as the
percentage of viable CD34+CD45+ cells in the graft after depletion will be determined.
4.2. Rationale for Study Design
For the following reasons no prospective, randomized study design for the present
study has been chosen: Firstly, it should be noted that currently no ‘standard of care’
exists for the treatment of the specific patient population receiving a haploidentical
stem cell transplantation. Approaches currently used within the scope of haplo-
identical stem cell transplantation are, for example, graft manipulations via
CliniMACS CD34+ enrichment or CliniMACS CD3/19+ depletion, the use of unmani-
pulated grafts in combination with high dose of cyclophosphamide post-transplan-
tation or the use of umbilical cord derived grafts. In general, approaches are chosen
by transplant centers according to their experiences, their preferences and to the
study protocols available. Thus, a comparison with an accepted ‘standard of care’ as
in the matched setting situation (with a 10/10 match) is not possible within the scope
of haploidentical stem cell transplantation. Thus, data currently available do not
suffice to allow the clear identification of a comparator therapy. However, transplan-
tation is the most promising and only potential curative treatment option for this
patient population. Secondly, it has to be considered that the present study is the first
prospective clinical study by this sponsor with TCRα/β and CD19-depleted PBSC
grafts in the haploidentical setting. Thus, the main goal, as a first step of clinical
development, is to verify that the potential beneficial accessory cells, particularly
CD3+TCRγ/δ+ T cells retained in the graft, are safe and do not show alloreactivity.
This is also reflected by the primary endpoint, which is aGvHD grade II–IV on
Day 100. Following these arguments, the conduct of a non-randomized, single arm,
prospective study was chosen to determine safety and tolerability of TCRα/β and
CD19-depleted PBSC grafts in the haploidentical setting.The study will be performed
as multi-center trial to reach sufficient patient numbers in the planned time because
of the low overall frequency of the conditions analyzed (Table 1).
Final 7.0 32 of 111 21. May 2015

Table 1: Incidences of oncological and non-malignant diseases considered in this

study
Incidence per year
[relative number of patients]
Hematological malignancy
ALL 1.5/100,000
AML 3/100,000
Hodgkin’s lymphoma 2–4/100,000
Non-Hodgkin’s lymphoma 5–10/100,000
MDS 4/100,000
MPS 4,6/100,000
CML 1–2/100,000
Multiple myeloma 4-6/100,000
Solid tumour
Neuroblastoma 1/7,000
Rhabdomyosarcoma 2/100,000
Ewing sarcoma 3/100,000
Peripheral primitive neuroectodermal tumor (PNET) 7/1,000,000
Non-malignant condition
Severe aplastic anemia 2/1,000,000
PNH 1–1.5/1,000,000
HLH 1.2/1,000,000
SCID 1/100,000 to 1/500,000
Chediak Higashi Syndrome unknown
Malignant osteopetrosis 1/100,000 to 1/500,000
Lysosomal storage diseases 1/5,000 to 1/10,000 overall
4.3. Study Population

Hematopoietic stem cell transplantation is often the only curative treatment option for
critically ill patients suffering from diseases of the hematopoietic system, including
numerous other diseases besides leukemia, which are listed in the indication
catalogue of the German working group for bone marrow- and blood stem cell
transplantation (http://www.dag-kbt.de). The first aim of this therapy is to eliminate a
bone marrow infiltrative process, such as leukemia, or to correct congenital
immunodeficiency disorders by destruction of the patient’s hematopoietic system
followed by its replacement through progenitor cells in the stem cell graft capable of
reconstituting normal bone marrow function. In this context toxicity of the
hematopoietic stem cell transplantation, especially incidence and severity of GVHD,
which will be primarily validated in the present study, does not depend mainly on the
underlying disease, but on the composition of the stem cell transplant itself. Following
these arguments patients with a multitude of underlying diseases are admitted to be
enrolled in this study. Even though it is expected that patients with high-risk leukemia
will form the majority of patients in this study, safety and feasibility of TCRα/β/CD19
depleted peripheral blood stem cells should also be validated in other common
indications for blood stem cell transplantation, as solid tumors and non-malignant
diseases. At the same time, patient inclusion is limited to the common indications for
hematopoietic stem cell transplantation. This is ensured since the indication for an
allogeneic blood stem cell transplantation has to be stated before a patient is enrolled
in this study. In addition enrolment is restricted to patients, for which a HLA-matched
donor is lacking. In this context it also has to be considered that the 41 individual
patients, who have already been treated in Tübingen and Rome with TCRα/β and
CD19-depleted haploidentical stem cell grafts suffered from numerous malignant and
Final 7.0 33 of 111 21. May 2015

non-malignant diseases. Independent from the underlying disease, the reported

results are promising. Based on these experiences a benefit for the patients may be
expected from transplantation with TCRα/β and CD19-depleted haploidentical stem
cell grafts. The extension of inclusion criteria to all common indications for stem cell
transplantation will allow critically ill patients with these indications to participate in
this study.
Both, adult as well as pediatric patients will be included. Altogether 41 patients, all of
which were children, have already been treated in Tübingen and Rome with TCRα/β
and CD19-depleted haploidentical stem cell grafts, either in the absence of other
therapeutic options or following a treatment failure. Some of the treated diseases do
not appear in the adult population, but in the pediatric population, only, and require a
stem cell transplantation. Such indications will be also evaluated in the present study,
including relapsed neuroblastoma, relapsed rhabdomyosarcoma, Ewing sarcoma,
haemaphagocytic lymphohistiocytosis, SCID, Chediak Higashi syndrome as well as
the storage diseases listed in the inclusion criteria. Furthermore it has to be
considered that first line treatment of the more common indications as AML, ALL,
Hodgin’s disease as well as non-Hodgin lymphoma in the pediatric population differs
strongly from that in the adult population. In case first line therapy fails, the disease
becomes refractory or a relapse occurs a stem cell transplantation is indicated. The
outcomes of the 41 pediatric patients, which have already been treated suggest a
direct benefit for the pediatric population, which is required by article 4g of the
Clinical Trial Directive 2001/20/EC. Furthermore, the patients’ interest prevails over
that of science and society, as required by article 4i of the Clinical Trial Directive
2001/20/EC. A rapid development of this promising treatment option is considered to
be in the best interest of children suffering from any of the indications included in this
study to ensure its becoming rapidly available to a larger population in case of
encouraging outcomes. In this context it also has to be noted that treatment success
in a pediatric population cannot be infered from treatment success in an adult
population. The same is true for side effects and toxicity profile, which will be
evaluated in this study. A clinical trial with adult patients, only, would prevent
prospective data collection and analysis in the pediatric population, which is needed
for an authorization according to §21a AMG. In this case the start of a study with
pediatric patients would require analyzed data from a study with adult patients. This
would result in a substantial delay in development of this therapeutic option in
pediatric patients. As a consequence it would not be available for children for the
next few years. Due to the reasons stated above, we conclude that this study fulfills
the criteria required for a trial with pediatric patients.
It is planned to enroll 60 patients, overall, in 13 centers, with 30 adult patients in
7 centers and 30 paediatric patients in 6 centers.
4.3.1. Selection Criteria
Patient Inclusion Criteria
A patient has to meet all of the following criteria to be eligible:
1. Male or female without childbearing potential (e.g. prepubertal child, post-
menopausal, surgically sterile) or using medically adequate contraception
2. Patient suffering from hematological malignancy, non-hematological malignancy
or non-malignant disease requiring and eligible for allogeneic stem cell
transplantation. Indications are:
Final 7.0 34 of 111 21. May 2015

• Hematological malignancy in complete remission (CR), partial remission (PR) or

with stable disease (defined according to current international recommendations
and guidelines)
- Patient with high-risk acute myeloid leukemia (AML) in CR1
Patient with relapsed or primary therapy-refractory AML
- Patient with high-risk acute lymphatic leukemia (ALL) in CR1
Patient with relapsed or primary refractory ALL
- Patient with relapsed or primary refractory Hodgkin’s disease
- Patient with relapsed or primary refractory non-Hodgkin’s lymphoma
- Patient with refractory myelodysplastic syndrome (MDS)/myeloproliferative
syndrome (MPS)
- Patient with relapsed or refractory multiple myeloma (MM)
• Non-hematological malignancy without curative treatment option, mainly
- Neuroblastoma:
Patient with relapsed metastatic nmyc-negative or –positive or local nmyc-
positive neuroblastoma
- Soft tissue sarcoma:
Patient with relapsed metastatic soft tissue sarcoma (rhabdomyosarcoma,
Ewing sarcoma, peripheral neuroectodermal tumor)
Patient >10 years with soft tissue sarcoma with primary bone metastases or
bone involvement
• Non-malignant disease with HSCT as treatment option, mainly
- Acquired or congenital hematological disease
o Severe aplastic anemia (patient not responding to immune
suppression),
o Paroxysmal nocturnal haemoglobinuria (PNH)
o Haemophagocytic lymphohistiocytosis (HLH)
- Congenital immunodeficiency
o Severe combined immune deficiency (SCID) or related disease
o Chediak Higashi Syndrome
- Congenital metabolic disorder
o Malignant osteopetrosis
o Lysosomal storage disorder (mucopolysaccharidoses, leukodystrophy,
glycoprotein disorder)
3. No HLA-identical stem cell donor available as determined by high-resolution
typing (maximum of 1 antigen or allelic mismatch are acceptable [9/10 match]),,
but eligible haploidentical donor with >1 antigenic or allelic mismatch (9/10-
match) identified and at call; Exception: Haplo-identical HSCT is medically
indicated even if an HLA-identical donor is available and decision for haplo-
identical HSCT has been made according to hospital routine prior to inclusion of
the patient into this study.
4. Patient aged ≥8 weeks to ≤65 years
5. Karnofsky (patient >16 years)/Lansky (patient ≤16 years) index >60%
6. Patient in good clinical condition without concomitant disease significantly
increasing the risk of transplantation, see exclusion criteria
7. Adult patient without active infection at the time of transplantation
8. Pediatric patient without uncontrollable, progressive infection at the time of
transplantation
9. Informed consent given (by patient or legal representative) prior to any
study-related procedures.
Final 7.0 35 of 111 21. May 2015

Patient Exclusion Criteria

A patient meeting any of the following criteria is not eligible for the present study:
1. Age >65 years or <8 weeks
2. Patients with progressive disease prior HCT
3. <3 months after preceding hematopoietic cell transplantation (HCT)
4. History of neurological impairment (active seizures, severe peripheral neuropathy,
signs of leukencephalopathy, active CNS infection)
Note: For patients with HLH or malignant osteopetrosis or other patients with
heavy pretreatment with irradiation or intrathecal chemotherapy pre-transplant
CNS MRI and neurological consultation are mandatory.
5. Fungal infections with radiological and clinical progression
6. Liver function abnormalities with bilirubin >2 mg/dL and elevation of
transaminases higher than 400 U/L
7. Chronic active viral hepatitis
8. Adult patients: ejection fraction <40% on echocardiography; pediatric patients:
ejection fraction <40% or shortening fraction <25% on echocardiography
9. Patients with > grade II hypertension (uncontrolled by Common Toxicity Criteria,
CTC)
10. Creatinine clearance below threshold defined for stem cell transplantation
according to local clinical standard
11. Respiratory failure necessitating supplemental oxygen
12. HIV infection
13. Female patient who is pregnant or breast feeding, or adult of reproductive
potential not willing to use an effective method of birth control during study
treatment and for at least 12 months thereafter.
Note: Women of childbearing potential must have a negative serum pregnancy
test at study entry.
14. Concurrent severe or uncontrolled medical disease (e.g. uncontrolled diabetes,
congestive heart failure, myocardial infarction within 6 months prior to the study,
unstable and uncontrolled hypertension, chronic renal disease, or active
uncontrolled infection) which by assessment of the treating physician could
compromise participation in the study
15. Patient with a history of psychiatric illness or a condition which could interfere with
his/her ability to understand the requirements of the study (this includes
alcoholism/drug addiction)
16. Patient unwilling or unable to comply with the protocol or unable to give informed
consent
17. Treatment with any investigational product within 4 weeks prior to study treatment
(transfusion of the IMP)
Donor Selection
1. Haploidentical family member previously identified as eligible donor by
donor/recipient cross-matching including HLA-typing.
Note: In case of positive cross-match results for donor-reactive anti HLA
antibodies an alternative donor with negative cross-match results should be
preferred, if available. If no alternative donor with negative cross-match results is
available, removal of anti HLA antibodies is highly recommended to prevent graft
rejection.
2. Donor age ≥16 years
Final 7.0 36 of 111 21. May 2015

Note: Positive evaluation for allogeneic hematopoietic cell donation has to have
been performed at the collection center according to local standard practice. Also
informed consent for mobilization and collection of peripheral blood stem cells
according to local institutional guidelines has to have been given in this context
independently of the present clinical study. Stem cell mobilization and collection
procedures are not part of this study and will be performed at the collection
center according to local standard procedures.
3. Study specific informed consent given.
4.3.2. Withdrawal and Replacement
Patient Withdrawal and Replacement

Patients or their legal representatives are free to withdraw consent to participate in
the study at any time without penalty and without stating any reason. This will not
prejudice the future medical care of the patient in any way. All patients have to be
closely monitored and treated as appropriate, according to the respective guidelines
that relate to patients with the malignant and non-malignant conditions included in
this study. The investigators are authorized to perform additional follow-up
examinations at their discretion.
Patients have to be withdrawn if any of the following events occur:
• The patient withdraws consent
• Pregnancy
If a patient has to be withdrawn because of pregnancy, she has to be followed-
up until after the delivery of the child and reported to the sponsor accordingly.
The Sponsor or the investigators are free to withdraw patients when this is
considered medically or otherwise necessary. Justified reasons are
• Adverse event(s)
• Violation of eligibility criteria
• Violation of the study protocol (e.g. conditioning or failure to attend study visits)
• Donor withdrawal.
Reason(s) for and the justifications of the withdrawal always have to be documented
on the electronic case report form (eCRF) in as much detail as possible. If a patient is
prematurely withdrawn from the study for any reason, the investigator will under all
circumstances try to perform all evaluations described for the early termination visit
(see section 5.5). Withdrawn patients can be replaced.
Screening failures (patients who had a screening examination (Visit 1), but who, for
any reason, did not receive the IMP) and patients, who receive hematopoietic cell
grafts with an insufficient number of CD34+CD45+ cells (<4.0 × 106/kg BW) may be
replaced to include at least 60 evaluable patients into the study.
Donor Replacement
If a patient withdraws study-specific consent prior to conditioning and transplantation
the blood sample of the donor will be destroyed in case it has already been collected.
Should a donor be no longer eligible for stem cell mobilization or apheresis according
to the responsible collection center’s judgement, the investigator will be informed
immediately. Reasons have to be stated and will be documented on the electronic
case report form (eCRF) of the recipient. An alternative donor for the patient should
be identified as soon as possible. If this is not possible the patient has to be
withdrawn from the study.
Final 7.0 37 of 111 21. May 2015

4.3.3. Patient Identification and Randomization

No randomization is planned for this single-arm, open-label study.
Patients will be approached for this study after the decision to proceed with
transplantation has been made, a HLA-matched donor is not available and a suitable
haploidentical donor has been identified. Transplant physicians will evaluate the
patient eligibility for assignment to this study. Only patients who comply with all entry
(in- and exclusion) criteria will receive a patient number and will be assigned to the
course of the study treatments. Eligibility criteria will be verified and ineligible patients
will proceed off-study and no further follow-up will be obtained.
Eligible patients willing to participate in the study will sign an approved consent form
prior to any study relevant treatment.
During the study, patients will be identified by an ascending 4-digit patient number,
consisting of a two digit site number followed by a two digit subject number (xx-yy).
Patient numbers will be assigned in a consecutive manner at each site in the order of
inclusion, e.g. the first patient at each site receives the first number (for example for
site 02: 02-01), the second patient receives the next number in sequence and so on.
This 4-digit patient number will be recorded on the electronic case report forms
(eCRFs) and all study-specific documents of a specific patient and in the site patient
file.
The investigator will keep a record with the names, the birth dates and the patient
number of the patients (subject identification list) to allow checking of data in the
clinical files of each patient, when required. This record will remain at the study site.
4.3.4. Protocol Violations

Except in the case of a medical emergency, no protocol violation is authorized
outside amendments established by Miltenyi Biotec GmbH and approved by an
IEC/IRB (Institutional Review Board). Protocol violations may affect the conduct of
the study from legal and ethical points of view and may influence the statistical
analysis and pertinence of the study. In case of a medical emergency event in a
patient leading to a protocol violation this will only be allowed for the single patient.
The investigator has to contact the sponsor to clarify if the patient may continue in
the study.
4.3.5. Premature Termination of the Study

If the investigator, the Sponsor or the clinical monitor becomes aware of conditions or
events suggesting a possible hazard to patients in case the study continues, the
study may be terminated after appropriate consultation between the relevant parties.
The study may also be terminated early at the Sponsor’s discretion in the absence of
such a finding.
Conditions that may warrant termination include, but are not limited to:
• The discovery of an unexpected, significant, or unacceptable risk to the patients
enrolled in the study
• Failure to enroll patients at an acceptable rate
• A decision on the part of the Sponsor to suspend or discontinue development of
the IMP.
Final 7.0 38 of 111 21. May 2015

4.4. Study Treatment

Patients will be treated in this study with individual hematopoietic PBSC grafts from
haploidentical donors depleted of TCRα/β+ and CD19+ cells. Prior to transplantation
patients will be prepared with different dose-reduced conditioning regimens.
Conditioning regimens for patients are described in more detail below (see section
5.1.2).
4.4.1. Preparation of the Hematopoietic Stem Cell Graft (IMP)
Stem cell mobilization and stem cell apheresis will be performed according to
standard of care following legal requirements in Germany and in the Netherlands. For
Germany this includes, but is not limited to the German transfusion law and relevant
guidelines [78]. In order to ensure that the TCRα/β and CD19-depleted stem cell graft
is available for the patient at the required time, coordination of the parties concerned
has to be performed according to clinical practice for haploidentical stem cell
transplantation with manipulated stem cell grafts following internal guidelines and
legal requirements (e.g. German guideline ‘Richtlinie zur Transplantation peripherer
Blutzellen’). It is the Investigator’s responsibility to ensure that a suitable stem cell
graft will be available for the patient in time.
The manufacturing process of the TCRα/β- and CD19-depleted cell grafts will be
performed in local GMP laboratories assigned by the sponsor (see Appendix 7). The
manufacturing process and quality control will be performed according to validated
procedures and documented in accordance with full GMP requirements.
The stem cell apheresis product (see sections 5.1.1) will be depleted of TCRα/β+ and
CD19+ cells by negative selection using the automated CliniMACS® Plus device as
described in the IMPD, the current version of the CliniMACS user manual and
according to institutional Standard Operating Procedures (SOPs) in place and
validated at the manufacturing sites.
The specification for the final formulation of the IMPs (TCRabCD19PBSC and
TCRabCD19PBSC_cryo) has been set according to findings from previous studies
[41, 42, 48, 50].
For transplantation the following graft composition is targeted
• Minimum percentage of viable CD34+CD45+ cells ≥95%
• Number of viable CD34+CD45+ cells ≥4 × 106 cells/kg BW of the patient
• Number of TCRα/β+ cells ≤25 × 103 cells/kg BW of the patient
• Number of CD20+ cells ≤1 × 105 cells/kg BW of the patient
As noted above the target graft cell doses following processing with the CliniMACS
TCRα/β/CD19 System are defined as both, a CD34+CD45+ cell count of ≥4 × 106
cells/kg BW of the recipient and a TCRα/β+ cell count of ≤25 × 103 cells/kg BW of the
recipient. It is not permitted to exceed the target cell number of 25 × 103 TCRαβ+
cells/kg BW unless necessary to reach the target cell number of
≥4 × 106 CD34+CD45+ cells/kg BW. The graft may not contain >1 × 105 TCRα/β+
cells. For any graft with TCRα/β+ cell content above this limit, a part of the graft will
be retained to adjust such grafts to the maximal T cell content ≤1 × 105 TCRα/β+
cells/kg BW, without falling below the limit of 4 × 106 CD34+CD45+ cells/kg BW of the
patient.
While the main target is a total number of CD34+CD45+ cells of ≥4 × 106 cells/kg BW,
simultaneously a total number of TCRα/β+ cells of ≤25 × 103 cells/kg BW is aimed for.
Final 7.0 39 of 111 21. May 2015

If after 3 runs of stem cell apheresis and TCRα/β- and CD19-depletion a minimum
cell dose of 4 × 106 CD34+CD45+ cells/kg BW of the patient is not reached a second
cycle of mobilization and cell collection is allowed. The resulting second graft may be
transfused additionally 2–3 weeks after the initial transplantation.
Since the potential result of stem cell mobilization varies with the age of the donor
(more reliable result is expected with younger donors) processing of grafts depends
on the age of the donor.
• In general it is expected that in donors <50 years, grafts reach sufficient cell
counts. Therefore patient conditioning normally starts immediately upon
enrolment and transplantation of the IMP ‘TCRabCD19PBSC’ will be performed
immediately after stem cell apheresis and depletion as described below.
• If poor mobilization is expected (e.g. in donors ≥50 years) the grafts may be
cryopreserved and pooled prior to transplantation. Thus a sufficient
CD34+CD45+ cell count may be confirmed before the patient starts with
conditioning. In this case individual grafts resulting from maximum three
subsequent cycles of stem cell apheresis and cell depletion will be
cryopreserved and pooled (IMP ‘TCRabCD19PBSC_cryo). The patient will be
allowed to enter into treatment when sufficient CD34+CD45+ cell counts have
been confirmed by the subsequent analysis, only.
It is up to the responsible investigator to decide case by case, which is the most
appropriate approach.
In case clarification is needed the coordinating investigator or the responsible leading
investigator should be contacted.
All specification parameters for IMP release will be documented in the ‘certificate of
analysis’ which will be provided to the study centers together with the IMP. Beside
the numbers of CD34+CD45+-, TCRα/β+- and CD20+- cells these are:
• Number of CD3+ cells (total and per kg BW of the patient)
• Number of CD45+ cells (total and per kg BW of the patient)
• The percentage of recovered viable CD45+ cells after TCRα/β and CD19
depletion: target value ≥90%
• Haematocrit in mL/mL erythrocytes
• Result of visual control (bags undamaged, no cell aggregates visible).
Sterility will be asserted and documented but is not relevant for release, since the
tests can be completed after the time of transplantation, only.
Log depletion of TCRα/β+ and CD19+ cells and CD56+CD16+ cell counts for
additional evaluation of the graft, which are not parameters of drug specification, will
also be documented in the ‘certificate of analysis
Quality control will be monitored as described in section 9.6.
4.4.2. Packaging, Labeling and Storage
Packaging
The IMP ‘TCRabCD19PBSC’ will be delivered in primary bags which are packed for
transport in an outer package (sterile overwrap packaging). The IMP
‘TCRabCD19PBSC_cryo’ will be packaged in Cryobags. Hard-case aluminum
cassettes or equivalent packaging will be used for secondary packaging.
Labeling
Final 7.0 40 of 111 21. May 2015
The bags/packaging of the IMPs will be labeled in accordance with the applicable
regulatory guidelines (for labeling details see IMPD and Investigators File). Labels
will be in local language, i.e. in German for German sites and in Dutch for the Dutch
Site. An English label will be appended to the IMPD.
Storage
The IMP ‘TCRabCD19PBSC’ is intended for direct administration after completion of
the preparation process with a shelf-life of 72 h calculated from end of apheresis,
with storage at 4±2°C.
The IMP ‘TCRabCD19PBSC_cryo’ will be stored in the vapour phase over liquid
nitrogen at the manufacturing site (shelf-life of ≤1 year) and thawed at bedside for
direct application of the cells according to procedures in clinical routine.
Further details regarding packaging, labeling and storage of the investigational
product are described in the IMPD. Temperature during storage, both of IMP
‘TCRabCD19PBSC and IMP ‘TCRabCD19PBSC_cryo’, will be controlled and
documented.
4.4.3. Transport of Investigational Product

The transport of the stem cell apheresis product to the GMP Manufacturing site will
follow the local standard practice and SOPs developed by the GMP Manufacturing
sites. The transport of the IMP from the GMP Manufacturing site to the local clinical
site for transplantation will follow the local standards of clinical practice for the
transport of allogeneic blood stem cell transplants following legal requirements and
relevant guidelines.
4.4.4. Administration of Investigational Product

For transplantation patients will receive the hematopoietic stem cell grafts
intravenously on Day 0, Day 1 and Day 2 as appropriate to reach the CD34+CD45+
target cell count according to the respective center’s institutional guidelines for
methods of infusion (see section 5.1.3).
4.5. Compliance
The study treatment will be administered by the investigator. Therefore, patient
compliance with study treatment will not be monitored.
Patients that are non-compliant with the study protocol (e.g. non-attendance at study
visits or refusal to undergo certain assessments) may be excluded at the discretion of
the investigator or sponsor (see section 4.3.2).
4.6. Graft and Device Accountability

The investigator is responsible for maintaining accurate accountability records
throughout the study. The administration of the cell grafts will be documented in the
CRF and in the patient’s medical file. The investigator of a German site is responsible
for ensuring that all unused or partially used cell grafts will be disposed of according
to the German legal regulations (AMG) and the local regulations for biological
products. The investigator of a Dutch site is responsible for ensuring that all unused
of partially used cells grafts will be disposed of according to The Netherlands law on
Medical Research Involving Human Subjects (WMO) and the relevant local
regulations for cellular products (e.g. WBMV).
Final 7.0 41 of 111 21. May 2015

Any stem cell graft which will not be used during the study for transplantation will be
stored in case it is needed for later treatment of the patient in agreement with the
patient information and the informed consent. Upon death of the patient it will be
destroyed. The manufacturer has to ensure, that disposal will follow the German
(AMG) and the Netherlands (WMO, WBMV) legal regulations, as applicable, and all
relevant regulations for biological or for cellular products, as relevant.
5. STUDY PROCEDURES
5.1. Study Treatment Plan
All patients enrolled in the study will undergo hematopoietic stem cell transplantation
from a haploidentical family donor. Donors will have been treated for mobilization of
hematopoietic stem cells prior to PBSC collection by stem cell apheresis according to
local standards at their collection center. Prior to infusion the donor-derived PBSC
grafts will be depleted of TCRα/β positive and CD19 positive cells using the
CliniMACS TCRα/β/CD19 System (see section 4.4.1.). Patients will undergo non-
myeloablative chemotherapy to enable the engraftment of hematopoietic stem cells
prior to the intravenous infusion of the individual stem cell grafts. The exact regimen
used for this conditioning will depend on the underlying disease and the clinical state
of the patient (see section 5.1.2). GVHD prophylaxis with mofetil-mycophenolate until
Day 30 post-transplantation is planned for all patients.
Patients will have to attend 14 visits from the baseline visit to primary endpoint on
day 100, 3 visits in the follow-up phase I (until Day 365) and 1 visit in the follow-up
phase II 2 years after the transplantation. Patients will presumably be hospitalized
for 28 days post-transplantation. Donors will have to attend the center for one visit to
give their informed consent and for collection of a blood sample.
A data safety monitoring board (DSMB) will be responsible for the overall safety of
the patients in the study (for further details see section 9.4). A continuous safety
monitoring will be performed throughout the study for the parameters acute GVHD
grade III–IV and NRM until Day 100 post transplantation. Statistical stopping
guidelines have been defined to guarantee the patients’ safety in the study and to
allow for immediate reaction in case of elevated incidence rates (see section 7.10.1).
5.1.1. Mobilization and Collection of Donor PBSC

In this study, haploidentical family donors will donate peripheral blood stem cells
according to legal requirements and local institutional practice. Hematopoietic stem
cell mobilization and collection of peripheral blood by stem cell apheresis will be
performed according to relevant clinical and regulatory guidelines for collection and
transplantation of PBSC grafts. Since stem cell transplantation is the only potentially
curative therapeutic option for the critically ill patients in this study it is planned
independently of this study. All procedures related to donors apart from an additional
taking of a blood sample are therefore performed according to clinical routine and
independent of this study.
The target stem cell dose is ≥4 × 106 CD34+CD45+ cells/kg BW of the recipient with
≥95% viable CD34+CD45+ cells in the graft.
Depending on the recipient’s body weight, it is expected, that a total of 1 to 2 stem
cell apheresis runs of the donor for paediatric patients and 2–3 stem cell apheresis
Final 7.0 42 of 111 21. May 2015
runs of the donor for adult patients will be needed to obtain the target number of
CD34+CD45+ cells in the graft. In donors with poor mobilizing, in which the minimum
CD34+CD45+ cell dose of ≥4 × 106 cells/kg BW is not achieved after 3 stem cell
apheresis runs and the following TCRα/β- and CD19-depletion procedure, it is at the
discretion of the transplant center to continue with a second cycle of mobilization and
PBSC collection or to decide about the most appropriate approach for the respective
patient case by case. Irrespectively of this decision, all patients enrolled will be
followed and analyzed in an intent-to-treat manner. Graft preparation procedures are
described in section 4.4.
Note: For Patients with non-malignant diseases generation and
cryopreservation of an autologous graft prior to conditioning of the patient is
recommended in order to have - in the case of graft failure - a back-up
treatment available.
5.1.2. Conditioning Regimen for Patients

The conditioning regimen used will depend on the underlying disease and the clinical
state of the patients. Depending on the donor’s age and the intended use of IMP
‘TCRabCD19PBSC’ or ‘TCRabCD19PBSC_cryo’, patient conditioning will either be
started prior to mobilization treatment of the donor (transfusion of IMP
‘TCRabCD19PBSC’ intended: time schedule as described in Table 2 and Table 3) or
after the graft has been collected and a sufficient CD34+CD45+ cell count has been
confirmed (see section 4.4.1). In the latter case, ‘TCRabCD19PBSC_cryo’ will be
used and time schedules of donor and recipient will be independent of each other:
The donor will receive mobilization treatment and grafts will be prepared before the
corresponding patient will be started on the conditioning treatment.
Patients with immunodeficiencies and patients with haematological or non-
haematological malignancies who are in complete remission (Minimal Residual
Disease [MRD] burden ≤104 or <5% blasts) prior to transplantation will receive a
dose-reduced conditioning regimen of Fludarabine, Thiotepa, Melphalan and ATG
Fresenius S (Table 2).
Fludarabine, Thiotepa and Melphalan are approved drugs which have been
previously used in various conditioning regimens for allogeneic HCT. ATG Fresenius
S is an approved drug for prophylaxis against solid organ transplant rejection and
since 2011 in Germany approved in the indication ‘prophylaxis of graft-versus-host
disease for unrelated stem cell transplant donors in adults.’ In pediatric patients ATG
will be used off-label in this protocol (see drug information in IB). Prophylactically,
before application of ATG Fresenius S, anti-allergic prophylaxis with corticosteroids
(e.g. prednisolone, methylprednisolone) and antihistamines (e.g. ranitidine,
dimetidine) will be administered according to hospital routine.
Therapy-refractory patients with hematological malignant disease (incomplete
remission, MRD burden >104 or >5% blasts) and for patients with an increased
risk of graft failure (independent of their state of remission) defined as patients
with non-malignant diseases and chemotherapy naïve patients except of
patients with immunodeficiencies will receive either the same dose-reduced
conditioning regimen of Fludarabine, Thiotepa, Melphalan and ATG Fresenius S
(Table 2) as patients in CR or as an alternative, no ATG Fresenius S, but instead TNI
(7 Gy on Day –1) (Table 3) according to hospital routine.
The responsible investigator has to decide case by case, about the most appropriate
Final 7.0 43 of 111 21. May 2015

In case the patients’ body mass index is below 18.5 or above 25 the conditioning
regimen should be adjusted according to the body surface area of the patient
conferring to hospital routine.
Table 2: Conditioning regimen with the use of ATG Fresenius

Day
Drug Dose –12 –11 –10 –9 –8 –7 –6 –5 –4 –3 –2 –1 0 +1 +2
Patients
2
Fludarabine 1 × 40 mg/m X X X X
BS/d
IV, inf. 30 min
Thiotepa 2 × 5 mg/kg BW/d X
IV, inf. 4 h at 8 am
and 8 pm
2
Melphalan 1 × 70 mg/m X X
BS/d
IV bolus
Anti-allergic prophylaxis X X X X
ATG 1 mg/kg/d, IV X
Fresenius 9 mg/kg/d, IV X
10 mg/kg/d, IV X X
IMP transplantation X (X) (X)
(X): if necessary
Table 3: Conditioning regimen with the use of TNI
Day
Drug Dose –12 –11 –10 –9 –8 –7 –6 –5 –4 –3 –2 –1 0 +1 +2
Patients
2
Fludarabine 1 × 40 mg/m X X X X
BS/d
IV, inf. 30 min
Thiotepa 2 × 5 mg/kg BW/d X
IV, inf. 4 h at 8 am
and 8 pm
2
Melphalan 1 × 70 mg/m X X
BS/d
IV bolus
TNI 7 Gy X
IMP transplantation X (X) (X)
TNI=total nodal irradiation; (X): if necessary
Note: Graft failure will be analysed after treatment of the first 10 patients with
incomplete remission and with TNI instead of ATG Fresenius S. If graft failure
occurred in <3 of the 10 patients with this treatment regimen all patients enrolled
afterwards can be transplanted with TNI instead of ATG in the conditioning regimen
independent of being in complete or incomplete remission.
The responsible Investigator has to decide case by case, about the most appropriate
5.1.3. PBSC Transplantation

The individually manufactured IMP (released according to the IMPD) will be
administered intravenously on Day 0 to all patients according to individual
institutional guidelines after appropriate processing and quantification has been
Final 7.0 44 of 111 21. May 2015

performed by the GMP manufacturing site. Additional transfusions on Day +1 and

Day +2 will be performed as necessary.
The number of transfusions will depend on the number of individual stem cell
apheresis cycles needed to reach a sufficient content of ≥4 × 106/kg BW
CD34+CD45+ cells for transplantation. Each individual graft may be administered
either immediately after processing as IMP ‘TCRabCD19PBSC’ with up to three
transfusions on three subsequent days or may be cryopreserved after processing for
subsequent single transfusion of the pooled product.
GVHD prophylaxis has to be performed by administration of MMF (see section
5.1.4.1).
5.1.4. Prophylaxis, Supportive Care and Concomitant Treatments

Any clinically necessary treatment according to the investigator’s decision and
general or center specific guidelines and standards is permitted.
Concomitant Medication
At baseline and each subsequent visit until Day 100, patients will be asked what
medications they are currently taking. Any medication that the patient receives or
takes other than the study drug has to be recorded as concomitant medication,
including herbal and other non-traditional remedies. Standard prophylactic
medication as described below (see section 5.1.4.1) will be documented with generic
or trade name, indication, route of administration, start and stop dates and maximum
daily dose with unit of measurement from Day -12 to Day 100. (Dose changes have
to be documented.) All other concomitant medications from baseline to Day 100 have
to be documented in the eCRF with generic or trade name, indication, route of
administration, maximum and minimum dose including unit of measurement and start
and end dates (before study or date; ongoing or date). (Dose changes will not be
documented.)
During the follow-up phases, that is after Day 100 until the end of the study
concomitant treatment with other cellular products (as e.g. DLI or other white blood
cells) will be documented, only. Additionally, medication at the time of development
of a SAR will be documented in detail on the SAE form.
5.1.4.1. Prophylaxis
Anti-allergic Prophylaxis
Prophylactically, before application of ATG Fresenius S, anti-allergic prophylaxis with
corticosteroids (e.g. prednisolone, methylprednisolone) and antihistamines (e.g.
Ranitidin, Dimetidin) will be administered according to hospital routine.
GVHD Prophylaxis
All patients will receive MMF 2 × 20 mg/kg BW/d from Day –1 until Day 30 post-
transplantation.
Prophylaxis of Viral, Bacterial and Fungal Infections

Following prophylactic treatment starting with the first day of the conditioning is
strongly recommended.
Adult Patients
• Levofloxacin (Tavanic®) 500 mg PO (1-0-0) or 500 mg IV (1-0-0)
Final 7.0 45 of 111 21. May 2015

• Ambisome 1 mg/kg BW/d during the conditioning regimen until Day –1

• Caspofungin 50 mg/m2/d or ambisome 1 mg/kg BW/d or micafungin 1 mg/kg
BW/d from Day 0 until end of conditioning or oral medication possible.
• Posaconazol (Noxafil®) 3 × 200 mg/d PO or voriconazol (Vfend®) 2 × 200 mg/d
(1-0-1) until Day 100 or until CD4 count is >100/µl
• Acyclovir (Zovirax®) 1-2-1 tablets of 400 mg or 250 mg IV until Day 180 or until
CD4 count is> 100/µl
• Trimethoprim 160 mg + sulfamethoxazol 800 mg (Cotrim forte®), 3 × 1 tbl/week
(1-0-0) or pentamidine inhalation every 3 weeks
• Polyclonal immunoglobulin 15 g IV every 3 weeks until Day 100.
The prophylaxis with acyclovir, pyrethamine and cotrim should be continued for the
first year after transplantation. The dose of acyclovir can be reduced to 200 mg BID
after neutrophil recovery. Furthermore, prophylaxis against infectious diseases
should be adjusted to the clinical situation, and to drug toxicities, the occurrence of
GVHD and the necessity of immunosuppressive treatment.
Paediatric Patients
• Caspofungin 50 mg/m2/d or ambisome 1 mg/kg BW/d from Day +1 until
discharge, followed by:
• Posaconazol (Noxafil®) 3 × 4mg/kg PO or voriconazol (Vfend®) 2 × 7mg/kg PO
until Day 180 or until CD4 count is >100/µl
• Acyclovir (Zovirax®) 250mg/m2 IV until discharge, followed by acyclovir PO
40mg/kg/d until Day 180 or until CD4 count is >100/µl
• Trimethoprim 160 mg + sulfamethoxazol 800 mg (Cotrim forte®) at a dosage of
5mg/kg Trimethoprim/day divided in 2 doses, 3x week; IV administration until
discharge, then PO until day 180 or pentamidine inhalation every 3 weeks
• Polyclonal immunoglobulin 200 mg/kg BW IV every 3 weeks until Day 100
• Penicillin G 25,000 U/kg BW/d for 2 years.
Furthermore, prophylaxis against infectious diseases should be adjusted to the
clinical situation and to drug toxicities, the occurrence of GVHD and the necessity of
immunosuppressive treatment.
Suspected/Confirmed viral Reactivation: Treatment in adult and paediatric patients
If CMV-, EBV-, ADV-reactivation is suspected because of positive PCR or antibody
assays, specific preemptive antiviral therapy is strongly recommended:
• ADV in stool or blood: cidofovir 5 mg/kg BW every 2 weeks; in case of
increasing ADV copy numbers in blood donation of ADV specific donor T cells
• CMV reactivation: start preemptive therapy with therapeutic dose ganciclovir or
foscavir immediately after the first positive PCR. If CMV DNA will be detected in
urine or throat prior to transplant, eradication with ganciclovir or foscavir should
be tried before start of the conditioning regimen
• EBV in blood: in case of increasing copy numbers or >1000 copies EBV/µL
rituximab 375 mg/m2 and cidofovir 5 mg/kg BW every 2 weeks.
5.1.4.2. Supportive Care

Institutional standards for general supportive care after transplantation should be
maintained and should include antimicrobial agents (see above), nutritional support
and blood product support as necessary, if not otherwise indicated previously.
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Note: Nutritional support and supplements as well as fluid replacement/volume

substitution solutions given according to institutional guidelines will not be
documented as concomitant medication.
Venous Access
Patients will have an appropriate long-term central venous access placed, as detailed
by institutional standard practice, prior to beginning the conditioning regimen.
Blood Products
All blood products, except the infused TCRα/β- and CD19-cells depleted PBSC, will
be irradiated in accordance with institutional standards. Recipients who are CMV
negative will receive either CMV negative blood products or leukocyte depleted blood
products from study entry.
Nutrition
A low microbial diet will be maintained while the recipient is in isolation. Parenteral
nutrition will be initiated depending on the patient’s needs.
Isolation
Recipients will be maintained in single occupancy rooms with protective isolation per
5.1.4.3. Other Concomitant Treatments

AB0 Incompatibility
All patients with AB0 incompatibility to their donor should be evaluated and treated
according to the individual center’s standard procedure.
Caution: It should be noted that cases of haemolysis in the recipient have been
documented in patients with even minor AB0 mismatch to their donor. Therefore,
such patients should be monitored closely and be treated aggressively immediately
on detection of any evidence of haemolysis.
Administration of G-CSF
Generally it is highly recommended not to administer granulocyte colony stimulating
factor (G-CSF) e.g. neupogen to shorten duration of neutropenia after
conditioning.However, in some exceptional cases it might be duly justified to
administer G-CSF. In such cases the coordinating investigator or the respective
leading investigator should be consulted for advice regarding administration of G-
CSF.
In case G-CSF has been administered a justification should be recorded in the
eCRF.
Positive cross-match results:

It is highly recommended to treat patients with positive cross-match results for donor
reactive anti HLA antibodies (and without an alternative donor with negative cross-
match results) before stem cell transplantation according to hospital routine (e.g. with
rituximab, plasmapheresis) to improve the chances for successful donor engraftment.
In case such patients were not treated to remove HLA antibodies the rational should
be recorded in the eCRF.
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Second Transplantation with PBSC

In case decision for a second HSCT from the same or an alternative donor for the
treatment of graft failure or relapse has been made the reconditioning has to be
documented as Concomitant Treatment and the new graft has to be documented in
the eCRF Appendix, section “Transfusion/treatment with cellular products. Such
patients who receive a second HSCT should perform the respective early termination
visit (see 5.5 Early Termination Visit). The patient remains in the trial with only safety
reporting on visit XIV to XVIII, but no other trial procedures completed that means
only information regarding SAEs, NRM, survival and relapse has to be documented.
In addition these patients must continue to be followed up for SAEs until the safety
follow-up visit after 2 years (see section 6.3.2).
Transfusions
Leukocyte-poor platelets will be administered when the platelet count is <10 × 109/L
or per institutional guidelines. If the platelets are refractory as documented by lacking
sequential platelet count rise within 30 min post platelet transfusion, platelets will be
administered only in case of active bleeding. Platelets must be transfused upon any
active bleeding. Packed red cell transfusions are recommended when haemoglobin
is <8 mg/dL or as clinically indicated.
Febrile Neutropenia
Broad-spectrum antibiotics will be administered intravenously according to
Management of bacterial infections
Antibiotic treatments will be administered per institutional guidelines.
Management of PTLD
Treatment of PTLD will be performed according to institutional standards at the
discretion of the treating physician.
Management of Acute GVHD
Recipients who develop acute GVHD will be treated according to local institutional
standards at the investigator’s discretion.
Management of Graft Failure
Primary or secondary graft failure as judged by neutrophil counts will be considered a
treatment failure. Patients will be treated according to institutional guidelines at the
investigator’s discretion. Management of graft failure should be discussed with the
coordinating investigator or the respective leading investigator.
5.1.4.3.1. Enrolment in Other Clinical Trials

All patients undergoing allogeneic hematopietic stem cell transplantation are at risk of
the same complications, namely GVHD, infection and relapse. Regardless of risk
classification and the underlying disease of the patient at the time of study enrolment
these complications may require additional experimental treatment after infusion of
the IMP. Patients with these complications are often not suitable for standard therapy
or unlikely to benefit from it, therefore enrolment in appropriate clinical studies might
be required according to the investigator’s judgment. Thus, inclusion in another
clinical trial after the primary endpoint of this study is reached (i.e. after Day 100) will
be allowed for indications described in the following. Enrollment in any other clinical
study up to Day 100 however, is not allowed according to this study protocol.
Final 7.0 48 of 111 21. May 2015

Patients, who will be included in an other study protocol before the primary endpoint
at Day 100 is reached will be excluded from the ‘per protocol analysis’.
Note: Enrolment in another clinical trial during the follow-up period of this study (up
to 2 years after infusion of the IMP) will be documented in the patient’s eCRF.
Treatment of Relapse/ Progression post Transplantation

The management and treatment of relapse or progression of disease is a significant
clinical problem following allogeneic hematopoietic stem cell transplantation but
therapeutic options are extremely limited. Where established standard therapies are
not available patients with recurrence of the original malignancy or disease
progression after infusion of the IMP will be treated according to individual treatment
decision.
Thus, patients with relapse (including molecular relapse, MRD+) as well as patients
with progression of the underlying disease, who are no longer eligible for approved
therapies are allowed to be included in appropriate clinical studies after the primary
endpoint of the present study has been reached (i.e., after Day 100), provided the
new study does not interfere with the endpoints of the present study according to the
investigator’s judgment. In all instances, treatment of relapse and progression and
selection of the clinical study will follow local guidelines of the study center and
national/international guidelines/recommendations as applicable.
Treatment of Chronic GVHD post Transplantation

While first-line therapy of chronic GVHD is based on randomized trials and consists
of prednisone with or without a calcineurin inhibitor, optimal secondary treatment has
not yet been established. Therefore, patients with steroid-refractory chronic GVHD
will be allowed to be included in appropriate clinical studies after the primary endpoint
of the this study has been reached (after Day 100) provided the new study does not
interfere with the endpoints of the present study according to the investigator’s
judgment.
Treatment of GVHD and selection of an appropriate clinical study will be performed
according to the guidelines of the local study center. It should be considered that
according to the recommendations of the consensus conference in clinical practice in
chronic GVHD initial secondary treatment should include agents with an adequate
safety profile and well-documented activity, whereas agents with significant side
effects should be reserved to third- or fourth-line treatment (79).
Treatment of Infections post Transplantation

Infections—bacterial, viral, fungal and parasitic—continue to be a substantial cause
of death after allogeneic hematopoietic stem cell transplantation. Novel approaches
are still required since standard therapies have not been established to date.
Accordingly, patients suffering from infections and failing standard therapy will be
allowed to be included in appropriate clinical studies after the primary endpoint of the
present study has been reached (after Day 100) provided the new study does not
interfere with the endpoints of the present study according to the investigator’s
judgment. Treatment of infections and selection of an appropriate clinical study will
follow local guidelines of the study center and the recommendations of the European
Conference on Infections in Leukemia
Final 7.0 49 of 111 21. May 2015

(http://www.leukemianet.org/content/treat_research/supportive_care/standards_sop_
and_recommendations/index_eng.html).
5.2. Study Assessments

5.2.1. Evaluation of Grafts
The cell content of PBSC grafts (the individual IMPs) after TCRα/β and CD19
depletion will be analysed to ensure the quality of the graft for transplantation and to
evaluate the performance of the CliniMACS TCRα/β/CD19 System. The following
parameters will be assessed:
• The percentage of recovered viable CD34+CD45+ cells after TCRα/β and CD19
depletion procedure: target value ≥95%.
• Log Depletion of TCRα/β+ cells
• Log Depletion of CD19+ cells
• Cell counts: CD34+CD45+ stem cells, CD20+ B cells, CD56+CD16+ NK cells,
TCRα/β+ and TCRγ/δ+ cells, CD3+ cells and CD45+/WBC cells analysed by flow
cytometry after processing prior to transplantation
• The percentage of recovered viable CD45+ cells after TCRα/β and CD19
depletion procedure: target value ≥90%
• Haematocrit value in graft in mL/mL erythrocytes
• Number of grafts containing ≥4 × 106 CD34+CD45+ cells/kg BW of the recipient
• Number of grafts containing ≤25 × 103 TCRα/β cells/kg BW of the recipient
• Number of grafts containing ≤1 × 105 CD20+ cells/kg BW of the recipient.
Additional parameters for IMP specification documented in the ‘certificate of analysis’
(see section 4.4.1) are
• Visual control (bags undamaged, no cell aggregates visible) and
• Sterility (assessed but not relevant for IMP release).
5.2.2. Evaluation of Donors
The donor will have been assessed regarding medical eligibility for stem cell donation
prior to and independent of all study procedures according to guidelines and
standards for stem cell donation. This will have been done at the individual collection
center. Furthermore, donors will have given their informed consent for stem cell
donation and all procedures related to it prior to and independent of the inclusion of
the respective patient (recipient) in this present study. Donors will be allocated an
individual donor ID by the collection center according to local standards.
Some of the data documented at the collection center during evaluation for stem cell
donation will be transferred to the recipient’s eCRF. Data transferred will comprise
• Donor ID
• Demographic characteristics (month and year of birth, relation to the patient,
ethnic origin, gender)
• Results of routine laboratory tests (see below for details): serology, PCR
analysis of virology, AB0 Rh blood typing and HLA-typing.
Prior to the start of conditioning (Visit II) of therespective patient the donor will be
asked to give his/her study related written informed consent and to visit the
transplantation center for one baseline visit. During this visit the study specific blood
samples will be collected (see below). No study specific procedures concerning the
Final 7.0 50 of 111 21. May 2015
donor including transfer of previously collected data will be performed without written
informed consent of the donor.
Blood sampling
The following blood samples will be collected at the transplantation center after the
donor’s signing of the study specific informed consent:
• 0.5 mL EDTA blood for DNA analysis (sample for KIR-Genotyping)
• 1.3 mL EDTA blood as reference sample for immunophenotyping to enable
differentiation between host and donor cells after transplantation
• 15 mL EDTA blood as reference sample for analysis of the TCR activity
In total, 17 mL of blood will be collected from the donor at the baseline visit.
Printed labels including all necessary information needed for clear and unique
identification of each sample will be provided.
Sample shipment
For sample handling and shipment please refer to the Laboratory Manual.
Assays (Additional Central Assessments)

Central Assessments of KIR genotyping and TCR activity will be performed at
Miltenyi Biotec GmbH (Germany).
Central Assessments of TCR spectratyping and TREC analysis will be perfomed at
the University Children’s Hospital Würzburg (Germany).
Immune cell phenotyping will be performed in the related GMP-laboratory (see
Appendix 7).
5.2.2.1. Donor Baseline Laboratory Data

The following data have to be entered in the eCRF for baseline evaluation of the
donor, as possible:
I. Serology
Titers assessed as positive or negative for each parameter.
• Anti HIV1/2 antibodies
• HBs-antigen
• Anti-HBc-IgG
• Anti HCV antibodies
• CMV antibodies
• Varicella zoster virus antibodies
• EBV antibodies
• HHV-6-antibodies
• HTLV 1/2 antibodies
• Treponema pallidum antibodies
• Toxoplasmosis antibodies.
II. PCR analyses
• HIV (blood)
• HCV (blood)
III. AB0 Rh blood typing and HLA-typing
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5.2.3. Evaluation of Patients

5.2.3.1. Examinations
The following examinations have to be performed as scheduled in section 5.3.4.
I. Demographic characteristics
Demographic details (height in cm, weight in kg, date of birth, ethnic origin, gender)
will be documented for each patient at baseline.
II. Medical history
The patient’s general medical history (‘medical history other than underlying
disease’) will be assessed and documented in the eCRF at baseline. Start and stop
date of any medically relevant disease and ongoing disease have to be stated.
Investigator’s will be asked to assess the clinical significance of ongoing diseases in
the context of the present study.
Note: If a condition is assessed to be ‘compromising participation in this study’ the
patient has to be rated as meeting exclusion criterion 14 and may not be enrolled!
The underlying disease will be specified as described in inclusion criterion 2. For
patients suffering from malignancies, information about previous relapses should be
documented.
Medically relevant previous therapies for diseases other than the underlying
disease will be documented at baseline with indication, generic or trade name, start
and stop date or ongoing, and current dose if ongoing. Furthermore, previous
therapies of the underlying disease will be documented as previous stem cell
transplantation, previous radio-/chemotherapy (chemotherapy: regimen and number
of cycles, radiotherapy: dose and start/stop date).
III. Physical examination
Physical examination of patients will be performed at each subsequent visit until
study end (Visit XVIII, Month 24). Results will be documented in the eCRF per body
system (general appearance, eyes, ENT, respiratory, cardiovascular, gastrointestinal,
urogenital, musculoskeletal/connective tissue, skin/mucosal, lymphatic and nervous
system) and pathological findings will be recorded. Patient’s performance indices
Karnofsky (adult patients, >16 years; [47]) or Lansky (paediatric patients, ≤16 years;
[52]) will be assessed.
IV. Vital signs
The following vital signs will be assessed and documented in the eCRF for patients
at each visit until study end as described in the patients’ study flow-chart (see Table
6).
• Measurement of supine systolic and diastolic blood pressure in mm Hg
• Resting heart rate in beats/min
• Body temperature in °C (aural)
• Body weight in kg
V. Additional examinations as appropriate at baseline
• Ultrasound examination of abdomen
• CT scan of chest
• ECG
• Echocardiography
• Lumbar puncture
Final 7.0 52 of 111 21. May 2015
• In case of neurological impairment, HLH, malignant osteopetrosis, previous

heavy chemotherapy: CNS MRI and neurological consultation.
Note: Results of additional examinations obtained up to 3 months prior to the
baseline assessments may be used according to the investigator’s judgment.
VI. Evaluation of Sorror Comorbidity Index [72] at baseline

(adult patients, only)
VII. Disease Status and Staging
The clinical state of patients pre- and post-transplantation will be determined
according to current international recommendations for the malignant indications
analysed in this study. The underlying malignancy will be assessed by the
investigator according to current international guidelines and documented in the
eCRF at baseline, Day 28, Day 100 and Month 12. Remission will be defined as
complete remission, partial remission, unchanged and progressive disease.
• Hematological malignancies will be assessed by laboratory staging, see below
section 5.2.3.2, VIII.
Definition of ‘complete remission’ (CR) and ‘partial remission’ (PR): Complete
remission (CR, morphologic and cytogenetic as appropriate) and partial
remission (PR) will be defined according to current international
recommendations and guidelines for the indications analysed in this study and
assessed by the investigator.
• All other malignant diseases will be staged according to the relevant inter-
national guidelines.
5.2.3.2. Laboratory examinations

Laboratory standard of care assessments (see below, I.–IX.) are routine parameters
for evaluation of patients prior to and post haploidentical stem cell transplantation
according to institutional and regulatory guidelines. These parameters will be
analysed at the respective certified local clinical laboratory normally used by each
study site. The analysis will be performed according to the local instructions and
guidelines.
Before starting the study, every investigator will supply the Sponsor with a list of
normal ranges and units of measurement and also with laboratory certificates.
Additional study-specific laboratory assessments of outcome parameters will be
performed centrally (see below, X.).
I. Small Panel
Laboratory analyses of the ‘small panel’ will be performed at each visit from Day 7 to
Day 100 (Visit IV to Visit XIV). They comprise the following parameters:
Complete blood count
• Haemoglobin
• Hematocrit
• Erythrocytes
• Leukocytes
• Thrombocytes
Differential Blood count
Substrates
Final 7.0 53 of 111 21. May 2015
• Total bilirubin
• Creatinine
• Urea
• Total protein
• C-reactive protein (CRP)
• Uric acid
Enzymes
• Alanine-aminotransferase (ALAT)
• Aspartate-aminotransferase (ASAT)
• Alkaline phosphatase (AP)
The laboratory parameters of the small panel may or may not be part of the standard
post-transplant monitoring at the site. However, all lab parameters of the small panel
must be perfomed as specified above for all patients enrolled in TCRalpha/beta-
Haplo2010 trial.
Note: It is expected that due to the conditioning therapy and after hematopoietic
stem cell transplantation laboratory values of the small panel will be outside the
normal ranges for an extended period of time. This is no study-specific feature but
expected consequence of any stem cell transplantation therapy. All abnormal
laboratory values have to be assessed by the investigator regarding clinical
significance in this context. All abnormal laboratory values, which according to the
investigator’s judgment are clinically significant in this specific therapeutic setting
have to be recorded as AEs.
II. Daily Panel

Laboratory analyses of the ‘daily panel’ will be performed daily from Day +4 to
Day 28 or the day of engraftment, whatever comes first, and at each subsequent visit
from Day 35 to Day 100 (Visit VIII to Visit XIV). If engraftment is prior to day 28,
laboratory assessments will be performed as per hospital routine until day 28. The
‘daily panel’ comprises the following three parameters:
• Leukocytes
• Thrombocytes
• Neutrophils
Note: It is expected that due to the conditioning therapy and after hematopoietic
stem cell transplantation laboratory values of the daily panel will be outside the
normal ranges for an extended period of time. This is no study-specific feature but
expected consequence of any stem cell transplantation therapy. All abnormal
laboratory values have to be assessed by the investigator regarding clinical
significance in this context. All abnormal laboratory values, which according to the
investigator’s judgment are clinically significant in this specific therapeutic setting
have to be recorded as AEs.
III. Big Panel

Laboratory analyses of the ‘big panel’ will be performed at baseline. They comprise
all analysis of the ‘small panel’ and the ‘daily panel’ and additionally:
Hematology
• Differential count for number of blasts (as applicable)
Serum pregnancy test in women of childbearing potential (β-HCG)
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The observations listed in the big panel may or may not be part of the standard pre-
transplant evaluation at the site; however, all laboratory parameters of the big panel
must be performed as specified above for patients enrolled in TCRalpha/beta-
Haplo2010 trial.
IV. Special Laboratory Analysis
Special analysis will be performed at Day 100 for paediatric patients, only. They
comprise:
• TSH
• fT3
• fT4
• Growth hormone
V. Serology
Serology (IgG) will be analysed at baseline, only. Previous results are accepted, if
obtained within 4 weeks prior to start of conditioning. Parameters will be assessed as
positive or negative.
• Anti HIV1/2 antibodies
• HBs-antigen
• Anti-HBc-IgG
• Anti HCV antibodies
• CMV antibodies
• Varicella zoster virus antibodies
• EBV antibodies
• HHV-6-antibodies
• HTLV 1/2 antibodies
• Treponema pallidum antibodies
• Toxoplasmosis antibodies.
Note: To be omitted in patients suffering from SCID or related diseases. Direct
detection methods may be used instead as per hospital routine. Positive results are
to be documented under "Other Disease -History" in the eCRF.
VI. PCR analyses

PCR analysis for fungal and viral reactivation/infection will be performed at baseline,
once to twice weekly from Day +4 to Day +28, weekly until Day 70 and at Day 100
(Visit VIII to Visit XIV). Additional PCRs should be performed after day 100 as per
hospital routine, until a cell count of 250 CD4+ cells/µl is reached.
• HIV (blood; at baseline, only)
• HCV (blood; at baseline, only)
• Aspergillus (blood): PCR or serology may be performed as per hospital routine
• CMV (blood). CMV in throat swab, urine and quantitative PCR in blood as per
hospital routine.
• ADV (blood, stool) mandatory for pediatric patients only. In adult patients as per
hospital routine
• EBV (blood; if positive, also quantitative PCR in blood)
• In case of fever quantitative PCR of VZV, HSV and of EBV is recommended.
VII. AB0 Rh blood typing and HLA-typing and cross matching (at baseline,
only)
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VIII. Laboratory staging (hematological diseases, only)

Staging and cytogenetic complete remission (CR) will be defined according to current
international recommendations and guidelines for the hematological indications
analysed in this study and assessed by the investigator. Laboratory staging will be
assessed based on results of
• Pathology, flow cytometry, cytogenetics, MRD
• Bone marrow aspirate and biopsy.
IX. Chimerism
Chimerism will be assessed at each study center’s local laboratory by PCR-analysis
of blood samples at each visit after transplantation until completion of follow-up
phase I (12 month follow-up), starting on the day where clinically relevant results are
expected by the treating physician and of bone marrow samples from Days 28 and
100 of patients with haematological malignancies compared to original samples from
donors and recipients prior to transplantation
• PCR peripheral blood (PB),
• PCR bone marrow biopsy (BM)
X. Immune reconstitution of T, B, NK-cells

Immune cell phenotyping will be performed according to institutional practice and will
be recorded in the eCRF at days 7, 14, 21, 28, 35, 42, 49, 56, 63, 70, 100, Month 6,
9 and 12 post-transplantation, if available.
5.2.3.3. Collection of blood samples for transplantation related evaluations

Standard of Care
Investigations which will be performed as standard of care investigations have been
described in detail above (see section 5.2.3.2).
Prior to conditioning and at defined visits (see study flow-chart, (Table 6) blood
samples and samples of bone marrow aspirate have to be collected for routine
monitoring of patients after blood stem cell transplantation. The approximate blood
volumes needed for these routine analyses are summarized in
Table 4.
Table 4: Blood and bone marrow sampling for standard of care routine monitoring,
performed at the local laboratories of the hospital
Volume
Parameter see collected
EDTA blood
a
CBC and hematology section 5.2.3.2, I., II. , III. 2 mL
Viral infection: PCR analysis section 5.2.3.2, VI. 10 mL
Analysis of chimerism section 5.2.3.2 IX 10 mL
Heparinized blood
a
Clinical chemistry section 5.2.3.2 I., II. , III. 2×10 mL
Serum
AB0 Rh blood group, HLA typing and cross-matching section 5.2.3.2, VII. 10 mL
Special laboratory analyses (paediatric patients) section 5.2.3.2, IV. 2.7 mL
Serology section 5.2.3.2, V. 10 mL
Citrate blood
a
Coagulation section 5.2.3.2, I., II. , III. 10 mL
Final 7.0 56 of 111 21. May 2015

Volume
Parameter see collected
Bone marrow (EDTA)
Analysis of chimerism, staging section 5.2.3.2, VIII., IX. 5 mL
section 5.2.6, XI.
Biopsy
Staging section 5.2.3.2, VIII. 5 mL
Additional central assessments

Investigations which will be performed centrally as additional assessments are
described in detail below (see section 5.2.6).
Prior to conditioning and at defined visits (see study flow-chart, Table 6) blood
samples have to be collected for additional monitoring of patients after blood stem
cell transplantation (immune cell phenotyping and analyses of immune recovery).
The approximate blood volumes needed for these additional analyses are
summarized in Table 5.
Table 5: Blood and bone marrow sampling for additional assessments: see section
5.2.3.2, X.
Volume
Parameter Analysis at collected
EDTA blood
Immune cell phenotyping GMP laboratory (Appendix 8) 1.3 mL
TCR Vbeta spectratyping University Children’s Hospital Würzburg 3 mL
TCR Vgamma/delta spetratyping University Children’s Hospital Würzburg 3 mL
TREC analysis (thymic function) University Children’s Hospital Würzburg 1 mL
KIR genotyping Miltenyi Biotec GmbH laboratory 0.5 mL
NK cell KIR repertoire: FACS analysis Miltenyi Biotec GmbH laboratory 6 mL
NK cell activity Miltenyi Biotec GmbH laboratory 12 mL
T cell activity: Virus-specific T cells Miltenyi Biotec GmbH laboratory 23 mL
T cell activity: T cell stimulation/proliferation Miltenyi Biotec GmbH laboratory 20 mL
5.2.4. Adverse Events

For definition of Adverse Events (AEs) please refer to Chapter 6. The methods for
assessing safety parameters consist of clinical routine methods (physical
examination, vital signs, laboratory and clinical evaluation) and measures for
observation of patients after haploidentical stem cell transplantation according to
institutional and regulatory guidelines. Laboratory testing will include analysis for
recurrence or newly occurring infectious diseases caused by cytomegalovirus (CMV),
adenovirus (ADV) and Epstein-Barr virus (EBV) and aspergillus.
Therapy-related toxicities are expected during the conditioning period and will be
rated as known and unknown therapy-related toxicity. Unknown therapy-related
toxicities will have to be documented as adverse event as described in section 6.1.1.
From Day 0 onwards no further distinction between known and unknown therapy-
related toxicities will be made. All Adverse events (AEs) will be documented from
Day 0 to Day 100. Serious adverse events (SAEs) as defined in section 6.1 will be
documented for patients in the eCRF from the baseline visit until the end of study.
AEs will be recorded in the eCRF and SAEs will be recorded on special SAE case
report forms on paper.
Safety outcome parameters (GVHD, NRM and infusional toxicity) will be assessed as
described below (see section 5.2.6).
Final 7.0 57 of 111 21. May 2015
5.2.5. Concomitant medication

Concomitant medication will be recorded from baseline until Day 100. During the
follow-up phases concomitant medication will be recorded on the SAE form in case of
an SAR. Furthermore, new treatment with cellular products as e.g. DLI or other white
blood cells as well as inclusion in another clinical study will be documented from
Day 100 until the end of the study.
5.2.6. Outcome Parameters

Safety/tolerability, feasibility and other outcome parameters are defined and will be
measured as described in the following:
Safety parameters
I. Graft-versus-host disease (GVHD):
• Incidence of acute GVHD grades II–IV on Day 100 post-transplantation is
defined as primary study objective. It will be expressed as ‘incidence of acute
GVHD grades II–IV’ and ‘time to occurrence of acute GVHD grades II–IV’.
• Incidence of acute GVHD grade I on Day 100 post-transplantation is defined
as secondary study objective. It will be expressed as ‘incidence of acute
GVHD grade I’ and ‘time to occurrence of acute GVHD grade I’.
• Incidence and severity of chronic GVHD after 1 and 2 years.
Statistical stopping guidelines referring to incidence of acute GVHD within
100 days after SCT have been defined to ensure patients’ safety throughout the
study (see section 7.10.1).
• Incidence and severity of acute GVHD will be graded according to the
Glucksberg Criteria (Appendix 1: Glucksberg Criteria for grading of acute
GVHD)
• Incidence and severity of chronic GVHD will be graded according to standard
criteria for grading of chronic GvHD (Appendix 2: Grading of chronic GVHD).
Note: Whenever possible, the diagnosis of GVHD should be confirmed by biopsy
of the organ involved and histological examination of the biopsy specimen by a
pathologist experienced in the diagnosis of GVHD.
II. Non-Relapse-Mortality (NRM):

NRM is defined as death occurring in a patient between day of first
transplantation (Day 0) and day of assessment (all visits throughout the study),
not due to disease relapse/recurrence. Statistical stopping guidelines referring
to incidence of NRM within 100 days after SCT have been defined to ensure
patients’ safety (see section 7.10.1).
III. Infusional toxicity:
Maximum infusional toxicity on the days of transfusion will be evaluated by
measuring the patient’s blood pressure, heart rate, respiration rate and
temperature one hour prior to the allograft infusion and then approximately 15
minutes, 30 minutes, 2 hours, and 4 hours post infusion.
IV. Graft failure:
Primary graft failure is defined as the failure to achieve an ANC >500 cells/µL by
Day +28. Secondary graft failure is defined as initial neutrophil engraftment
followed by subsequent decline in neutrophil counts <500 cells/µL, unresponsive
to growth factor therapy.
Feasibility parameters
Final 7.0 58 of 111 21. May 2015

V. Engraftment:
• Neutrophil cell counts will be determined by flow cytometry and time to
neutrophil engraftment will be measured by determining the first of three
consecutive measurements of ANC ≥500/µL following conditioning regimen
induced nadir, starting from the day of the first stem cell transplantation
(Day 0) until Day 28.
• Platelet counts will be determined by flow cytometry and time to platelet
engraftment will be measured by determining the first of three consecutive
measurements of platelet count ≥20,000/µL without platelet transfusion
support for seven days, starting from the day of the first stem cell
transplantation (Day 0) until Day 28.
VI. Survival:
• Overall survival rate (OS) is defined as time from transplantation to death or
last follow-up and will be assessed at Day 100 and after 1 year and 2 years.
• Disease-free survival (DFS) is defined as the minimum time to relapse/
recurrence, to death or to the last follow-up, from the time of transplantation
and will be assessed at Day 100 and after 1 year and 2 years.
VII. Transfusion requirement:
• Number of thrombocyte infusions needed after transplantation and time to
last thrombocyte infusion starting from Day 0 until Day 100.
• Number of erythrocyte infusions needed after transplantation and time to last
erythrocyte infusion starting from Day 0 until Day 100.
• Number of infusions of other blood products needed after transplantation
and time to last infusion of other blood products starting from Day 0 until
Day 100.
VIII. Relapse rate:
The number of patients with relapse as defined below at the time of assessment
(Day 100 and after 1 year and 2 years) will be assessed. Time to relapse will be
calculated from the time of transplantation to evidence of relapse as defined
above.
Definition of Relapse (Morphologic and Cytogenetic)
Relapse will be defined according to current international recommendations and
guidelines for the malignant indications analysed in this study and will be
assessed by the investigator (compare section 5.2.3.1 VI and section 5.2.3.2 VIII:
staging).
IX. Hospitalization/rehospitalization:
Number of days that patients had to be hospitalized until discharge after
transplantation, and after any subsequent occurrence of an event leading to
rehospitalization assessed at Day 28 and Day 100.
X. Quality of life:
Patients will be asked to answer EQ-5D [60] (patients ≥18 years) or PedsQL
(patients <18 years) and FACT-BMT (patients ≥18 years) quality of life
questionnaires at baseline, at Day 100 and after 1 year and 2 years. Results will
be assessed by determining respective total score values.
Laboratory parameters (routine local assessments)
XI. Cell chimerism after transplantation
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The percentage of donor cells present in peripheral blood at days 7, 14, 21, 28,
35, 42, 49, 56, 63, 70, 100, Month 6, 9 and 12 post-transplantation, and in bone
marrow at Days 28 and 100 of patients with haematological malignancies.
Laboratory parameters (additional central assessments)
XII. Immune cell phenotyping (analysis at GMP laboratory)
Immune cell phenotyping will be performed at the central GMP Laboratory
(Appendix 7).
Immune cell phenotyping of T, B, NK and T regulatory (Treg) cell subsets will be
analysed by cell counting of CD3+, CD4+, CD8+, CD3+CD56+, CD3+TCRα/β+,
CD3+TCRγ/δ+ T cells, naive CD4+TCRα/β+, memory CD4+TCRα/β+, naive
CD8+TCRα/β+, memory CD8+TCRα/β+, DN TCRα/β+, TCR Vd2+TCRγ/δ+, TCR
Vd2–TCRγ/δ+, naive Tregs, ‘memory’ Tregs, B cells, monocytes, NK cells,
neutrophils, eosinophils on Days 7, 14, 21, 28, 63, 100 and Month 6 and 12 post-
transplantation.
XIII. Reconstitution of immune system (analysis at central laboratory)
The following parameters will be analysed centrally at Miltenyi Biotec GmbH
laboratory or at the University Children’s Hospital Würzburg.
• Reconstitution of T Vbeta repertoire by TCR V beta spectratyping (PCR
analysis) on Days 28, 63 and 100 post-transplantation; at University Children’s
Hospital Würzburg
• Reconstitution of T Vgamma/delta reptoire by TCR V gamma/delta spectra-
typing on Days 28, 63 and 100 post-transplantation; at University Hospital
Würzburg
• Thymic function by TREC analysis (PCR analysis), samples collected on
Days 28, 63 and 100 post-transplantation; at University Children’s Hospital
Würzburg
• KIR genotyping of donor and recipient from baseline sample; at Miltenyi Biotec
GmbH
• Reconstitution of NK cell KIR repertoire by FACS analysis, samples collected
on Days 28, 63 and 100 post-transplantation; at Miltenyi Biotec GmbH
• Analysis of T cell activity by analysis for virus-specific T cells reactive against
CMV, EBV and ADV and by analysis of T cell stimulation and proliferation,
samples collected on Days 28, 63 and 100 post-transplantation; at Miltenyi
Biotec GmbH
• Analysis of NK cell activity, samples collected on Days 28, 63 and 100 post-
transplantation; at Miltenyi Biotec GmbH.
Printed labels including all necessary information needed for clear and unique
identification of each sample will be provided.
Sample shipment
For sample handling and shipment please refer to the Laboratory Manual.
5.3. Study Visits Schedule

The present study is an open label, prospective, single arm, multicenter phase I/II
clinical study investigating the effects of TCRα/β and CD19 depleted hematopoietic
stem cell grafts from G-CSF mobilized donors in the treatment of adult and paediatric
Final 7.0 60 of 111 21. May 2015
patients suffering from hematological and non-hematological malignancies with high-

risk of relapse or non-malignant diseases and with an indication for PBSC
transplantation. Patients will receive the TCRα/β and CD19 depleted PBSC grafts as
stem cell transfusion.
Patients will be required to attend 18 visits during the study: 14 visits until Primary
Endpoint and 4 visits after Primary Endpoint (follow-up phase I) until the final follow-
up visit two years after study enrolment (follow-up phase II).
Donors will have to come to the transplantation center for one visit.
5.3.1. Donors’ visit schedule

Baseline Visit
Donors will have to visit the transplantation center at least once to give their study
specific informed consent and for collection of study-specific reference blood
samples (see section 5.2.2). This visit might be splitted as necessary. The blood
samples should be collected prior to the first administration of medication for stem
cell mobilization at the collection center, if possible, but latest until start of apheresis.
(Note: treatment with stem cell mobilizing medication and apheresis will be
performed according to medical routine for stem cell donation and not part of the
study-specific procedures).
5.3.2. Patients’ visits schedule

Baseline (within four weeks prior to start of conditioning, Day –40 to Day –14)
• Visit I
Pre-transplantation (conditioning; Day –12 to –1)
• Visit II
Transplantation (Day 0 to +2)
• Visit IIIa (Day 0): First transplantation
• Visit IIIb (Day 1): Second transplantation, if necessary
• Visit IIIc (Day 2): Third transplantation, if necessary
Post-transplantation phase I: after IMP infusion (Day +1 to +28, ±2 days)
• Visit IV: Day +7 after first IMP infusion
• Visit V: Day +14 after first IMP infusion
• Visit VI: Day +21 after first IMP infusion
• Visit VII: Day +28 after first IMP infusion
Post-transplantation phase II: after Visit VII to primary study endpoint at Day +100
(Day +35 to +70, ±2 days; Day +100 ±5 days)
• Visit VIII: Day +35 after first IMP infusion
• Visit IX: Day +42 after first IMP infusion
• Visit X: Day +49 after first IMP infusion
• Visit XI: Day +56 after first IMP infusion
• Visit XII: Day +63 after first IMP infusion
• Visit XIII: Day +70 after first IMP infusion
• Visit XIV: Day +100 after first IMP infusion, day of primary study
endpoint
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Follow-up phase I: after Day 100 to 12 months (, ±2 weeks)

• Visit XV: +6 months after first IMP infusion
• Visit XVI: +9 months after first IMP infusion
• Visit XVII: +12 months after first IMP infusion
Follow-up phase II: after 24 months (±4 weeks)
• Additional follow-up visit at Month 24 after first IMP infusion (after data closure,
Visit XVIII).
5.3.3. Donors’ Assessments

Baseline visit
Donors will be informed about the study at the baseline visit; this will be recorded and
documented appropriately. Study specific written informed consent has to be
obtained at this visit, date of consent will be recorded in the eCRF. No study related
procedures will be performed before written consent has been obtained.
The baseline visit has to be scheduled prior to start of conditioning (Visit II) of the
patient (see above). After obtaining written informed consent and confirmation of in-
and exclusion criteria, samples of peripheral blood will be collected as specified
above (see section 5.2.2). The samples will be processed and stored under
appropriate conditions, until shipment to Miltenyi Biotec GmbH, Germany, for
subsequent analysis (for sample handling, please refer to the Laboratory Manual).
5.3.4. Patients’ Assessments by Visit

Physical condition including vital signs and Karnofsky and Lansky index, acute
GVHD, NRM, infections and other adverse events and concomitant medication will
be assessed post-transplantation continuously throughout the study until Day 100.
Documentation will include all serious adverse events during the entire study period
from baseline onwards.
Patients’ assessments are summarized in Table 6.
Patients will be handed a study patient card (‘Studienpass’) at hospital discharge
(Visit VII) which will provide additional information and will lead them through post-
transplantation phase II and the follow-up phases of the study.
Visit I: Baseline Assessments (once, optionally at Day –40 to Day –14)

Baseline assessments may be performed up to 4 weeks prior to start of conditioning.
Before performing any study-related assessments, the investigator will inform the
patient both by word of mouth and in writing about all aspects of the study including
potential benefit and risks associated with the participation in this study. No study-
specific assessment will be performed unless the patient has given her/his written
informed consent. Baseline assessments can be performed during a 2–3 day period
as necessary for the patient’s comfort and convenience.
Note: Straining examinations needed for evaluation of inclusion/exclusion criteria
(e.g. CT, lumbar puncture) which have been performed on the patient recently do not
have to be repeated for baseline assessment. Instead previous results obtained up to
3 months prior to the baseline assessments may be used according to the
investigator’s judgment and if agreed by the patient.
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Written informed consent has to be obtained at Visit I. For children an age-related

informed consent/assent will have to be signed as appropriate, furthermore written
informed consent of the child’s legal representative has to be obtained. The date of
consent will be recorded in the eCRF.
When the patient has consented to participate in the study, the following parameters
will be assessed at the baseline visit (see section 5.2.3).
• Inclusion and exclusion criteria
Concomitant medication
• Documentation of previous therapy of underlying disease
• Documentation of relevant previous and ongoing medication
Outcome parameters: feasibility
• Completion of quality of life (EQ-5D or PedsQL and FACT-BMT) questionnaires
Pre-transplant evaluations should include
• Demographic characteristics
• Medical history
• Physical examination, including Karnofsky/Lansky performance status
• Vital signs
• Additional examinations as considered appropriate by the investigator
Ultrasound
CT chest
ECHO/ECG
Cardiac performance (LVEF) determined by MUGA or echocardiogram
Lumbar puncture if prior history of CNS disease
Note: Results obtained up to 3 months prior to the baseline assessments may
be used according to the investigator’s judgement.
• Evaluation of Sorror Comorbidity Index (in adult patients only) [72]
• Disease status, clinical staging (patients with non-haematological malignant
disease)
Laboratory standard of care
• Collection of blood for laboratory analysis (‘big panel’, see section 5.2.3.2, III.)
• Collection of blood sample for serology analyses
• Collection of blood sample for PCR analysis to detect various viral infections
(HIV, HCV, CMV, ADV, EBV) and a blood sample for PCR or serology to detect
aspergillus infections
• Collection of blood sample for AB0 Rh blood typing and HLA matching
• Bone marrow puncture for laboratory staging (patients with haematological
malignancies only)
Outcome parameters: laboratory
• Collection of reference sample of peripheral blood and of bone marrow aspirate
for chimerism studies according to institutional standards: a pre-transplantation
sample is collected to allow identification of genetic differences between patient
and donor for subsequent analyses. In case suitable pre-transplantation
samples (DNA) for chimerism analyses of the patient already exist, a repeated
collection of the reference sample may be omitted. It has to be noted that - in
case of previous transplantations – the pre-transplantation sample is only
allowed to be used as reference sample, if collected after the last
transplantation (patients with haematological malignancies only).
Final 7.0 63 of 111 21. May 2015
• Collection of blood sample for KIR-Genotyping

Visit II: conditioning (Days –12 to –1)
The conditioning will start at Day –12 and will be performed as described above
(see section 5.1.2). The following evaluations will be performed during conditioning
therapy (Day –12 to Day –1). They are considered standard of care.
Examinations
• Vital signs
Adverse events and concomitant medication
• Documentation of new concomitant medication
• Documentation of therapy-related toxicities (known and unknown)
• Documentation of serious adverse events
Note: Known therapy-related toxicities which meet the SAE definition but are
not life-threatening will be documented in the eCRF as SAE, but will not be
recorded on special SAE report forms on paper and will not be reported as
SAE irrespective of duration of hospitalization.
Visit III: transplantation (Day 0 to Day +2)

At visit IIIa, IIIb and—if necessary—IIIc transplantation will be performed and acute
toxicity of the cell infusion will be monitored. The following evaluations will occur on
the days of transplantation (Day 0 to Day +2). They are considered standard of care.
Prior to transplantation
Examinations
Post-transplantation
• Monitoring of adverse events/serious adverse events
Note: Infections which meet the SAE definition but are not life-threatening will
be documented in the eCRF as SAE, but will not be recorded on special SAE
report forms on paper and will not be reported as SAE irrespective of duration
of hospitalization.
• Documentation of new concomitant medication
Outcome parameters: safety
• Monitoring of acute infusional toxicity: vital signs (see section 5.2.3.1, IV.).
Vital signs before HHCT within 1 hour prior to infusion
Vital signs after HHCT: 15, 30, 120 and 240 minutes post-infusion
Visit IV (Day +7), Visit V (Day +14) and Visit VI (Day +21):
post-transplantation phase I (each visit ±2 days)
While patients remain hospitalized after transplantation in a setting according to
institutional guidelines, the following assessments are planned in weekly intervals for
visits IV to VI to evaluate engraftment and immune reconstitution of the patients:
Examinations
• Vital signs
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• Laboratory assessments, ‘small panel’ (see section 5.2.3.2, I.)
Note: Blood samples for analyses of laboratory parameters of the ‘small panel’
will be collected weekly during the post-transplantation phase I (until Day +28)
according to institutional standards
Laboratory assessments, ‘daily panel’: the daily panel of laboratory parameters
(see section 5.2.3.2, II.) will be analysed daily from Day +4 to Day +28 or the
day of engraftment, whatever comes first. If engraftment is prior to day 28,
laboratory assessments will be performed as per hospital routine until day 28.
In addition, the parameters of the ‘daily panel’ will be measured at each visit
from Day 35 to Day 100 (Visit VIII to Visit XIV).
• Collection of blood sample for PCR analysis to detect CMV, ADV and EBV
infections and a blood sample for PCR or serology to detect aspergillus
infections; samples will be collected and analysed once to twice weekly until
Day +28
• Documentation of immune cell phenotyping results as measured routinely..
• Documentation of adverse events/serious adverse events
of hospitalization.
• Documentation of concomitant medication
• Assessment of acute GVHD
• Monitoring of NRM
• Assessment of engraftment
• Documentation of transfusions (thrombocytes, erythrocytes, other)
Outcome parameters: laboratory (routine local assessments)
• Collection of sample of peripheral blood for analysis of chimerism, if clinically
relevant results are expected by the treating physician
Outcome parameters: laboratory (additional central assessments)
• Collection of blood sample for immune cell phenotyping (flow cytometry).
Visit VII (Day +28±2 days): post-transplantation phase I, discharge visit
All assessments described above for Visits IV to VI will also be performed at the
discharge visit VII. Additionally, the following parameters will be evaluated:
Examinations
• Disease status, clinical staging (patients with malignant disease)
• Bone marrow puncture for laboratory staging (patients with hematological
malignancies only)
• Graft failure (primary, secondary)
• Monitoring of duration of hospitalization
Final 7.0 65 of 111 21. May 2015
• Collection of sample of bone marrow for analyses of chimerism (patients with

hematological malignancies only)
• Collection of blood sample for analyses of reconstitution of immune system
Visit VIII (Day +35±2 days) to Visit XIII (Day +70±2 days):
post-transplantation phase II
At all visits of post-transplantation phase II (Visit VIII [Day +35] to Visit XII [Day +70])
the stability of the donor chimerism will be evaluated in weekly intervals. The
following assessments are planned:
Examinations
• Vital signs
• Laboratory assessments, ‘small panel’ and ‘daily panel’ (see section 5.2.3.2, I
and II.)
• Collection of blood sample for PCR analysis to detect CMV, ADV and EBV
infections and a blood sample for PCR or serology to detect aspergillus
infections;
• Documentation of immune cell phenotyping results as measured routinely.
of hospitalization.
• Collection of samples of peripheral blood for analyses of chimerism, if clinically
• Additionally at Visit XII (Day +63):
Collection of blood sample for immune cell phenotyping
Collection of blood sample for analysis of reconstitution of the immune
system.
Visit XIV (Day +100±5 days): post-transplantation phase II, primary study endpoint
The following assessments are planned for visit XIV at Day +100:
Examinations
• Vital signs
Final 7.0 66 of 111 21. May 2015

• Collection of blood sample for special laboratory analyses in paediatric patients
• Collection of blood sample for PCR analysis of CMV, ADV and EBV infections
and a blood sample for PCR or serology to detect aspergillus infections;
• Bone marrow puncture for laboratory staging (hematological malignancies)
of hospitalization.
• Assessment of acute GVHD: primary study objective
• Monitoring of overall and disease-free survival
• Monitoring of relapse rate
• Monitoring of occurrence and duration of rehospitalization
• Collection of samples of peripheral blood for analyses of chimerism, for patients
with haematolocial maignancies additionally samples of bone marrow are
collected if clinically relevant results are expected by the treating physician
• Collection of blood sample for immune cell phenotyping
• Collection of blood samples for analysis of reconstitution of the immune system.
Visit XV (Month +6±2 weeks) and Visit XVI (Month +9±2 weeks): follow-up phase I
The following assessments are planned for visits during the follow-up phase I:
Examinations
• Vital signs
Serious adverse events
• Documentation of serious adverse events in the eCRF
Serious adverse reactions
• Reporting of serious adverse reactions
• Documentation of concomitant medication in case of a SAR
Documentation of new treatment with cellular products as e.g. DLI or other white
blood cells and inclusion in other clinical study
Final 7.0 67 of 111 21. May 2015


• Collection of blood sample for immune cell phenotyping (at Visit XV [Month +6],
only).
Visit XVII (Month +12±2 weeks): follow-up phase I
The following assessments are planned for Visit XVII:
Examinations
• Vital signs
• Disease status, clinical staging (for malignant disease; at Visit XVII [Month +12])
• Assessment of chronic GVHD
Outcome parameters: laboratory (routine local)
• Collection of blood sample for immune cell phenotyping.
Visit XVIII (Month +24±4 weeks): follow-up phase II, data closure
An additional follow-up visit and data closure are planned for 24 months post-
transplantation. The following assessments will be performed:
Examinations
• Vital signs
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• Completion of quality of life (EQ-5D or PedsQL and FACT-BMT) questionnaires.
Final 7.0 69 of 111 21. May 2015

Table 6: Evaluations and assessments of patients: flow-chart

Visits I II III IV V VI VII
Condition- Transplan- Post Transplantation I
Baseline ing* tation* (before or until discharge)
Day Day Days Day (±2 days)
Assessments
–40 to –14 –12 to –1 0 to +2 +7 +14 +21 +28
Patient informed consent x
Inclusion / Exclusion Criteria x
Examinations (see section 5.2.3.1)
Demographic characteristics (I.) x
Medical History (II.) x
a
Physical Examination (III.) x x x x x x x
Vital Signs (IV.) x x x x x x x
b
Additional Examinations (V.) x
Sorror Comorbidity Index (VI.) x
c
Clinical Staging (VII.) x x
Laboratory standard of care (see section 5.2.3.2)
d e e e e
Laboratory Analysis (I., II., III.) x x x x x
Special laboratory analysis (IV.)
Serology (V.) x
PCR analyses (VI.) xf, g xg, h xg, h xg, h xg, h
AB0 Rh blood group/HLA typing x
(VII.)
i BM BM
Laboratory staging (VIII.) x x
Adverse Events (see section 5.2.4)
Reporting of SAEs x x x x x x x
j
Documentation of AEs x x x x x x
Reporting of SARs only
Concomitant Medication x x x x x x x
Outcome Parameters (see section 5.2.6)
Safety Parameters
l l l l
GVHD (I.) x x x x
NRM (II.) x x x x
Infusional toxicity (III.) x
Graft failure (IV.) x
Engraftment (V.) x x x x
Survival (VI.)
n
Transfusions (VII.) x x x x
Relapse rate (VIII.)
Days of hospitalization (IX.) x
Quality of life questionnaires (X.) x
Laboratory parameters
R: PB; BM,i PB PB PB PB; BM,i
Chimerism analysis (XI.) x x x x x
o
Immune cell phenotyping (XII.) x x x x
p q
Immune system reconstitution x x
(XIII.)
a b
including Karnofsky/Lansky index; Ultrasound, CT chest, ECHO/ECG, LVEF, DLCO, lumbar puncture if
appropriate; non-hematological malignancies; ‘big panel’, including pregnancy test (β-HCG) for female
c d
e f
recipients; ‘small panel’ weekly, daily panel from Day 4 until Day +28 daily, then weekly with ‘small panel’; HIV
g h
and HCV (baseline only); CMV, ADV, EBV and aspergillus, once to twice weekly up to Day 28, then weekly;
i
twice weekly from Day 4 to Day 28; hematological malignancies only, within 30 days prior to HHCT and as
j k
indicated after HHCT; therapy-related toxicity, only; only in case of occurrence of a SAR or treatment with cellular
l m n
product (DLI, Ag-specific T cells infusion) and inclusion in other clinical study; acute GVHD; chronic GVHD;
o
platelets, erythrocytes etc.; documentation of data, if available as per hospital routine, and additional analysis at
p
GMP laboratory: flow cytometric analysis of immune cell phenotypes (T, B, NK cells and Tregs); additional
analysis at central laboratory: donor/recipient KIR genotyping for baseline, other parameters (T Vbeta repertoire, T
q
gamma/delta repertoire, TREC analysis, KIR repertoire, T cell and NK cell activity) as indicated; reference sample
PB BM R
for KIR genotyping; peripheral blood; bone marrow; Reference specimen
Roman numerals in brackets: see corresponding paragraphs in sections 5.2.3.1, 5.2.3.2 and 5.2.6
Final 7.0 70 of 111 21. May 2015

Table 6: Evaluation and assessments of patients: flow-chart (continued)

VIII IX X XI XII XIII XIV XV XVI XVII XVIII
Follow-
Post Transplantation II Follow-up
up phase
(after discharge until primary endpoint) phase I
Visits II
Day Month (±4
Assessments (±2 days) (±5 days) (±2 weeks) weeks)
+35 +42 +49 +56 +63 +70 +100* +6 +9 +12 +24
Patient informed consent
Inclusion / Exclusion Criteria
Examinations(see section 5.2.3.1)
Demographic Characteristics (I.)
Medical History (II.)
a
Physical Examination (III.) x x x x x x x x x x x
Vital Signs (IV.) x x x x x x x x x x x
b
Additional Examinations (V.)
Sorror Comorbidity Index (VI.)
c
Clinical Staging (VII.) x x
Laboratory standard of care (see section 5.2.3.2)
e e e e e e e
Laboratory analysis (I., II., III.) x x x x x x x
Special laboratory analysis (IV.) x
Serology (V.)
g g g
PCR analyses (VI.) x xg x xg x xg xg
AB0 Rh blood group/HLA typing (VII.)
i
Laboratory Staging (VIII.) xBM
Adverse Events (see section 5.2.4)
Reporting of SAEs x x x x x x x
Documentation of AEs x x x x x x x x x x x
Reporting of SARs only x x x x
k k k k
Concomitant Medication x x x x x x x x x x x
Outcome Parameters (see section 5.2.6)
Safety Parameters
l l l l l l l m m
GVHD (I.) x x x x x x x x x
NRM (II.) x x x x x x x x x x x
Infusional toxicity (III.)
Graft failure (IV.)
Engraftment (V.)
Survival (VI.) x x x
n
Transfusions (VII.) x x x x x x x
Relapse rate (VIII.) x x x
Days of hospitalization (IX.) x
Quality of life questionnaires (X.) x x x
Laboratory parameters
Chimerism analysis (XI.) xPB xPB xPB xPB xPB xPB xPB;BM,i xPB xPB xPB
o
Immune cell phenotyping (XII.) x x x x
p
Immune system reonstitution (XIII.) x x
a b
including Karnofsky/Lansky index; Ultrasound, CT chest, ECHO/ECG, LVEF, DLCO, lumbar puncture if
appropriate; non-hematological malignancies; ‘big panel’, including pregnancy test (β-HCG) for female
c d
e f
recipients; ‘small panel’ weekly, daily panel from Day +4 until Day +28 daily, then weekly with ‘small panel’; HIV
g h
and HCV (baseline only); CMV, ADV, EBV and aspergillus once to twice weekly up to Day 28, then weekly;
i
twice weekly from Day 4 to Day 28; hematological malignancies only, within 30 days prior to HHCT and as
j k
indicated after HHCT; therapy-related toxicity, only; only in case of occurrence of a SAR or treatment with cellular
l m n
product (DLI, Ag-specific T cells infusion) and inclusion in other clinical study; acute GVHD; chronic GVHD;
platelets, erythrocytes etc.;
o
documentation of data, if available as per hospital routine, and additional analysis at GMP laboratory: flow
p
cytometric analysis of immune cell phenotypes (T, B, NK cells and Tregs); additional analysis at central
laboratory: donor/recipient KIR genotyping for baseline, other parameters (T Vbeta repertoire, T gamma/delta
PB BM
repertoire, TREC analysis, KIR repertoire, T cell and NK cell activity) as indicated; peripheral blood; bone
marrow; * primary study endpoint
Roman numerals in brackets: see corresponding paragraphs in sections 5.2.3.1, 5.2.3.2 and 5.2.6
Final 7.0 71 of 111 21. May 2015

5.4. Unscheduled visits

It is at the discretion of the investigator to appoint additional visits as medically
necessary. Measures and assessments performed and the date and reason for such
a visit have to be documented in detail in the eCRF.
5.5. Early Termination Visit (ETV)

If a patient prematurely discontinues the study because of any reason before
Day 100 all assessments planned for the regular Visit XIV (Day 100) should be
performed at an early termination visit (‘Early Termination Visit [ETV] A’). The
following examintations have to be performed at the ETV A:
Examinations
• Vital signs
• Collection of blood sample for special laboratory analyses in paediatric patients
• Collection of blood sample for PCR analysis of CMV, ADV and EBV infections
and a blood sample for PCR or serology to detect aspergillus infections;
• Bone marrow puncture for laboratory staging (hematological malignancies only)
of hospitalization.
• Monitoring of occurrence and duration of rehospitalization
• Collection of blood sample for immune cell phenotyping
• Collection of blood samples for analysis of reconstitution of the immune system.
If a patient prematurely discontinues the study because of any reason after Day 100
all assessments planned for the regular Visit XVII (Month 12) should be performed at
Final 7.0 72 of 111 21. May 2015

an ‘Early Termination Visit [ETV] B’. The reason for premature discontinuation has to
be documented in the eCRF. The following examinations have to be performed at the
ETV B:
Examinations
• Vital signs
• Disease status, clinical staging (for malignant disease; at Visit XVII [Month +12])
• Documentation of immune cell phenotyping results as measured according to
institutional routine.
• Documentation of serious adverse events (concomitant medication will be
documented in case of an SAE on the SAE form)
of hospitalization.
• Documentation of new treatment with cellular products as e.g. DLI or other
white blood cells and inclusion in other clinical study.
Outcome parameters: laboratory (additional central asessments)
• Collection of blood sample for immune cell phenotyping.
6. ADVERSE EVENTS
6.1. Definitions
6.1.1. Adverse event (AE)
The term adverse event describes any untoward medical occurrence in a patient or
clinical investigation subject administered an investigational medicinal product (IMP).
It does not necessarily have a causal relationship with this study 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 an
investigational medicinal product (IMP).
This definition includes:
• Any sign or symptom occurring during the study
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• Any event or disease present on the day of inclusion in the study with
symptoms that worsens during the study
• Any accident
• Any significant change in laboratory parameters
• Any reaction to drug withdrawal
• Any drug interaction
• Any effect related to overdose, abuse or dependence.
6.1.2. Adverse Reaction (AR)

Adverse reactions (ARs) include all untoward and unintended responses to an IMP
related to any dose administered. All AEs judged by either the reporting investigator
or the Sponsor as having a reasonable possibility of a causal relationship between
the event and the IMP qualify as ARs. This means that there are facts (evidence) or
arguments to suggest a causal relationship between the event and the IMP. An AR is
defined as unexpected when its nature, severity or outcome is not consistent with the
information that has been obtained from previous observations and investigational
trials.
6.1.3. Serious Adverse Events (SAEs)

A SAE is any untoward medical occurrence that at any dose:
• results in death,
• is life-threatening (an event in which the patient was at risk of death at the time
of the event; however, it does not refer to an event, which, hypothetically, might
have caused death if it had been more serious.),
• requires inpatient hospitalization or prolongation of existing hospitalization,
• results in persistent or significant disability/incapacity,
• is a congenital abnormality/birth defect
• is another important medical event that may not be immediately life threatening
or result in death or hospitalization but, based upon appropriate medical
judgment is thought to jeopardize the patient or subject or require medical or
surgical intervention to prevent one of the outcomes defining a SAE.
Furthermore the following are considered to be serious adverse events even if they
do not comply with the definition:
• Any pregnancy discovered during the study.
Note: Pregnancy is an exclusion criterion. Hence contraceptive measures are
obligatory in woman of childbearing potential throughout the study. If pregnancy
should occur during the study the pregnancy has to be followed to term. The
outcome for mother and child has to be documented.
Study-specific definition
In the context of this study any occurrence of GVHD grade III–IV until Day 100 has
to be reported according to the standard procedure of SAE reporting, irrespective
whether or not fulfilling the SAE criteria defined above (for further details see section
6.4)
In the context of this study (seriously ill patients undergoing conditioning treatment
leading to immune dysfunction) the following events meeting the above definition will
be documented in the eCRF as SAE, but will not be recorded on special SAE report
forms on paper and will not be reported as SAE:
Final 7.0 74 of 111 21. May 2015

• An infection requiring inpatient hospitalization or prolongation of hospitalization

which is not life-threatening
• A known therapy-related toxicity requiring inpatient hospitalization or
prolongation of hospitalization which is not life-threatening.
Note: Planned hospitalization for a preexisting condition or hospitalization due to a
procedure required as per study protocol is not considered to be a serious adverse
event.
6.1.4. Serious Adverse Reactions (SARs) and Suspected Unexpected Serious

Adverse Reactions (SUSARs)
An AR that meets seriousness criteria, as defined above, is defined as a serious
adverse reaction (SAR). A suspected unexpected (unlisted) serious adverse reaction
(SUSAR) is a SAR, the nature or severity of which is not consistent with the
applicable product information (Investigator’s brochure [IB]).
6.2. Assessment of Adverse Events/Therapy-Related Toxicities

6.2.1. Severity
The intensity of the severity of adverse events must be assessed by using the
following categories: mild, moderate and severe. This assessment is subjective and
medical judgment should be used to compare the reported adverse events to similar
types of events observed in clinical practice. It is important to recognize that severity
is not equivalent to event seriousness. Guidelines for severity assessments are listed
below:
• Mild: Awareness of a sign or symptom barely noticeable to the patient
or does not make the patient uncomfortable; the AE does not
cause a limitation of the usual activities.
• Moderate: Symptom with enough discomfort to cause interference with
normal activities. Treatment of symptom may be needed.
• Severe: Symptom of a sufficient severity to cause the patient severe
discomfort and prevent performance of normal activities;
resistant to conventional symptomatic treatment.
6.2.2. Causality
The causality of AEs refers to the relationship of the AE to study treatment. When
completing the eCRF, the investigator will be asked to assess the causality of the
event. Investigators will be asked to assess causality as ‘IMP-related’, ‘concomitant-
medication-related’ or ‘other’. Causality will then be categorized by using a simple
binary decision for causality according to the following criteria for all three parameters
as applicable:
• Not related:
An adverse event for which there is no reasonable possibility of a causal
relationship with the IMP.
• Related:
An adverse event for which there is a reasonable possibility of a causal
relationship with the IMP. This means that there are facts (evidence) or
arguments to suggest a causal relationship.
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The assessment of whether there is a reasonable possibility of a causal relationship

is usually made by the investigator. The causality assessment given by the
investigator should not be downgraded by the sponsor. If the sponsor disagrees with
the investigator’s causality assessment, the opinion of both, the investigator and the
sponsor, should be provided with the report. The following points should be taken
into account during causality assessment of adverse events:
• Timing of the event between administration of the drug and the onset of the
adverse event
• Drug levels and evidence, if any, of overdose
• A known or expected response pattern to the suspect drug including previous
experience with the drug and whether the adverse event is known to have
occurred with the drug, physiological effects of the drug, known adverse event
related to drugs belonging to the same or a similar class, can be explained by
the pharmacological action or with regard to findings in animals or a specific
genetic predisposition of the patient.
6.3. Monitoring, Recording and Reporting of Adverse Events

6.3.1. General Requirements
During the course of the study, SAEs will be recorded and reported on separate
paper SAE forms from baseline until the primary endpoint is reached (Visit XIV)
irrespective of the relation to the study drug. During the follow-up phase (after Visit
XIV until Visit XVIII) SAEs will be recorded and reported on separate paper SAE
forms only, if assessed as related to the study drug.. AEs irrespective of relation to
study drug will be recorded from Day 0 to Day 100. The investigator will be
responsible for ensuring that correct information concerning AEs as specified above
is included on the appropriate eCRF forms.
The following data will be recorded:
• Description of the AE: nature, frequency, intensity, time and date of onset and
resolution, outcome, and causal relationship to the study drug according to the
investigator)
• Whether the AE has to be considered as an SAE
• Action taken.
All AEs occurring during the study must be followed-up until the AE has completely
resolved, stabilized or the sequelae can be assessed by the investigator. In case of
withdrawal the respective eCRF has to be completed. The results of additional
diagnostic measures as the result of an AE, such as laboratory tests, ECG,
angiogram, echocardiogram and MRI must be available at site and eventually copied
on request.
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6.3.2. Reporting of Serious Adverse Events
The investigator has to report all SAEs within 24 h to

will enter all data in a
pharmacovigilance database and will immediately (within 1 working day) notify the
DSMB in case of graft failure, NRM or acute GVHD grade III–IV.
All SAEs that occur during the period of observation from baseline until primary
endpoint (Visit XIV), and all SAEs occurring up to 30 days after receiving the stem
cell injection (study drug) in case of withdrawal, whether considered to be associated
with the study drug or not, have to be reported by the investigator.
During follow-up phases I and II (until Visit XVIII) only SARs have to be reported by
the investigator.
An SAE/SAR report has to be faxed or emailed within 24 hours after knowledge of

the event by the investigator to
under the following contact:
SAE/SAR Fax:
Or
SAE/SAR email:
The investigator should not wait for additional information to fully document the event
before notifying the appointed safety officer of an SAE, though additional information
may be requested. The minimum information required for an initial report is:
• Sender of report (name, address and phone number of investigator)
• Date of initial report
• Patient identification (patient number)
• Protocol number
• Investigational medicinal product
• Description of event with outcome, if available, and criteria for seriousness,
expectedness and causality.
Where applicable, information from relevant laboratory results, hospital case records
and autopsy reports should be obtained. The investigator is also required to submit
follow-up reports within 24 hours to until the
SAE has resolved or—in the case of permanent impairment—until the SAE
stabilizes. In addition, the event will be documented in the eCRF.
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The results of additional diagnostic measures as the result of an SAE, such as

laboratory tests, ECG, angiogram, echocardiogram and MRI must be available at site
and eventually copied on request.
will report all serious and unexpected adverse
drug reactions (SUSARs) and SAEs as required by local legal regulations within the
required time to the responsible Competent Authorities, the Independent Ethics
Committees (IECs) in Germany and the Netherlands, the Sponsor, the DSMB and to
all investigators concerned. Regulatory agencies will be notified as soon as possible
but no later than 7 days after first knowledge of fatal or life-threatening unexpected
AR and no later than 15 days after knowledge of any other unexpected SAR.
Note: Known therapy-related toxicities (TRT) and infections which meet the SAE
definition, but are not life-threatening will be documented in the eCRF as SAE, but
will not be recorded on special SAE report forms on paper and will not be reported as
SAE irrespective of duration of hospitalization.
6.3.3. Other Procedures for Recording and Reporting Adverse Events

All Adverse Events will be analysed at the end of the study period.
During this study, there will be standard pharmacovigilance monitoring for SARs and
SUSARs to allow for annual safety reporting procedures.
will be responsible for safety updates to regulatory authorities in
accordance with guidelines and guidance and the German Medicines Act (AMG) or
the Wet medisch-wetenschappelijk onderzoek met mensen (WMO) and the Wet op
bijzondere medische verrichtingen (WBMV) and for updating IECs (Independent
Ethics Committees) in accordance with the guidelines.
6.4. Adverse Events of Specific Interest

GVHD grade III and grade IV, NRM and graft failure are defined as adverse events of
specific interest for the purpose of this study and, will be monitored in detail. Adverse
events of specific interest will be continuously documented and have to be reported
according to the standard procedure of SAE reporting described above until Visit XIV,
Day +100.
Adverse Events of Specific Interest will be reported to the DSMB. Details of the
DSMB’s responsibilities and working procedures are provided below, see section 9.4.
6.5. Follow-Up of Adverse Events

All AEs experienced by a patient, irrespective of the suspected causality, will be
monitored:
• until the event has resolved or in case of a clinically significant abnormal
laboratory value has returned to baseline or stabilized at an acceptable level
according to the investigator and the clinical monitor,
• until there is a satisfactory explanation for the changes observed, or
• until the patient is lost to follow-up.
The institutions/physicians following the patients during and after convalescence will
be informed about the patient’s study participation and advised to increase their level
of attention. The study center investigator will conduct safety-related follow-up
Final 7.0 78 of 111 21. May 2015

examinations on an outpatient basis for a minimum of 6 months or until resolution of

SAEs occurring documented at Visit XVIII (end of follow-up phase II).
6.6. Therapy Toxicities of Conditioning
Therapy-related toxicities are known during the conditioning period and will have to
be rated as known and unknown therapy-related toxicity. Unknown therapy-related
toxicities will have to be documented as adverse event as described in section 6.1.1.
From Day 0 onwards no further distinction between known and unknown therapy-
related toxicities will be made. Known therapy-related toxicities are to be found in the
respective SmPCs. Adverse events will be documented from Day 0 to Day 100.
6.7. Unblinding of treatment / Emergency identification

Not applicable.
7. STATISTICS
The data will be statistically analysed by the statistical department of
as identified in the Section ‘Contact Addresses and
Responsibilities’. Before start of the statistical analysis a separate statistical analysis
plan (SAP) will be finalized, providing detailed methods for the analyses outlined
below. Analyses will be performed after database lock.
Any deviations from the planned analyses will be described and justified in the final
integrated study report. Statistical programming and analyses will be performed using
the validated computer software package SAS® or other validated statistical software
as required.
7.1. General Considerations

The statistical analyses in this study will be exploratory since the study is not
powered to address any pre-defined statements but to generate valid hypotheses on
safety/tolerability and feasibility issues. A formal sample size calculation was
therefore not done. Thus, all resulting p-values and confidence intervals are to be
interpreted in the exploratory sense only.
Data will be appropriately summarized and analysed using tabulation and graphs for
demographic and baseline characteristics, safety/tolerability and feasibility
observations and measurements. Standard descriptive summary statistics (i.e. n,
arithmetic mean, standard deviation, median, lower/upper quartiles, and
minimum/maximum values) will be calculated for continuous variables. Categorical
data will be presented in frequency tables using counts and percentages. Data for
the adult and paediatric patients will be presented separately in summary tables.
Subgroup analyses will be performed by the type of indication, as appropriate. For all
data collected during the study, listings of the individual raw data will be provided.
Individual patient data listings will be presented per parameter and will be sorted by
study group, center, patient number and study visit, if applicable.
The main analysis will be performed after completion of the 100 days post-
transplantation visit (end of post-transplantation phase II), i.e. when all patients have
either completed the 100 days period after transplantation, are lost to follow-up or
have died within this period. Additional analyses will be done on the 1-year post-
transplantation (end of follow-up phase I) and on the 2-year follow-up data (end of
Final 7.0 79 of 111 21. May 2015
follow-up phase II), i.e. when all patients have completed the 1-year or 2-year period
after transplantation, are lost to follow-up or have died within these periods.
7.2. Determination of Sample Size

No formal estimation of the sample size was performed. It is planned to enroll a total
of n = 60 patients, overall, in 13 centers, with 30 adult patients in 7 centers and 30
paediatric patients in 6 centers.
7.3. Analysis Sets

7.3.1. Definitions
The statistical evaluation will be based on separate, hierarchically organized,
analysis sets as defined below:
• Safety analysis set: The safety analysis set will consist of all patients enrolled.
• Full analysis set: The full analysis set will consist of all patients who underwent
transplantation with the IMP.
• Per-protocol analysis set: The per-protocol analysis set will consist of all patients
of the full analysis set for whom no major protocol violations have been recorded.
Demographic and baseline characteristics will be analysed for all three analysis sets.
General safety data (AEs including infections) will be evaluated using the safety
analysis set. Outcome parameters (safety [GVHD, NRM and infusional toxicity],
feasibility and laboratory) will be evaluated using the full analysis set and the per-
protocol analysis set.
7.3.2. Eligibility, Protocol Deviations

Each patient’s allocation to the different analysis sets will be identified and mutually
agreed on between the Sponsor and the CRO prior to database lock. Deviations from
the protocol including violations of inclusion/exclusion criteria will be assessed as
‘minor’ or ‘major’ based on the combined decisions of the CRO and the Sponsor
according to the specifications outlined in the statistical analysis plan or a separate
document. Enrollment in another clinical study after Day 100 possibly interfering with
the endpoints of the present study will be considered a major protocol violation.
Major deviations from the protocol will lead to the exclusion of a patient from the per-
protocol analysis set. Listings will be prepared to show the eligibility of all patients for
the different analysis sets and a summary will be given on the number of patients per
analysis set. A detailed description of the criteria used for data sort out will be
provided before start of the evaluations.
7.4. Graft Evaluation

Total TCRα/β and CD20 cell counts and the recovery of viable CD34+CD45+ cells
after depletion as well as cell counts of lymphocyte subpopulations of the individual
grafts after processing and prior to transplantation will be analysed and presented
using standard descriptive summary statistics (i.e., n [number of grafts], arithmetic
mean, standard deviation, median, lower/upper quartiles, and minimum/maximum
values). Graphical presentation will be given by means of box and whisker plots and
bar charts, as appropriate. Furthermore the log depletion of TCRα/β cells, of CD19
cells, the number of grafts containing ≥4 × 106 CD34+CD45+ stem cells/kg BW, the
number of grafts containing ≤25 × 103 TCRα/β cells/kg BW and the number of grafts
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containing ≤1 × 105 CD20+ cells/kg BW and their relative proportions will be

calculated. This analysis will be performed for all grafts processed for transplantation.
7.5. Demographics and Baseline Characteristics

Demographics and baseline characteristics for patients and donors will be
summarized by means of descriptive statistics for continuous data or frequency
tables for categorical data. The use of concomitant medications will be tabulated by
generic drug name and Anatomical Therapeutic Chemical (ATC) code according to
the WHO-DD terminology. Findings concerning medical history and concomitant
diseases will be presented in MedDRA terminology categorized by primary System
Organ Class (SOC) and Preferred Term.
Demographic data and other baseline characteristics will be analysed for all three
analysis sets.
7.6. Treatment and Study Compliance

Treatment will be administered by study personnel hence patient compliance with
study treatment will not be monitored and analysed.
Patients who fail to attend study visits or refuse to undergo certain assessments are
non-compliant with the study protocol. Listings and frequency tables of those patients
will be provided.
7.7. Analyses of the Primary Endpoint

The primary endpoint will be the incidence and severity of acute GVHD grade II–IV
within 100 days post-transplantation. The acute GVHD will be assessed and graded
according to the Seattle (Glucksberg) criteria (Appendix 1: Glucksberg Criteria for
grading of acute GVHD).
The exploratory analysis of the primary endpoint will be performed for the FAS and
the PP analysis sets.
Frequency tabulations of the number and percentage of patients with acute GVHD by
severity (i.e. the ‘crude incidence rates’) will be presented and displayed graphically
together with the two-sided 95% confidence interval (Clopper-Pearson).
In addition, time to acute GVHD will be evaluated to assess the incidence and
severity of acute GVHD from day of transplant. The first day of acute GVHD onset at
a certain grade will be used to calculate a cumulative incidence curve for that acute
GVHD grade. An overall cumulative incidence curve will be computed along with a
95% confidence interval at 100 days post-transplant with death considered as a
competing risk.
7.8. Analyses of the Secondary Endpoints

Analyses of the secondary endpoints, which are described in section 3.1, will be
performed for the FAS and the PP analysis sets. All inferential analyses for the
secondary safety, feasibility and laboratory outcome variables will be interpreted in
the exploratory sense.
Standard descriptive summary statistics (i.e., n, arithmetic mean, standard deviation,
median, lower/upper quartiles, and minimum/maximum values) will be calculated for
continuous variables. Categorical data will be presented in frequency tables using
counts and percentages. Graphical presentation will be given by means of box and
whisker plots and bar charts, as appropriate.
Final 7.0 81 of 111 21. May 2015
Survival distributions will be estimated using the Kaplan-Meier method. Binomial

proportions will be estimated using the observed proportion and Clopper-Pearson
interval estimator. Incidence rates will also be estimated using the cumulative
incidence function.
A detailed description of the planned analyses for the secondary endpoints will be
included in the statistical analysis plan.
7.9. Safety Analyses

The safety analysis set will be used to evaluate issues of general safety, i.e., adverse
events (including infections), serology, physical examinations, vital signs and
concomitant medication.
7.9.1. Adverse Events

Adverse events will be processed in the statistical analysis after classification into
standardized medical terminology from the verbatim description (Investigator term)
using the Medical Dictionary for Regulatory Activities (MedDRA). Adverse event data
will be summarized and presented using primary MedDRA System Organ Classes
(SOCs) and Preferred Terms (PTs), overall and classified by maximum severity and
maximum relationship to the IMP.
Special attention will be given to those patients who died, who have discontinued the
study due to an adverse event or who experienced serious adverse events. In
addition, the recurrence or new occurrence of infectious diseases, mainly caused by
viral/bacterial/fungal agents will be summarized. GVHD, NRM and acute infusional
toxicity after infusion of the IMP are secondary outcome parameters and will be
assessed as described in section 7.8.
All adverse events data will be listed in the individual patient data listings, including
all information documented on the adverse event form. Separate listings will be
provided likewise for serious adverse events, adverse events in subjects who died,
and for adverse events leading to discontinuation of the study. Concomitant
medication documented in relation to the occurrence of a SAR after Day 100 on SAE
forms will be listed separately as ‘Concomitant medication as documented on SAE
forms after Day 100’.
7.9.2. Laboratory
Laboratory values for standard of care parameters (e.g. clinical chemistry, serology
and CBCs) will be summarized by the type of laboratory test. For each quantitative
laboratory parameter descriptive summary statistics (for absolute values and
changes from baseline) will be displayed by study visits. Laboratory values of the
daily panel from Day 0 to Day 28 and results of PCR analysis of viral reactivation will
be displayed by day of assessment.
Laboratory baseline parameters as e.g. results of donor/recipient matching, AB0
blood typing and HLA phenotyping, analyses of allo-antibodies etc. will be presented
in individual patient data listings.
The results of all laboratory tests evaluated continuously will be presented in
individual patient data listings including the center specific reference ranges, the
investigator’s assessment for values outside the reference range and the
investigator’s comment. Abnormal values will be identified by flagging all values
below and above the reference range. Separate listings will be provided for clinically
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significant laboratory values as assessed by the investigator. Furthermore, separate

listings will be provided for values outside the reference ranges documented for the
follow-up phases (after Day 100, Visit XV to Visit VXIII).
7.9.3. Physical Examination

Results from physical examinations will be summarized for each scheduled study
visit by means of a frequency table.
7.9.4. Vital Signs

Vital signs will be presented in tabulated statistical summaries of the raw data for
each scheduled time. In addition absolute changes from the baseline values
(baseline/pre-infusion) will be provided.
7.10. Safety Monitoring of Key Safety Endpoints

Patient safety will be assessed continuously throughout the study by monitoring
incidence and severity of acute GVHD and NRM. Each case of GVHD grade III–IV
and each case of NRM have to be reported immediately and will be announced to the
DSMB (see section 9.4 and section 6.4).
Graft failure will also be analysed and assessed by the DSMB for decision on the
conditioning regimen after treatment of the first 10 patients who have not received
ATG Fresenius. If graft failure occurred in less than 3 patients treated without the use
of ATG Fresenius, all patients enrolled after this analysis—both, patients in CR and
patients with therapy-refractory disease—can be transplanted without the use of ATG
Fresenius in the conditioning regimen.
7.10.1. Statistical Stopping Guidelines

Acute GVHD grade III–IV and NRM will be monitored and incidence rates will be
supervised by the DSMB continuously throughout the study. Additionally, statistical
monitoring of key safety endpoints will be performed separately for the cohorts of
paediatric and adult patients. Key safety endpoints will be assessed in 5 interim looks
after a total of 6 patients within each cohort have completed the 100 days post-
transplantation visit (end of post-transplantation phase II) successively, i.e. when all 6
patients at each stage have either completed the 100 days period after
transplantation, are lost to follow-up or have died within this period. These interim
looks will be forwarded to the Sponsor and the DSMB. If rates significantly exceed
pre-set statistical thresholds at the interim looks, further recruitment will be stopped
and the Sponsor will decide about further study continuation after consultation with
the DSMB (see section 9.4). We expect that the probability of experiencing GVHD
grade III–IV will be about 0.10 and of experiencing NRM will be about 0.10
(paediatric patients) or 0.20 (adult patients).
The statistical stopping guidelines presented here are to serve as a trigger for
initiating consultation with the DSMB for additional review. They are not intended as
formal ‘stopping rules’ that would mandate automatic closure of study enrollment.
• Acute GVHD grades III–IV will be monitored based on a binomial test of
proportion, testing the null hypothesis, i.e. the acute GVHD rate at Day 100 post-
transplantation is less than or equal to 10%. A Kim and DeMets class α spending
function with ρ = 0.40 will be used for the group sequential test with the null
hypothesis H0 : p = p0 = 0.10 vs. the alternative H1 : p = p1 = 0.30.
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• Likewise the rate of NRM will be monitored based on a binomial test of proportion,
testing the null hypothesis, i.e. the NRM rate of Day 100 post-transplantation is
less than or equal to 10% (paediatric patients) or 20% (adult patients) vs. the
alternative proportion of 30% (paediatric patients) or 50% (adult patients).
A group sequential design with a maximum of K = 5 stages will be chosen. The
critical values and the test characteristics of the group sequential test design were
calculated for a Kim and DeMets class α spending function with ρ = 0.40. The
calculations were performed with the statistical software program ADDPLAN 6.0
(Adaptive Designs – Plans and Analyses®). The following tables (Table 7 and Table
8) provide statistical information on the stopping boundaries for the two key safety
endpoints based on the standardized test statistic together with values of α spent and
power achieved for each of the equally sized five stages with n = 6 patients per stage
within each cohort. If at analysis stage k the cumulative number of patients with
events is equal or greater than the critical value, the null hypothesis can be rejected
and consultation with the DSMB for additional review will be initiated.
Table 7: Interim safety monitoring stopping boundaries for aGVHD grade III–IV at
n patients
Bounds to Sign. level Power
Stage α spent Critical
reject H0 one-sided achieved per Stage Total
value
Paediatric patients
1 1.939 0.0263 0.0263 0.2691 6 6 3
2 2.184 0.0145 0.0347 0.4425 6 12 5
3 2.228 0.0130 0.0408 0.5996 6 18 6
4 2.238 0.0126 0.0457 0.7259 6 24 7
5 2.238 0.0126 0.0500 0.8195 6 30 8
Adult patients
1 1.939 0.0263 0.0263 0.2691 6 6 3
2 2.184 0.0145 0.0347 0.4425 6 12 5
3 2.228 0.0130 0.0408 0.5996 6 18 6
4 2.238 0.0126 0.0457 0.7259 6 24 7
5 2.238 0.0126 0.0500 0.8195 6 30 8
H0 : p = p0 = 0.10 vs. H1 : p = p1 = 0.30; Kim and DeMets class α spending function with ρ = 0.40
Table 8: Interim safety monitoring stopping boundaries for NRM at Day 100 post-
transplantation
n patients
Bounds to Sign. level Power
Stage α spent Critical
reject H0 one-sided achieved per Stage Total
value
Paediatric patients
1 1.939 0.0263 0.0263 0.2691 6 6 3
2 2.184 0.0145 0.0347 0.4425 6 12 5
3 2.228 0.0130 0.0408 0.5996 6 18 6
4 2.238 0.0126 0.0457 0.7259 6 24 7
5 2.238 0.0126 0.0500 0.8195 6 30 8
Adult patients
Final 7.0 84 of 111 21. May 2015

1 1.939 0.0263 0.0263 0.3737 6 6 4

2 2.184 0.0145 0.0347 0.5981 6 12 7
3 2.228 0.0130 0.0408 0.7684 6 18 9
4 2.238 0.0126 0.0457 0.8767 6 24 11
5 2.238 0.0126 0.0500 0.9383 6 30 12
H0 : p = p0 = 0.10/0.20 vs. H1 : p = p1 = 0.30/0.50; Kim and DeMets class α spending function with
ρ = 0.40.
7.11. Interim Analysis

No formal interim analyses are planned for this study.
7.12. Data Handling

Data processing will be performed by
appointed by Miltenyi GmbH, using the CFR 21 part 11 compliant EDC-System
MARVIN.
Data will be entered into a database specifically created for this study either remote
by the trained site staff or via electronic data load for data not documented in the
eCRF, e.g., central laboratory data. Changes made to the data will be documented in
an audit trail. The study database and the eCRF will be created according to the
format and content of the study protocol and the study-specific details laid down in
the data management plan.
The following coding dictionaries will be used in their current installed version at the
beginning of coding activities, ensuring that there is only one version used during the
study:
• Diseases: Medical Dictionary for Regulatory Activities (MedDRA)
• Adverse events: Medical Dictionary for Regulatory Activities (MedDRA)
• Drugs: WHO-DRL dictionary
All available data will be included in the analyses and will be summarized as far as
possible. Unless otherwise specified, no substitution of missing data will take place,
i.e., missing data will not be replaced but will be handled as 'missing' in the statistical
evaluation.
In the case of a premature termination, the last available observation will be used for
analysis (LOCF approach), if applicable and considered reasonable. Missing
baseline data will not be replaced. A more detailed description of the handling of
missing data will be provided in the statistical analysis plan after review of the data.
8. ETHICAL ASPECTS
The procedures set out in this study protocol are designed to ensure that the
Sponsor and investigators abide by the principles of the Good Clinical Practice
guidelines of the ICH, and of the Declaration of Helsinki. The study also will be
carried out in keeping with local legal requirements.
8.1. Independent Ethics Committee Approval

Before implementing this study, the protocol, the proposed informed consent form
and other information to subjects, must be reviewed by a properly constituted
Final 7.0 85 of 111 21. May 2015

Independent Ethics Committee / Institutional Review Board (IEC/IRB). A signed and

dated statement that the protocol and informed consent have been approved by the
IEC/IRB must be available at Miltenyi Biotec GmbH before study initiation. The name
and occupation of the chairman and the members of the IEC/IRB must be present at
Miltenyi Biotec GmbH.
8.2. Informed Consent

Patients who are eligible for enrolment into the study will be informed by the
investigator in detail about the study. Donors will be informed about additional
requirements (data transfer and collection and genetic analysis of one blood sample)
and that donor’s consent to these requirements is needed to allow the patient to be
treated within this study. Information for donors does not involve routine procedures
of stem cell mobilization and apheresis because donors will already have agreed to
these procedures prior to being invited to donate stem cells for transplantation in the
context of the present study. For information about these requirements, for giving
consent and for collection of the blood sample the donor and where applicable their
legally authorized representative has to visit a study center once. Patients will be
allowed adequate time for consideration and making an informed decision, at least
24 hours. During this period patients will have the opportunity to discuss questions
and concerns with their treating physician. If patients are willing to participate in the
study, informed consent will be obtained from them or their legally authorized
representative according to the regulatory and legal requirements of Germany and of
The Netherlands. The consent form will be dated and retained by the investigator as
part of the study records. The investigator will not undertake any investigation
specifically required for this clinical study until valid consent has been obtained. The
date when consent was obtained will be documented in the eCRF.
According to the German Medicines Act, under-age patients personally have to give
informed consent in addition to their legal representatives, provided they are able to
understand the information given to them in this study.
The explicit wish of a minor or a mentally incapacitated adult, who is capable of
forming an opinion and assessing the study information, to refuse participation in or
to be withdrawn from the study at any time has to be considered by the investigator.
Patients/donors will be asked expressly to give their consent for use of their samples
in genetic analysis (additional assessments of transplantation success). If a patient
denies his/her consent to this analysis he/she cannot be enrolled in the study,
because the additional assessments are obligatory for determination of study
endpoints.
Patients can withdraw their consent at any time during the study period without
having to give a reason and without prejudice regarding their future medical
treatment.
If a protocol amendment is required, the informed consent form may need to be
revised to reflect the changes to the protocol. If the consent form is revised, it has to
be reviewed and approved by the appropriate IEC/Competent Authority (i.e. PEI and
Ministry of Health, Welfare and Sports, respectively), and signed by all patients
subsequently enrolled in the study as well as those currently enrolled in the study.
Final 7.0 86 of 111 21. May 2015

8.3. Data confidentiality

All study findings and documents will be regarded as confidential. The investigator
and members of his research team are not allowed to disclose such information
without prior written approval from the Sponsor.
The anonymity of participating patients has to be maintained. Patients will be
identified on the eCRF submitted to the Data Base by their patient number, not by
name. Documents not to be submitted to the Data Base that identify the patient (e.g.,
the signed informed consent) must be maintained in confidence by the Investigator.
8.4. Liability and Insurance

The insurance for study-related claims will be covered by Chubb Insurance Company
of Europe SE, Germany. The Sponsor will take out reasonable third-party liability
insurance cover in accordance with all local legal requirements. The civil liability of
the investigator, the persons instructed by him and the hospital, practice or institute in
which they are employed and the liability of the Sponsor with respect to financial loss
due to personal injury and other damage that may arise as a result of the execution
of this study are governed by the applicable law.
The Sponsor will arrange for patients participating in this study to be insured against
financial loss due to personal injury caused by the IMP tested or by medical steps
taken in the course of the study. If concomitant enrolment of a patient in another
clinical study after Day 100 is planned according to the investigator’s medical advice,
the insurance companies of the present and of the new study will have to be
informed accordingly before enrolment.
9. ADMINISTRATIVE PROCEDURES
By signing the study protocol, the investigator accepts to comply with all of the
following points:
9.1. Regulatory aspects

The study will be conducted according to the requirements of the declaration of
Helsinki (current version and in compliance with the current provisions of the German
Drug Law and the Wet medisch-wetenschappelijk onderzoek met mensen (WMO)
and the Wet op bijzondere medische verrichtingen (WBMV), the respective decrees
and the European Clinical Trial Directive. Regulatory reporting requirements will be
agreed on by the parties in the investigator contract.
9.2. Protocol Approval and Amendment

Before the start of the study, the study protocol and other relevant documents will be
approved by the IECs and Competent Authorities (PEI and Ministry of Health,
Welfare and Sports), in accordance with German and The Netherlands legal
requirements. The Sponsor must ensure that all ethical and legal requirements have
been met before the first patient is enrolled in the study.
This protocol is to be followed exactly. For any alteration of the protocol,
amendments must be written, receive approval from the appropriate personnel, and
receive IEC and Competent Authority (PEI and Ministry of Health, Welfare and
Sports) approval prior to implementation in Germany or The Netherlands, if
Final 7.0 87 of 111 21. May 2015

appropriate. Administrative changes not affecting the patient benefit/risk ratio may be
established without the need for a formal amendment.
All amendments will be distributed to all protocol recipients, with appropriate
instructions.
9.3. Duration of the Study

The maximum duration of the study for each patient will be approximately 2 years.
The study will close when all patients have completed the additional safety follow-up
visit at Month 24. The main analysis will be performed when all patients have
completed Day 100 (Visit XIV). Additional analyses are planned after completion of
the 1-year visit (end of follow-up phase I with Visit XVII) and after completion of the
safety follow-up visit 2 years after study enrolment (follow-up phase II with
Visit XVIII).
9.4. Data Safety Monitoring Board (DSMB)

A Data Safety Monitoring Board (DSMB) will be convened for this study. The DSMB
will consist of at least three medical experts. To ensure absolute autonomy of the
DSMB the members will be independent from the Sponsor and the participating
clinical study sites, and in no way involved in the performance of the study.
The primary responsibilities of the DSMB are to closely review and evaluate the
following study data for participant safety:
• Key safety endpoints (see section 6.4) immediately upon occurrence
• Interim analyses of key safety endpoints (see section 7.10.1)
• SUSARs
• Graft failure
and make, after meeting, recommendations to the Sponsor concerning the
continuation, modification, or termination of the trial. The DSMB considers study-
specific data as well as relevant background knowledge about disease, IMP and
patient population under study. The board members will confer and conclude their
assessments seeking for consensus and announcing a joint decision. The DSMB
chair person will then inform the CRO and the Sponsor about the board’s
recommendations. All decisions about the conduct of the study will rest solely with
the Sponsor in close cooperation with the coordinating investigator.
A DSMB meeting will be conducted due to reasons given below (but not limited to):
• If GVHD rates exceed pre-set statistical thresholds at the interim looks (see
section 7.10.1)
• If NRM rates exceed pre-set statistical thresholds at the interim look (see
section 7.10.1)
DSMB duties and procedures will be described in detail in the DSMB Charter. This
has to be finalized prior to study start, affirmed by each board member and approved
by the Sponsor. It will define functions, responsibilities and working procedures of the
DSMB, the frequency and modalities of telephone conferences and face-to-face
meetings, the provision of clinical data for case assessments, statistical stopping
guidelines, the decision process in case of premature study termination and
documentation of decisions reached, and any necessary functionalities and
processes.
Final 7.0 88 of 111 21. May 2015

Competent authorities, ethics committees and investigators will be informed without

delay in case of premature study termination.
9.5. Other Ethical and Regulatory Issues

During the course of the study, the Sponsor is obliged to submit to the IEC the
following:
• Substantial Amendments to the protocol
• Serious and unexpected AEs (SUSARs) and their outcomes
• Development safety update report.
If a significant safety issue is identified, either from an ICSR (individual case safety
report) or from review of aggregate data, then the Sponsor will issue prompt
notification to all parties involved, i.e. regulatory authorities, investigators and IECs.
A significant safety issue is one that has a significant impact on the course of the
clinical study or program (including the potential for suspension of the study program
or amendments to protocols) or warrants immediate update of informed consent.
9.6. Data Quality Assurance

The Sponsor will conduct a site visit to verify the qualifications of each investigator,
inspect the site facilities, and inform the investigators of responsibilities and
procedures for ensuring adequate and correct documentation.
The investigators are required to prepare and maintain adequate and accurate case
histories designed to record all observations and other data pertinent to the study for
each study participant. All information recorded in the eCRF for this study must be
consistent with the patient’s source documentation (i.e. medical records).
To ensure data quality and completeness the following have to be observed:
• Individual Case Safety Reports (ICSRs) fully documented in the data base
• Diligent follow-up of each case
• Retaining of investigator’s verbatim AE terms (documenting any Sponsor
differences)
• Consistent and accurate codification of reported terms.
The responsible person in the respective stem cell laboratories will be primarily
responsible for the quality and safety of the cell product. The quality control and
safety data will be subject to secondary post-hoc monitoring by the clinical monitor.
Should the clinical monitor detect a violation of the quality control and product
release standards, he/she will immediately inform the sponsor.
9.7. Case Report Forms and Source Documentation

All relevant data collected during the study for all of the patients enrolled in the study
will be recorded in the eCRF. The data will be entered by the responsible investigator
or a person authorized by him in a timely manner. For the safety follow-up visit at
Month 24 (Visit XVIII) additional paper CRF forms may be used if necessary due to
logistical reasons.
The physician will confirm the completeness, correctness and plausibility of the data
by his signature. All source documents from which eCRF entries are derived should
be placed in the patient’s medical records.
Final 7.0 89 of 111 21. May 2015

The original data in the eCRF for each patient will be checked against source
documents at the study site by the clinical monitor. Additions and corrections will be
dated and signed by the responsible physician or an authorized person. Reasons
must be given for corrections that are not self-explanatory.
All data are stored in a central database. Instances of uninterpretable data will be
discussed with the investigator for resolution. If corrections or additions are needed
after checking the eCRF, a corresponding query must be formulated and forwarded
to the physician for his response.
9.8. Access to Source Data

During the course of the study, a clinical monitor will perform site visits to review
protocol compliance, compare eCRF and individual patient’s medical records, assess
graft and device accountability, and ensure that the study is being conducted
according to pertinent regulatory requirements. eCRF entries will be verified with
source documentation.
Checking of the eCRF for completeness and clarity, and cross-checking with source
documents will be required to monitor the progress of the study. Regulatory
authorities, IECs and other authorized persons (e.g. CRAs monitoring the study) may
wish to carry out source data checks or on-site audit inspections. Direct access to
source data will be required for these inspections and audits. They will be performed
giving due consideration to data protection and medical confidentiality. The
investigator agrees to ensure all necessary support at all times.
9.9. Data Processing

All data will be entered in a central data base. The data review and data handling
document has to be developed during the initiation phase of the study. It will include
specifications for consistency and plausibility checks on data and will also include
data-handling rules for obvious data errors.
9.10. Archiving Study Records

According to ICH guidelines, essential documents should be retained for a minimum
of 2 years after the last approval of a marketing application in an ICH region and until
there are no pending or contemplated marketing applications in an ICH region or until
at least 2 years have elapsed since the formal discontinuation of clinical development
of the investigational product.
The essential documents of this study will be retained for a longer period if required
by the applicable local legal requirements.
9.11. Publication Policy

All information concerning this study and not previously published is considered
confidential information. All study results remain property of the Sponsor
The results of the clinical study will be published after complete data collection and
evaluation. Prof. R. Handgretinger, as the Coordinating Investigator of the study, will
be the senior author of the main publication(s). Prof. P. Lang as leading investigator
for pediatric patients and Prof. W. Bethge as leading investigator for adult patients
will be the first author(s) of the main publication(s). Trial site principal investigators
will be represented as authors of the publication according to the number of patients
Final 7.0 90 of 111 21. May 2015
recruited and followed. Central laboratories will be particularly considered, if data

analysed in these labs will be used within the publication. All further details will be
regulated by the respective agreement between the investigator and the sponsor.
Any publication has to be approved of by the Coordinating Investigator and the
Sponsor.
Final 7.0 91 of 111 21. May 2015

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Final 7.0 96 of 111 21. May 2015

A. Appendices
Appendix Title(s)
No.
Appendix 1 Glucksberg Criteria for grading of acute GvHD
Appendix 2 Grading of Chronic GvHD
Appendix 3 Karnofsy/Lansky Performance Indeces
Appendix 4 Sorror Comorbidity Index

Appendix 5 List of Study Personnel
Appendix 6 List of Clinical Study Sites and Principal Investigator
Appendix 7 List of Manufacturing Sites of the IMP
Appendix 8 List of Central Laboratories
Appendix 9 Independent Physician (NL)
Final 7.0 97 of 111 21. May 2015

Appendix 1: Glucksberg Criteria for grading of acute GVHD

Staging and grading of acute GVHD according to the Glucksberg criteria [31]:
Stage Skin Liver Gut
1 rash <25% body surface bilirubin 35–50 µM/L diarrhoea >500 mL/d to <1 L/day
2 rash 25–50% body surface bilirubin 51–100 µM/L diarrhoea/d 1–1.5 L/day
3 generalized erythema bilirubin 101–250 µM/L diarrhoea >1.5 L/day
4 generalized erythema with bilirubin >250 µM/L Severe abdominal pain with or
bullous formation and without ileus
desquamation
Grade Skin stage Liver stage Gut stage Clinical performance Comments
I 1–2 0 0 no decrease
II 1–3 1 1 mild decrease Gut or liver stage 1 or both
III 2–3 2–3 2–3 marked decrease Gut or liver stage 2–3 or both
IV 2–4 2–4 2–4 extreme decrease
Final 7.0 98 of 111 21. May 2015

Appendix 2: Grading of Chronic GvHD

Chronic GVHD Grading according to Greinix et al. [35]
1. Classification at the Time of Initial Presentation

The purpose of this classification is to identify patients with cGVHD who need long-term systemic
immunosuppression according to clinical and laboratory findings and risk factors at the time of initial
diagnosis. Long-term treatment with immunosuppression is indicated for patients with clinical
extensive cGVHD and for patients with limited cGVHD and high-risk features (i.e. platelets count
<100,000 or on-going steroids).
CLINICALLY LIMITED cGVHD
1. Oral abnormalities consistent with cGVHD, a positive skin or lip biopsy, and no other
manifestation of cGVHD
2. Mild liver test abnormalities (alkaline phosphatase ≤2 × upper limit of normal, AST or ALT ≤3 ×
upper limit of normal and total bilirubin ≤1.6 mg/100 mL) with positive skin or lip biopsy, and no
other manifestation of cGVHD
3. Less than 6 papulosquamous plaques or limited skin rash or dyspigmentation (<20% of the
body surface), positive skin biopsy, and no other manifestation of cGVHD
4. Ocular sicca (Schirmer’s test ≤5 mm), positive skin or lip biopsy, and no other manifestation of
cGVHD
5. Vaginal or vulvar abnormalities with positive biopsy, and no other manifestation of cGVHD
CLINICALLY EXTENSIVE cGVHD

1. Involvement of two or more organs with symptoms or signs of cGVHD, with biopsy
documentation of cGVHD in any organ, >15% baseline body weight loss not due to other
causes
2. Skin involvement more extensive than defined for clinically limited cGVHD, confirmed by
biopsy
3. Scleroderma or morphea
4. Onycholysis or onychodystrophy thought to represent cGVHD, with documentation of cGVHD
in any organ
5. Decreased range of motion in wrist or ankle extension due to fasciitis caused by cGVHD
6. Contractures thought to represent cGVHD
7. Bronchiolitis obliterans not due to other causes
8. Positive liver biopsy; abnormal liver function tests not due to other causes with alkaline
phosphatase >2 x upper limit of normal, AST or ALT >3 x upper limit of normal, or total
bilirubin >1.6, and documentation of cGVHD in any organ
9. Positive upper or lower GI biopsy
2. MANIFESTATIONS OF CHRONIC GVHD (DEFINITIONS OF SIGNS AND

SYMPTOMS)
In all cases, concomitant processes (i.e. infections or drug reactions) must be ruled out.
A Karnofsky Clinical Performance score [47] <60% (adult patients) or a Lansky performance score [52]
60% (children), >15% weight loss and recurrent infections are usually signs of clinically extensive
cGVHD. Abnormalities that could indicate chronic GVHD are categorized by organ system as listed
below.
Skin Erythema, dryness, pruritis, pigmentary changes (i.e. hyperpigmentation, vitiligo),
mottling, papulosquamous plaques, nodules, exfoliation, macular-papular or urticarial
rash, scleroderma, morphea (one or several circumscribed, indurated and shiny lesions)
Nails Ridging, onichodystrophy, onycholysis
Final 7.0 99 of 111 21. May 2015

Hair Premature graying (scalp hair, eyelashes, eyebrows), thinning scalp hair, alopecia,
decreased body hair
Mouth Dryness, burning, gingivitis, mucositis, striae, atrophy, erythema, lichenoid changes,
ulcers, labial atrophy or pigmentary changes, tooth decay, tightness around the mouth
Eyes Dryness, burning, blurring, gritty eyes, photophobia, pain
Vagina/vulva Dryness, dyspareunia, stricture or stenosis, erythema, atrophy or lichenoid changes not
induced by ovarian failure
Liver Elevated liver function tests not due to other causes (alkaline phosphatase ≥3 × upper
limit of normal, AST or ALT >4 × upper limit of normal or total serum bilirubin
≥2.5 mg/100 mL;
In the absence of cGVHD involving other organs, liver biopsy is required to confirm
the diagnosis!
Lung Bronchiolitis obliterans (see below, ‘diagnostic indicators’), cough, wheezing, dyspnea
on exertion, history of recurrent bronchitis or sinusitis
GI Anorexia, nausea, vomiting, weight loss, diarrhea, dysphagia, odynophagia,
malabsorption
Fasciitis Stiffness and tightness with restriction of movement, occasionally with swelling, pain,
cramping, erythema and induration (most commonly affecting the forearms, wrists and
hands, ankles, legs and feet), inablility to extend the wrists without flexing the fingers or
the elbows, contractures
Muscle Proximal muscle weakness, cramping
Skeletal Arthralgia of large proximal girdle joints and sometimes smaller joints.
LABORATORY TESTING AND DIAGNOSTIC INDICATORS OF CHRONIC GVHD

Eye Schirmer’s test with a mean value <5 mm at 5 min, or symptomatic with values of 6–
10 mm, or keratitis detected by slit lamp examination
Liver Elevated liver function tests not due to other causes (see above, definition of clinically
limited and clinically extensive cGVHD)
Lung New obstructive lung defect defined as an FEV1 <80% of predicted with either an
FEF25–75 <65% of predicted or RV >120% of predicted, or a decrease of FEV1/FVC by
>12% within a period of less than 1 year
A diagnosis of bronchiolitis obliterans requires negative microbiological tests from
bronchoalveloar lavage and evidence of air trapping by high resolution end-expiratory
and end-inspiratory CT scans of the chest.
A thoracoscopic lung biopsy may be necessary in order to confirm the diagnosis of
bronchiolitis obliterans in patients who have obstructive lung disease without air
trapping when cGVHD involving other organs is absent.
Esophagus Esophageal web formation, stricture or dysmotility demonstrated by barium swallow,
endoscopy or manometry
Muscle Elevated CPK or aldolase, EMG findings consistent with myositis
Blood Thrombocytopenia (usually 20,000–100,000 / µL), eosinophilia, hypogamma-
globulinemia, hypergammaglobulinemia and autoantibodies occur in some cases.
Final 7.0 100 of 111 21. May 2015

3. CURRENT STAGING OF CHRONIC GVHD
Organ scoring
Stage 0 I II III
__ General condition __ Asymp- __ Symptomatic, fully __ Symptomatic, __ Symptomatic;
tomatic and ambulatory; ambulatory; limited self-care;
fully active restricted only in capable of self- >50% of waking
(ECOG 0, physically care; >50% of hours in bed
KPS/Lansky strenous activity waking hours (KPS <60%)
100%) (KPS 80–90%) out of bed
(KPS 60–70%)
Skin __ No __ <20% BSA with __ 21–50% BSA __ >50% BSA
__ Maculopapular symptoms disease signs but OR involvement OR deep sclerotic
rash NO sclerotic with superficial features
__ Lichen-planus features slerotic features, ‘hidebound’
like features not ‘hidebound' (unable to pinch)
__ Papulosquamous (able to pinch) OR impaired
lichens or mobility,
icthyosis ulceration or
__ Hyperpigmen- severe pruritus
tation
__ Hypopigmen-
tation
__ Keratosis pilaris
__ Erythema
__ Erythroderma
__ Polikiloderma
__ Sclerotic features
__ Pruritus
__ Hair involvement
____________%BSA
Mouth __ No __ Mild symptoms; __ Moderate __ Severe symptoms
symptoms with disease signs symptoms with with disease
but not limiting signs with partial signs on
oral intake Iimitation of oral examination with
significantly intake major Iimitation of
oral intake
Eyes __ No __ Mild dry eyes __ Moderate dry __ Severe dry eyes,
symptoms symptoms not eyes, symptoms symptoms
Mean tear test: affecting ADL partially significantly
__ >10 (requiring affecting ADL affecting ADL
__ 6–10 eyedrops ≤3× per (requiring (special eyeware
__ ≤5 day) OR eyedops >3× to relieve pain)
__ not done asymptomatic per day or OR unable to
signs of kerato- punctal plugs) work because of
conjunctivitis sicca WITHOUT ocular symptoms
vision OR loss of vision
impairment caused by kerato-
conjunctivitis
sicca
GI tract __ No __ Symptoms such __ Symptoms __ Symptoms
symptoms as dysphagia, associated with associated with
weight: ___ kg anorexia, nausea, mild to moderate significant weight
vomiting, weigth loss (5– loss (>15%),
abdominal pain or 15%) requires nutri-
diarrhoea without tional supplement
significant weight for most calorie
loss (<5%) needs OR
esophageal
dilation
Final 7.0 101 of 111 21. May 2015

Organ scoring (continued)

Stage 0 I II III
Genital tract __ No symptoms __ Symptomatic wilth __ Symptomatic __ symptomatic
mild distinct signs wilh distinct WITH advanced
on examination signs on signs (stricture,
AND no effect on examination Iabia aggluti-
coitus and minimal AND painfull nation or severe
discomfort with effect on coitus ulceration) and
GYN examination or discomfort severe pain with
GYN coitus or inability
examination to insert vaginal
speculum
Liver __ Normal LFT __ Elevated bilirubin, __ Bilirubin __ Enzymes >5×
ALT, AST or AP >3 mg/dL or ULN
<2× ULN enzymes 2–4×
ULN
Lungs __ No symptoms __ Mild symptoms __ Moderate __ Severe
(shortness of symptoms symptoms;
FEV1: _______% __ FEV1 <80% breath after (shortness of (shortness of
DLCO: ________% OR LFS =2 climbing one flight breath after breath at rest;
of steps) walking on flat requiring O2)
ground)
__ FEV1 60–79% OR __ FEV1 ≤39 OR
LFS 3–5 __ FEV1 40–59% LFS 10–12
OR LFS 6–9
Joints / Fascia __ No symptoms __ Mild tightness of __ Tightness of __ Contractures
arms or legs, arms or legs OR WITH significant
normal or mild joint con- decrease of ROM
decreased range tractures, ery- AND significant
of motion AND not thema thought limitation of ADL
affecting ADL due to fasciitis, (unable to tie
moderate de- shoes, button
crease ROM shirts dress self
AND mild to etc.)
moderate Iimi-
tation of ADL
ADL: Activity of daily Iife. Pulmonary scoring should be performed using both the symptom and pulmonary
function testing (PFT) scale whenever possible. When discrepancies exist between pulmonary symptom and PFT
scores the higher value should be used for final scoring. Scoring used for the Lung Function Score (LFS) is
preferred, but if DLCO is not available, grading using FEV1 should be used. The LFS is a global assessment of
lung function after the diagnosis of bronchiolitis obliterans has already been established. The % predicted FEV1
and DLCO (adjusted for haematocrit but not alveolar volume) should be converted to a numeric score as follows:
>80% = 1; 70–79% = 2; 60–69% = 3; 50–59% = 4; 40–49% = 5; <40% = 6.
The LFS = FEV1 score + DLCO score with possible range of 2–12.
Other organ attendance: ____________________________________________________________

(the heaviness is rated by damage of ADL (mild = 1, moderate = 2, severe = 3)
Other·associated symptoms, organ attendances or complications

(please mark where applicable).
_________________________________________________________________________________
Severity code by grade of attendance (mild = 1, moderate = 2, severe = 3):
Final 7.0 102 of 111 21. May 2015

Appendix 3: Karnofsky / Lansky Performance Indeces

KARNOFSKY PERFORMANCE SCALE: PATIENTS >16 YEARS [47]
100% Normal, no complaints, no signs of disease
90% Capable of normal activity, few symptoms or signs of disease
80% Normal activity with some difficulty, some symptoms or signs
70% Caring for self, not capable of normal activity or work
60% Requiring some help, can take care of most personal requirements
50% Requiring help often, requiring frequent medical care
40% Disabled, requires special care and help
30% Severely disabled, hospital admission indicated but no risk of death
20% Very ill, urgently requiring admission, requires supportive measures or treatment
10% Moribund, rapidly progressive fatal disease processes
0% Death
LANSKY PLAY-PERFORMANCE SCALE: PATIENTS ≤16 YEARS) [52]
100% Fully active, normal
90% Minor restrictions in physically strenuous activity
80% Active, but tires more quickly
70% Both greater restriction of and less time spent in play activity
60% Up and around, but minimal active play; keeps busy with quieter activities
Gets dressed but lies around much of the day, no active play, able to participate in
50%
all quiet play and activities
40% Mostly in bed; participates in quiet activities
30% In bed; needs assistance even for quiet play
20% Often sleeping; play entirely limited to very passive activities
10% No play; does not get out of bed
0% Unresponsive
Final 7.0 103 of 111 21. May 2015

Appendix 4: Sorror Comorbidity Index [72]
Comorbidities Definitions HCT-CI

weighted
scores
Arrhythmia Atrial fibrillation or flutter, sick sinus syndrome, or 1
ventricular arryhthmias
Cardiac Coronary artery disease*, congestive heart failure, 1
myocardial infarction, or EF of <50%
Inflammatory bowel Crohn’s disease or ulcerative colitis 1
disease
Diabetes Requiring treatment with insulin or oral 1
hypoglycemic, but not controlled with diet alone
Cerebro-vascular Transient ischemic attacks or cerebro-vascular 1
disease accident
Psychiatric disturbance Depression/anxiety requiring psychiatric 1
consultation or treatment at the time of HCT
Hepatic – mild Chronic hepatitis, bilirubin >1.5×ULN, or AST/ALT 1
>2.5×ULN
Obesity Patients with a BMI of >35 for adults or with BMI- 1
for-age percentile of >95th percentile for children
Infection Documented infection or fever of unknown etiology 1
requiring anti-microbial treatment before, during
and after the start of conditioning regimen
Rheumatology SLE, RA, polymyositis, mixed CTD, and 2
polymyalgia rheumatica
Peptic ulcer Requiring treatment 2
Moderate/severe renal Serum creatinine >2 mg/dL, on dialysis, or prior to 2
renal transplantation
Moderate pulmonary DLCO or FEV1 66%–80% 2
or dyspnea on slight activity
Prior solid malignancy Treated at any time in the patient’s past history, 3
excluding non-melanoma skin cancer
Heart valve disease Except asymptomatic mitral valve prolapse 3
Severe pulmonary DLCO or FEV1 65% 3
or dyspnea at rest or requiring oxygen
Moderate/severe Liver cirrhosis, bilirubin >1.5×ULN, or AST/ALT 3
hepatic >2.5×ULN
Total Score
= ___________
*One or more vessel-coronary artery stenosis, requiring medical treatment, stent, or bypass graft.
EF = ejection fraction, BMI = body mass index, ULN = upper limit of normal, SLE = systemic lupus
erythmatosis, RA = rheumatoid arthritis, CTD = connective tissue disease, DLCO = diffusion capacity of
carbon monoxide, FEV1 = forced expiratory volume in one second, AST = aspartate aminotransferase,
ALT = alanine aminotransferase
Final 7.0 104 of 111 21. May 2015

Appendix 5: List of Study Personnel

Sponsor Miltenyi Biotec GmbH
Germany
Study Responsible Dr. med. Liane Preußner
Physician Phone: +49 (2204) – 8306-3950
Fax: +49 (2204) – 8306-6699
Trial Management of Dr. Sandra Karitzky
the Sponsor Phone: +49 (2204) – 8306-6560
Fax: +49 (2204) – 8306-6699
E-mail: sandrak@miltenyibiotec.de
Coordinating Professor Dr. med. Rupert Handgretinger

Investigator University Children’s Hospital
University Clinics Tübingen
Hoppe-Seyler-Str. 1
72076 Tübingen
Germany
Phone: +49 (7071) – 298-4744
Fax: +49 (7071) – 29-3966
E-mail: rupert.handgretinger@med.uni-tuebingen.de
National Coordinating Professor Dr. med. Rupert Handgretinger

Investigator, (Leiter
der klinischen
Prüfung [‚LKP‘], AMG)
Serious Adverse
Event Reporting
Statistician
Project Management
Final 7.0 105 of 111 21. May 2015

Data Management
Monitoring
Pharmacovigilance Miltenyi Biotec GmbH

Germany
Dr. med. Liane Preußner
Phone: +49 (2204) – 8306-3950
Fax: +49 (2204) – 8306-6699
Final 7.0 106 of 111 21. May 2015

Appendix 6: List of Clinical Study Sites and Principal Investigators

Site No. 1: Prof. Dr. med. Peter Lang
Tübingen (DE) Leading investigator (children)
University Children’s Hospital Tübingen
Department 1
Hoppe-Seyler-Str. 1
D-72076 Tübingen, Germany
Site No. 2: Prof. Dr. med. Wolfgang Bethge

Leading investigator (adults)
Tübingen (DE)
University Clinic Tübingen II
Department Hematology, Oncology, Immunology and Rheumatology
Otfried-Müller Str. 10
Site No. 3: Prof. Dr. med. Paul-Gerhardt Schlegel

University Clinics Würzburg
Würzburg (DE)
Pediactric Hematology and Oncology
Josef-Schneider-Str. 2
D-97070 Würzburg, Germany
Site No. 4: Prof. Dr. Stephan Mielke

University Würzburg
Würzburg (DE)
Medizinische Klinik and Poliklinik II
Oberdürrbacher Straße 6
Site No. 5: Prof. Dr. med. Ansgar Schulz

Universitätskinderklinik Ulm
Ulm (DE)
Department for Oncology, Stem Cell Transplantation
Eythstraße 24
D-89075 Ulm, Germany
Site No. 6: Prof. Dr. med. Donald Bunjes

Medizinische Universitätsklinik Ulm
Ulm (DE)
Klinik für Innere Medizin III
Albert-Einstein-Allee 23
Site No. 7: PD Dr. med. Johann Greil

Universitätskinderklinik Heidelberg
Heidelberg (DE)
Zentrum für Kinder- und Jugendmedizin
Angelika-Lautenschläger-Klinik, Klinik für Kinderheilkunde III
Im Neuenheimer Feld 430
D-69120 Heidelberg, Germany
Site No. 8: Prof. Dr. med. Niederwieser

Universitätsklinikum Leipzig AöR
Leipzig (DE)
Abt. Hämatologie/Onkologie
Johannisallee 32a
04103 Leipzig, Germany
Final 7.0 107 of 111 21. May 2015

Site No. 9: Dr. Dr. Lambros Kordelas

Universitätsklinikum Essen (AöR)
Essen (DE)
Hufelandstr. 55
45147 Essen, Germany
Site No. 10 Prof. Dr. med. Peter Bader

Universitätsklinikum Frankfurt, Goethe Universität
Frankfurt (DE)
Klinik für Kinder- und Jugendmedizin
Theodor-Stern-Kai 7
60590 Frankfurt am Main, Germany
Site No. 11 Prof. Dr. med. Roland Meisel

Universitätsklinik Düsseldorf
Düsseldorf (DE)
Klinik für Kinder-Onkologie, -Hämatologie und klinische Immunologie
Moorenstr. 5
40225 Düsseldorf, Germany
Site No. 12 PD Dr. Gernot Stuhler

Wiesbaden (DE) DKD HELIOS Klinik Wiesbaden
Fachbereich Stammzelltransplantation
Aukammallee 33
65191 Wiesbaden, Germany
Site No. 21 Prof. Dr. Jürgen Kuball

Utrecht (NL) Dept. of Hematology
UMC Utrecht
Dept. of Hematology (HP Q05.4.300)
Heidelberglaan 100
3584 CX Utrecht, The Netherlands
Additional sites may be added during the study, as necessary.
Final 7.0 108 of 111 21. May 2015

Appendix 7: List of Manufacturing Sites of the IMP / GMP

Laboratories
Dr. Michael Schumm
Tübingen* (GE)
Universitätsklinikum Tübingen
Gemeinsames Stammzelllabor
Hoppe-Seyler-Str. 1
Würzburg (GE) Prof. Dr. Matthias Eyrich

Universitätsklinikum Würzburg
Gemeinsames Stammzelllabor
Josef-Schneider-Straße 2, D30
Ulm (GE) Dr. Markus Wiesneth

Institut für klinische Transfusionsmedizin und Immungenetik Ulm
Helmholtzstr. 10
Frankfurt (GE) Dr. Halvard Bönig

DRK Blutspendedienst Baden-Württemberg - Hessen
Institut für Transfusionsmedizin und Immunhämatologie
Sandhofstraße 1
D-60528 Frankfurt am Main, Germany
Leipzig (GE) Dr. Vladan Vucinic

Universitätsklinikum Leipzig AöR
Internistische Onkologie und Hämatologie
Johannisallee 32A
04103 Leipzig, Germany
Utrecht (NL) Dr. Ineke Slaper-Cortenbach

Cell Therapy Facility
UMC Utrecht
Cell Therapy Facility (HP F03.821)
Heidelberglaan 100
3584 CX Utrecht, The Netherlands
During the course of the study additional manufacturers might be added.

* Tübingen will be the manufacturing site and the GMP laboratory for the study centers in Tübingen,
Düsseldorf, Heidelberg and Wiesbaden throughout the study.
Final 7.0 109 of 111 21. May 2015

Appendix 8: List of Central Laboratories
Additional assessments Prof.Dr. Matthias Eyrich

(TCR-Spectratyping, Universitätskinderklinik Würzburg
TREC Analysis) Hämatologisch-Onkologisches Labor, D31 UG
Josef-Schneider-Strasse 2
Additional Assessments Dr. Christiane Siewert

(T-cell activity, NK-cell activity, KIR- Miltenyi Biotec GmbH
genotyping and analysis of KIR- Friedrich-Ebert-Straße 68
repertoire) 51429 Bergisch Gladbach, Germany
Dr. Michael Schumm

Additional Assessments
Universitätsklinikum Tübingen
(MACSQuant CoreLab for immune Gemeinsames Stammzelllabor
phenotyping) Hoppe-Seyler-Str. 1
Additional Assessments Prof. Dr. Matthias Eyrich

Universitätsklinikum Würzburg
(MACSQuant CoreLab for immune Gemeinsames Stammzelllabor
phenotyping) Josef-Schneider-Straße 2, D30
During the course of the study additional central laboratories may be added.
Final 7.0 110 of 111 21. May 2015

Appendix 9: Independent Physician (NL)
Independent Physician
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Miltenyi Biotec GmbH Clinical Study Protocol

M2011-238; EudraCT No. 2011-005562-38 TCRalpha/beta-Haplo2010

CLINICAL TRIAL PROTOCOL

Final 7.0 1 of 111 21. May 2015

Liane Preußner, MD (Signature) (Date)

Declaration of the Coordinating Investigator (incl. National Coordinating

Prof. R. Handgretinger, MD (Signature) (Date)

Prof. P. Lang, MD (Signature) (Date)

Prof. W. Bethge, MD (Signature) (Date)

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A multi-center phase I/II safety and feasibility study using

Name (Investigator) (Signature) (Place, Date)

Name (Deputy) (Signature) (Place, Date)

Name (Deputy) (Signature) (Place, Date)

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3. STUDY OBJECTIVES AND ENDPOINTS...................................................................... 29

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4.5. Compliance ................................................................................................................... 41

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7.3. Analysis Sets ................................................................................................................. 80

Appendix 4: Sorror Comorbidity Index [72] .............................................................................104

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Contact Addresses and Responsibilities

List of Sponsors Study Personnel: see Appendix 5

Coordinating Investigator: see Appendix 5

National Coordinating Investigator: see Appendix 5

Safety Monitoring Board: see Appendix 5

Statistician: see Appendix 5

Project Management: see Appendix 5

Monitoring: see Appendix 5

Data Management: see Appendix 5

List of Clinical Study Sites and Principal see Appendix 6

List of Manufacturing Sites of IMP: see Appendix 7

List of Laboratories: see Appendix 8

Independent Physician (NL): see Appendix 9

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List of Abbreviations and Definitions of Terms

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HIV Human immunodeficiency virus

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Targeted Cell number ≤1 × 105/kg BW

Additional patient inclusion criteria:

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TCRγ/δ , naive Tregs, ‘memory’ Tregs, B-cells, monocytes,

NK cells , neutrophils, eosinophils;

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­ Number of grafts with ≤1 × 10 CD20 cells/kg BW

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Follow-up for patients:

2. BACKGROUND AND RATIONALE

CD34+ Enriched Stem Cell Grafts

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CD3+ and CD19+ Depleted Stem Cell Grafts

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TCRα/β+ and CD19+ Depleted Stem Cell Grafts

First clinical experiences in haploidentical HCT using TCRα/β- and CD19-depleted

Number of grafts with ≤1 × 10 CD20 cells/kg BW