A Phase I/II Study of Allogeneic Umbilical Cord Blood and Umbilical Cord Tissue-Derived Mesenchymal Stromal Cell Infusions in Children With Cerebral Palsy
Phase: PHASE1
Conditions: Cerebral Palsy
Sponsor: Joanne Kurtzberg, MD

 
 
 
April 1, 2019 
 
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A Phase I/II Study of Allogeneic Umbilical Cord Blood and Umbilical 
Cord Tissue-Derived Mesenchymal Stromal Cell Infusions in Children 
with Cerebral Palsy (IND 17921) 
 
Principal Investigators: 
Jessica Sun, MD 
Robertson Clinical & Translational Cell Therapy Program 
DTRI/Duke University 
DUMC Box 3350 
Phone:  919-668-1102 
 
Joanne Kurtzberg, MD 
Robertson Clinical & Translational Cell Therapy Program 
DTRI/Duke University 
DUMC Box 3350 
Phone:  919-668-1102 
 
 

 
 
 
April 1, 2019 
 
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Additional Investigators: 
Laura Case, DPT, MS, PCS, C/NDT 
DUMC Box 104002 
Durham, NC  27705 
Phone:  919-681-9930 
 
Kathryn Gustafson, PhD 
DUMC Box 2739 
Durham, NC  27710 
Phone:  919-684-8148 
 
Joan Jasien, MD 
DUMC Box 3986 
Durham, NC 27710 
Phone:  919-668-0477 
 
Colleen McLaughlin, DNP, CPNP 
DUMC Box 3677 
Durham, NC  27710 
Phone:  919-668-2657 
 
Mohamad Mikati, MD 
DUMC Box 3936 
Durham, NC  27710 
Phone:  919-668-0477 
 
Gordon Worley, MD 
DUMC Box 3120 
Durham, NC 27710 
 
 
 
Statistician: 
Jesse Troy, PhD 
DUMC Box 3350 
Durham, NC  27710 
Phone:  919-668-2932 
 
 
 
STCL Lab Supervisor: 
Barbara Waters-Pick, BS, MT (ASCP) 
DUMC Box 3350 
2400 Pratt Street, Suite 1300 
Durham, NC  27705 
Phone:  919-668-1178  
 
 
GMP Cell Processing Lab Supervisor: 
Norin Meadows 
DUMC Box 3350 
Durham, NC  27710 
Phone:  919-668-8956  
 
 
 
 
 
 

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TABLE OF CONTENTS 
 
INVESTIGATOR SIGNATURE PAGE .................................................................................................... 6 
PROTOCOL SUMMARY .......................................................................................................................... 7 
ABBREVIATIONS ..................................................................................................................................... 8 
1.0 
PURPOSE ...................................................................................................................................... 9 
2.0 
BACKGROUND AND HYPOTHESIS ........................................................................................ 9 
2.1 
OVERVIEW ................................................................................................................................... 9 
2.2  
CEREBRAL PALSY ..................................................................................................................... 10 
2.3  
RATIONALE FOR CELLULAR THERAPY ..................................................................................... 10 
2.4 
CELL TYPES UTILIZED IN THIS TRIAL ....................................................................................... 11 
2.4.1 
Allogeneic Umbilical Cord Blood (CB) ............................................................................... 11 
2.4.2 
Mesenchymal Stromal Cells (MSCs) ................................................................................... 12 
2.5 
ANIMAL STUDIES OF CELL THERAPY IN BRAIN INJURIES ........................................................ 14 
2.5.1 
Animal Studies in Hypoxic/Ischemic Brain Injury ............................................................. 14 
2.5.2 
Animal Studies in Stroke ...................................................................................................... 15 
2.5.3 
Animal Studies in Intraventricular Hemorrhage (IVH) ..................................................... 15 
2.5.4 
Summary ............................................................................................................................... 15 
2.6 
HUMAN STUDIES OF CELL THERAPY IN NEUROLOGIC CONDITIONS ...................................... 16 
2.6.1 
Umbilical Cord Blood in the Treatment of Inherited Metabolic Diseases ......................... 16 
2.6.2 
Safety of Autologous Cord Blood in Children with Brain Injuries .................................... 17 
2.6.3 
CB Infusion in Babies with Hypoxic Ischemic Encephalopathy (HIE) ............................ 17 
2.6.4 
CP-AC Study: A Phase II Randomized, Placebo-Controlled, Crossover Study of 
Autologous CB Infusion in Children with Cerebral Palsy .............................................................. 18 
2.6.5 
Allogeneic Sibling CB in Cerebral Palsy ............................................................................. 22 
2.6.6 
Allogeneic Unrelated Donor CB in Children with Cerebral Palsy ..................................... 23 
2.6.7 
MSCs in Cerebral Palsy ....................................................................................................... 24 
2.6.8 
Allogeneic CB in Stroke ....................................................................................................... 24 
2.6.9 
MSCs in Stroke ..................................................................................................................... 25 
2.6.10  
Unrelated Donor Cells in Autism Spectrum Disorder (ASD) ......................................... 25 
2.6.11  
MSCs in Other Neurologic Conditions ........................................................................... 26 
2.7 
SOURCE OF UNRELATED CB UNITS FOR THIS TRIAL .............................................................. 26 
2.8 
SPECIFICATIONS FOR QUALIFICATION CB UNITS ................................................................... 26 
2.9 
SOURCE OF MSCS FOR THIS STUDY:  HCT-MSC .................................................................. 27 
2.10 
STUDY RATIONALE AND HYPOTHESES .................................................................................... 30 
2.11 
STUDY DESIGN .......................................................................................................................... 30 
2.12 
RISKS AND BENEFITS ................................................................................................................ 31 
3.0 
STUDY OBJECTIVES................................................................................................................ 31 
4.0 
STUDY DESIGN ......................................................................................................................... 32 
4.1 
GENERAL DESIGN ..................................................................................................................... 32 
4.2 
STUDY FLOW CHART ................................................................................................................ 32 
4.3 
STUDY ENDPOINTS ................................................................................................................... 33 
5.0 
RESEARCH PARTICIPANT SELECTION AND WITHDRAWAL ...................................... 33 

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5.1 
STUDY POPULATION ................................................................................................................. 33 
5.2 
INCLUSION CRITERIA ................................................................................................................ 33 
5.3 
EXCLUSION CRITERIA ............................................................................................................... 33 
5.4 
RESEARCH PARTICIPANT RECRUITMENT AND SCREENING .................................................... 34 
5.5 
EARLY WITHDRAWAL OF RESEARCH PARTICIPANTS .............................................................. 34 
6.0 
STUDY PRODUCTS .................................................................................................................. 34 
6.1 
ALLOGENEIC UMBILICAL CORD BLOOD ................................................................................... 34 
6.2 
HUMAN UMBILICAL CORD TISSUE-DERIVED MESENCHYMAL STROMAL CELLS (HCT-MSC)36 
6.3 
DONOR SCREENING FOR CB AND HCT-MSC ......................................................................... 37 
6.4 
PACKAGING OF STUDY PRODUCTS .......................................................................................... 38 
6.5 
ADMINISTRATION OF STUDY PRODUCT ................................................................................... 38 
6.6 
SAFETY FOLLOW-UP ................................................................................................................. 38 
7.0 
STUDY PLAN .............................................................................................................................. 38 
7.1 
OVERVIEW ................................................................................................................................. 38 
7.2  
PATIENT SCREENING ................................................................................................................ 39 
7.3 
CB UNIT SELECTION ................................................................................................................. 39 
7.4 
STUDY PRODUCT INFUSION ..................................................................................................... 39 
7.5 
CARE DURING UNEXPECTED EVENTS ..................................................................................... 40 
7.6  
MOTOR ASSESSMENTS ............................................................................................................ 40 
7.7 
FUNCTIONAL AND QUALITY OF LIFE ASSESSMENTS ............................................................... 41 
7.8 
IMAGING ASSESSMENTS ........................................................................................................... 41 
7.9 
REQUIRED EVALUATIONS ......................................................................................................... 42 
8.0 
STATISTICAL CONSIDERATIONS ........................................................................................ 43 
8.1 
STUDY DESIGN .......................................................................................................................... 43 
8.2 
ACCRUAL ................................................................................................................................... 43 
8.3 
STUDY DURATION ..................................................................................................................... 43 
8.4 
PRIMARY AND SECONDARY ENDPOINTS ................................................................................. 43 
8.5 
SAMPLE SIZE AND POWER CALCULATIONS ............................................................................. 43 
8.6  
ANALYSIS PLAN ......................................................................................................................... 45 
8.6.1 Analysis Populations ................................................................................................................ 45 
8.6.2 Timing of Analyses ................................................................................................................... 45 
8.6.3 Demographics, Baseline Characteristics, and Disposition ..................................................... 45 
8.6.4 Analysis of the Primary and Secondary Endpoints ................................................................. 45 
9.0 
SAFETY AND ADVERSE EVENT REPORTING .................................................................. 46 
9.1 
DEFINITIONS .............................................................................................................................. 46 
9.2 
ADVERSE EVENT REPORTING .................................................................................................. 46 
9.3 
SERIOUS ADVERSE EVENT REPORTING .................................................................................. 47 
9.4 
ELICITING ADVERSE EVENT INFORMATION ............................................................................. 47 
9.5 
STOPPING GUIDELINES ............................................................................................................ 47 
9.6 
SUBJECT REPLACEMENT .......................................................................................................... 47 
10.0 
DATA SAFETY MONITORING BOARD (DSMB) ................................................................. 47 
11.0 
DATA HANDLING AND QUALITY ASSURANCE................................................................ 47 
11.1 
CASE REPORT FORMS ............................................................................................................. 47 
11.2 
VIDEO RECORDINGS ................................................................................................................. 48 
11.3 
INSPECTION OF RECORDS ........................................................................................................ 48 
11.4 
STUDY RECORD RETENTION .................................................................................................... 48 
12.0 
ADMINISTRATIVE ASPECTS .................................................................................................. 48 

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12.1 
CONFIDENTIALITY...................................................................................................................... 48 
12.2 
INSTITUTIONAL REVIEW BOARD APPROVAL ............................................................................ 49 
12.3 
MODIFICATION OF THE PROTOCOL .......................................................................................... 49 
12.4 
INFORMED CONSENT ................................................................................................................ 49 
12.5 
PROTOCOL VIOLATIONS AND DEVIATIONS .............................................................................. 49 
12.6 
STUDY REPORTING REQUIREMENTS ....................................................................................... 50 
12.7 
STUDY CONDUCT ...................................................................................................................... 50 
12.8 
PUBLICATIONS ........................................................................................................................... 50 
13.0 
REFERENCES ............................................................................................................................ 51 
14.0 
APPENDICIES ............................................................................................................................ 58 
 
 
 

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INVESTIGATOR SIGNATURE PAGE 
 
 
 
A Phase I/II Study of Allogeneic Umbilical Cord Blood and Umbilical Cord 
Tissue-Derived Mesenchymal Stromal Cell Infusions in Children with 
Cerebral Palsy 
 
 
I agree to conduct and supervise this clinical study in accordance with the 
design and specific provisions of this protocol; modifications to the study 
or protocol are acceptable only with a mutually agreed upon protocol 
amendment except when necessary to protect the safety of subjects.  I agree 
to await IRB approval for the protocol and informed consent before initiating 
the study, to obtain informed consent from subjects prior to their enrollment 
in the study.  I agree to report to responsible regulatory agencies and the 
IRB (when necessary) adverse events that occur in the course of this 
investigation.  I agree to maintain accurate and adequate records in the case 
report forms as required by this protocol and maintain those records for the 
period of time required.  I will make the study documentation available for 
safety oversight committee review and/or for other inspections as required.  
I agree to maintain study documentation for the period of time required.  I 
agree to comply with all other requirements regarding the obligations of 
clinical investigators according to FDA regulations and guidance.  I agree to 
ensure that all people assisting in the conduct of this study are informed in 
meeting the above commitments. 
 
 
 
 
(Investigator’s printed name) 
 
 
 
(Investigator’s signed name) 
 
 
 
 
 
 
 

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PROTOCOL SUMMARY 
 
Protocol Number: 
 
 
Title: 
 
A Phase I/II Study of Allogeneic Umbilical Cord Blood and 
Umbilical Cord Tissue-Derived Mesenchymal Stromal Cell 
Infusions in Children with Cerebral Palsy 
 
Study Phase: 
I/II 
 
Study Site: 
Single site; Duke University, Durham NC 
 
Study Therapy, Dosage, and 
Route of Administration: 
 
(1) Single intravenous infusion of a maximum of 
10x107/kg allogeneic umbilical cord blood (CB) cells  
(2) Three intravenous infusions of 2x106/kg human 
umbilical cord tissue cells (hCT-MSC), manufactured from 
allogeneic umbilical cord donors 
Objectives: 
Primary Objective(s):   
1. To determine the efficacy of repeated intravenous 
doses of hCT-MSC in children with cerebral palsy 
2. To determine the effect size of change in GMFM-
66 scores in subjects treated with allogeneic CB or 
hCT-MSC 
Secondary Objective(s): 
1. To determine changes in brain connectivity via 
MRI imaging in these children and whether they 
correlate with clinical response 
Research Participant Population: 
90 evaluable children ages 2-5 years with hypertonic 
cerebral palsy due to stroke, periventricular leukomalacia 
or hypoxic ischemic encephalopathy 
Study Design: 
 
Randomized, open label  
Safety Assessments/Endpoints: 
Incidence of infusion reactions, bloodstream infections, 
graft versus host disease, and alloimmunization 
 
 
 
 

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ABBREVIATIONS 
 
 
 
AE 
 
Adverse Event 
AlloCB 
 
Allogeneic Umbilical Cord Blood 
ASD 
 
Autism Spectrum Disorder 
CB 
 
Umbilical Cord Blood 
CBC 
 
Complete Blood Count 
CBU 
 
Umbilical Cord Blood Unit 
CFR 
 
Code of Federal Regulations 
CFU 
 
Colony Forming Unit 
CMV 
 
Cytomegalovirus 
CRF 
 
Case Report Form 
DMSO 
 
Dimethyl Sulfoxide 
DTI 
 
Diffusion Tensor Imaging 
EMG 
 
Electromyography 
FDA 
 
Federal Drug Administration 
fMRI 
 
Functional Magnetic Resonance Imaging 
GMP 
 
Good Manufacturing Practice 
GMFCS 
 
Gross Motor Function Classification System 
GMFM-66 
 
Gross Motor Function Measure - 66 
GvHD 
 
Graft versus Host Disease 
hCT-MSC 
 
Human Cord Tissue-Derived Mesenchymal Stromal Cells 
HIE 
 
Hypoxic Ischemic Encephalopathy 
HIV 
 
Human Immunodeficiency Virus 
HLA 
 
Human Leukocyte Antigen 
HTLV 
 
Human T-Cell Lymphotropic Virus 
ICH 
 
International Conference on Harmonisation 
IRB 
 
Internal Review Board 
IV 
 
Intravenous 
IVH 
 
Intraventricular Hemorrhage 
MCA 
 
Middle Cerebral Artery 
MRI 
 
Magnetic Resonance Imaging 
MSCs 
 
Mesenchymal Stromal Cells 
PVL 
 
Periventricular Leukomalacia 
SAE 
 
Serious Adverse Event 
STCL 
 
Stem Cell Laboratory 
SOP 
 
Standard Operating Procedure 
TNCC 
 
Total Nucleated Cell Count 
WBC 
 
White Blood Cell 

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1.0 
PURPOSE 
The main purpose of this study is to estimate change in motor function 12 months after 
treatment with a single dose of allogeneic umbilical cord blood (AlloCB) or repeated 
doses of umbilical cord tissue-derived mesenchymal stromal cells (hCT-MSC) in children 
with cerebral palsy.  In addition, this study will contribute much needed data to the 
clinical trials community on the natural history of the motor function in CP over short-
term (less than 1 year) time periods relevant to the conduct of clinical trials and assess 
the safety of AlloCB and hCT-MSC infusion in children with cerebral palsy. 
2.0 
BACKGROUND AND HYPOTHESIS 
2.1 
Overview 
Children with cerebral palsy face a lifetime of disability, resulting in enormous physical, 
emotional, and financial burdens to affected patients, their parents, and society at large.  
Typically caused by an in utero or perinatal injury to the developing brain, cerebral palsy 
is the most common – and most costly – chronic motor disorder of childhood.  The 
cornerstone of cerebral palsy treatment relies on countless hours of physical and 
occupational therapies that are entirely supportive.  There is no treatment available to 
repair the brain damage that caused the disabilities.  Thus, a novel therapy that could 
promote repair of damaged brain tissue has potential to reduce societal burden and to 
greatly improve survival, function, and quality of life for patients with cerebral palsy.   
 
Umbilical cord blood (CB) and mesenchymal stromal cells (MSCs) lessen the clinical 
and radiographic impact of hypoxic brain injury and stroke in animal models.  In patients 
with leukodystrophies undergoing unrelated donor CB transplantation, donor cells have 
been observed engrafted in the brain post-transplant.  In addition, we have observed 
ongoing myelination in the brains of these patients after initial demyelination caused by 
the underlying disease.  The central hypothesis of our work is that CB cells or human 
cord tissue-derived MSCs (hCT-MSC), acting primarily through paracrine mechanisms, 
could serve as vehicles for emerging cellular therapies in patients with brain injuries.  We 
conducted safety studies and recently completed phase II, randomized, double blind, 
placebo-controlled trial of autologous CB in children with cerebral palsy.  In that study, 
children who were infused with ≥2x107 cells/kg exhibited a greater degree of motor 
improvement than children who received lower doses or placebo.  That study was limited 
by small sample size since many children with cerebral palsy do not have a banked 
autologous cord blood unit and by the inclusion of children 1-2 years of age for whom 
analysis of the predicted motor change score was not possible.  We also conducted a 
phase I safety study of sibling CB infusion in 15 patients with cerebral palsy, indicating 
that allogeneic partially HLA-matched CB infusion is safe in this patient population.   
 
Therefore, this study is a phase I/II trial of higher doses of allogeneic CB or hCT-MSC 
to confirm our previous findings and eliminate the restriction of having an autologous CB 
unit.  The hypothesis of this trial is that adequately dosed allogeneic CB cells and hCT-
MSC administered intravenously will, via cellular or trophic effects, induce repair of brain 
injury in children with cerebral palsy, thereby improving their functional abilities.  In 
addition, data from the prior study indicates that brain connectivity as measured by MRI 
with diffusion tensor imaging (DTI) correlates with functional status and motor 
improvement in children with cerebral palsy, and therefore may be useful as a biomarker 
and predictor of response.  This relationship will be further explored in this clinical trial. 

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2.2  
Cerebral Palsy 
Cerebral palsy is a form of acquired brain injury defined as a group of disorders of 
posture and movement attributed to non-progressive disturbances in the fetal or 
neonatal brain.1  It is the most prevalent motor disorder of childhood, affecting 2 to 3 of 
every 1,000 live births.2  In the United States, approximately 10,000 babies and infants 
are diagnosed with cerebral palsy each year.  According to the United Cerebral Palsy 
Research and Education Foundation, approximately 764,000 children and adults in the 
United States have cerebral palsy (http://ucp.org/wp-content/uploads/2013/02/cp-fact-
sheet.pdf).  While some infants at risk can be identified at birth, many cases are silent in 
the newborn and only detected when the child exhibits paresis, spasticity and 
developmental delay in the first few years of life.  Affected children have varying degrees 
of functional impairments ranging from mild limitations in advanced gross motor skills to 
severely limited self-mobility despite the use of assistive technology.  Though cerebral 
palsy is a motor disorder, it can be associated with cognitive and sensory deficits and 
can have a detrimental impact on global childhood development by affecting the ability to 
explore, communicate, learn, and achieve independence.3 
 
Cerebral palsy is one of the most disabling and costly chronic conditions of childhood, 
with medical costs for children with cerebral palsy estimated at 10 times higher than 
children without cerebral palsy4 and an estimated annual cost of care of $8.2 billion for 
affected children in 2002.5  Unfortunately, no curative therapy exists.  Current treatment 
modalities focus on maximizing functional abilities and quality of life through several 
approaches.  These include non-pharmacologic measures such as physical, 
occupational and speech therapies, orthotics, optimal nutrition, constraint therapy and 
use of adaptive devices; pharmacologic interventions such as oral pharmacological 
agents, botulinum toxin injections, and oral or intrathecal baclofen; and surgical options 
including dorsal rhizotomy and various orthopedic procedures.  Most children with 
cerebral palsy require lifelong supportive care, adaptive therapy and equipment, and 
special education, utilizing greater shares of medical and societal resources as 
compared to typically developing children.  
Both full-term and premature infants can develop cerebral palsy as a result of brain 
injury sustained during the prenatal, perinatal, or postnatal periods.  Most affected full-
term infants have sustained a prenatal neurologic insult, such as an in-utero stroke, or a 
hypoxic-ischemic injury at the time of delivery.  The incidence of cerebral palsy is much 
higher in premature babies, affecting more than ten percent of very low birth weight 
infants (<1500g).6  In this population, periventricular leukomalacia, which can be 
associated with ischemic injury or vasculitis is a common cause.  Still, some children 
have no identifiable risk factors or etiology. 
2.3  
Rationale for Cellular Therapy  
Human brain development is a complex and long process, beginning in the third week of 
gestation and extending through at least adolescence, probably into adulthood.  While 
the majority of neurogenesis occurs in utero, synaptogenesis and myelination continue 
postnatally, with substantial growth and development during the first six years of life.  
Injury to the developing brain not only results in loss of normal brain cells and tissue, but 
also alters the neuro-environment, disrupting the intricate cellular and environmental 
interactions essential for healthy brain development.  There is growing evidence that cell 
therapies have the ability to influence tissue damage and repair by signaling and 
activation of host cells via trophic and/or paracrine effects.  While the exact mechanisms 
of neural sparing and/or recovery remain the subject of preclinical investigations, several 

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mechanisms have been hypothesized.7,8  The survival potential of host neural cells may 
be enhanced by the delivery of trophic factors from infused and/or transplanted cells that 
provide anti-inflammatory and neuroprotective effects.9-12  Brain plasticity may be 
increased by enhancing synaptogenesis, instigating endogenous repair mechanisms, 
stimulating angiogenesis resulting in neovascularization, and inducing migration and 
proliferation of endogenous neural stem cells.13-15 Additionally, many neurologic 
conditions involve activation of proapoptotic signal transduction, which could be 
harnessed to attract cells to brain lesions in those diseases.  Thus, CB/cord tissue-
derived cells could also potentially act as a vehicle to deliver neuroprotective and 
restorative factors or signal endogenous cells to act in a targeted way toward damaged 
brain tissue.   
 
In children with cerebral palsy, reorganization of central motor pathways has been 
demonstrated using transcranial magnetic stimulation (TMS), electromyography (EMG), 
and functional MRI (fMRI).  In patients with a unilateral lesion involving the motor tracks, 
motor control of the affected limbs is primarily provided by abnormal corticospinal 
projections from either adjacent areas in the affected hemisphere or distant regions in 
the ipsilateral, healthy hemisphere,16-18 demonstrating that an unaffected part of the brain 
is compensating for the injured portion.  Changes in the primary source of cortical 
activation on fMRI have also been described in individual patients after virtual reality 
therapy 19 and constraint-induced movement therapy, in which the unaffected limb is 
restrained while the affected limb undergoes intensive motor training.20  These 
observations indicate that neural plasticity plays an important role in the ability of 
children with cerebral palsy to recover from early brain injury.  Modulating that 
plasticity via cellular therapy may enhance such recovery.     
2.4 
Cell Types Utilized in this Trial 
2.4.1 Allogeneic Umbilical Cord Blood (CB) 
Many children who might benefit from CB therapies will not have their autologous CB 
banked. In order to extend these therapies to all patients, use of an allogeneic product 
will be necessary. Allogeneic CB has been used extensively in the field of hematopoietic 
transplantation. More recently, based on substantial preclinical data, CB has been 
investigated as a potential therapy for patients with brain injuries and neurological 
conditions. As most children and adults do not have access to autologous CB, allogeneic 
cells are frequently utilized in clinical trials of cellular therapies for neurologic conditions.  
 
Allogeneic human CB as a source of cells for hematopoietic reconstitution after 
myeloablative therapy has a proven track record of over 30 years of use in the clinic, 
with over 35,000 transplants performed.21  Allogeneic CB transplantation has been 
shown to be safe and has not been shown to cause tumors or cellular dysregulation.  
Compared to other cell sources, CB has the following advantages: 
1. CB is an abundantly available source of stem cells that can be harvested at no risk 
to the mother or infant.  It is routinely collected, cryopreserved, and banked.  
2. Public CB donors are screened for risks of transmitting infectious agents through 
blood per CFR1271, subpart C donor screening regulations.  Important infectious 
agents, particularly cytomegalovirus (CMV), are much less common in the newborn 
than adults, and are less likely to contaminate CB units. 
3. CB units, cryopreserved and banked, are available on demand, and can be easily 
shipped and thawed for use when needed, eliminating delays and uncertainties that 
complicate bone marrow collection from unrelated donors. 

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4.  CB lymphocytes are more immunotolerant of a new host.  Thus, the intensity of 
graft-versus-host reactivity of fetal lymphocytes is less than that of adult cells, so 
transplantation of CB after myeloablation and immunosuppression results in less 
graft versus host disease (GvHD) than transplantation of bone marrow or other adult 
hematopoietic stem cell sources. 
5. CB contains pluripotent stem cells that have demonstrated the ability to differentiate 
into numerous types of cells throughout the body, including in the brain.  Thus CB 
may provide a source of cells for non-hematopoietic tissue repair or regeneration. 
 
In this study, we propose to use allogeneic CB donor without the use of chemotherapy or 
other immunosuppressive therapies. 
2.4.2 Mesenchymal Stromal Cells (MSCs) 
MSCs are a heterogeneous group of undifferentiated, pluripotent cells that can be 
isolated from several different tissues including bone marrow, adipose tissue, and birth 
tissues (umbilical cord tissue, placenta).  While MSCs can give rise to mesodermal 
tissue types including bone, cartilage, and fat, their primary mechanism of action is 
thought to result from immunomodulatory and other paracrine effects.  MSCs have 
demonstrated a multitude of immunomodulatory effects on both humoral and cell-
mediated immune responses.  These include, but are not limited to, inhibiting B-, T-, NK, 
dendritic-cell, and microglial proliferation, decreasing pro-inflammatory cytokine 
production, and blocking neutrophil recruitment.  In addition, numerous preclinical 
studies using MSCs transplantation for diseases of the central nervous system suggest 
that MSCs can act through release of different neurotrophic, anti-inflammatory, and anti-
apoptotic factors to promote recovery the injured area and prevent further damage.22-26   
 
Despite their ability to modulate the immune response, MSCs themselves have low 
immunogenicity.  MSCs express low levels of MHC class I molecules on their surface 
and lack the expression of MHC class II and several costimulatory molecules.  This 
allows MSCs to be used in the allogeneic setting across HLA barriers, without the need 
for donor-recipient HLA matching.  In fact, in a review of 13 human studies of 
intravenous allogeneic MSC administration, including 1,012 mostly adult patients, there 
were no reports of infusional toxicity,27 supporting the notion that MSCs are “immune-
privileged” and can avoid immunological allorecognition.  When utilized as a therapeutic 
cell, MSCs exert effects via trophic signaling.  It is estimated that after infusion, MSCs 
survive in the recipient for up to 4 months.  MSCs do not engraft in the recipient. 
 
Safety of MSCs: 
MSCs manufactured from bone marrow and adipose tissue have been studied in 
hundreds of clinical trials worldwide involving thousands of individuals and a wide variety 
of human conditions.  Their safety has been demonstrated repeatedly, and is 
summarized in a 2012 systematic review and meta-analysis27 of 36 clinical trials in 14 
countries using MSCs in over 1,000 recipients with cardiovascular, neurological, 
oncologic, metabolic, gastrointestinal, and post-transplant conditions.  Based on a 
combination of randomized controlled trials and uncontrolled studies with follow-up 
periods of weeks to up to 5 years, there was no association between MSC treatment and 
acute infusional toxicity, organ system complications, infection, death, or malignancy.  
The only side effect associated with MSC treatment was transient fever, which did not 
cause any long-term sequelae.  Importantly, no malignancies were reported in patients 
without a prior history of cancer.   
 

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After intravenous administration, MSCs initially distribute to the lungs.  While there is a 
theoretical risk of respiratory complications, the only randomized trial in which acute 
pulmonary reactions have been observed was conducted in patients with chronic 
ischemic heart failure who received intracoronary MSCs.28  Thus, this risk seems to be 
limited to patients whose underlying condition makes them susceptible to developing 
pulmonary edema.  Although a few cardiovascular studies delivering MSC via 
intramuscular injection have reported arrhythmias, the incidence was not statistically 
different from control groups in meta-analysis.27  
 
Experience with MSCs in Children: 
The most well-studied MSC product given to children to date is a bone marrow-derived 
product originally manufactured by Osiris as Prochymal® and subsequently acquired by 
Mesoblast as Remestemcel-L used to treat patients with GvHD after a hematopoietic 
stem cell transplant.  Prochymal® was approved in Canada in 2012 for use in children 
with acute GvHD who have failed to respond to steroid treatment.  A recent study of the 
product enrolled 241 children (median age 9.6 years) with refractory GvHD from 2007-
2014.29  Eight MSC doses (2x106 cells/kg/dose) were given intravenously over a four 
week time period, with four additional weekly infusions in patients who demonstrated a 
partial or mixed response.  There were no incidences of ectopic tissue formation in 2434 
total doses, with follow-up of 2-9 years.  The most frequent severe adverse events were 
infections (24%) and respiratory disorders (16%), common issues in post-transplant 
patients.  There were only 11 severe adverse events and one infusion reaction related to 
MSC treatment.  Patients with primary steroid-refractory GvHD who had only been 
treated with steroids demonstrated an 81% response rate to MSCs.  Overall, 65% of 
patients demonstrated an improvement in their GvHD, with higher response rates in 
patients with less severe disease.  This demonstrates that repeated doses of MSCs are 
safe and well-tolerated in children, and suggests that they provide anti-inflammatory and 
immunomodulatory effects in the setting of GvHD. 
 
In a recent study, a German group pooled bone marrow cells from eight healthy donors 
to create an MSC cell bank.30  They treated 26 children, ages 1-19 years old with 
steroid-refractory GvHD, with a total of 81 doses of MSCs from this cell bank (median 3 
doses per patient, median 2.2x106/kg MSCs IV per dose).  Treatment was well-tolerated, 
with only one reported headache and one episode of nausea across all doses.   Six 
patients died from causes unrelated to MSC treatment (disease progression, GvHD, 
sepsis, thromboembolism).  Seventy-seven percent of patients responded to therapy, 
defined as a complete or partial remission of their GvHD.  
 
Others are investigating the use of MSCs in patients with other conditions.  There have 
been case reports of intracoronary administration of autologous bone marrow-derived 
MSCs in children, ages 4 months to 14 years, with severe dilated cardiomyopathy and/or 
heart failure with subsequent short- and intermediate-term improvement in clinical 
condition and B-type natriuretic peptide values.31-33  Studies have also been conducted 
in patients with type 1 diabetes.34  One such study administered umbilical cord-derived 
MSCs intravenously in 29 patients, including several children, with newly diagnosed type 
1 diabetes.35  Treatments were well-tolerated, and moderate improvements in some 
metabolic measures were observed.  Early phase studies of MSCs are also underway in 
preterm infants with bronchopulmonary dysplasia.  In the only published study, cells 
were delivered intratracheally and no significant side effects were reported in these 
young babies.36 None of these early phase studies have reported toxicity related to the 
MSC therapy.   

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2.5 
Animal Studies of Cell Therapy in Brain Injuries  
Numerous animal models have demonstrated both neurological and survival benefits of 
cell therapies in the setting of stroke, ischemia, and intracranial hemorrhage.37-40 These 
injuries differ in that some are focal (i.e. stroke) and others are global (i.e. hypoxia), but 
all are typically characterized by immediate damage to all neural cell types within the 
affected region accelerating a cascade of events that lead to demyelination and necrosis 
of brain tissue.  Inflammation, apoptosis, neuronal and oligodendrocyte death, and 
astrocytosis are all operative in mediating damage resulting from these insults. 
Neuroprotection, neovascularization, and neuronal regeneration have all been 
demonstrated after cell administration in various models.15,37,41  
2.5.1 Animal Studies in Hypoxic/Ischemic Brain Injury 
Cellular therapy has been studied in models of hypoxic ischemic encephalopathy (HIE). 
In a neonatal rat model that results in severe cerebral damage and contralateral spastic 
paresis after unilateral carotid artery ligation on day seven of life, intraperitoneal CB 
mononuclear cells administered one day after the hypoxic event migrate to the area of 
brain damage and persist for at least two weeks.  Although the extent of morphologic 
injury on gross pathology was not altered, animals who received CB mononuclear cells 
did not develop spastic paresis, indicating functional recovery.39  In a baby rabbit model 
of HIE via uterine artery occluion,42 Tan demonstrated that human CB administered nine 
days after the injury improved gross motor function in functional assays (figure 1).43  A 
possible dose effect was also observed, with less improvement detected at lower doses.  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Figure 1:  Rabbit model of cerebral palsy and intravenous CB administration. 
Panel A: Experimental model.  The uterine artery is occluded 9 days prior to delivery, resulting in a cerebral palsy 
phenotype.  Kits are delivered just prior to term and receive IV CB cells on the day of birth.  Motor assessments 
are performed on days 1, 5, and 11. Panel B: CB administration improved neurobehavioral scores at days 5 and 
11 in the severe group compared to saline and media. Panel C: CB administration improved neurobehavioral 
scores at days 5 and 11 in the mild group.  *p<0.05, **p<0.01.  (Drobyshevsky et al, Dev Neurosci, 2015) 
  A 
*
**
*
*
*
**
**
**
**
*
*
*
*
**
*
C 
B 

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In a murine model of HIE, bone marrow-derived MSCs delivered into the brain 
parenchyma resulted in reduced lesion size and improved behavioral outcomes even 
when treatment was started 10 days after the insult.  Evidence of increased endogenous 
cell proliferation and decreased microglial proliferation was observed.44 
2.5.2 Animal Studies in Stroke 
The most extensively studied models involve brain damage resulting from permanent 
middle cerebral artery occlusion (MCAO) in adult rats or transient occlusion 
accompanied with hypoxia in neonatal rats or mice.  Intravenous injection of CB can 
greatly mitigate the damage caused by such acute hypoxic/ischemic brain injury.45  
 
In addition to evaluating treatment response and further elucidating mechanism of 
action, many preclinical studies have attempted to address other critical aspects of cell 
administration, including dose, timing, and route of delivery.  In addition to functional 
outcomes, Vendrame examined anatomic measures of infarct volume after MCAO in the 
presence of increasing doses of CB cells.37  At 4 weeks after infusion, infarct volume 
measurements revealed an inverse relationship between CB cell dose and damage 
volume, reaching significance at a dose of 107 cells, thereby demonstrating a dose-
dependent relationship between CB cell dose, behavioral improvement, and neuronal 
sparing.  Also in an MCAO model, Chen showed a greater improvement in 
somatosensory behavior and neurologic dysfunction when CB or bone marrow stromal 
cells were administered at one day versus seven days after the injury, suggesting that 
earlier may be better.38  In fact, the majority of stroke models have evaluated stem cell 
therapy in the acute or subacute setting, immediately to several days after the insult. 
However, Shen demonstrated improvement in functional outcomes in rats treated 
intravenously with bone marrow MSCs one month after MCAO stroke.  In this model, 
scar tissue was reduced and the number of proliferating cells and oligodendrocyte 
precursors in the area of injury were increased, possibly indicating neurogenesis and 
myelination.46  This suggests that although the ideal timing of cellular therapy for stroke 
or other brain injury is still unknown, benefits may be attainable long after the injury is 
sustained.  Models delivering cells via the intravenous, intra-arterial, intracerebral, and 
intranasal routes have not demonstrated that any one mode of delivery is significantly 
superior in terms of functional outcomes.47-49  
2.5.3 Animal Studies in Intraventricular Hemorrhage (IVH) 
Intraventricular hemorrhage (IVH) is common in preterm babies and often results in 
periventricular leukomalacia and subsequent development of cerebral palsy.  Ballabh 
and colleagues developed a rabbit model of IVH by administering glycerol 
intraperitoneally to premature rabbit pups.50  In this model, IVH is followed by the 
development of hydrocephalus and subsequent white matter demyelination.  
Intraventricular administration of human CB cells 24 and 72 hours after glycerol failed to 
prevent the hydrocephalus, but did reduce subsequent demyelination (Ballabh, personal 
communication, 2014).  In a neonatal rat model of IVH induced by intraventricular 
injection of autologous blood, mice given umbilical cord tissue-derived MSCs 
intravenously two days after IVH exhibited less reactive gliosis and hypomyelination and 
significant behavioral improvement compared to control animals.51  
 
2.5.4 Summary  
In summary, xenogeneic infusion of human CB cells and cord tissue-derived cells in 
small animals following brain injury results in improved survival and functional outcomes.  

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Optimal dose, timing, and route of administration as well as specifics regarding 
mechanism of action are still the subject of preclinical investigations, and may vary 
based on the type, degree, and time interval since the insult.  Nonetheless, these studies 
suggest that dose may play an important role, that intervention in the chronic phase of 
injury can still be beneficial, and that intravenous administration may be adequate. 
2.6 
Human Studies of Cell Therapy in Neurologic Conditions 
2.6.1 Umbilical Cord Blood in the Treatment of Inherited Metabolic Diseases 
Allogeneic transplantation of human CB in patients with genetic lysosomal and 
peroxisomal storage diseases is effective in preventing or ameliorating the associated 
neurological damage.52-55  The engraftment of donor cells into a patient with an inherited 
metabolic disease provides a constant source of enzyme replacement, thereby slowing 
or halting the progression of disease.  Patients with these diseases, ranging in age from 
newborns to young adults, transplanted early in the course of their disease derive 
extensive benefits from the transplant procedure, which both extends life (for decades) 
and greatly improves neurologic functioning.56-58  Infusion of CB cells in these cases are 
proceeded by immunosuppression to support the goal of durable engraftment.   
 
In collaboration with Dr. Evan Snyder and with informed consent of her parents, we 
examined the brain of a 20-month-old female baby who died after sex-mismatched CB 
transplant for symptomatic Krabbe disease.  Immunocytochemical staining for cell type-
specific markers were combined with FISH using probes for X and Y chromosomes and 
DAPI staining to affirm host versus donor cells within the brain.  Numerous male cells 
were found distributed throughout the forebrain, brainstem, cerebellum and cortex.  
Donor cells were also seen within blood vessels and transversing through and 
incorporated into the choroids plexus.  Many stained with macrophage markers and a 
few donor-derived oligodendrocytes were seen.  Globoid bodies, the pathological 
perivascular signature of Krabbe disease, were eliminated.59  This observation, as well 
as animal studies that demonstrate donor-derived cells in the brain and multiple other 
organs after hematopoietic stem cell transplant, indicates that the transplanted stem 
cells are capable of repopulating more than just the hematopoietic system and are not 
limited by the blood-brain barrier.60 
 
Cortical White Matter 
 
Choroid plexus
Figure 2:  Brain sections of a female Krabbe patient who died 1o months post UCBT. XY Fish demonstrates 
male (donor, blue dots) cells engrafting in and differentiating in brain. 

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2.6.2 Safety of Autologous Cord Blood in Children with Brain Injuries  
At our institution, we have already treated several hundred children ages one day to nine 
years with or at risk for cerebral palsy with autologous CB.  The first 184 patients were 
described in 2010 as a Phase I study.61  Between 3/2004 and 12/2009, 184 infants and 
children with HIE, cerebral palsy, congenital hydrocephalus and other brain injuries 
received 198 autologous cord blood infusions at Duke (14 patients received two 
infusions based on availability of larger cell doses).  Autologous CBUs were obtained 
from 24 different cord blood banks:  149 (81%) CBUs came from 11 private U.S. banks 
(113 from two of the larger private U.S. banks), 13 (7%) CBUs from 11 international 
banks, and 22 (12%) CBUs from 2 public banks.  The majority of parents had elected to 
store their child’s cord blood privately when they were born.  The median volume of cord 
blood collected was 60 mL (range 5-180 mL) and median TNC contained in the cord 
blood, as reported by the cord blood bank at the time of cryopreservation, was 4.7x108 
(range 0.3-33.8x108) total nucleated cells and 1.8 x106 (range 0-19.1 x106) CD34 cells.  
All infusions were administered through a peripheral IV after premedication with oral 
Tylenol 15mg/kg, IV Benadryl 0.5mg/kg and IV Solumedrol 0.5mg/kg.  Median post thaw 
recovery of TNC was 82% (range 13-200%), and patients were dosed with a median of 
2.0x107 TNC/kg (range 0.1-13.3x107), 0.7x105 CD34+ cells/kg (range 0.04-6.4x105), and 
6.5x104 CFU/kg (range 0-315x104).   
 
Three patients (1.5%) experienced hypersensitivity reactions during their CB infusion 
characterized by wheezing with or without urticaria two to ten minutes after the IV 
infusion was initiated. The reactions resolved after discontinuation of the infusion and 
treatment with additional IV Benadryl and bronchodilators. The remainder of the CB cells 
were discarded for two of the infusions stopped prior to completion; one patient was able 
to restart and complete the CB infusion. One patient’s mother experienced an allergic 
reaction consisting of urticaria, presumably due to contact with DMSO exhaled onto her 
face and neck by her child receiving a CB infusion. The reaction resolved with oral 
Benadryl. No adverse events have been reported.  Specifically, no infections, 
autoimmune diseases, tumors, or other adverse events have been observed.   
 
This series demonstrated safety and feasibility of autologous CB infusion, and anecdotal 
reports of improved function were common.  However, there was potential for a 
considerable placebo effect as parents often endorsed dramatic benefit even shortly 
after infusion. Therefore, a randomized, double blind, placebo-controlled trial was 
performed to evaluate objective functional outcomes. This study is described in detail 
below. 
2.6.3 CB Infusion in Babies with Hypoxic Ischemic Encephalopathy (HIE) 
In phase I trial of newborns with HIE at birth conducted at Duke, fresh, non-
cryopreserved autologous CB processed on Sepax 1 (Biosafe, Geneva) was infused in 
1, 2, or 4 doses within the first 72 hours of life in babies with moderate-to-severe 
encephalopathy qualifying for systemic hypothermia.62  These babies (n=39) were 
compared to a concomitant group of babies who were also cooled at Duke but did not 
receive CB cells (n=146).  Infusions were found to be safe in these critically ill babies, 
and babies receiving cells had increased survival rates to discharge (100% vs. 89%, 
p=0.03).  Of the 25 cell recipients with known one-year outcomes, 16 (64%) survived 
with Bayley III scores ≥ 85 in all three domains, and of the 63 cooled-only infants with 
known one year outcomes, 25 (40%) survived with Bayley III scores ≥ 85 in all three 

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domains (p = 0.04).  A phase II randomized, placebo controlled, multicenter trial is 
currently underway.  
2.6.4 CP-AC Study: A Phase II Randomized, Placebo-Controlled, Crossover 
Study of Autologous CB Infusion in Children with Cerebral Palsy 
Table 1:  Characteristics of 
Patients and Autologous CB 
Units 
Autologous 
CB Group 
(N=32) 
Placebo 
Group 
(N=31) 
Patient Characteristics 
Age, years – median (range) 
2.1 (1.1-6.2) 
2.3 (1.1-7.0) 
Sex – no. (%) 
 
 
    Male  
20 (62.5) 
22 (71) 
Race – no. (%) 
 
 
  Caucasian 
27 (84.4) 
28 (90.3) 
Type of Cerebral Palsy – no. (%) 
 
 
    Hypotonic Quadraplegia 
1 (3.1) 
3 (9.7) 
    Spastic Diplegia 
6 (18.8) 
6 (19.4) 
    Spastic Hemiplegia 
15 (46.9) 
15 (48.4) 
    Spastic Quadraplegia 
10 (31.3) 
7 (22.6) 
GMFCS Level* – no. (%) 
 
 
    I/II 
21 (65.6) 
21 (67.7) 
    III/IV 
11 (34.4) 
10 (32.3) 
Bayley Cognitive Score (n=43) Τ – 
median (range) 
85 (55-110) 
90 (55-110) 
Cord Blood Characteristics – median (range) 
Collection Volume, mL 
66 (4.5-146) 
 
Pre-Cryo TNCC, x108 
4.4 (1.1-15.5) 
 
Pre-Cryo Cell Dose, x107/kg 
3.00 (1.11-8.68) 
 
Cell Dose Infused, x107/kg 
1.98 (0.75-4.83) 
 

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The CP-AC study was a 
prospective, randomized, 
double blind, placebo controlled 
crossover study of a single intravenous infusion of autologous CB in children ages 1-6 
years with cerebral palsy.63 The Gross Motor Classification System (GMFCS) was 
utilized to classify the level of motor function at study entry and follow-up. Children were 
eligible if they were (1) GMFCS level 2-4 or (2) GMFCS level 1 with hemiplegia if they 
used their affected hand as an assist only. Children with known genetic conditions, 
intractable seizures, or severe microcephaly were ineligible. Autologous CB units had to 
have a documented precryopreservation total cell dose of 1-5x107/kg, negative sterility 
culture, and negative maternal infectious disease screening. Subjects were evaluated at 
baseline, one-, and two-years with motor evaluations and brain MRI. They were 
randomized to the order in which they received CB and placebo infusions (given one 
year apart). The primary endpoint was change in Gross Motor Function Measure-66 
(GMFM-66) score one year after the initial infusion (CB or placebo). Infusions, dosed at 
1-5x107/kg based on the precryopreservation total nucleated cell count (TNCC), were 
administered intravenously over 5-10 minutes in the outpatient setting after 
premedication with Tylenol, Benadryl, and Solumedrol.  Subjects received IV fluids and 
were monitored for 2-4 hours post-infusion.  
 
Results:  Sixty-three patients were enrolled and randomized to an initial infusion of 
autologous CB (n=32) or placebo (n=31). Median age at initial infusion was 2.1 years.  
Etiology of CP was classified into the following categories: periventricular leukomalacia 
(n=17), in utero stroke or bleed (n=27), ischemic injury (n=7), or other (n=12). One-third 
of patients had moderately severe GMFCS levels (III-IV) at study entry. The two 
treatment groups were balanced with respect to age, gender, race, type and severity of 
cerebral palsy. Patient and CB characteristics are shown in Table 1. Despite negative 
pre-cryopreservation cultures, one CB unit grew β-hemolytic strep at the time of thaw. 
There were no clinical infections. One subject had transient infusion reactions consisting 
of hives +/- low-grade fever after each infusion; an additional dose of Benadryl was 
administered after the first reaction.   
 
Analysis of the 63 patients at one year showed no difference in GMFM-66 change 
scores between placebo and treated groups (6.9 vs. 7.5, p=0.72).  However, treated 
subjects who received above the median infused cell dose of 1.98x107/kg demonstrated 
improvement in GMFM-66 change scores compared to subjects who received lower cell 
doses (p=0.05) (Figure 3).  The infused cell dose was not correlated with age (p=0.70) or 
type (p=0.32) or severity (p=0.46) of cerebral palsy.   
 
CD34+ Dose Infused, x105/kg 
0.60 (0.11-3.90) 
 
CFU Dose Infused, x105/kg 
3.91 (0.04-36.21) 
 
Figure 3:  GMFM-66 Scores by Randomized Treatment Assignment and Infused Cell Dose 
Panel A: Distribution of GMFM-66 score at baseline and 1 year in patients randomized to placebo 
and autologous cord blood.  Lines connect the group means (circles) over time. Panel B: GMFM-
66 change scores based on median cell doses (Precryopreservation doses: Low, <3x107/kg vs. 
High, >=3x107/kg; Infused doses: < 1.98x107/kg vs. High: >= 1.98x107/kg). Panel C: One year 
Observed-Expected GMFM-66 scores in patients >2 years of age at baseline based on infused 
cell dose. Panel D: PDMS-2 gross motor quotient change scores based on infused cell dose. 

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Using a subject’s baseline GMFM-66 score, GMFCS level, age, and published 
percentiles,64 we calculated the expected one-year GMFM-66 score for each patient.  
Since such percentile values are only available for children ages ≥2 years, one-year old 
subjects (n=25) were excluded from this analysis.  In the evaluable patients (n=38), the 
difference between the actual one-year GMFM-66 score and the expected one-year 
GMFM-66 score was then calculated.  In the entire autologous CB group, the median 
actual-expected difference in GMFM-66 scores was 1.7 (range -6.1-14.5), versus 2.2 
(range -6.2-12.6) in the placebo group.  When the CB group was analyzed by infused 
cell dose, subjects who received above the median infused cell dose of 1.98x107/kg 
(n=9) improved a median of 4.3 points greater than expected (p=0.05 vs placebo), 
whereas subjects who received below the median infused dose improved a median of 
1.9 points less than expected (p=0.02 vs high dose; p=0.07 vs placebo, figure 3C). 
 
We then used the two year data to further explore the effect of cell dose by comparing 
the difference between observed and expected GMFM-66 scores one year after CB 
infusion in all subjects who were ≥2 years old when they were treated (n=46), regardless 
of when the infusion was given (baseline or one year). In this analysis, the dose 
relationship from the primary analysis was confirmed:  subjects who received ≥2x107 
cells/kg (n=23) improved a median 3.6 points (IQR -0.4 to 4.5) greater than expected, 
whereas subjects who received <2x107/kg did not improve beyond expectation (median  
-1.1, IQR -3.7 to 1.3, p=0.003, figure 4A). 
 

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Fifty patients were also eligible for analysis of the Peabody Developmental Motor 
Scales-2 (PDMS-2), which assesses motor skills in children from birth through age 5, 
one year post initial treatment. The median one-year change from baseline in the Gross 
Motor Quotient was 1.0 in the autologous CB group and -0.5 in the placebo group 
(p=0.39).  When the subjects treated with autologous CB were analyzed by infused cell 
dose (>/< 1.98x107/kg), a significant change was detected in the Gross Motor Quotient 
both at one year post-randomization (figure 3D) and one year post-CB infusion, 
regardless of the timing of the infusion (3.0 vs. 0, p= 0.02, figure 4B).  
 
 
 
Imaging Results:  MRI data was analyzed to explore relationships between change in 
GMFM-66 scores, total brain connectivity, and cell dose.  Accurate anatomical image 
parcellation could not be obtained in approximately one-third of subjects due to 
substantial morphologic brain abnormalities, leaving 23 treated patients and 15 placebo 
patients with usable connectivity data.  There were no statistically significant differences 
in type of cerebral palsy, GMFCS level, or age between patients with and without 
analyzable images. As previously described,65 there was a moderate correlation 
between change in GMFM-66 score and total connectivity at one-year in all analyzable 
subjects (n=38, Spearman r=0.53; 95% CI: 0.25, 0.73; p<0.001). Total connectivity 
change was not related to baseline GMFCS level, typography of cerebral palsy, or 
gender, but was inversely correlated with age (Spearman r=-0.52; 95% CI: -0.72, -0.23; 
p=0.001).  In the two-year analysis when all evaluable subjects were examined by cell 
dose, patients who received ≥2x107 TNCC/kg (n=19) demonstrated a statistically 
significant greater increase in normalized whole brain connectivity one year after 
treatment than children who received lower doses (n=19; p=0.04, figure 5).  In the 
sensorimotor network, nodes with significant increases in connectivity that correlated 
with improvement in GMFM-66 scores included the pre- and post-central gyri, basal 
ganglia, and brain stem. 
 
Figure 4:  Gross Motor Function One Year after CB Infusion by Cell Dose 
High dose >=2x107/kg, low dose <2x107/kg.  Panel A: Observed-Expected GMFM-66 scores one 
year after treatment in patients >2 years of age at the time of CB infusion. Panel B: PDMS-2 gross 
motor quotient change scores based on infused cell dose. 

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Results of this trial suggest that when administered in sufficient doses (>1.98x107/kg), 
autologous CB may improve motor function in young children with cerebral palsy.  The 
study was limited by small sample size of the dosing groups and the inclusion of children 
1-2 years of age for whom analysis of the predicted motor change score was not 
possible.   
2.6.5 Allogeneic Sibling CB in Cerebral Palsy   
Given the benefit in motor improvement observed in the CP-AC study utilizing 
autologous CB, and recognizing that most children do not have a suitably qualified 
autologous CB unit available, we conducted a phase I clinical trial to evaluate the safety 
of fully HLA-matched or haploidentical allogeneic sibling CB infusion in children with 
cerebral palsy (ClinicalTrials.gov ID NCT02599207).  Fifteen children (9 female, 5 male), 
ages 1-6, with spastic CP were enrolled and treated. Children were eligible if they were 
(1) GMFCS level 2-4 or (2) GMFCS level 1 with hemiplegia if they used their affected 
hand as an assist only. Children with known genetic conditions, intractable seizures or 
severe microcephaly were ineligible. Sibling CB units had to have a precryopreservation 
total nucleated cell count (pTNCC) ≥2.5x107/kg, negative sterility cultures, negative 
maternal infectious disease screening and be a ≥4/8 HLA match with the participant. 
 
Figure 5: Brain Connectivity via MRI/DTI   
Panel A: Change in normalized whole brain connectivity one year after treatment. Panel B: 
Connectome representation. The nodes and edges included are those that demonstrated 
significantly increased improvement in children receiving high doses compared to those receiving 
low doses, as indicated by the color chart, with insignificant nodes shown in gray. 
D
A 
B 

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Participants were evaluated at baseline and 6 months with functional evaluations 
(GMFM, Peabody), brain CT, and laboratory studies. On the day of infusion, sibling CB 
units were thawed and washed. After premedication with Tylenol, Benadryl and 
Solumedrol, participants received a dose of ≥2.5x107/kg cells, based on the pTNCC, 
intravenously over 5-10 minutes in the outpatient setting. Participants received IV fluids 
and were monitored for 1-2 hours post-infusion. Safety assessments were conducted 24 
hours, 2 weeks, and 3, 6 and 12 months post-infusion. 
 
The median baseline age of participants was 3.7 years (range 1.4-6). Sibling CB 
infusions had a median TNCC of 4.3x107/kg (range 1.8-5.2) and median CD34 dose of 
0.6x105/kg (range 0.1–1.8). Four CB infusions were full HLA matches, 11 were 
haploidentical. All CB units had negative post-thaw sterility cultures. There were no 
acute infusion reactions and no unexpected imaging findings. No platelet antibodies, 
donor-specific HLA antibodies, or donor cells were detected in peripheral blood six 
months after infusion. With a median follow-up of 11 months, there were a total of 34 
adverse events in 13 participants. Most (19/34) were attributed to common childhood 
infections, and none were related to the CB infusion. The only serious adverse event 
was unrelated to the infusion and occurred in a participant with a history of seizures who 
was hospitalized for a prolonged febrile seizure 5 months after CB infusion. Six months 
after infusion, participants improved a mean of 4.8 (SD 2.5) points on the GMFM-66 and 
1 (SD 2.9) point on the Peabody Gross Motor Quotient. This study confirms that partially 
or fully-HLA matched allogenenic sibling CB infusion is safe and feasible in young 
children with CP.  
2.6.6 Allogeneic Unrelated Donor CB in Children with Cerebral Palsy   
A study of allogeneic CB was conducted in 105 Korean children with cerebral palsy.66  
This study had three groups:  allogeneic CB + cyclosporine + erythropoietin; 
erythropoietin alone; and a control group.  One severely affected patient died in her 
sleep 14 weeks after CB administration, and this was determined to be unrelated to the 
CB infusion.  Eight other patients experienced serious adverse events requiring 
hospitalization (pneumonia–4, seizure–1, influenza–2, urinary tract infection–1), but the 
distribution did not differ between groups.  Non-serious adverse events that were more 
common in the CB group were pneumonia and irritability.  At one year of follow-up, there 
were no reported prolonged or delayed serious adverse events.  The authors reported 
greater improvements in cognitive and select motor functions in children who received 
CB and erythropoietin versus controls.  There was no CB-only group for comparison.   
  
The same group subsequently conducted a study of allogeneic CB versus placebo in 36 
children ages 6 months to 20 years with cerebral palsy.67  In this study, the CB group 
showed greater improvements in gross motor performance at 6 months compared to the 
placebo group, and motor outcomes were positively correlated with the dose of CB cells.   
 
In a Russian study, 80 patients ages 1-12 years with cerebral palsy received 1-6 
intravenous infusions of allogeneic CB with an average dose of 2.5x108 viable cells per 
infusion.68  Most patients who received four or more infusions showed improvement in 
tone, motor, and/or cognitive function, but there was no control group for comparison.  In 
their series, factors that impacted treatment response included age, severity of brain 
damage, and number of CB infusions. 
 

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In summary, data from early phase clinical trials demonstrate that CB infusion is safe in 
young children with cerebral palsy and suggest that it may be effective in improving 
motor function when given intravenously at adequate doses. 
2.6.7 MSCs in Cerebral Palsy 
MSCs derived from both bone marrow and umbilical cord tissue have been investigated 
in cerebral palsy in a few small studies in China.69-71  In one study, autologous bone 
marrow-MSCs were injected intrathecally and/or into brain parenchyma of 46 children 
ages six months to 15 years with cerebral palsy.70  The only side effects reported were 
transient low-grade fever and wound aches.  That study demonstrated a statistically 
significant change in GMFM-66 percentile at one, six and 18-months post-treatment, 
though there was no placebo or natural history cohort for comparison.      
 
In a case report, one patient with cerebral palsy was treated with seven doses of 5-
10x106 MSCs generated from the umbilical cord tissue of her younger sibling and 
administered via the intravenous and intrathecal routes.69  This patient also experienced 
temporary low-grade fever after the intrathecal injections but no other side effects.  She 
was followed for 28 months after the final treatment, and demonstrated motor 
improvements during that time.   
 
In another study, eight twin pairs of children with spastic cerebral palsy received four 
intrathecal injections of allogeneic, unrelated umbilical cord tissue-derived MSCs.71  
Improvements in GMFM-88 scores were noted six months after treatment.  Interestingly, 
the improvement within each member of the twin pair was correlated, suggesting a 
possible hereditary component to the response to therapy.    
2.6.8 Allogeneic CB in Stroke 
The CoBIS study was an IRB approved, FDA IND sponsored, prospective, open-label, 
multi-center, Phase 1 safety study of a single intravenous infusion of non-HLA matched, 
ABO matched, allogeneic CB in 10 adults ages 18-80 years old.  CB units were 
selected by ethnicity, blood type, and ability to supply a dose of 0.5 – 1.5 x 107 TNCC/kg.  
Eligible patients included those experiencing a recent, acute cortical, hemispheric, 
ischemic stroke in the middle cerebral artery (MCA) distribution as detected by MRI as a 
diffusion weighted abnormality and are enrolled if their National Institutes of Health 
Stroke Scale (NIHSS) is 8-15 (right hemisphere) or 8-18 (left hemisphere).  Participants 
who received tPA or undergo mechanical perfusion are eligible for inclusion. Participants 
were not pre-treated with immunosuppressive drugs. The primary endpoint was safety 
as assessed by the frequency and severity of adverse events within 24 hours of CB 
infusion and during the 12 months post CB infusion.  Secondary outcome measures 
included Modified Rankin Scale (mRS), NIHSS, the Barthel Index (BI), and European 
Quality of Life (EQ-5D-3L), Patient Health Questionnaire Scale (PHQ8), Telephone 
Interview for Cognitive Status (TICS), and a self-reporting survey of rehabilitation 
therapy.  MRI was used to evaluate changes in the brain 3 months post infusion.  
  
There were no serious adverse events in any of the 10 participants enrolled and treated 
with allogeneic unrelated donor non-HLA matched, ABO matched CB within 10 days of 
their stroke. This safety data suggests intravenous infusion of unmatched, allogeneic, 
CB cells is well tolerated.  A randomized, placebo controlled Phase 2 study is underway 
with the goal of using unrelated non-HLA matched CB to down-regulate inflammation 
and promote neuroprotection and neurorepair in patients with ischemic stroke.    

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2.6.9 MSCs in Stroke 
Several small studies using systemically administered autologous bone marrow-derived 
MSCs have been conducted in adult patients with acute or chronic stroke, with no 
significant side effects.72  A phase II study of an allogeneic MSC product (MultiStem) 
was recently conducted in 126 adult patients with stroke (65 treated, 61 placebo).73  
MSC therapy was well-tolerated.  While there was no difference between placebo and 
treated patients in measures of stroke recovery, the treatment group had a lower rate of 
mortality and infections, associated with down regulation of inflammatory biomarkers 
including IL-6.  Patients who received MSCs earlier (24-36 hours post-stroke vs. 36-48 
hours post-stroke) demonstrated more favorable recovery than patients who received 
later treatment or placebo.     
2.6.10  Unrelated Donor Cells in Autism Spectrum Disorder (ASD) 
At Duke, we are currently conducting a phase II randomized, double-blind, placebo 
controlled trial of autologous or allogeneic CB infusion in young children with ASD 
(DukeACT).  Participants are randomized to the order in which they receive CB and 
placebo infusions, given six months apart. Children who have an adequate autologous 
CB unit stored are assigned to receive an autologous CB infusion; those without an 
available autologous CB unit are assigned to receive an infusion of allogeneic, unrelated 
donor CB from the Carolinas Cord Blood Bank with the same donor screening and HLA 
matching requirements set forth in this protocol.  
 
A blinded safety analysis was performed after the first 100 participants were enrolled in 
the DukeACT study.  Patients who enroll in DukeACT are randomized in a 2:1 ratio to 
CB (autologous or allogeneic) or placebo.  With up to 12 months of follow-up, the safety 
profile of CB infusions has been similar to our prior experience.  There have been no 
Serious Adverse Events, graft versus host disease, or product-related infections.  As of 
August 2017, there were a total of 166 Adverse Events (AEs) reported in 100 
participants. One hundred and twenty-two events (73.5%) were Mild, 42 (25.3%) were 
Moderate, and 2 (1.2%) were Severe. Six events (3.6%) in 3 subjects (3.0% of enrolled 
patients, 4.4% of subjects reporting events) were Related to study product. The six 
related events are coded as Immune System Disorders/Allergic Reaction to Excipient.  
One Severe event reported as “Hypoxia, Transient” was related to an allergic reaction to 
the study product. The other reported Severe event was a “Fall” Unrelated to the study 
product.  Overall, 22/38 subjects (57.9%) with an autologous cell source had AEs and 
40/62 (64.5%) of subjects with an allogeneic cell source had AEs, the vast majority of 
which were related to common childhood ailments. The most frequently occurring 
Unrelated events were: Pyrexia (16 events), Upper Respiratory Infection (11 events), 
Otitis Media (8 events), and Aggression (8 events).  
In China, 37 autistic children, ages 3-12 years, were enrolled in a study utilizing 
allogeneic CB mononuclear cells (CBMNC) and/or umbilical cord-derived mesenchymal 
stem cells (UCMSC).74  The children were divided into three groups:  CBMNC (n=14), 
CBMNC+UCMSC (n=9), and control (n=14).  All children received standard rehabilitation 
therapy.  Cells were given in four doses, 5-7 days apart, via intravenous and/or 
intrathecal administration.  In the cell recipients, transient fever (5/23) was the only 
reported adverse event related to the therapy.  Compared to control patients at six 
months, they observed greater improvements in the Childhood Autism Rating Scale 
(CARS) in the CBMNC+UCMSC group and in the Clinical Global Impression scale (CGI) 
in both cell groups.  This was a small, non-randomized study designed primarily to 
assess safety.  Nonetheless, the functional data suggests the potential for benefit.  

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2.6.11  MSCs in Other Neurologic Conditions 
A few clinical trials of autologous MSC therapy have been conducted in patients with 
multiple sclerosis.  Of 25 patients with progressive multiple sclerosis treated with a single 
intrathecal injection of autologous bone marrow-derived MSCs, the disease stabilized in 
half of patients over a one-year time period.75  Side effects, all transient and self-limited 
including low-grade fever, nausea/vomiting, lower limb weakness, and headache, were 
likely related to the intrathecal route of administration and were consistent with those 
reported from other small studies of intrathecal MSC treatment in patients with multiple 
sclerosis.76,77 Another study that treated 10 patients with progressive visual deficits due 
to multiple sclerosis with a single intravenous dose of autologous bone marrow-derived 
MSCs demonstrated improvements in visual acuity, visual evoked response latency, and 
optic nerve area.78  
 
One clinical trial has been conducted in seven patients with Parkinson’s Disease who 
received a single dose of autologous bone marrow-derived MSCs transplanted to the 
subventricular zone using stereotaxic surgery.  With a follow-up of 10 to 36 months, 
three patients demonstrated an improvement in disease symptoms. Two patients also 
reported subjective improvement of symptoms and reduction in drug dosage.79  
2.7 
Source of Unrelated CB Units for this Trial 
The Carolinas Cord Blood Bank (CCBB) is one of the largest public cord blood banks in 
the nation. Established in 1998 with support from the National Heart and Blood Institute 
of the NIH, the CCBB has over 30,000 CB units in inventory and has distributed over 
2,500 CB units for transplant to date. In 2012 the CCBB received approval from the FDA 
for its BLA application to market DUCORD, a stem cell product derived from umbilical 
cord blood, for use in transplants between unrelated donors and recipients. DUCORD is 
approved for use in hematopoietic stem cell reconstitution for patients with disorders 
affecting the hematopoietic system that are inherited, acquired, or result from 
myeloablative treatment. The CCBB currently collects from 10 hospital sites (8 in North 
Carolina, 1 in Atlanta, GA and 1 in Boston, MA). It also accepts CB donations from 
mothers delivering in any hospital in North Carolina and Atlanta through a kit donation 
program. 
2.8 
Specifications for Qualification CB Units  
Based on established criteria utilizing allogeneic CB for hematopoietic stem cell 
transplantation and our experience in treating more than 600 children with autologous 
CB for neurological conditions, we have established the following criteria to qualify 
banked CB units for cell therapy studies. CB units utilized for this current study will be 
obtained from the Carolinas Cord Blood Bank.  It is the first choice to utilize CB units 
from the Carolinas Cord Blood Bank.  However, if a qualifying CB unit cannot be found 
from the Carolinas Cord Blood Bank, another accredited bank may be used to locate a 
matching CB unit with the following criteria listed below:      
 
The CB unit must have: 
 
1. Pre-cryopreservation total nucleated cell count (TNCC) documented and at least 
12 x 107/kg.  We target CB units that are at least 12 x 107/kg, however, we will 
accept units that are at least 10 x 107/kg.       
2. Pre-cryopreservation viability ≥85% of total cells and ≥70% of CD34+ cells (if 
performed)  
3. Pre-cryopreservation sterility culture performed and negative 

AlloCB and hCT-MSC in Cerebral Palsy 
 
April 1, 2019 
 
Page 27 of 61 
4. Maternal infectious disease screening as follows:  Testing must include negative 
results for Hepatitis B, Hepatitis C, HIV, HTLV, and syphilis.  Additional 
screening, which is dependent on the timing of the CB collection, may be 
performed based on local and national regulations. Units from mothers who have 
a positive CMV antibody screen may be used.   
5. Test sample available for identity confirmation and potency testing 
6. HLA typing performed and meets study-specific parameters 
7. CD45+ viability ≥40% and CD34+ viability ≥70% on thawed test sample 
 
These same criteria will be utilized for this clinical trial and, along with procedures for CB 
administration, are detailed again in section 6.0. 
2.9 
Source of MSCs for this Study:  hCT-MSC 
hCT-MSC is a third party MSC product manufactured from allogeneic donor digested 
umbilical cord tissue that is expanded for two passages in culture, cryopreserved, stored 
in the vapor phase of liquid nitrogen, and banked.  The umbilical cord tissue is donated 
by healthy mothers delivering healthy full term babies after a normal pregnancy with 
written informed consent.  The cells are manufactured, cryopreserved and stored in the 
Robertson CT2 GMP laboratory (Duke University, Durham, NC).  
 
Umbilical cord tissue is an attractive source of MSCs as it is readily available and easily 
obtained without consequence to the donor, is non-controversial, has a higher 
proliferative potential than MSCs from other postnatal sources.80  Numerous preclinical 
studies have not demonstrated any evidence of tumorigenicity or toxicity of cord tissue-
derived MSCs.81  A summary of early phase clinical trials published in English that 
utilized cord tissue-derived MSCs is shown in Table 1.  Among these 36 studies 
including 695 patients and at least 1,416 doses of cord tissue-derived MSCs with follow-
up ranging from three months up to six years, no severe adverse events were reported.  
Several more clinical trials of cord-tissue derived MSCs in various disease conditions are 
underway (clinicaltrials.gov). 
 
 

AlloCB and hCT-MSC in Cerebral Palsy 
 
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Table 1:  Umbilical Cord Tissue-Derived MSCs Studied in Humans 
Report 
Indication 
# of 
treated 
patients 
Ages, yr 
(range 
or mean) 
Route of 
administration 
Dose 
# of doses 
(interval) 
Duration 
of f/u 
(months) 
AEs due to 
CT-MSCs 
Li, 201674,82 
Autism 
20 
3-14 
IT 
1x106/kg 
2 
(5-7 days) 
6 
transient fever 
(n=4) 
Xie, 201683 
Hypoxic Ischemic 
Encephalopathy 
12 
45-68 
IV 
1x108 
1 
6 
none 
Wang, 201684 
Juvenile Idiopathic 
Arthritis 
10 
2-15 
IV 
4x107 
2 
(3 months) 
12 
none 
Chen, 201685 
Psoriasis 
2 
26-35 
IV 
1x106/kg 
1, 5 
48, 60 
none 
Cai, 201634  
Type I Diabetes 
21 
5-28 
Intra-arterial 
(pancreatic artery) 
1.1x106/kg 
1 
12 
none 
Hu, 201686 
Type 2 Diabetes 
31 
42-63 
IV 
1x106/kg 
2 
(monthly) 
36 
none 
Hua, 201687 
Spinal Cord Injury 
1 
25 
IT 
1x107 
4 
(3 days) 
12 
none 
Kim, 201688 
Atopic Dermatitis 
34 
29 
Subcutaneous 
2.5-5x107 
1 
3 
none 
Qin, 201689 
Diabetic Foot 
28 
N/A 
Endovascularly 
N/A 
N/A 
3 
none 
Wang, 201690,91 
Lupus 
40 
17-54 
IV 
1x106/kg 
2 
(weekly) 
12-72 
 
none 
Zhao, 201592 
Heart Failure 
30 
53.2 
Intracoronary 
NR 
1 
6 
none 
Liang, 201593 
Radiation Myelitis 
1 
37 
IV, IT 
5.2x107 IV, 
1.1x107 IT 
1 
18 
none 
Wang, 201571 
Cerebral Palsy 
16 
4-12 
IT 
1-1.5x107 
4 
6 
NR 
Li, 201594 
Coronary Artery 
Occlusion 
15 
N/A 
Intracoronary 
3-5x106 
1 
24 
none 
Rajput, 201595 
Duchenne Muscular 
Dystrophy 
11 
5-18 
IV & IM 
1x106/kg 
4 
(monthly) 
36 
none 
Gu, 201596 
Lupus Nephritis 
58 
12-55 
IV 
1x106/kg 
1 
12 
none 
Pan, 201597 
Bone Marrow Necrosis 
1 
11 
Intrabone 
2x107 
1 
38 
none 
Wu, 201598 
GvHD 
24 
14-44 
IV 
0.5-1x106/kg 
1 
1-24 
none 
Zhu, 201599 
Leukemia, undergoing 
haplo HSCT 
25 
4-17 
IV 
1-1.4x106/kg 
1 
3-25 
none 
Miao, 2015100 
Neurological disorders 
88 
2-68 
IT 
NR 
4-6 
(5-7 days) 
NR 
transient HA, 
fever, back/leg 
pain 

AlloCB and hCT-MSC in Cerebral Palsy 
 
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Report 
Indication 
# of 
treated 
patients 
Ages, yr 
(range 
or mean) 
Route of 
administration 
Dose 
# of doses 
(interval) 
Duration 
of f/u 
(months) 
AEs due to 
CT-MSCs 
Li, 2015101 
Becker Muscular 
Dystrophy 
3 
6-46 
IV 
3-5x107 
1 
3 
none 
Wang, 2015102 
Hemorrhagic Cystitis 
7 
11-38 
IV 
0.8-
1.6x106/kg 
1-3 
NR 
none 
Cheng, 2014103 
Spinal Cord Injury 
10 
35.3 
Spinal Cord 
Injection 
2x107 
2 
(10 days) 
6 
none 
Chang, 201336 
Bronchopulmonary 
Dysplasia 
9 
10 days 
Intratracheal 
1-3x107 
1 
3 
none 
Li, 2014104 
Multiple Sclerosis 
13 
42 
IV 
4x106/kg 
3 
(2 weeks) 
12 
none 
Kong, 2014105 
Type 2 Diabetes 
18 
 
IV 
N/A 
3 
6 
slight fever 
Wang, 2013106 
Traumatic Brain Injury 
20 
5-48 
IT 
1x107 
4 
(5-7 days) 
6 
HA, dizziness 
Wang, 2013107 
Primary Biliary 
Cirrhosis 
7 
33-58 
IV 
0.5x106/kg 
3 
(monthly) 
12 
none 
Zhang, 2013108 
HIV 
7 
26-49 
IV 
0.5x106/kg 
3 
(monthly) 
12 
none 
Jin, 2013109 
Spinocerebellar Ataxia 
16 
 
IV & IT 
N/A 
N/A 
12 
none 
Wu, 2013110 
Leukemia, undergoing 
CBT 
8 
3-12 
IV 
2-10x106/kg 
1 
8-27 
none 
Jiang, 2013111 
Stroke 
4 
40-59 
Intra-arterial (MCA) 
2x107 
1 
6 
none 
Hu, 201335 
Type I Diabetes 
15 
17.6 
IV 
1.5-3.2x107 
1 
24 
none 
Shi, 2012112 
Liver Failure 
24 
24-59 
IV 
0.5x106/kg 
3 
(monthly) 
18 
none 
Zhang, 2012113 
Liver Cirrhosis 
30 
25-64 
IV 
0.5x106/kg 
3 
(monthly) 
12 
none 
Qu, 2009114 
Bone Nonunion 
36 
36 
Intrabone 
1x106 - 
1x107 
1 
13 
none 
 
 
 
 
 
 
 
 
 
Totals:   
    36 studies 
 
695 
0-68 
IV:  362 
IT:  162 
Other:  199 
 
 >1,416  
3-72 
4 studies 
(3 IT route) 
 
Abbreviations:  HA-headache, IM-intramuscular, IT-intrathecal, IV-intravenous, N/A-not available, NR- not recorded 
 

AlloCB and hCT-MSC in Cerebral Palsy 
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2.10 Study Rationale and Hypotheses 
As described above, previous studies suggest that adequately dosed autologous CB 
infusion can improve motor function in children with cerebral palsy. As it is not feasible 
that every child with cerebral palsy will have access to their autologous CB, this study 
will assess efficacy of two allogeneic sources of cells that can be available to all patients 
in need. The major goal of this study is to investigate change in motor function 12 
months after treatment with two allogeneic cell sources, allogeneic CB and hCT-MSCs. 
This study will generate important data regarding the effect size of change in motor 
function of these two cell sources and a natural history cohort to aid in the planning of 
future trials. The rationale for the study and for the potential benefit of cell therapy in 
cerebral palsy is based upon the following hypotheses: 
 
 
 
We have demonstrated safety and preliminary evidence of potential dose-dependent 
efficacy of autologous CB infusions in children with cerebral palsy. 
 
It is possible that different cell types, e.g. cord blood mononuclear cells versus cord 
tissue MSCs, may influence brain connectivity by different mechanisms. 
 
Multiple doses of cells may be superior to a single dose of cells. 
 
The developing brain exhibits remarkable plasticity, making young children ideal 
candidates for deriving maximal therapeutic benefit from restorative therapies, 
including CB.  
 
CB cells, acting through paracrine mechanisms, may facilitate endogenous repair 
mechanisms and promote formation of new neural connections the motor cortex 
resulting in significant clinical improvements. 
 
Brain connectivity plays an important role in the pathophysiology, and potentially 
mechanism of repair, of brain injury in children with cerebral palsy.  Specifically, we 
hypothesize that (1) impairments in brain connectivity account for the motor deficits 
in children with cerebral palsy, (2) increases in brain connectivity have a direct 
impact on functional improvements, (3) children with cerebral palsy who receive CB 
infusions will exhibit greater increases in brain connectivity than children who receive 
placebo infusions, and (4) the severity of baseline brain connectivity abnormalities 
predict the potential for benefit of CB therapy.   
2.11 Study Design 
This study is a phase I/II, prospective, randomized, open-label trial designed to assess 
the effect size of change in GMFM-66 score in subjects treated with hCT-MSC or 
allogeneic CB and assess the safety of repeated doses of hCT-MSC in young children 
with cerebral palsy.  Children ages 2-5 years with cerebral palsy due to hypoxic ischemic 
encephalopathy, stroke, or periventricular leukomalacia may be eligible to participate.  
All participants will ultimately be treated with an allogeneic cell product at some point 
during the study.  Participants will be randomized to one of three arms:  (1) the “AlloCB” 
arm will receive one allogeneic CB infusion at the baseline visit; (2) the “MSC” arm will 
receive three hCT-MSC infusions, one each at baseline, three months, and six months; 
(3) the “natural history” arm will not receive an infusion at baseline but will receive an 
allogeneic CB infusion at 12 months.  All participants will have an initial clinical 
evaluation to verify and classify the diagnosis of cerebral palsy and determine eligibility.  
They will return for study visits an additional two (AlloCB and natural history arms) or 
three (MSC arm) times.  Outcome measures, described in detail in section 7.6, will be 
assessed at baseline, six-months, and one-year time points.  Additional safety endpoints 
will be assessed remotely for 12 months after the final in-person visit. 

AlloCB and hCT-MSC in Cerebral Palsy 
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2.12 Risks and Benefits  
CB cells will be thawed, washed, and infused using standard operating procedures that 
have been used in over 35,000 individuals worldwide.  The potential risks associated 
with infusion of allogeneic CB cells include a hypersensitivity reaction to the product 
(rash, shortness of breath, wheezing, difficulty breathing, hypotension, swelling around 
the mouth, throat or eyes, tachycardia, diaphoresis) to the product or transmission of 
infection.  Additional risks associated with infusion of allogeneic CB cells include 
hypertension, bradycardia, anaphylaxis, hematuria, acute hemolytic reaction, rejection of 
cells, immune dysregulation (develop of HLA directed antibodies), and development of 
graft-versus-host disease.  All CB units are screened for the risk of transmitting 
infectious agents and must meet release criteria prior to infusion, as described below.  
Participants must have normal immune function and will not receive immunosuppressive 
therapy prior to or after infusion of CB or hCT-MSC.   
 
Potential risks associated with MSC infusion include a reaction to the product (rash, 
shortness of breath, wheezing, difficulty breathing, hypotension, swelling of the mouth, 
throat or eyes, tachycardia, diaphoresis), infection transmission, and HLA sensitization.  
Theoretical risks that must be considered but have not been associated with MSC 
administration include the possibility of immune suppression and ectopic tissue 
formation.  CB collected with the cord tissue used to manufacture hCT-MSC is screened 
for the risk of transmitting infectious agents, and the product must meet release criteria 
prior to infusion, as described below.  Participants must have normal immune function 
and will not receive immunosuppressive therapy prior to or after infusion of hCT-MSC.   
 
Participants who undergo a brain MRI may require moderate sedation or, rarely, general 
anesthesia, in order to complete the MRI and infusion.  Risks associated with anesthesia 
include nausea, vomiting, blood vessel injury, nerve injury, lung injury, heart attack, 
allergy to medications, brain damage, respiratory insufficiency, hypoxia, hypotension, 
and anaphylaxis, and death.  These associated risks are described in detail in the 
consent form. Risks associated with blood draws and IVs include momentary discomfort 
or pain, bruising, infection, bleeding, clotting, and fainting.  Risks associated with genetic 
testing include medical, psychosocial, and economic risks, effects on insurability and 
employability, limits on educational options, and social stigma.      
 
Potential benefits of this intervention include the possibility that the cells may, via direct 
or indirect mechanisms, induce changes in vivo that result in motor improvement. 
3.0 
STUDY OBJECTIVES 
Primary Objective:  To determine the effect size of change in GMFM-66 score in children 
with cerebral palsy treated with a single dose of approximately 10 x107 cells/kg of 
allogeneic CB or three doses of 2x106 cells/kg of hCT-MSC.  
 
Secondary Objective:  To assess the safety of repeated doses of hCT-MSC in children 
with cerebral palsy. 
 
Exploratory Objectives:   (1) To determine the change in the Peabody Developmental 
Motor Scale-2 (PDMS-2) score at 6 and 12 months in children treated with allogeneic 
CB or hCT-MSC.  (2) To analyze the change in normalized total brain connectivity, as 
measured by brain MRI with DTI, from baseline to 12 months.  (3) To assess changes 
functional and quality of life measures at 6 and 12 months. 

AlloCB and hCT-MSC in Cerebral Palsy 
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4.0 
STUDY DESIGN 
4.1 
General Design 
This study is a phase I/II, prospective, randomized, open-label trial designed to determine the 
effect size of change in GMFM-66 score in subjects treated with hCT-MSC or allogeneic CB and 
assess the safety of repeated doses of hCT-MSC in children with cerebral palsy.  Children ages 
2-5 years with cerebral palsy due to hypoxic ischemic encephalopathy, stroke, or periventricular 
leukomalacia may be eligible to participate.  All participants will ultimately be treated with an 
allogeneic cell product at some point during the study.  Participants will be randomized to one of 
three arms:  (1) the “AlloCB” arm will receive one allogeneic CB infusion at the baseline visit; (2) 
the “MSC” arm will receive three hCT-MSC infusions, one each at baseline, three months, and 
six months; (3) the “natural history” arm will not receive an infusion at baseline but will receive 
an allogeneic CB infusion at 12 months.  Motor outcome measures will be assessed at baseline, 
six-months, and one-year time points.  Safety will be evaluated at each infusion visit and 
remotely for an additional 12 months after the final visit.  Duration of study participation will be 
24 months from the time of baseline visit.  Randomization to treatment arms will be stratified by 
GMFCS level at study entry and etiology of CP (Stroke vs. Other). . 
 
4.2 
Study Flow Chart 
 
*MRI will be performed in a subset of participants 
24 Month Safety 
Evaluations 
12 Month Evaluations     
+/- Infusion 
6 Month Evaluations        
+/- Infusion 
3 Month Infusion 
Baseline Infusion & 
Evaluations  
Randomization 
Enrollment 
CBU Screening 
Patient Screening 
Review 
Records, Labs, 
Videos, Photos 
Review Report, 
HLA & Potency 
testing 
Consent 
AlloCB 
CB 
Motor, MRI* 
No infusion 
Motor Eval 
No infusion 
Motor, MRI* 
MSC 
hCT-MSC 
Motor, MRI* 
hCT-MSC 
hCT-MSC 
Motor Eval 
No Infusion 
Motor, MRI* 
Natural 
History 
No infusion 
Motor, MRI* 
No infusion 
Motor Eval 
CB 
Motor, MRI* 
Remote Safety Evaluation 
Qualifying Visit 

AlloCB and hCT-MSC in Cerebral Palsy 
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4.3 
Study Endpoints 
Primary Endpoint:   
The primary endpoint is the difference between a participant’s observed and expected 
changes in GMFM-66 score 12 months after the initial study infusion. Interval estimates 
will be reported separately for the hCT-MSC, AlloCB, and Natural History arms. 
Expected GMFM-66 scores at 12 months will be calculated based on the participant’s 
baseline age, GMFCS level, and GMFM-66 score at study entry using published 
reference percentiles.64 
 
Secondary Endpoints:   
The secondary endpoint is the number of adverse events occurring over the 12-month 
period post-infusion with hCT-MSC or AlloCB. 
 
Exploratory Analyses: 
 
Observed GMFM-66 score at baseline, 6, and 12 months 
 
Change in the Peabody Developmental Motor Scale-2 (PDMS-2) score at 6 and 
12 months. 
 
Change in normalized total brain connectivity, as measured by brain MRI with 
DTI, from baseline to 12 months. 
 
Change in functional and quality of life measures at 6 and 12 months. 
5.0 
RESEARCH PARTICIPANT SELECTION AND WITHDRAWAL 
5.1 
Study Population 
Ninety evaluable children ages 2-5 years with hypertonic cerebral palsy. 
5.2 
Inclusion Criteria 
1. Age ≥24 months and ≤60 months adjusted age at the time of enrollment.   
2. Diagnosis:  Unilateral or bilateral hypertonic cerebral palsy secondary to in utero or 
perinatal stroke/hemorrhage, hypoxic ischemic encephalopathy (including, but not 
limited to, birth asphyxia), and/or periventricular leukomalacia. 
3. Performance status:  Gross Motor Function Classification Score levels I – IV 
4. Review of brain imaging (obtained as standard of care prior to study entry) does not 
suggest a genetic condition or brain malformation. 
5. Legal authorized representative consent. 
5.3 
Exclusion Criteria 
1. Available qualified autologous cord blood unit. 
2. Hypotonic or ataxic cerebral palsy without spasticity. 
3. Autism and autistic spectrum disorders. 
4. Hypsarrhythmia. 
5. Legally blind. 
6. Intractable seizures causing epileptic encephalopathy. 
7. Evidence of a progressive neurologic disease. 
8. Has an active, uncontrolled systemic infection or documentation of HIV+ status. 
9. Known genetic disease or phenotypic evidence of a genetic disease on physical 
exam. 
10. Concurrent genetic or acquired disease or comorbidity(ies) that could require a future 
allogeneic stem cell transplant.  

AlloCB and hCT-MSC in Cerebral Palsy 
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Page 34 of 61 
11. Requires ventilatory support, including home ventilator, CPAP, BiPAP, or 
supplemental oxygen. 
12. Impaired renal or liver function as determined by serum creatinine >1.5mg/dL and/or 
total bilirubin >1.3mg/dL except in patients with known Gilbert’s disease. 
13. Possible immunosuppression, defined as WBC <3,000 cells/mL or absolute 
lymphocyte count (ALC) <1500 with abnormal T-cell subsets. 
14. Patient’s medical condition does not permit safe travel. 
15. Previously received any form of cellular therapy. 
5.4 
Research Participant Recruitment and Screening 
Patients may be recruited through IRB-approved advertising for the study on the 
websites of CB banks, parent sponsored websites, the NMDP website, selected cerebral 
palsy societies, local medical providers, and through a record of inquiries for previous 
studies (brain injury database. Separate IRB approval will be obtained for any 
advertisements.  
Screening for this study is conducted under a separate, IRB-approved screening 
protocol (Pro00063563). Under this protocol, after written informed consent is obtained 
from a parent/guardian, the patient’s medical records, videos, and results of brain 
imaging are obtained and reviewed. The medical review is conducted by a team of 
pediatric nurses, nurse practitioners, and physicians to identify the presence of any 
exclusion criteria. If no exclusion criteria are identified, screening labs are performed and 
a search may be conducted to identify a suitably matched CB unit (see section 6.1). 
5.5 
Early Withdrawal of Research Participants 
Criteria for Removal from Protocol Therapy:  
1. Diagnosis of a genetic disease while under evaluation or on study.   
2. Change in medical condition that precludes study participation.   
 
Patients who are off protocol therapy are to be followed until they meet off-study criteria 
(see below). Follow-up data will be obtained on off-protocol participants unless consent 
is withdrawn. Participants that are taken off study prior to infusion of the CB will be 
considered not evaluable and can be replaced with another participant.  
 
Off-Study Criteria:  
1. Death.   
2. Lost to follow-up.   
3. Withdrawal of consent for any further data collection.   
4. Completion of the final study visit.   
6.0 
STUDY PRODUCTS 
6.1 
Allogeneic Umbilical Cord Blood 
Allogeneic unrelated donor CB units utilized for this trial will be obtained from the 
Carolinas Cord Blood Bank, an FDA licensed Public Cord Blood Bank at Duke University 
Medical Center.  However, if a qualifying CB unit cannot be found from the Carolinas 
Cord Blood Bank, another accredited bank may be used to locate a matching CB unit 
with the matching criteria.  CB donors must be eligible for donation to a public cord blood 
bank for allogeneic use.  Donor eligibility screening via questionnaires is performed in 
accordance with CFR 1271.75 and infectious disease testing is performed in accordance 

AlloCB and hCT-MSC in Cerebral Palsy 
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with CFR 1271.80 and 1271.85.  The unit must also have an appropriate degree of HLA 
matching and meet product specifications as detailed below.   
 
HLA Matching 
All potential study participants will undergo high resolution HLA typing at HLA-A, B, and 
HLA-DRB1 via blood or buccal swab. Patients receiving allogeneic CB will have HLA 
typing performed on two separate samples for confirmation. Allogeneic units that are 
potential matches will initially be identified from a search of the Carolinas Cord Blood 
Bank, or another accredited bank  The best available HLA-matched (≥4/6), using 
intermediate level matching at HLA Class I A and B and high resolution-allele level 
matching at HLA Class II, DRB1, CB unit with a pre-cryopreservation nucleated cell dose 
≥12 x 107 cells/kg will be selected (we target CB units that are at least 12 x 107/kg, 
however, we will accept units that are ≥10 x 107/kg.)  Once a unit is selected, HLA typing 
will be used to confirm the original HLA typing and to select the best matching unit. 
When possible, at least 1 match at each HLA loci will be prioritized.  A CB unit must be 
at least 4/6 HLA-matched with the patient.   
ABO/Rh Compatibility 
Recipients’ ABO/Rh blood typing will be obtained.  CB units will not be selected based 
on ABO typing.  However, an Rh negative CB unit will be selected for Rh negative 
female participants to avoid Rh sensitization in young females.   
CB Unit Characterization 
Results of initial testing at the cord blood bank must include a pre-cryopreservation 
TNCC, viability and sterility culture.  Pre-cryopreservation TNCC must be ≥12x107/kg 
(we will accept units that are ≥10 x 107/kg) to target administration of 10x107 cells/kg 
post thaw, sterility cultures must have been negative, total viability must have been 
≥85%, and CD34+ cell viability (if performed) must have been ≥70%.  
 
A test vial or segment must be available from each CB unit for potency testing and 
confirmatory HLA typing. The segment will be detached from the candidate unit and 
tested for potency and identity (HLA-confirmatory typing) per Standard Operating 
Procedures in the CCBB at Duke.  Units will be deemed acceptable for the trial if viability 
of the CD45 cell population is ≥40% and viability of the CD34 cell population is ≥70%.  
CFU growth, expression of aldehydehydrogenase and CD34 will be described but will 
not be a specification for study enrollment.   
 
Prior to the patients’ arrival, their designated CB unit will be transferred from the 
Carolinas Cord Blood Bank (or other accredited bank) to the Duke STCL, located in the 
same building, where it will be stored in a liquid nitrogen freezer until the day of infusion. 
On the infusion day, the CB will be thawed and washed in dextran/albumin and 
resuspended in an appropriate volume based on recipient weight for administration to 
the patient the standard fashion115 per SOP STCL-PROC-036.  At the time of thawing, 
standard studies listed (see table) will be performed.  Only TNCC is utilized for release.  
A maximum dose of 10x107 TNC/kg will be prepared for infusion in a syringe or bag and 
infused over 2-25 minutes.  

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Post-Thaw Cord Blood Unit Testing  
Test 
Specifications 
Total Nucleated Cell Count (TNCC) 
Report; used to calculate final dose 
Viability 
Report 
Viability of the CD34+ population* 
≥70% 
Viability of the CD45+ population 
≥40% 
Sterility** 
No Growth 
Colony Forming Unit (CFU) growth 
Report 
ALDHbr as a percentage of CD45+ cells 
Report 
*Viability of the CD34+ cells post-thaw was previously tested on a segment and required to meet 
the specification of ≥70%. Therefore, for the clinical product, we will report but not use the post-
thaw viability as a release criteria. 
**If a positive culture is obtained after product administration, a plan is put into effect to notify the 
clinical and study teams and treat the patient if indicated. 
 
6.2 
Human Umbilical Cord Tissue-derived Mesenchymal Stromal Cells (hCT-MSC) 
hCT-MSCs are a product of allogeneic cells manufactured from digested umbilical cord 
tissue that is expanded in culture, cryopreserved and banked (complete composition and 
manufacturing details available in IND 17313).  hCT-MSCs are manufactured in the 
Duke CT2 GMP cell manufacturing lab from umbilical cord tissue donated to the 
Carolinas Cord Blood Bank, an FDA-licensed, FACT-accredited, public cord blood bank 
at Duke University Medical Center, after written informed consent from the donor baby’s 
mother.  Cord tissue is harvested from the placentas of male babies delivered by 
elective C-section after a normal, full-term pregnancy.   Donor screening questionnaires 
are completed by the maternal donor, and maternal blood is tested for communicable 
diseases by the CLIA-certified donor screening laboratory in Charlotte, NC.  Donors 
must be eligible for donation to a public cord blood bank for allogeneic use.  After 
delivery of the placenta and cord, the cord blood is aseptically drained from the 
placenta.  Then the cord is dried and cleaned with chloropreps, separated from the base 
of the placenta, placed in a sterile bottle containing Plasmalyte A, and transported to the 
Robertson CT2 GMP cell processing laboratory at room temperature in a validated 
container.  
  
In the clean room manufacturing suite, in a biosafety cabinet, the cord tissue is removed 
from the media, placed in sterile dishes, cut into small pieces and then minced and 
digested in the Miltenyi Biotec GentleMacs Octo Dissociator with GMP-grade 
enzymes:  hyaluronidase, DNase, collagenase, papain.  The resultant cell suspension is 
placed in culture in Prime XV MSC Expansion XSFM (Irvine Scientific) media with 1% 
platelet lysate and grown to confluence (~7-14 days) to establish the P0 culture.  To 
establish the master cell bank, P0 is harvested and cryopreserved in cryovials with 
Cryostor 10 media (BioLife), and stored in the vapor phase of liquid nitrogen.  P1 and P2 
cultures are grown under similar conditions, in HYPERFlasks or HYPERStacks without 
platelet lysate, as needed to create the working cell bank and product for administration, 
respectively. Cells from P1 and P2 are removed from plastic cultureware using TrypLE 
(Gibco). The final product is derived from the P2 cultures which are harvested into 

AlloCB and hCT-MSC in Cerebral Palsy 
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plasmalyte with 5% human serum albumin, washed and cryopreserved in compartment 
cryobags containing 50-100 million cells in a final concentration of 10% DMSO with 
dextran (Akron Scientific).  On the day of administration, one compartment is thawed, 
diluted in 10-40 mLs of plasmalyte IV solution, placed in a syringe or bag and 
transported to the bedside for administration over 30-60 minutes.    
  
At each passage, the cell product is characterized by assessing cell surface phenotype 
by flow cytometry and functional assays via T-cell proliferation and organotypic models 
of microglial activation.  Each lot, prior to cryopreservation of P2, will also be tested for 
sterility, endotoxin and mycoplasma and these tests must meet specifications.  For 
dosing, release testing after thaw and dilution will include TNCC and viability via 
cellometer.  Patients will be dosed with 2x106 hCT-MSCs/kg based on the post thaw 
count. 
 
Process and Final Formulation 
hCT-MSC is manufactured from a single umbilical cord tissue in a series of three steps 
that generate a master cell bank, a working cell bank, and the study product.  The 
product for each step is cryopreserved in a controlled rate freezer and stored in the 
vapor phase of liquid nitrogen. At P2, a representative cryobag is thawed and qualified 
prior to the treatment of any patients with that lot of product. Testing for product release 
includes total nucleated cell count, viability, phenotype, functional assays, endotoxin, 
mycoplasma, gram stain and sterility. Each lot of cells is also tested for adventitial 
viruses prior to cryopreservation.   
 
On the day of treatment, cells are thawed per SOP STCLAOP-028 JA2 and then diluted 
in 10-40 mLs of plasmalyte A + 5% human serum albumin (HSA).  An aliquot is removed 
for cell count, viability, and sterility culture.  If the cells are ≥70% viable, the final product 
volume is adjusted to deliver 2x106 cells/kg to the study subject.  The cells are delivered 
to the bedside in a syringe containing plasmalyte A, 5% HSA, and residual DMSO.  Any 
removed cell suspension is inoculated into aerobic and anaerobic culture bottles for 
sterility testing. The cells have a four-hour expiry at room temperature post thaw. 
 
The hCT-MSC final product will be released conditionally for administration to the patient 
after testing a post thaw cell count and viability.  Final release will occur after the 14-day 
sterility culture period for the study product.  In the event that a sterility culture turns 
positive after administration of the product, the organism will be identified and antibiotic 
sensitivities performed.  The patient’s family will be contacted to determine if they are 
symptomatic (i.e. fever or other signs of infection).  Asymptomatic patients will be 
observed but will not be treated with antibiotics.  Symptomatic patients will be evaluated 
and treated accordingly, with blood cultures and antibiotics as appropriate.  All patients 
receiving a product with subsequent positive sterility test will be followed with daily 
contact by a study nurse for 14 days after the positive sterility test is noted. 
6.3 
Donor Screening for CB and hCT-MSC 
Donor screening and testing is performed per Carolinas Cord Blood Bank standard 
operating procedures to meet all requirements in 21CFR Part 1271.  The screening and 
testing is current with recommendations and is approved by the FDA under biological 
license number 1870. Maternal donors of umbilical cord blood are screened and tested 
for HIV-1, HIV-2, HIV-O, hepatitis B virus (HBV, surface antigen and core antibody), 
hepatitis C virus (HCV) antibody, Treponema pallidum (syphilis), Creutzfelds-Jakob 
Disease (CJD, screening only), Chagas Disease, human T-lymphotropic virus types 1 

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and 2 (HTLV-1, HTLV-2) and total antibodies against CMV. Nucleic acid testing for HIV-
1/2/O, HBV, West Nile Virus and HCV are also performed on maternal blood. Screening 
for Zika virus may also be performed. 
 
 
Because the cord tissue used for this study will be obtained from donors consented for 
cord blood donation to the Carolinas Cord Blood Bank, they will undergo donor 
screening and infectious disease testing per Carolinas Cord Blood Bank standard 
operating procedures.  The cord blood-associated maternal samples and cord tissue 
MSC samples will be retained as reference samples for future testing as part of this 
study. 
6.4 
Packaging of Study Products 
All cellular products receive a unique identification number (ISBT Demand 128 bar code) 
to ensure product integrity and maintain chain of custody. The clinical site or cord blood 
bank assigns an ISBT Demand 128 bar code label to the CB unit or hCT-MSC product, 
which is placed on the product bag/syringe directly or via tie tag. Products are 
transported from the STCL to the infusion site in a validated cooler by a trained courier. 
6.5 
Administration of Study Product 
Patients will be arrive in clinic on the morning of their scheduled infusion.  A peripheral 
IV will be placed either by an anesthesiologist, clinical staff or study staff and 
premedication with Benadryl 0.5mg/kg/dose IV and Solumedrol 0.5-1mg/kg IV will be 
administered. Allogeneic CB products will be administered intravenously over 5 to 25 
minutes under direct physician supervision. hCT-MSC products will be administered 
intravenously over 30-60 minutes under direct supervision.  Vital signs (heart rate, blood 
pressure, temperature, respiratory rate) will be checked upon arrival to the clinic and as 
clinically indicated.  Pulse oximetry will be monitored continuously throughout the 
infusion and for at least 5 minutes post infusion.  Patients will be hydrated with standard 
intravenous fluids as tolerated and observed for at least one hour post infusion.   
6.6 
Safety Follow-up 
On Day 1 following the infusion, the participant will be seen by study staff to assess for 
any infusion related adverse reactions or complications. At ~2 weeks post-infusion, a 
member of the study team will contact the parent or guardian via phone or email to 
assess patient status and any adverse events. In-person safety assessments will occur 
at each study visit.  In addition, a questionnaire (see Appendix 1) will be administered at 
6, 12, and 24 months post baseline visit to assess for serious adverse events.  
7.0 
STUDY PLAN 
7.1 
Overview  
Parents/Guardians who have previously contacted our program and have a child who 
may meet eligibility criteria for this study will be notified that this study is available. After 
initial contact, parents/guardians of potential research participants will have an initial 
phone interview with study personnel to describe the study, verify basic eligibility criteria, 
and confirm their interest in participation. The participant’s eligibility will then be 
screened through review of medical records, video, laboratory testing, and imaging 
under a separate screening protocol.  
Once all screening is complete and the patient is likely to meet study criteria, a suitable 

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unrelated donor CB unit will be identified at the Carolinas Cord Blood Bank, or other 
accredited cord blood bank. The CB unit will be screened as described in section 6.  
Participants will then travel to Duke for their first visit. On day 1, written informed consent 
will be obtained.  Patient eligibility will be confirmed by a physical observation and 
verification of cerebral palsy diagnosis and GMFCS level.  If no exclusion criteria are 
realized, the participant will be randomized to a treatment arm.  During their first visit, all 
participants will have physical therapy evaluations, and a subset of patients will undergo 
brain MRI.  Participants will have study infusions as determined by their assigned 
treatment arm (at baseline only for AlloCB; at 12-months only for Natural History; at 
baseline, 3-, and 6-months for MSCs).  Participants will be evaluated the day after each 
infusion, and parents will be contacted for phone or email follow-up ~2 weeks after each 
infusion. All participants will return to Duke six (motor assessments) and 12 months 
(motor assessments and brain MRI) after the baseline visit.  Participants on the MSC 
arm will also return at three months for an hCT-MSC infusion. A remote safety 
assessment will be performed via phone or email at 24 months post-infusion.  
7.2  
Patient Screening 
Initial patient screening will be conducted with informed consent under a separate 
protocol and will include a review of medical records, videos, and initial laboratory 
testing. If no exclusion criteria are identified, informed consent will be obtained, and the 
patient will be randomized to a treatment arm.  If indicated (AlloCB and Natural History 
arms), an unrelated donor CB unit will be identified at the Carolinas Cord Blood Bank, or 
other accredited cord blood bank.  Participants will travel to Duke for initial evaluation. 
Evaluations and treatments will be conducted in the outpatient setting. A physical exam 
and baseline GMFCS assessment will be conducted to confirm eligibility, and the 
participant undergo the remainder of the study evaluations.   
7.3 
CB Unit Selection 
For participants randomized to the AlloCB and Natural History arms, an allogeneic 
unrelated donor CB unit will be identified at the Carolinas Cord Blood Bank, or other 
accredited cord blood bank. HLA typing will be obtained on the patient, and the best 
available HLA-matched CB unit with a pre-cryopreservation nucleated cell dose ≥12x107 
cells/kg will be chosen.  We target CB units that are at least 12x107 cells/kg, however, 
we will accept units that are at least 10x107 cells/kg.  When possible, at least 1 match at 
each HLA loci will be prioritized.  An Rh negative CB unit will be selected for Rh 
negative female participants to avoid Rh sensitization in young females.   
Once a suitable allogeneic CB unit has been deemed an acceptable match, a sample of 
the CB unit will be tested for potency in the Duke STCL. If results of these tests are 
satisfactory, the CB unit will be delivered to the Duke STCL in the frozen state.  
7.4 
Study Product Infusion 
On the day of infusion, CB cells or hCT-MSC product will be prepared by the STCL and 
provided for infusion of the patient in the outpatient clinic under the supervision of the 
study team and Pediatric Blood and Marrow Transplant Program staff.  A peripheral IV 
will be placed by clinical staff, anesthesia or a member of the study team.  Prior to the 
study infusion, premedications (Benadryl and Solumedrol) will be administered.  CB cells 
will have a four-hour expiry at room temperature post-thaw. 
 
Allo CB infusion will be given over approximately 5-25 minutes and hCT-MSC infusions 
over 30-60 minutes using standard practices.  The child will receive 1-1.5x maintenance 

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IV fluids as described below and be observed in the clinic for a minimum of one hour 
after the infusion.  Patients will be discharged from clinic after at least one hour providing 
all vital signs are at their baseline and they are awake and asymptomatic with no 
evidence of toxicity.  Patients will be evaluated by study staff the day after the infusion to 
assess for any infusion-related adverse reactions or complications.  A phone call to 
parents/guardians by study staff to assess safety of the infusion will be conducted two 
weeks after the infusion. 
 
Maintenance IV Fluid Rate (Holliday-Segar Method from Harriet Lane Handbook) 
Body weight:  
mL/kg per day 
1st 10 kg 
 
100 
 
divided by 24hr/day 
2nd 10 kg 
 
 50  
 
divided by 24 hr/day 
each add'l kg  
 20 
 
divided by 24 hr/day 
 
If a patient has evidence of illness on the day of planned infusion, including but not 
limited to fever >38.5 C, vomiting, diarrhea, or respiratory distress, the infusion will be 
postponed. 
7.5 
Care During Unexpected Events 
In the event that a patient develops signs or symptoms of anaphylaxis including urticaria, 
difficulty breathing, cough, wheezing, or vomiting during their CB infusion, the infusion 
will be terminated and appropriate medical therapy initiated.  
7.6  
Motor Assessments 
Gross Motor Function Measurement-66 (GMFM-66):  The GMFM-66 is a standardized 
observational instrument designed and validated to measure change in gross motor 
function over time in children with cerebral palsy.  Developmental curves of expected 
progression have been published for children ages 2 – 12 years,64,116 allowing for the 
calculation of future expected scores based on the baseline age, GMFCS level, and 
GMFM-66 score.  The GMFM-66 consists of 66 items, divided into five categories: lying 
and rolling, sitting, crawling and kneeling, standing, and walking, running, and jumping. 
Each item is scored on a four-point Likert scale.  The GMFM-66 is a subset of the 
GMFM-88, which contains an additional 22 items, primarily in the lying and rolling 
category.  Both measures have been validated in children with cerebral palsy from 
5 months to 16 years of age.  A 5-year old child without motor disabilities is able to reach 
the maximum score.117  A computer program, the Gross Motor Ability Estimator, is used 
to calculate the GMFM-66 total scores.  The primary endpoint of this study is the 
difference between a child’s actual and expected changes in GMFM-66 score 12 months 
after the initial study infusion.  Control (placebo) and treated patients will be compared.  
When possible, the entire GMFM-88 will be performed, and subsets may be analyzed as 
exploratory endpoints. 
 
Peabody Developmental Motor Scales (PDMS-2):  The PDMS-II is a standardized 
assessment of early childhood motor development that evaluates both gross and fine 
motor skills. It is designed for children from birth through 5 years of age. The 
assessment is composed of six subtests that measure interrelated motor abilities that 
develop early in life (i.e., reflexes, stationary, locomotion, object manipulation, grasping, 
and visual-motor integration). Gross Motor Quotient, Fine Motor Quotient, and Total 
Motor Quotient composite scores are obtained.  For this study, the Gross Motor Quotient 
will be obtained and analyzed as a secondary endpoint. 

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7.7 
Functional Assessment 
Pediatric Evaluation of Disability Inventory-Computer Adaptive Test (PEDI-CAT):  The 
PEDI-CAT measures abilities in three functional domains:  Daily Activities, Mobility, and 
Social/Cognitive.  The computerized adaptive version is intended to provide an accurate 
and precise assessment of a child’s abilities while increasing efficiency and reducing 
respondent burden by utilizing item response theory statistical models to determent 
which items are assessed within each domain based on responses to prior items.     
7.8 
Imaging Assessments 
Participants’ brain imaging obtained previously as standard of care will be reviewed by a 
member of the Brain Imaging Analysis Center (BIAC) team to determine if accurate 
anatomical image parcellation would be likely on a brain MRI.  Those participants for 
whom usable data is likely to be obtained (estimated as approximately two-thirds of 
eligible participants) will undergo brain MRI with diffusion tensor imaging (DTI).  Diffusion 
weighted images will be acquired on a 3 Tesla GE scanner (Waukesha, WI). T1-
weighted images will be obtained with an inversion-prepared 3D fast spoiled-gradient-
recalled (FSPGR) pulse sequence.  These images will be analyzed to obtain measures 
of whole brain connectivity. 

AlloCB and hCT-MSC in Cerebral Palsy 
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7.9 
Required Evaluations 
 
Time Points# 
Screening 
Baseline 
 
3 months 
6 months 
12 months 
24 months 
All Participants 
 
 
 
 
 
 
CBCD*, CMP*, Type & Screen*, Patient HLA typing  
X 
 
 
 
 
 
Patient DNA sample for chimerism 
 
X 
 
 
 
 
CBCD*, CMP*, Type & Screen*, Direct Coombs, HLA 
Antibody Screen (PRA), Immune Reconstitution Panel, 
Humoral Immune Profile 
 
X 
 
 
X 
 
Motor Assessments (GMFM-66, PDMS-2) 
 
X 
 
X 
X 
 
Brain MRI^ 
 
X 
 
 
X 
 
Functional Assessment (PEDI-CAT) 
 
X 
 
X 
X 
 
Safety Assessment – remote  
 
 
 
 
 
X 
AlloCB Participants 
 
 
 
 
 
 
CBU potency; HLA confirmatory typing 
X 
 
 
 
 
 
History & Physical 
 
X 
 
X 
X 
 
Donor Referral Panel 
 
X 
 
 
 
 
CBU:  TNCC, viable  CD34+ count, CFU, sterility 
culture, donor DNA sample for chimerism 
 
X 
 
 
 
 
Safety Assessment – in-person, day after infusion 
 
X 
 
 
 
 
Natural History Participants 
 
 
 
 
 
 
CBU potency; HLA confirmatory typing 
X 
 
 
 
 
 
History & Physical 
 
X 
 
X 
X 
 
Donor Referral Panel 
 
 
 
 
X 
 
CBU:  TNCC, viable CD34+ count, CFU, sterility culture, 
donor DNA sample for chimerism 
 
 
 
 
X 
 
Safety Assessment – in-person, day after infusion 
 
 
 
 
X 
 
MSC Participants 
 
 
 
 
 
 
History & Physical 
 
X 
X 
X 
X 
 
Donor Referral Panel 
 
X 
 
 
 
 
CBCD, CMP 
 
 
X 
X 
 
 
Safety Assessment – in-person, day after infusion 
 
X 
X 
X 
 
 
*CBCD and CMP may be obtained at baseline or within 6 months prior to enrollment 
^Brain MRI will be performed on patients with eligible screening neuroimaging 
#Safety and return assessments should be performed within a month of the indicated time point. 
Abbreviations:  CBCD-Complete Blood Count and Differential, CMP-Complete Metabolic Panel, HLA-Human Leukocyte Antigen, GMFM-66-Gross Motor Function 
Measure-66, PDMS-2-Peabody Developmental Motor Scale-2, MRI-Magnetic Resonance Imaging, CBU-Cord Blood Unit, TNCC-Total Nucleated Cell Count, CFU-
Colony Forming Units  

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8.0 
STATISTICAL CONSIDERATIONS 
8.1 
Study Design 
This study is a phase I/II, prospective, randomized, open-label trial designed to provide 
interval estimates of the 12-month change in motor function after treatment with AlloCB 
and hCT-MSC, provide additional data to the clinical trials community on the natural 
history of the motor function in CP over short-term (less than 1 year) time periods 
relevant to conduct of clinical trials, and assess the safety of repeated doses of hCT-
MSC and a single dose of AlloCB in children with cerebral palsy. 
Children ages 2-5 years with cerebral palsy due to hypoxic ischemic encephalopathy, 
stroke, or periventricular leukomalacia will be eligible to participate.  All participants will 
ultimately be treated with an allogeneic cell product at some point during the study.  
Participants will be randomized (1:1:1) to one of three arms:  (1) the “AlloCB” arm will 
receive one allogeneic CB infusion at the baseline visit; (2) the “MSC” arm will receive 
three hCT-MSC infusions, one each at baseline, three months, and six months; the 
“natural history” arm will not receive an infusion at baseline but will receive an allogeneic 
CB infusion at 12 months. The occurrence of adverse events will be evaluated at 3, 6, 
12, and 24 months post-randomization in all participants. Motor function outcome 
measures will be assessed at baseline, six-months, and one-year time points in all 
participants.  Duration of study participation will be 24 months from the time of the 
baseline visit.  Randomization will be stratified by GMFCS Level (I/II or III/IV) and 
etiology of CP (Stroke vs. Other).  
8.2 
Accrual 
It is estimated that up to 8-12 research participants will be enrolled each month and that 
approximately 12-15 months of accrual will be necessary to enroll 90 evaluable 
participants.   
8.3 
Study Duration 
Each subject’s participation in the study will be 24 months, with clinic visits occurring 
during the first 12 months and a remote safety assessment at 24 months. Given that 
accrual will take up to 15 months it is estimated that the remote safety assessment will 
be conducted on that last patient 39 months (3.25 years) after the study opens. 
8.4 
Primary and Secondary Endpoints 
The primary endpoint of this study is the difference between a child’s observed and 
expected changes in GMFM-66 score 12 months after the initial study infusion.  This 
study will provide separate interval estimates of the mean of this outcome measure in 
patients assigned to the hC-MSC, AlloCB, and Natural History arms at 12-months. The 
secondary endpoint of this study is the number of adverse events occurring over a 12-
month period post-treatment with hCT-MSC or AlloCB.  
8.5 
Sample Size and Power Calculations 
The sample size for this study was selected to provide a high level of precision for 
estimating the mean of the observed minus expected 12-month change on the GMFM-
66 in each of the study arms, and to provide a high probability of detecting commonly 
occurring adverse events after infusion with AlloCB or hCT-MSC. 
 

AlloCB and hCT-MSC in Cerebral Palsy 
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As shown in the table below, a sample size of 30 patients per group provides a 95.8% 
probability of detecting common adverse events that occur in 10% of infusions (with 
hCT-MSC or AlloCB). This sample size also provides a 78.5% probability of observing 
events that occur in 5% of infusions, and a 26.0% probability of observing rare events 
that occur in 1% of infusions. 
 
Probability of Observing One or More Events with Various Sample Sizes* 
 
 
Probability (%)* 
N=20 N=30 N=40 N=50 
True Probability of an Event (%) 
 
 
 
 
1 
18.2 
26.0 
33.1 
39.5 
5 
64.2 
78.5 
87.1 
92.3 
10 
87.8 
95.8 
98.5 
99.5 
20 
98.8 
99.9 
100.0 
100.0 
50 
100.0 
100.0 
100.0 
100.0 
 
*Binomial probability of 1 or more independent events. 
 
The sample size for this study must also support estimation of the mean observed-
minus-expected GMFM-66 change score at 12 months post-intervention with MSC, 
AUCB, and in the Natural History arm. Thus, three interval estimates will be constructed 
using the t-distribution as follows. 
(𝑥̅ −𝑡𝛼2
⁄ ∗𝑠
√𝑛, 𝑥̅ + 𝑡𝛼2
⁄ ∗𝑠
√𝑛)  
 
The margin of error E is the confidence interval half-width:  
𝐸= 𝑡𝛼2
⁄ ∗𝑠
√𝑛 
 
The margin of error for this study was selected as 2 points with a confidence level of 
95%. The following formula was solved iteratively to obtain the sample size for each 
treatment group. 
 
𝑁= (
𝑡𝛼2
⁄ ∗𝑠
𝐸
)
2
 
 
The standard deviation, s, was estimated using 36 participants in the CP-AC trial who 
met age and GMFCS inclusion criteria for the present study: 5.16 (95% CI: 4.18, 6.13). 
 
Starting with a sample size of 20, and assuming a standard deviation of 5.16, a total of 3 
iterations were required to reach a final group sample size of 28 as shown in the table 
below. 
Iteration # Starting N Degrees of Freedom 𝒕𝜶𝟐
⁄  
Ending N 
1 
20 
19 
2.093 29 
2 
29 
28 
2.048 28 
3 
28 
27 
2.052 28 

AlloCB and hCT-MSC in Cerebral Palsy 
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Page 45 of 61 
 
Therefore, a group size of 28 patients allows for 95% confidence in the estimation of the 
mean 12-month observed-minus-expected GMFM-66 change score in one of the study 
arms (Natural History, MSC or AUCB) with a margin of error of no more than 2. This 
sample size is also concordant with what is required (N=30) for reasonable probability of 
detecting commonly occurring adverse events, as described above. Finally, if the 
standard deviation of the secondary outcome measure is as high as that indicated by the 
upper limit of the 95% confidence interval from the CP-AC study (6.13 points) then a 
sample of 126 patients allows for a margin of error no larger than ~2.5 points for each of 
the three interval estimates.   
 
The total sample size for this study is therefore set at 90 evaluable patients (30 per 
group). 
8.6  
Analysis Plan 
8.6.1 Analysis Populations 
The following populations are defined to support analyses of the primary and secondary 
endpoints. 
 
Intention to Treat Population 
This population will include all enrolled and randomized participants according to their 
assigned treatment. The primary endpoint will be evaluated in this population. 
 
Safety Population 
The safety population defines the patients in whom the secondary endpoint will be 
evaluated and will include all subjects who received at least 1 infusion. Analyses of the 
Safety Population will be conducted using an as-treated approach, which considers each 
patient according the treatment actually received rather than the treatment they were 
assigned. 
8.6.2 Timing of Analyses 
The analysis of the primary and secondary outcome measures will be conducted when 
the last patient reaches their 12-month visit. An update will be made to the safety 
analyses when the last patient reaches their 24-month visit. 
8.6.3 Demographics, Baseline Characteristics, and Disposition 
Demographics and baseline characteristics will be summarized for all research 
participants and separately by randomized assignment. Characteristics to be examined 
include age, sex, race/ethnicity, baseline GMFM-66 score, GMFCS level, and etiology of 
CP. The number of participants entering and completing the study will be diagrammed 
using the CONSORT guidelines. 
8.6.4 Analysis of the Primary and Secondary Endpoints 
The occurrence of adverse events in the Safety Population will be summarized 
descriptively in tables and figures for all subjects and separately by treatment received. 
Estimates of the mean observed-minus-expected GMFM-66 change score at 12 months 
will be reported in the Intention to Treat Population along with 95% confidence intervals 
as described above.  

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Page 46 of 61 
9.0 
SAFETY AND ADVERSE EVENT REPORTING 
9.1 
Definitions 
Adverse Event (AE):  An adverse event is any untoward medical occurrence associated 
with the use of the investigational product regardless of whether it is considered related 
to the investigational product.   
 
Serious Adverse Event (SAE):  An adverse event or suspected adverse reaction is 
considered “serious” if, in the view of either the investigator or sponsor, it results in any 
of the following outcomes:  death, a life threatening adverse event, inpatient 
hospitalization or prolongation of existing hospitalization, a persistent or significant 
incapacity or substantial disruption of the ability to conduct normal life functions, or a 
congenital anomaly/birth defect.  Important medical events that may not result in death, 
be life-threatening, or require hospitalization may be considered serious when, based 
upon appropriate medical judgment, they may jeopardize the patient or subject and may 
require medical or surgical intervention to prevent one of the outcomes listed in this 
definition. 
 
Grade/Severity:  Grade/severity will be assessed according to CTCAE v4.0 guidelines. 
 
Suspected Adverse Reaction:  A suspected adverse reaction is any adverse event for 
which there is a reasonable possibility that the investigational product caused the 
adverse event.  “Reasonable possibility” means there is evidence to suggest a causal 
relationship between the investigational product and the adverse event. 
 
Causality:  The investigator will use the following question when assessing causality of 
an adverse event to the investigational product:  “Is there a reasonable possibility that 
the investigational product caused the event?”  An affirmative answer designates the 
event as a suspected adverse reaction. 
9.2 
Adverse Event Reporting 
All adverse events reported or observed during the study beginning at the time of 
enrollment must be recorded.  Information to be reported includes when the site became 
aware of the event, investigator-specified assessment of severity and relationship to 
study therapy, whether there is an alternative etiology, seriousness, as well as any 
required treatment or evaluations, and outcome.  In general, investigators should report 
adverse events as diseases or syndromes whenever possible, instead of reporting 
individual component symptoms, signs, laboratory abnormalities, and sequelae. 
Severe adverse infusion reactions (fatal, life-threatening or requiring hospitalization) will 
be reported within seven calendar days of receipt of the information.   All fatal or life 
threatening SAEs will be reported by the investigator or its representatives to the FDA by 
telephone or fax within seven calendar days after receipt of the information, following 
FDA guidelines.  All serious and unexpected AEs will be reported to the FDA via a 
written report within 15 days of receipt of the information (21 CFR 312.32).  If the 
principal investigator assesses an event to be unrelated to the study, then the event will 
not require expedited reporting but will be included in the annual summary report. 
The following events within 24 hours of study product infusion will be also be recorded in 
the e-CRF: allergic reaction/hypersensitivity, sinus bradycardia, sinus tachycardia, 

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Page 47 of 61 
hypertension, hypotension, fever in the absence of neutropenia, rigors/chills, nausea, 
vomiting, infection with unknown ANC, dyspnea, hypoxia, and hemoglobinuria. 
9.3 
Serious Adverse Event Reporting 
The Principal Investigator or its representative will be responsible for telephone or fax 
reporting of any unexpected SAEs to the FDA.  The Principal Investigator or its 
representative will notify the FDA by telephone or fax of any fatal or life threatening 
experience (expedited report) associated with the use of the study therapy as soon as 
possible but no later than seven calendar days after receipt of the information.  Initial 
notification will be followed by a written report within 15 calendar days.  For SAEs  
associated with the use of the study therapy, the Principal Investigator will notify the FDA 
as soon as possible, but no later than 15 days, of the initial receipt of the information.  
The Principal Investigator or Sub-Investigator is responsible for informing the Institutional 
Review Board (IRB) and DSMB of any study related and unexpected SAEs.   
9.4 
Eliciting Adverse Event Information 
In addition to research participant observations, AEs will be documented from any data 
collected throughout the study including clinically significant laboratory values or physical 
exam findings. 
9.5 
Stopping Guidelines 
The following stopping guidelines will be monitored during the duration of the study.  The 
stopping guidelines will be monitored by the CRO, The EMMES Corporation, and are to 
be used to indicate boundaries requiring discussion by the investigators and DSMB.  
The study will be stopped for a safety review if: 
 
Any subject experiences a grade 4-5 infusion reactions within 24 hours of infusion; 
OR 
 
Two or more grade 4-5 adverse events determined to be temporally related by the 
medical safety monitor and/or the DSMB occur; 
OR 
 
Any subject experiences a blood stream infection within 6 months of infusion; 
OR 
 
Any death. 
9.6 
Subject Replacement 
 
The sponsor may replace any subject who has not been dosed.   
10.0 DATA SAFETY MONITORING BOARD (DSMB)  
A DSMB will be formed and a charter established.  Members of the DSMB will be 
independent of Duke University.  The DSMB will be notified immediately for all SAEs 
directly related to the study product throughout the study.  A total safety assessment will 
be prepared on an annual basis and forwarded to the DSMB for review as well.  Policies 
of the DSMB will be described in the DSMB charter and signed by all members. 
11.0 DATA HANDLING AND QUALITY ASSURANCE 
11.1 Case Report Forms 
As part of the responsibilities assumed by participating in the study, the Principal 
Investigator or Sub-Investigators agree to maintain adequate case histories of the 

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Page 48 of 61 
research participants treated as part of the research under this protocol.  The Principal 
Investigator or Sub-Investigator agrees to maintain accurate CRFs and source 
documentation as part of the case histories. 
EMMES will supply the CRF electronically (eCRF) through an electronic data entry 
system.  All eCRFs should be completed by the clinical site.  An authorized user at the 
clinical center completes the initial screening by entering in the research participant 
demographics and inclusion/exclusion criteria on the eligibility form.  Upon enrollment, a 
form submission schedule is generated for each participant, and displayed as a grid of 
forms by study visit, that permits direct access to each eCRF for data entry.  All CRF 
information is to be filled in.  If an item is not available or is not applicable, an exception 
can be requested through the data system.  Corrections to data fields can be made in 
the eCRF which maintains a data audit trail.   
11.2 Video Recordings 
Video recordings of potential subjects from parents and guardians may be submitted and 
used for determining study eligibility.  Video recordings may also be obtained of portions 
of the motor evaluations if indicated with parental consent, and may include full facial 
features.  The recordings will be used solely for analysis by the research team.  They will 
be stored electronically on a password-protected server.   
11.3 Inspection of Records 
The Principal Investigator or Sub-Investigators and institutions involved in the study will 
permit study-related monitoring, audits, IRB review, and regulatory inspection(s) by 
providing direct access to all study records.  In the event of an audit, the Principal 
Investigator or Sub-Investigator agrees to allow the Food and Drug Administration 
(FDA), or other regulatory agency access to all study records.  The Principal Investigator 
or Sub-Investigators should promptly notify all relevant parties of any audits scheduled 
by any regulatory authorities and promptly forward copies of any audit reports received 
to the both. 
11.4 Study Record Retention 
Study results will be retained in the patient’s research record for six years after the study 
is completed or until the patient reaches the age of 21, whichever is longer.  Essential 
documents should be retained until at least two years after the last approval of a 
marketing application in an International Conference on Harmonisation (ICH) region and 
until there are no pending or contemplated marketing applications in an ICH region or at 
least two years have elapsed since the formal discontinuation of clinical development of 
the investigational product.  These documents should be retained for a longer period, 
however, if required by the applicable regulatory requirements.  
12.0 ADMINISTRATIVE ASPECTS 
The following administrative items are meant to guide the Principal Investigator or Sub-
Investigator in the conduct of the study but may be subject to change based on industry 
and government Standard Operating Procedures or Working Practice Documents or 
Guidelines.   
12.1 Confidentiality 
All laboratory specimens, evaluation forms, reports, and other records will be identified in 
a manner designed to maintain research participant confidentiality.  All records will be 

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Page 49 of 61 
kept in a secure storage area with limited access.  Clinical information will not be 
released without the written permission of the research participant’s guardian except as 
necessary for monitoring and auditing. 
The Principal Investigator or Sub-Investigator and all employees and coworkers involved 
with this study may not disclose or use for any purpose other than performance of the 
study, any data, record, or other unpublished, confidential information disclosed to those 
individuals for the purpose of the study.   
12.2 Institutional Review Board Approval 
Federal regulations and the ICH guidelines require that approval be obtained from an 
IRB prior to participation of human research participants in research studies.  Prior to the 
study onset, the protocol, informed consent, any advertisement used to recruit study 
patients, and any other written information regarding this study to be provided to the 
research participant or the research participant’s legal guardian must be approved by the 
IRB.   
All IRB approvals should be signed by the IRB Chairman or designee and must identify 
the IRB name and address, the clinical protocol by title and/or protocol number, and the 
date the approval and/or favorable opinion was granted. 
The Principal Investigator or Sub-Investigator is responsible for obtaining continued 
review of the clinical research at intervals not exceeding one year or otherwise specified 
by the IRB.  The Principal Investigator or Sub-Investigator must supply the Sponsor or its 
designee with written documentation of continued review of the clinical research. 
12.3 Modification of the Protocol 
Any changes in this research activity, except those necessary to remove an apparent, 
immediate hazard to the research participant, must be reviewed and approved by the 
IRB.   
12.4 Informed Consent 
A written informed consent in compliance with Part 50 of Title 21 of the Code of Federal 
Regulations (CFR) and Institutional IRB shall be obtained from each research participant 
prior to entering the study or performing any unusual or non-routine procedure that 
involves risk to the research participant.   
Before enrollment, each prospective research participant and/or his/her legal guardian 
will be given a full explanation of the study and be allowed to read the approved 
informed consent form.  Once the Principal Investigator or Sub-Investigator is assured 
that the research participant/legal guardian understands the implications of participating 
in the study, the research participant/legal guardian will be asked to give consent to 
participate in the study by signing the informed consent form. 
The Principal Investigator or Sub-Investigator shall provide a signed/dated copy of the 
signed informed consent to the research participant and/or legal guardian.   
12.5 Protocol Violations and Deviations 
The Principal Investigator or Sub-Investigator or designee must document and explain in 
the research participant’s source documentation any deviation from the approved 
protocol.  The Principal Investigator or Sub-Investigator may implement a deviation from, 

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Page 50 of 61 
or a change of, the protocol to eliminate an immediate hazard to study research 
participants without prior IRB approval.  As soon as possible after such an occurrence, 
the implemented deviation or change, the reasons for it, and any proposed protocol 
amendment(s) should be submitted to the IRB for review and approval, to the Sponsor 
for agreement, and to the regulatory authorities, if required. 
A deviation from the protocol is an unintended and/or unanticipated departure from the 
procedures and/or processes approved by the Sponsor and the IRB and agreed to by 
the Principal Investigator or Sub-Investigator.  Deviations usually have an impact on 
individual research participants or a small group of research participants and do not 
involve inclusion/exclusion or primary endpoint criteria.  A protocol violation occurs when 
there is nonadherence to the protocol that results in a significant, additional risk to the 
research participant, when the research participant or Principal Investigator or Sub-
Investigator has failed to adhere to significant protocol requirements (inclusion/exclusion 
criteria) and the research participant was enrolled without prior Sponsor approval, or 
when there is nonadherence to FDA regulations and/or ICH GCP guidelines. 
The IRB should be notified of all protocol violations and deviations in a timely manner as 
required by the site’s IRB. 
12.6 Study Reporting Requirements 
By participating in the study, the Principal Investigator or Sub-Investigator agrees to 
submit reports of serious adverse events according to the timeline and method outlined 
in the protocol.  In addition, the Principal Investigator or Sub-Investigator agrees to 
submit annual reports to his/her IRB as appropriate.  The Principal Investigator or Sub-
Investigator also agrees to provide the Sponsor with an adequate report shortly after 
completion of the Principal Investigator’s or Sub-Investigator’s participation in the study. 
12.7 Study Conduct 
The Principal Investigator agrees that the study will be conducted according to the 
principles of the ICH E6 Guideline on GCP and the principles of the World Medical 
Association Declaration of Helsinki.  The Principal Investigator will conduct all aspects of 
this study in accordance with all national, state, and local laws or regulations. 
12.8 Publications 
Following completion of the study, the data may be considered for reporting at a 
scientific meeting or for publication in a scientific journal.  In these cases, Duke 
University will be responsible to determine how the manuscript is written and edited, the 
number and order of authors, the publication to which it will be submitted, and other 
related issues. 
 
 

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14.0 APPENDICIES 
 
 
Appendix 1:  Safety Questionnaire  
 
 
 
 
 
 
 
 
 

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SAFETY QUESTIONNAIRE 
Patient Name:  __________________ 
Date completed:  __ __ /__ __/__ __ __ __ 
                             m   m      d     d     y     y     y     y 
 
Please answer the following items regarding your child to the best of your ability.  Some of the questions 
may be the same as the questions you answered previously.   
 
Since your child’s last study visit, did any of the following occur? 
1.  Was your child hospitalized? 
 Yes; if so, why? _________________________________________________ 
 No 
2.  Did your child develop a tumor? 
 Yes 
 No 
3.  Did your child develop abnormal skin lesions? 
 Yes 
 No 
4.  Was your child diagnosed with cancer? 
 Yes 
 No 
5.  Did your child have a serious infection? 
 Yes 
 No 
6.  Was your child diagnosed with an autoimmune disease? 
 Yes 
 No 
7.  Did your child require a blood transfusion? 
 Yes 
 No 
8.  Has your child developed any other health problems not described above? 
 Yes; if so, what? _________________________________________________ 
 No 
9.  Has your child developed any new onset seizures? 
 Yes 
 No 
10. Has your child started any new medications? 

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 Yes; if so, which ones? _____________________________________________ 
 No 
11. Has your child’s diet changed (i.e. started/stopped a special type of diet)? 
 Yes; if so, how? _____________________________________________ 
 No 
12. Have you noticed an increase in any of the following in your child since their last visit?  Select 
all that apply. 
 Appetite (increased or decreased) 
 Yes       No 
 Fatigue 
 Yes       No 
 Headache 
 Yes       No 
 Restlessness 
 Yes       No 
 Tremor 
 Yes       No 
13. Has your child participated in any of the following:  
 Other cell therapy 
 Yes       No  
 Immune therapy (i.e. IVIG, steroids) 
 Yes       No 
 Other clinical trials 
 Yes       No 
 If yes, please describe:  ___________________________________________ 
 
 
 
 
Please describe any changes you have seen in your child since receiving their first infusion: 
 
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________
_____________________________________________________________________________________ 

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Please rate your child’s development in the following domains since receiving their first infusion 
(Circle the number that corresponds best.): 
 
1 
2 
3 
4 
5 
Slowed  
down 
Stayed 
about the 
same 
Improved 
a little 
more than 
expected 
Improved 
more than 
expected 
Improved 
much 
more than 
expected 
 
 
1.  Fine Motor (use of hands/fingers for activities like holding toys, playing, drawing, eating, etc) 
 
1 
2 
3 
4 
5 
 
 
2.  Gross Motor (use of big muscles for activities like head control, rolling, crawling, walking, 
running or climbing) 
 
1 
2 
3 
4 
5 
 
 
3.  Receptive Language (language listening and understanding, following spoken requests, etc.)  
 
1 
2 
3 
4 
5 
 
 
4.  Expressive Language (speaking skills: use of signs, words, sentences, etc.)  
 
1 
2 
3 
4 
5 
 
 
5.  Cognitive Skills (thinking, learning, reasoning, and problem solving)  
 
1 
2 
3 
4 
5 
 
 
6.  Adaptive and Self Help Skills (feeding, dressing, toileting, etc.)  
 
1 
2 
3 
4 
5 
 
  
7.  Social Skills and Behavior (interpersonal skills, peer skills, emotions, behaviors)  
 
1 
2 
3 
4 
5