Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne Muscular Dystrophy
Phase: PHASE1
Conditions: Duchenne Muscular Dystrophy
Sponsor: Megan Waldrop

 
Protocol version 4.0 (18Nov2021) 
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Phase I/IIa Systemic Gene Delivery Clinical Trial of 
scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne 
muscular dystrophy 
CLINICAL PROTOCOL 
Version:  V4.0 (18Nov2021) 
Sponsor-Investigator:  Megan Waldrop, M.D. 
Co-Investigators: Kevin M. Flanigan, M.D. 
Kim McBride, M.D. 
Nicolas Wein, Ph.D. 
Ramkumar Krishnamurthy, Ph.D. 
Linda P. Lowes, Ph.D. 
Maryann Kaler, FNP 
 
 
 
 
 
 
 
 
The Research Institute at Nationwide Children’s Hospital                                
Center for Gene Therapy 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
Protocol version 4.0 (18Nov2021) 
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SIGNATURE PAGE 
 
Study Title: 
Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA 
for Exon 2 Duplication-Associated Duchenne muscular dystrophy  
 
Study Sponsor 
Megan A. Waldrop, MD 
 
Financial Sponsor 
Audentes Therapeutics 
 
Protocol Version 
v4.0 18Nov2021 
 
As Sponsor-Investigator, I agree to personally conduct this study in compliance with Principles 
of Good Clinical Practice as defined by federal, state, and local laws and regulations. I will abide 
by the current protocol (provided herein), including any protocol amendments that are approved 
by our institutional review board (IRB). 
 
 
___________________________ 
Megan A. Waldrop, MD 
Sponsor-Investigator 
Nationwide Children’s Hospital 
  
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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TABLE OF CONTENTS 
1 
Protocol Synopsis.................................................................................................................... 7 
2 
Administrative Information .................................................................................................. 10 
2.1 
Document History .......................................................................................................... 10 
2.2 
Summary of Changes ..................................................................................................... 10 
2.3 
Contact Information ....................................................................................................... 12 
3 
List of Abbreviations and definition of terms ....................................................................... 15 
4 
Introduction: Key Roles, Background, and Scientific Rationale .......................................... 17 
4.1 
Clinical Trial, Sponsor Investigator and Co-Investigators ............................................. 17 
4.2 
Abstract .......................................................................................................................... 18 
4.3 
Background and Significance......................................................................................... 18 
4.3.1 
Disease Characteristics ........................................................................................... 18 
4.3.2 
Disease Pathogenesis .............................................................................................. 19 
4.3.3 
Current Treatments for DMD ................................................................................. 19 
4.4 
Rationale......................................................................................................................... 19 
4.4.1 
Rationale for Gene Therapy using U7snRNA exon skipping ................................. 19 
4.4.2 
Rationale for therapeutic skipping of exon 2 .......................................................... 20 
5 
Objectives and Purpose ......................................................................................................... 21 
5.1 
Primary Outcome ........................................................................................................... 21 
5.2 
Secondary Outcome ....................................................................................................... 21 
5.3 
Exploratory Outcomes.................................................................................................... 22 
6 
Study Design and Endpoints ................................................................................................. 23 
6.1 
Overall Study Design ..................................................................................................... 23 
6.2 
Study Population ............................................................................................................ 23 
6.3 
Number of Subjects ........................................................................................................ 23 
6.4 
Dosing plan .................................................................................................................... 23 
6.5 
Criteria for Study Termination ....................................................................................... 24 
6.6 
Establish Subject Identification Number........................................................................ 24 
7 
Study Enrollment and Withdrawal........................................................................................ 24 
7.1 
Inclusion/Exclusion ........................................................................................................ 24 
7.2 
Subject Withdrawal Criteria ........................................................................................... 26 
8 
Study Agent .......................................................................................................................... 26 
8.1 
Description of the Study Drug ....................................................................................... 27 
9 
Study Procedures and Schedule ............................................................................................ 27 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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9.1 
Study Visits .................................................................................................................... 27 
9.1.1 
Screening (day -45 to day -1).................................................................................. 27 
9.1.2 
Gene Transfer Visit ................................................................................................. 30 
9.1.3 
Post Gene Transfer Monitoring Plan ...................................................................... 33 
9.2 
Schedule of Events ......................................................................................................... 40 
9.3 
Muscle Strength Test Schedule ...................................................................................... 42 
9.4 
Long Term Follow up .................................................................................................... 42 
10 Procedure Criteria ................................................................................................................. 43 
10.1 
Muscle biopsy ............................................................................................................. 43 
10.2 
Magnetic Resonance Imaging (MRI) ......................................................................... 43 
10.3 
Pulmonary functional tests (PFTs) ............................................................................. 43 
10.4 
Vector Shedding Studies ............................................................................................ 43 
10.5 
Height Measurements ................................................................................................. 43 
10.6 
Concomitant Medications ........................................................................................... 43 
10.6.1 Prophylactic Administration of Prednisone/Prednisolone ...................................... 44 
11 Assessment of Safety ............................................................................................................ 44 
11.1 
Adverse and Serious Adverse Event .......................................................................... 44 
11.1.1 Definition of an Adverse Event .............................................................................. 44 
11.1.2 Unexpected Adverse Events ................................................................................... 45 
11.1.3 Serious adverse event (SAE) .................................................................................. 46 
11.1.4 Life-threatening (21 CFR 312.32(a)) ...................................................................... 46 
11.2 
Relationship to Study Drug ........................................................................................ 46 
11.3 
Time Period and Frequency for Event Assessment and Follow-Up .......................... 47 
11.3.1 Obligations of the Investigator ................................................................................ 47 
11.3.2 Safety Reporting ..................................................................................................... 47 
11.3.3 Safety Reporting: Content and Format ................................................................... 48 
11.3.4 Adverse Event Reporting from Primary Care Physician ........................................ 48 
11.3.5 Follow-up of Adverse Events ................................................................................. 49 
11.4 
Dose limiting toxicity (DLT) ...................................................................................... 49 
11.4.1 Expected Adverse Events Related to Disease Progression ..................................... 50 
11.4.2 Anticipated Adverse Event Lab Findings related to Intervention .......................... 50 
11.5 
Discontinuation Rules................................................................................................. 51 
12 Clinical Monitoring ............................................................................................................... 51 
12.1 
Data and Safety Monitoring Plan ............................................................................... 51 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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12.1.1 DSMB Membership ................................................................................................ 51 
12.1.2 DSMB Responsibilities ........................................................................................... 51 
12.1.3 DSMB Reporting and Meetings.............................................................................. 52 
12.2 
Institutional Review Board Monitoring ...................................................................... 53 
13 Statistical Considerations ...................................................................................................... 53 
14 Source Documents and Access to Source Data/Documents ................................................. 53 
14.1 
Data Management and Study Forms .......................................................................... 53 
14.2 
Electronic Case Report Forms .................................................................................... 54 
14.3 
Inspection of Records ................................................................................................. 54 
15 Quality Assurance and Quality Control ................................................................................ 54 
15.1 
Study Monitoring ........................................................................................................ 54 
16 Ethics/Protection of Human Subjects ................................................................................... 55 
16.1 
Ethics Review ............................................................................................................. 55 
16.2 
Ethical Conduct of the Study ...................................................................................... 55 
16.3 
Written Informed Consent .......................................................................................... 55 
17 Data Handling and Record Keeping ..................................................................................... 56 
17.1 
Retention of Records .................................................................................................. 56 
17.2 
Retention of Samples .................................................................................................. 56 
17.3 
Study Reports ............................................................................................................. 57 
17.3.1 Annual Study Reports ............................................................................................. 57 
18 References ............................................................................................................................. 58 
 
 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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List of Tables 
Table 1: Document Record ......................................................................................................... 10 
Table 2: Revision Record ............................................................................................................ 10 
Table 3: Study Team.................................................................................................................... 12 
Table 4: Data Safety Monitoring Board .................................................................................... 13 
Table 5: Study Vendor Listing ................................................................................................... 15 
Table 6: Abbreviations and Specialist Terms ........................................................................... 15 
Table 7: Normal and abnormal laboratory values at Nationwide Children’s Hospital 
Laboratory ................................................................................................................................... 25 
Table 8: Schedule of Events ........................................................................................................ 40 
Table 9: Muscle Strength Testing Schedule .............................................................................. 42 
Table 10: Adverse Event Classification ..................................................................................... 49 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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1 PROTOCOL SYNOPSIS 
Name of Sponsor 
Megan A. Waldrop, M.D. 
Name of 
Investigational 
Product 
scAAV9.U7.ACCA 
Study Title 
Phase I/IIa Systemic Gene Delivery Clinical Trial of 
scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne 
muscular dystrophy 
Study Number 
N = 1-3, 3.0 x 1013 vg/kg 
Sponsor Investigator 
Megan A. Waldrop, M.D. 
Clinical Study Phase 
Phase I/IIa 
Number of Centers 
Single site (Nationwide Children’s Hospital) 
Study Duration 
We will evaluate short-term safety over a two-year period.  
 
Subjects will be evaluated at screening, during gene transfer, and for 
return follow up visits on days 7, 14, 30, 60, 90, 180 and months 12, 
18 and 24. Additional immune studies will continue with blood 
samples collected locally and shipped to NCH at days 45, 75, 120, 
150, and two weeks following steroid dose reduction. 
Study Design 
Open-label study of scAAV9.U7.ACCA 
Subject Population 
Inclusion Criteria: 
1. Age greater than 6 months and less than 14 years 
2. Confirmed duplication of exon 2 in the DMD gene using a 
clinically accepted technique that completely defines the 
mutation 
3. Pre-ambulant (not yet walking) or ambulant (as defined by 
the ability to walk 10 meters without assistance) 
4. Males of any ethnic group will be eligible 
5. Ability to cooperate with muscle testing 
6. In subjects age 4 and above, stable dose and regimen of 
corticosteroid therapy (prednisone, deflazacort, or their 
generic forms) for at least 12 weeks prior to gene transfer. 
 
Exclusion Criteria: 
1. Active viral infection based on clinical observations 
2. Symptoms or signs of cardiomyopathy, including: 
a. Dyspnea on exertion, pedal edema, shortness of 
breath upon lying flat, or rales at the base of the lungs 
b. Echocardiogram with ejection fraction below 40% 
3. Serological evidence of HIV infection, or Hepatitis B or C 
infection 
4. Diagnosis of (or ongoing treatment for) an autoimmune 
disease 
5. Persistent leukopenia or leukocytosis (WBC ≤ 3.5 K/µL or ≥ 
20.0 K/µL) or an absolute neutrophil count < 1.5K/µL 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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6. Concomitant illness or requirement for chronic drug 
treatment that in the opinion of the SI creates unnecessary 
risks for gene transfer 
7. AAV9 binding antibody titers ≥ 1:400 as determined by 
ELISA immunoassay 
8. Abnormal laboratory values in the clinically significant range 
as listed in Table 7, based upon normal values in the 
Nationwide Children’s Hospital Laboratory. 
Study Procedures 
The vector will be delivered via peripheral limb vein. Study subjects 
will receive sedation if deemed necessary for the procedure following 
Nationwide Children’s Hospital dosing protocol.  
Primary Outcome 
Safety is the primary outcome for this clinical gene transfer trial. 
Secondary Outcomes We propose the following secondary efficacy outcome measures in 
leg muscles, each in comparison to baseline values: 
• Expression of dystrophin as demonstrated by 
immunofluorescent staining with a dystrophin C-terminus 
antibody and the exon-1 encoded epitope antibody (Manex 
1A) in muscle biopsy sections. 
• Dystrophin protein expression quantified by western blot 
using a dystrophin C-terminus antibody and the exon-1 
encoded epitope antibody (Manex 1A) and assessed by 
densitometry in muscle biopsy tissue. 
• Percentage of exon 2 exclusion from the dystrophin mRNA 
transcript as measured by RT-PCR analysis. 
• Transduction efficiency will be measured by qPCR of a U7-
targeting sequence primer pair amplifying the DNA 
containing the U7snRNA, and expressed as vector genomes 
normalized to a single-copy genomic DNA control in each 
muscle sample. 
Exploratory 
Outcomes 
We propose the following exploratory clinical outcome measures, 
each in comparison to baseline values:  
1) For Muscle Strength: 
• In subjects enrolling at 36 months of age or younger: 
improvement or stabilization of Bayley-IV scores (Bayley 
Scales of Infant and Toddler Development) 
• In subjects enrolling at greater than 36 months of age but 
younger than 5 years: improvement or stabilization of the 
North Star Ambulatory Assessment (NSAA) score, time to 
climb four stairs and amount of time to walk 100 meters 
(100MWT) 
• In subjects enrolling at age 5 years or greater: improvement or 
statistically significant stabilization in: 
o North Star Ambulatory Assessment (NSAA) score 
o Amount of time required to walk 100 meters (100MWT) 
o Time to climb four stairs 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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o Force generated in knee flexion and extension in 
maximum voluntary isometric contraction (MVICT).   
 
2) As an exploratory efficacy measure, muscle MRI will be assessed 
using T2-weighted multi-slice spin echo (SE) axial images and single 
voxel 1H-MRS from both the lower leg and thigh to measure lipid 
fraction and (1)H2O T2 signal.  These will be performed at baseline 
and at 6, 12, 18 and 24 months post infusion and data will be assessed 
using published techniques1 and compared to baseline in 
collaboration with Dr. Glenn Walter, University of Florida. 
 
*An MRI will not be performed if sedation is required. See Section 
10.2 for more details.  
 
3) Percentage of exon 2 exclusion from the dystrophin mRNA 
transcript as measured by RNA-Seq analysis. 
 
Sample Size 
DMD subjects will receive scAAV9.U7.ACCA at the minimal 
efficacious dose (MED). 
N = 1-3, 3.0 x 1013 vg/kg 
Statistical Analysis 
For the secondary outcome measure of dystrophin expression, 
quantification of the number of fibers expressing dystrophin 
(determined by quantification of the number of fibers), measurement 
of the intensity of dystrophin staining at the sarcolemma (based on 
quantitative image analysis), and the degree of exon 2 skipping (as 
determined by RT-PCR) will be performed for each patient at each 
time point in a blinded fashion, and intra-patient comparisons of pre- 
and post-treatment values will be analyzed using repeated measures 
ANOVA with a Dunnett correction for multiple comparisons vs. 
baseline. For detecting change, we will have 80% power to detect a 
very large effect size (SMD=1.4-3.3, for sample sizes from 3-6). 
Long-term follow-up 
Safety follow-up will continue over a two-year period that 
incorporates the active phase of the protocol. Subjects will then 
transfer to an annual monitoring program where data will be collected 
from annual standard of care visits for up to 5 years. 
 
 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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2 
ADMINISTRATIVE INFORMATION 
2.1 
Document History 
Table 1: Document Record 
Document 
Version and Date 
Submission Purpose 
U7.ACCA Clinical Protocol 
DRAFT - v0.7 15Jan2019 
FDA Initial Submission S/N 0000 
U7.ACCA Clinical Protocol 
v1.0 25Oct2019 
Response to FDA hold S/N 0001 
U7.ACCA Clinical Protocol 
v1.0 13Dec2019 
Changes requested by the DSMB, 
sent to FDA as S/N 0002 
U7.ACCA Clinical Protocol 
V2.0 20Jan2020 
Protocol and Consent Changes 
U7.ACCA Clinical Protocol  
V3.0 25Mar2020 
Protocol and Consent changes 
requested by the DSMB 
U7.ACCA Clinical Protocol 
V4.0 18Nov2021 
Protocol and Consent Changes 
 
2.2 
Summary of Changes 
The section below highlights content changes in this version of the protocol. Language deleted 
from the previous version of the protocol appears in red/strikethrough. Language added to the 
previous version of the protocol appears in bold. 
Table 2: Revision Record 
Protocol Version 
Description of Changes 
V1.0 25Oct2019 
• Removed control group and dose escalation 
from trial. 
• Added RT-PCR measurement of exon 2 
exclusion from dystrophin mRNA transcript 
to secondary outcomes. 
• Moved RNASeq analysis of exon 2 exclusion 
from dystrophin mRNA transcript to exploratory 
outcomes. 
• Changed inclusion criteria to greater than 6 
months and less than 14 years.   
• Removed Western Blot analysis at days 90 and 
180 as well as 12 months for dystrophin 
associated protein complex. 
• Removed Leukocyte marker analysis at days 90, 
180, and 12 months.  
• Added language to indicate report of non-life 
threatening SAEs and DLTs to the IRB 
within 15 calendar days. 
• Added possibility of moving blood tests to 
other days of the visit if the child is under 
10kg and may have too much blood drawn. 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
Protocol version 4.0 (18Nov2021) 
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• Added language to clarify the expected 
elevations in AST/ALT levels for the DMD 
population. 
• Clarified that Dose-Limiting Toxicity events 
must be unexpected to be considered as such. 
• Added Hepatology to the fields of expertise of 
the DSMB members.  
• Added language to allow for weekly blood 
tests past 30 days post-infusion if the 
subject’s AST/ALT levels have not returned 
to baseline and the prednisone/prednisolone 
dose is maintained rather than decreased 
back to the standard clinical dose. 
• Removed specification of the antibody used for 
immunofluorescence staining and western blot. 
• Updated CTCAE referenced to v5.0. 
• Changed Megan Waldrop from Principal 
Investigator to Sponsor Investigator. 
• Changed the N from N=3 to N=1-3. 
V1.0 13Dec2019 
• Added labs for Total IgG, D-Dimer, Direct 
Bilirubin, SC5b-9, C3, and C4 at screening 
• Added Total IgG at Day 30, 60, 90, 120, 150, 
180, Month 12, Month 18, Month 24 
• Added Direct Bilirubin to Day -1, 1, 7, 14, 30, 
45, 60, 75, 90, 2 weeks post steroid, 120, 150, 
180, Month 12, Month 18, Month 24 
• Removed redundancies in the Safety Reporting 
section. 
• Added Nicolas Abreu, Linda Lowes, 
Ramkumar Krishnamurthy, and Cassandra 
Karingada as Co-Investigators. 
V2.0 20Jan2020 
• Clarified Physical therapy testing schedule.  
Added climb 4 stairs test and 100MWT to 3 
to 5 year olds.  Added PFT testing to begin at 
age 5 years. 
• Removed PT functional testing assessment age 
related change 
• Clarified timing of screening blood work 
• Added language for addition and 
discontinuation of proton pump inhibitor 
(PPI) with increased prednisolone dosage. 
• Clarified timing of vital sign monitoring 
during gene transfer with and without 
sedation. 
• Added Maryann Kaler as Co-Investigator. 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
Protocol version 4.0 (18Nov2021) 
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V3.0 25Mar2020 
• Added CH50 lab test to Screening labs 
• Added C3, C4, Ch50, and Sc5b-9 to Day 7 
labs 
• Changed volume of BD Luer-Lok used for 
investigational product for gene transfer to 
50mL from 60mL (60mL no longer available) 
• Changed vitals monitoring post gene transfer to 
every 15 minutes for 4 hours post and then 
hourly until transfer to the Neurology floor, 
then every four hours until discharge.  
V4.0 18Nov2021 
• Removed Cassandra Karingada, Nicolas Abreu, 
and Johan Harris. 
• Added Sharmada Subramanian and Kristen 
Brown. 
• Updated IQVIA monitor information. 
• Changed exploratory functional outcome for 
subjects under 36 months of age from Bayley III 
to Bayley-IV. 
• Added visit windows for follow up visits. 
• Clarified how the total vector genome dose 
will be rounded based on subject weight. 
2.3 
Contact Information 
Table 3: Study Team 
Role in Study Name 
Address and Telephone Number 
Sponsor 
Investigator 
Megan Waldrop, MD 
WA3013 700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-722-2231  
megan.waldrop@nationwidechildrens.org 
Co-Investigator 
Kevin Flanigan, MD 
WA3011 700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-722-2231  
Kevin.flanigan@nationwidechildrens.org 
Co-Investigator 
Kim McBride, MD 
700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-355-5734 
Kim.mcbride@nationwidechildrens.org 
Co-Investigator 
Ramkumar 
Krishnamurthy, PhD 
700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-722-2611 
Ramkumar.Krishnamurthy@nationwidechildrens.org 
Co-Investigator 
Nicolas Wein, PhD 
WA3020 700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-722-2678 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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Nicolas.wein@nationwidechildrens.org 
Co-Investigator 
Linda P. Lowes, Ph.D. 
700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-722-2849 
Linda.lowes@nationwidechildrens.org 
Co-Investigator 
Maryann Kaler, FNP 
WA2022 700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-355-3572 
Maryann.kaler@nationwidechildrens.org 
Program 
Manager 
Tabatha Simmons, PhD WA2015 700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-722-6921 
Tabatha.simmons@nationwidechildrens.org 
Clinical 
Research 
Coordinator 
Sharmada Subramanian 700 Children’s Drive 
Columbus, OH 43205 
Ph: 614-722-2231 
Sharmada.Subramanian@nationwidechildrens.org  
Clinical 
Research 
Coordinator 
Team Lead 
Kristen Brown  
WA2014 700 Children’s Dr 
Columbus, OH 43205 
Ph: 614-722-6918 
 Kristen.Brown@nationwidechildrens.org  
Regulatory 
Specialist 
Rachel Manthe-Gross, 
PhD 
W178 700 Children’s Dr 
Columbus, OH 43204 
Ph: 614-355-2681 
Rachel.manthe-gross@nationwidechildrens.org 
 
 
Table 4: Data Safety Monitoring Board 
Name 
Address and Telephone Number 
Edward C. Smith, M.D. 
Duke University Medical Center 
3000 Erwin Road 
Durham, NC 27705 
Ph: 919-668-0477 
Edward.smith@duke.edu 
Barry J. Byrne, M.D., Ph.D. 
PO Box 100296 
Gainesville, FL 32610-0296 
Ph: 352-294-8968 
Barry.byrne@ufl.edu 
Arash Grakoui, Ph.D. 
Emory University 
Room 3020  
954 Gatewood Road NE 
Atlanta, GA 30322 
Ph: 404-727-7822 
Arash.grakoui@emory.edu 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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Robert Squires, M.D. 
UPMC Children’s Hospital Pittsburgh 
4401 Penn Avenue 
Pittsburgh, PA 15224 
Ph: 413-692-8648 
Squiresr@upmc.edu 
Volker Straub, M.D., Ph.D. 
Newcastle University and Newcastle Hospitals 
NHS Foundation Trust 
Central Parkway 
Newcastle upon Tyne, NE1 3BZ, UK 
Ph: +44(0)1912418762/8655 
Volker.straub@ncl.ac.uk 
 
 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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Table 5: Study Vendor Listing 
Role in Study 
Name 
Address and Telephone 
Number 
Monitor 
IQVIA Biotech 
Heather Pindel  
580 N. 4th Street, Suite 270 
Columbus, OH 43215 
Ph: 216-233-6184 
 
Heather.Pindel@iqvia.com  
EDC CRO 
Medrio 
 
Jessica Dvotsky 
 
 
Ashton Choi 
345 California St, Ste 325 
San Francisco, CA 94104 
Ph: 215-919-1368 
jdvotsky@medrio.com 
 
Ph: 415-276-9223 
achoi@medrio.com 
 
3 LIST OF ABBREVIATIONS AND DEFINITION OF TERMS 
The following abbreviations and specialist terms are used in this study protocol. 
 
Table 6: Abbreviations and Specialist Terms 
Abbreviation of Specialist Term 
Definition 
100MWT 
100 Meter Walk Test 
ABD1 
Actin Binding Domain 1 
ABD2 
Actin Binding Domain 2 
AE 
Adverse Event 
ALT 
Alanine Transaminase 
AST 
Aspartate Aminotransferase 
BA 
Bioavailability 
BE 
Bioequivalence 
BMD 
Becker Muscular Dystrophy 
BUN 
Blood Urea Nitrogen 
CBC 
Complete Blood cell Count 
CBER 
Center for Biologics Evaluation and Research 
CGH 
Comparative Genomic Hybridization  
CH1 
Calponin Homology Domain 1  
CK 
Creatinine Kinase 
CMP 
Complete Metabolic Panel 
CTCAE 
Common Terminology Criteria for Adverse Events 
DLT 
Dose Limiting Toxicity 
DMD 
Duchenne Muscular Dystrophy 
DSMB 
Data Safety Monitoring Board 
ECG 
Electrocardiogram 
ECHO 
Echocardiogram 
eCRF 
electronic Case Report Form 
ELISA 
Enzyme-Linked Immunosorbent Assay 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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Abbreviation of Specialist Term 
Definition 
ELISpot 
Enzyme-Linked ImmunoSpot 
FDA 
Food and Drug Agency 
GALGT2 
beta-1,4-N-acetyl-galactosaminyltransferase 2 
GCP 
Good Clinical Practice 
GeMCRIS 
Genetic Modification Clinical Research Information Service 
GGT 
Gamma-Glutamyl Transpeptidase 
HIV 
Human Immunodeficiency Virus 
ICF 
Informed Consent Form 
IDS 
Investigational Drug Services 
IEC 
Institutional Ethics Committee 
IFN 
Interferon 
IND 
Investigational New Drug 
IRB 
Institutional Review Board 
IRES 
Internal Ribosome Entry Site 
IV 
Intravascular 
kg 
Kilogram 
L 
Liter 
MAPH 
Multiplex Amplifiable Probe Hybridization  
min 
Minute 
mL 
Milliliter 
MLPA 
Multiplex Ligation-Dependent Probe Amplification 
MOP 
Manual of Operating Procedures 
mOsmol 
Milliosmols  
MRI 
Magnetic Resonance Imaging 
mRNA 
messenger RNA 
MVICT 
Maximum Voluntary Isometric Contraction 
NAGLU 
α-N-acetylglucosaminidase 
NCH 
Nationwide Children's Hospital 
NIH 
National Institutes of Health 
NPO 
Nothing by mouth 
NSAA 
North Start Ambulatory Assessment 
OSU 
Ohio State University 
PBMC 
Peripheral Blood Mononuclear Cell 
PCR 
Polymerase Chain Reaction 
PFT 
Pulmonary Function Test 
PI 
Principal Investigator 
PT 
Prothrombin Time 
PTT 
activated Partial Thromboplastin Time 
qPCR 
quantitative PCR 
SAE 
Serious Adverse Event 
SFC 
Spot Forming Colonies 
SGSH 
N-sulfoglucosamine sulfohydrolase 
SI 
Sponsor Investigator 
snRNA 
small nuclear RNA 
vg 
Vector Genomes 
 
 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
Protocol version 4.0 (18Nov2021) 
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4 INTRODUCTION: KEY ROLES, BACKGROUND, AND SCIENTIFIC 
RATIONALE 
4.1 Clinical Trial, Sponsor Investigator and Co-Investigators 
The study will be carried out at The Abigail Wexner Research Institute at Nationwide Children’s 
Hospital (NCH) in Columbus, Ohio, and Megan Waldrop, M.D., will serve as Sponsor 
Investigator. Dr. Waldrop is a board-certified Pediatric and Neuromuscular Neurologist and a 
faculty member of the NCH Center for Gene Therapy.  She is also an Assistant Professor of 
Pediatrics and Neurology at The Ohio State University and an attending physician in the MDA 
and SMA Clinics at NCH.  She completed fellowship training in Neuromuscular Medicine at 
Nationwide Children’s Hospital/The Ohio State University and completed a fellowship in 
Neuromuscular Genetic Therapeutics (Gene Therapy) at Nationwide Children’s Hospital.  She is 
currently a co-investigator on three active gene therapy trials at NCH
 
 
 
 
 
 
Dr. Kevin Flanigan, M.D., will serve as Co-Investigator.  Dr. Flanigan, Professor of Pediatrics 
and Neurology at the Ohio State University (OSU), is the Director of the Center for Gene 
Therapy at The Abigail Wexner Research Institute of NCH, where he holds the Robert F. & 
Edgar T. Wolfe Foundation Endowed Chair in Neuromuscular Research. He trained in 
Neurology (residency) and Neuromuscular Medicine (fellowship) at the Johns Hopkins Hospital, 
followed by postdoctoral training in Human and Molecular Genetics at the University of Utah 
before joining the faculty at NCH.  Dr. Flanigan has extensive experience as a clinical 
neuromuscular specialist and molecular geneticist.  He has participated as an investigator in 
multiple clinical trials in muscular dystrophies, including trials of nonsense suppression, exon 
skipping, and myostatin inhibition. 
 
. He is the 
principal investigator and original IND initiating investigator of two ongoing trials:
 
 
 
 
 
 
 
 
 
   
Dr. Kim McBride, M.D., M.S., will be a Co-Investigator. He is an Associate Professor in the 
Department of Pediatrics at The Ohio State University College of Medicine, the chief of division 
of medical genetics, and an Investigator in the Center for Cardiovascular and Pulmonary 
Research at The Abigail Wexner Research Institute at Nationwide Children’s Hospital. He 
received fellowship training in clinical genetics and biochemical genetics at Baylor College of 
Medicine, and is board certified in pediatrics, clinical genetics and clinical biochemical genetics. 
He has a special interest in inborn errors of metabolism and lysosomal storage diseases. He has 
participated in trials of enzyme replacement therapies, including Hunter4 and Pompe,5 and is 
active in natural history studies of Hunter syndrome,6  phenylketonuria7 and maternal 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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phenylketonuria8. More recently he has been a co-investigator on two gene transfer clinical trials: 
 
 
  Dr. McBride is also the Sponsor-Investigator of an additional gene transfer clinical 
trial: 
 
 
Dr. Wein, Co-Investigator, is an Assistant Professor of Pediatrics at OSU and a Principal 
Investigator within the Center for Gene Therapy at NCH.  He has a long-standing interest in 
genetic therapies for muscular dystrophies. He was the first author on the Nature Medicine paper 
describing the function of the IRES, and played a critical role in the generation of the vector and 
in performing preclinical studies2.  
4.2 Abstract 
The proposed clinical trial is a systemic (intravenous) delivery of scAAV9.U7.ACCA for DMD 
patients with a duplication of exon 2 in the DMD gene.  Preclinical data shows that the small 
nuclear RNA (snRNA) construct delivered by the scAAV9.U7.ACCA vector causes significant 
skipping of exon 2, resulting in exclusion of the exon from the mature messenger RNA (mRNA) 
with a high degree of efficiency, leading to mRNA containing only a single exon 2 (wild type 
[WT] mRNA) or no copies of exon 2 (Del2 mRNA).  Translation of the wild-type mRNA results 
in entirely normal dystrophin protein, whereas translation of the Del2 mRNA via translational 
initiation of an internal ribosome entry sequence, or IRES) results in a highly functional isoform 
expressed in patients known to walk into their eighth decade2. 
The study is designed as an open-label trial to assess safety and obtain preliminary efficacy data.  
scAAV9.U7.ACCA will be delivered to the systemic circulation via peripheral limb vein.  The 
subjects will receive a gene transfer dose of 3 x 1013 vector genomes per kilogram (vg/kg).   
This is a phase I/IIa safety and tolerability study.  Muscle biopsies will be performed by open 
biopsy at baseline and three months post-injection in the quadriceps muscles, and (using the 
needle biopsy approach outlined below) at 6 and 12 months post-injection in different quadriceps 
muscles (details of the muscle biopsy protocols, sites, and muscles chosen are found below in 
Section 10.1).  
4.3 Background and Significance 
4.3.1 Disease Characteristics 
DMD is the most common childhood form of severe muscular dystrophy.  Inheritance is X-
linked recessive with a birth prevalence estimate of 1 in 5000 live male births9.  Approximately 
one-third of cases represent new mutations of the DMD gene while the remainder are inherited 
on the X chromosome from a carrier mother.  Symptoms of gross motor delay may present 
around age 2 or 3 and prompt the need for diagnostic evaluation.  DMD is progressive, with loss 
of ambulation occurring, in the pre-corticosteroid treatment era, by age 1210.  Historically 
patients died from respiratory complications, but improved pulmonary and ventilatory measures 
(including nocturnal ventilatory support) have further improved survival11 making dilated 
cardiomyopathy a more common cause of morbidity and mortality. 
   
 
 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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4.3.2 Disease Pathogenesis 
The DMD gene was cloned over 30 years ago, which led to defining the molecular basis of the 
disease12 and the identification of dystrophin as the deficient protein in muscles of DMD 
patients13.  Dystrophin is a 427kDa cytoskeletal protein required for muscle fiber stability.  Loss 
of this protein results in susceptibility to repeated cycles of necrosis and regeneration with 
satellite cell depletion and diminished regenerative capacity of the muscle which ends in fat and 
connective tissue replacement (fibrosis). The mutational spectrum within the DMD gene reveals 
that deletions of one or more exons are found in ~65% of cases clustered in two hotspot 
regions14.  Originally, multiplex PCR kits were developed that were able to detect 95%-98% of 
all deletions15, 16, but improved molecular methods of diagnosis – including multiplex ligation-
dependent probe amplification (MLPA)17,  multiplex amplifiable probe hybridization (MAPH)18, 
and comparative genomic hybridization (CGH) allow screening of all exons for copy number 
variation, facilitating detection of deletions and duplications.  Methods of direct sequence 
analysis of the entire coding region are now in routine use, allowing detection of nearly all 
clinically relevant disease mutations19.  
4.3.3 Current Treatments for DMD 
The only therapeutic approach that has clearly demonstrated unequivocal efficacy for treatment 
of DMD is glucocorticoids (prednisone or deflazacort).  Treatment with one of these agents has 
been repeatedly shown to result in increased muscle strength and delayed loss of ambulation20-22.  
Deflazacort has recently gained FDA approval for treatment of boys with DMD, based largely on 
a slightly more favorable side effect profile23.  However, treatment with either of these drugs can 
be associated with significant side effects, including weight gain, Cushingoid features, 
hypertension, cataract formation, loss of bone density, vertebral compression fractures, long 
bone fractures, and behavioral problems.  
Molecular therapies, such a gene replacement, exon skipping, and nonsense mutation 
suppression provide a promising alternative to glucocorticoids.  Exon skipping utilizes antisense 
oligomers to induce altered pre-mRNA splicing to restore an open reading frame, and eteplirsen 
has received FDA approval, based largely on a small but statistically significant increase in 
dystrophin expression24, 25.  Nonsense suppression utilizes an oxadiazole to enable ribosomal 
readthrough of nonsense mutations to theoretically make full length dystrophin (PMID 
28728956).   However, both exon skipping and the as-yet unapproved nonsense suppression 
approach are only beneficial to small subsets of patients with specific mutations, 13% and 15% 
respectively26-28.  
4.4 Rationale 
4.4.1 Rationale for Gene Therapy using U7snRNA exon skipping 
Two exon splicing modification therapies have been shown to be effective and are FDA-
approved in neuromuscular diseases, each using antisense oligomers: nusinersen in spinal 
muscular atrophy29, and eteplirsen in DMD30.  The U7 gene therapy approach uses a non-
spliceosomal snRNA targeted to a specific exon via an antisense sequence.  The U7snRNA 
interferes with spliceosome assembly with high efficiency, resulting in the exclusion of the target 
exon from the mRNA31. In contrast to antisense oligomer therapies, long term skipping can be 
achieved with a single dose, and this approach has been effective in improving muscle function 
in animal models of DMD32, 33.   
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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4.4.2 Rationale for therapeutic skipping of exon 2 
Duplications of one or more exons account for 6% to 11% of all DMD-associated mutations34, 35, 
and duplications of exon 2 are the most common, accounting for 10% of all duplication 
mutations.   Dr. Flanigan’s laboratory has developed an adeno-associated virus vector 
(scAAV9.U7.ACCA) in which four copies of U7snRNA are contained within the viral genome.  
Two copies are targeted toward the exon 2 splice donor site, and two are targeted toward the 
splice acceptor site.   
 
In this study, scAAV9.U7.ACCA will be utilized to skip one or both of the duplicated exon 2 in 
the DMD gene.  Viral-based skipping of exon 2 is a viable therapeutic approach because of an 
unusual feature of the DMD gene that provides an enormous therapeutic window.  Specifically, if 
only one exon is skipped, wild-type dystrophin will be produced, but if both exons 2 are skipped, 
protein translation will utilize the IRES in exon 5 to produce a highly functional N-truncated 
dystrophin2.  
 
The most relevant preclinical studies supporting this have all been published by the Flanigan 
laboratory over the past eight years.  The feasibility of this approach stems from the 
identification of the first North American founder allele in DMD, a predicted nonsense mutation 
within exon 1 (c.9G>A; p.Trp3X), which resulted in very mild Becker muscular dystrophy.  It 
was first identified in several families in a cohort of 1,100 dystrophinopathy patients35, who 
along with other families were then shown to share a 3.7 million base pair region extending 
across exon 1, confirming that it was a founder allele in this population 36. Despite the fact that a 
nonsense mutation should result in expression of no dystrophin, symptoms were minimal.  Most 
patients had only myalgias with exertion; the most severely affected individual stopped walking 
at age 62, while others had no symptoms into their eighth decade36. 
 
It was demonstrated that this mild phenotype was due to the expression of significant amounts of 
an isoform of dystrophin resulting from translational initiation from an AUG codon within exon 
6 of DMD37.  The resulting isoform is approximately 413 kiloDaltons (kD) in size, and lacks the 
calponin homology domain 1 (CH1) that makes up the first half of the actin binding domain 1 
(ABD1) at the N-terminus of dystrophin.  Despite the canonical view that an intact ABD1 is 
required for dystrophin functionality, the human “experiment” – the presence of the founder 
allele in humans without significant symptoms, and clearly no effect on reproductive fitness – 
argues that the dystrophin rod-domain-located actin binding domain 2 (ABD2) is sufficient for 
significant preservation of muscle function. 
 
Using a dual-luciferase reporter approach, the IRES activity was mapped to exon 5, and showed 
that patients with other 5’ mutations express the ΔCH1 isoform 37,2.  This work, published in 
Nature Medicine, established the presence of a glucocorticoid-responsive IRES consisting of 
nearly the entire exon.  In order to establish the potential clinical utility of activation of this 
IRES,  a new mouse model of DMD was made with a duplication of mouse exon 2 that 
essentially mimics the phenotypic features of the standard mdx model (which contains a 
nonsense mutation in exon 23) 3. It was noted that duplications of exon 2 were nearly always 
associated with DMD, even though the duplication resulted in an altered reading frame and 
hence a premature termination codon in the second copy.  At the same time, it was noted that 
deletions of exon 2, which are similarly out-of-frame, had never been reported in either Dr. 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
Protocol version 4.0 (18Nov2021) 
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Flanigan’s large cohort or in other exhaustive dystrophinopathy mutational databases available34, 
38.  Assuming that such a mutation had never been ascertained because of IRES activity, it was 
hypothesized that the IRES was active in the absence of exon 2 but not in the presence of a 
duplicated exon 2.  Consistent with this hypothesis, studies in the dual luciferase reporter system 
showed this to be the case, and this was confirmed by the subsequent clinical identification of an 
essentially asymptomatic patient with a deletion of exon 22.          
 
The immediate therapeutic implication of these results is that skipping of exon 2 would be 
expected to result in IRES activation.  Furthermore, it is important to emphasize that for 
patients with DMD due to a duplication of exon 2, the presence of a functional IRES provides a 
wide margin of safety, as boys with exon 2 duplications cannot be “overtreated” to be made 
worse.  Exon 2 skipping will result in either the wild-type transcript containing one copy of exon 
2, or in the complete exclusion of exon 2, resulting in IRES activation and the expression of the 
ΔCH1 isoform which – as is demonstrated by patients who express it – is highly protective.   
 
5 OBJECTIVES AND PURPOSE 
5.1 Primary Outcome 
Safety is the primary outcome for this clinical gene transfer trial.  Stopping criteria are based on 
development of unacceptable toxicity defined as the occurrence of two or more unexpected 
Grade III or higher treatment-related toxicities. 
5.2 Secondary Outcome 
Secondary efficacy outcome measures: Muscle biopsy analysis for safety and dystrophin 
expression will be studied using multiple serial sections.  
 
a) Expression of dystrophin as demonstrated by immunofluorescent in staining with a 
dystrophin C-terminus antibody and the exon 1-encoded epitope antibody (Manex 1A) in 
muscle biopsy sections. 
b) Dystrophin protein expression quantified by western blot using a dystrophin C-terminus 
antibody and the exon 1-encoded epitope antibody (Manex 1A) and assessed by 
densitometry in muscle biopsy tissue. 
c) Percentage of exon 2 exclusion from the dystrophin mRNA transcript as measured by 
RT-PCR analysis. 
d) Transduction efficiency will be measured by qPCR of a U7-targeting sequence primer 
pair amplifying the DNA containing the U7snRNA, and expressed as vector genomes 
normalized to a single-copy genomic DNA control in each muscle sample. 
 
For immunofluorescence, multiple serial sections will be studied. For each of these measures, 
statistical analysis based on differences between pre- and post-gene transfer muscle specimens 
will be analyzed using a paired t test (p < 0.05).  Sections and western blots will be prepared by a 
single technician, who will blind the sample identifier; images will be obtained and expression 
assessed by another technician blinded to the sample time point.  Quantification of 
immunofluorescent staining will be performed using a published protocol39 that has been used 
for comparisons between multiple labs with highly reproducible results40, with the results 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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reported as membrane-localized dystrophin staining intensity relative to normal muscle sections.  
The muscle analysis of gene expression will occur without breaking the blind. 
Splicing of exon 2 will be assessed by RT-PCR analysis, which will be performed under the 
same blinding regimen as used for protein expression studies.  RT-PCR and data analysis will be 
performed under standardized protocols, resulting in quantification of unskipped (Dup2), wild-
type (WT), and exon 2 deleted (Del2) transcripts.  Notably, both WT and Del2 transcripts are 
therapeutic. 
Transduction efficiency will be measured by qPCR of a U7-targeting sequence primer pair 
amplifying the DNA containing the U7snRNA, and expressed as vector genomes normalized to a 
single-copy genomic DNA control in each muscle sample. 
5.3 Exploratory Outcomes 
Exploratory functional outcome measures will be performed on the schedule as described. 
 
A) Muscle Function at 6 and 12 months post gene transfer in comparison to baseline values as 
follows$: 
a) In subjects enrolling under at 36 months of age or younger: 
• Improvement or statistically significant stabilization of Bayley-IV scores (Bayley 
Scales of Infant and Toddler Development) 
b) In subjects enrolling at greater than 36 months of age and younger than 5 years of age: 
• Improvement or statistically significant stabilization of the North Star Ambulatory 
Assessment (NSAA) score. 
• Time to climb 4 stairs 
• The amount of time required to walk 100 meters (100MWT) 
 
c) In subjects enrolling at 5 years or greater, improvement or statistically significant of the 
following: 
• North Star Ambulatory Assessment (NSAA) score. 
• Time to climb 4 stairs 
• The amount of time required to walk 100 meters (100MWT) 
• The force generated in knee flexion and extension in the maximum voluntary 
isometric contraction (MVICT) 
$Subjects will be tested using age appropriate assessments. 
B) As an exploratory efficacy measure, muscle MRI will be assessed using T2-weighted multi-
slice spin echo (SE) axial images and single voxel 1H-MRS from both the lower leg and 
thigh to measure lipid fraction and (1)H2O T2 signal.  These will be performed at baseline 
and at 6, 12, 18 and 24 months post infusion, but only if sedation is not required.  Data will 
be assessed using published techniques1 and compared to baseline values in collaboration 
with Dr. Glenn Walter, University of Florida (see Section 10.2 for more details). 
  
C) As an exploratory efficacy measure, the percentage of exon 2 exclusion from the dystrophin 
mRNA transcript will be measured by RNA-Seq analysis. 
 
D) Additional analyses performed on muscle biopsy tissue will include: 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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a) Immune staining at 90 and 180 days as well as 12 months will be performed on biopsy 
specimens for members of the dystrophin-associated protein complex, including the 
sarcoglycans and β-dystroglycan.   
b) Muscle morphometrics will also be performed, including fiber size histograms and 
quantification of central nucleation at 90 and 180 days as well as 12 months. 
 
6 STUDY DESIGN AND ENDPOINTS 
6.1 Overall Study Design 
The proposed phase I/IIa clinical trial is an open-label trial where subjects will take part in a 
single injection study of self-complementary AAV9 carrying the ACCA snRNA under the 
control of a U7 promoter (scAAV9.U7.ACCA) delivered one time through a peripheral vein of 
the DMD subjects. 
The comparison of muscle tissue from baseline and 90 days post-gene transfer will be the basis 
of the secondary efficacy analysis. Upon completion of the 2-year study period, subjects will be 
monitored annually as per standard of care for up to 5 years.  
There will be at least 6 weeks between dosing of the first and second subjects within the cohort.  
The allowance of 6 weeks between dosing of the first and second subjects in the cohort provides 
time for an DSMB review of the safety analysis from five time points (Days 1, 2, 7, 14 and 30) 
prior to dosing of the next subject.   
Notably, we have not required a 30 day time point between subsequent subjects because we wish 
to accommodate the realistic possibility that siblings may both enroll in the study, and viral 
shedding might possibly expose the second of each sibling pair to the vector, generating 
antibodies that are exclusionary to the trial.  We would consider the burden on families of living 
apart for six weeks to be excessive, and the prospect of making parents choose between one or 
another of their sons to enroll to be ethically untenable. 
6.2 Study Population  
All subjects will be enrolled at Nationwide Children’s Hospital. 
DMD subjects aged between 6 months and less than 14 years of age, with proven exon 2 
duplication mutations of the DMD gene, will be eligible for enrollment in the gene transfer 
study. Subjects of any ethnic or racial background will be enrolled. 
6.3 Number of Subjects 
Up to 3 DMD subjects matching the inclusion/exclusion criteria will be enrolled. Enrollment will 
be proceed as outlined in Section 6.4. 
6.4 Dosing plan 
The proposed phase I/IIa clinical trial is an open-label single injection study of systemic 
(intravenous) delivery of scAAV9.U7.ACCA for DMD subjects with associated duplications of 
exon 2 in the DMD gene.  Preclinical data shows that the small nuclear RNA (snRNA) construct 
delivered by the scAAV9.U7.ACCA vector causes significant skipping of exon 2, resulting in 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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exclusion of one (wild type) or both copies (Del2) of the exon from the mature messenger RNA 
(mRNA) with a high degree of efficiency.  Translation of either the wild-type mRNA or N-
truncated mRNA (via IRES translational initiation) results in a highly functional isoform. The 
dose selection is based on toxicology-biodistribution studies and predicted by pre-clinical studies 
in non-human primates. 
This protocol is designed to deliver the vector via a one-time infusion through a peripheral vein 
of the DMD subjects. The subjects will receive the minimal efficacious dose (3 x 1013 vg/kg) as 
identified in murine studies. The total vector genome dose for subjects who weigh ≥ 20kg will be 
adjusted by rounding the subject body weight up to the closest whole kilogram. The total vector 
genome dose for subjects who weigh <20kg will be adjusted by rounding the subject body 
weight to the nearest tenth of a kilogram. 
Dose (vg/kg) 
N 
3.0 x 1013 
1-3 
 
6.5 Criteria for Study Termination 
An independent Data Safety Monitoring Board (DSMB) will be established. Safety data will be 
monitored on a continual basis throughout the trial.  The DSMB can recommend early 
termination of the trial for reasons of safety.  Study enrollment will be halted by the investigators 
when any subject experiences two or more Grade 3, or higher adverse events that are 
unanticipated and possibly, probably, or definitely related to the study drug.  This will 
include any subject death, important clinical laboratory finding, or any severe local complication 
in the injected area related to administration of the study agent.  If after review by the DSMB, the 
decision is made to continue, the study will proceed according to Section 6.4 of this protocol. 
6.6 Establish Subject Identification Number 
All subjects will be given a unique sequentially assigned subject number. Subjects will be 
identified by code only to protect identity. This will be done prior to any research procedures 
when the subject is identified as a potential candidate.  
The subject identification number will be generated as described below: 
1) U7 (study identifier) 
2) DUP2 (to indicate a DMD gene transfer trial) 
3) The last two digits will represent the sequential number of candidates screened 
Example: Subject ID: U7-DUP2-01 
7 STUDY ENROLLMENT AND WITHDRAWAL 
All subjects will be enrolled at Nationwide Children’s Hospital.  Subjects will be identified from 
among the muscular dystrophy clinic population, as well as among a large number of treatment-
amenable subjects who have self-identified themselves to the investigators.    
7.1 Inclusion/Exclusion 
Inclusion Criteria 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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1. Age greater than 6 months and less than 14 years.   
2. Confirmed duplication of exon 2 the DMD gene using a clinically accepted technique 
that completely defines the mutation19, 35. 
3. Pre-ambulant (not yet walking) or ambulant (as defined by the ability to walk 10 meters 
without assistance). 
4. Males of any ethnic group will be eligible 
5. Ability to cooperate with muscle testing. 
6. In subjects age 4 and above, stable dose and regimen of corticosteroid therapy (including 
either prednisone or deflazacort and their generic forms) for at least 12 weeks prior to 
gene transfer. 
 
Exclusion Criteria 
1. Active viral infection based on clinical observations. 
2. Symptoms or signs of cardiomyopathy, including:  
a. Dyspnea on exertion, pedal edema, shortness of breath upon lying flat, or rales at 
the base of the lungs 
b. Echocardiogram with ejection fraction below 40% 
3. Serological evidence of HIV infection, or Hepatitis B or C infection 
4. Diagnosis of (or ongoing treatment for) an autoimmune disease  
5. Persistent leukopenia or leukocytosis (WBC ≤ 3.5 K/µL or ≥ 20.0 K/µL) or an absolute 
neutrophil count < 1.5K/µL 
6. Concomitant illness or requirement for chronic drug treatment that in the opinion of the 
SI creates unnecessary risks for gene transfer 
7. AAV9 binding antibody titers ≥ 1:400 as determined by ELISA immunoassay 
8. Abnormal laboratory values in the clinically significant range in Table 7, based upon 
normal values in the Nationwide Children’s Hospital Laboratory 
 
Table 7: Normal and abnormal laboratory values at Nationwide Children’s Hospital 
Laboratory 
System 
Assay 
Normal Range 
Abnormal 
(Clinically Significant) 
Liver Function 
GGT 
8-80 U/L 
>80 U/L 
Total Bilirubin 
0.1-1 mg/dL 
≥3 mg/dL 
Direct Bilirubin 
<0.6 mg/dL 
>2 mg/dL 
Renal Function 
Cystatin C 
0.3-1.3 mg/dL 
>1.8 mg/dL 
Hematologic 
Hemoglobin 
(g/dL) 
2 – 6 yrs: 11.5-13.5 
6 – 12 yrs: 11.5-15.5 
13-18 years: 13-15 
For all ages: 
≤ 8 or ≥ 18 g/dL 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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Hematologic 
White Blood 
Cells 
4 – 6 yrs: 5.5-15.5 
6 – 10 yrs: 5-14.5 
10 – 21 yrs: 4.5-13.5 
For all ages: 
White blood cell count  
≤ 3.5 or ≥ 20 x 103 
cells/mL 
OR 
Absolute neutrophil count 
of ≤ 1.5 x 103 cells/mL 
*  For Hemoglobin, subjects will be excluded if they have a value less than the lower limit of 
normal for age at screening regardless of age.  Abnormal but not clinically significant 
hemoglobin levels may be repeated during the screening period, and the subject accepted 
for enrollment if the results are not less than the lower limit of normal. 
7.2 Subject Withdrawal Criteria 
If a subject chooses to withdraw at any point BEFORE gene transfer, no further follow up will be 
required, other than routine post-operative care if a muscle biopsy was completed.  
If a subject chooses to withdraw at any point AFTER gene transfer, all safety laboratory studies 
will still need to be completed. The subject will not be required to undergo any further muscle 
biopsies, muscle MRIs (if applicable) or functional assessments.  
8 STUDY AGENT 
Preparation of the scAAV9.U7.ACCA gene vector will be done by the Nationwide Children’s 
Hospital Investigational Drug Services (IDS) according to the Manual of Operating Procedures 
(MOP). The vector will be delivered undiluted to the gene transfer venue at Nationwide 
Children’s Hospital in 50mL Becton Dickinson Luer-Lok polypropylene syringe(s).  It will be 
delivered at room temperature (not frozen) and administered to the subject within 24 hours of 
preparation.   Handling of scAAV9.U7.ACCA vector will follow compliance standards for 
Biosafety Level 1 vectors: (http://osp.od.nih.gov/sites/default/files/NIH_Guidelines.html)  
 
The infusion rate will not be slower than the slowest rate at which the infusion set up 
compatibility testing was performed. The infusion rate will not approach or exceed 2 mL/kg/min 
(a rapid bolus rate) for any pediatric subject. The infusion will be given over approximately 60 to 
90 minutes, incorporating competing concerns including maximizing uptake in the musculature 
and limiting sedation time versus limiting risk for IV infiltration and infusing slowly enough to 
observe for evidence of infusion reaction before entire dose is given. Depending on vector lot 
concentration, and other factors, infusion rate is expected to be around 0.03-0.08 mL/kg/min. 
 
The vector will be flushed from the infusion tubing using 20 mL Lactated Ringer’s/Hartmann’s 
solution at the end of the infusion. The vector request, shipment, storage, preparation and 
management will be documented by the IDS. 
 
 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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8.1 Description of the Study Drug 
 
Investigational Product 
Product Name 
scAAV9.U7.ACCA 
Dosage Form 
The vector titer is based off linear qPCR and is stored in 2.0mL 
vials, filled with 1.0 mL per vial. 
All vials are 2.0mL screwcap polypropylene vials 
Unit Dose 
N = 1-3, 3.0 x 1013 vg/kg 
Route of Administration 
Intravenous injection of scAAV9.U7.ACCA via peripheral limb 
vein 
Physical Description 
scAAV9.U7.ACCA is provided frozen in vials. Once thawed, 
scAAV9.U7.ACCA is a clear liquid 
Manufacturer 
Nationwide Children’s Hospital 
Clinical Manufacturing Facility 
575 Children’s Crossroads 
Columbus, OH 43215 
 
9 STUDY PROCEDURES AND SCHEDULE 
Subjects will complete 12 onsite visits and up to 5 local blood draws until 2 years post-transfer. 
9.1 Study Visits 
9.1.1 Screening (day -45 to day -1) 
Informed consent will be obtained prior to the collection of any data and any study related 
procedures. Baseline measures from subjects will be obtained prior to gene transfer and will 
occur over two visits (up to 3 weeks apart). Because a pre-existing antibody response would be 
exclusionary, ELISA and ELISpot testing will be performed at the first visit, allowing the results 
to be reviewed prior to the open muscle biopsy and its subsequent risks (including sedation). The 
weight obtained at screening will be used to calculate the viral vector dose.  
The following assessments will occur at these visits: 
• Informed Consent 
• Assent, if applicable 
• Medical history 
• Concomitant Medications 
• Physical Exam 
• Vital Signs (temperature, respiratory rate, heart rate, blood pressure, height (see Section 
10.5 for more details), and weight) 
• MRI of the lower legs (see Section 10.2 for more details) 
• ECHO 
• ECG 
• Functional tests (variable depending on age of enrollment) 
o Enrollment age: ≤ 36 months 
▪ Bayley-IV scores (Bayley Scales of Infant and Toddler Development) 
o Enrollment age: > 36 months and < 5 years of age 
▪ North Star Ambulatory Assessment (NSAA) 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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▪ Time to climb four stairs 
▪ Amount of time required to walk 100 meters (100MWT) 
 
o Enrollment age:  ≥ 5 years old until Non-Ambulant 
▪ North Star Ambulatory Assessment (NSAA) score 
▪ Time to climb four stairs 
▪ Amount of time required to walk 100 meters (100MWT) 
▪ Force generated in knee flexion and extension in maximum voluntary 
isometric contraction (MVICT).   
• Pulmonary functional tests (PFTs), including forced vital capacity.  (See Section 10.3 for 
more details.) 
• Open Muscle Biopsy (For details, see Section 10.2)  
The following blood lab work will occur at this visit.  (Lab work may be drawn over several days 
during visit if deemed necessary by study team except as noted above for ELISA and ELISpot.) 
• Complete blood cell count (CBC) and differential, platelets 
• Aspartate aminotransferase (AST)/alanine transaminase (ALT) 
• Gamma-glutamyl transpeptidase (GGT)* 
• A Comprehensive Metabolic Panel (CMP) that includes: 
o Albumin/Serum total protein 
o Serum total bilirubin 
o Electrolytes 
o Calcium 
o Creatinine/Blood Urea Nitrogen (BUN) 
o Serum glucose 
o Alkaline phosphatase  
• Prothrombin time (PT)/activated partial thromboplastin time (PTT)/INR 
• Amylase 
• Creatine kinase (CK) 
• Cystatin C 
• Total IgG 
• D-Dimer** 
• Direct Bilirubin 
• SC5b-9** 
• C3, C4** 
• CH50** 
• Antibody testing for Hepatitis B, C and HIV 
• ELISA for detection of total antibodies to AAV9 
• ELISpots for T-cell responses to AAV9 and Dystrophin 
• Blood for vector shedding studies 
 
* For subjects taking drugs that potentially induce GGT synthesis through the cytochrome P450 system, the 
clinically significant range will be estimated starting at two times the levels obtained from the baseline screening 
test and up. 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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** These laboratory tests have been recommended by the DSMB to be obtained at screening. These will not 
necessarily be exclusionary but having a baseline value will be helpful for comparison if a subject were to have any 
platelet or renal abnormalities that may be immune mediated after gene transfer.  
 
The following urine lab work will occur at this visit: 
• Urinalysis 
• Urine for biobanking  
• Urine for vector shedding studies 
 
The following additional samples will be collected for vector shedding studies: 
• Saliva 
• Feces 
 
For children under 10 kg in weight, the blood draw schedule will be reviewed to limit blood 
draw volumes to ≤ 3% of total body volume over a 24 hour period and ≤ 10% of total body 
volume over a 30 day period.  The study team may deem it necessary to move or remove non-
safety-related blood draws to other days if the volume drawn in one 24-hour period will be too 
near to 3% of total body volume.   
Repeat testing of laboratory abnormalities within the screening period:  Given the biologic 
variability inherent in some laboratory testing results, we will retest subjects with minimal and 
not clinically significant abnormalities resulting in values outside the NCH lab’s normal range as 
defined by Table 7, such as minimal abnormalities of hemoglobin or CBC values.  Similarly, 
subjects with anti-AAV9 endpoint titers at (but not above) the cutoff level of 1:400 will be re-
tested for confirmation of titer.  Normal values on the second of these tests will be considered to 
be acceptable for enrollment. 
 
 

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9.1.2 Gene Transfer Visit 
9.1.2.1 Pre Infusion Visit (Day -1) 
Subjects will arrive at the Nationwide Children’s Hospital within 24 hours prior to gene transfer.  
The following assessments will occur at this visit: 
• Medical history 
• Concomitant Medications 
• Adverse Events 
• Physical Exam 
• Vital Signs (heart rate, respiratory rate, temperature, blood pressure, height (see Section 
10.5 for more details) and weight) 
 
The following blood lab work will occur at this visit: 
• Complete blood cell count (CBC) and differential, platelets 
• Aspartate aminotransferase (AST)/alanine transaminase (ALT) 
• Gamma-glutamyl transpeptidase (GGT)* 
• A Comprehensive Metabolic Panel (CMP) that includes: 
o Albumin/Serum total protein 
o Serum total bilirubin 
o Electrolytes 
o Calcium 
o Creatinine/Blood Urea Nitrogen (BUN) 
o Serum glucose 
o Alkaline phosphatase  
• Prothrombin time (PT)/activated partial thromboplastin time (PTT)/INR 
• Creatine kinase (CK) 
• Direct Bilirubin 
• ELISpots for T-cell responses to AAV9 and Dystrophin 
• Whole blood for biobanking 
* For subjects taking drugs that potentially induce GGT synthesis through the cytochrome P450 system, the 
clinically significant range will be estimated starting at two times the levels obtained from the baseline screening 
test and up. 
 
In previous gene therapy studies at NCH, there has been an observed antigen specific T-cell 
response to the AAV vectors. This is an expected response between 2 - 4 weeks following gene 
transfer. One possible consequence to such antigen specific T-cell responses is clearance of the 
transduced cells and loss of transgene expression.  All subjects (ages 4 years and older) will 
already be on prednisone, prednisolone, or deflazacort for 12 weeks at time of enrollment as part 
of the inclusion criteria.  The standard clinical dose of prednisone or prednisolone is 0.75 
mg/kg/day, and the standard clinical dose of deflazacort is 0.9 mg/kg/day. In order to dampen the 
host immune response to our AAV-based therapy, for each subject the glucocorticoid dose will 
be increased to prednisone or prednisolone 1.0 mg/kg/day, not to exceed 60 mg/day; subjects on 
deflazacort at enrollment will be placed on prednisone or prednisolone for the immediate peri-
injection period. For subjects younger than 4 years of age who may not be on a corticosteroid, 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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prednisolone will be prescribed for the purposes of dampening the host immune response.  This 
treatment will begin one day prior to the gene transfer and only after passing all inclusion and 
exclusion criteria (in the event that a measure is repeated on Day -1).  All subjects will also be 
prescribed a proton pump inhibitor (PPI) for the purposes of dampening potential gastrointestinal 
upset related to the increased corticosteroid dosage.  This corticosteroid and PPI regimen will be 
continued after gene transfer as discussed in Section 10.6.1, followed by a return to the pre-
treatment corticosteroid regimen.  
9.1.2.2 Day of Gene Transfer (Day 0) 
 
The venue for gene transfer will be determined during the screening visit.  There is a range of 
subject ages, from 6 months through age 14 years.  For the youngest subjects, sitting still during 
the gene transfer (up to 90 minutes) may be problematic.  For these youngest subjects, gene 
transfer will be performed in the hospital’s Procedure Center under sedation, and they will be 
monitored in the Pediatric Intensive Care Unit after the gene transfer.  For older subjects – 
anticipated to be any subject above the age of 3 years – the gene transfer will be performed 
within the Pediatric Intensive Care Unit itself, followed by monitoring in the same unit. Transfer 
out of the PICU may be undertaken after the initial 24 hours of post-infusion monitoring, if the 
SI has no concerns. 
 
Prior to Gene Transfer, the following assessments will occur at this visit: 
• Medical history 
• Concomitant Medications 
• Adverse Events 
• Physical Assessment (physical exam will consist of the anesthesia evaluation prior to 
sedation or by the study team if sedation is not required)) 
• Vital Signs (temperature, respiratory rate, heart rate, and blood pressure) 
 
The following procedures work will occur at this visit: 
• Picture of Injection Site, both prior to and after Gene Transfer 
• Gene Transfer 
 
The following samples will be collected for vector shedding studies 4 and 8 hours post gene 
transfer (see Section 10.4 for details): 
• Blood 
• Urine 
• Saliva 
• Feces 
 
Prior to vector infusion, an assessment will be performed of vitals to be collected for clinical 
purposes by either anesthesia staff or the study team, depending on the planned use of anesthesia 
or not.  
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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If sedation is required for subjects, they will continue on their usual diet until eight hours prior to 
gene transfer, after which they will have no solid food; clear liquids will be allowed up until two 
hours prior to gene transfer, after which they will be fully NPO. They will resume their usual diet 
after they have returned to pre-sedation baseline. The specifics of the gene transfer itself follow 
below. If no sedation is required, subjects will be allowed a normal diet until 2 hours prior to the 
gene transfer after which they will need to be fully NPO. 
If the subject appears inadequately hydrated in the judgment of the SI, bolus(es) of 10-20 mL/kg 
normal saline may be given during the time between either admission or Procedure Center 
check-in and gene transfer. 
If sedation is not required, the subject will become NPO 2 hours prior to gene transfer and will 
remain NPO for 2 hours after gene transfer is completed. D5 1/2NS will be infused in the second 
IV during the NPO period. 
Gene transfer will be performed under sterile conditions within an appropriate inpatient facility. 
If deemed necessary by the SI, the gene transfer will occur under light to moderate sedation 
under the direction of a qualified anesthesiologist. If required, the subjects will be given sedation 
utilizing NCH anesthesia protocols that minimize the risk of anesthetic reaction in muscular 
dystrophy. 
 
Sedation is being utilized in this instance to minimize anxiety or uncooperative behavior in the 
subjects, with an aim of maximizing potential study participation benefit to the pediatric subject 
by preventing study agent subcutaneous infiltration or IV loss with study agent spillage (the 
subjects will receive the minimal efficacious dose identified in the animal model). In those 
subjects who in the opinion of the SI (and in consultation with the anesthesiologist and parents) 
are determined to not need sedation in order to safely deliver the vector, sedation may be 
deferred, and the vector delivered in the PICU. 
 
All subjects participating in this trial will receive an intravenous injection of scAAV9.U7.ACCA 
via peripheral limb vein. Dosing is the minimum efficacious dose as established in murine studies.   
 
• 
n= 1-3 – 3.0 X 1013 vg/kg  
 
The total vector genome dose for subjects who weigh ≥ 20kg will be adjusted by rounding up to 
the closest whole kilogram. The total vector genome dose for subjects who weigh < 20kg will be 
adjusted by rounding the subject body weight to the nearest tenth of a kilogram. Dosing is 
extrapolated from efficacy observed in preclinical studies as the minimal efficacious dose 
identified in the animal model.  
 
Subjects will be closely monitored for side effects during the infusion, including continuous 
heart rate, respiratory rate, and pulse oximetry; and intermittent blood pressure monitoring. Heart 
rate, respiratory rate, pulse oximetry (only if sedation is being utilized), temperature, and blood 
pressure will be measured before and immediately after the infusion, and at least every five 
minutes during the infusion if sedated, and every 15 minutes if not sedated. Monitoring will be 
repeated at 15 minutes post-infusion, every 15 minutes for 4 hours following the infusion, and 
then every hour until transfer from the PICU to the Neurology floor.  On the Neurology floor, 
vital signs will be measured every 4 hours until discharge.  

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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Infusion reactions:  Infusion will be terminated for evidence of an allergic reaction of grade 2 or 
greater, including anaphylaxis, based upon CTCAE v5.0 criteria, and reported using these 
criteria: 
 
Grade 1: Transient flushing or rash, drug fever <38° C (<100.4° F); intervention not indicated  
Grade 2: Rash, flushing, urticaria, dyspnea, drug fever >38° C:  
 
Intervention or infusion interruption indicated; responds promptly to symptomatic 
treatment (e.g., antihistamines, NSAIDS, narcotics); prophylactic medications 
indicated for ≤24 hrs  
Grade 3:  Prolonged (e.g., not rapidly responsive to symptomatic medication and/or brief 
interruption of infusion); recurrence of symptoms following initial improvement; 
hospitalization indicated for clinical sequelae (e.g., renal impairment, pulmonary 
infiltrates) 
Grade 4:  Life-threatening consequences; urgent intervention indicated 
Grade 5: Death 
 
Under CTCAE v5.0 criteria, anaphylaxis is by definition grade 3 (Symptomatic bronchospasm 
with or without urticaria; allergy-related edema/angioedema, hypotension), and would result in 
infusion termination and systemic treatment. 
 
Subjects will remain in an independent hospital room following gene transfer and remain 
hospitalized for up to 48 hours post gene transfer. 
9.1.3 Post Gene Transfer Monitoring Plan 
To facilitate these scheduled visits, all subjects will remain in the Columbus metropolitan area 
for a minimum of two weeks following gene transfer.  If there is a systemic immune response 
that leads to acute respiratory distress syndrome, which implies a severe inflammation of the 
lung parenchyma, the subject would be hospitalized and treated aggressively with fluid 
replacement for hypotension if necessary and receive pulse methylprednisolone treatment (1 
gram per day) over 3-5 days.   
9.1.3.1 Day 1 
The following assessments will occur at this visit: 
• Medical history 
• Concomitant Medications 
• Adverse Events 
• Physical Exam 
• Vital Signs (blood pressure, heart rate, respiratory rate, and temperature) 
 
The following blood lab work will occur at this visit: 
• Complete blood cell count (CBC) and differential, platelets 
• Aspartate aminotransferase (AST)/alanine transaminase (ALT) 
• Gamma-glutamyl transpeptidase (GGT)* 
• A Comprehensive Metabolic Panel (CMP) that includes: 
o Albumin/Serum total protein 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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o Serum total bilirubin 
o Electrolytes 
o Calcium 
o Creatinine/Blood Urea Nitrogen (BUN) 
o Serum glucose 
o Alkaline phosphatase  
• Prothrombin time (PT)/activated partial thromboplastin time (PTT)/INR 
• Creatine kinase (CK) 
• Direct Bilirubin 
• Blood for vector shedding studies (see Section 10.4 for details) 
* For subjects taking drugs that potentially induce GGT synthesis through the cytochrome P450 system, the 
clinically significant range will be estimated starting at two times the levels obtained from the baseline screening 
test and up. 
 
The following urine lab work will occur at this visit: 
• Urinalysis 
• Urine for biobanking  
• Urine for vector shedding studies (see Section 10.4 for details) 
 
The following additional samples will be collected for vector shedding studies (see Section 10.4 
for details): 
• Saliva 
• Feces 
 
The following procedures will occur at this visit: 
• Picture of Injection Site 
 
9.1.3.2 Day 2 
On the second day after gene transfer (Day 2) the subject will be discharged from the hospital 
following approval from by a study team doctor. 
The following assessments will occur at this visit: 
• Medical history 
• Concomitant Medications 
• Adverse Events 
• Physical Exam 
• Vital Signs (blood pressure, heart rate, respiratory rate, and temperature) 
 
Blood will be collected for vector shedding studies (see Section 10.4 for details): 
The following urine lab work will occur at this visit: 
• Urine for biobanking  
• Urine for vector shedding studies (see Section 10.4 for details) 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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The following additional samples will be collected for vector shedding studies (see Section 10.4 
for details): 
• Saliva 
• Feces 
 
9.1.3.3 Days 7 (± 2 days), 14 (± 2 days), and 30 (± 3 days) 
Subjects will return for follow up visits on Days 7, 14, and 30 after treatment, with repeat 
examinations, blood and urine work as outlined below.  
 
The following assessments will occur at this visit: 
• Medical history 
• Concomitant Medications 
• Adverse Events 
• Physical Exam 
• Vital Signs (heart rate, respiratory rate, temperature, weight, height (see Section 10.5 for 
more details), and blood pressure) 
 
The following blood lab work will occur at this visit: 
• Complete blood cell count (CBC) and differential, platelets 
• Aspartate aminotransferase (AST)/alanine transaminase (ALT) 
• Gamma-glutamyl transpeptidase (GGT)* 
• A Comprehensive Metabolic Panel (CMP) that includes: 
o Albumin/Serum total protein 
o Serum total bilirubin 
o Electrolytes 
o Calcium 
o Creatinine/Blood Urea Nitrogen (BUN) 
o Serum glucose 
o Alkaline phosphatase  
• Prothrombin time (PT)/activated partial thromboplastin time (PTT)/INR 
• Amylase 
• Creatine kinase (CK) 
• Total IgG (Day 30 only) 
• Direct Bilirubin 
• ELISpots for T-cell responses to AAV9 and Dystrophin 
• Blood for vector shedding studies (see Section 10.4 for details) 
* For subjects taking drugs that potentially induce GGT synthesis through the cytochrome P450 system, the 
clinically significant range will be estimated starting at two times the levels obtained from the baseline screening 
test and up. 
The following urine lab work will occur at this visit: 
• Urinalysis 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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• Urine for biobanking  
• Urine for vector shedding studies (see Section 10.4 for details) 
 
The following additional samples will be collected for vector shedding studies (see Section 10.4 
for details): 
• Saliva 
• Feces 
 
On Day 7, additional tests will include complement studies, consisting of Sc5b-9, C3, C4, and 
CH50. 
 
On Day 30, additional tests will include an ELISA for detection of total antibodies to AAV9, 
Cystatin C, and whole blood banking.  
 
9.1.3.4 Days 45, 75, 120, 150, and 2 weeks after return of prednisone dose to baseline (± 3 
days) 
For convenience, for subjects outside of the Columbus area, these may be locally drawn and 
shipped to the NCH clinical laboratory.   
The following blood lab work will occur at this visit: 
• Aspartate aminotransferase (AST)/alanine transaminase (ALT) 
• Gamma-glutamyl transpeptidase (GGT)* 
• A Comprehensive Metabolic Panel (CMP) that includes: 
o Albumin/Serum total protein 
o Serum total bilirubin 
o Electrolytes 
o Calcium 
o Creatinine/Blood Urea Nitrogen (BUN) 
o Serum glucose 
o Alkaline phosphatase  
• Prothrombin time (PT)/activated partial thromboplastin time (PTT)/INR 
• Total IgG (Day 120, 150) 
• Direct Bilirubin 
* For subjects taking drugs that potentially induce GGT synthesis through the cytochrome P450 system, the 
clinically significant range will be estimated starting at two times the levels obtained from the baseline screening 
test and up. 
 
9.1.3.5 Day 60 (± 7 days) 
The following assessments will occur at this visit: 
• Medical history 
• Concomitant Medications 
• Adverse Events 
• Physical Exam 
• Vital Signs (heart rate, respiratory rate, temperature, weight, height (see Section 10.5 for 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
Sponsor-Investigator: Megan A. Waldrop, M.D. 
 
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more details) and blood pressure) 
 
The following blood lab work will occur at this visit: 
• Complete blood cell count (CBC) and differential, platelets 
• Aspartate aminotransferase (AST)/alanine transaminase (ALT) 
• Gamma-glutamyl transpeptidase (GGT)* 
• A Comprehensive Metabolic Panel (CMP) that includes: 
o Albumin/Serum total protein 
o Serum total bilirubin 
o Electrolytes 
o Calcium 
o Creatinine/Blood Urea Nitrogen (BUN) 
o Serum glucose 
o Alkaline phosphatase  
• Prothrombin time (PT)/activated partial thromboplastin time (PTT)/INR 
• Creatine kinase (CK) 
• Total IgG 
• Direct Bilirubin 
• ELISA for detection of total antibodies to AAV9 
• ELISpots for T-cell responses to AAV9 and Dystrophin 
• Whole blood for biobanking 
• Blood for vector shedding studies (see Section 10.4 for details) 
* For subjects taking drugs that potentially induce GGT synthesis through the cytochrome P450 system, the 
clinically significant range will be estimated starting at two times the levels obtained from the baseline screening 
test and up. 
 
The following urine lab work will occur at this visit: 
• Urinalysis 
• Urine for biobanking  
• Urine for vector shedding studies (see Section 10.4 for details) 
 
The following additional samples will be collected for vector shedding studies (see Section 10.4 
for details): 
• Saliva 
• Feces 
 
9.1.3.6 Day 90 (± 7 days) and 180 (± 14 days), and Months 12 (± 14 days), 18 (± 14 days), and 
24 (± 14 days) visits 
 
The following assessments will occur at these visits: 
• Medical history 
• Concomitant Medications 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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• Adverse Events 
• Physical Exam 
• Vital Signs (heart rate, respiratory rate, temperature, weight, and blood pressure, height 
(see Section 10.5 for more details)) 
• ECHO 
• ECG 
• Functional tests (variable depending on age of enrollment) 
o Enrollment age: ≤ 36 months 
▪ Bayley-IV scores (Bayley Scales of Infant and Toddler Development) 
o Enrollment age: > 36 months and < 5 years of age 
▪ North Star Ambulatory Assessment (NSAA) 
▪ Time to climb 4 stairs 
▪ Amount of time required to walk 100 meters (100MWT) 
▪  
o Enrollment age: ≥ 5 years old until Non-Ambulant 
▪ North Star Ambulatory Assessment (NSAA) score 
▪ Time to climb 4 stairs 
▪ Amount of time required to walk 100 meters (100MWT) 
▪ Force generated in knee flexion and extension in maximum voluntary 
isometric contraction (MVICT).   
• Pulmonary functional test (PFT) (see Section 10.3 for more details) 
The following blood lab work will occur at this visit: 
• Complete blood cell count (CBC) and differential, platelets 
• Aspartate aminotransferase (AST)/alanine transaminase (ALT) 
• Gamma-glutamyl transpeptidase (GGT)* 
• A Comprehensive Metabolic Panel (CMP) that includes: 
o Albumin/Serum total protein 
o Serum total bilirubin 
o Electrolytes 
o Calcium 
o Creatinine/Blood Urea Nitrogen (BUN) 
o Serum glucose 
o Alkaline phosphatase  
• Prothrombin time (PT)/activated partial thromboplastin time (PTT)/INR 
• Creatine kinase (CK) 
• Cystatin C 
• Total IgG 
• Direct Bilirubin 
• ELISA for detection of total antibodies to AAV9 
• ELISpots for T-cell responses to AAV9 and Dystrophin 
• Blood for vector shedding studies (see Section 10.4 for details) 
* For subjects taking drugs that potentially induce GGT synthesis through the cytochrome P450 system, the 
clinically significant range will be estimated starting at two times the levels obtained from the baseline screening 
test and up. 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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The following urine lab work will occur at this visit: 
• Urinalysis 
• Urine for biobanking 
• Urine for vector shedding studies (see Section 10.4 for details) 
 
The following additional samples will be collected for vector shedding studies (see Section 10.4 
for details): 
• Saliva 
• Feces 
 
At the Day 90 and 180 and the Month 12, either an open or needle muscle biopsy will occur 
(see Section 10.1 for more details).  Additionally, at the Day 180, and Months 12, 18 and 24 
month visits, MRI imaging will also be performed (see Section 10.2 for more details) 
 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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9.2 Schedule of Events 
Table 8: Schedule of Events 
 

Phase I/IIa Systemic Gene Delivery Clinical Trial of scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy 
 
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TABLE 8 FOOTNOTES
15. For children under 10 kg in weight, the blood draw schedule will be reviewed to limit blood draw volumes to ≤ 3% of total body weight over a 24 hour period and ≤ 10% of total body weight over a 30 day 
period. 
9. Depending on the age of subject, there may be sedation required for gene therapy and needle biopsies. This determination will be at the discretion of the PI.
7. Day 0 Vital Signs (Heart rate, respiratory rate, pulse oximetry (only if sedated), temperature, and blood pressure) will be measured as per Section 9.1.2.2. 
1. If there are changes to the study, parents or subjects will be re-consented or re-assented at their next visit.
8. Samples scheduled in between f/u visits will be collected locally and shipped to us. 
2. Visit will occur over multiple days. 
3. Muscle strength testing at baseline (days -45 to -1) and visits 8-10 will be performed at least 1 day preceding the biopsy. 
4. An open muscle biopsy will be peformed on one leg at the baseline, and on the other leg at 3 months post gene transfer. Needle muscle biopsy will be performed in either leg at 6 months and the 
contralateral at 12 months. Biopsies must be performed after the strength evaluation and lab tests. See section 12.1 for more details.
5. Prophylactic prednisolone/prednisone as well as Proton Pump Inhibitor (PPI) begins on Day -1 and is tapered according to AST, ALT, and ELISpot results starting at Day 30.  Anticipate that most subjects will be 
on prednisolone for up to 120 days.
6. Day 0 physical exam will consist of the anesthesia evaluation prior to sedation or by study team if sedation is not required.
10. The functional testing administered will be determind by the age of the subject.
11. In order to avoid the risk of sedation for an exploratory efficacy measure, muscle MRI will not be performed in any subjects who are unable to remain still for a MRI without sedation. To evaluate if a subject 
will be able to remain still for a MRI, a mock MRI will be given at the screening visit.
14. If the subject is too young to reliably rest these values, it will not be performed and will not be recorded as a protocol deviation. 
12. Vector shedding analysis post dosing will be performed on DNA isolated from different biological fluids (plasma, urine, saliva and feces) until three consecutive samples are negative for the presence of viral 
DNA for each specimen type.
13. Vector Shedding on day 0 will be collected at 4 and 8 hours post gene transfer.

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9.3 Muscle Strength Test Schedule 
Table 9: Muscle Strength Testing Schedule 
 
9.4 Long Term Follow up 
We will follow the most recent FDA guidelines with regard to long-term subject follow-up 
following gene transfer. As discussed and based on prior experiences with rAAV or transgene, 
there is a very low probability of gene transfer-related adverse events. We will, however, 
evaluate short-term safety over a two-year period that incorporates the active phase of the 
protocol. Subjects will then transfer to an annual monitoring program where data will be 
collected from annual standard of care visits for up to 5 years.  
 
If newly identified risks are associated with our product, or if the subjects suffer any adverse 
events during this period, we will initiate a long-term follow-up according to the FDA 
guidelines. We will, of course, notify CBER if there is any indication of need to extend follow-
up period. All subjects will be provided with written instructions on how to contact the Sponsor 
Investigator or study coordinator if they experience any serious adverse event that they consider 
possibly related to study treatment or study participation. This information will also be included 
in the Informed Consent document. All subjects will be instructed to notify the Sponsor 
Investigator of a change of address or contact information. 
 
The final results of the clinical trial will be shared with the participants at the completion of the 
study when all data has been collected, analyzed, and published. However, if significant findings 
become available that might increase the risk of the subjects or might affect their decision to 
remain in the study, then information will be made available as soon as it is available.  
 
At the time of death, no matter the cause, permission for an autopsy will be requested of their 
families. Subjects will be asked to advise their families of this request and of its scientific and 
medical importance. 
 
 
 

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10 PROCEDURE CRITERIA  
10.1 Muscle biopsy 
The baseline open muscle biopsy will be performed in one quadriceps muscle, or other 
appropriate muscle as determined by the SI.  Another open muscle biopsy of the quadriceps, or 
other muscle as determined by the SI (biopsy will be in the contralateral leg to the pre-treatment 
biopsy, unless otherwise determined by the SI), will be performed 3 months post-infusion.   
All open muscle biopsies will be performed under anesthesia, as per NCH standard clinical 
protocols. 
Needle muscle biopsies of the quadriceps muscles (using the Vacora device; C.R. Bard, Inc.) will 
be performed at 6 and 12 months.  In order to avoid resampling of the previous biopsy site, the 
open biopsies are anticipated to be performed in the vastus lateralis muscles, and the needle 
biopsies in the vastus medialis muscles.  Every effort will be made to sample these same muscles 
in all subjects; however, in subjects (particularly older ambulant subjects) with limited 
quadriceps musculature, alternate muscle biopsy sites may be selected at the SI’s discretion 
(including biceps, gastrocnemius, tibialis anterior or deltoid muscles).  
10.2 Magnetic Resonance Imaging (MRI) 
In order to avoid the risk of sedation for an exploratory efficacy measure, muscle MRI will not 
be performed in any subjects who are unable to remain still for a MRI without sedation. To 
evaluate if a subject will be able to remain still, a mock-MRI will be performed in coordination 
with Child Life during the screening visit. If the mock-MRI is unable to be completed, the 
subject will not be required to provide a clinical MRI during the trial. If the MRI is unable to be 
performed without sedation, it will not be recorded as a protocol deviation. 
10.3 Pulmonary functional tests (PFTs) 
PFT testing will be performed in subjects 5 years or older.  If the subject is too young to reliably 
test these values, it will not be performed and will not be recorded as a protocol deviation.  
10.4 Vector Shedding Studies 
Vector shedding analysis post dosing will be performed on DNA isolated from different 
biological fluids (plasma, urine, saliva and feces) until three consecutive samples are negative for 
the presence of viral DNA for each specimen type. 
10.5 Height Measurements 
Height will be measured with the patient standing when ambulatory. If the patient is non-
ambulatory, ulna length measurement will be used to calculate height. 
10.6 Concomitant Medications 
Prior and concomitant medications will be captured in the eCRF form two weeks prior to study 
enrollment through the last study visit. The SI will encourage participants to maintain the 
medication and supplements they are on at enrollment through the course of the study.  Subjects 
on aspirin or drugs that could affect coagulation will continue their medication as indicated.  

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Several investigations show that preoperative aspirin ingestion and intravenous heparin therapy 
can be administered safely without concerns about the risk of postoperative bleeding and should 
not lead to modification or cessation of such therapy.41-43 Subjects 4 years old or greater must 
already be on a stable dose of prednisone, prednisolone, or deflazacort for 12 weeks at time of 
enrollment as part of the inclusion criteria. 
10.6.1 Prophylactic Administration of Prednisone/Prednisolone  
 
All subjects will be discharged from the hospital on oral prednisone or prednisolone at 1 
mg/kg/day, which will be adjusted according to IFN-gamma T-cell studies in the following 
weeks. Subjects will also continue on a PPI until they return to their baseline corticosteroid dose 
or sooner based on the subject’s symptoms and discretion of the sponsor investigator and/or 
parent. 
If the 30 day post-infusion IFN-gamma T-cell responses to AAV9 and dystrophin show ≤50 SFC 
per 1 x 106 PBMCs, the subject’s prednisone or prednisolone dose will be decreased back to the 
standard clinical dose of 0.75 mg/kg/day. Those subjects who had been on deflazacort may be 
switched back to the standard clinical dose of 0.9 mg/kg day.  For either regimen, 
approximations of these target doses may be allowed to account for pill formulations (for 
prednisone and deflazacort), as in clinical practice.  If either the AST or ALT exceeds >2.5X the 
subject’s baseline values, and results are confirmed on a follow-up blood test, the prednisone or 
prednisolone regimen will be maintained at 1 mg/kg/day until the enzyme levels fall within a 
range, 2.5X the baseline value.  Subjects may be asked to undergo a weekly blood test outside of 
prescheduled visits should the SI deem it necessary to monitor the T-cell response or AST/ALT 
levels more closely. 
 
If the IFN-gamma T-cell responses show ≥50 SFC per 1 x 106 PBMCs, we may increase the dose 
to approximately 2 mg/kg/day depending on T-cell response measured by ELISpot assay and 
prolong a subsequent tapering protocol based on the individual subject’s immune response 
profile as assessed by subsequent ELISpot assays (as on the schedule above). Based upon other 
studies performed at NCH, we anticipate that oral prednisone or prednisolone may be 
administered for up to 120 days post gene transfer prior to the return to the standard clinical 
corticosteroid regimen of each subject. 
 
11 ASSESSMENT OF SAFETY 
The primary outcome for this clinical trial is safety.  
11.1 Adverse and Serious Adverse Event 
11.1.1 Definition of an Adverse Event 
As stated above this protocol will follow the final regulations issued by the Food and Drug 
Administration addressing the safety reporting requirements for investigational new drug 
applications (INDs) found in 21 CFR part 312 and for bioavailability and bioequivalence studies 
found in 21 CFR part 320. “Safety Reporting Requirements for INDs and BA/BE Studies”. 
 
http://www.fda.gov/downloads/Drugs/GuidanceComplianceRegulatoryInformation/Guidances/U
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Adverse Event (AE):  Adverse event means any untoward medical occurrence associated with 
the use of a drug in humans, whether or not considered drug related. 
Adverse events will be graded by the investigator accordingly (as indicated in Section 11.4):  
1 = mild 
2 = moderate 
3 = severe 
4 = life threatening or debilitating 
5 = fatal  
 
Association or relatedness to the study agent will be graded as follows:  
5 = unrelated 
4 = unlikely 
3 = possibly 
2 = probably 
1 = definitely related.  
 
Adverse reaction: An adverse reaction means any adverse event caused by a drug. Adverse 
reactions are a subset of all suspected adverse reactions for which there is reason to conclude that 
the drug caused the event. 
Suspected adverse reaction (21 CFR 312.32(a)) Suspected adverse reaction (SAR) means any 
adverse event for which there is a reasonable possibility that the drug caused the adverse event. 
For the purposes of IND safety reporting, ‘reasonable possibility’ means there is evidence to 
suggest a causal relationship between the drug and the adverse event. A suspected adverse 
reaction implies a lesser degree of certainty about causality than adverse reaction, which means 
any adverse event caused by a drug. 
11.1.2 Unexpected Adverse Events 
Unexpected adverse events are those which are not previously reported with recombinant AAV 
vectors, commonly not seen in association with the subject's underlying disease or with the 
procedures to be used in this study, or are related to a known toxicity but differ because of 
greater severity or specificity.   
Potential expected AEs include localized injection site reactions, which might also be related to 
the procedure and be independent of the AAV vector itself.  Asymptomatic elevations in 
transaminases are a feature of Duchenne muscular dystrophy, as they are related to release of 
AST and ALT from muscle tissue and correlate with levels of creatine kinase.  These are a 
feature of the disease itself.  Levels of AST (up to 12.3X the upper limit of normal) and ALT (up 
to 22.6X the upper limit of normal) are expected to be seen at baseline, and in the setting of 
normal GGT function are not indicative of muscle injury44.  They are thus not exclusionary for 
enrollment and will not be recorded as adverse events prior to gene transfer.   
Systemic delivery of AAV vectors have resulted in AEs including asymptomatic transient 
transaminitis (up to levels of 2.5x the subject’s baseline value and 3X the upper limit of normal 
for DMD subjects for AST and ALT) with preserved liver synthetic function and no significant 
elevation in GGT) following gene transfer (see Section 11.4.2). 

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11.1.3 Serious adverse event (SAE)  
An adverse event or suspected adverse reaction is considered “serious” if, in the view of the 
investigator/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. Examples of such 
medical events include allergic bronchospasm requiring intensive treatment in an emergency 
room or at home, blood dyscrasias or convulsions that do not result in inpatient hospitalization, 
or the development of drug dependency or drug abuse.  
 
To reiterate, an SAE is an event in categories 3, 4, and 5.   
 
Category 3:   Severe adverse event; inability to carry on normal activities; required professional 
medical attention 
Category 4:   Life-threatening or permanently disabling adverse event 
Category 5:   Fatal adverse event 
 
11.1.4 Life-threatening (21 CFR 312.32(a))  
An adverse event or suspected adverse reaction is considered “life-threatening” if, in the view of 
either the investigator or sponsor, its occurrence places the patient or subject at immediate risk of 
death. It does not include an adverse event or suspected adverse reaction that, had it occurred in a 
more severe form, might have caused death. 
The SI will fulfill the reporting responsibilities to FDA on behalf of Nationwide Children’s 
Hospital using the web-based Adverse Event reporting system (GeMCRIS).   
11.2 Relationship to Study Drug 
Association or relatedness to the study agent, study procedures and the subject's pre-existing 
disease will be graded as follows:  
 
1. Definitely related 
2. Probably related 
3. Possibly related 
4. Unlikely related 
5. Unrelated 
 
If the relationship between the AE/SAE and the investigational product is determined to be 
“possible” or “probable”, the event will be considered to be related to the investigational product 
for the purposes of expedited regulatory reporting. 
 
 

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11.3 Time Period and Frequency for Event Assessment and Follow-Up 
11.3.1 Obligations of the Investigator  
The Sponsor Investigator will submit an IND safety report to the FDA on any serious adverse 
event that is both unexpected and associated with the use of the gene transfer product (i.e., there 
is reasonable possibility that the event may have been caused by the use of the product; the 
investigator will not await definitive proof of association before reporting such events); as well 
as a written report on any finding from tests in laboratory animals that suggests a significant risk 
for human research participants including reports of mutagenicity, teratogenicity, or 
carcinogenicity.  The report will be clearly labeled as a “Safety Report” and will be submitted to 
the FDA and to the local Institutional Review Board within the timeframes set forth in Section 
11.3.2. 
The Sponsor Investigator will adhere to any other serious adverse event reporting requirements 
in accordance with federal regulations, state laws, and the local institutional policies and 
procedures, as applicable. 
The Sponsor Investigator will be responsible for ensuring that the reporting requirements are 
fulfilled and will be held accountable for any reporting lapses. 
 
11.3.2 Safety Reporting  
The investigator or his designee will report all serious and unexpected adverse events to the IRB, 
CBER/FDA and DSMB according to regulatory requirements described as follows: 
IRB: All Serious Adverse Events (SAEs) and Dose Limiting Toxicities (DLTs), regardless of 
expectedness, relatedness, or if they meet the definition for unexpected problems in the 
clinical trial will be reported to the IRB as soon as possible, but not later than 15 calendar 
days after the sponsor’s initial receipt of the information.  
DSMB: All Serious Adverse Events (SAEs) and Dose Limiting Toxicities (DLTs) that are life-
threatening will be reported to the DSMB within 48 hours of notification, regardless of 
expectedness, relatedness, or if they meet the definition for unexpected problems to the 
clinical trial. 
FDA: Any serious adverse event that is fatal or life-threatening, that is unexpected, and associated 
with the use of the gene transfer product will be reported by the study investigator to the 
FDA as soon as possible, but not later than 7 calendar days after the sponsor’s initial 
receipt of the information.  
Serious adverse events that are unexpected and associated with the use of the gene transfer 
product, but are not fatal or life-threatening, will be reported to the FDA as soon as 
possible, but not later than 15 calendar days after the sponsor’s initial receipt of the 
information.  
Changes in this schedule will be permitted only where, under the FDA IND regulations [21 CFR 
312(c) (3)], changes in this reporting schedule have been approved by the FDA and are reflected 
in the protocol. 
If, after further evaluation, an adverse event initially considered not to be associated with the use 
of the gene transfer product is subsequently determined to be associated, then the event will be 

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reported to the FDA as soon as possible, but in no case later than 15 calendar days after the 
determination is made. 
Relevant additional clinical and laboratory data will become available following the initial serious 
adverse event report. Relevant follow-up information to an IND safety report will be submitted 
concurrently to the FDA/IRB and the DSMB as soon as the information is available and will be 
identified as such, i.e., "Follow-up IND Safety Report.”   If a serious adverse event occurs after 
the end of a clinical trial and is determined to be associated with the use of the gene transfer 
product, that event will be reported concurrently to the FDA within 15 calendar days of the 
determination. 
Any finding from tests in laboratory animals that suggests a significant risk for human research 
participants including reports of mutagenicity, teratogenicity, or carcinogenicity will be reported 
to FDA and the DSMB as soon as possible, but not later than 15 calendar days after the 
sponsor’s initial receipt of the information. 
Should a serious adverse event deemed possibly, probably or definitely related to the study agent 
occur during administration, the study agent will be discontinued, appropriate treatment will be 
given under medical supervision and the subject will be examined as frequently as necessary 
thereafter until symptoms cease or stabilize. 
11.3.3 Safety Reporting: Content and Format   
The serious adverse event report will include, but need not be limited to: (1) the date of the 
event; (2) designation of the report as an initial report or a follow-up report, identification of all 
safety reports previously filed for the clinical protocol concerning a similar adverse event, and an 
analysis of the significance of the adverse event in light of previous similar reports; (3) clinical 
site; (4) the Sponsor Investigator; (5) FDA’s Investigational New Drug (IND) application 
number; (6) vector type , e.g., adeno-associated virus; (7) vector subtype, if relevant; (8) gene 
delivery method, e.g., in vivo transduction; (9) route of administration, e.g., intramuscular; (10) 
dosing schedule; (11) a complete description of the event; (12) relevant clinical observations; 
(13) relevant clinical history; (14) relevant tests that were or are planned to be conducted; (15) 
date of any treatment of the event; and (16) the suspected cause of the event.  These items will be 
reported through an FDA safety report by using the recommended Adverse Event Reporting 
Template available on NIH’s web site at: 
http://osp.od.nih.gov/sites/default/files/resources/Adverse_Event_Template_.docx 
11.3.4 Adverse Event Reporting from Primary Care Physician 
Close communication will be established with the primary care physician of all study 
participants and will be maintained throughout the study. The important hallmarks of the study 
along with the proposed reporting plan will be explained.  We will request that the primary care 
physician provide information regarding every routine visit and any intercurrent event taking 
place during the two years following gene transfer.   Laboratory reports, hospitalizations, clinical 
notes and any other relevant medical records will be requested at the time of their occurrence.   If 
non-routine visits are reported to us by the primary care physician, the study investigator will 
initiate an investigation to determine the possibility of an adverse event related to the gene 
transfer and will adhere to the adverse event reporting requirements in accordance with federal 
regulations, state laws, and the local institutional policies and procedures, as applicable. 
During the consent process, the study investigator will emphasize the importance of subject 
communication with our study team.  Any routine or non-routine doctor’s visits or medical care 

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received during the two years following gene transfer should be reported to the study team.  The 
study doctor will explain to the participant that copies of any relevant medical records of those 
visits will be requested from their medical care provider. 
11.3.5 Follow-up of Adverse Events 
All adverse events will be followed until resolution or stabilization. 
11.4 Dose limiting toxicity (DLT)  
Dose limiting toxicity is defined as any adverse event that is possibly, probably, or definitely 
related to the study agent, and is unexpected.  This would include any grade 3 event, according to 
the Common Terminology Criteria for Adverse Events (CTCAE) version 5.0; these 
classifications are outlined in Table 10.  
 
Note: Elevations in transaminases and transient alterations in WBCs, as outlined in Section 
12.4.2, above, are expected, and will not be considered as dose limiting when within the ranges 
noted therein.   
 
 
Table 10: Adverse Event Classification 
1 
Mild adverse event; did not require treatment 
2 
Moderate adverse event; resolved with treatment 
3 
Severe adverse event; inability to carry on normal activities; required professional 
medical attention 
4 
Life-threatening or permanently disabling adverse event 
5 
Fatal adverse event 
 
Study enrollment will be halted by the investigators when any subject experiences two or more 
Grade 3, or higher adverse event toxicity that are unexpected and possibly, probably, or 
definitely related to the study drug. The event will then be reviewed by the Data Safety 
Monitoring Board (DSMB) before making a determination to continue enrollment as described 
in Section 0. 
Laboratory tests with values within the clinically significant range will be repeated during the 
same visit whenever possible (refer to CTCAE v5.0 for a complete list of clinically significant 
range values). If the test result returns after the subject leaves the clinic, they will be immediately 
contacted.  For local residents they will be asked to return to the outpatient clinic for a repeat 
test.  For non-local residents, arrangements will be made to have the blood test redrawn in a 
laboratory close to home or by their primary care physician.  To avoid any confusion for the 
primary care physician, they will be informed (with permission from the subject) of their 
participation in the study at the time of gene transfer.  If the AE requires treatment, this will be 
carried out by the primary care physician or a doctor of choice selected by the subject.  We will 
obtain copies of repeat laboratory tests and any relevant medical records that will be added to the 
subject’s research chart.  
The SI will fulfill the reporting responsibilities under 21 CFR 312.32(c), to notify FDA in an 
IND safety report of potentially serious risks, as soon as possible, but no later than 15 calendar 
days after the investigator receives the safety information and determines that the information 

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qualifies for reporting. The investigator will confer with the DSMB and FDA before continuing 
to enroll. 
11.4.1 Expected Adverse Events Related to Disease Progression 
Subjects enrolled under this clinical protocol are expected to present clinically with adverse 
events related to natural progression of the disease. The draft guidance entitled “Duchenne 
Muscular Dystrophy Developing Drugs for Treatment over the Spectrum of Disease” 
(http://www.parentprojectmd.org/site/DocServer/Guidance_Document_Submission_-
_Duchenne_Muscular_Dystrop.pdf?docID=15283) provides the basis of expected disease-related 
adverse events. 
Adverse events determined to be due to the underlying disease progression will be recorded but 
will not be subject to the expedited reporting requirements outlined in Section 11.3.2. All AEs 
related to the disease and unrelated to the gene therapy administration will be reported annually to 
the FDA and IRB. 
11.4.2 Anticipated Adverse Event Lab Findings related to Intervention 
 
• Asymptomatic elevations in transaminases are a feature of Duchenne muscular 
dystrophy, as they are related to release of AST and ALT from muscle tissue and 
correlate with levels of creatine kinase.  These are a feature of the disease itself.  Levels 
of AST (up to 12.3X the upper limit of normal) and ALT (up to 22.6X the upper limit 
of normal) are expected to be seen at baseline, and in the setting of normal GGT function 
are not indicative of muscle injury44.  They are thus not exclusionary for enrollment, and 
will not be recorded as adverse events prior to gene transfer.   
Following gene transfer, transient transaminases (up to levels of 2.5x the subject’s 
baseline value) with preserved liver synthetic function and no significant elevation in 
GGT are anticipated, and will be recorded as Grade 1 adverse events, but not considered 
as adverse reaction in the presence of a normal GGT value. Mild decreases in leukocytes 
and lymphocyte counts within the first 30 days after gene transfer have been observed 
in other trials of AAV mediated gene transfer.  Such transient decreases in lymphocytes 
or leukocytes will be recorded as adverse events but considered anticipated.   
• Leukocytosis and neutrophilia during prednisone or prednisolone treatment: 
An elevation of up to 5000 cells/mm3 above the subject’s baseline value is expected in 
the neutrophil count (the primary granulocyte in circulation) and, consequently, in the 
leukocyte count within 5 hours of initiating prednisone or prednisolone therapy, based 
upon studies in healthy adult volunteers45. This occurs secondary to release of 
granulocytes from the marginated pool into the circulation accompanied by an increase 
in the size of the marginated granulocyte pool due to steroids.46 This steroid effect may 
be seen with increased steroid dosing.   
 
 

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11.5 Discontinuation Rules 
An independent Data Safety Monitoring Board (DSMB) will be established. Safety data will be 
monitored on a continual basis throughout the trial.  The DSMB can recommend early 
termination of the trial for reasons of safety.  Study enrollment will be halted by the investigators 
when any subject experiences two or more Grade 3, or higher adverse events that are 
unexpected and possibly, probably, or definitely related to the study drug.  This will include 
any subject death, important clinical laboratory finding, or any severe local complication in the 
injected area related to administration of the study agent.  If after review by the DSMB, the 
decision is made to continue, the study will proceed according to Section 6.4 of this protocol. 
 
The investigators will confer with the DSMB on all Grade 3 or higher adverse events that are 
possibly, probably, or definitely related to the study agent before continuing enrollment.  
12 CLINICAL MONITORING 
12.1 Data and Safety Monitoring Plan 
The Data and Safety Monitoring Board (DSMB) will act in an advisory capacity to review 
participant safety and study progress of Phase I/IIa Systemic Gene Delivery Clinical Trial of 
scAAV9.U7.ACCA for Exon 2 Duplication-Associated Duchenne muscular dystrophy.  
 
12.1.1 DSMB Membership 
The DSMB membership consists of persons completely independent of the investigator who have 
no financial, scientific, or other conflicts of interest with the trial.  Current or past collaborators or 
associates of Dr. Waldrop must note any conflict of interest before their eligibility to serve on the 
DSMB is approved.  
 
The DSMB will include experts in or representatives of the fields of:  
 
• Pediatric Neurology and Neuromuscular Diseases  
• Immunology  
• Gene Therapy  
• Hepatology 
• Muscular Dystrophy Clinical Care  
• Clinical Research and Clinical Trials 
 
Individuals invited to serve on the DSMB as either voting or non-voting members must disclose 
any potential conflicts of interest, whether real or perceived.  Conflicts of interest can include 
professional, proprietary, and miscellaneous interests as described in the NIH Grant Policy 
Statement and 45 CFR Part 94.  Potential conflicts that develop during a member’s tenure on a 
DSMB must also be disclosed.  Written documentation attesting to an absence of conflict of 
interest is required annually.  
12.1.2 DSMB Responsibilities 
Responsibilities of the DSMB are to:  

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• Review the research protocol, informed consent documents and plans for data and safety 
monitoring; 
• Evaluate the progress of the trial, including periodic assessments of data quality and 
timelines, participant recruitment, accrual and retention, participant risk versus benefit, 
trial site performance, and other factors that can affect study outcome; 
• Consider factors external to the study when relevant information becomes available, such 
as scientific or therapeutic developments that may have an impact on participant safety or 
the ethics of the trials; 
• Review study performance, make recommendations and assist in the resolution of 
problems reported by the Sponsor Investigator; 
• Protect the safety of the study participants; 
• Ensure the confidentiality of the trial data and the results of monitoring; and  
• Assist by commenting on any problems with study conduct, enrollment, and sample size 
and/or data collection. 
 
12.1.3 DSMB Reporting and Meetings 
Reports describing the status of the study will be prepared by the Sponsor Investigator’s staff and 
sent to the DSMB prior to a meeting, or at the DSMB’s request.   
 
A meeting (either by teleconference or webcast) with the DSMB will be scheduled prior to study 
initiation, after the Day 30 visit of the first subject and then approximately every 6 months, or at 
the DSMB’s request.  Reports will be submitted prior to a scheduled meeting for review by the 
DSMB.  
 
Reports will include the following: 
• 
A brief narrative of the study status, including the target enrollment, current and projected 
time to completing enrollment. Any significant events and/or difficulties should be briefly 
described in this narrative. 
• 
A brief narrative for each participant describing gender, age, race and ethnicity and other 
relevant demographic characteristics. The narrative for each participant should briefly 
describe his/her study status (i.e., dose level, visit number, adverse event information); 
• 
A timeline outlining the study progress relative to visit number for each participant, as 
well as time points for each SAE/Dose Limiting Toxicity. A total for Adverse Events 
(AEs) for each participant should be included.  
• 
A summary of AEs by severity levels; 
• 
A listing of AE details grouped by participant; 
• 
A listing of SAE details grouped by participant; 
• 
A listing of deaths 
• 
A summary of clinically significant laboratory test results 
 
 
 

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12.2 Institutional Review Board Monitoring 
The Sponsor Investigator must obtain IRB approval for the investigation.  Initial IRB approval, 
and all materials approved by the IRB for this study including the patient consent form and 
recruitment materials must be maintained by the Investigator and made available for inspection. 
Amendments will be submitted to the Nationwide Children’s Hospital IRB for their review and 
approval prior to implementation.  When an amendment to a protocol substantially alters the 
study design or increases potential risk to the study subject, the Informed Consent form will be 
revised and if applicable, subject's consent to continue participation will again be obtained. 
 
13 STATISTICAL CONSIDERATIONS 
For the secondary outcome measure of dystrophin expression, quantification of the number of 
fibers expressing dystrophin (determined by quantification of the number of fibers), 
measurement of the intensity of dystrophin staining at the sarcolemma (based on quantitative 
image analysis), and the degree of exon 2 skipping (as determined by RT-PCR) will be 
performed for each patient at each time point in a blinded fashion, and intra-patient comparisons 
of pre- and post-treatment values will be analyzed using repeated measures ANOVA with a 
Dunnett correction for multiple comparisons vs. baseline. For detecting change, we will have 
80% power to detect a very large effect size (SMD=1.4-3.3, for sample sizes from 3-6). 
 
14 SOURCE DOCUMENTS AND ACCESS TO SOURCE 
DATA/DOCUMENTS 
14.1 Data Management and Study Forms 
 
An outside contracted monitor of the study called a “CRO” will also monitor the study on a 
regular basis to make sure the study is conducted in compliance with all regulatory aspects of the 
protocol. 
 
All data and observations will be documented on electronic Case Report Forms (CRF) by source 
documentation using the Medrio Electronic Data Capture designed for the study.  A Safety 
Monitor will have access to the data to monitor adherence to the protocol and to applicable FDA 
regulations, and the maintenance of adequate and accurate clinical records.  An electronic Case 
Report Form will be completed for every subject that was registered for participation in the study.  
The Case Report Form will be reviewed in detail.  Case Report Forms will be completed as 
information becomes available. 
 
Case Report Forms will be reviewed in detail by the Safety Monitor in a regular basis for which 
the Safety Monitor will have access to subject medical records, laboratory data, and other source 
documentation.  Safety monitor will make a decision as to the data acceptability.  If errors or 
omissions are found in the course of a data audit, or if clarification of data is required, the 
electronic Case Report Form(s) in question will be corrected by the SI or their designee.  Data 
Resolution may be generated on omissions or clarifications, to be completed, electronically 
signed and dated, and maintained as a part of the eCRF.  The SI will sign and accept the indicated 

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electronic Case Report Form.  This signature will indicate that thorough inspection of the data 
therein has been made and will thereby certify the contents of the form.   
14.2 Electronic Case Report Forms 
Adequate and accurate case records will be maintained and all relevant observations and data 
related to the study will be recorded.  This will include medical history, physical examination, 
concomitant medications, adverse events, vitals, MRI, ECHO, ECG, muscle strength tests, PFTs, 
muscle biopsies, hematology, clinical chemistry, serology, coagulation, and immunologic results.  
 
Electronic CRFs will be used in this study.  The eCRF will be electronically signed and dated by 
the Sponsor Investigator or his/her designee after his/her review.  After the completion of the 
study, completed eCRFs will be retained in the archives. 
 
Completed eCRFs will be reviewed by the study monitor against the source documentation for 
accuracy and completeness.   
 
14.3 Inspection of Records 
The Investigator agrees to allow the monitor to inspect the drug storage area, study drug stocks, 
drug accountability records, subject charts, study source documents, and other records relative to 
study conduct. 
 
15 QUALITY ASSURANCE AND QUALITY CONTROL 
15.1 Study Monitoring 
The study will be monitored in compliance with the relevant parts of 21 CFR and according to 
the ICH GCP Guidelines.  
The procedures outlined in the protocol and case report forms will be carefully reviewed by the SI 
and staff prior to study initiation to ensure appropriate interpretation and implementation.  No 
deviations from the protocol shall be made except in emergency situations where alternative 
treatment is necessary for the protection, proper care and well-being of subjects. 
 
During the study, a monitor will have regular contact with the investigational site for the 
following: 
 
• Provide information and support to the investigator(s); 
• Confirm that facilities remain acceptable; 
• Confirm that the investigational team is adhering to the protocol and ICH/GCP 
guidelines, that data are being accurately recorded in the case report forms, and that 
investigational product accountability checks are being performed; 
• Perform source data verification. This includes comparison of the data in the case report 
forms with the patient’s medical records at the hospital or practice, and other records 
relevant to the study. This will require direct access to all original records for each patient 

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(e.g. clinic charts) including access to the site’s Electronic Medical Record (EMR) and 
any videos captured during specified visit assessments [edit as applicable to study]; 
• Records and report any protocol deviations not previously reported; 
• Confirms AEs and SAEs have been properly documents on eCRFs and confirm those 
SAEs that met criteria for reporting have been forwarded to the IRB. 
 
The monitor will be available between visits if the investigator(s) or other staff needs 
information or advice. 
 
16 ETHICS/PROTECTION OF HUMAN SUBJECTS 
16.1 Ethics Review 
The final study protocol, including the final version of the Informed Consent Form, must be 
approved or given a favorable opinion in writing by an IRB or IEC as appropriate. 
The Sponsor Investigator is responsible for informing the IRB of any amendment to the protocol 
in accordance with Nationwide Children’s Hospital requirements.  The protocol must be re-
approved by the IRB upon receipt of amendments and annually, as required by Nationwide 
Children’s Hospital. When an amendment to a protocol substantially alters the study design or 
increases potential risk to the study subject, the Informed Consent form will be revised and if 
applicable, subject's consent to continue participation will again be obtained. 
16.2 Ethical Conduct of the Study 
The study will be performed in accordance with ethical principles that have their origin in the 
Declaration of Helsinki and are consistent with ICH/Good Clinical Practice, and applicable 
regulatory requirements. 
16.3 Written Informed Consent 
Legally effective and properly executed written informed consent, in compliance with 21 CFR 
50 and the International Conference on Harmonization (ICH) guidelines, will be obtained from 
each subject before the subject is entered into the trial or before any unusual or non-routine 
procedure is performed that involves risk to the subject. The Sponsor Investigator(s) will ensure 
that the patient is given full and adequate oral and written information about the nature, purpose, 
possible risk and benefit of the study.  Subjects must also be notified that they are free to 
discontinue from the study at any time.  The patient should be given the opportunity to ask 
questions and allowed time to consider the information provided. 
 
Attention will be directed to the basic elements that are required for incorporation into the 
informed consent under US Federal Regulations for Protection of Human Subjects [21CFR 
50.25(a)]. If new information related to the study arises, subjects will be asked to sign a revised 
document. Signed consent forms will remain in each subject's research chart and be available for 
the verification by study monitors at any time. For this study, only the Sponsor Investigator will 
be obtaining initial consent. Reconsenting due to changes in the Informed Consent may be done 
by other study staff as delegated on the Delegation of Authority Log. The Sponsor 
Investigator(s) must maintain the original, signed Informed Consent Form. Subjects will be given 
a signed, dated copy of their consent form documents. 

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Per the National Institutes of Health’s Guidelines for Research Involving Recombinant or 
Synthetic Nucleic Acid Molecules (https://osp.od.nih.gov), an autopsy must be requested should 
a study participant die following participation in a gene transfer trial, no matter what the cause of 
death.  An autopsy is requested to obtain vital information about the safety and efficacy of gene 
transfer.  Subjects’ parent(s)/legal guardians will be asked to provide consent for an autopsy in 
advance of any death.  Consent is requested at the beginning of the study to relieve the burden of 
making such a decision at the time of death should such an event occur. 
 
17 DATA HANDLING AND RECORD KEEPING 
17.1 Retention of Records 
All primary data that are a result of the original observations and activities of the study and that 
are necessary for the reconstruction and evaluation of any study report will be retained in a 
secure archive at the study site for a period not less than 2 years after the last approval of a 
marketing application in an ICH region, and until there are no pending or contemplated 
marketing applications in an ICH region, or until at least 2 years have lapsed since the formal 
discontinuation of the clinical development of the investigational product. 
The site will maintain a Regulatory Binder.  In this binder, there will be tabbed sections for study 
documents including the following: study personnel identification and signature list, subject 
screening records, subject roster (names omitted), protocol and amendments or administrative 
changes, FDA Form 1572 (if required), study staff Curricula Vitae, IRB documentation, an 
approved sample ICF, correspondence, site monitoring reports, blank Data Documentation form, 
and lab accreditations and normal values.  The site must keep this binder current and available 
for review by the IRB and/or FDA. 
17.2 Retention of Samples 
The identified storage laboratory will be responsible for arranging storage of any remaining or 
unused biological samples as well as properly documenting the storage procedures, once all 
study-required analyses are complete, as per sample processing requirements until such time that 
the IND holder provides external storage vendor transfer or destruction instructions. 
 
 

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17.3 Study Reports 
17.3.1 Annual Study Reports 
Within 60 days after the one-year anniversary of the date on which the investigational new drug 
(IND) application went into effect, and after each subsequent anniversary until the trial is 
completed, the Sponsor Investigator will submit information set forth as follows:  
a) Clinical Trial Information.  This will be a brief summary of the status of the trial in 
progress or completed during the previous year.  The summary will include the following 
information for the trial: (1) the title and purpose of the trial; (2) clinical site; (3) the 
Sponsor Investigator; (4) clinical protocol identifiers, including the IRB and IBCSC 
protocol numbers, and the FDA IND application number; (5) participant population (such 
as disease indication and general age group); (6) the total number of participants planned 
for inclusion in the trial; the number entered into the trial to date; the number whose 
participation in the trial was completed; and the number who dropped out of the trial with 
a brief description of the reasons; (7) the status of the trial, e.g., open to accrual of 
subjects, closed but data collection ongoing, or fully completed, and (8) if the trial has 
been completed, a brief description of any study results. 
 
b) Progress Report and Data Analysis.  Information obtained during the previous year's 
clinical and non-clinical investigations, including:  (1) a narrative or tabular summary 
showing the most frequent and most serious adverse experiences by body system; (2) a 
summary of all serious adverse events submitted during the past year; (3) a summary of 
serious adverse events that were expected or considered to have causes not associated 
with the use of the gene transfer product such as disease progression or concurrent 
medications; (4) if any deaths have occurred, the number of participants who died during 
participation in the investigation and causes of death; and (5) a brief description of any 
information obtained that is pertinent to an understanding of the gene transfer product’s 
action, including, for example, information about dose-response, information from 
controlled trials, and information about bioavailability.  
 
c) A copy of the updated clinical protocol including a technical and non-technical abstract. 
 
 

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