A Phase I/II, Open Label, Multicentre, Ascending Single Dose, Safety Study of a Novel Adeno- Associated Viral Vector (FLT180a) in Patients With Haemophilia B
Phase: PHASE1
Conditions: Hemophilia B
Sponsor: University College, London

 
PRS record reference: NCT03369444  
 
Trial Title: A Phase I/II, Open label, Multicentre, Ascending Single Dose, Safety 
Study of a Novel Adenoassociated Viral Vector (FLT180a) in Patients With 
Haemophilia B  
 
This document contains the following study related documents:  
15/0552 - Protocol Version 8.1, 21 Apr 2020 (final) 

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
Sponsor Code: 15/0552 
Page 1 of 118 
 
Version 8.1 
CONFIDENTIAL 
21 Apr 2020 
 
 
 
Full title of trial: 
 
A Phase I/II, Open label, Multicentre, Ascending 
Single Dose, Safety Study of a Novel Adeno-
associated Viral Vector (FLT180a) in Patients With 
Haemophilia B 
 
Short title: 
 
A Factor IX Gene Therapy Study (FIX-GT) 
 
Version and date of protocol: 
Previous Versions: 
Final Version 8.1 21 Apr 2020 
Version 1.0 12 Apr 2017 
Version 2.0 26 Sep 2017 
Version 3.0 14 May 2018 
Version 3.1 22 May 2018 
Version 4.0 21 Nov 2018 
Version 5.0 12 Mar 2019 
Version 6.0 30 Apr 2019 
Version 7.0 19 Jul 2019 
Version 7.1 07 Aug 2019 
Version 8.0 29 Feb 2020 
Sponsor: 
Sponsor protocol number: 
University College London (UCL) 
15/0552 
 
Funder: 
Funder Reference: 
Freeline Therapeutics Ltd. 
FLT180a-01 
 
EudraCT no: 
IND #: 
Active IMP 
Phase of Trial: 
Site(s): 
2017-000852-24  
018311 
FLT180a  
Phase I/II 
International multicentre study 
 
Chief Investigator: 
Pratima Chowdary 
The Katharine Dormandy Haemophilia and Thrombosis Centre 
Royal Free Hampstead NHS Trust 
Royal Free Hospital, Pond Street  
Hampstead, London NW3 2QG 
Sponsor Representative: 
Joint Research Office 
UCL 
Gower Street 
London WC1E 6BT 
 
 
 
 
 

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
Sponsor Code: 15/0552 
Page 3 of 118 
 
Version 8.1 
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1 
Emergency Contact Information 
In the event of a serious adverse event (SAE), the investigator must complete the Clinical Trial 
SAE Form and ensure its transmission within 24 hours to the contract research organisation 
(CRO) pharmacovigilance department. Applicable fax numbers and e-mail addresses can be 
found on the SAE form (sent under separate cover).  
For emergency protocol- or safety-related issues, the investigator must contact the CRO 
Medical Monitor: 
Or if unavailable, contact the central out-of-hours group at

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
Sponsor Code: 15/0552 
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Contents 
1 
Emergency Contact Information .................................................................................... 3 
2 
List of Abbreviations .................................................................................................... 10 
3 
Trial Personnel ............................................................................................................ 13 
4 
Summary ..................................................................................................................... 14 
5 
Introduction ................................................................................................................. 28 
5.1 
Background ................................................................................................. 28 
5.2 
Nonclinical Data .......................................................................................... 29 
5.3 
Clinical Data ................................................................................................ 29 
5.4 
Rationale and Risks/Benefits ....................................................................... 30 
5.4.1 
Rationale ................................................................................................. 30 
5.4.2 
Potential Risks and Benefits .................................................................... 34 
5.4.3 
Assessment and Management of Risk ..................................................... 40 
6 
Objectives ................................................................................................................... 40 
6.1 
Primary........................................................................................................ 40 
6.2 
Secondary ................................................................................................... 40 
6.3 
Exploratory .................................................................................................. 40 
7 
Trial design ................................................................................................................. 41 
7.1 
Overall Design ............................................................................................. 41 
7.2 
Endpoints .................................................................................................... 43 
7.2.1 
Primary Endpoints ................................................................................... 43 
7.2.2 
Secondary Endpoints ............................................................................... 43 
7.2.3 
Exploratory Endpoints .............................................................................. 44 
8 
Selection of Patients ................................................................................................... 45 
8.1 
Inclusion Criteria ......................................................................................... 45 
8.2 
Exclusion Criteria ........................................................................................ 45 
9 
Investigational Medicinal Products and Non-Investigational Medicinal Products.......... 46 
9.1 
Name and Description of ATIMP(s) and IMP(s) ........................................... 46 
9.2 
Source of ATIMP/IMP, Manufacture, Distribution and Storage .................... 47 
9.2.1 
Manufacture............................................................................................. 47 

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9.2.2 
Manufacture and Shipment ...................................................................... 47 
9.2.3 
Importation into the UK, QP Certification and Storage Prior to Shipment to 
Investigational Site ..................................................................................................... 48 
9.2.4 
Shipment of IMP to Infusion Sites ............................................................ 48 
9.2.5 
Infusion Sites ........................................................................................... 48 
9.2.6 
Receipt Storage and Handling of ATIMP(s)/IMP(s) at Site ....................... 48 
9.2.7 
Accountability and Traceability of ATIMP(s)/IMP(s) ................................. 49 
9.3 
Name and Description of Non-Investigational Medicinal Product ................. 49 
9.4 
Implementation of the Immunosuppressants Regimen ................................ 49 
10 
Concomitant Medication .............................................................................................. 51 
10.1 
Prior Treatment ........................................................................................... 51 
10.2 
Concomitant Treatment ............................................................................... 51 
10.3 
Withdrawal From Existing FIX Concentrate Therapy ................................... 52 
11 
Study Procedures ........................................................................................................ 52 
11.1 
Study Schedule ........................................................................................... 52 
11.1.1 
Screening ................................................................................................ 52 
11.1.2 
Vector Infusion (Day -1 to Day +4) ........................................................... 54 
11.2 
Follow-up Visits (Weeks 1-25) ..................................................................... 57 
11.2.1 
Week 1-5 Assessments ........................................................................... 57 
11.2.2 
Week 6-12 Assessments ......................................................................... 58 
11.2.3 
Week 13 Assessments ............................................................................ 59 
11.2.4 
Week 14 Assessments ............................................................................ 59 
11.2.5 
Week 15 Assessments ............................................................................ 60 
11.2.6 
Week 16 Assessments ............................................................................ 60 
11.2.7 
Week 17-19 Assessments ....................................................................... 61 
11.2.8 
Week 20 Assessments ............................................................................ 62 
11.2.9 
Week 21-25 Assessments ....................................................................... 63 
11.2.10 
Frequent Local Laboratory Assessments ................................................. 63 
11.3 
Week 26/End-of-Study Visit ......................................................................... 63 
11.4 
Study Procedures/Evaluations .................................................................... 65 
11.4.1 
Demographic and Baseline Assessments ................................................ 65 

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11.4.2 
Efficacy .................................................................................................... 66 
11.4.3 
Safety ...................................................................................................... 68 
11.4.4 
Vector Shedding ...................................................................................... 70 
11.4.5 
Health Economic Assessments................................................................ 70 
11.4.6 
Exploratory .............................................................................................. 71 
11.5 
Volume of Blood to be Drawn From Each Patient ........................................ 72 
11.6 
Discontinuation/Withdrawal of Participants .................................................. 74 
11.6.1 
Withdrawal of Consent to Data Collection ................................................ 74 
11.6.2 
Loss to Follow-up..................................................................................... 74 
11.7 
Definition of End of Trial .............................................................................. 74 
12 
Dose Progression, Dose Escalation and Stopping Rules ............................................ 75 
12.1 
Oversight Committees ................................................................................. 75 
12.1.1 
Trial Management Group (TMG) .............................................................. 75 
12.1.2 
Independent Data Monitoring Committee (DMC) ..................................... 75 
12.1.3 
Trial Steering Committee (TSC) ............................................................... 75 
12.2 
Dose-Limiting Toxicity ................................................................................. 75 
12.3 
Dose Escalation .......................................................................................... 76 
12.3.1 
Dose Interval Between Patients ............................................................... 76 
12.3.2 
Rules Applicable to Evaluation of Safety in Each Dose Cohort ................ 76 
12.3.3 
Rules Applicable to Assessment of FIX Activity in Each Dose Cohort ...... 77 
12.4 
Temporary Halt ........................................................................................... 79 
12.5 
Stopping Rules ............................................................................................ 79 
13 
Recording and Reporting of Adverse Events ............................................................... 79 
13.1 
Definitions ................................................................................................... 80 
13.2 
Assessments of Adverse Events ................................................................. 81 
13.2.1 
Severity ................................................................................................... 81 
13.2.2 
Causality .................................................................................................. 81 
13.2.3 
Expectedness .......................................................................................... 82 
13.2.4 
Seriousness ............................................................................................. 82 
13.2.5 
Serious Adverse Event Reporting Procedures ......................................... 82 

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13.2.6 
Serious Adverse Event Collection Timeframe .......................................... 82 
13.2.7 
Serious Adverse Event Onset and Resolution Dates ............................... 82 
13.2.8 
Fatal Outcome ......................................................................................... 83 
13.2.9 
Important Medical Events ........................................................................ 83 
13.2.10 
Pregnancy ............................................................................................... 83 
13.2.11 
Regulatory Agency, Ethics Committee, and Site Reporting ...................... 84 
13.2.12 
Expedited Reporting of Other Safety Events ............................................ 84 
13.2.13 
Reporting Urgent Safety Measures .......................................................... 84 
14 
Statistical Considerations ............................................................................................ 85 
14.1 
Statistical Analysis Plan .............................................................................. 85 
14.2 
Procedures for Reporting Any Deviation(s) From the Original Statistical 
Plan………. ................................................................................................................ 85 
14.3 
Number of Patients ..................................................................................... 85 
14.3.1 
Choice of Sample Size ............................................................................ 85 
14.4 
Demographic and Baseline Characteristics ................................................. 85 
14.5 
Study Populations ....................................................................................... 86 
14.5.1 
Analysis Population ................................................................................. 86 
14.5.2 
The Per-protocol Population .................................................................... 86 
14.6 
Primary Endpoint ......................................................................................... 86 
14.6.1 
Safety ...................................................................................................... 86 
14.6.2 
Efficacy .................................................................................................... 86 
14.7 
Secondary Endpoints .................................................................................. 87 
14.7.1 
Safety ...................................................................................................... 87 
14.7.2 
Endogenous FIX Production .................................................................... 87 
14.7.3 
Haemostatic Effectiveness ....................................................................... 87 
14.7.4 
Shedding ................................................................................................. 88 
14.7.5 
Immune Response ................................................................................... 88 
14.8 
Exploratory Endpoints ................................................................................. 88 
14.8.1 
Haemostatic Effectiveness ....................................................................... 88 
14.8.2 
Immune Response ................................................................................... 88 
14.8.3 
Health Economics .................................................................................... 89 

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14.9 
Subgroup Analysis ...................................................................................... 89 
14.10 
Handling of Missing Data ............................................................................ 89 
14.11 
Protocol Deviations ..................................................................................... 90 
15 
Sponsor and Investigator Responsibilities ................................................................... 90 
15.1 
Sponsor’s Responsibilities........................................................................... 90 
15.1.1 
Good Clinical Practice Compliance .......................................................... 90 
15.1.2 
Monitoring Requirement for the Trial ........................................................ 91 
15.1.3 
Insurance ................................................................................................. 91 
15.1.4 
Public Posting of Study Information ......................................................... 92 
15.1.5 
Submission of Summary of Clinical Study Report to Competent Authorities and 
Ethics Committees ...................................................................................................... 92 
15.1.6 
Study Suspension, Termination, and Completion .................................... 92 
15.1.7 
Notification of Serious Breaches to Good Clinical Practice and/or the 
Protocol……. .............................................................................................................. 92 
15.2 
Investigator’s Responsibilities ..................................................................... 93 
15.2.1 
Good Clinical Practice Compliance .......................................................... 93 
15.2.2 
Protocol Adherence and Investigator Agreement ..................................... 93 
15.2.3 
Data Collection Tools and Source Document Identification ...................... 93 
15.2.4 
Completing Case Report Forms ............................................................... 94 
15.2.5 
Data Handling and Analysis ..................................................................... 94 
15.2.6 
Monitoring ................................................................................................ 94 
15.2.7 
Audit/Inspection ....................................................................................... 94 
15.3 
Ethical and Regulatory Considerations ........................................................ 94 
15.3.1 
Informed Consent .................................................................................... 94 
15.3.2 
Institutional Review Board or Ethics Committee ....................................... 95 
15.3.3 
Protocol Amendments ............................................................................. 95 
15.4 
Record Keeping and Archiving .................................................................... 95 
15.4.1 
Archiving Essential Trial Documentation Relating to Traceability ............. 95 
15.5 
Privacy and Confidentiality .......................................................................... 97 
16 
Publication Policy ........................................................................................................ 97 
17 
Funding ....................................................................................................................... 97 

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
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18 
Statement of Compliance ............................................................................................ 98 
19 
References .................................................................................................................. 99 
Appendix 1: Treatment Management Plans ....................................................................... 102 
1.1 
Management of Thrombosis ...................................................................... 102 
1.2 
Management of Anaphylaxis ..................................................................... 102 
1.3 
Management of immune complex disease ................................................ 102 
1.4 
Management of Inhibitors .......................................................................... 102 
1.5 
Management of Infusion Reaction ............................................................. 103 
Appendix 2: Management Guidelines for CMV Reactivation .............................................. 104 
Appendix 3: Management Guidelines for Tacrolimus Dosing ............................................. 106 
Appendix 4: Summary of Protocol Changes ....................................................................... 109 
 
 
 

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
Sponsor Code: 15/0552 
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2 
List of Abbreviations 
AAV 
Adeno-associated virus 
AE 
Adverse Event 
AR 
Adverse Reaction 
ALT 
Alanine aminotransferase 
AST 
Aspartate aminotransferase  
ATIMP 
Advance Therapy Investigational Medicinal Product 
CBC 
Clinical Biotechnology Centre 
CMV 
Cytomegalovirus 
CRO 
Contract Research Organisation 
CRP 
C-reactive protein 
DLT 
Dose-limiting toxicity 
DMC 
Data monitoring committee 
EC 
Ethics Committee 
eCRF 
Electronic case report form 
ECG 
Electrocardiogram 
EOS 
End of Study 
EU 
European Union 
EudraCT 
European Clinical Trials Database 
GCP 
Good Clinical Practice 
GFR 
Glomerular filtration rate 
GGT 
Gamma-glutamyl transpeptidase 
GMP 
Good Manufacturing Practice  
GTMP 
Gene Therapy Medicinal Product 
FIX 
Factor IX 
HAL 
Haemophilia Activities List 
HB 
Haemophilia B 
HBV 
Hepatitis B virus 
HBsAg 
Hepatitis B surface antigen 
HCV 
Hepatitis C virus 

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hFIX 
Human Factor IX 
HIV 
Human immunodeficiency virus 
HJHS 
Haemophilia Joint Health Score 
 
 
ICF 
Informed consent form 
ICH 
International Conference on Harmonisation 
IMP 
Investigational medicinal product 
IRB 
Institutional Review Board 
IU 
International unit 
IV 
Intravenous(ly) 
LFT(s) 
Liver function test(s) 
MRI 
Magnetic Resonance Imaging 
NCI CTCAE 
National Cancer Institute Common Terminology Criteria for 
Adverse Events 
NIMP 
Non-investigational medicinal product 
PCR 
Polymerase chain reaction 
PI 
Principal Investigator 
PROBE 
Patient Reported Outcomes, Burdens and Experiences 
QoL 
Quality of life 
QP 
Qualified person (for release of trial drug) 
rcAAV 
Replication competent adeno-associated virus 
SAE 
Serious adverse event 
SAR 
Serious adverse reaction 
SAP 
Statistical Analysis Plan 
SUSAR 
Suspected unexpected serious adverse reaction  
TAT 
Thrombin-antithrombin complex  
TEAE 
Treatment Emergent Adverse Event 
TMG 
Trial Management Group 
TSC 
Trial Steering Committee 
UCL 
University College London 
US 
United States 

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vg 
Vector genomes 
WFH 
World Federation of Hemophilia 
WHODAS 
World Health Organisation Disability Assessment Schedule 

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
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3 
Trial Personnel 
Chief Investigator 
Pratima Chowdary 
 
 
 
Sponsor’s representative 
UCL Joint Research Office 
 
 
 
CRO Project Manager 
Syneos Health, LLC 
 

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4 
Summary 
Protocol number: 15/0552 
Drug: FLT180a (AAV2/S3-
-Ti-FIXco1)  
Title of the study: A Phase I/II, Open label, Multicentre, Ascending Single Dose, Safety Study of a Novel Adeno-
associated Viral Vector (FLT180a) in Patients with Haemophilia B  
Number of patients: 
Up to 24 patients may be enrolled into the study; the actual number of patients will depend on the criteria for 
dose escalation.  
14 patients will be enrolled at the terminal dose level.  
Investigator(s): This is a multicentre study. 
Site(s) and region(s):  
The study will be conducted at sites across Europe, Africa and North America. 
Clinical phase: I/II 
Objectives: 
Primary:  
Safety 
To assess the safety of systemic administration of FLT180a in adults with haemophilia B (HB) at up to 3 different 
dose cohorts. 
Efficacy 
To assess Factor IX (FIX) levels following systemic administration of FLT180a, at the terminal dose level. 
Secondary:  
• To investigate the endogenous production of FIX following systemic administration of FLT180a at up to 3 
different dose cohorts. 
• To investigate the effectiveness of a single administration of FLT180a on annualised bleeding rate and 
exogenous FIX consumption. 
• To assess the immune response to the FIX transgene product following systemic administration of FLT180a. 
• To assess viral shedding in various body fluids after systemic administration of FLT180a. 
Exploratory: 
• To assess the immune response to the adeno-associated virus (AAV)-S3 capsid proteins following systemic 
administration of FLT180a. 
Following a single administration of FLT180a: 
• To investigate the impact of endogenous production of FIX on functional status and disability in HB. 
• To investigate the impact of endogenous production of FIX on quality of life (QoL) in HB. 
• To investigate the impact of endogenous production of FIX on physical activity in HB. 
• To investigate the impact of endogenous production of FIX on haemophilia health status in HB. 
• To investigate the impact of endogenous production of FIX on joint health in HB. 
• To investigate the impact of endogenous production of FIX on health resource utilisation in HB. 
Rationale: 
Haemophilia B is an X-linked recessive bleeding disorder that affects approximately 1 in 30,000 males. The 
disorder results from a defect in the f9 gene and is characterised by frequent spontaneous bleeding into joints 
and soft tissues which, without adequate treatment, causes a chronic debilitating arthropathy. 
Current treatment for HB in the Western World involves intravenous (IV) infusions of plasma-derived or 
recombinant factor concentrates at the time of a bleed (on-demand therapy) or as regular prophylactic infusion 
of factor concentrate. On-demand treatment is effective at arresting haemorrhage but cannot prevent chronic 
damage that ensues after a bleed. In severely affected patients, prophylactic therapy to prevent bleeding 
episodes can greatly reduce such bleeding when plasma FIX levels are maintained at or above only 1% of 
normal.1 The relatively short half-life of standard FIX necessitates frequent IV administration of factor 
concentrates (2-3 times a week) for prophylaxis against haemorrhage, which is invasive, inconvenient, and 
 
1 Ljung RC. Prophylactic infusion regimens in the management of hemophilia. Thromb.Haemost. 1999; 82:525-530. 

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problematic for children. Even with extended half-life (EHL) factor concentrates frequent infusions are required 
for the life time of the patient. 
Continuous synthesis of FIX by the host cells after a single administration of gene therapy offers a real 
opportunity to prevent bleeding episodes and eliminate the need for regular infusions. In addition, current 
experience suggests that the level of efficacy seen with gene therapy is likely to be superior to that seen with 
prophylactic treatment and, when considering the most recent data, may even offer the prospect of a functional 
cure with a lack of haemorrhage even in response to trauma or surgery. 
This study aims to characterise the safety of a new gene therapy (FLT180a) and investigate its potential to 
alter the disease phenotype from severe to mild/normal through the endogenous production of FIX following a 
single administration of vector. 
Investigational product, dose, and mode of administration: 
FLT180a will be given as a single dose, slow IV infusion. 
The dose escalation scheme proposed: 
• Cohort 1 (low dose): 6 x 1011 vector genomes (vg)/kilogram (kg) (of body weight) 
• Cohort 2 (intermediate dose): 2 x 1012 vg/kg 
• Cohort 3 (high dose): 4 x 1012 vg/kg 
Based on the observed FIX response, the dose level within a cohort may be reduced to avoid exposing patients 
to levels that exceed the normal physiological range, see Protocol Section 12.3.1. 
Methodology: 
This is a phase I/II, open label, multicentre, ascending single dose, safety study of FLT180a in patients with 
severe (FIX activity <1%) or moderately severe (FIX activity 1-2% with severe bleeding phenotype) HB. Patients 
who provide consent to participate in this study will be screened for eligibility and will have historical data on 
bleeding and FIX consumption documented from the previous 3 years’ medical notes. During the screening 
period patients will complete a diary to prospectively record on-going bleeding events and FIX consumption.  
On the day prior to infusion (Day -1), the patient will attend the investigational site and final eligibility assessments 
will be conducted. On Day 0, FLT180a will be administered as a single dose, slow IV infusion into a peripheral 
vein, and the patient will be monitored closely. Subject to satisfactory results from safety evaluations, the patient 
will be discharged from the investigational site but will continue to be monitored through a comprehensive battery 
of safety assessments at outpatient visits for a period of 26 weeks, following which the patient will enter a period 
of long-term follow-up conducted under a separate extension protocol.  
Up to 3 dose cohorts of vector (low, intermediate, and high) will be tested in the dose escalation. Two patients 
will be tested at each dose cohort with an additional patient added in the event of a dose-limiting toxicity (DLT) 
(2 + 1 design). Safety outcomes (as determined by adverse event [AE] reporting and laboratory tests) and results 
of FIX activity will be overseen by a trial management group and a data monitoring committee consisting of 
independent medical experts. Dose escalation may occur provided there is no more than 1 DLT at any dose 
cohort and if the resulting FIX activity fails to reach the target level. The goal for FIX response is to maximise the 
number of patients who achieve a FIX activity in the range 70-150% whilst minimising the risk of overshooting 
the normal physiological range. In order to reduce the risk of overshooting the normal physiological range, dose 
reduction within a dose cohort may occur if FIX activity in the initial patients in that dose cohort exceeds levels 
predefined in the protocol. Where a dose reduction occurs the 2+1 design will apply at that new dose level within 
the cohort. Dose escalation/reductions will be overseen by the trial management group and data monitoring 
committee. Additional patients may be added to a dose cohort to ensure adequate characterisation of any DLT, 
safety issues not meeting DLT criteria, or the FIX response at the request of the trial management group and 
data monitoring committee and at the discretion of the Sponsor. 
All patients will be provided with a take-home pack of immunosuppressants which will only be used at the 
direction of the investigator and liver function tests will be monitored intensively post infusion. From the week 3 
visit, all patients will take a course of immunosuppressants. At all other times, if patients develop liver 
inflammation, they will be instructed by the investigator to initiate treatment with a short course of 
immunosuppressants.  
The Sponsor, trial management group and data monitoring committee will select the terminal dose level based 
on patient FIX activity levels with the aim of ensuring the majority of patients will reach a FIX activity within normal 
limits in the absence of dose-limiting AEs. This terminal dose level will be expanded to 14 patients. 
On completion of the study, patients will be followed for 15 years under a separate extension protocol. 
Inclusion and exclusion criteria:  
Inclusion criteria: 
1. 
Adults males, ≥ 18 years of age;  
2. 
Confirmed diagnosis of HB defined as one of the following:  

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(a) Documented severe FIX deficiency with plasma FIX activity of <1% of normal, or  
(b) moderately severe FIX deficiency with plasma FIX activity level between ≥1% and ≤2% and a severe 
bleeding phenotype defined by one of the following: 
(i)  On prophylaxis for a history of bleeding, or  
(ii) On-demand therapy with a history of 4 or more bleeding episodes/year on average over the past 3 years, 
or  
(iii) Evidence of chronic haemophilic arthropathy (pain, joint destruction, and loss of range of motion). 
3. 
Able to give full informed consent and able to comply with all requirements of the trial including 15-year 
long-term follow-up;  
4. 
Willing to practice barrier contraception until at least three consecutive semen samples after vector 
administration are negative for vector sequences;  
5. 
Lack of neutralising anti-AAV-S3 antibodies using an in vivo transduction inhibition assay within 4 weeks 
of vector administration; 
6. 
At least 150 exposure days to FIX concentrates.  
Exclusion criteria: 
1. 
Presence of neutralising anti-human FIX antibodies (inhibitor, determined by the Bethesda inhibitor assay) 
at the time of enrolment or a previous history of FIX inhibitor; 
2. 
Patients at high risk of thromboembolic events (high risk patients would include those with a history of 
arterial or venous thromboembolism (e.g. deep vein thrombosis, pulmonary embolism, non-haemorrhagic 
stroke, arterial embolus) and those with acquired thrombophilia including conditions such as atrial 
fibrilation);  
3. 
Use of investigational therapy for haemophilia within 30 days before enrolment;  
4. 
Patients with active hepatitis B or C, and HBsAg or HCV RNA viral load positivity, respectively, or currently 
on antiviral therapy for hepatitis B or C. Negative viral assays in 2 samples, collected at least 6 months 
apart, will be required to be considered negative. Both natural clearers and those who have cleared HCV 
on antiviral therapy are eligible.; 
5. 
Serological evidence of HIV-1;  
6. 
Evidence 
of 
liver 
dysfunction 
(persistently 
elevated 
alanine 
aminotransaminase, 
aspartate 
aminotransferase, bilirubin >1.5 x upper limit of normal); 
7. 
Platelet count <50 x 109/L;  
8. 
Uncontrolled glaucoma, diabetes mellitus, or hypertension;  
9. 
Malignancy requiring treatment;  
10. Patients with uncontrolled cardiac failure, unstable angina or myocardial infarction in the past 6 months;  
11. Poor performance status (World Health Organization score >1); 
12. Prior treatment with any gene transfer medicinal product; 
13. Known or suspected intolerance, hypersensitivity or contraindication to the investigational product and 
non-investigational medicinal products or their excipients; 
14. Planned major elective surgery prior to the end of trial. 
15. Current or relavant history of a physical or psychiatric illness or any medical condition that in the opinion 
of the investigator could affect the patients safety or interfere with the study assessments. 
16. CMV IgG postive patients who are CMV PCR positive at screening. 
Maximum duration of patient involvement in the study: 
• Planned duration of screening period: Up to 52 weeks  
• Planned duration of follow-up period post infusion: 26 weeks  
• Planned duration of long-term follow-up: 15 years (under a separate extension protocol)  
Endpoints and statistical analysis:  
Endpoints: 
Primary endpoints 
Safety 
Safety as assessed by the reporting of AEs according to Common Terminology Criteria for Adverse Events 
(CTCAE) v5.0. 
 
 

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
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Efficacy 
The following primary endpoints will be analysed: 
• 
The proportion of patients achieving clinical FIX response at Week 26, at the terminal dose level. A clinical 
FIX response is defined as achieving FIX activity of 5% to 150%. 
• 
The proportion of patients also achieving normalised FIX response at Week 26, at the terminal dose level. 
A normalised FIX response is defined as achieving FIX activity of in the normal range (50-150%).2 
Secondary endpoints 
Safety 
Safety as assessed by reporting of abnormal or change from baseline findings from safety assessments 
including, laboratory assessments, vital signs, ECG, physical exam and liver ultrasound. 
Endogenous FIX production  
• The proportion of patients achieving FIX activity at or above 5%, 15%, 30%, 40%, 50% and 70% but no 
more than 150% of normal, at each scheduled visit. 
• The proportion of patients achieving FIX activity at or above 5%, 15%, 30%, 40%, 50%, 70% and 150% of 
normal, at each scheduled visit. 
• Absolute change from baseline in FIX activity. 
Haemostatic effectiveness 
• Change from baseline in annualised bleeding rate. 
• Change from baseline in FIX concentrate consumption. 
In order to ensure enough time has elapsed for the patient to have endogenous FIX activity to protect the patient 
from spontaneous bleeding episodes, the calculation period for haemostatic effectiveness will be from day 15 
inclusive. 
Immune response 
Immune response to the FIX transgene product (i.e., development of inhibitors) will be assessed by 
measurement of the level of inhibitors. 
Shedding 
Clearance of vector genomes in plasma, urine, saliva, stool, and semen. 
Exploratory endpoints 
Haemostatic effectiveness 
Exploration of the correlation between FIX levels and bleeding events over time. 
Immune response 
• Immune response to the AAV-S3 capsid will be assessed by measurement of the S3 neutralising antibody 
titre. 
• T-cell responses to AAV-S3 capsid in peripheral blood mononuclear cells. 
Disability status 
Change from baseline in World Health Organization Disability Assessment Schedule 2.0 score. 
Physical activity  
Change from baseline in Haemophilia Activities List (HAL) 2005. 
Health-related quality of life  
Change from baseline in the EQ-5D-5L and the Haem-A-QoL score. 
Haemophilia health status 
Change from baseline in the PROBE score. 
Assessment of joint health/function  
Change from baseline in the Haemophilia Joint Health Score (HJHS). 
Health resource utilisation 
Number of haemophilia related medical appointments and medical activities. 
Number of emergency room visits. 
Number of hospitalisations related to haemophilia. 
Length of hospital stay. 
 
2 https://www.wfh.org/en/page.aspx?pid=643 

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
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Number of days lost from education or work by patients and caregivers due to bleeding episodes. 
Number of physiotherapy sessions, specialist consultations and appointments with professional caregivers. 
 
Statistical analysis: 
Statistical analysis plan 
All study data will be listed and all relevant data will be tabulated and summarised by dose cohort and overall. A 
Statistical Analysis Plan (SAP) will be written and finalised prior to database lock. This plan will give a detailed 
description of all summaries and analyses that will be presented. 
Demographic and baseline characteristics 
Continuous variables will be summarised using number of observations, mean and standard deviation, median, 
and minimum and maximum values. Categorical values will be summarised using number of observations and 
percentages. Medical history will be summarised using number of observations and percentages of patients 
reporting each category. 
Primary endpoint 
Safety 
Adverse events will be coded using CTCAE version 5.0. Frequency of treatment-emergent AEs (TEAEs) will be 
calculated for each body system and preferred term, and by dose cohort, for number of events and number of 
patients reporting the event. The severity of the TEAEs and the relationship to study medication will be 
summarised for each body system and preferred term by dose level.  
Efficacy 
The FIX response will be derived for each patient based on the FIX activity level measured at the central 
laboratory. Clinical FIX response is defined as achieving FIX activity of 5-150% and normalised FIX response is 
defined as achieving FIX activity in the normal range (50-150%). 
The two primary endpoints will be analysed on the full analysis set. The proportion of patients achieving clinical 
FIX response at Week 26, will be summarised for the patients who received the terminal dose level. For the 
primary analysis, last-observation-carried-forward (LOCF) methodology will be used to impute any missing FIX 
value at Week 26, by the last non-missing FIX value.  For the primary analysis, FIX response will be assessed 
whether or not the patient is receiving or has been receiving immunosuppressants due to transaminitis outside 
the period of prophylactic immunosuppressant treatment. The proportion of patients also achieving normalisation 
will be summarised in a similar manner. 
Secondary endpoints 
Safety 
Descriptive statistics (number of observations, mean, standard deviation, minimum, median and maximum 
values) will be calculated for clinical laboratory tests at applicable visits. Changes from baseline will also be 
presented. For laboratory tests, abnormal values will be flagged in the data listings. In particular, patients 
achieving FIX activity above 150% of normal will be summarised by dose and overall. 
Vital signs (systolic and diastolic blood pressure, temperature, and pulse) and physical examination will be 
summarised by dose cohort using appropriate descriptive statistics. Continuous variables will be summarised 
using number of observations, mean, standard deviation, minimum, median, and maximum values. Categorical 
values will be summarised using number of observations and percentages. 
Endogenous FIX production  
• The proportion of patients achieving FIX activity at or above 5%, 15%, 30%, 40%, 50% and 70% but no 
more than 150% of normal, at each scheduled visit will be summarised by dose and overall. 
• The proportion of patients achieving FIX activity at or above 5%, 15%, 30%, 40%, 50%, 70% and 150% of 
normal, at each scheduled visit will be summarised by dose and overall. 
• Change from baseline in FIX activity as a percentage of normal values will be summarised in tabular and 
graphical format for each patient, by dose and overall. 
Haemostatic effectiveness 
Annualised bleeding rate 
The number of breakthrough bleeding episodes (spontaneous and traumatic) following FLT180a infusion will be 
annualised and compared with the patient’s own baseline bleeding history (the annualised mean of 3 years of 
historical bleeding records, and where possible, prospective data collected during the screening period will be 
used). 
Factor IX concentrate consumption 
The dose (international unit [IU]/kg) of factor concentrate used overall and by type of bleeding episode (i.e., 
spontaneous or traumatic) and location of bleeding episode (i.e., joint, soft tissue, or muscle) will be summarised 
with descriptive statistics. The total units of annualised factor consumption will be calculated and compared with 

Short title: A Factor IX Gene Therapy Study (FIX-GT)  
 
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the patient’s own baseline factor concentrate history (the annualised mean of 3 years’ historical factor 
concentrate consumption records, and where possible, prospective data collected during the screening period 
will be used). 
 
In order to ensure enough time has elapsed for the patient to have endogenous FIX activity to protect the patient 
from spontaneous bleeding episodes, the calculation period for haemostatic effectiveness will be from day 15 
inclusive to the date of completion of the last diary. More details will be provided in the SAP. 
Immune response 
FIX 
Immune response to the FIX transgene product (i.e., development of inhibitors) will be assessed by 
measurement of the level of inhibitors. 
Descriptive statistics (number of observations, mean, standard deviation, minimum, median and maximum 
values) will be calculated for immune response laboratory tests at applicable visits. 
Shedding 
Clearance of vector genomes in plasma, saliva, urine, stool, and semen will be summarised. The time to 
unquantifiable result by body fluid will be summarized and listed. 
 

Short title: A Factor IX Gene Therapy Study (FIX-GT)   
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Table 1: Schedule of Assessments 
Procedure 
Week 
Day 
Week* 
 
-52 to -1A 
-1B 
0 
+1 
+2 
+4 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
Visit Window 
 
 
 
 
 
+ / - 1 day 
Informed Consent 
X 
 
For Days -1 to +4 
please see Table 2: Detailed 
Schedule of Assessments for 
Infusion Week 
 
 
 
 
 
 
 
 
 
 
Demographics and Medical History 
X 
 
 
 
 
 
 
 
 
 
 
Bleeding History (incl. Target Joint 
Assessment) 
X 
 
 
 
 
 
 
 
 
 
 
Prior and Concomitant Medications 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Physical ExaminationC 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Vital SignsD 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
12-lead ECG  
X 
 
X 
 
X 
 
 
 
X 
 
 
Adverse Event Assessment 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Joint Evaluation (HJHS) 
X 
 
 
 
 
 
 
 
 
 
 
Liver UltrasoundE 
X 
 
 
 
 
 
 
 
 
 
 
AAV Antibody ScreenF 
XG 
 
 
 
 
 
 
 
 
 
 
FIX Antigen 
X 
 
 
 
 
 
 
 
 
 
 
FIX GenotypeH 
X 
 
 
 
 
 
 
 
 
 
 
HBV, HCV, HIV CMV ScreenI 
X 
 
 
 
 
 
 
 
 
 
 
Baseline FIX ActivityJ   
X 
 
 
 
 
 
 
 
 
 
 
FIX Activity Trough 
X 
 
 
 
 
 
 
 
 
 
 
Diary Completion for Bleeding 
Events and FIX Consumption 
 
 
 
 
 
 
 
 
 
 
 
QoL (EQ-5D-5L and Haem-A-QoL), 
Disability (WHODAS 2.0), Physical 
Activity (HAL 2005), Haemophilia 
Health Status (PROBE) and Health 
Resource UtilisationK 
X 
 
 
 
 
 
 
 
 
 
 
Liver Function Test (Local)L, M 
 
XN 
XN 
XN 
XN 
XN 
XO 
XO 
XO 
XO 
XO 
FIX Activity Level (Local)M 
 
XN 
XN 
XN 
XN 
XN 
XO 
XO 
XO 
XO 
XO 
Haematology, Chemistry incl. CRP, 
Coagulation Screen (Local)P 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Haematology, Chemistry incl. CRP, 
Coagulation Screen (Central) 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Liver Function Test (Central) 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
FIX Activity Level (Central)F 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
FIX Inhibitor Level 
X 
X 
X 
X 
 
 
X 
 
 
X 
 
AAV-S3 Antibody TitreF 
 
X 
X 
 
 
 
X 
 
 
 
 
Mononuclear Cells (Elispot) 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 

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Procedure 
Week 
Day 
Week* 
 
-52 to -1A 
-1B 
0 
+1 
+2 
+4 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 
Visit Window 
 
 
 
 
 
+ / - 1 day 
Mononuclear Cells (Research) - 
Optional 
 
 
For Days -1 to +4 
please see Table 2: Detailed 
Schedule of Assessments for 
Infusion Week 
 
 
 
 
 
 
 
 
 
 
Research Plasma Samples - 
Optional 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
FIX Activity Research Plasma 
Samples 
 
X 
 
 
X 
 
 
 
 
 
 
Immune Response Research 
Plasma Samples 
 
XQ 
XQ 
XQ 
XQ 
XQ 
XQ 
XQ 
XQ 
XQ 
XQ 
Prophylactic ImmunosuppressantsR 
 
 
 
 
 
 
 
 
 
 
 
Test for Reactivation of HepatitisS 
 
 
 
 
X 
 
X 
 
X 
 
X 
Test for CMVT 
 
 
 
X 
X 
X 
X 
X 
X 
X 
X 
Test for Tacrolimus LevelU 
 
 
 
X 
X 
X 
X 
X 
X 
X 
X 
PCR of Vector Genomes in Plasma, 
Saliva, Urine, Stool and Semen 
 
XV 
XV 
XV 
XV 
XV 
XV 
XV 
XV 
XV 
XV 
 
 

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Procedure 
Week* 
 
11 
12 
13W 
14 
15W 
16 
17W 
18W 
19W 
20 
21W 
22W 
23W 
24W 
25W 
26/EOSX 
Visit Window 
+ / - 1 day 
+ / - 3 days 
Informed Consent 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Demographics and Medical History 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Bleeding History (incl. Target Joint 
Assessment) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Prior and Concomitant Medications 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Physical ExaminationC 
X 
X 
 
X 
 
X 
 
 
 
X 
 
 
 
 
 
X 
Vital SignsD 
X 
X 
 
X 
 
X 
 
 
 
X 
 
 
 
 
 
X 
12-lead ECG  
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Adverse Event Assessment 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Joint Evaluation (HJHS) 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Liver UltrasoundE 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
AAV Antibody ScreenF 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
FIX Antigen 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
FIX GenotypeH 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
HBV, HCV, HIV CMV ScreenI 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Baseline FIX ActivityJ   
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
FIX Activity Trough 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Diary Completion for Bleeding 
Events and FIX Consumption 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
QoL (EQ-5D-5L and Haem-A-QoL), 
Disability (WHODAS 2.0), Physical 
Activity (HAL 2005), Haemophilia 
Health Status (PROBE) and Health 
Resource UtilisationK 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Liver Function Test (Local)L, M 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
X 
FIX Activity Level (Local)M 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
XO 
X 
Haematology, Chemistry incl. CRP, 
Coagulation Screen (Local)P 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Haematology, Chemistry incl. CRP, 
Coagulation Screen (Central) 
X 
X 
 
X 
 
X 
 
 
 
X 
 
 
 
 
 
X 
Liver Function Test (Central) 
X 
X 
 
X 
 
X 
 
 
 
X 
 
 
 
 
 
X 
FIX Activity Level (Central)F 
X 
X 
 
X 
 
X 
 
 
 
X 
 
 
 
 
 
X 
FIX Inhibitor Level 
 
X 
 
 
 
X 
 
 
 
X 
 
 
 
 
 
X 
AAV-S3 Antibody TitreF 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Mononuclear Cells (Elispot) 
X 
X 
 
X 
 
X 
 
 
 
X 
 
 
 
 
 
X 

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Procedure 
Week* 
 
11 
12 
13W 
14 
15W 
16 
17W 
18W 
19W 
20 
21W 
22W 
23W 
24W 
25W 
26/EOSX 
Visit Window 
+ / - 1 day 
+ / - 3 days 
Mononuclear Cells (Research) - 
Optional 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Research Plasma Samples - 
Optional 
X 
X 
 
X 
 
X 
 
 
 
X 
 
 
 
 
 
X 
FIX Activity Research Plasma 
Samples 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
Immune Response Research 
Plasma Samples 
XQ 
XQ 
 
XQ 
 
XQ 
 
 
 
XQ 
 
 
 
 
 
X 
Prophylactic ImmunosuppressantsR 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Test for Reactivation of HepatitisS 
 
X 
 
X 
 
X 
 
X 
 
X 
 
 
 
 
 
 
Test for CMVT 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
 
 
 
 
Test for Tacrolimus LevelU 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
 
 
 
 
 
 
PCR of Vector Genomes in 
Plasma, Saliva, Urine, Stool and 
Semen 
XV 
XV 
 
XV 
 
XV 
 
 
 
XV 
 
 
 
 
 
XV 
 
Abbreviations: AAV = adeno-associated virus; CMV = cytomegalovirus; CRP = C-reactive protein; ECG = electrocardiogram; EOS = end of study; FIX = Factor IX; HAL = 
haemophilia activity list; HBV = hepatitis B virus; HCV = hepatitis C virus; HIV = human immunodeficiency virus; HJHS = Haemophilia Joint Health Score; PCR = 
polymerase chain reaction; QoL = quality of life; WHODAS = World Health Organisation Disability Assessment Schedule. 
 
*  Week 1 visit is Day +7, Week 2 visit is Day +14, Week 3 visit is Day +21, Week 4 visit is Day +28 etc. 
 
A  The screening period can be up to 52 weeks to allow for controlled prospective collection of bleeding event and FIX consumption data via the patient diary. A patient can 
move forward to Day -1 assessments and dosing as dosing slots are available. There is no minimum time frame for the data collection. ALL screening assessments should 
be completed in a timely manner (~2 weeks) following consent to confirm patient eligibility. The patient should be contacted ~every 4 weeks through the screening period to 
check on diary completion and collect details on any AEs. A repeat AAV Antibody screen is mandated within 4 weeks of dosing. If the screening period for a patient is longer 
than 16 weeks then a repeat of the central laboratory screening bloods (FIX Antigen/ HBV, HCV, HIV, CMV Screen/ Haematology, Chemistry incl. CRP, Coagulation 
Screen/ Liver Function Test/ FIX Activity Level/ FIX Inhibitor Level) will be required within 4 weeks of dosing and the patient reported outcomes should also be repeated at 
Day -1. 
B  The Day -1 assessments can be conducted as early as Day -3 for logistical reasons, if required. 
C  Height (screening only) and weight (screening) will also be measured. Waist circumference (cm), hip circumference (cm), neck circumference (cm) and bioimpedance will 
be measured at screening. 
D  Vital signs include pulse, blood pressure.  
E  Dependent on results of liver ultrasound further investigations including fibroscan/MRI/elastography may be carried out. 
F  Two aliquots will be taken at each timepoint. 
G  A negative assay (transduction inhibition assay) outcome must be documented within 4 weeks of dosing. Repeat testing may be indicated due to the long screening window. 
H  Blood sample to be taken and sent to the central laboratory for testing if not already documented in the medical records. 
I  HCV antibody testing followed by HCV RNA load. HCV RNA viral load only indicated for patients with history of HCV and positive HCV antibody test. CMV IgG testing 
followed by CMV PCR. CMV PCR only indicated for a positive CMV IgG. 

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J  Established following 5 days washout or from documented medical records. 
K  For Health Resource Utilisation at Screening a 6-month history will be collected and at EOS all Health Resource Utilisation since Screening (or Day -1) will be collected. 
L  Albumin, alkaline phosphatase, direct bilirubin, indirect bilirubin, total bilirubin, alanine aminotransferase, aspartate aminotransferase. 
M  Frequency to be increased in response to an upward trend in LFTs and in line with guidance in section 9.4. 
N  Three times per week. The tests should be as evenly spaced through the week as possible, for example: Monday, Wednesday and Friday. On days where other clinic 
assessments are not required these samples may be taken at study site or at an alternative location (e.g. patient’s home). 
O  Twice weekly. The tests should be as evenly spaced through the week as possible, for example: Monday and Thursday or Tuesday and Friday. On days when other clinic 
assessments are not required these samples may be taken at study site or at an alternative location (e.g. patient’s home). 
P  Complete blood count with differentials and platelets. Chemistry incl. CRP to include sodium, potassium, phosphate, blood urea nitrogen or urea, serum creatinine (and 
estimated GFR), CRP. Coagulation screen to include prothrombin time and activated partial thromboplastin time.  
Q  To be drawn only during the course of investigation or treatment of breakthrough transaminitis. 
R  A weight based immunosuppressant regimen will be initiated at the Week 3 visit. For details see section 9.4.  
S  Local testing. Only required for patients with history of Hepatitis B or C, commencing at Week 4 and continuing every 2 weeks until prophylactic immunosuppressant regimen 
completed. Tests should be undertaken to coincide with use of immunosuppression if outside of the prophylactic regimen. On days where other clinic assessments are not 
required these samples may be taken at study site or at an alternative location (e.g. patient’s home). 
T  Local testing. For patients positive for CMV (IgG) at screening weekly CMV PCR testing for the duration of the immunosuppressant regimen. Tests should be undertaken 
to coincide with use of immunosuppression if outside of the prophylactic regimen. On days where other clinic assessments are not required these samples may be taken at 
study site or at alternative location (e.g. patient’s home). Management guidelines in the case of CMV reactivation can be found in Appendix 2.  
U  Local testing. Tacrolimus levels should be tested at each blood draw until levels are in the therapeutic range and weekly thereafter for the duration the patient is taking 
tacrolimus in the immunosuppressant regimen. Tests should be undertaken to coincide with use of tacrolimus if outside of the prophylactic regimen. On days where other 
clinic assessments are not required these samples may be taken at study site or at an alternative location (e.g. patient’s home). Management guidelines for tacrolimus dosing 
can be found in Appendix 3. 
V  Three times within 7-10 days following vector infusion then weekly until 3 consecutive samples are negative. 
W  Blood draws may be taken either at study site or at alternative location (e.g. patient’s home). The intention should be for at least one of the two blood draws per week to 
be conducted at study site. 
X  In the event of patient discontinuation / withdrawal every effort should be made to complete week 26/end of study procedures. 
 
 
 

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Table 2: Detailed Schedule of Assessments for Infusion Week  
 
Procedure 
Day -1a 
Day 0 (Infusion Day) 
Day 
 
Pre-
doseb 
Timepoint (minutes) during 
Infusion 
Timepoint (hours) from End of Infusion 
 
 
0 
+15 
+30 
+45 
+60 
+1 
+2 
+3 
+4 
+5 
+6 
+8 
+10 
+12 
+16 
+20 
+1 
+2 
+4 
Window 
- 2 days 
 
 
+/- 5 mins 
+/- 10 mins 
+/- 15 mins 
 
 
+/- 1 
day 
Informed Consentc 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Physical 
Examinationd 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
X 
Vital Signse 
X 
X 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Xf 
Xf 
X 
X 
X 
12-lead ECG  
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
QoL (EQ-5D-5L and 
Haem-A-QoL), 
Disability 
(WHODAS 2.0), 
Physical Activity 
(HAL 2005), 
Haemophilia Health 
Status (PROBE) 
and Health 
Resource 
Utilisationg 
Xh 
FLT180a  
Administration 
 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Diary Completion 
for Bleeding Events 
and FIX 
Consumption 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Prior and 
Concomitant 
Medications 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Adverse Events 
Assessment 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 

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Procedure 
Day -1a 
Day 0 (Infusion Day) 
Day 
 
Pre-
doseb 
Timepoint (minutes) during 
Infusion 
Timepoint (hours) from End of Infusion 
 
 
0 
+15 
+30 
+45 
+60 
+1 
+2 
+3 
+4 
+5 
+6 
+8 
+10 
+12 
+16 
+20 
+1 
+2 
+4 
Window 
- 2 days 
 
 
+/- 5 mins 
+/- 10 mins 
+/- 15 mins 
 
 
+/- 1 
day 
Local Lab Tests 
Haematology, 
Chemistry incl. 
CRP, Coagulation 
Screeni 
X 
X 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
X 
X 
X 
Liver Function Testj 
X 
X 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
X 
X 
X 
FIX Activity Level  
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
Central Lab Tests 
Haematology, 
Chemistry incl. 
CRP, Coagulation 
Screen 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
X 
Liver Function Test 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
FIX Activity Levelk  
X 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
AAV Antibody 
Screenk 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Mononuclear Cells 
(Elispot) 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Mononuclear Cells 
(Research) - 
Optional 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Research Plasma 
Sample - Optional 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Immune Response 
Research Plasma 
Sample 
X 
PCR of Vector 
Genomes in 
Plasma, Saliva, 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Xl 
Xl 

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Procedure 
Day -1a 
Day 0 (Infusion Day) 
Day 
 
Pre-
doseb 
Timepoint (minutes) during 
Infusion 
Timepoint (hours) from End of Infusion 
 
 
0 
+15 
+30 
+45 
+60 
+1 
+2 
+3 
+4 
+5 
+6 
+8 
+10 
+12 
+16 
+20 
+1 
+2 
+4 
Window 
- 2 days 
 
 
+/- 5 mins 
+/- 10 mins 
+/- 15 mins 
 
 
+/- 1 
day 
Urine, Stool and 
Semen 
 
Abbreviations: AAV = adeno-associated virus; CRP = C-reactive protein; ECG = electrocardiogram; PCR = polymerase chain reaction. 
 
a  The Day -1 assessments can be conducted as early as Day -3 for logistical reasons, if required. 
b  Approximately 1 hour before infusion. 
c  The ‘pre-infusion’ dosing stage informed consent may occur prior to day -1. 
d  Weight (Day -1) will also be measured. Waist circumference (cm), hip circumference (cm), neck circumference (cm) and bioimpedance will be measured at Day -1. 
e  Vital signs from day -1 to day +4 include pulse, blood pressure, respiration rate and temperature. 
f  16 and 20 hour timepoints are only required for the first two patients in each cohort. 
g  For Health Resource Utilisation at Day -1 all Health Resource Utilisation since Screening will be collected. 
h  Only to be completed if patients had been in the screening period for more than 16 weeks. 
i  Complete blood count with differentials and platelets. Chemistry incl. CRP to include sodium, potassium, phosphate, blood urea nitrogen or urea, serum creatinine (and 
estimated GFR), CRP. Coagulation screen to include prothrombin time, activated partial thromboplastin time. 
j  Albumin, alkaline phosphatase, direct bilirubin, indirect bilirubin, total bilirubin, alanine aminotransferase, aspartate aminotransferase. 
k  Two aliquots will be taken at each timepoint. 
l  Three times within 7-10 days following vector infusion then weekly until 3 consecutive samples are negative. 
 

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5 
Introduction 
5.1 
Background 
Haemophilia B (HB) is an X-linked recessive bleeding disorder that affects approximately 1 in 
30,000 males. This disorder results from a defect in the Factor IX (f9) gene, which encodes a 
serine protease, critical for appropriate fibrin clot formation. Approximately 50% of individuals 
with HB have a severe disorder characterised by functional FIX levels of less than 1% of 
normal (<1 U/dL = 50 ng/mL). An additional “moderately severe” patient group also exists that 
is characterised by FIX levels between 1-2% whilst retaining the severe bleeding phenotype. 
Clinically the disease is characterised by frequent spontaneous bleeding into joints and soft 
tissues which, without adequate treatment, causes a chronic debilitating arthropathy. 
Spontaneous bleeding into closed spaces such as the brain occurs in a significant proportion 
of patients with catastrophic manifestations including death. Current treatment for HB in the 
Western World may be either on demand treatment or prophylactic treatment. On-demand 
treatment involves intravenous (IV) infusions of plasma-derived or recombinant factor 
concentrates at the time of a bleed. This is highly effective at arresting haemorrhage but cannot 
totally abrogate chronic damage that ensues after a bleed. In severely affected patients, 
bleeding episodes can be dramatically reduced with prophylactic treatment when plasma FIX 
levels are maintained at or above only 1% of normal by regular administration of FIX protein 
concentrates.1 The relatively short half-life of standard FIX concentrates requires infusions to 
be frequent at 2-3 times a week. Even extended half-life (EHL) FIX concentrates require 
regular infusions. The need for regular infusions is invasive, inconvenient, and highly 
problematic for children. Central lines are often required in children which may cause 
significant morbidity due to catheter-related complications. 
The cost of prophylactic treatment for an adult is in excess of $150,000 per year, a significant 
cost burden for all healthcare systems even within the developed world. In the context of 
haemophilia worldwide, the cheaper plasma-derived FIX concentrates, if used for treatment 
alone rather than prophylaxis, still remain unaffordable by the majority of people worldwide 
who have haemophilia B.2 Formation of neutralising antibodies (inhibitors) which occur in 3% 
of HB patients resulting in poor response to factor concentrates, often with fatal outcomes, is 
a rare but significant complication of protein replacement therapy. Additionally, the 
psychological burden associated with the constant but unpredictable risk of bleeding in this 
chronic disorder is, perhaps, not fully appreciated. Therefore, replacement therapy with FIX 
products continues to be less than ideal both in the context of the developed as well as the 
developing world. 
These unmet needs have fuelled interest in gene therapy of HB which has several fundamental 
advantages over other single-gene disorders. The clinical manifestations represent a simple 
cause and effect relationship which is attributable to the lack of a single gene product that 
circulates in minute amounts in the plasma. In addition, tightly regulated control of gene 
expression is not essential since a wide range of FIX is expected to be beneficial and nontoxic. 
The availability of animal models including FIX-knockout mice3,4,5 and HB dogs6,7 has facilitated 
extensive preclinical evaluation of gene therapy strategies. Perhaps the most important single 

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aspect of HB that makes it such an excellent target for gene therapy is the fact that even a 
small rise (1% of physiological levels) in circulating clotting factor would have a significant 
beneficial therapeutic effect which could transform the lives of patients with severe 
haemophilia. Small changes in FIX can be easily assessed using both clinical and laboratory 
evaluation. Continuous synthesis of FIX by the host cells after gene therapy provides a real 
opportunity to prevent bleeding episodes rather than simply treating the bleeds after they have 
occurred. Additionally, the risks and the inconvenience of regular infusion of protein 
concentrates would be avoided following gene therapy. Importantly endogenous synthesis of 
FIX within hepatocytes after liver-targeted delivery of vector may also lead to immunological 
tolerance to the transgene product thereby potentially reducing the risk of neutralising FIX 
antibody formation.8,9,10 Finally, and crucially, endogenously synthesised FIX is maintained at 
a constant level after gene transfer, thus avoiding the “saw tooth” pattern produced by 
intermittent infusion with its inherent troughs, where break through bleeding is most likely to 
occur.  
The results from gene therapy studies conducted to date offers the prospect of sustained FIX 
expression in excess of levels that can be achieved with exogenous prophylactic treatment. 
The achievement of superior efficacy suggests there is potential for a functional cure with a 
lack of haemorrhage even in response to trauma or surgery. 
5.2 
Nonclinical Data 
The overall nonclinical development programme consists of in vitro and in vivo 
pharmacodynamic 
studies 
and 
single-dose 
Good 
Laboratory 
Practice-compliant 
toxicology/biodistribution studies in Rhesus Macaques.  
In the text of this document, reference is made to FLT180 and FLT180a. FLT180a is the 
proposed investigational medicinal product and is an evolution of FLT180 which was the 
construct on which initial pre-clinical testing was carried out. The only difference between 
FLT180 and FLT180a is that FLT180a incorporates a change to the codon optimisation that 
was used in FLT180. The codon optimisation is in exons 2 to 8 of the f9 gene and represents 
a change of <20% of the overall sequence. The capsid, promotor, intron, exons 1, poly A tail 
and the amino acid sequence of the factor IX produced are all unchanged and are identical to 
FLT180. Because the change does not affect the transgene product and only affects a small 
part of the overall vector sequence, the change is not expected to affect product quality 
attributes other than protein expression levels. On this basis, non-clinical work carried out on 
FLT180 is considered to be highly supportive of FLT180a. 
Please refer to the FLT180a Investigator Brochure for full details of the non-clinical data 
including pharmacodynamics, toxicology and biodistribution. 
5.3 
Clinical Data 
Currently a number of gene therapy medicinal products for HB are in development. These 
products are building on the work conducted by Nathwani et al, where 10 patients were treated 
with a first-generation AAV gene therapy.11 In this study, a single IV infusion of AAV2/8-LP1-
hFIXco resulted in an increase in plasma FIX activity from baseline values that ranged from 

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less than 1% of the normal value to steady-state levels of 1 to 6% of the normal value in all 10 
patients. Patients lacked antibodies to the capsid that was used in the study and response was 
linked to the dose patients received. Among patients receiving a low (2 x 1011 vg/kg) or 
intermediate (6 x 1011 vg/kg) dose of vector, the increase in FIX activity was modest (1 to 3% 
of the normal value). In contrast, remarkably consistent FIX expression (mean, 5.1 ±1.7% of 
the normal value) was observed in the 6 patients receiving the high dose of vector. Following 
treatment AAV8-mediated FIX expression remained stable over a period of >6 years, in 
keeping with the stability of transgene expression in animal models, including nonhuman 
primates.12  
Of the 7 patients who were receiving prophylaxis with FIX concentrate before gene transfer, 4 
were able to stop regular FIX replacement therapy, and most of the others were able to 
increase the interval between prophylactic infusions. Despite the reduction in the use of FIX 
concentrate, the average annual number of bleeding episodes was consistently lower after 
gene transfer, most notably in patients in the high-dose cohort, for whom the mean 
steady-state FIX expression was around 5% of the normal value. This reduction in the number 
of annual bleeding episodes occurred despite higher overall physical activity levels reported 
by all patients, including participation in sports activities that had previously been associated 
with bleeding. This improvement in the bleeding phenotype among patients in the high-dose 
cohort is consistent with the natural bleeding tendency in patients with mild haemophilia who 
have plasma FIX levels of 5 to 40%. Such patients have either no or very few spontaneous 
bleeding episodes but are at risk for excessive haemorrhage after trauma or surgery.13  
The major vector-related adverse event (AE) was a dose-dependent, asymptomatic increase 
in the serum alanine aminotransferase (ALT) level associated with a decline in FIX levels, 
suggesting a loss of transduced hepatocytes. Expansion of the high-dose cohort showed that 
this AE was common, occurring in 4 of the 6 patients but was well managed through the 
administration of prednisolone. The increase in serum ALT levels occurred consistently at 7 to 
10 weeks after gene transfer, thus defining the critical period of monitoring and pharmacologic 
intervention. In other studies using a similar construct (George et al NEJM 2017), the same 
asymptomatic increase in ALT requiring steroid treatment has been seen as early as 4 weeks 
post treatment with a dose of 5 x 1011 vg/kg.14 
5.4 
Rationale and Risks/Benefits 
5.4.1 
Rationale 
Haemophilia B is an X-linked recessive bleeding disorder that affects approximately 1 in 
30,000 males. The disorder results from a defect in the f9 gene and is characterised by 
frequent spontaneous bleeding into joints and soft tissues which, without adequate treatment, 
causes a chronic debilitating arthropathy. 
Current treatment for HB in the developed world involves either IV infusions of plasma-derived 
or recombinant factor concentrates at the time of a bleed (on-demand therapy) or regular 
infusions of clotting factors to prevent bleeds (prophylactic treatment). On-demand treatment 
is highly effective at arresting haemorrhage but cannot totally abrogate chronic damage that 

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ensues after a bleed. In severely affected patients, bleeding episodes can be dramatically 
reduced by prophylactic treatment when plasma FIX levels are maintained at or above only 
1% of normal by regular administration of FIX protein concentrates.1 The relatively short half-
life of FIX necessitates frequent IV administration of factor concentrates (2-3 times a week) 
which is invasive, inconvenient and problematic for children. 
Following administration of AAV gene therapy, the target cells are transduced. AAV gene 
therapy is poor at integrating into the host genome and the transgene remains episomally in 
those transduced cells. It is from this position that the transgene is expressed and, in the case 
of FIX, this is a steady, continuous expression. This continuous synthesis of FIX by the host 
cells after gene therapy offers a real opportunity to prevent bleeding episodes and eliminating 
the need for regular infusions. In addition, current experience suggests that the level of efficacy 
seen with gene therapy is likely to be superior to that seen with prophylactic treatment and, 
when considering the most recent data, may even offer the prospect of a functional cure and 
a lack of haemorrhage even in response to trauma or surgery.  
This study aims to investigate the safety of a new gene therapy (FLT180a) and investigate its 
potential to alter the disease phenotype from severe to mild/normal through the endogenous 
production of FIX following a single administration of vector. 
Dose Selection 
The objective is to select an initial dose of FLT180a, based on studies in non-human primates 
and extrapolation from results in previous studies that will result in FIX levels that would, at a 
minimum, reduce disease severity whilst balancing the risk of toxicity.  
In the study conducted by Nathwani et al, IV infusion of AAV2/8-LP1-hFIXco at low (2 x 1011 
vector genomes [vg]/kilogram [kg]), intermediate (6 x 1011 vg/kg), or high dose (2 x 1012 vg/kg) 
resulted in in a 1.5%, 2%, and 5% level of hFIX respectively. Asymptomatic, transient 
transaminitis occurred in 4 of 6 patients at high dose level (at 7-10 weeks) but was effectively 
managed by administration of a short course of prednisolone with liver enzymes normalised 
and hFIX levels retained in the 2-4% range.  
The FLT180a (AAV2/S3-
-Ti-FIXco1) investigational medicinal product (IMP) has been 
designed with the following potential enhancements which are hypothesised to confer an 
increase in FIX activity over and above what was achieved with the previous construct: 
(1) 
A new small synthetic liver promoter (
) 
(2) 
An enhanced transgene including 
a. codon optimised FIX gene 
b. a gain-of-function Padua mutation (R338L)  
c. truncated intron in a natural position 
(3) 
A novel engineered capsid (AAV-S3) developed from wild-type AAV3 and AAV8 through 
capsid shuffling. This enables the vector to transduce human hepatocytes with high 
efficiency. 

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In vivo data in mice models are not predictive of transgene expression in humans and therefore 
the proposed doses have been estimated based on extrapolation of in vitro data in human cells 
comparing the performance of elements of the AAV2/8-LP1-hFIXco construct used in the 
previous study by Nathwani et al. and those incorporated in FLT180a.11,15,16,17 Please refer to 
the investigator brochure for full details. 
The planned low cohort dose of FLT180a in this study (low dose: 6 × 1011 vg/kg of body weight) 
will be in line with the intermediate dose utilised in the study conducted by Nathwani et al. 
Table 3: FIX Activity Projections illustrates projections for FIX activity in the current study based 
on putative enhancements in the new construct. 
 
Table 3: FIX Activity Projections 
Dose (vg/kg) 
FIX levels (% of normal) 
achieved in published FIX 
trial (Nathwani et al) 
(Using AAV2/8-LP1-hFIXco) 
FIX levels (% of normal) 
projected for FLT180a in 
this study 
2 x 1011 
1.5 
- 
6 x 1011 
2 
30 
2 x 1012  
5 
75 
4 x 1012 
Not studied 
>75 
Abbreviations: FIX = Factor IX; kg = kilogram; vg = vector genomes. 
 
Depending on safety outcome (including AE reporting and laboratory tests) and results of FIX 
levels, dose cohort escalation may occur provided there is no more than 1 dose-limiting toxicity 
(DLT) at any dose level and if the resulting FIX activity fails to reach the target range. A DLT 
is defined as any grade 3 or greater AE at least possibly related to FLT180a. The goal for FIX 
response is to maximise the number of patients who achieve a FIX activity in the range 70-
150% whilst minimising the risk of overshooting the normal physiological range. A FIX activity 
in the range 70-150% represents normalisation to a level at which patients may undergo 
surgical procedures without the requirement for additional exogenous factor concentrates.18 
The dose will be escalated in increments to a maximum dose of 4 x 1012 vg/kg (high dose).  In 
order to reduce the risk of overshooting the normal physiological range, reduction of the dose 
level within a dose cohort may occur if FIX activity exceeds pre-defined levels in the initial 
patients. Where a dose reduction occurs the 2+1 design will apply at that new dose level within 
the cohort. Additional patients may be added to a dose cohort to ensure adequate 
characterisation of any DLT, safety issues not meeting DLT criteria, or the FIX response, at 
the request of the trial management group and data monitoring committee and at the discretion 
of the Sponsor (see Section 12).  

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Two Good Laboratory Practice toxicology studies using FLT180 and FLT180a have been 
conducted in Rhesus Macaques.  
In the first study using FLT180 and FLT180a, animals received doses of 3.34 x 1013 vg/kg and 
1.235 x 1012 vg/kg, respectively. For FLT180, this is 50-fold greater than the starting dose in 
cohort 1, and approximately 8-fold greater than the dose in cohort 3. For FLT180a this is 2-
fold the starting dose in cohort 1. There were no adverse findings in this study during the 8 
and 4 week assessment period respectively.  
In a supplementary toxicology study conducted with FLT180a at a dose of 3.41 x 1013 vg/kg 
(50-fold greater than the starting dose in cohort 1 and approximately 8 times the maximum 
human dose proposed) there were adverse histopathological findings in one animal that were 
likely associated with high hFIX levels (>12-times normal) 8 weeks post dose and considered 
an exaggerated pharmacological effect, thrombotic effects have been seen in one non-human 
primate. The occurrence of antibodies to hFIX in this animal may have resulted in an under 
estimation of the hFIX levels and therefore higher levels of hFIX may have been produced in 
the animal with the lung lesion.   
Peak hFIX levels up to 170% and sustained hFIX levels of 110% of normal have not resulted 
in FLT180a associated adverse effects and in this context the nonclinical data is considered 
to support dosing in this study. This protocol mandates rigorous monitoring of FIX levels in 
patients and includes strict controls around inter patient interval and stopping rules that are 
designed to minimise risk in the clinic associated with supraphysiological FIX levels.  
Dosing and Dose Escalation Interval 
In order to monitor for acutely occurring, dose-limiting AEs and to minimise the risk of 
overshooting the normal physiological range for FIX, a 4-week interval between dosing 
patients will be enforced during the dose escalation phase to monitor for acutely occurring 
dose-limiting AEs. This interval can be extended should there be any safety concerns (see 
Section 12.3.2 and Section 12.3.3). 
Analysis of the data from the studies conducted by Nathwani et al11,17 has shown a marked 
effect of corticosteroid treatment on FIX activity levels. An increase of 2.5 to 3-fold in FIX levels 
have been observed in parallel with the prednisolone regimen (unpublished data), these return 
to pre-steroid levels approximately 5 weeks following cessation. The steroid response with the 
smaller 
 promoter used in FLT180a is expected to be less profound but not absent and 
therefore pre-steroid FIX levels (week 3) will be used to guide dose escalation decisions. 
Based on data from a similar single stranded AAV vector currently in development (George et 
al NEJM 2017) FIX activity levels at 3 weeks represent approximately 65% of those achieved 
at plateau.14 Cohort data review will utilise the 65% FIX level at week 3 to project an expected 
steady state and will guide dose escalation/reduction decisions. 
As this will be a first time in human study for FLT180a, an extended 6-week interval will be 
observed following dosing of the first patient in the study and prior to dosing the second patient 
to minimise risk to subsequent patients of unanticipated AEs. 

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The main AE from previous studies has been an asymptomatic increase in serum ALT level 
associated with a decline in FIX levels occurring typically at week 7-10 following dosing in the 
study by Nathwani et al.11 To date these events have been effectively managed with rapid 
introduction of a short course of corticosteroids. In other studies using a similar construct 
(George et al NEJM 2017), the same asymptomatic increase in ALT requiring steroid treatment 
has been seen as early as 4 weeks post treatment with a dose of 5 x 1011 vg/kg.14 
All patients will be given a take-home pack of immunosuppressants to be taken under the 
direction of the investigator, which will allow rapid intervention if transaminase elevations are 
observed. In addition, during the anticipated critical time period all patients will receive a course 
of immunosuppressants commencing at the week 3 visit. 
As patient follow-up will potentially be carried out across multiple sites, it will be a requirement 
for all investigators with active patients who have received FLT180a to confirm the absence of 
safety issues in patients under their care that would prevent further patient dosing/dose 
escalation. Dosing of subsequent patients will not be allowed until this confirmation is received. 
5.4.2 
Potential Risks and Benefits 
The following are potential risks associated with participation in this study.  
Humoral Immune Response to AAV  
Experience in animal models indicates that administration of AAV results in a serotype specific 
stimulation of neutralising antibody production, and that such antibodies preclude 
re-administration of AAV of the same serotype although other serotypes can be successfully 
re-administered. It is therefore highly probable that patients who receive AAV-S3 whilst 
participating in this study will develop neutralising antibodies and that any future administration 
of vector of this serotype would be ineffective. This event becomes more problematic if the 
individual patient receives a sub-therapeutic dose. The sponsors objective is therefore to 
select an initial dose, based on studies in non-human primates and extrapolation from results 
in previous studies that may result in FIX levels that would, at a minimum, reduce disease 
severity (see section on Dose Selection). Experience from the first-generation FIX study 
conducted by Nathwani et al demonstrates that activity remains stable (>6 years) but natural 
turnover of liver cells would suggest that it is possible for levels to decline over time. An initial 
therapeutic level therefore might become sub-therapeutic in the future increasing the 
importance of careful evaluation of benefit/risk in dose selection.  
A major mitigating factor against this risk is that in both murine and non-human primate models, 
vector particles of an alternative serotype can be administered with subsequent predictable 
expression of FIX.1 Thus, if patients enrol into this trial and receive a subtherapeutic dose, they 
could be considered for future trials of AAV vectors of an alternate serotype with a potentially 
therapeutic benefit despite the presence of antibodies to AAV-S3.  
Screening for Liver Abnormalities 
Patients with evidence of liver dysfunction (persistently elevated ALT, AST, bilirubin >1.5xULN) 
are excluded from the study. All patients will undergo a liver ultrasound. Based on patient 

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history and outcome of the ultrasound, additional tests like fibroscan, MRI and elastography 
may be requested to exclude significant fibrosis prior to inclusion in the study with the input of 
the Chief Investigator. Results of these tests should be discussed with the Chief Investigator 
and Sponsor in order to confirm eligibility for the study. 
Vector-induced Hepatitis and Loss of FIX Expression  
In the study conducted by Nathwani et al, the major vector-related AE was a dose-dependent, 
asymptomatic increase in the serum ALT level associated with a decline in FIX levels, 
suggesting a loss of transduced hepatocytes. Expansion of the high-dose cohort showed that 
this AE was common, occurring in 4 of the 6 patients but was well managed through the 
administration of prednisolone. The increase in serum ALT levels occurred consistently at 7 to 
10 weeks after gene transfer, thus defining the critical period of monitoring and pharmacologic 
intervention. In other studies using a similar construct (George et al NEJM 2017), the same 
asymptomatic increase in ALT requiring steroid treatment has been seen as early as 4 weeks 
post treatment with a dose of 5 x 1011 vg/kg.14 
Patients will be given a take-home pack of immunosuppressants to be used in the event of 
transaminase elevation. Patients will be monitored intensively over the course of the study for 
evidence of immune hepatitis and a short course of immunosuppressants will be mandated for 
all patients commencing at the week 3 visit. Outside of this time, patients will be instructed not 
to take these immunosuppressants unless instructed to do so by the investigator however 
availability of immunosuppressants will facilitate rapid initiation of treatment if necessary.  
Immunosuppressants to Suppress Immune Hepatitis 
The long-term use of high dose corticosteroids may be associated with the development of 
side effects. These include high blood pressure, elevation in blood sugar, weight gain with 
increased appetite and fluid retention. Osteoporosis is generally considered a long-term side 
effect and other bone complications include aseptic necrosis or pathological fractures. 
Cramps and joint pain have also been described. The occurrence of glaucoma and cataracts 
has been described in patients taking long-term corticosteroids. In addition, gastrointestinal 
irritation may occur and could result in gastrointestinal haemorrhage. Emotional disturbances 
and mood changes are also described.  
For patients entering the study with a history of hepatitis, there is a risk of disease reactivation 
associated with immune suppression. Monitoring for hepatitis B/C reactivation will be 
implemented for patients with a history of hepatitis B and C in parallel with the introduction of 
the immunosuppression regimen. 
The use of tacrolimus may be associated with the development of side effects. These include 
lymphoma and other malignancies, susceptibility to infections including polyoma virus and 
cytomegalovirus, nephrotoxicity, neurotoxicity, hyperkalaemia, hypertension, hyperglycaemia, 
insomnia, visual disturbances, headaches, hyperphosphatemia, tremor, myocardial 
hypertrophy, pure red cell aplasia, haemolytic uremic syndrome, posterior reversible 
encephalopathy syndrome. Use of live vaccines for immunisation should be avoided. 

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Patients should also avoid prolonged exposure to UV light and sunlight by wearing protective 
clothing and sunscreen with a high protection factor (minimum SPF30) and avoid grapefruit 
juice/ grapefruit during use of tacrolimus. 
The risk of side effects to the immunosuppressants will be minimised by careful monitoring, 
utilisation of the lowest effective dose, and tapering/cessation in the prophylaxis regimen and 
in response to break through transaminitis as soon as the evidence of immune hepatitis begins 
to subside.  
A management guideline for tacrolimus dosing is available in Appendix 3.  
Monitoring for CMV reactivation will be implemented for all patients positive for CMV (IgG) at 
screening in parallel with the introduction of the immunosuppression regimen.  In the instance 
of CMV reactivation a management guideline for CMV reactivation is available in Appendix 2. 
Patients with pre-existing contraindications to the immunosuppressants are excluded from the 
trial.  
Transduction of Extra-hepatic Tissues  
Following systemic administration of AAV-S3, there is potential for the spread of vector 
particles to non-hepatic tissues including the gonads. Therefore, shedding will be assessed in 
samples taken 3 times over the first 10 days of the study. Thereafter samples will be taken 
weekly until week 12 and then at visits 14, 16, 20 and 26 as required until no further evidence 
of FLT180a is detectable (as determined by 3 consecutive clear samples). As a precaution, 
only patients willing to practice a reliable barrier method of contraception during the study will 
be enrolled to avoid the possibility of horizontal, and potentially vertical transfer of vector.  
Development of Neutralising Anti-human FIX Antibodies  
A significant concern of this study, as with previous studies, is that research subjects may 
develop an immune response to FIX protein following systemic administration of FLT180a. In 
macaques, neutralising antibodies have been observed in a small proportion of animals 
exposed to human GTMP, including FLT180a. Investigations suggest that these antibodies 
are provoked by expression of a xenoprotein in non-human primates.1,13 However, several 
studies indicate that endogenous synthesis of FIX in the liver reduces the risk of developing 
inhibitors.8,9,10 To enhance the safety of AAV-mediated FIX expression, a liver-specific 
regulatory element (
) is contained within the expression cassette which limits transgene 
expression to hepatocytes.  
All patients eligible for the study will have received >150 exposure days to factor concentrates 
and are therefore less likely to develop inhibitors following AAV infusion and liver 
transduction.19  
Patients who have an inhibitor or a history of inhibitor after protein replacement therapy will be 
excluded from the study.  
 
 

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Insertional mutagenesis and/or tumorigenesis  
Insertional mutagenesis leading to oncogenesis is a recognised safety concern of 
vector-based gene therapy medicinal products. For vectors that do not efficiently integrate, 
such as AAV vectors modified to avoid integrations, insertions into the genome represent 
unintended and potentially rare events.  
Deep sequencing studies in murine and nonhuman primate models have shown that 
integration of the AAV genome can occur in the liver.20,21,22 Additionally, studies in mice have 
reported an increased incidence of hepatocellular carcinoma following perinatal or neonatal 
gene transfer.23,24 Studies in other murine models of bleeding diathesis however have failed to 
replicate this finding and collectively the available data in mice as well as larger animal models 
suggest that AAV has an extremely low risk of tumourigenesis.24,25  
Liver biopsies taken from six of nine adult patients (age range: 33-62 years) one year after 
receiving AAV2/5 hybrid vector (with a liver specific human α-1 antitrypsin promoter), for the 
treatment of acute intermittent porphyria, have shown a scarce random integration of vector at 
non-specific locations with no evidence of malignant transformation. Patients with acute of 
chronic liver disease were excluded from this study.26,27 
Conversely, AAV infection has been indicated as having a protective role against cancer.28 
Overall the risk of insertional mutagenesis following AAV-mediated gene transfer has been 
assessed to be low. Proviral DNA is maintained predominantly episomally in the transduced 
cell and the assessment of low risk is consistent with the fact that wild-type AAV infection in 
humans, although common, is not associated with oncogenesis. Patients will be followed for 
up to 15 years and will be fully informed regarding risk of mutagenesis prior to study entry.  
Risk of Blood Tests  
The risk of blood tests from a vein includes temporary discomfort at the site of puncture, 
possibly bruising and swelling around the puncture site, occasionally infection and bleeding 
into the surrounding muscle and tissues. These complications are likely to be exceedingly rare 
as many severe HB patients routinely self-administer factor concentrates IV without significant 
procedure related side effects. Standard procedures that are well established in 
comprehensive haemophilia treatment centres in Europe, Africa and North America will be 
followed to ensure the safe and uneventful administration of vector IV and during venepuncture 
for blood tests. Bleeding in the soft tissues around the venepuncture site will be treated with 
FIX concentrates, if necessary. 
Allergic Reactions or Anaphylaxis 
Allergic-type reactions, including anaphylaxis are a rare consequence of administration of 
biologicals. Onset of anaphylaxis is usually rapid following injection of an allergen with 90% 
beginning within 40 minutes.29 Patients should be informed of early symptoms and signs of 
hypersensitivity reactions, including hives, generalized urticaria, angioedema, chest tightness, 
dyspnoea, wheezing, faintness, hypotension, tachycardia, and anaphylaxis. Patients will 
remain under observation in the investigational centre for at least 12 hours following infusion 

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to minimise the risk associated with acute allergic reaction. Vital signs (pulse, blood pressure, 
respiration rate, and temperature) will be monitored at 15 minute intervals from the start of 
infusion. Vital signs will be monitored hourly for 6 hours following infusion and then every 2 
hours for 6 hours. 
Thrombogenicity 
The aim of the protocol is to reduce disease severity in HB patients dosed with FLT180a by 
increasing FIX expression. This is a first time in man protocol, and FIX expression 
approximations may have been underestimated.  
The expression cassette incorporates the gain-of-function Padua mutation. This is a naturally 
occurring mutation and originates from an Italian family in which, despite normal levels of FIX 
antigen, the hemizygous mother had 337% of the normal FIX activity level, a younger son (pre-
puberty) 551% of the normal activity level and an older son (23 years) 776% of the normal 
level of activity.30 Despite the very high FIX activities seen in this family, there was no family 
history of venous thrombosis until the older son (FIX activity 776% of normal) presented with 
a diagnosis of occlusive femoral–popliteal deep-vein thrombosis in the right leg.  
In the supplementary toxicology study conducted with FLT180a at a dose of 3.41 x 1013 vg/kg 
(50-fold greater than the starting dose in cohort 1) there were adverse histopathological 
findings in one animal that were likely associated with high hFIX levels (>1200%) and 
considered an exaggerated pharmacological effect.   
The World Federation of Hemophilia (WFH) regard 150% as the upper limit of normal for Factor 
IX activity. 
Should FIX activity achieved exceed the normal physiological range, there is a risk of 
thrombogenicity. The risk is considered to be very low based on the magnitude of FIX activity 
with apparent lack of symptomatology for many years in the family where the Padua mutation 
was first described and on studies in mice and dogs expressing the Padua mutation. In one 
study, sustained hFIX expression reached approximately 200% hFIX activity and 30% of 
antigen levels respectively with no clinical or laboratory evidence of thrombosis.31  
Patients will however be informed of early symptoms and signs of thrombotic phenomena, 
including pain and/or tenderness along a vein, swelling of an arm or leg without pain or 
tenderness, redness along a vein, low fever without any known reason (such as a cold or flu), 
sudden shortness of breath or difficulty breathing or coughing, sudden chest pain, sudden 
severe headache or changes in vision, and numbness or tingling in arms or legs. If such an 
event occurs whilst the patient is at home, the patient will be instructed to seek immediate 
medical care.  
In the instance of supratherapeutic FIX levels guidelines for management and 
thromboprophylaxis are included in section 12.3.3.  
Patients at high risk of thromboembolic events are excluded from the trial. 
  

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Myocarditis 
Two sets of toxicity studies with a caesium chloride purified vector preparation were performed 
in mice to support clinical trials with AAV2/8-LP1-hFIXco, the first-generation FIX gene therapy 
trial that were conducted by Nathwani et al. Animals in the first cohort received vector 
preparation without modification. These animals experienced both cardiac thrombosis and 
myocarditis. The second cohort received vector which was 10-fold concentrated. Thrombosis 
and myocarditis were again seen, but this cohort had pronounced hepatic necrosis. All the 
mice that exhibited toxicity had hFIX levels greater than 300 µg/mL, which is 60 times the 
upper limit of normal in humans (FIX activity approximately 6000%). Myocardial toxicity in 
transgenic mice secondary to high levels of FIX has been described previously.32 Toxicity in 
mice was only observed at very high levels of hFIX, therefore these effects are not expected 
to occur in study participants whose FIX levels are anticipated to remain in the normal range 
after gene transfer. Cardiac monitoring with electrocardiogram (ECG) is however implemented 
as part of the scheduled assessments and should be followed by echocardiogram if necessary. 
Risks associated with DNA impurities  
DNA impurities in AAV vector preparations, derived from production plasmids used for 
transient transfection, are well described.33,34,35 
The plasmid system used in manufacture of FLT180a reduces the likelihood of replication-
competent AAV (rcAAV) generation compared with previous products.  The incidence of rcAAV 
in FLT180a has been shown to be very low at less than 10 rcAAV in 1 x 1010 vector genomes, 
i.e. below the limit of detection of the validated assay method.  
Packaging of prokaryotic DNA impurities, including the kanamycin antibiotic resistance gene, 
derived from production plasmids has also been detected at levels ranging from 1.2 to 6.3% 
in AAV vectors generated by transfection of HEK293 cells or by helper-virus infection of stable 
producer cell lines. Kanamycin sequences were present in the FLT180a GMP batch 17-086 at 
a level of 0.74% vector genomes. No other batches will be used for this study.  
Therefore, the administration to humans of the clinical lot of FLT180a is not considered to 
result in toxicity due to the presence of DNA impurities. Their role in the initiation of liver 
transaminitis however cannot be excluded especially as bacterial DNA itself can generate an 
immune response in humans but we believe that transient immunosuppression with 
immunosuppressants will suppress liver cytotoxicity and preserve long-term transgene 
expression from hepatocytes. 
Unknown Risks Associated With First Time in Human ATIMP 
This is a first time in human clinical trial of an advanced therapy and as such there may be as 
yet uncharacterised risks that have not been evident in preclinical species. A comprehensive 
battery of safety assessments is included in the protocol but in order to mitigate against as yet 
unknown risks, the following additional steps will be taken. 
The first 2 patients at each dose level will remain as inpatients at the infusion centre for 
24 hours following dosing. All infusion centres will have access to emergency facilities.  

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An extended 6-week interval will be observed following dosing of the first patient in the study 
and prior to dosing the second patient to minimise risk to subsequent patients of unanticipated 
delayed AEs. 
5.4.3 
Assessment and Management of Risk 
This trial is categorised as: Type C = markedly higher than the risk of standard medical care.  
Management plans for occurrence of certain events described above can be found in Appendix 
1. 
 
6 
Objectives 
6.1 
Primary  
Safety 
To assess the safety of systemic administration of FLT180a in adults with HB at up to 
3 different dose cohorts. 
Efficacy 
To assess FIX levels following systemic administration of FLT180a, at the terminal dose level.  
6.2 
Secondary  
• 
To investigate the endogenous production of FIX following systemic administration of 
FLT180a at up to 3 different dose cohorts. 
• 
To investigate the effectiveness of a single administration of FLT180a on annualised 
bleeding rate and exogenous FIX consumption. 
• 
To assess the immune response to the FIX transgene product following systemic 
administration of FLT180a. 
• 
To assess viral shedding in various body fluids after systemic administration of FLT180a. 
6.3 
Exploratory 
• 
To assess the immune response to the AAV-S3 capsid proteins following systemic 
administration of FLT180a. 
Following a single administration of FLT180a: 
• 
To investigate the impact of endogenous production of FIX on functional status and 
disability in HB. 
• 
To investigate the impact of endogenous production of FIX on quality of life (QoL) in HB. 
• 
To investigate the impact of endogenous production of FIX on physical activity in HB. 
• 
To investigate the impact of endogenous production of FIX on haemophilia health status 
in HB. 
• 
To investigate the impact of endogenous production of FIX on joint health in HB. 

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• 
To investigate the impact of endogenous production of FIX on health resource utilisation 
in HB. 
 
7 
Trial design 
7.1 
Overall Design 
This is a phase I/II, open label, multicentre, ascending single dose, safety study of FLT180a in 
patients with severe HB.  
Patients will undergo the screening assessments described in Section 11.1.1 and the 
Schedule of Assessments (Table 1: Schedule of Assessments) up to 52 weeks prior to Study 
Day 0 (gene therapy infusion). Due to the risk of bleeding in this patient group, a washout from 
the patients FIX concentrate regimen is not mandated as part of this protocol. The investigator 
must however be able to demonstrate, from the patients’ medical records, a documented FIX 
activity level of <1% for severe patients or <2% for moderately severe patients. If (at the 
investigators discretion) a FIX concentrate washout is undertaken during the screening period, 
a minimum of 5 days washout is required. 
Treatment-eligible patients will report to the study site on the day prior to receiving the gene 
therapy infusion (Day -1, NB. Day -1 assessments may be conducted as early as Day -3 for 
logistical reasons). On Day 0, FLT180a will be administered as a single dose, slow IV infusion 
into a peripheral vein, and the patient will remain in the study centre for at least 12 hours and 
until the investigator has deemed the patient as fit to be discharged. The first 2 patients treated 
at each dose level will remain at the study centre for 24 hours following infusion prior to 
discharge.  
Patients who are on prophylactic therapy with FIX concentrates will remain on their usual 
dosing schedule and will be closely monitored for the FIX activity levels after screening and 
administration of FLT180a. If FIX activity levels ≥3% are reached then prophylaxis will be held 
pending a repeat analysis within a period of 72 hours. If the FIX activity levels are ≥3% at that 
time then prophylaxis will be stopped with continued/regular assessment of FIX activity levels 
and occurrence of spontaneous bleeding.  
Patients will be required to undergo study evaluations at intervals over the 26-week post 
treatment period. These will take place either at the study infusion site or at their normal 
haemophilia treatment centre. In order to monitor for shedding of vector genome sequences, 
patients will be required to provide plasma, saliva, urine, stool and semen samples until 
3 successive samples are shown to be clear. 
This is a first-in-human trial and as such an ascending-dose design has been implemented to 
enable dose evaluation in a step-wise manner. 3 dose cohorts of vector (low, intermediate, 
and high) will be tested in the dose escalation. Two patients will be tested at each dose level 
with an additional patient added in the event of a DLT (2 + 1 design). Dose escalation may 
occur provided there is no more than 1 DLT at any dose cohort and if the resulting FIX activity 
fails to reach the target level. A reduction of the dose level within a cohort may occur if FIX 

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activity exceeds defined levels in order to reduce the risk of overshooting the normal 
physiological range. Where a dose reduction occurs the 2 + 1 design will apply at that new 
dose level within the cohort. At the discretion of the Sponsor following advice from the trial 
management group (TMG) and independent data monitoring committee (DMC), additional 
patients may be added to any cohort to ensure adequate characterisation of either safety or 
the FIX response prior to dose escalation/reductions. The Sponsor, trial management group 
and data monitoring committee will select the terminal dose level based on the patient FIX 
activity levels with the aim of ensuring the majority of patients will reach a FIX activity within 
normal limits and in the absence of dose-limiting AEs, the terminal dose level will be expanded 
to 14 patients. This design minimises the number of patients that would need to be dosed at 
suboptimal levels whilst allowing evaluation of safety with the option to expand a group on 
observation of dose-limiting AEs. An extended 6-week interval will be observed between the 
first and second patient on study to monitor for any unanticipated delayed AEs. Subsequently, 
whilst dose escalation is ongoing, the study mandates a minimum 4-week interval between 
patients during which time efficacy and safety will be reviewed prior to a decision to dose the 
next patient. Once the terminal dose level is agreed and cohort expansion initiated, the dosing 
interval between patients will be reduced to 48 hours. 
The main risk in this study is considered to be a dose-dependent, asymptomatic increase in 
the serum ALT level associated with a decline in FIX levels, suggesting a loss of transduced 
hepatocytes. In the study conducted by Nathwani et al, transaminase elevations tended to 
occur between weeks 7-10 and were effectively managed by a short course of corticosteroids. 
In the study by George et al (NEJM 2017) the same asymptomatic increase in ALT requiring 
steroid treatment has been seen as early as 4 weeks post treatment with a dose of 5 x 1011 
vg/kg.14 In this study, all patients will be given a take-home pack of immunosuppressants 
following infusion and transaminase levels will be monitored closely. During the critical time 
period, from the week 3 visit, all patients will receive immunosuppressants, at all other times 
patients will be instructed to start the course of immunosuppressants only if told to do so by 
the investigator. 
The main efficacy endpoint will be based on an analysis of the proportion of patients achieving 
a clinical or normalised FIX response at 26 weeks. A clinical FIX response is defined as 
achieving a FIX activity of 5% to 150% of normal. Five percent has been selected as the 
threshold for a clinical FIX response because using gene therapy in haemophilia B patients to 
increase FIX activity from <1% to 5% has been shown to lead to a highly clinically significant 
improvement in annualised bleed rates and exogenous factor consumption.11,17 A normalised 
FIX response is defined as achieving a FIX activity level in the normal range (50-150% - per 
the World Federation of Haemophilia).36 The normal range has been selected as the threshold 
level for a normalised FIX response because reaching this level will modify the patient 
phenotype from severe at study start to normal at which point patients would not experience 
spontaneous bleeds.  
The choice of a 26-week endpoint is based on previous experience with AAV gene therapy for 
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therapy. Based on this, it is anticipated that the patients FIX activity will have reached a stable 
level by 26 weeks, thus this is an appropriate point at which to measure activity. 
7.2 
Endpoints 
7.2.1 
Primary Endpoints 
Safety 
Safety as assessed by the reporting of AEs according to Common Terminology Criteria for 
Adverse Events (CTCAE) version 5.0. 
Efficacy 
The following primary endpoints will be analysed: 
• 
The proportion of patients achieving clinical FIX response at Week 26, at the terminal 
dose level. A clinical FIX response is defined as achieving a FIX activity of 5% to 150%. 
• 
The proportion of patients also achieving normalised FIX response at Week 26, at the 
terminal dose level. A normalised FIX response is defined as achieving FIX activity in 
the normal range (50-150%).38 
7.2.2 
Secondary Endpoints 
Safety 
Safety as assessed by reporting of abnormal or change from baseline findings from routine 
safety assessments including, laboratory assessments, vital signs, ECG, physical exam and 
liver ultrasound. 
Endogenous FIX Production  
• 
The proportion of patients achieving FIX activity at or above 5%, 15%, 30%, 40%, 50% 
and 70% but no more than 150% of normal, at each scheduled visit. 
• 
The proportion of patients achieving FIX activity at or above 5%, 15%, 30%, 40%, 50%, 
70% and 150% of normal, at each scheduled visit. 
• 
Absolute change from baseline in FIX activity. 
Haemostatic Effectiveness 
• 
Change from baseline in annualised bleeding rate. 
• 
Change from baseline in FIX concentrate consumption. 
In order to ensure enough time has elapsed for the patient to have endogenous FIX activity to 
protect the patient from spontaneous bleeding episodes, the calculation period for haemostatic 
effectiveness will be from day 15 inclusive. 
Immune Response 
Immune response to the FIX transgene product (i.e., development of inhibitors) will be 
assessed by measurement of the level of inhibitors. 

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Shedding 
Clearance of vg in plasma, saliva, urine, stool, and semen. 
7.2.3 
Exploratory Endpoints 
Haemostatic Effectiveness 
Exploration of the correlation between FIX levels and bleeding events over time. 
Immune Response 
• 
Immune response to the AAV-S3 capsid will be assessed by measurement of the S3 
neutralising antibody titre. 
• 
T-cell responses to AAV-S3 capsid in peripheral blood mononuclear cells. 
Disability Status 
Change from baseline in World Health Organization Disability Assessment Schedule 2.0 
(WHODAS 2.0) score. 
Physical Activity  
Change from baseline in Haemophilia Activities List (HAL) 2005. 
Health Related Quality of Life  
Change from baseline in the EQ-5D-5L and the Haem-A-QoL score. 
Haemophilia Health Status 
Change from baseline in the PROBE score. 
Assessment of Joint Health/Function  
Change from baseline in the Haemophilia Joint Health Score (HJHS). 
Health Resource Utilisation 
• 
Number of haemophilia related medical appointments and medical activities. 
• 
Number of visits at site. 
• 
Number of emergency room visits. 
• 
Number of hospitalisations related to haemophilia. 
• 
Length of hospital stay. 
• 
Number of days lost from education or work by patients and caregivers due to bleeding 
episodes. 
• 
Number of physiotherapy sessions, specialist consultations and appointments with 
professional caregivers. 
 

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8 
Selection of Patients 
Patient enrolment at a site will only commence once the trial has: 
• 
documented Ethics Committee, Competent Authority and Local Institution approval. 
• 
been initiated on behalf of the sponsor. 
• 
been issued with a site activation letter on behalf of the sponsor. 
Patients may only be enrolled at approved trial sites.  
8.1 
Inclusion Criteria 
1. Adults males, ≥ 18 years of age.  
2. Confirmed diagnosis of HB defined as one of the following:  
(a) Documented severe FIX deficiency with plasma FIX activity of <1% of normal, 
or 
(b) moderately severe FIX deficiency with plasma FIX activity level between ≥1% 
and ≤2% and a severe bleeding phenotype defined by one of the following:  
i. On prophylaxis for a history of bleeding, or  
ii. On demand therapy with a history of 4 or more bleeding episodes/year 
on average over the past 3 years, or  
iii. evidence of chronic haemophilic arthropathy (pain, joint destruction, and 
loss of range of motion). 
3. Able to give full informed consent and able to comply with all requirements of the trial 
including 15-year long-term follow-up.  
4. Willing to practice barrier contraception until at least 3 consecutive semen samples after 
vector administration are negative for vector sequences.  
5. Lack of neutralising anti-AAV-S3 antibodies using an in vivo transduction inhibition assay 
within 4 weeks of vector administration.  
6. At least 150 exposure days to FIX concentrates. 
8.2 
Exclusion Criteria 
1. Presence of neutralising antihuman FIX antibodies (inhibitor, determined by the Bethesda 
inhibitor assay) at the time of enrolment or a previous history of FIX inhibitor.  
2. Patients at high risk of thromboembolic events (high risk patients would include those with 
a history of arterial or venous thromboembolism (e.g. deep vein thrombosis, pulmonary 
embolism, non-haemorrhagic stroke, arterial embolus) and those with acquired 
thrombophilia including conditions such as atrial fibrilation). 
3. Use of investigational therapy for haemophilia within 30 days before enrolment.  

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4. Patients with active hepatitis B or C, and HBsAg or hepatitis C virus (HCV) RNA viral load 
positivity, respectively, or currently on antiviral therapy for hepatitis B or C. Negative viral 
assays in 2 samples, collected at least 6 months apart, will be required to be considered 
negative. Both natural clearers and those who have cleared HCV on antiviral therapy are 
eligible. 
5. Serological evidence of HIV-1.  
6. Evidence of liver dysfunction (persistently elevated alanine aminotransferase, aspartate 
aminotransferase, bilirubin >1.5 x upper limit of normal). 
7. Platelet count <50 x 109/L.  
8. Uncontrolled glaucoma, diabetes mellitus, or hypertension.  
9. Malignancy requiring treatment.  
10. Patients with uncontrolled cardiac failure, unstable angina or myocardial infarction in the 
past 6 months.  
11. Poor performance status (World Health Organization score >1). 
12. Prior treatment with any gene transfer medicinal product. 
13. Known or suspected intolerance, hypersensitivity or contrainidication to the investigational 
product and non-investigational medicinal products or their excipients. 
14. Planned major elective surgery prior to the end of trial. 
15. Current or relavant history of a physical or psychiatric illness or any medicial condition that 
in the opinion of the investigator could affect the patients safety or interfere with the study 
assessments. 
16. CMV IgG postive patients who are CMV PCR positive at screening. 
 
9 
Investigational Medicinal Products and Non-Investigational Medicinal 
Products 
9.1 
Name and Description of ATIMP(s) and IMP(s) 
FLT180a 
(AAV2/S3-
-Ti-FIXco1) 
is 
a 
replication-incompetent 
single-stranded 
recombinant AAV vector. The vector is composed of a single-stranded DNA genome 
encapsidated in an AAV-derived protein capsid. FLT180a possesses 3 novel features:  
(1) 
A new small synthetic liver promoter (
) 
(2) 
An enhanced transgene including 
a. codon optimised FIX gene 
b. a gain-of-function Padua mutation (R338L)  
c. truncated intron in a natural position 

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(3) 
A novel engineered capsid (AAV-S3) developed from wild-type AAV3 and AAV8 through 
capsid shuffling. This enables the vector to transduce human hepatocytes with high 
efficiency. 
FLT180a is formulated 
 
 The drug product is presented as a sterile, aqueous 
concentrate for solution for infusion, supplied in 10 mL vials. Each vial contains 5 mL of 
FLT180a Concentrate for Solution for Infusion. The vials are sealed with rubber stoppers and 
aluminium seals with plastic flip tops. 
FLT180a Concentrate for Solution for Infusion is stored at <-60oC and must remain frozen until 
thawed prior to use. 
FLT180a is classified as an advanced therapy investigational medicinal product (ATIMP) and 
specifically a gene therapy medicinal product. 
9.2 
Source of ATIMP/IMP, Manufacture, Distribution and Storage 
9.2.1 
Manufacture 
FLT180a Concentrate for Solution for Infusion is manufactured according to Good 
Manufacturing Practice (GMP) by: 
Children’s GMP LLC  
St. Jude Children’s Research Hospital  
Memphis, Tennessee 
USA 
 
An audit of Children’s GMP LLC has been performed by a Qualified Person (QP) from the 
Clinical Biotechnology Centre who are responsible for importation into the UK and QP 
certification (Section 9.2.3). 
9.2.2 
Manufacture and Shipment 
A clinical batch of FLT180a Concentrate for Solution for Infusion will be manufactured 
according to GMP and tested according to a product specification. The genomic titre will be 
determined as part of the product release testing and will be stated on the Certificate of 
Analysis and included on the product labelling. The genomic titre stated on the labelling and 
Certificate of Analysis should be used for all dose calculations.  
Investigational medicinal product will be labelled in accordance with local regulatory 
requirements. 
Shipment of the IMP will be undertaken by a company specialising in cold chain transportation 
of biopharmaceuticals. Vials will be maintained at <-60oC during shipment. All shipments will 
include temperature loggers so that the temperature control during transit can be confirmed. 

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9.2.3 
Importation into the UK, QP Certification and Storage Prior to Shipment to 
Investigational Site 
Importation of the IMP into the UK will be performed by: 
Clinical Biotechnology Centre (CBC) 
University of Bristol 
Langford House  
Lower Langford 
Bristol 
BS40 5DU 
 
Qualified Person (QP) certification of the batch will be performed by an appropriately 
experienced QP from CBC. The batch manufacturing record, test results and the certificate of 
analysis for the clinical batch will be provided to the QP for the purposes of batch certification.  
In the event that IMP cannot be certified by the QP, it will be retained under quarantine at CBC. 
CBC will also perform secondary packaging and labelling in the UK, including labelling with 
expiry date. Investigational medicinal product will be stored at CBC at <-60oC until shipped to 
the investigational sites for administration.  
9.2.4 
Shipment of IMP to Infusion Sites 
FLT180a Concentrate for Solution for Infusion will be shipped from CBC to the UK infusion 
site. The appropriate number of vials required for dosing will be determined based on the 
weight of each patient. The vials will be shipped to the infusion site prior to the scheduled 
dosing day by a company specialising in cold chain transportation of biopharmaceuticals. All 
shipments will contain temperature monitoring devices.  
FLT180a Concentrate for Solution for Infusion required for dosing in the US will be retained at 
the site of manufacturing which is also the infusion site. No shipment is therefore required. 
9.2.5 
Infusion Sites 
Administration of IMP will be restricted to centralised infusion sites. 
 
Royal Free Hospital 
Pond Street  
Hampstead 
London NW3 2QG 
 
St. Jude Children’s Research Hospital 
262 Danny Thomas Place 
Memphis, TN 38105  
USA 
 
9.2.6 
Receipt Storage and Handling of ATIMP(s)/IMP(s) at Site 
All IMP aspects of the trial at participating sites are the responsibility of the Principal 
Investigator (PI), who may delegate this responsibility to the local pharmacist or other 

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appropriately trained personnel. The delegation of duties must be recorded on the Staff 
Signature and Delegation of Tasks Form. 
FLT180a will be received at the investigational site by the PI or designee. The vials will be 
inspected to ensure that they have remained frozen in transit and will be transferred to a 
restricted access freezer maintaining a temperature of <-60oC. 
Once predose eligibility checks have been confirmed on Day 0, the vector will be thawed and 
diluted. All procedures will be performed in a standard biological safety cabinet which has been 
cleaned by each site’s standard cleaning procedure.  
Detailed instructions on receipt, storage and handling of FLT180a Concentrate for Solution for 
Infusion are contained in the ATIMP management plan. Instructions for the preparation of 
FLT180a infusion solutions are also included in the ATIMP management plan. All local 
requirements related to activities involving genetically modified organisms will be adhered to.    
9.2.7 
Accountability and Traceability of ATIMP(s)/IMP(s) 
There is a system set-up to ensure the traceability of each ATIMP from the starting material, 
through to administration to the participant and destruction or final transfer. A comprehensive 
ATIMP management plan and associated Standard Operating Procedures and forms are in 
place to ensure that the required accountability and traceability data is collected and retained.  
The requirement for the manufacturer and Investigator/clinical trial site(s) to retain their part of 
the traceability information is set out in the relevant contractual agreements with the Sponsor. 
9.3 
Name and Description of Non-Investigational Medicinal Product  
Immunosuppressants will be provided as commercially available locally. 
Prednisolone and Prednisone are used in the same manner and are equally effective. 
Therefore, the recommended schedule for prednisolone, equally applies to prednisone. 
Prednisolone (prednisone), methylprednisolone and tacrolimus are considered to be non-
investigational medicinal product (NIMP) in this trial and will be sourced from stocks at the 
infusion centre or local haemophilia treatment centre. 
Investigational sites are responsible for maintaining a system which allows adequate 
reconstruction of NIMP movements and evaluation of patient compliance. 
Risks associated with use of NIMP as described in their Summary of Product Characteristics 
should be communicated to the patients by the investigator. 
9.4 
Implementation of the Immunosuppressants Regimen 
Patients will be instructed to only take treatment as instructed by the investigator. 
  
 
 
 
 
 

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If break through transaminitis occurs, follow the instructions below. 
Break through transaminitis 
• 
When a patient has an elevation in ALT beyond the upper limit of normal (ULN) or the 
trends of LFT and/or Factor IX levels suggest potential transaminitis or a lack of 
response to the immunosuppression regimen: 
o Evaluate to exclude other causes of transaminitis. 
o Assume it is vector induced transaminitis after reasonable attempts to exclude 
other causes. Record/report the event accordingly in the source documentation 
and eCRF. 
o Contact the Chief Investigator immediately. 
o Administer methylprednisolone 1g IV for 3 days; 
Check tacrolimus levels and if required adjust dose to ensure therapeutic levels. 
If tacrolimus is not currently in use, initiate immediately. Management 
guidelines for tacrolimus dosing are contained within Appendix 3. 

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Following methylprednisolone give prednisolone at 1mg/kg/day. 
o Monitor LFTs daily initially to establish response to treatment by evaluating the 
trend in ALT levels. Once a response to treatment has been established 
continue prednisolone 1mg/kg/day with regular monitoring of the LFTs, two or 
3 times a week, until the LFTs are within normal range and continue for an 
additional week and then initiate prednisolone taper as described above. 
Tacrolimus can be discontinued at any time in consultation with the Chief 
Investigator and Medical Monitor. 
• 
The Investigator, in consultation with the Chief Investigator and Medical Monitor, may 
use clinical judgement to assess the appropriate course of treatment and monitoring in 
response to ANY rise in ALT or if any other cause (suspected or confirmed) is identified. 
 
At all times, Investigators should be vigilant of a downward trend in FIX activity level, which 
alongside an elevation in ALT, may be an early indicator of transaminitis. 
 
Where there is contact or consultation with the Chief Investigator, the Sponsor should be kept 
informed. 
 
10 
Concomitant Medication 
All non-study treatment received within 30 days prior of initial informed consent and for the 
duration of the study must be recorded on the appropriate electronic case report form (eCRF) 
page. Concomitant medication includes prescription and non-prescription medication and 
herbal treatments.  
10.1 
Prior Treatment 
Prior treatment includes all treatment received from 30 days prior to the initial informed consent 
up until the time of FLT180a infusion. Prior treatment information must be recorded on the 
appropriate eCRF page.  
10.2 
Concomitant Treatment  
Concomitant treatment refers to all treatment taken between the date of investigational product 
infusion and the week 26/end of study visit, inclusive. All concomitant treatments information 
must be recorded on the appropriate eCRF page. 
Patients should be instructed not to start taking any new medications, including non-
prescription drugs and herbal preparations, unless they have received permission from the 
investigator, time allowing, i.e. investigator permission not required for emergency medical 
care. 
Other therapy considered necessary for the patient’s welfare may be given at the discretion of 
the investigator. All such therapy must be recorded in the eCRF. No other IMP may be used 

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concomitantly with the study treatment. The patients are not allowed to participate concurrently 
in another clinical study. 
10.3 
Withdrawal From Existing FIX Concentrate Therapy 
Patients who are on prophylactic therapy with FIX concentrates will remain on their usual 
dosing schedule and will be closely monitored for the FIX activity levels after screening and 
administration of FLT180a. If FIX activity levels ≥3% are reached then prophylaxis will be held 
pending a repeat analysis within a period of 72 hours. If the FIX activity levels are ≥3% at that 
time then prophylaxis will be stopped with continued/regular assessment of FIX activity levels 
and occurrence of spontaneous bleeding. 
 
11 
Study Procedures  
11.1 
Study Schedule 
11.1.1 
Screening 
The following trial-specific procedures will be carried out after the initial informed consent form 
(ICF) is signed to assess the participant’s eligibility. Screening evaluations may be performed 
at the infusion study centre or in the patient’s haemophilia treatment centre. A window of up to 
52 weeks (from initial consent to day -1) is allowed for screening to allow for controlled 
prospective collection of bleeding event and FIX consumption data via the patient diary. A 
patient can move forward to Day -1 assessments and dosing as dosing slots are available. 
There is no minimum time frame for the data collection. ALL screening assessments should 
be completed in a timely manner (~2 weeks) following consent to confirm patient eligibility and 
may take place across multiple visits to complete all assessments. During screening the patient 
should be contacted ~every 4 weeks to check on diary completion and collect details on any 
AEs. A repeat AAV Antibody screen is mandated within 4 weeks of dosing. If the screening 
period for a patient is longer than 16 weeks then a repeat of the central laboratory screening 
bloods (FIX Antigen/ HBV, HCV, HIV, CMV Screen/ Haematology, Chemistry incl. CRP, 
Coagulation Screen/ Liver Function Test/ FIX Activity Level/ FIX Inhibitor Level) will be required 
within 4 weeks of dosing and the patient reported outcomes should also be repeated at Day -
1. Patients may be rescreened should this time window elapse. 
• 
Demographics will be recorded. 
• 
Medical history will be documented. 
• 
WHO performance status will be documented. 
• 
Prior and concomitant medication will be documented. 
• 
A bleeding and FIX concentrate utilisation history will be documented (3-year history) 
and any target joints will be recorded. 
• 
A physical examination will be conducted including measurement of the patient’s height 
and weight. Waist circumference, hip circumference, neck circumference and 
bioimpedance will also be measured. 

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• 
A 12-lead ECG will be conducted. 
• 
Vital signs (pulse, blood pressure) will be measured. 
• 
A clinical assessment of joints will be undertaken using the HJHS. 
• 
A liver ultrasound scan will be conducted. 
• 
AAV-S3 antibody screening will be conducted. Note: A blood sample drawn within 
4 weeks of dosing must be confirmed as negative before a patient can be considered for 
dosing. It is acceptable to take 2 samples during the screening period. Should there be 
any problems with the testing it is acceptable to take repeat samples as necessary. 
• 
Blood samples will be taken for HIV, HCV (anti-HCV antibodies and HCV RNA viral load, 
HCV RNA viral load only indicated for patients with history of HCV and positive HCV 
antibody test), HBV, CMV (CMV IgG test and CMV PCR; CMV PCR only indicated for 
patients with positive CMV IgG) screening. Note: a repeat blood sample drawn within 4 
weeks of dosing for testing will required if the patient is in the screening period for longer 
than 16 weeks. 
• 
Blood samples will be taken for haematology, chemistry incl. CRP, coagulation screen 
and LFTs (central lab). Note: a repeat blood sample drawn within 4 weeks of dosing for 
testing will required if the patient is in the screening period for longer than 16 weeks. 
• 
FIX activity level (trough). 
- 
For patients on on-demand therapy, a blood sample should be taken following 5 days 
free from factor concentrates. 
- 
For patients on conventional prophylactic therapy, a blood sample should be taken at 
the end of the longest acceptable interval between doses. 
- 
For patients taking EHL prophylactic therapy, a blood sample should be taken within 
the 48-hour period before the next planned dose. 
• 
Baseline FIX activity (following 5 days washout or obtained from documented medical 
records). 
• 
A blood sample will be taken to evaluate FIX inhibitor level. Note: a repeat blood sample 
drawn within 4 weeks of dosing for testing will required if the patient is in the screening 
period for longer than 16 weeks. 
• 
A blood sample will be taken to evaluate FIX antigen level. Note: a repeat blood sample 
drawn within 4 weeks of dosing for testing will required if the patient is in the screening 
period for longer than 16 weeks. 
• 
FIX genotype will be recorded from the patients’ medical notes. (if not available this test 
may be conducted as part of the trial) 
• 
Quality of life will be evaluated (EQ-5D-5L and Haem-A-QoL). 
• 
Disability status will be assessed (WHODAS 2.0). 

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• 
Physical activity status will be assessed (HAL 2005). 
• 
Haemophilia health status will be assessed (PROBE). 
• 
Health resource utilisation will be documented (6-month history). 
• 
A bleeding/FIX consumption diary will be issued to the patient with instructions on 
completion. 
• 
Adverse events will be recorded during the screening period from the time of initial 
consent. 
ALT levels will guide treatment for breakthrough transaminitis during the study and alcohol 
intake can cause ALT levels to rise, making it difficult to determine whether transaminitis is 
alcohol or vector related. As such, a patient’s alcohol intake for the course of the trial should 
also be discussed at the screening visit with a recommendation to moderate their intake for 
the duration of the trial. Further, patients will be advised to abstain from alcohol from 1 week 
prior to vector infusion to approximately 3 months following vector infusion. Discussion around 
a patient’s alcohol intake should be ongoing throughout the trial. 
11.1.2 
Vector Infusion (Day -1 to Day +4) 
Vector infusion will only occur once all screening assessments have been conducted and the 
patient is deemed to be eligible for the study. Eligible patients will attend a qualified infusion 
centre prior to the day of vector infusion and an additional ‘pre-infusion’ dosing site informed 
consent will be obtained from each patient. On the day prior to vector infusion, local safety 
laboratory tests will be utilised for management of patients to ensure rapid turnaround of 
clinical results. Duplicate safety blood samples will be drawn at day -1 to provide a baseline 
for both central and local laboratories. For all consenting patients the following procedures will 
be undertaken: 
Day -1 
• 
A second informed consent will be taken – this may occur prior to Day -1. 
• 
A physical examination will be conducted (including confirmation of weight). Waist 
circumference, hip circumference, neck circumference and bioimpedance will also be 
measured. 
• 
Vital signs (pulse, blood pressure, respiration rate, and temperature) will be measured. 
• 
A 12-lead ECG will be conducted. 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab). For list of required tests please see Table 2: Detailed Schedule of 
Assessments for Infusion Week and Section 11.4.3.2. 
• 
Blood samples will be taken for haematology, chemistry incl. CRP, coagulation screen 
and LFTs (central lab). 
• 
Blood will be taken for evaluation of FIX activity level (sample for both local and central 
lab). 

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• 
Blood will be taken for collection of peripheral blood mononuclear cells (Elispot and 
Research). The mononuclear (Research) sample is optional. 
• 
A blood sample will be taken and stored. This sample may be used to retrospectively 
analyse AAV-S3 antibody status immediately prior to dosing.  
• 
PCR of vg in plasma, saliva, urine, stool, and semen. 
• 
Adverse event and concomitant medication status will be recorded. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  
• 
Plasma samples will be drawn and frozen for research purposes. These samples are 
optional. 
• 
Plasma samples will be drawn and frozen for immune response research purposes. 
If the screening period for a patient is longer than 16 weeks then the following will also be 
completed on Day -1: 
• 
Quality of life will be evaluated (EQ-5D-5L and Haem-A-QoL). 
• 
Disability status will be assessed (WHODAS 2.0). 
• 
Physical activity status will be assessed (HAL 2005). 
• 
Haemophilia health status will be assessed (PROBE). 
• 
Health resource utilisation will be documented (since screening). 
Day 0 
• 
Intravenous catheter insertion into a suitable peripheral vein (e.g., the median cubital 
vein). 
• 
The IV catheter will be flushed with saline. 
• 
The vector will be thawed and diluted in accordance with instructions in the ATIMP 
management plan. 
• 
Prepared drug will be kept at room temperature prior to administration.  
• 
At approximately 1 hour prior to vector infusion, the investigator or designee will assess 
the patient’s vital signs and blood samples will be taken for haematology, chemistry incl. 
CRP, coagulation screen and LFTs (local lab). For list of required tests please see Table 
2: Detailed Schedule of Assessments for Infusion Week and Section 11.4.3.2. Blood will 
also be taken for evaluation of FIX activity level (sample for both local and central lab). 
• 
The calculated FLT180a vector dose will be infused over 1 hour through the catheter 
using an appropriate infusion pump. 
• 
Vital signs (pulse, blood pressure, respiration rate, and temperature) will be monitored at 
15 minute intervals (+/-5 mins) from the start to completion of infusion.  
• 
On completion of the infusion, the catheter will again be flushed with saline.  

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• 
Vital signs will be monitored hourly for 6 hours (+/- 10 mins) from the end of infusion and 
then every 2 hours (+/-15 mins) for a further 6 hours.  
• 
Laboratory safety blood samples, haematology, chemistry incl. CRP, coagulation screen 
and LFTs will be repeated at 8 hours after the end of the infusion. For list of required 
tests please see Table 2: Detailed Schedule of Assessments for Infusion Week and 
Section 11.4.3.2. 
• 
Adverse event and concomitant medication status will be recorded. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  
• 
Patients will remain hospitalised for 12 hours to observe for any immediate toxicity of the 
procedure. If the vital signs are stable, the catheter will be removed prior to patient 
discharge from the centre. 
• 
The first 2 patients at each dose level will remain at the study centre for 24 hours following 
infusion with additional vital signs monitoring at 16 and 20 hours (+/-15 mins) following 
infusion. 
• 
All patients will be discharged with a medical alert card. 
Day +1, +2, and +4 
During the first week following infusion, evaluations will be performed exclusively at the study 
centre where the participant received their vector infusion by the investigator or designee.  
At each visit: 
• 
A physical examination will be conducted. 
• 
Vital signs (pulse, blood pressure, respiration rate, and temperature) will be measured. 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab). For list of required tests please see Table 2: Detailed Schedule of 
Assessments for Infusion Week and Section 11.4.3.2. 
• 
Blood will be taken for haematology, chemistry incl. CRP and coagulation screen (central 
lab) on day 2 and 4.  
• 
Blood will be taken for evaluation of FIX activity level on day 2 and 4 (local lab). 
• 
Adverse event and concomitant medication status will be recorded. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  
• 
Plasma, saliva, urine, stool and semen samples will be taken for polymerase chain 
reaction (PCR) of vg on day 2 and 4.  
• 
In addition, on Day +1, a 12-lead ECG will be conducted. 

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11.2 
Follow-up Visits (Weeks 1-25) 
After the first week, evaluations may be performed in the patient’s regular haemophilia 
treatment centre or at the patient’s home (in the case of routine blood sampling and vital signs 
monitoring). 
11.2.1 
Week 1-5 Assessments 
The following assessments will be conducted at the main visit each week (or as specified): 
• 
A physical examination will be conducted. 
• 
Vital signs (pulse, blood pressure) will be measured. 
• 
A 12-lead ECG will be conducted at week 2 and 4. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(central lab). 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab). 
• 
Blood will be taken for collection of mononuclear cells for immune studies (Elispot). 
• 
Blood will be taken for evaluation of FIX activity levels (central and local lab). 
• 
Blood will be taken for evaluation of FIX inhibitor levels (weeks 1, 2, 3). 
• 
Commencing at the week 3 visit, patients will receive a course of immunosuppressants 
(see Section 9.4).  
• 
For any patients with a history of HBV and HCV, local testing for hepatitis B or C 
reactivation should be conducted at week 4. 
• 
From week 3 CMV testing (CMV PCR; in patients with CMV positive (IgG) at screening) 
should be conducted weekly in conjunction with immunosuppressant dosing. 
Management guidelines in the case of CMV reactivation can be found in Appendix 2. 
• 
From week 3 tacrolimus levels should be measured at each blood draw until therapeutic 
levels are established and weekly thereafter in conjunction with immunosuppressant 
dosing. Management guidelines for tacrolimus dosing can be found in Appendix 3. 
• 
Blood will be taken for evaluation of AAV-S3 antibody titre (weeks 1, 2). 
• 
Plasma, saliva, urine, stool, and semen samples will be taken for PCR of vg (three times 
within 7-10 days of vector infusion and weekly thereafter). These collections will be done 
unless there have been 3 prior consecutive specimens that were negative.  
• 
Adverse event and concomitant medication status will be recorded at each visit. 
• 
Plasma samples will be drawn and frozen for research purposes. These samples are 
optional. 

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• 
Plasma samples will be drawn and frozen for FIX activity assay research purposes 
(weeks 1 and 4). 
• 
Plasma samples will be drawn and frozen for immune response research purposes only 
during the course of investigations or treatment of breakthrough transaminitis. 
Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 2 
further separate occasions each week during week 1-5 (see Section 11.2.10).  
11.2.2 
Week 6-12 Assessments 
The following assessments will be conducted at the main visit each week (or as specified): 
• 
A physical examination will be conducted. 
• 
Vital signs (pulse, blood pressure) will be measured. 
• 
A 12-lead ECG will be conducted at week 8. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(central lab). 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab). 
• 
Blood will be taken for collection of mononuclear cells for immune studies (Elispot). 
• 
Blood will be taken for evaluation of FIX activity levels (central and local lab). 
• 
Blood will be taken for evaluation of FIX antigen levels (week 12). 
• 
Blood will be taken for evaluation of FIX inhibitor levels (weeks 6, 9, 12). 
• 
Patients will continue to receive a course of immunosuppressants (see Section 9.4).  
• 
For any patients with a history of HBV and HCV, local testing for hepatitis B or C 
reactivation should be conducted at week 6, 8, 10, 12.  
• 
CMV testing (CMV PCR; in patients with CMV positive (IgG) at screening) should be 
conducted weekly in conjunction with immunosuppressant dosing. Management 
guidelines in the case of CMV reactivation can be found in Appendix 2. 
• 
From week 3 tacrolimus levels should be measured at each blood draw until therapeutic 
levels are established and weekly thereafter in conjunction with immunosuppressant 
dosing. Management guidelines for tacrolimus dosing can be found in Appendix 3. 
• 
Blood will be taken for evaluation of AAV-S3 antibody titre (week 6, 12). 
• 
Plasma, saliva, urine, stool, and semen samples will be taken for PCR of vg weekly. 
These collections will be done unless there have been 3 prior consecutive specimens 
that were negative.  
• 
Adverse event and concomitant medication status will be recorded at each visit. 

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• 
Plasma samples will be drawn and frozen for research purposes. These samples are 
optional. 
• 
Plasma samples will be drawn and frozen for immune response research purposes only 
during the course of investigations or treatment of breakthrough transaminitis. 
Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 1 
further separate occasion each week during week 6-12 (see Section 11.2.10). 
11.2.3 
Week 13 Assessments 
• 
Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 
2 separate occasions during week 13 (see Section 11.2.10). Blood draws may be taken 
either at study site or at alternative location (e.g. patient’s home). The intention should 
be for at least one of the two blood draws per week to be conducted at study site. 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab), once in week 13. Blood draw may be taken either at study site or at alternative 
location (e.g. patient’s home). 
• 
CMV testing (CMV PCR; in patients with CMV positive (IgG) at screening) should be 
conducted in conjunction with immunosuppressant dosing, as required. 
• 
Tacrolimus level should be conducted in conjunction with immunosuppressant dosing, 
as required. 
• 
Adverse event and concomitant medication status will be recorded and ongoing diary 
completion assessed. 
11.2.4 
Week 14 Assessments 
The following assessments will be conducted at the main visit: 
• 
A physical examination will be conducted. 
• 
Vital signs (pulse, blood pressure) will be measured. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(central lab). 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab). 
• 
Blood will be taken for collection of mononuclear cells for immune studies (Elispot). 
• 
Blood will be taken for evaluation of FIX activity levels (central and local lab). 
• 
Patients will continue to receive a course of immunosuppressants (see Section 9.4).  
• 
For any patients with a history of HBV and HCV, local testing for hepatitis B or C 
reactivation should be conducted.  

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• 
CMV testing (CMV PCR; in patients with CMV positive (IgG) at screening) should be 
conducted in conjunction with immunosuppressant dosing. Management guidelines in 
the case of CMV reactivation can be found in Appendix 2. 
• 
Tacrolimus levels should be measured in conjunction with immunosuppressant dosing. 
Management guidelines for tacrolimus dosing can be found in Appendix 3. 
• 
Plasma, saliva, urine, stool, and semen samples will be taken for PCR of vg. These 
collections will be done unless there have been 3 prior consecutive specimens that were 
negative.  
• 
Adverse event and concomitant medication status will be recorded at each visit. 
• 
Plasma samples will be drawn and frozen for research purposes. These samples are 
optional. 
• 
Plasma samples will be drawn and frozen for immune response research purposes only 
during the course of investigations or treatment of breakthrough transaminitis. 
Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 1 
further separate occasion during week 14 (see Section 11.2.10). 
11.2.5 
Week 15 Assessments 
• 
Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 
2 separate occasions during week 15 (see Section 11.2.10). Blood draws may be taken 
either at study site or at alternative location (e.g. patient’s home). The intention should 
be for at least one of the two blood draws per week to be conducted at study site. 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab), once in week 15. Blood draw may be taken either at study site or at alternative 
location (e.g. patient’s home). 
• 
CMV testing (CMV PCR; in patients with CMV positive (IgG) at screening) should be 
conducted in conjunction with immunosuppressant dosing, as required. 
• 
Tacrolimus level should be conducted in conjunction with immunosuppressant dosing, 
as required. 
• 
Adverse event and concomitant medication status will be recorded and ongoing diary 
completion assessed. 
11.2.6 
Week 16 Assessments 
The following assessments will be conducted at the main visit: 
• 
A physical examination will be conducted. 
• 
Vital signs (pulse, blood pressure) will be measured. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  

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• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(central lab). 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab). 
• 
Blood will be taken for collection of mononuclear cells for immune studies (Elispot). 
• 
Blood will be taken for evaluation of FIX activity levels (central and local lab). 
• 
Blood will be taken for evaluation of FIX inhibitor levels. 
• 
Patients will continue to receive a course of immunosuppressants (see Section 9.4).  
• 
For any patients with a history of HBV and HCV, local testing for hepatitis B or C 
reactivation should be conducted.  
• 
CMV testing (CMV PCR; in patients with CMV positive (IgG) at screening) should be 
conducted in conjunction with immunosuppressant dosing. Management guidelines in 
the case of CMV reactivation can be found in Appendix 2. 
• 
Tacrolimus levels should be measured in conjunction with immunosuppressant dosing. 
Management guidelines for tacrolimus dosing can be found in Appendix 3. 
• 
Plasma, saliva, urine, stool, and semen samples will be taken for PCR of vg. These 
collections will be done unless there have been 3 prior consecutive specimens that were 
negative.  
• 
Adverse event and concomitant medication status will be recorded at each visit. 
• 
Plasma samples will be drawn and frozen for research purposes. These samples are 
optional. 
• 
Plasma samples will be drawn and frozen for immune response research purposes only 
during the course of investigations or treatment of breakthrough transaminitis. 
Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 1 
further separate occasion during week 16 (see Section 11.2.10). 
11.2.7 
Week 17-19 Assessments 
• 
Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 
2 separate occasions each week during week 17-19 (see Section 11.2.10). Blood draws 
may be taken either at study site or at alternative location (e.g. patient’s home). The 
intention should be for at least one of the two blood draws per week to be conducted at 
study site. 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab), once a week, weeks 17-19. Blood draw may be taken either at study site or 
at alternative location (e.g. patient’s home). 
• 
For any patients with a history of HBV and HCV, local testing for hepatitis B or C 
reactivation should be conducted at week 18. 

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• 
CMV testing (CMV PCR; in patients with CMV positive (IgG) at screening) should be 
conducted in conjunction with immunosuppressant dosing, as required. 
• 
Tacrolimus level should be conducted in conjunction with immunosuppressant dosing, 
as required. 
• 
Adverse event and concomitant medication status will be recorded and ongoing diary 
completion assessed. 
11.2.8 
Week 20 Assessments 
The following assessments will be conducted at the main visit: 
• 
A physical examination will be conducted. 
• 
Vital signs (pulse, blood pressure) will be measured. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(central lab). 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab). 
• 
Blood will be taken for collection of mononuclear cells for immune studies (Elispot). 
• 
Blood will be taken for evaluation of FIX activity levels (central and local lab). 
• 
Blood will be taken for evaluation of FIX inhibitor levels. 
• 
Patients will discontinue the course of immunosuppressants (see Section 9.4).  
• 
For any patients with a history of HBV and HCV, local testing for hepatitis B or C 
reactivation should be conducted.  
• 
CMV testing (CMV PCR; in patients with CMV positive (IgG) at screening) should be 
conducted in conjunction with immunosuppressant dosing. Management guidelines in 
the case of CMV reactivation can be found in Appendix 2. 
• 
Tacrolimus levels should be measured in conjunction with immunosuppressant dosing. 
Management guidelines for tacrolimus dosing can be found in Appendix 3. 
• 
Plasma, saliva, urine, stool, and semen samples will be taken for PCR of vg. These 
collections will be done unless there have been 3 prior consecutive specimens that were 
negative.  
• 
Adverse event and concomitant medication status will be recorded at each visit. 
• 
Plasma samples will be drawn and frozen for research purposes. These samples are 
optional. 
• 
Plasma samples will be drawn and frozen for immune response research purposes only 
during the course of investigations or treatment of breakthrough transaminitis. 

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Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 1 
further separate occasion during week 20 (see Section 11.2.10). 
11.2.9 
Week 21-25 Assessments  
• 
Frequent local laboratory assessments (FIX activity level and LFTs) will be conducted on 
2 separate occasions each week during week 21-25 (see Section 11.2.10). Blood draws 
may be taken either at study site or at alternative location (e.g. patient’s home). The 
intention should be for at least one of the two blood draws per week to be conducted at 
study site. 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab), once a week, weeks 21-25. Blood draw may be taken either at study site or 
at alternative location (e.g. patient’s home). 
• 
Adverse event and concomitant medication status will be recorded and ongoing diary 
completion assessed. 
11.2.10 
Frequent Local Laboratory Assessments 
In addition to the main weekly visit schedule, the following laboratory assessments will be 
conducted an additional 2 times per week (3 times in total) between week 1-5 and an additional 
1 time per week (2 times in total) between week 6-12, week 14, week 16 and week 20. For 
week 13, 15, 17, 18, 19, 21, 22, 23, 24 and 25 the following laboratory assessments will be 
conducted 2 times per week. The tests should be as evenly spaced through the week as 
possible, for example: 3 times per week – Monday, Wednesday and Friday; 2 times per week 
– Monday and Thursday or Tuesday and Friday. These samples may be taken at study site or 
at an alternative location (e.g. patient’s home) by a home nursing vendor. For week 13, 15, 17, 
18, 19, 21, 22, 23, 24 and 25 the intention should be for at least one of the two blood draws 
per week to be conducted at study site. To facilitate rapid evaluation of results and intervention 
where necessary, local laboratory analysis should be used for assessments during this period:  
• 
FIX activity level.  
• 
LFTs - for list of required tests please see Table 1: Schedule of Assessments and Section 
11.4.3.2. 
In response to an upward trend in ALT, frequency of LFT and FIX activity level assessments 
should be increased in line with the guidance in Section 9.4. 
Where required to meet the requested schedule of testing these frequent assessments should 
include sample for reactivation of hepatitis, tacrolimus levels and CMV testing as per Table 1: 
Scheduled of Assessments. These samples may be taken at study site or at an alternative 
location (e.g. patient’s home) by a home nursing vendor.   
11.3 
Week 26/End-of-Study Visit 
The following assessments will be conducted at week 26. Should the patient discontinue the 
study at an earlier time point, every attempt should be made to conduct the week 26 
assessments to ensure adequate safety follow-up. 

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• 
A physical examination will be conducted. 
• 
Vital signs (pulse, blood pressure) will be measured. 
• 
A 12-lead ECG will be conducted. 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(central lab). 
• 
Blood will be taken for haematology, chemistry incl. CRP, coagulation screen and LFTs 
(local lab). 
• 
Blood will be taken for collection of mononuclear cells for immune studies (Elispot and 
Research). The mononuclear (Research) sample is optional 
• 
Blood will be taken for evaluation of FIX activity levels (central and local lab). 
• 
Blood will be taken for evaluation of FIX antigen levels. 
• 
Blood will be taken for evaluation of FIX inhibitor levels. 
• 
Blood will be taken for evaluation of AAV-S3 antibody titre. 
• 
Plasma, saliva, urine, stool, and semen samples will be taken for PCR of vg. These 
collections will be done unless there have been 3 prior consecutive specimens that were 
negative. 
• 
Plasma samples will be drawn and frozen for research purposes. These samples are 
optional. 
• 
Plasma samples will be drawn and frozen for FIX activity assay research purposes. 
• 
Plasma samples will be drawn and frozen for immune response research purposes. 
• 
Adverse event and concomitant medication status will be recorded. 
• 
Assessment of bleeding episodes and FIX usage based on patient diaries and direct 
questioning.  
• 
A clinical assessment of joints will be undertaken using the HJHS. 
• 
A liver ultrasound scan will be conducted. 
• 
Quality of life will be evaluated (EQ-5D-5L and Haem-A-QoL). 
• 
Disability status will be assessed (WHODAS 2.0). 
• 
Physical activity status will be assessed (HAL 2005). 
• 
Haemophilia health status will be assessed (PROBE). 
• 
Health resource utilisation will be evaluated since screening or Day -1. 

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11.4 
Study Procedures/Evaluations 
11.4.1 
Demographic and Baseline Assessments 
11.4.1.1 
Informed Consent Procedure  
The informed consent process will be conducted twice during the study. Initially at screening 
and then again once eligibility has been confirmed but prior to FLT180a infusion. 
It is the responsibility of the investigator, or a suitably qualified co-investigator delegated by 
the investigator, to obtain written informed consent from each patient before any trial related 
procedures are undertaken. The investigator or designee will explain the aims, methods 
anticipated benefits, and potential hazards of the trial, that the patients are under no obligation 
to participate and that they can withdraw at any time, without having to give a reason. 
A mandatory meeting with an independent patient advocate, who will be tasked with focusing 
on the risks associated with the trial, will be arranged prior to the second ‘pre-infusion’ dosing 
site informed consent to ensure that the participant fully comprehends the risks. The right to 
withdraw at any time during the study without prejudice will also be explained again. The 
requirement for a meeting with an independent patient advocate will not apply once the 
terminal dose level is selected. 
A copy of the signed ICF will be given to the participant. The original signed ICF will be retained 
at the study site and a copy placed in the medical notes.  
As part of the consent process, patients will be informed that in the event of death, no matter 
what cause, permission for an autopsy will be requested of their families. Patients will be asked 
to advise their families of this request and of its scientific and medical importance should they 
choose to participate. 
11.4.1.2 
Patient Identification Code 
On enrolment to the study, the patient will be given a unique patient identification code 
consisting of: a 2-digit country number (e.g., 44), a 2-digit site number (e.g., 03), a 1-digit study 
number (for this study 1) and a 3-digit patient number (e.g., 004). Using these specifications, 
the fourth patient enrolled at site 03 in the United Kingdom (country code 44) will have the 
unique patient identification code 44031004. Allocation of the patient identification code will be 
handled through the eCRF. 
11.4.1.3 
Factor IX Mutation Genotype 
Factor IX mutation genotype should be transcribed onto the eCRF from the patients’ medical 
records during the screening period. Where genotype has not been established, a blood 
sample will be sent to the central laboratory for analysis. 
11.4.1.4 
Medical History 
A 5-year medical history should be taken during the screening period. Details of any clinically 
relevant abnormalities should be noted on the eCRF. 

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11.4.1.5 
Bleeding History 
The investigator should document the patients bleeding and FIX utilisation history over the 
preceding 3 years. 
From this data target joint assessment will be recorded as per the criteria below38:  
A target joint is defined as one in which there have been three or more spontaneous bleeds 
within a consecutive 6-month period. Where there have been ≤2 bleeds into the joint within a 
consecutive 12-month period the joint is no longer considered a target joint. 
11.4.1.6 
AAV-S3 Neutralising Antibody Screening 
During the screening period, a blood sample will be taken from the patient to determine the 
presence or absence of neutralising antibodies to the AAV-S3 serotype. 
A negative result from a transduction inhibition assay within 4 weeks of dosing is required to 
confirm eligibility. It is acceptable to take 2 samples during the screening period. Should there 
be any problems with the testing it is acceptable to take repeat samples as necessary. At each 
time-point a primary sample will be drawn and plasma sent for analysis. A secondary sample 
will be drawn and plasma frozen locally as a back-up sample. 
Procedures for collection, processing, storing, and transporting of samples to the central 
laboratory are fully described in the study Laboratory Manual.  
11.4.1.7 
HIV, HCV, HBV, CMV Screening 
A blood sample will be taken at screening to assess the following: 
• 
Hepatitis B virus surface antigen (HBsAg). 
• 
Anti-hepatitis C virus (anti-HCV) antibodies and Hepatitis C RNA viral load. HCV RNA 
viral load only indicated for patients with history of HCV and positive HCV antibody test. 
• 
Human immunodeficiency virus 1 and 2 (anti-HIV1/2) antibodies. 
• 
Cytomegalovirus IgG antibodies and Cytomegalovirus PCR. CMV PCR only indicated 
if patients are positive on CMV IgG. 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual. 
11.4.2 
Efficacy 
11.4.2.1 
Baseline FIX Activity 
Baseline FIX activity must be established for all patients. 
Due to the risk of bleeding in this patient group, a washout from the patients FIX concentrate 
regimen is not mandated as part of this protocol. The investigator must however be able to 
demonstrate a documented FIX activity level of <1% for severe patients or <2% for moderately 
severe patients from the patients’ historical medical records.  

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For patients where a washout is undertaken as part of this protocol, a minimum of 5 days off 
therapy is required. In this case the FIX activity trough sample will also be used as the baseline 
value. 
11.4.2.2 
FIX Activity Trough 
A trough level of FIX activity should be established during the screening period. 
• 
For patients on on-demand therapy, a blood sample should be taken following 5 days 
free from factor concentrates. 
• 
For patients on conventional prophylactic therapy, a blood sample should be taken at 
the end of the longest acceptable interval between doses. 
• 
For patients taking EHL prophylactic therapy, a blood sample should be taken within 
the 48-hour period before the next planned dose. 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual. 
11.4.2.3 
FIX Activity 
Blood samples for assessment of FIX activity will be drawn in accordance with Table 1: 
Schedule of Assessments and Table 2: Detailed Schedule of Assessments for Infusion Week. 
Samples at the various time-points will be analysed at a central laboratory (efficacy) and local 
laboratory (efficacy and safety) as indicated.  
Timepoints for a central laboratory sample: 
A primary sample will be drawn and plasma sent for analysis of FIX activity. 
A secondary sample will be drawn and plasma frozen. The secondary sample will be used to 
establish FIX activity in an alternative investigational assays. 
Procedures for collection, processing, storing, and transporting to the laboratory are fully 
described in the study Laboratory Manual. 
Timepoints for a local laboratory sample: 
Local blood samples will be taken Day -1, Day 0 (pre-dose), Day +2, Day +4, 3 times per week 
between weeks 1-5, 2 times per week between weeks 6-25 and once in week 26/EOS. These 
samples will be analysed locally. On days where other clinic assessments are not required 
these samples may be taken at study site or at an alternative location (e.g. patient’s home). 
Local laboratory analysis will be utilised to ensure rapid turnaround of results in this exception. 
11.4.2.4 
FIX Antigen 
Blood samples for assessment of FIX antigen will be drawn in accordance with Table 1: 
Schedule of Assessments. 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual.  

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11.4.2.5 
Bleeding Episodes 
Bleeding episodes will be entered into the patient’s diary and will include: 
• 
Number 
• 
Location 
• 
Aetiology (spontaneous, traumatic, etc) 
• 
Total units of FIX concentrate required to resolve the bleed 
Diary data will be reviewed with the investigator at the next available visit. 
11.4.2.6 
Factor IX Concentrate Usage  
For each dose of FIX concentrate administered the patient will record the concentrate, dose 
given in total international unit (IU), the reason for administration (prevention, bleeding 
episode), and the frequency of administration in the patient diary.  
Diary data will be reviewed with the investigator at the next available visit.  
11.4.3 
Safety 
11.4.3.1 
Physical Examination 
The following sites will be examined: head, neck, ears, nose, throat, eyes, chest, lungs, heart, 
abdomen, skin, and lymph nodes. 
The following systems will be assessed: musculoskeletal and neurological. 
The patient’s height (screening only) and weight will also be measured. Waist circumference 
(cm), hip circumference (cm), neck circumference (cm) and bioimpedance will be measured at 
screening and Day -1 only. 
11.4.3.2 
Laboratory Safety Assessments 
Blood samples for safety assessments will be drawn in accordance with Table 1: Schedule of 
Assessments and Table 2: Detailed Schedule of Assessments for Infusion Week. 
The following laboratory tests will be performed at a central laboratory to assess safety.  
• 
Haematology: complete blood count with differential, platelet count. 
• 
Chemistry incl. CRP: sodium, potassium, calcium, chloride, phosphate, CO2, glucose, 
blood urea nitrogen, serum creatinine, C-reactive protein. 
• 
Coagulation screen: prothrombin time, activated partial thromboplastin time, 
prothrombin split fragments 1+2 (F1+2), D-dimer, TAT. 
• 
Liver Function Tests: albumin, alkaline phosphatase, direct bilirubin, indirect bilirubin, 
total bilirubin, alanine aminotransferase, aspartate aminotransferase, total protein, 
GGT. 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual. 
The following laboratory tests will be performed at a local laboratory to assess safety. 

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• 
Haematology: complete blood count with differential, platelet count. 
• 
Chemistry incl. CRP: sodium, potassium, phosphate, blood urea nitrogen or urea, 
serum creatinine (and estimated GFR), C-reactive protein. 
• 
Coagulation screen: prothrombin time, activated partial thromboplastin time. 
• 
Liver Function Tests: albumin, alkaline phosphatase, direct bilirubin, indirect bilirubin, 
total bilirubin, alanine aminotransferase, aspartate aminotransferase. 
• 
FIX activity level. 
Full local laboratory safety blood samples will be taken Day -1, Day 0 (pre-dose and +8 hours), 
Day +1, Day +2, Day +4. Samples for LFT’s and FIX activity level will be taken 3 times per 
week between weeks 1-5, 2 times per week between weeks 6-25 and once at week 26/EOS. 
These samples will be analysed locally. 
Local laboratory analysis will be utilised to ensure rapid turnaround of results in this exception. 
On days where other clinic assessments are not required these samples may be taken at study 
site or at an alternative location (e.g. patient’s home). 
Blood samples for reactivation of hepatitis, tacrolimus levels and CMV testing will be drawn in 
accordance with Table 1: Schedule of Assessments and Section 9.4 and analysed locally. On 
days where other clinic assessments are not required these samples may be taken at study 
site or at an alternative location (e.g. patient’s home). Management guidelines in the case of 
CMV reactivation can be found in Appendix 2. Management guidelines for tacrolimus dosing 
can be found in Appendix 3. 
If immunosuppressants are implemented at any other time, monitoring for reactivation of 
hepatitis B/C, tacrolimus levels and CMV testing should be undertaken in conjunction with the 
immunosuppressant dosing. 
11.4.3.3 
FIX Inhibitor  
Blood samples for assessment of FIX neutralising antibody development (inhibitor) will be 
drawn in accordance with Table 1: Schedule of Assessments. 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual.  
11.4.3.4 
AAV-S3 Antibody Titre 
Blood samples for assessment of AAV-S3 Antibodies will be drawn in accordance with Table 
1: Schedule of Assessments. A primary sample will be drawn and plasma sent for analysis. A 
secondary sample will be drawn and plasma frozen locally as a back-up sample. 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual.  
11.4.3.5 
Vital Signs 
Measurements of vital signs (blood pressure, pulse, temperature, and respiratory rate) will be 
performed according to the study schedule (Table 1: Schedule of Assessments and Table 2: 

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Detailed Schedule of Assessments for Infusion Week). Blood pressure and pulse will be 
determined in the sitting position (for 5 minutes). 
Blood pressure should be determined by cuff (manual or automated is acceptable although 
the same method should be used throughout the study). 
Any clinically significant deviations from vital signs at Day -1 should be reported as an AE. 
11.4.3.6 
ECG 
A 12-lead ECG will be conducted in accordance with the study schedule (Table 1: Schedule 
of Assessments and Table 2: Detailed Schedule of Assessments for Infusion Week). 
Any clinically significant deviations from Day -1 should be reported as an AE. 
11.4.3.7 
Liver Ultrasound 
A liver ultrasound will be conducted in accordance with the study schedule (Table 1: Schedule 
of Assessments). 
Any clinically significant deviations from screening should be reported as an AE. 
11.4.3.8 
Adverse Event Collection 
Patients will be questioned in a general way at each study visit to establish whether AEs have 
occurred since the previous visit (e.g., “How have you been feeling since your last visit?”). 
Additionally, the investigator will evaluate other collected data (e.g., patient diaries, 
questionnaires, clinical evaluations) to ascertain whether an AE has occurred. Adverse events 
are collected from the time initial informed consent is signed until week 26. Please refer to 
Recording and Reporting of Adverse Events in Section 13. 
11.4.4 
Vector Shedding 
Plasma, saliva, urine, stool and semen samples for PCR of vector genome will be taken in 
accordance with Table 1: Schedule of Assessments and Table 2: Detailed Schedule of 
Assessments for Infusion Week. These collections will occur until there have been 3 
consecutive samples that are negative. 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual. 
11.4.5 
Health Economic Assessments 
Assessments will be conducted in accordance with the study schedule (Table 1: Schedule of 
Assessments and Table 2: Detailed Schedule of Assessments for Infusion Week). 
11.4.5.1 
Quality of Life  
Quality of life will be evaluated using the EQ-5D-5L and Haem-A-QoL. 
11.4.5.2 
Disability 
Disability will be assessed using the WHODAS 2.0. 
11.4.5.3 
Physical Activity  
Physical activity will be assessed using the HAL 2005. 

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11.4.5.4 
Haemophilia Health Status 
Haemophilia health status will be assessed using PROBE. 
11.4.5.5 
Health Resource Utilisation 
The following items will be recorded in the eCRF: 
• 
Number of haemophilia related medical appointments and medical activities 
• 
Number of emergency room visits 
• 
Number of hospitalisations related to haemophilia 
• 
Length of hospital stay 
• 
Number of physiotherapy sessions, specialist consultations and appointments with 
professional caregivers. 
The following items will be captured in the patient diary: 
Number of days lost from education or work by patients and caregivers due to bleeding 
episodes. 
11.4.6 
Exploratory 
11.4.6.1 
Joint Health/Function 
Joint health/function will be conducted in accordance with the study schedule (Table 1: 
Schedule of Assessments). 
Joint health/function will be assessed using the HJHS score. 
11.4.6.2 
Mononuclear Cells (Immune Studies) 
Blood samples will be collected for immune studies in accordance with Table 1: Schedule of 
Assessments and Table 2: Detailed Schedule of Assessments for Infusion Week.  
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual.  
11.4.6.3 
Mononuclear Cells (Research) 
At Day -1 and week 26 a second mononuclear cell sample will be taken for research purposes. 
These samples are optional. Research samples will be retained beyond the end of the study 
and may be analysed alongside samples taken as part of the long-term follow-up study or 
pooled with other studies using similar constructs to generate a larger sample for analysis. Any 
analysis will be conduct independent of the study report. The samples may therefore be held 
for up to 15 years and analysed as part of future research which has been ethically approved.  
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual.  
11.4.6.4 
Research Plasma  
Research plasma samples will be collected in accordance with Table 1: Schedule of 
Assessments and Table 2: Detailed Schedule of Assessments for Infusion Week for research 

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purposes. These samples are optional. Research samples will be retained beyond the end of 
the study and may be analysed alongside samples taken as part of the long-term follow-up 
study or pooled with other studies using similar constructs to generate a larger sample for 
analysis. Any analysis will be conduct independent of the study report. The samples may 
therefore be held for up to 15 years and analysed as part of future research which has been 
ethically approved.  
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual. 
11.4.6.5 
FIX Activity Research Plasma 
Blood samples will be collected for FIX activity research studies in accordance with Table 1: 
Schedule of Assessments. The intent of drawing these samples is to investigate the variability 
of the FIX activity assay in accordance with the draft FDA guidance for gene therapy in 
haemophilia.39 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual. 
11.4.6.6 
Immune Response Research Plasma 
Blood samples will be collected for immune response research studies in response to 
elevations in LFT’s and in accordance with Table 1: Schedule of Assessments and Table 2: 
Detailed Schedule of Assessments for Infusion Week. The intent of drawing these samples is 
to investigate immune response biomarkers at time-points when events of breakthrough 
transaminitis are observed. 
Procedures for collection, processing, storing, and transporting to the central laboratory are 
fully described in the study Laboratory Manual. 
11.5 
Volume of Blood to be Drawn From Each Patient 
Table 4: 
Volume of Blood to be Drawn From Each Patient for Central Laboratory 
Testing 
Assessment 
Sample Volume 
(mL) 
Number of 
Samples 
Taken 
Total Volume (mL) 
AAV-S3 antibody screen/titre 
4.5 
7 
31.5 
Genotyping 
2 
1 
2 
Safety 
Chemistry 
(including CRP and 
LFTs where 
combined and 
serology - HBV, 
3.5 or 5 (where 
serology included)  
20 of which 1 
serology 
71.5 

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Table 4: 
Volume of Blood to be Drawn From Each Patient for Central Laboratory 
Testing 
Assessment 
Sample Volume 
(mL) 
Number of 
Samples 
Taken 
Total Volume (mL) 
HCV, HIV, CMV as 
required) 
HCV RNA viral 
load 
5 
1 
5 
CMV PCR 
5 
1 
5 
Haematology 
2 
20 
40 
Coagulation 
4.5 
20 
90 
FIX activity 
9 
20 
180 
FIX inhibitor 
4.5 
10 
45 
FIX antigen 
4.5 
3 
13.5 
Viral shedding 
2 
5 (min) to 19 
(max) 
10-38 
Mononuclear Cells (Elispot) 
20 
17 
340 
Mononuclear Cells (Research) 
20 
2 
40 
Research Plasma Sample 
4.5 
17 
76.5 
FIX Activity Research Plasma 
Sample 
20 
3 
60 
Immune Response Research 
Plasma Sample 
4.5 
2 (min) to 17 
(max) 
4.5-76.5 
Total mL (Approximate) 
 
1015-1135 
Abbreviations: AAV = adeno-associated virus; CMV = cytomegalovirus; CRP = C-reactive protein; FIX = Factor IX; 
HBV = hepatitis B virus; HCV = hepatitis C virus, HIV = human immunodeficiency virus, max = maximum, min = 
minimum.  
 
During this study, it is expected that approximately 1015-1135mL of blood will be drawn from 
all patients for central laboratory assessments. 
The protocol allows for monitoring of certain parameters through local laboratories. Blood 
volume requirements are likely to vary between centres, however, it is estimated that 

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approximately 1000mL will be drawn for local laboratory assessments over the course of the 
study. 
Note: The amount of blood to be drawn for each assessment is an approximation. The amount 
of blood to be drawn may vary according to the instructions provided by the manufacturer or 
laboratory for an individual assessment.  
11.6 
Discontinuation/Withdrawal of Participants  
The reason for discontinuation/withdrawal must be determined by the investigator and 
recorded in the patient’s medical record and on the eCRF. If a patient is withdrawn for more 
than one reason, each reason should be documented in the source document and the most 
clinically relevant reason should be entered on the eCRF. 
Reasons for discontinuation include but are not limited to: 
• 
Protocol violation 
• 
Withdrawal by patient 
• 
Death  
• 
Lost to follow-up 
• 
Other (If “other” is selected, the investigator must specify the reason on the eCRF). 
If a patient expresses their wish to withdraw from the trial, the investigator should explain the 
importance of remaining on trial follow-up, or failing this, of allowing routine follow-up data to 
be used for trial purposes.  
In the event of early withdrawal/discontinuation, every effort should be made to collect data in 
line with week 26/end-of-study assessments. 
11.6.1 
Withdrawal of Consent to Data Collection  
If a patient explicitly states they do not wish to contribute further data to the trial, their decision 
must be respected and recorded in the eCRF and medical notes.  
11.6.2 
Loss to Follow-up 
If a patient moves from the area, every effort should be made for the patient to be followed up 
at another participating trial site and for this new site to take over the responsibility for the 
patient, or for follow-up via the patient’s general practitioner.  
If a patient is lost to follow-up at a site, every effort should be made to contact the patient’s 
general practitioner to obtain information on the patient’s status.  
11.7 
Definition of End of Trial 
The end of the trial will be defined as the last visit by the last participant. 
 

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12 
Dose Progression, Dose Escalation and Stopping Rules  
The sponsor will retain final responsibility for decision making on all aspects of dose 
progression, dose escalation/reduction and stopping rules. Progression of dosing within a 
cohort, dose escalation/reduction and stopping rules are detailed in a study specific plan. The 
following elements will be included.  
12.1 
Oversight Committees  
12.1.1 
Trial Management Group (TMG) 
The Trial Management Group (TMG) will include the chief investigator and trial staff, PIs, and 
sponsor personnel. The TMG will be responsible for overseeing the trial to ensure that the 
protocol is adhered to and will take appropriate action to safeguard participants and the quality 
of the trial itself. The group will meet at least monthly and will send updates to PIs who are not 
in attendance.  
The TMG will review trial progress, substantial amendments, patient eligibility and ongoing 
oversight of patient enrolment within each dose level. 
Written approval from the TMG will be mandatory prior to dosing of each patient.    
12.1.2 
Independent Data Monitoring Committee (DMC)  
The role of the DMC is to provide independent advice on data and safety aspects of the trial.  
Meetings of the committee will be held to review patient data prior to a dose escalation 
decision, in the case a DLT occurs, selection of the terminal dose or any other issue or safety 
concern the TMG feel they need advice on from the DMC (i.e. dose reduction). The DMC is 
advisory to the sponsor and can recommend premature closure of the trial. The DMC will be 
governed by a charter that dictates constitution and decision making on dose escalation and 
trial stopping rules. 
A positive recommendation from the DMC will be mandatory prior to dose escalation and 
selection of the terminal dose cohort.  
12.1.3 
Trial Steering Committee (TSC) 
The role of the Trial Steering Committee (TSC) is to provide overall supervision of the trial. 
The TSC will review the recommendations of the independent DMC and, on consideration of 
this information, recommend any appropriate amendments/actions for the trial as necessary.  
The TSC acts on behalf of the funder and Sponsor. The TSC is governed by a charter which 
dictates constitution and terms of reference. 
12.2 
Dose-Limiting Toxicity 
A DLT is defined as any grade 3 or greater AE at least possibly related to FLT180a except for 
increases in ALT or AST that are not associated with increases in bilirubin. 
Any occurrence of a DLT should be reported as an SAE. 
The DMC will review any occurrence of a DLT and further dosing will be suspended until the 
DMC provides a recommendation to continue.  

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The trial may be stopped before completion at any time on the recommendation of the DMC 
or decision by the sponsor.  
12.3 
Dose Escalation 
12.3.1 
Dose Interval Between Patients  
FLT180a will be given as a single dose, slow IV infusion.  
The planned dose escalation scheme is as follows: 
• 
Cohort 1 (low dose): 6 x 1011 vg/kg of body weight 
• 
Cohort 2 (intermediate dose): 2 x 1012 vg/kg 
• 
Cohort 3 (high dose): 4 x 1012 vg/kg. 
The dose level within a cohort may be reduced based on observed FIX activity levels, see 
Section 12.3.3. 
• 
Cohort 2 (intermediate dose): 2 x 1012 vg/kg can be reduced to: 
- 
1.5 x 1012 vg/kg 
- 
1.3 x 1012 vg/kg 
- 
1 x 1012 vg/kg 
- 
8 x 1011 vg/kg 
• 
Cohort 3 (high dose): 4 x 1012 vg/kg can be reduced to: 
- 
3 x 1012 vg/kg 
As this is a first-in-human study, an extended 6-week interval will be observed between the 
first and second patient to monitor for any unanticipated delayed AEs. 
When applying the rules below, safety will be assessed first (see Section 12.3.2) followed by 
evaluation of FIX activity (see Section 12.3.3). 
During the dose escalation phase of the trial, review of FIX data at a dose of 1.3 x1012 vg/kg, 
is suggestive of the impact of body weight on expression levels.  A dose capping at 90 kg is 
being introduced at this dose (1.3 x1012 vg/kg) to ensure that patients FIX activity levels, at 
steady state are ideally in the target range. 
12.3.2 
Rules Applicable to Evaluation of Safety in Each Dose Cohort 
Dose first patient: 
If no DLT is observed, evaluate FIX activity before progressing to dose the second patient. 
If DLT arises, then consult DMC prior to dosing second patient and plan for a third patient 
at the dose level. 
 
 

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Dose second patient: 
If no DLT has occurred in either patient at the dose level, evaluate FIX activity before dose 
escalation (see Section 12.3.3). 
If DLT is observed in second patient (with no DLT observed in first patient), consult DMC 
and plan for addition of third patient at same dose. 
If DLT is observed in both patients 1 and 2, apply temporary halt to trial and consult 
competent authority. 
Dose third patient (if necessary): 
If no DLT occurs, evaluate FIX activity before dose escalation (see Section 12.3.3). 
If DLT observed in third patient and this is second patient in which a DLT has been 
observed, apply temporary halt to trial and consult competent authority. 
The TMG will review safety data for each patient prior to further dosing in line with the rules 
above. The TMG may recommend enrolment of additional patients be added to any cohort to 
characterise any safety concern that has not met DLT criteria. Where consulted the DMC 
must review data and provide recommendation to TMG and Sponsor before another patient 
is dosed. 
Should additional patients be added and 2 DLTs occur at any given dose level then apply 
temporary halt to trial and consult competent authority. 
12.3.3 
Rules Applicable to Assessment of FIX Activity in Each Dose Cohort 
Factor IX activity is expected to follow a pattern similar to that observed with other single-
stranded gene therapies, with initial response appearing within a few days and rising to 
approximately 50% of peak levels over the first 2 weeks post infusion. Following this, trajectory 
is expected to slow reaching approximately 75% of peak at 4 weeks and achieving steady-
state levels at approximately week 12. During steroid administration (from week 3) FIX levels 
may be exaggerated. As such, week 3 FIX levels (prior to steroid administration) will be used 
to guide decision making on progression between dose levels. 
The target range for FIX activity level at steady state is 70-150% 
The TMG will review FIX activity data for each patient prior to further dosing. 
The DMC will review FIX activity data for each patient prior to dose escalation, selection of the 
terminal dose and where requested by the TMG. 
The TMG and DMC are in place to protect patient safety and trial validity and will review all 
data, and projections of anticipated FIX trajectory can be considered, in order to provide 
recommendations to the Sponsor for dose escalation, terminal dose level selection and dose 
reductions.  
The following rules should be observed to assess dosing decisions in relation to FIX activity 
and anticipated trajectory: 

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• 
FIX activity levels up to and including week 3 (~65% of steady state) will be used to 
guide decision making for progression between dose levels. The following FIX activity 
level ranges will be applied and the oversight committees involved in decision making 
are indicated in brackets: 
- 
< 50% – dose escalate (TMG and DMC) 
- 
≥ 50% and < 60% – consider dose escalation (TMG and DMC) 
- 
≥ 60% and < 105% – continue at that dose level (TMG) 
- 
≥ 105% and < 150% – TMG evaluates the risk benefit of the dose reduction and 
recommends the dose for the next patient in keeping with the dose cohorts 
(TMG and DMC) 
- 
> 150% – stop dosing further patients until FIX activity levels have peaked*. 
TMG and DMC will convene to discuss next steps e.g. finding alternate dosing 
strategies.  
* The attainment of peak FIX activity levels will be judged by the TMG but will 
generally require evidence over a two-week period showing stability or evidence of 
continued decline. 
• 
If any patient exceeds 400% at any time stop dosing further patients until FIX activity 
levels have peaked. TMG and DMC will convene to discuss next steps. 
• 
If any patient FIX levels exceed 200%, consider thromboprophylaxis after individual 
risk assessment in conjunction with the Chief Investigator. Patients with known risk 
factors (obesity, smoker, heart failure, older than 60 years age etc.) should be offered 
thromboprophylaxis (e.g. apixaban) if FIX levels exceed 200%. Patients with no risk 
factors should be offered thromboprophylaxis if FIX levels exceed 300%. Patients 
should be closely monitored by the Investigator for signs and symptoms of potential 
thrombosis. Patients on thromboprophylaxis need to be carefully observed for signs of 
bleeding. While on thromboprophylaxis, ongoing individual risk assessment of the 
patient’s status including review of FIX levels should be conducted in conjunction with 
the Chief Investigator. If the risk assessment is favourable and FIX levels return below 
threshold levels, consideration should be given to stopping thromboprophylaxis.     
• 
If any patient exhibits a positive inhibitor result at any time stop dosing further patients 
until confirmation of presence of an inhibitor, or not, from a repeat test(s) (see section 
12.5 Stopping Rules). TMG and DMC will convene to discuss next steps. 
• 
A minimum of 2 patients treated at any given dose level will be required for dose 
escalation and selection of terminal dose level decisions. 
• 
Dose reduction decisions (where necessary) can be implemented based on data from 
1 patient treated at any given dose level. 
• 
The TMG and DMC may recommend enrolment of additional patients at any dose to 
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• 
For selection of the terminal dose level, the TMG and DMC will aim to select a dose 
that enable the majority of patients to attain a FIX activity level in the range 70-150% 
at steady state. The TMG and DMC will utilise all safety and efficacy data available at 
the time of decision making. Projection data of FIX activity level from week 4 can be 
considered where necessary. As such selection of the terminal dose level would be 
discussed when 2 patients are in the range 50-150% at week 3. 
• 
Once the terminal dose level is selected, 14 patients in total will be dosed at this level. 
Patients may be dosed at 48-hour intervals. If any patient exceeds 400% at any time 
stop dosing further patients until FIX activity levels have peaked (see above for 
definition of peak). TMG and DMC will convene to discuss next steps.  
• 
The TMG and/or DMC will have latitude in the recommendation they make from the 
start point for discussion and would utilise all safety and efficacy data currently 
available and projection of anticipated FIX activity at steady state at the dose.  
Where more than 2 patients have been dosed at a dose under review it would be expected 
that the range where the majority of patients FIX activity level falls should be the start point for 
discussions on the recommendation. 
12.4 
Temporary Halt 
Once a temporary halt is applied to the trial (via submission of a substantial amendment) an 
internal safety review by the DMC will take place. If the Sponsor deems it appropriate to restart 
the trial; this can be done only following approval by the relevant competent authorities, ECs 
and along with any local approvals required. 
12.5 
Stopping Rules 
Further enrolment into the study will be put on hold in the event of any of the following and a 
substantial amendment will be submitted to temporarily halt the trial: 
• 
Death of a patient related to FLT180a 
• 
Development of malignancy related to FLT180a 
• 
Development of an inhibitor (Anti-FIX antibody) – a positive result must be confirmed 
by the result of a second sample taken within 1-4 weeks after the initial sample, 
exhibiting inhibitor, using the Bethesda assay. 
Participants who have already received the vector infusion will continue to be followed per the 
protocol. If, following an internal safety review by the independent DMC, the Sponsor deems 
it appropriate to restart the trial; this can only be done following approval by all relevant 
competent authorities, ECs and along with any local approvals required.  
 
13 
Recording and Reporting of Adverse Events  
An AE is any untoward medical occurrence in a clinical investigation subject administered a 
pharmaceutical product and that does not necessarily have a causal relationship with this 

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In addition, any signs or symptoms experienced by the patient after signing the ICF, or leading 
up to the onset date of the SAE, or following the resolution date of the SAE, must be recorded 
as an AE, if appropriate. 
13.2.8 
Fatal Outcome 
Any SAE that results in the patient’s death (i.e., the SAE was noted as the primary cause of 
death) must have fatal checked as an outcome with the date of death recorded as the 
resolution date. For all other events ongoing at the time of death that did not contribute to the 
patient’s death, the outcome should be considered not resolved, without a resolution date 
recorded. 
For any SAE that results in the patient’s death or any ongoing events at the time of death, the 
action taken with the investigational product should be recorded as “dose not changed” or “not 
applicable” (if the patient never received investigational product). 
13.2.9 
Important Medical Events 
For the purpose of this study the following will be considered as important medical events and 
will be reported in line with SAE reporting procedures. 
• 
Vector-induced hepatitis (transaminase elevations related to FLT180a). 
• 
Development of an inhibitor. 
• 
A patient develops a grade 2 or greater allergic reaction related to administration of 
FLT180a. 
• 
Thrombosis.  
Other events may be reported at the discretion of the Investigator. 
13.2.10 
Pregnancy  
The population under study is male; therefore, pregnancies are not applicable.  
A prerequisite to entry into the trial is a willingness to practice a reliable barrier method of 
contraception until 3 negative semen samples have been obtained after vector administration. 
In the previous study conducted by Nathwani et al this occurred within 6 weeks in all 
participants. However, if the partner of a male participant becomes pregnant, before 3 negative 
semen specimens have been obtained from the patient, the site team should request for 
permission to keep in touch with their partner to follow-up the pregnancy to its conclusion.   
Consent to report information regarding the pregnancy must be obtained from the participant’s 
pregnant partner. A trial-specific pregnancy monitoring information sheet and informed 
consent form for partners of trial participants must be used for this purpose. 
The pregnancy should be reported on a pregnancy reporting form following consent and 
submitted to the CRO pharmacovigilance department.  
The CRO pharmacovigilance department must be kept informed of any new developments 
involving the pregnancy. 

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13.2.11 
Regulatory Agency, Ethics Committee, and Site Reporting 
The CRO is responsible for notifying the relevant regulatory authorities and Central Ethics 
Committees (ECs)/Institutional Review Boards (IRBs) of related, unexpected SAEs (SUSARs) 
in accordance with the regulatory requirements. SUSARs that are fatal or life-threatening must 
be notified with 7 days from the CRO learning of them with any additional relevant information 
reported within a further 8 days. Other SUSARs must be reported within 15 days from the CRO 
learning of them. The CRO will also inform all participating sites. 
The investigator is responsible for notifying the local ECs/IRBs or the relevant local regulatory 
authority of all SAEs that occur at his or her site as required. 
The CRO will submit an annual development safety update report to relevant regulatory 
authorities. 
13.2.12 
Expedited Reporting of Other Safety Events 
New events related to the conduct of the trial or the development of the ATIMP which are likely 
to affect the safety of the participants should be reported according to the existing timelines 
for expedited reporting. 
This includes: 
• 
SAEs which could be associated with the trial procedures and which could require 
modification of the conduct of the trial. 
• 
Events which pose significant hazard to the participant population. 
• 
SAEs related to mandatory concomitant medication, product application process 
(surgical or other) and product failure (including lack of efficacy) should all be 
considered.  
• 
Events (as described above) which are fatal or life-threatening, will be notified to the 
EC and competent authority by the CRO within 7 days of the CRO learning of them. 
Events falling into one of the other categories of serious (see Section 13.1 for definition) 
will be reported by the CRO to the EC and CA within 15 days after the CRO has learned 
of them.   
Expedited reporting and reporting of adverse events will be in accordance with EudraLex 
Volume 10, Chapter II (Detailed Guidance on the collection, verification and presentation of 
adverse event/reaction reports arising from clinical trials on medicinal products for human use 
(‘CT-3’; 2011/C 172/01), June 2011) in the EU and with 21 CFR Part 312 to the FDA.  
Should an event also be considered as urgent safety measures, it must be reported in line with 
the requirements set out below. 
13.2.13 
Reporting Urgent Safety Measures 
Urgent safety measures can be put in place with immediate effect without needing to gain prior 
authorisation by the EC (and competent authority where applicable), in order to protect clinical 
trial participants from any immediate hazard to their health and safety. 

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Implementation of urgent safety measures should be notified immediately to the sponsor and 
CRO who will be responsible for onward reporting to the EC and competent authority.  
 
14 
Statistical Considerations  
14.1 
Statistical Analysis Plan 
All study data will be listed and all relevant data will be tabulated and summarised by dose 
level and overall where appropriate. A Statistical Analysis Plan (SAP) will be written and 
finalised prior to database lock. This plan will give a detailed description of all summaries and 
analyses that will be presented. 
Statistical analysis will be performed using SAS® (SAS Institute Inc., Cary, NC, USA) statistical 
software (Release 9.3 or later). 
14.2 
Procedures for Reporting Any Deviation(s) From the Original Statistical Plan  
Any deviations from the analyses planned in the protocol will be detailed in the SAP and 
deviations from the original statistical plan will be captured in the clinical study report. 
14.3 
Number of Patients 
The number of patients enrolled in the dose-escalation phase I/II study will depend on the 
number of DLTs observed in each cohort and the number of dose levels evaluated. The study 
will include a minimum of 14 and a maximum of 24 patients. A total of 14 patients will be 
enrolled at the selected terminal dose level. 
14.3.1 
Choice of Sample Size 
Because of the limited number of patients in the HB population (an estimated incidence of 1 in 
30,000 male births), the sample size of this study is based on pragmatic rather than statistical 
considerations.  
The 2 + 1 dose escalation aims to minimise the number of patients that would need to be 
dosed at suboptimal levels whilst allowing evaluation of safety, with the option to expand a 
group on observation of dose-limiting AEs.  
14.4 
Demographic and Baseline Characteristics 
Continuous variables will be summarised using number of observations, mean, and standard 
deviation, median, minimum, and maximum values. Categorical values will be summarised 
using number of observations and percentages. 
Medical history will be summarised using number of observations and percentages of patients 
reporting each category. 
Exposure to investigational product (i.e., total amount of study drug received) will be listed for 
all patients by dose level. 

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14.5 
Study Populations  
14.5.1 
Analysis Population 
The Full Analysis Set will include all patients who received FLT180a. This will be the primary 
population for all analyses of safety, efficacy and baseline characteristics. 
14.5.2 
The Per-protocol Population 
The Per-protocol Population will include all patients from the Full Analysis Set, excluding those 
patients with major protocol deviations (following the process described in section 14.11). This 
data will be used for sensitivity analyses.  
The Screened Set will include all patients screened. This set will be used for the listing and 
summaries of patient disposition and protocol deviations. 
14.6 
Primary Endpoint 
14.6.1 
Safety 
Adverse events will be coded using CTCAE version 5.0. Frequency of treatment-emergent 
AEs will be calculated for each body system and preferred term, and by dose level, for number 
of events and number of patients reporting the event. The severity of the TEAEs and the 
relationship to study medication will be summarised for each body system and preferred term 
by dose level.  
Withdrawals from the study will be summarised by dose level. 
Narratives will be presented for all deaths and patients reporting SAEs. 
Toxicities will be graded according to the NCI CTCAE version 5.0. 
14.6.2 
Efficacy 
The FIX response will be derived for each patient based on the FIX activity level measured at 
the central laboratory. Clinical FIX response is defined as achieving FIX activity of 5-150% and 
normalised FIX response is defined as achieving FIX activity in the normal range (50-150%). 
The two primary endpoints will be analysed on the full analysis set. The proportion of patients 
achieving clinical FIX response at Week 26, will be summarised for the patients who received 
the terminal dose level. For the primary analysis, last-observation-carried forward (LOCF) 
methodology will be used to impute any missing FIX value at Week 26, by the last non-missing 
FIX value.  For the primary analysis, FIX response will be assessed whether or not the patient 
is receiving or has been receiving immunosuppressants due to transaminitis outside the period 
of prophylactic immunosuppressant treatment. The proportion of patients also achieving 
normalization will be summarised on a similar manner. 
Sensitivity analyses of the primary endpoint will be performed: 
• 
The proportion of patients achieving clinical or normalised FIX response at Week 26 
with patients with a missing FIX response at Week 26 being considered as a non-
responder. 

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• 
The proportion of patients achieving clinical or normalised FIX response at Week 26 
excluding those patients who have received immunosuppressants in the previous 28 
days (4 weeks) before the Week 26 assessment, will also be summarised. 
• 
The proportion of patients achieving clinical or normalised FIX response at Week 26 
excluding those patients with major protocol deviations (per-protocol set) will also be 
summarised. 
More details and additional sensitivity analyses may be provided in the statistical analysis plan.  
14.7 
Secondary Endpoints 
14.7.1 
Safety 
Descriptive statistics (number of observations, mean, standard deviation, minimum, median, 
and maximum values) will be calculated for clinical laboratory tests at applicable visits. 
Changes from baseline will also be presented. For laboratory data, abnormal values will be 
flagged in the data listings. In particular, patients achieving FIX activity above 150% of normal 
will be summarised by dose and overall. 
Vital signs (systolic and diastolic blood pressure, temperature, and pulse) and physical 
examination will be summarised by dose level using appropriate descriptive statistics. 
Continuous variables will be summarised using number of observations, mean, standard 
deviation, minimum, median, and maximum values. Categorical values will be summarised 
using number of observations and percentages.  
14.7.2 
Endogenous FIX Production  
The FIX response will be derived for each patient based on FIX activity level measured at the 
central laboratory. Baseline FIX activity level will be defined as described in section 11.4.2.1. 
The proportion of patients achieving FIX activity at or above 5%, 15%, 30%, 40%, 50% and 
70% but no more than 150% of normal, at each scheduled visit, will be summarised by dose 
and overall. 
The proportion of patients achieving FIX activity at or above 5%, 15%, 30%, 40%, 50%, 70% 
and 150% of normal, at each scheduled visit, will be summarised by dose and overall. 
Change from baseline in FIX activity as a percentage of normal values will be summarised in 
tabular and graphical format for each patient, by dose and overall. 
14.7.3 
Haemostatic Effectiveness 
Annualised Bleeding Rate (ABR) 
The number of breakthrough bleeding episodes (spontaneous and traumatic) following 
FLT180a infusion will be annualised, and compared with the patient’s own baseline bleeding 
history (the annualised mean of 3 years of historical bleeding records, and, where possible, 
prospective data collected during the screening period will be used). 

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Factor IX Concentrate Consumption 
The dose (IU/kg) of factor concentrate used overall and by type of bleeding episode (i.e., 
spontaneous or traumatic) and location of bleeding episode (i.e., joint, soft tissue, or muscle) 
will be summarised with descriptive statistics. The total units of annualised factor consumption 
will be calculated and compared with the patient’s own baseline factor concentrate history (the 
annualised mean of 3 years’ historical factor concentrate consumption records, and, where 
possible, prospective data collected during the screening period will be used). 
In order to ensure enough time has elapsed for the patient to have endogenous FIX activity to 
protect the patient from spontaneous bleeding episodes, the calculation period for haemostatic 
effectiveness will be from day 15 inclusive to the date of completion of the last diary. More 
details will be provided in the SAP. 
14.7.4 
Shedding 
Clearance of vgs in plasma, saliva, urine, stool, and semen will be summarised. The time to 
unquantifiable result by body fluid will be summarized and listed. 
14.7.5 
Immune Response 
FIX 
Immune response to the FIX transgene product (i.e., development of inhibitors) will be 
assessed by measurement of the level of inhibitors. 
Descriptive statistics (number of observations, mean, standard deviation, minimum, median, 
and maximum values) will be calculated for immune response laboratory tests at applicable 
visits. 
14.8 
Exploratory Endpoints 
14.8.1 
Haemostatic Effectiveness 
If data permits, explorations of the correlation between FIX levels and bleeding events will be 
performed. For example, the proportion of bleeding events (all, treated, spontaneous) by FIX 
activity range of interest: <5%. 5-30%, 30-50% and >50% will be summarised and graphical 
displays produced. 
14.8.2 
Immune Response 
AAV-S3 Capsid 
Immune response to the AAV-S3 capsid will be assessed by measurement of the S3 
neutralising antibody titre. 
T-cell Responses to AAV-S3 Capsid in Peripheral Blood Mononuclear Cells 
Descriptive statistics (number of observations, mean, standard deviation, minimum, median, 
and maximum values) will be calculated for immune response laboratory tests at applicable 
visits. 

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14.8.3 
Health Economics 
Quality of life will be evaluated using the EQ-5D-5L and Haem-A-QoL.  
In addition, disability status will be assessed using the WHODAS 2.0 and the physical activity 
will be assessed using the HAL 2005. Haemophilia health status will be assessed using 
PROBE. Joint health and function will be evaluated using the HJHS score. 
Descriptive statistics will be applied to study the changes in the scores from baseline to end-of-
treatment in each dose level. 
Descriptive analyses of the PROBE questionnaire data will be performed separately. 
14.9 
Subgroup Analysis 
Subgroup analysis will be performed where appropriate including: 
• 
Patients receiving FIX on-demand therapy 
• 
Patients on conventional FIX prophylaxis therapy 
• 
Patients on extended half-life FIX prophylaxis therapy 
• 
Baseline FIX level 
• 
Baseline HJHS score. 
Additional subgroups may be defined in the SAP.  
A DMC will meet to review the safety results from a given cohort before escalating to the next 
dose in a new cohort. With the agreement or at the recommendation of the DMC, additional 
patients may be added to any cohort to characterise a DLT or any safety concern that has not 
met DLT stopping rules or the FIX response. 
14.10 
Handling of Missing Data  
Missing data is expected due to the patient population, and general features of clinical studies: 
• 
(D) 
patient died 
• 
(W) 
patient withdrew informed consent or became lost to follow-up 
• 
(M) 
assessment (may be a complete visit) could not be performed for a reason not 
covered by one of the reasons above. 
Number and percentage (based on the number of treated patients) of missing values and 
reasons (categories as defined above) will be summarised by visit for FIX levels. 
LOCF methodology will be used for the primary efficacy analysis, irrespective of the reason 
why the data is missing, if known. Sensitivity analysis will be performed as described in section 
14.6.2. Analyses will be performed considering all data observed for the respective analysis 
sets. For missing AEs and concomitant medication start and end dates the followings rules will 
be applied: 
Partial/missing start date: 

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• 
Missing day: impute the first of the month unless month is same as month of first dose 
of study drug, then impute first dose date. 
• 
Missing day and month: impute 1st January unless year is the same as first dose date, 
then impute first dose date. 
• 
Completely missing: impute first dose date unless the end date suggests it could have 
started before this, in which case impute the 1st January of the same year as the end 
date. 
Partial/missing end date: 
• 
Missing day: impute the last day of the month unless month is same as month of end of 
study visit, then impute day prior to end of study visit. 
• 
Missing day and month: impute 31st December unless year is the same as end of study 
visit, then impute day prior to end of study visit. 
14.11 
Protocol Deviations 
Protocol deviations will be addressed at monitoring on an ongoing basis. All protocol deviations 
are to be recorded with the indication of whether they are major as determined by the study 
management team, in cooperation with data management, medical monitoring, and 
the sponsor. These data will be imported into SAS and listed only, including the assignment of 
minor or major. A review of the protocol deviations will be performed before database lock. 
  
15 
Sponsor and Investigator Responsibilities 
This study is conducted in accordance with current applicable regulations, ICH, EU Directive 
2001/20/EC and its updates; The Medicines for Human Use (Clinical Trials) Regulation 
SI.2004/1031 and its updates; Advanced Therapy Medicinal Product Regulation 
1394/2007/EC or 21 CFR Part 312 in the United States and local ethical and legal 
requirements.  
The name and address of each third-party vendor (e.g., CRO) used in this study will be 
maintained in the investigator’s and sponsor’s files, as appropriate. 
15.1 
Sponsor’s Responsibilities 
15.1.1 
Good Clinical Practice Compliance 
The study sponsor and any third party to whom aspects of the study management or 
monitoring have been delegated will undertake their assigned roles for this study in compliance 
with all applicable industry regulations, ICH Good Clinical Practice (GCP) Guideline E6 (R2) 
2017, EU Directive 2001/20/EC, The Medicines for Human Use (Clinical Trials) Regulation 
SI.2004/1031 as amended, as well as all applicable national and local laws and regulations. 
Visits to sites are conducted by representatives of the study sponsor and/or the company 
organising/managing the research on behalf of the sponsor to inspect study data, patients’ 
medical records, and eCRFs in accordance with current GCP and the respective local and 

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international government regulations and guidelines. Records and data may additionally be 
reviewed by auditors or by regulatory authorities. 
The sponsor ensures that local regulatory authority requirements are met before the start of 
the study. The sponsor (or a nominated designee) is responsible for the preparation, 
submission, and confirmation of receipt of any regulatory authority approvals required prior to 
release of investigational product for shipment to the site. 
15.1.2 
Monitoring Requirement for the Trial 
The sponsor will determine the appropriate level and nature of monitoring required for the trial. 
Risk will be assessed on an ongoing basis and adjustments made accordingly. 
The degree of on-site monitoring (if required) will be proportionate to the objective, purpose, 
phase, design, size, complexity, blinding, endpoints and risks associated with the trial. 
A trial-specific monitoring plan will be established for studies. The trial will be monitored in 
accordance with the agreed monitoring plan which may include elements of remote monitoring 
in cases where appropriate. 
15.1.3 
Insurance 
The following statement applies to all sites excluding USA:  
University College London (UCL) holds insurance against claims from participants for injury 
caused by their participation in the clinical trial. Participants may be able to claim compensation 
if they can prove that UCL has been negligent. However, as this clinical trial is being carried 
out in a hospital, the hospital continues to have a duty of care to the participant of the clinical 
trial. University College London does not accept liability for any breach in the hospital’s duty 
of care, or any negligence on the part of hospital employees. This applies whether the hospital 
is an NHS Trust or otherwise.   
Participants may also be able to claim compensation for injury caused by participation in this 
clinical trial without the need to prove negligence on the part of UCL or another 
party.  Participants who sustain injury and wish to make a claim for compensation should be 
advised to do so in writing in the first instance to the Chief Investigator, who will pass the claim 
to the Sponsor’s Insurers, via the Sponsor’s office. 
Hospitals selected to participate in this clinical trial shall provide clinical negligence insurance 
cover for harm caused by their employees and a copy of the relevant insurance policy or 
summary shall be provided to University College London, upon request. 
The following statement applies to sites in USA: 
University College London holds insurance against claims from participants for injury caused 
by their participation in this clinical trial. Participants may be able to claim compensation if they 
can prove that UCL has been negligent. However, if this clinical trial is being carried out in a 
hospital, the hospital continues to have a duty of care to the participant of the clinical trial. 
University College London does not accept liability for any breach in the hospital’s duty of care, 

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or any negligence on the part of hospital employees. This applies whether the hospital is an 
NHS Trust or otherwise.   
Hospitals selected to participate in this clinical trial shall provide clinical negligence insurance 
cover for harm caused by their employees and a copy of the relevant insurance policy or 
summary shall be provided to UCL, upon request.  
15.1.4 
Public Posting of Study Information 
The sponsor is responsible for posting appropriate study information on applicable websites. 
Information included in clinical study registries may include participating investigators’ names 
and contact information.  
15.1.5 
Submission of Summary of Clinical Study Report to Competent Authorities 
and Ethics Committees 
The CRO on behalf of the sponsor will provide a summary of the clinical study report to the 
competent authority and, where applicable, the Independent EC of the member state(s) 
concerned as required by regulatory requirement(s) and to comply with the community 
guideline on GCP. This requirement will be fulfilled within 6 months of the end of the study 
completion date for paediatric studies and within 1 year for non-paediatric studies as per 
guidance.  
15.1.6 
Study Suspension, Termination, and Completion 
The sponsor may suspend or terminate the study at any time for any reason. If the study is 
suspended or terminated, the sponsor will ensure that applicable sites, regulatory agencies 
and IRBs/ECs are notified as appropriate. Additionally, the discontinuation of a registered 
clinical study which has been posted to a designated public website will be updated 
accordingly. 
The CRO will make an end-of-study declaration to the relevant competent authority as required 
by Article 10 (c) of EU Directive 2001/20/EC and The Medicines for Human Use (Clinical Trials) 
Regulation SI.2004/1031 as amended. 
15.1.7 
Notification of Serious Breaches to Good Clinical Practice and/or the 
Protocol  
A “serious breach” is a breach which is likely to effect to a significant degree: 
(a) the safety or physical or mental integrity of the patients of the trial; or 
(b) the scientific value of the trial. 
The sponsor of a clinical trial shall notify the licensing authority in writing of any serious breach 
of: 
(a) the conditions and principles of GCP in connection with that trial; or  
(b) the protocol relating to that trial, as amended from time to time, within 7 days of becoming 
aware of that breach. 

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The sponsor and CRO will be notified immediately of any case where the above definition 
applies during the trial conduct phase.   
15.2 
Investigator’s Responsibilities 
15.2.1 
Good Clinical Practice Compliance 
The investigator must undertake to perform the study in accordance with ICH GCP Guideline 
E6 (R2) 2017, EU Directive 2001/20/EC, The Medicines for Human Use (Clinical Trials) 
Regulation SI.2004/1031 as amended and applicable regulatory requirements and guidelines. 
It is the investigator’s responsibility to ensure that adequate time and appropriately trained 
personnel are available at the site prior to commitment to participate in this study. The 
investigator will maintain a list of appropriately qualified persons to whom the investigator has 
delegated significant study-related tasks, and shall, upon request of the sponsor, provide 
documented evidence of any licenses and certifications necessary to demonstrate such 
qualification. Curriculum vitae for investigators and sub-investigators are provided to the study 
sponsor (or designee) before starting the study. 
If a potential research patient has a primary care physician, the investigator should, with the 
patient’s consent, inform them of the patient’s participation in the study. 
15.2.2 
Protocol Adherence and Investigator Agreement 
The investigator and any co-investigators must adhere to the protocol as detailed in this 
document. The investigator is responsible for enrolling only those patients who have met 
protocol eligibility criteria. Investigators are required to sign an investigator agreement to 
confirm acceptance and willingness to comply with the study protocol. 
If the investigator suspends or terminates the study at their site, the investigator will promptly 
inform the sponsor and the IRB/EC and provide them with a detailed written explanation. The 
investigator will also return all study materials to the sponsor. Upon study completion, the 
investigator will provide the sponsor, IRB/EC, and regulatory agency with final reports and 
summaries as required by (inter)national regulations. 
Communication with local IRBs/ECs, to ensure accurate and timely information is provided at 
all phases during the study, may be done by the sponsor, applicable CRO, investigator, or for 
multicentre studies, the chief Investigator according to national provisions and will be 
documented in the investigator agreement. 
15.2.3 
Data Collection Tools and Source Document Identification 
Data will be collected from sites on trial-specific eCRFs.   
Source data are contained in source documents and must be accurately transcribed on to the 
eCRF. Examples of source documents are hospital records which include laboratory and other 
clinical reports etc. 
A source document identification list will be implemented prior to the start of the trial to identify: 
• 
which data is to be recorded directly onto the eCRF;  

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• 
which data is recorded firstly into source documents, such as medical notes, and then 
transcribed into the eCRF; and   
• 
which data is not to be recorded in the eCRF but only recorded in source documents, 
e.g., participant questionnaires and diary cards.  
It is the responsibility of the PI to ensure the accuracy of all data entered in the eCRF. The 
delegation log will identify all those personnel with responsibilities for data collection and 
handling. 
15.2.4 
Completing Case Report Forms  
All eCRFs must be completed and signed by staff that are listed on the site staff delegation 
log and authorised by the PI to perform this duty. The PI is responsible for the accuracy of all 
data reported in the eCRF. 
15.2.5 
Data Handling and Analysis 
A trial-specific data management plan will be in place for the trial. This will contain details of 
the software to be used for the database, the process of database design, coding, data entry, 
data quality checks, data queries, data security, database lock and data transfers. 
15.2.6 
Monitoring 
The investigator must permit authorised representatives of the sponsor, the respective 
national, local, or foreign regulatory authorities, the IRB/EC, and auditors to inspect facilities 
and to have direct access to original source records relevant to this study, regardless of media. 
The clinical research associate/study monitor (and auditors, IRB/EC or regulatory inspectors) 
may check the eCRF entries against the source documents.  
15.2.7 
Audit/Inspection 
To ensure compliance with relevant regulations, data generated by this study must be available 
for inspection upon request by representatives of, for example, the US Food and Drug 
Administration (as well as other US national and local regulatory authorities), the European 
Medicines Agency the Medicines and Healthcare Products Regulatory Agency, other 
regulatory authorities, the sponsor or its representatives, and the IRB/EC for each site. 
15.3 
Ethical and Regulatory Considerations 
15.3.1 
Informed Consent 
It is the responsibility of the investigator to ensure that appropriate informed consent has been 
obtained from all study patients prior to any study-related procedures. Signed ICFs must 
remain in each patient’s study file and must be available for verification at any time. 
The PI will provide the sponsor with a copy of the ICF that was reviewed by the IRB/EC and 
which received their favourable opinion/approval. A copy of the IRB/EC’s written favourable 
opinion/approval of these documents must be provided to the sponsor, prior to the start of the 
study unless it is agreed to and documented (abiding by regulatory guidelines and national 
provisions) prior to study start that another party (i.e., sponsor or chief Investigator) is 

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responsible for this action. Additionally, if the IRB/EC requires modification of the sample 
patient information and ICF provided by the sponsor, the documentation supporting this 
requirement must be provided to the sponsor. 
15.3.2 
Institutional Review Board or Ethics Committee 
It is the responsibility of the PI(s) to ensure that all aspects of institutional review are conducted 
in accordance with current governmental regulations. 
The sponsor must receive a letter documenting EC approval, which specifically identifies the 
protocol, protocol number, and ICF, prior to the initiation of the study. Protocol amendments 
will be subject to the same requirements as the original protocol. 
A progress report must be submitted to the EC at required intervals and not less than annually. 
At the completion or termination of the study, the investigational site must submit a close-out 
letter to the EC and the sponsor. 
15.3.3 
Protocol Amendments 
All proposed protocol amendments will be assessed by the sponsor and classified as 
substantial or non-substantial. Substantial amendments will require review and approval by 
the relevant IRB/EC and/or regulatory authority (depending on the nature of the amendment) 
prior to being implemented, except where required to prevent or ameliorate immediate harm 
to participants, see Section 13.2.13 Reporting Urgent Safety Measures. Other local or 
national approvals may be required and will be sought accordingly prior to implementation.  
15.4 
Record Keeping and Archiving 
At the end of the trial, all essential documentation must be archived securely by the chief 
investigator and trial sites according to ICH GCP requirements.  
Essential documents are those which enable both the conduct of the trial and the quality of the 
data produced to be evaluated and show whether the site complied with the principles of GCP 
and all applicable regulatory requirements.  
In accordance with regulations for ATIMPs, certain items must be archived for a 30-year period 
(see Section 15.4.1). All archived documents must continue to be available for inspection by 
appropriate authorities upon request 
Essential documents must be maintained and may not be destroyed without written permission 
from the sponsor. 
15.4.1 
Archiving Essential Trial Documentation Relating to Traceability  
In accordance with the Advanced Therapy Regulations (1394/2007/EC), all parties (the 
sponsor of the trial, the tissue establishments/procurement organisation (if applicable), the 
animal facility (if applicable), the manufacturer and the investigator/institution where the ATIMP 
is used) will keep their parts of the traceability records for a minimum of 30 years after the 
expiry date of the ATIMP, or longer if required by the terms of the clinical trial authorisation or 

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by the agreement with the sponsor. These requirements will be set out in contractual 
agreements between the parties and the sponsor. 
To comply with the regulatory requirements, each responsible party must ensure that the 
information relating to the traceability and accountability, from the production of ATIMPs to the 
participant receiving the ATIMPS are archived for a minimum 30 years after the expiry date of 
the ATIMP.  
The following essential documents/traceability data will be retained by the investigator and 
institution responsible for the human application of the ATIMP: 
Shipping Records for IMP: 
• 
Certificate of analysis of the IMP  
• 
Treatment allocation and decoding documentation  
• 
Participant identification code list  
• 
IMP accountability at the site including final disposition of both used and unused 
product. 
These records contain relevant information for traceability purposes and at least the following 
minimum data set from these records will be kept for 30 years after the expiry date of the 
product, or longer if required by the terms of the clinical trial authorisation or by the agreement 
with the sponsor: 
• 
Identification of the investigator/institution  
• 
Identification of the sponsor  
• 
Identification of the manufacturing site  
• 
Product name/code  
• 
Pharmaceutical form, route of administration, quantity of dosage units and strength  
• 
Batch and/or code number  
• 
Trial reference code  
• 
Trial participant code  
• 
Participant identification code list (links name of recipient to the trial participant code)  
• 
Product expiry/retest date  
• 
Date of administration  
• 
Records of any product that was unused or destroyed at site and its final status  
The participant medical records will contain the product name/code, the trial reference code, 
trial participant code and administration dates and dose in order to ensure that a link can be 
made back to the identity of the product and the further traceability records of the investigator 
and sponsor. 

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15.5 
Privacy and Confidentiality 
The sponsor and/or its representatives accessing the records and data will take all reasonable 
precautions in accordance with applicable laws, regulations, and guidelines to maintain the 
confidentiality of patients’ identities. UCL as Sponsor is the Data Controller as defined under 
the EU General Data Protection Regulation 2016 and Data Protection Act 2018. 
Patients are assigned a unique identifying number.  
The results of studies – containing patients’ unique identifying number and relevant medical 
records will be recorded. They may be transferred to, and used in, other countries which may 
not afford the same level of protection that applies within the countries where this study is 
conducted. The purpose of any such transfer would include: to support regulatory submissions, 
to conduct new data analyses to publish or present the study results, or to answer questions 
asked by regulatory or health authorities.  
 
16 
Publication Policy 
The sponsor intends to publish the results of this study in accordance with external guidelines 
regardless of whether the outcomes are perceived as positive, neutral, or negative.  
All publications relating to the sponsor’s projects must undergo appropriate technical and 
intellectual property review, and require sponsor agreement to publish prior to release of 
information. The review is aimed at protecting the sponsor’s proprietary information existing 
either at the commencement of the study or generated during the study.  
If the study is part of a multicentre study, the first publication of the study results shall be made 
by the sponsor in the form of a multicentre publication of the compiled and analysed study 
results. If such a multicentre publication is not submitted to a journal for publication by the 
sponsor within an 18-month period after conclusion, abandonment, or termination of the study 
at all sites, or after the sponsor confirms there shall be no multicentre study publication of the 
study results, an investigator may individually publish the study results from the specific site in 
accordance with this section. The investigator must, however, acknowledge in the publication 
the limitations of the single site data being presented. 
No publication that incorporates the sponsor’s confidential information shall be submitted for 
publication without the sponsor’s prior written agreement to publish, and shall be given to the 
sponsor for review at least 60 days prior to submission for publication. If requested in writing 
by the sponsor, the institution and PI shall withhold submission of such publication for up to an 
additional 60 days to allow for filing of a patent application.  
 
17 
Funding 
The trial is fully funded by Freeline Therapeutics Ltd. 
 

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18 
Statement of Compliance 
The trial will be conducted in compliance with the approved protocol, ICH GCP and applicable 
regulatory requirement(s). 
 

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19 
References 
_____________________________________________________________________________________________________ 
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3 Kundu RK, Sangiorgi F, Wu LY et al. Targeted inactivation of the coagulation factor IX gene causes 
hemophilia B in mice. Blood. 1998; 92:168-174. 
4 Bi L, Lawler AM, Antonarakis SE et al. Targeted disruption of the mouse factor VIII gene produces a 
model of haemophilia A. Nat.Genet. 1995; 10:119-121. 
5 Wang L, Zoppe M, Hackeng TM et al. A factor IX-deficient mouse model for hemophilia B gene therapy. 
Proc.Natl.Acad.Sci.USA. 1997; 94:11563-11566. 
6 Giles AR, Tinlin S, Hoogendoorn H et al. Development of factor VIII:C antibodies in dogs with 
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7 Evans JP, Brinkhous KM, Brayer GD et al. Canine hemophilia B resulting from a point mutation with 
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8 Nathwani AC, Davidoff A, Hanawa H et al. Factors influencing in-vivo transduction by recombinant 
adeno-associated viral vectors expressing the human factor IX cDNA. Blood. 2001; 97:1258-1265. 
9 Mingozzi F, Liu YL, Dobrzynski E et al. Induction of immune tolerance to coagulation factor IX antigen 
by in vivo hepatic gene transfer. J.Clin.Invest. 2003; 111:1347-1356. 
10 Limmer A, Ohl J, Kurts C et al. Efficient presentation of exogenous antigen by liver endothelial cells 
to CD8+ T cells results in antigen-specific T-cell tolerance. Nat.Med. 2000; 6:1348-1354. 
11 Nathwani AC, Reiss U.M, Tuddenham E.G.D et al. Long-Term Safety and Efficacy of Factor IX Gene 
Therapy in Hemophilia B. N.Engl.J.Med. 2014; 371:1994-2004. 
12 Nathwani AC, Rosales C, McIntosh J, et al. Long-term safety and efficacy following systemic 
administration of a self-complementary AAV vector encoding human FIX pseudotyped with serotype 5 
and 8 capsid proteins. Mol.Ther. 2011; 19:876-85. 
13 den Uijl IE, Fischer K, Van Der Bom JG, Grobbee DE, Rosendaal FR, Plug I. Analysis of low frequency 
bleeding data: the association of joint bleeds according to baseline FVIII activity levels. Haemophilia. 
2011; 17:41-4. 
14 George LA, Sullivan SK, Giermasz A et al. Hemophilia B Gene Therapy with a High-Specific-Activity 
Factor IX Variant. N.Engl.J.Med. 2017; 377:2215-2227. 
15 Nathwani AC, Gray JT, McIntosh J, et al. Safe and efficient transduction of the liver after peripheral 
vein infusion of self-complementary AAV vector result in stable therapeutic expression of human FIX in 
non-human primates. Blood. 2007; 109:1414-1421. 

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16 Nathwani AC, Gray JT, Ng CYC, et al. Self-complementary adeno-associated virus vectors containing 
a novel liver-specific human factor IX expression cassette enable highly efficient transduction of murine 
and non-human primate liver. Blood; 2006; 107:2653-2661. 
17 Nathwani AC, Tuddenham EGD, Rangarajan S, et al. Adenovirus-Associated Virus Vector-Mediated 
Gene Transfer in Hemophilia B. N.Engl.J.Med. 2011; 365:2357-2365. 
18 Tables 7.1 and 7.2 - World Federation of Hemophilia: Guidelines for the Management of Hemophilia. 
2nd Edition. 06 July 2012. https://www.wfh.org/en/resources/wfh-treatment-guidelines 
19 Committee for medicinal products for human use (CHMP) Guideline on clinical investigation of 
recombinant and human plasma-derived factor IX products. 21 July 2011. 
20 Nowrouzi A, Penaud-Budloo M, Kaeppel C et al. Integration frequency and intermolecular 
recombination of rAAV vectors in non-human primate skeletal muscle and liver. Mol.Ther. 2012; 
20:1177-1186. 
21 Li H, Malani N, Hamilton SR et al. Assessing the potential for AAV vector genotoxicity in a murine 
model. Blood. 2011; 117:3311-3319. 
22 Nault J-C, Datta S, Imbeaud S et al. Recurrent AAV2-realted insertional mutageneis in human 
hepatocellular carcinomas. Nat.Genet. 2015; 47:1181-1193. 
23 Donsante A, Miller DG, Li Y et al. AAV vector integration sites in mouse hepatocellular carcinoma. 
Science. 2007; 317:477. 
24 Chandler RJ, LaFave MC, Varshney GK et al. Vector design influences hepatic genotoxicity after 
adeno-associated virus gene therapy. J.Clin.Invest. 2015 ;125:870-880. 
25 Kay MA. AAV vectors and tumorigenicity. Nat.Biotechnol. 2007; 25:1111-1113. 
26 Gil-Farina I, Fronza R, Kaeppel C et al. Recombinant AAV intergration is not associated with hepatic 
genotoxicity in nonhuman primates and patients. Mol.Ther. 2016; 24:1100-1105. 
27 D’Avola D, Lopez-Franco E, Sangro B et al. Phase I open label liver-directed gene therapy clinical 
trial for acure intermittent prophyria. J.Hepatol. 2016; 65:776-783. 
28 Srivastava A and Carter BJ. AAV Infection: Protection for Cancer. Hum.Gene.Ther. 2017; 28:323-
327. 
29 Golden DB. Patterns of anaphylaxis: acute and late phase features of allergic reactions. Novartis 
Found.Symp. 2004; 257:101-10. 
30 Simioni P et al. X-Linked Thrombophilia with a Mutant Factor IX (Factor IX Padua). N.Engl.J.Med. 
2009 361:1671-1675 October 22, 2009. 
31 Crudele JM, Finn JD, Siner JI, et al. AAV liver expression of FIX-Padua prevents and eradicates FIX 
inhibitor without increasing thrombogenicity in hemophilia B dogs and mice. Blood. 2015; 125:10:1553-
1561. 
32 Ameri A, Kurachi S, Sueishi K, Kuwahara M, Kurachi K. Myocardial fibrosis in mice with 
overexpression of human blood coagulation factor IX. Blood. 2003; 101:1871-1873. 

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33 Chadeuf G, Favre D, Tessier J et al. Efficient recombinant adeno-associated virus production by a 
stable rep- cap HeLa cell line correlates with adenovirus-induced amplification of the integrated rep-cap 
genome [In Process Citation]. J.Gene.Med. 2000; 2:260-268. 
34 Nony P, Chadeuf G, Tessier J, Moullier P, Salvetti A. Evidence for packaging of rep-cap sequences 
into adeno-associated virus (AAV) type 2 capsids in the absence of inverted terminal repeats: a model 
for generation of rep-positive AAV particles. J.Virol. 2003; 77:776-781. 
35 Hauck B, Murphy SL, Smith PH et al. Undetectable transcription of cap in a clinical AAV vector: 
implications for preformed capsid in immune responses. Mol.Ther. 2009; 17:144-152. 
36 https://www.wfh.org/en/page.aspx?pid=643 
37 Manns MP, Czaja AJ, Gorham JD et al. Diagnosis and management of autoimmune hepatitis. 
Hepatology. 2010; 51:2193-2213. 
38 Blanchette VS, Key NS, Ljung LR et al. Definitions in haemophilia: communication from the SSC of 
the ISTH. J.Thromb.Haemost. 2014; 12:1935-1939. 
39 Human Gene Therapy for Hemophilia – Draft Guidance for Industry, U.S. Department of Health and 
Human Services, Food and Drug Administration, Center for Biologics Evaluation and Research, 
July2018 
https://www.fda.gov/downloads/BiologicsBloodVaccines/GuidanceComplianceRegulatoryInformation/ 
Guidances/CellularandGeneTherapy/UCM610801.pdf 
 
 

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Appendix 1: Treatment Management Plans 
_____________________________________________________________________________________________________ 
1.1 
Management of Thrombosis 
Patients will be informed in advance of potential early symptoms and signs of thrombotic 
phenomena, including pain and/or tenderness along a vein, swelling of an arm or leg without 
pain or tenderness, redness along a vein, low fever without any known reason (such as a cold 
or flu), sudden shortness of breath or difficulty breathing or coughing, sudden chest pain, 
sudden severe headache or changes in vision, and numbness or tingling in arms or legs. If 
such an event occurs while the patient is at home, the patient will be instructed to seek 
immediate medical care.  
In the event of a thrombotic event, it will be managed according to established guidelines: 
Prevention and treatment of venous thromboembolism: international consensus statement 
(guidelines according to scientific evidence). Clin. Appl. Thromb. Hemost. 2013 Mar-
Apr;19(2):116-231.  
1.2 
Management of Anaphylaxis 
Patients will be informed in advance of potential early symptoms and signs of hypersensitivity 
reactions, including hives, generalized urticaria, angioedema, chest tightness, dyspnea, 
wheezing, faintness, hypotension, tachycardia, and anaphylaxis. Patients will remain under 
observation in the investigational centre for at least 12 hours following infusion to minimise the 
risk associated with acute allergic reaction. Vital signs (pulse, blood pressure, respiration rate, 
and temperature) will be monitored at 15-minute intervals from the start of infusion. Vital signs 
will be monitored hourly for 6 hours following infusion and then every 2 hours for 6 hours. 
Should anaphylaxis occur, then initial management will consist of standard acute control 
measures - epinephrine, antihistamine and steroid, followed by intensification of immune 
suppression by means of continued steroid, cyclosporine A and cyclophosphamide.   
1.3 
Management of immune complex disease 
Immune complex disease will be managed by immune suppression with agents detailed in the 
following review: 
Houssiau FA, Lauwerys BR. Current management of lupus nephritis Best Practice & Research. 
Clinical Rheumatology. 27(2013):319–328. 
1.4 
Management of Inhibitors 
Studies in non-human primates have shown that immune modulatory treatment with rituximab  
and cyclophosphamide is effective at managing neutralising anti-human FIX antibodies 
following AAV mediated gene transfer.‡ This finding has been replicated in recent studies 
resulting in complete eradication of the neutralizing anti-hFVIII antibody in two animals and a 
partial reduction in antibody titre in another. Therefore, should a neutralising antibody response 

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to the transgenic protein be observed in patients in this study a rituximab based 
immunosuppressive regimen will be implemented.   
‡Nathwani AC, Gray JT, McIntosh J et al. Safe and efficient transduction of the liver after 
peripheral vein infusion of self complementary AAV vector results in stable therapeutic 
expression of human FIX in nonhuman primates. Blood. 2007;109:1414-1421. 
1.5 
Management of Infusion Reaction 
Whilst acute infusion reaction is deemed highly unlikely in view of its absence in over 50 
infusions of varying concentrations and amounts of AAV vector for gene therapy, patients will 
remain under observation in the investigational centre for at least 12 hours following infusion. 
Vital signs (pulse, blood pressure, respiration rate, and temperature) will be monitored at 15-
minute intervals from the start of infusion. Vital signs will be monitored hourly for 6 hours 
following infusion and then every 2 hours for 6 hours. Should it occur, infusion reaction will be 
managed in the same way as transfusion associated reaction.  
Faed J, 2014, www.nzblood.co.nz; Guidelines for management of adverse transfusion 
reactions. 
 
 
 

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Appendix 2: Management Guidelines for CMV Reactivation 
__________________________________________________________________________ 
Human cytomegalovirus (CMV) can result in opportunistic infection in patients requiring 
immunosuppression for their underlying immune disorders such as systemic lupus 
erythematosus but at a much lower frequency than organ transplantation and haematopoietic 
stem cell transplantation (Atabani et al., 2012; Lim et al., 2019). Nevertheless, surveillance for 
CMV reactivation is necessary in these individuals so that treatment can be instituted early. 
Natural history studies have demonstrated a correlation between replication kinetics, peak and 
cumulative viral load with CMV end-organ disease (Atabani et al., 2012). Systematic review 
and meta-analysis have also demonstrated the validity of viral load, as determined by real-
time PCR (qPCR), as an appropriate surrogate endpoint for predicting the development of 
CMV disease and guiding pre-emptive therapeutic intervention for the prevention of CMV 
disease (Griffiths et al., 2016; Natori et al., 2018).   
A CMV PCR titre > 3000 genomes/ml (or locally agreed on cut-off) should lead to firstly a re-
evaluation of the immunosuppression regimen, regardless of any observed clinical symptoms. 
In clinical study UCL15/0552, this would prompt a taper of corticosteroids. We would maintain 
treatment with tacrolimus to prevent recurrence of transaminitis. Secondly, treatment is 
clinically indicated if there are any symptoms suggesting CMV infection (Ljungman et al., 
2017). In which case we suggest one of the following: Valganciclovir 900 mg bd orally, 
Ganciclovir 5mg/Kg bd iv or Foscarnet 60mg/Kg tds IV. Treatment is continued until 2 negative 
DNA results are obtained. If a subsequent episode is detected after treatment has been 
stopped, treatment will be reinitiated only if viral load is greater than 3000 genomes/ml.  
 
Atabani, S. F., Smith, C., Atkinson, C., Aldridge, R. W., Rodriguez-Perálvarez, M., Rolando, 
N., Griffiths, P. D. (2012). Cytomegalovirus replication kinetics in solid organ transplant 
recipients managed by preemptive therapy. American journal of transplantation : official 
journal of the American Society of Transplantation and the American Society of Transplant 
Surgeons, 12(9), 2457-2464. doi:10.1111/j.1600-6143.2012.04087.x 
 
Griffiths, P. D., Rothwell, E., Raza, M., Wilmore, S., Doyle, T., Harber, M., Emery, V. C. 
(2016). Randomized Controlled Trials to Define Viral Load Thresholds for Cytomegalovirus 
Pre-Emptive Therapy. PLoS One, 11(9), e0163722. doi:10.1371/journal.pone.0163722 
 
Lim, C. C., Tan, B. H., Tung, Y. T., Huang, H., Hao, Y., Mok, I. Y. J., Choo, J. C. J. (2019). 
Risk-stratified approach to anti-viral prophylaxis against cytomegalovirus disease in 
glomerulonephritis and renal vasculitis treated with potent immunosuppressants. Infectious 
diseases (London, England), 51(10), 745-752. doi:10.1080/23744235.2019.1648855 
 
Ljungman, P., Boeckh, M., Hirsch, H. H., Josephson, F., Lundgren, J., Nichols, G.,  Disease 
Definitions Working Group of the Cytomegalovirus Drug Development, F. (2017). Definitions 
of Cytomegalovirus Infection and Disease in Transplant Patients for Use in Clinical Trials. 
Clin Infect Dis, 64(1), 87-91. doi:10.1093/cid/ciw668 
 
Natori, Y., Alghamdi, A., Tazari, M., Miller, V., Husain, S., Komatsu, T., Forum, C. M. V. C. 
(2018). Use of Viral Load as a Surrogate Marker in Clinical Studies of Cytomegalovirus in 

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Solid Organ Transplantation: A Systematic Review and Meta-analysis. Clin Infect Dis, 66(4), 
617-631. doi:10.1093/cid/cix793 
 
 
 

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Appendix 3: Management Guidelines for Tacrolimus Dosing 
__________________________________________________________________________ 
Introduction 
Tacrolimus, is a macrolide immunosuppressant produced by Streptomyces tsukubaensis and 
is classified as a calcineurin inhibitor (CNI). Tacrolimus is an active ingredient in different oral 
formulations and is available as immediate release (IR) capsules (Prograf) or as granules for 
suspension (Modigraf) or as prolonged release (PR) capsules (Advagraf). Alternative 
formulations available in Europe include, generic IR capsules Adoport and Tacni, or generic 
PR capsules, Tacforius. In addition, PR formulations are also available as Tablets (Envarsus). 
Prograf and Advagraf are used in >90% of patients in Europe for both de-novo and 
maintenance therapy. Patients should be maintained on a single formulation of tacrolimus with 
the corresponding dosing regimen (i.e. prescribing by brand). Inadvertent, unintentional or 
unsupervised switching of formulations is associated with clinically relevant differences in 
systemic exposure to tacrolimus, and can lead to increased incidence of side effects, including 
under- or over immunosuppression. 
 
Target tacrolimus whole blood trough level 
Target trough levels (equivalent AUC of 200 ng.h/mL) are approximately 15 ng/ml.  
 
Timing of trough level for tacrolimus dose modifications 
For tacrolimus dose modifications, tacrolimus blood concentrations samples should be 
determined at the end of dosing interval as follows: 
• 
Tacrolimus IR (administered twice daily, morning and evening): Blood samples should be 
taken at 12 hours in the morning after the previous days evening dose.  
• 
Tacrolimus PR (administered once daily in the morning): Blood samples should be taken 
at 24 hours after previous days dose.  
For both formulations, blood samples should be taken as close as possible to 12 and 24 hours 
after dosing for IR and PR respectively. A maximum time window of +/- 1 hour is acceptable.  
Patients attending the clinic for collection of trough blood samples, MUST be advised NOT to 
take their morning dose tacrolimus prior to attending the clinic. Patients should bring their 
tacrolimus medication with them, and should take their morning dose AFTER blood sample 
collection. 
 
Tacrolimus oral dose 
Initiation of tacrolimus 
Tacrolimus therapy should be initiated at a dose of 0.2 mg/kg/day given orally:  

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• 
IR: administered in two divided doses, morning and evening, 12 hours apart 
• 
PR: administered once daily administration at 24 hour intervals.  
(Note: PR must be taken in the morning, except in subjects who work night shift).  
Daily doses should be rounded to the nearest 0.5 mg. In case of IR, if the total daily dose is 
not divisible into two equal doses, then the higher dose should be administered in the morning 
(e.g. patient requiring 16.5 mg, IR 8.5 mg should be administered in the morning and IR 8.0 
mg in the evening). This is to account for a lower and more variable oral bioavailability of the 
evening dose (diurnal variation).  
The capsules should be swallowed with fluid, preferably water on an empty stomach or at least 
one hour before or two to three hours after a meal. Grapefruit juice or mixed fruit juice should 
not be taken with tacrolimus because it is an enzyme inhibitor that can interfere with the 
metabolism of tacrolimus. 
 
Dose adjustments 
The investigator should adjust the patient’s dose and modify the tacrolimus dose regimen to 
maintain tacrolimus trough levels targeted to 15 ng/mL (recommended range 10 to 15 ng/mL).  
Blood levels should be monitored frequently, at every blood draw, with the frequency of 
monitoring reduced to weekly once the desired exposure has been reached and once other, 
potentially interacting treatments, are being used at stable doses. Owing to long half-life of 
tacrolimus (approximately 1 day in adults), it is recommended that dose adjustments are 
limited to twice per week as changes in trough levels will occur slowly, usually stabilising 48 to 
72 hours after the dose adjustment is made. In clinical practice, first opportunity for dose 
adjustment occurs on Day 2 to 3 after initiation of therapy. Ranges of expected blood levels in 
Caucasian subjects initiated at a dose of 0.2 mg/kg/day are as follows: 
 
Table 1: Proportion of patients achieving tacrolimus whole blood trough levels in target range 
of 10 – 15 ng/ml (expected in Caucasian patients). 
Formulation 
Day after 
start of 
tacrolimus 
Approximate number of patients with blood levels 
(% of patients) 
< 5 
(ng/mL) 
≥5-<10 
(ng/mL) 
≥10-<15 
(ng/mL) 
>15 - ≤20 
 (ng/mL) 
>20 - ≤30 
 (ng/mL) 
>30 
(ng/mL) 
IR 
2 
5 
30 
25 
20 
15 
5 
3 
5 
20 
25 
15 
30 
5 
4 
5 
10 
40 
30 
10 
5 
PR 
2 
5 
35 
30 
5 
15 
10 
3 
5 
35 
20 
20 
10 
10 
4 
5 
20 
40 
25 
15 
5 
 
Recommended dose adjustments based on trough levels 

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Dose adjustments (increase or decrease) should be made to maintain tacrolimus level within 
the recommended target range. The extent of dose changes should be based on observed 
tacrolimus levels on Days 2 and 3 after start of therapy as described below: 
 
Table 2: Suggested dose adjustments based on trough levels Day 2/3. 
Observed blood levels 
(ng/mL) 
Dose adjustment 
≤ 5 ng/mL 
Increase dose by 50%. Subsequent dose adjustment should be 
considered after 3 days if necessary, taking into consideration 
new trough levels 
>5 to ≤10 ng/mL 
Increase dose by 30%. Subsequent dose adjustment should be 
considered after 3 days if necessary, taking into consideration 
new trough levels 
>10 to ≤15 ng/mL 
No change in dose required 
>15 to ≤20 ng/mL 
Decrease dose by 10-20%. Subsequent dose decrease should 
be considered after 3 days if tacrolimus trough levels remain 
above 15 ng/mL 
>20 to ≤30 ng/mL 
Decrease dose by 20%. Subsequent dose decrease should be 
considered after 3 days if tacrolimus trough levels remain above 
20 ng/mL 
>30 ng/mL 
Decrease dose by 30%. Subsequent dose decrease should be 
considered after 3 days if tacrolimus trough levels remain above 
30 ng/mL 
 
 

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Appendix 4: Summary of Protocol Changes 
_____________________________________________________________________________________________________ 
Version 1.0 to 2.0 
Section  
Page 
Number(s) 
Summary of Change 
5.2 
21 
Detailed text for non-clinical summary has been deleted. Full reference for non-
clinical data now resides in the investigator brochure. 
5.3 
22 
Administrative changes to phrasing of text for clarity. 
5.4.1 
24-25 
Amended text related to dose justification. 
5.4.2 
29 
 
Under Heading Thrombogenicity. Inclusion of data relating to findings in GLP 
toxicology program. 
26-30 
Administrative changes to phrasing of text for clarity. 
30 
Under Heading Risks associated with DNA impurities. Removal of redundant text 
related to non-clinical studies. 
 
Version 2.0 to Version 3.0 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number and addresses of CI and Sponsor. 
2 
9, 10 
List of Abbreviations updated where required in line with content of protocol. 
3 
11 
Trial Personnel contact details updated where required. 
4, 14.3 
12, 69 
Increase of maximum number of patients that maybe enrolled from ‘18’ to ‘24’. 
4, 12.3.3, 
14.3 
12, 14, 62, 
63, 69 
Increase in number of patients to be enrolled at the terminal dose level from ‘6’ to 
‘14’. 
4, 12.3.2, 
12.3.3, 
14.9 
12, 13, 62, 
63, 72 
Removal of the specified maximum of 6 patients can be treated at a single dose 
level. Additional patients (number unspecified) can be added at any dose level at 
the request of the trial management group and data monitoring committee and at 
discretion of the Sponsor. 
4, 5.1 
13, 23 
Addition of sentence to Rationale ‘Even with extended half-life (EHL) factor 
concentrates frequent infusions are required for the life time of the patient.’ 
4, 5.4.1 
13, 26 
Update in potential disease phenotype from ‘milder forms’ to ‘mild/normal’. 
4, Table 3, 
12.3.1 
13, 27, 61 
Update in dose of Cohort 3 (high dose) from ‘6 x1012 vg/kg’ to ‘4 x 1012 vg/kg’.  
4, 5.4.1, 
7.1, 12.3.1, 
12.3.3 
13, 27, 35, 
61, 62, 63 
Update to reflect that the dose level of cohort maybe reduced dependent on the 
FIX response seen, to avoid exposing patients to levels exceeding the normal 
physiological range. Dose reduction steps are specified. 
4, 5.4.1, 
7.1, 12.1.2, 
12.3.2, 
12.3.3 
13, 14, 27, 
35, 36, 60, 
62, 63 
Update to provide clarity and correct omissions that safety and efficacy outcomes 
and dose escalation/reduction and selection of terminal dose level will be over 
seen by ‘a trial management group and a data monitoring committee’ not just ‘a 
data monitoring committee’. 
4, 5.4.1, 
12.3.3 
13, 27, 62, 
63 
Update to the goal for FIX activity from ‘40-100%’ to ‘70-150%’. FIX activity of 70-
150% representing normalisation and a level at which patients may undergo 
surgical procedures without a requirement for exogenous factor concentrates. 
4, Table 1, 
5.4.1, 
5.4.2, 7.1, 
9.3, 9.4, 
11.2.1 
13, 18, 19, 
20, 28, 29, 
30, 36, 42, 
43, 44, 49 
Update to the corticosteroid regimen from ‘6’ to ‘8’ weeks and a change from 
‘weeks 6-12’ to weeks ‘weeks 4-12’. 
4, 8.1 
14, 38 
Update in Inclusion Criteria 2, a, ii from ‘On-demand therapy with current a or past 
history of 4 or more bleeding episodes/year, or’ to ‘On-demand therapy with a 
history of 4 or more bleeding episodes/year on average over the past 3 years, or’. 
4, 8.1 
14, 38 
Update in Inclusion Criteria 5 from ‘Lack of neutralising anti-AAV-S3 antibodies 
using an in vivo transduction inhibition assay’ to ‘Lack of neutralising anti-AAV-S3 
antibodies using an in vivo transduction inhibition assay within 4 weeks of vector 
administration’. 
4, 5.4.2, 
8.2 
14, 32, 39 
Addition of an Exclusion Criteria 2 (previous Exclusion Criteria #’s from 2 onwards, 
increased by 1) ‘Patients at high risk of thromboembolic events (high risk patients 
would include those with a history of arterial or venous thromboembolism (e.g. 
deep vein thrombosis, pulmonary embolism, non-haemorrhagic stroke, arterial 

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embolus) and those with known inherited or acquired thrombophilia including 
conditions such as atrial fibrilation)’. 
4, 8.2 
14, 39 
Update in Exclusion Criteria 6 (previously 5) from ‘Evidence of liver dysfunction 
(persistently elevated alanine transaminase >1.5 x upper limit of normal)’ to 
‘Evidence of liver dysfunction (persistently elevated alanine aminotransaminase, 
aspartate aminotransferase, bilirubin >1.5 x upper limit of normal)’. 
4, 8.2 
14, 39 
Update in Exclusion Criteria 10 (previously 9) from ‘Patients with uncontrolled 
cardiac failure or unstable angina’ to ‘Patients with uncontrolled cardiac failure, 
unstable angina or myocardial infarction in the past 6 months’. 
4, 5.4.2, 
8.2 
15, 30, 39 
Update in Exclusion Criteria 13 (previously 12) from ‘Known or suspected 
intolerance or hypersensitivity to the investigational product of its excipients’ to 
‘Known or suspected intolerance, hypersensitivity or contraindication to the 
investigational product and non-investigational medicinal products or their 
excipients’. 
4, 8.2 
15, 39 
Addition of an Exclusion Criteria 14 ‘Planned major elective surgery prior to the 
end of trial’. 
4, 7.2.2, 
14.7.1 
15, 16, 37, 
71 
Addition of Secondary endpoint, Endogenous (hFIX) production of ‘The proportion 
of patients achieving hFIX activity at or above 70% of normal at week 26’ and 
updates to applicable sections of statistical analysis section. 
4, Table 1, 
7.2.3, 
11.1.1, 
11.4.4.1, 
14.8.1 
15, 18, 19, 
20, 37, 46, 
56, 72 
Addition of the ‘EQ-5D-5L’ patient report outcome to the study. Reflected in 
Scheduled of Assessments, trial design, study procedures and statistical analysis 
sections. 
Table 1, 
11.1.1, 
11.1.2, 
11.2.1, 
11.2.1.1, 
11.3, 
11.4.2.3, 
11.4.3.2, 
11.4.3.5, 
11.4.3.6 
18, 19, 20, 
21, 22, 45, 
46, 47, 48, 
49, 50, 51, 
53, 54, 55 
Updates to ‘Table 1: Schedule of Assessments’ to split out the requirements for 
the Infusion Week (Day -1 to Day +4) by creation of a second table ‘Table 2: 
Detailed Schedule of Assessments for Infusion Week’. Wider formatting/order/ 
rationalisation of assessments listed and adjustment of table footnotes for 
accordingly. Implicated changes in section 11, Study Procedures. 
Table 1, 
Table 2, 
7.1, 11.1.2 
18, 19, 20, 
21, 22, 34, 
46 
Addition of a window to the Day -1 visit which may now be conducted as early as 
Day -3 for logistical reasons. 
Table 1, 
Table 2 
18, 19, 20, 
21, 22 
Addition of a window to the Day +4 visit which may now be conducted +/- 1 day. 
Table 1, 
11.1.1, 
11.4.1.5 
18, 19, 20, 
45, 52 
Addition of Target Joint Assessment at Screening visit. 
Table 1, 
11.2.1, 
11.3 
11.4.3.2 
18, 19, 20, 
49, 50, 55 
Addition of Liver Function Test (Local) at Week 14, Week 16, Week 20 and Week 
26/EOS visits. 
Table 1, 
11.2.1, 
11.3, 
11.4.2.3 
18, 19, 20, 
49, 50, 53 
Addition of hFIX Activity Level (Local) at Week 14, Week 16, Week 20 and Week 
26/EOS visits. 
Table 1, 
Table 2, 
11.4.1.6, 
11.4.3.4 
18, 19, 20, 
21, 22, 52, 
55 
Addition of requirement for primary and back-up samples for AAV Antibody 
Screen, hFIX Activity Level (Central) and AAV-S3 Antibody Titre. 
Table 1, 
11.1.1, 
11.4.1.6 
18, 19, 20, 
45, 52 
Confirmation that AAV Antibody Screen is via transduction inhibition assay and 
that repeat testing may be indicated during long screening period.  
Table 1, 
Table 2 
18, 19, 20, 
21, 22 
Liver Function Test (Local) to include ‘Albumin, alkaline phosphatase, direct 
bilirubin, indirect bilirubin, total bilirubin, alanine aminotransferase, aspartate 
aminotransferase’ added to Schedule of Assessments for clarity. 
Table 1, 
11.2.1.1 
18, 19, 20, 
49, 50 
During intensive monitoring period for Liver Function Tests (Local) and hFIX 
Activity Level (Local) a recommendation on how the test should be distributed 
through the week. 
‘Three times per week. The tests should be as evenly spaced through the week as 
possible, for example: Monday, Wednesday and Friday.’ 

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‘Twice weekly. The tests should be as evenly spaced through the week as 
possible, for example: Monday and Thursday or Tuesday and Friday.’ 
Table 2, 
11.1.2 
21, 22, 46 
Clarification that the pre-infusion consent may occur prior to Day -1. 
Table 2 
21, 22 
Haematology, Chemistry incl. CRP, Coagulation Screen (Local) to include 
‘Complete blood count with differentials, sodium, potassium, phosphate, blood 
urea nitrogen, serum creatinine, CRP, prothrombin time, activated partial 
thromboplastin time’ added to Schedule of Assessments for clarity. 
5.3, 5.4.1, 
5.4.2, 7.1, 
12.3.3 
25, 28, 35, 
36, 62, 63 
Addition of text describing outcomes from the George et al, NEJM 2017 paper 
around the asymptomatic increase in ALT been seen as early as week and the 
FIX activity levels at week 4 representing ~75% of levels seen at steady state. 
5.4.1 
27 
Addition of ‘In vivo data in mice models are not predictive of transgene expression 
in humans and therefore the proposed doses have been estimated based on 
extrapolation of in vitro data in human cells comparing the performance of 
elements of the AAV2/8-LP1-hFIXco construct used in the previous study by 
Nathwani et al. and those incorporated in FLT180a.11,15,16,17 Please refer to the 
investigator brochure for full details.’ to support rational for dose selection. 
5.4.1 
28 
Supplementary updates to the data reported in the protocol from the pre-clinical 
work. 
5.4.1, 7.1., 
12.3.3 
27, 28, 35, 
36, 62, 63 
Updates to the dose escalation rules, selection of target dose criteria and criteria 
for dose reduction. 
5.4.1 
28 
Addition of ‘Analysis of the data from the studies conducted by Nathwani et al11,17 
has shown a marked effect of corticosteroid treatment on FIX activity levels. An 
increase of 2.5 to 3-fold in FIX levels have been observed in parallel with the 
prednisolone regimen (unpublished data), these return to pre-steroid levels 
approximately 5 weeks following cessation. The steroid response with the smaller 
 promoter used in FLT180a is expected to be less profound but not absent 
and therefore pre-steroid FIX levels (week 4) will be used to guide dose escalation 
decisions.’ to support rationale for dose escalation. 
5.4.2 
32 
Addition of ‘The World Federation of Hemophilia (WFH) regard 150% as the upper 
limit of normal for Factor IX activity.’ to support risk/benefit review for 
thrombogenicity. 
5.4.2 
33 
Supplementary updates to the data reported to support risk/benefit review for Risk 
associated with DNA impurities. 
7.1, 10.3 
35, 45 
Update in guidance on when prophylactic factor concentrate maybe ceased in line 
with clinical practice, from ‘Only after FIX activity levels ≥3% are reached at 2 
consecutive measurements will the patients be asked to stop their prophylactic 
FIX treatment.’ to ‘If FIX activity levels ≥3% are reached then prophylaxis will be 
held pending a repeat analysis within a period of 72 hours. If the FIX activity levels 
are ≥3% at that time then prophylaxis will be stopped with continued/regular 
assessment of FIX activity levels and occurrence of spontaneous bleeding.’ 
Figure 1 
36 
Removal of Figure 1: Study Schematic. 
9.2.4 
41 
Removal of ‘on a per patient basis’ so number of amount of IMP that can be 
shipped to infusion site is not limited to allow some flexibility for scheduling. 
9.3 
42 
Update in tablet doses available of prednisolone. 
10.2 
44 
Addition of ‘time allowing, i.e. investigator permission not required for emergency 
medical care.’ as previous wording could have implied this was not allowed. 
11.4.1.2 
51 
Update to reflect correct method of derivation of the patient number. 
11.5 
57, 58, 59 
Volume of blood to be drawn from each patient updated to reflect updated 
amounts that will be required. 
12 
59 
Addition of ‘The sponsor will retain final responsibility for decision making on all 
aspects of dose progression, dose escalation/reduction and stopping rules.’ 
12.3.2 
62 
Update from ‘If DLT observed in third patient, apply temporary halt to trial and 
consult competent authority.’ to ‘If DLT observed in third patient and this is second 
patient in which a DLT has been observed, apply temporary halt to trial and 
consult competent authority.’ for clarification. 
12.5 
64 
Removal of stopping rule ‘Persistence of vgs in semen >3 months’. 
13.2.11 
68 
Update from ‘The CRO is responsible for notifying the relevant regulatory 
authorities and Central Ethics Committees(ECs)/Institutional Review Boards 
(IRBs) of related, unexpected SAEs.’ to ‘The CRO is responsible for notifying the 
relevant regulatory authorities and Central Ethics Committees(ECs)/Institutional 
Review Boards (IRBs) of related, unexpected SAEs (SUSARs) in accordance with 
the regulatory requirements. SUSARs that are fatal or life-threatening must be 
notified with 7 days from the CRO learning of them with any additional relevant 
information reported within a further 8 days. Other SUSARs must be reported 

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within 15 days from the CRO learning of them. The CRO will also inform all 
participating sites.’ 
14.10 
72 
Update from  
‘Partial/missing end date: 
• 
Missing day: impute the last day of the month unless month is same as month 
of last dose of study drug, then last dose date. 
• 
Missing day and month: impute 31st December unless year is the same as 
end of study visit, then impute day prior to end of study visit.’ to 
‘Partial/missing end date: 
• 
Missing day: impute the last day of the month unless month is same as month 
of end of study visit, then impute day prior to end of study visit. 
• 
Missing day and month: impute 31st December unless year is the same as 
end of study visit, then impute day prior to end of study visit.’ to correct 
inaccuracies. 
15 
73 
Addition of ‘Advanced Therapy Medicinal Product Regulation (1394/2007/EC) or 
21 CFR Part 312 in the United States’. 
15.3.3 
77, 78 
Addition of section 
15.3.3  Protocol Amendments 
All proposed protocol amendments will be assessed by the sponsor and classified 
as substantial or non-substantial. Substantial amendments will require review and 
approval by the relevant IRB/EC and/or regulatory authority (depending on the 
nature of the amendment) prior to being implemented, except where required to 
prevent or ameliorate immediate harm to participants, see Section 13.2.13 Urgent 
Safety Measures. Other local or national approvals may be required and will be 
sought accordingly prior to implementation.  
15.5 
79 
Addition of ‘UCL as Sponsor is the Data Controller as defined under the EU 
General Data Protection Regulations 2016’ and removal of any references that 
initials or date of birth will or might be collected. 
19 
81, 82, 83 
Duplication of references noted and updated, numbering addressed. Due to 
Endnote autoformatting issues, difficult to retain integrity of structure of document 
with full tracked changes of updates to numbering. All new references and deleted 
references indicated as such in tracked changes version.  
All 
All 
Appropriate spelling, grammatical, formatting and minor updates and 
addition/removal/update of single words for clarity/consistency of text throughout 
protocol. 
 
Version 3.0 to Version 3.1 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number. 
4, 8.2 
14, 39 
Update in Exclusion Criteria 2 from ‘Patients at high risk of thromboembolic events 
(high risk patients would include those with a history of arterial or venous 
thromboembolism (e.g. deep vein thrombosis, pulmonary embolism, non-
haemorrhagic stroke, arterial embolus) and those with known inherited or acquired 
thrombophilia including conditions such as atrial fibrilation)’ to ‘Patients at high risk 
of thromboembolic events (high risk patients would include those with a history of 
arterial or venous thromboembolism (e.g. deep vein thrombosis, pulmonary 
embolism, non-haemorrhagic stroke, arterial embolus) and those with acquired 
thrombophilia including conditions such as atrial fibrilation)’. 
 
Version 3.1 to Version 4.0 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number/date and addition of IND#. 
4, Table 1, 
5.4.1, 
5.4.2, 7.1, 
9.3, 9.4, 
11.2.1, 
11.2.1.1 
13, 17, 18, 
19, 28, 29, 
32, 35, 41, 
42, 43, 44, 
49, 50 
Extensive updates to required corticosteroid prophylaxis regimen and actions in 
response to elevations in ALT rises. Associated updated to LFT monitoring and 
test of reactivation of hepatitis. 

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Table 1, 
11.2.1, 
11.3, 
11.4.6.5, 
11.5 
17, 18, 19, 
49, 50, 57, 
58 
Addition of samples for ‘FIX activity research sample’ and applicable updates to 
study schedule and study procedures/assessments sections. 
Table 2, 
11.4.3.2 
20, 21, 54 
Correction of error to reinstate ‘platelets’ as a specified test within the 
haematology local laboratory panel. 
5.4.1 
29 
Update of sentence ‘In macaques, neutralising antibodies have been observed in 
a small proportion of animals exposed to human GTMP’ to ‘In macaques, 
neutralising antibodies have been observed in a small proportion of animals 
exposed to human GTMP, including FLT180a.’ 
9.2.2 
39 
Update to correct referenced regulation from ‘Annex 13 (EudraLex Volume 4) in 
the EU’ to ‘(EudraLex Volume 4 GMP, Part III Annex 13 and Part IV in the EU)’. 
11.1.1 
46 
Addition of ‘A patients alcohol intake for the course of the trial should also be 
discussed at the screening visit with a recommendation to moderate their intake 
for the duration of the trial. This discussion should be ongoing throughout the 
trial.’. 
11.1.2 
47 
Correction in error which specifies a 200 mL infusion volume of the vector. 
Adjusted to ‘Calculated FLT180a vector dose…’. 
11.4.4 
56 
Addition of section on ‘Vector Shedding’ to study procedures and assessments 
section which has been previously omitted in error. 
11.5 
58 
Adjustment of ‘Volume of blood to be drawn from each patient’ to reflect addition 
of FIX activity research plasma sample and back-up sample collection for routine 
FIX activity sampling which was omitted in error. 
12.3.3, 
12.5, 
13.2.9 
63, 64, 68 
Updates to clarify definition with regards to confirming inhibitor and associated 
impact should an inhibitor be identified. 
13.2.12 
69 
Addition of the following ‘Expedited reporting and reporting of adverse events will 
be in accordance with EudraLex Volume 10, Chapter II (Detailed Guidance on the 
collection, verification and presentation of adverse event/reaction reports arising 
from clinical trials on medicinal products for human use (‘CT-3’; 2011/C 172/01), 
June 2011) in the EU and with 21 CFR Part 312 to the FDA.’. 
19 
81, 82, 83 
Addition of supporting references.  
All 
All 
Appropriate spelling, grammatical, formatting and minor updates and 
addition/removal/update of single words for clarity/consistency of text throughout 
protocol. 
 
Version 4.0 to Version 5.0 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number. 
4, Table 1, 
5.4.1, 7.1, 
9.4, 11.2.1, 
12.3.3 
13, 17, 18, 
19, 20, 28, 
29, 30, 36, 
43, 44, 50, 
63, 64 
Update to required corticosteroid prophylaxis regimen to commence at Week 3 
and test for reactivation of hepatitis. Update to actions to be taken in response to 
break-through transaminitis. 
Table 1, 
11.2.1.1, 
11.4.3.2, 
11.5 
17, 18, 19, 
20, 50, 55, 
59, 60 
Addition of local haematology samples to be conducted in line with regular local 
LFT monitoring. 
Table 1, 
11.2.1, 
11.3, 
11.4.6.4, 
11.5 
17, 18, 19, 
20, 50, 51, 
58, 59, 60 
Update to reflect collection of Research Plasma Samples at study visits with no 
dependency on an elevation in ALT. 
12.2 
61 
Update to definition for DLT from ‘A DLT is defined as any grade 3 or greater AE 
at least possibly related to FLT180a.’ to ‘A DLT is defined as any grade 3 or 
greater AE at least possibly related to FLT180a except for increases in ALT or 
AST that are not associated with increases in bilirubin. 
5.4.1, 
12.3.3 
28, 63, 64 
Updates to reflect that Week 3 values (last point pre-steroid from V5 of protocol) 
represents ~65% of steady state values and proportional updates to the guide 
values provided in the dosing rules. 

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All 
All 
Appropriate spelling, grammatical, formatting and minor updates and 
addition/removal/update of single words for clarity/consistency of text throughout 
protocol. 
 
Version 5.0 to Version 6.0 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number. 
4, 6.1, 7.1, 
7.2.1, 
14.6.2 
12, 14, 15, 
16, 37, 39, 
40, 76, 77 
Addition of a primary efficacy objective ‘To assess Factor IX FIX levels following 
systemic administration of FLT180a, at the terminal dose level’ and applicable 
associated updates to overall design, endpoints and statistical sections. 
4, 6.2, 6.3, 
7.2.2, 
7.2.3, 
14.7.5, 
14.8.1 
12, 15, 17, 
37, 40, 78 
Removal from secondary objective and addition to exploratory objective ‘To 
assess the immune response to the adeno-associated virus (AAV)-S3 capsid 
proteins following systemic administration of FLT180a’ and applicable associated 
updates to endpoints and statistical sections. Correction to provide clarification in 
ambiguity in previous versions of protocol of where this objective should sit. 
4, Table 1, 
Table 2, 
6.3, 7.2.3, 
11.1.1, 
11.1.2, 
11.3, 
11.4.5.4, 
14.8.2 
12, 15, 18, 
19, 20, 21, 
22, 23, 24, 
37, 41, 50, 
51, 55, 61, 
78 
Addition of the PROBE patient reported outcome to the study and applicable 
associated updates to objectives, endpoints, study schedule, study procedures/ 
assessments and statistical sections. 
4, Table 1, 
Table 2, 
7.1, 11.1.1, 
11.1.2 
13, 14, 18, 
19, 20, 21, 
22, 23, 24, 
38, 48, 49, 
50, 51, 52 
Extension of the screening period from ‘up to 14 weeks’ to ‘up to 52 weeks’ and 
applicable associated updates to study schedule and study procedures/ 
assessments sections 
4, 7.2.2, 
14.6.1, 
14.7.1 
15, 16, 40, 
76, 77 
Update of general safety endpoints from primary to secondary and applicable 
associated updates to statistical section. Correction to provide clarification in error 
in previous versions of protocol. 
4, 7.2.2, 
14.7.2 
15, 16, 40, 
77 
Update to endogenous FIX production secondary endpoints and applicable 
associated updates to statistical section. Updates further to addition of primary 
efficacy endpoint.   
4, 8.2 
14, 43 
Addition of exclusion criteria 15 ‘Current or relavant history of a physical of 
psychiatric illness or any medical condition that in the opinion of the investigator 
could affect the patients safety or interfere with the study assessments’. 
4, 13.2.1, 
14.6.1 
14, 16, 71, 
76 
Update to CTCAE version from V4.03 to V5.0. 
4, Table 1, 
Table 2, 
7.2.3, 
11.1.2, 
11.3, 
11.4.5.5 
15, 18, 19, 
20, 21, 22, 
23, 24, 41, 
51, 55, 61 
Update to Health resource utilisation endpoints from ‘Number of haemophilia 
related medical appointments’ to ‘Number of haemophilia related medical 
appointments and medical activities’ and Number of physiotherapy sessions’ to 
‘Number of physiotherapy sessions, specialist consultations and appointments 
with professional caregivers’ and applicable associated updates to the study 
procedures/assessments section. Clarifications also made to confirm when health 
resource utilisation data should be collected and the periods under review at each 
time point with updates to the study schedule, study procedures/assessments 
sections. 
Table 1, 
5.4.2 
18, 19, 20, 
21, 31, 32 
Updates to requirements around confirmation of liver health prior to dosing with 
FLT180a and applicable associated updates to the study schedule and risks and 
benefits sections. 
Table 1, 
11.1.1, 
11.4.1.7, 
11.5 
18, 19, 20, 
21, 49, 57, 
63, 64 
Update to include HCV RNA viral load testing at screening as required with 
updates to the study schedule and study procedures/assessments sections. 
Table 1, 
Table 2, 
11.1.2, 
11.2.1, 
11.3, 
11.4.6.3, 
11.4.6.4 
18, 19, 20, 
21, 22, 23, 
24, 51, 54, 
55, 62 
Updates to provide clarity that the Mononuclear Cells (Research) and Research 
Plasma Samples are ‘Optional’ samples in the trial with updates to the study 
schedule and study procedures/assessments sections.  

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Table 1, 
Table 2, 
11.1.2, 
11.2.1, 
11.3, 
11.4.6.6, 
11.5 
18, 19, 20, 
21, 22, 23, 
24, 51, 54, 
63, 64 
Addition of Immune Response Research Plasma Samples at Day -1 and EOS and 
to be taken throughout the trial in response to breakthrough transaminitis with 
updates to the study schedule and study procedures/assessments sections. 
Table 1, 
Table 2, 
5.4.2, 
11.2.1, 
11.3, 
11.4.4 
18, 19, 20, 
21, 22, 23, 
24, 33, 54, 
55, 61 
Update in requirement of negative vector shedding samples required per sample 
type from ‘2’ to ‘3’ and associated updates in study schedule, risk/benefit and 
study procedures/assessments sections. 
Table 1, 
5.4.2, 9.3, 
9.4, 11.2.1, 
11.4.3.2 
18, 19, 20, 
21, 32, 33, 
46, 47, 54, 
60 
Updates around prophylactic regimen and actions in response to breakthrough 
transaminitis to include: Updates from ‘corticosteroid(s)’ to 
‘immunosuppressant(s)’ where required; confirmation of time points for HCV 
reactivation testing; addition of risk language and precautions in relation to the use 
of tacrolimus; adjustments to when tacrolimus should be utilised in response to 
breakthrough transaminitis (both initiation and cessation); CMV PCR testing and 
tacrolimus level testing during tacrolimus treatment. Updates with study schedule, 
risk and benefits, IMP/NIMP and study procedures/assessments sections. 
5.4.1 
30 
Updated language in relation to preclinical data and related to updates included in 
Investigator Brochure V3 Addendum 1.  
5.4.2 
34 
Update in language around ‘Insertional mutagenesis and/or tumorigenesis’ to 
provide background to patient information sheet language requested by St Jude’s, 
USA IRB. 
11.1.1 
51 
Update to language around alcohol consumption to advise on period of abstinence 
alongside the moderation already noted. 
13.2.9 
73 
Addition of thrombosis as an Important Medical Event. 
14.5.2 
76 
Clarification of wording to define per protocol population. 
14.9 
78, 79 
Addition of subgroups for analysis. 
14.10 
79 
Addition of update process for handling of missing data. 
19 
88, 89, 90 
Addition of supporting references.  
All 
All 
Appropriate spelling, grammatical, formatting and minor updates and 
addition/removal/update of words for clarity/consistency of text throughout 
protocol. 
 
Version 6.0 to Version 7.0 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number. 
4, Table 1, 
5.4.1, 7.1, 
9.4, 11.2.1, 
14.6.2 
13, 16, 18, 
19, 20, 21, 
31, 32, 36, 
39, 46, 47, 
48, 54, 78 
Updates around prophylactic regimen and actions in response to breakthrough 
transaminitis to include: Updates from ‘corticosteroid(s)’ to 
‘immunosuppressant(s)’ where required resulting from inclusion of tacrolimus in 
the prophylaxis regimen; confirmation of time points for HCV reactivation testing; 
adjustments to CMV testing and tacrolimus level testing during tacrolimus 
treatment. Updates with study schedule, risk and benefits, IMP/NIMP and study 
procedures/assessments sections. 
4, 8.2 
14, 42 
Update in Exclusion Criteria 5 from ‘Serological evidence of HIV-1 who have CD4 
counts ≤200/mm3. Patients who are HIV-positive and stable, with an adequate 
CD4 count (>200/mm3) and undetectable viral load measured twice in the 
6 months prior to enrolment, on an antiretroviral drug regimen are eligible to enrol’ 
to ‘Serological evidence of HIV-1’. 
4, 8.2 
14, 43 
Addition of exclusion criteria 16 ‘CMV IgG postive patients who are CMV PCR 
positive at screening’. 
Table 1, 
11.1.1, 
11.4.1.7, 
11.5 
18, 19, 20, 
21, 49, 50, 
57, 58, 64, 
65 
Addition of CMV testing at screening in order to asses exclusion criteria 16. 
Table 2 
22, 23, 24 
Adjustment of timepoint of ‘+1 hour after end of infusion’ to schedule of 
assessments which was omitted in error. Procedures section already reflects 
timepoint. 

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Table 2, 
11.4.3.2 
22, 23, 24, 
60 
Update to required chemistry test from ‘blood urea nitrogen’ to ‘blood urea 
nitrogen or urea’ to allow flexibility and facilitate local site practices. 
7.1, 
13.2.10 
38, 75 
Update to correct omissions when updates were made from V5.0 to V6.0 to 
change from ‘2’ to ‘3’ in requirement of negative vector shedding samples. 
7.1, 12.3.3 
39, 70 
Adjustment to interval window once the terminal dose is selected from ‘2 weeks’ to 
’48-hours’. 
12.2 
67 
Addition of text to confirm that any DLT should be reported as an SAE. 
12.3 
67 
Addition of dose of 1.5x1012 vg/kg a dose reduction level in the intermediate dose 
cohort. 
12.3.3 
69, 70 
Adjustment to dose escalation criteria review criteria, clarification on wording 
around stabilisation, peak and acceptable upper limit values of FIX activity level. 
12.3.3 
70 
Addition of guidance around consideration of potential thromboprophylaxis when a 
threshold of FIX activity level is reached, 
13.2.9 
74 
Addition of the following text ‘Other events maybe reported at the discretion of the 
Investigator.’ under the definitions for Important Medical Events. 
All 
All 
Appropriate spelling, grammatical, formatting and minor updates and 
addition/removal/update of words for clarity/consistency of text throughout 
protocol. 
 
Version 7.0 to Version 7.1 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number. 
Table 1, 
9.4, 11.2.1, 
11.4.3.2 
18, 19, 20, 
32, 48, 54, 
60 
Update to include the following text ‘Local LFT and FIX activity level monitoring 
should occur at least weekly while patient is on immunosuppression (prophylaxis 
or treatment of breakthrough) and for two weeks following cessation.’. 
12.3.1 
67 
Update to include a dose of 1.3x1012 vg/kg following a request by the Data 
Monitoring Committee. 
 
Version 7.1 to Version 8.0 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number. 
4, 8.1 
16, 45 
Correction of number stated in inclusion criteria #4 from ‘two’ to ‘three’. Item not 
updated in error in previous versions of protocol. 
4, 7.2.2, 
14.7.3 
17, 19, 43, 
88 
Addition of wording to clarify start point of analysis for secondary endpoints for 
haemostatic effectiveness. Updates across endpoints and statistics sections. 
4, 7.2.3, 
14.8.1 
17, 44, 88 
Addition of an exploratory endpoint ‘Haemostatic effectiveness - Exploration of the 
correlation between FIX levels and bleeding events over time.’ Updates across 
endpoints and statistics sections. 
4, 14.7.4 
19, 88 
Addition of wording around statistical analysis for secondary endpoint around 
vector shedding. Correction of an omission from previous versions of protocol. 
Table 1, 
Table 2, 
11.2, 11.3, 
11.4.3.2 
20, 21, 22, 
23, 24, 25, 
26, 27, 57, 
58, 59, 60, 
61, 62, 63, 
64, 68, 69 
Adjustment to the frequency and requirements for local haematology samples 
tested locally. Addition of chemistry incl. CRP and coagulation screen samples to 
be tested locally at the same time points. Further includes addition of estimated 
GFR to the local chemistry panel. Updates in study schedule and study 
procedures/assessments sections. 
Table 1, 
5.4.2, 9.4, 
11.2, 
11.4.3.2, 
Appendix 3 
20, 21, 22, 
23, 24, 36, 
49, 50, 51, 
57, 58, 59, 
60, 61, 62, 
63, 68, 69, 
106, 107, 
108 
Adjustment to the prophylaxis regimen taking tacrolimus out to Week 20. 
Associated updates include a set of ‘Management guidelines for tacrolimus 
dosing’ included as Appendix 3 to the protocol. Other applicable updates include 
adjustments to patient monitoring to be line with tacrolimus dosing. Updates in 
study schedule, risk benefits and study procedures/assessments sections. 
Table 1, 
11.2, 
11.4.2.3, 
11.4.3.2 
20, 21, 22, 
23, 24, 57, 
58, 59, 60, 
Addition of monitoring visits at Weeks 13, 15, 17, 18, 19, 21, 22, 23, 24, 25. 
Monitoring to occur twice weekly. Monitoring is focused on patient FIX levels and 
LFT’s. Also includes once weekly haematology, chemistry incl. CRP and 
coagulation screen. Hepatitis testing, tacrolimus level and CMV testing as needed 

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61, 62, 63, 
67, 68, 69 
up to week 20 only. Updates in study schedule and study 
procedures/assessments sections. 
Table 1, 
Table 2, 
11.1.1, 
11.1.2, 
11.4.3.1 
20, 21, 22, 
23, 24, 25, 
26, 27, 52, 
54, 68 
Addition of neck, waist and hip circumference measurements along with 
bioimpedance to understand patient body morphology. Updates in study schedule 
and study procedures/assessments sections. 
Table 1, 
5.4.2 
20, 21, 22, 
23, 24, 35 
Addition of ‘elastography’ as an optional test should further assessment of the liver 
be required. 
Table 1, 
11.2, 
11.4.2.3, 
11.4.3.2 
20, 21, 22, 
23, 24, 57, 
58, 59, 60, 
61, 62, 63, 
67, 68, 69 
Addition of wording to allow flexibility in location of blood sampling for frequent 
laboratory assessments (FIX levels and LFT’s) and assessments at additional 
visits (Weeks 13, 15, 17, 18, 19, 21, 22, 23, 24, 25 - FIX levels, LFTs, hepatitis 
reactivation, tacrolimus level and CMV testing) to be covered by home nursing. 
Updates in study schedule and study procedures/assessments sections. 
Table 1, 
5.4.2, 9.4, 
11.2, 
11.4.3.2, 
Appendix 2 
20, 21, 22, 
23, 24, 36, 
49, 50, 51, 
57, 58, 59, 
60, 61, 62, 
63, 68, 69, 
104, 105 
Clarification of requirement for CMV testing to reflect only patients with a positive 
test (IgG) at screening need ongoing testing through trial and not all patients. Also 
included in the instance of CMV reactivation are a set of ‘Management guidelines 
for CMV reactivation’ included as Appendix 2. Updates in study schedule, risk 
benefits and study procedures/assessments sections. 
 
Table 2, 
11.1.2 
25, 26, 27, 
56 
Adjustment of post-dosing monitoring of vital signs to note that the 10 and 12 hour 
time-points are only required for the first 2 patients at any given dose level and 
adjustment of the catch all statement to say that patients remain ‘hospitalised for 8 
hours’. Updates in study schedule, risk benefits and study 
procedures/assessments sections. 
5.4.2 
35 
Correction of an error in risk text around risk factors of use of corticosteroids. 
5.4.2 
36 
Update to provide clarification of sampling scheduled for vg sampling in line with 
rest of protocol. 
5.4.2, 
12.3.3 
38, 78 
Addition of wording around on-going risk management and oversight of 
supratherapeutic levels of FIX and thromboprophylaxis. 
9.4 
51 
Removal of wording around steroid sparing regimens to replace the 
immunosuppressant prophylactic regimen where contraindicated. 
9.4 
51 
Removal of wording around the need for regular monitoring while on 
immunosuppressant and for two weeks after. This is now covered in the updates 
to study schedule. 
11.2 
57, 58, 59, 
60, 61, 62, 
63 
Update to complete section to provide now details as sub-sections for visits as 
they occur in schedule with required assessments. Visits grouped where 
appropriate. 
11.4.1.1 
65 
Addition of text to indicate the need for a meeting with a patient advocate is not 
required for patients once the terminal dose levels has been selected. 
11.4.3.2 
68, 69 
Addition of calcium, APTT and GGT to the chemistry, coagulation screen and LFT 
panels for central lab testing. 
12.3.1 
76 
Addition of text introducing a cap to the dose that can be administered in the trial. 
Text added: ‘During the dose escalation phase of the trial, review of FIX data at a 
dose of 1.3 x1012 vg/kg, is suggestive of the impact of body weight on expression 
levels.  A dose capping at 90 kg is being introduced at this dose (1.3 x1012 vg/kg) 
to ensure that patients FIX activity levels, at steady state are ideally in the target 
range.’ 
All 
All 
Appropriate spelling, grammatical, formatting and minor updates and 
addition/removal/update of words for clarity/consistency of text throughout 
protocol. 
 
Version 8.0 to Version 8.1 
Section(s) 
Page 
Number(s) 
Summary of Change 
Title Page 
1 
Update to version number. 
Table 2, 
11.1.2 
25, 26, 27, 
56 
Reversion of changes in V8.0 to adjustment of post-dosing monitoring of vital 
signs to note that the 10 and 12 hour time-points are only required for the first 2 
patients at any given dose level and adjustment of the catch all statement to say 
that patients remain ‘hospitalised for 8 hours’. ALL patients will have vital signs 
taken to 12 hours post end of infusion and will remain hospitalised for 12 hours. 

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Updates in study schedule, risk benefits and study procedures/assessments 
sections.