Clinical trial NCT01839656
Conditions: Spinal Muscular Atrophy
Keywords: Spinal Muscular Atrophy SMA SMN SMNRx ISIS-SMNRx ISIS 396443 IONIS-SMNRx Spinraza Nusinersen
Sponsor: Biogen
Start date: 2013-05-08; Completion date: 2017-08-21
Enrollment: 21 participants; Allocation: NON_RANDOMIZED; Intervention model: PARALLEL

Brief summary:
The primary objective is to examine the clinical efficacy of multiple doses of nusinersen (ISIS 396443) administered intrathecally to participants with Infantile-Onset Spinal Muscular Atrophy (SMA). The secondary objectives are to examine the safety and tolerability of multiple doses of nusinersen administered intrathecally to participants with infantile-onset SMA and to examine the cerebral spinal fluid (CSF) and plasma Pharmacokinetics (PK) of multiple doses of nusinersen administered intrathecally to participants with infantile-onset SMA.

Detailed description:
This study was conducted and the protocol was registered by Ionis Pharmaceuticals, Inc.. In August 2016, sponsorship of the trial was transferred to Biogen.

Arms: arm 1: None arm 2: None
Interventions: intervention 1: nusinersen
Intervention descriptions: intervention 1: Administered by intrathecal (IT) injection

Protocol documents:
Prot_000.pdf: 
 
 
 
 
IONIS PHARMACEUTICALS, INC. 
 
 
ISIS 396443-CS3A 
 
A Study to Assess the Efficacy, Safety, Tolerability, and 
Pharmacokinetics of Multiple Doses of Nusinersen (ISIS 396443) 
Delivered Intrathecally to Patients with Infantile-Onset Spinal 
Muscular Atrophy 
 
Protocol Amendment 6 – 25 January 2016 
 
 
 
 
 
Sponsor: 
Ionis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 


ISIS 396443-CS3A 
Protocol 
CONFIDENTIAL 
ISIS 396443-CS3A 
Amendment 6 
25 January 2016 
A Study to Assess the Efficacy, Safety, Tolerability, and 
Pharmacokinetics of Multiple Doses of Nusinersen (ISIS 396443) 
Delivered Intrathecally to Patients with Infantile-Onset Spinal 
Muscular Atrophy 
Protocol Amendment 6-25 January 2016 
Protocol History: 
Original Protocol: 
Protocol Amendment 1: 
Protocol Amendment 2: 
Protocol Amendment 3: 
Protocol Amendment 4: 
Protocol Amendment 5: 
Sponsor: 
Ionis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA 92010 
4 March 2013 
14 August 2013 
18 January 2014 
14 March 2014 
19 May 2014 
29 May 2015 
2 
PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 6  
Protocol 
 
25 January 2016 
 
3 
 
ISIS 396443 
Ionis Protocol Number ISIS 396443-CS3A 
Protocol Amendment 6 
 
Clinical Phase:  2 
A Study to Assess the Efficacy, Safety, Tolerability, and 
Pharmacokinetics of Multiple Doses of Nusinersen (ISIS 396443) 
Delivered Intrathecally to Patients with Infantile-Onset Spinal 
Muscular Atrophy 
 
Trial Sponsor: 
Ionis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 
Phone: + 01 760 931 9200 
Fax: 
+ 01 760 603 2700 
Key Sponsor Contact: 
 
 
 
Phone: + 
 
Fax: 
+ 
 
Date: 
25 January 2016 
 
 
Confidentiality Statement 
This document contains confidential information of Ionis Pharmaceuticals, Inc. that must not be 
disclosed to anyone other than the recipient study staff and members of the institutional review 
board or authorized regulatory agencies.  This information cannot be used for any purpose other 
than the evaluation or conduct of the clinical investigation without the prior written consent of 
Ionis Pharmaceuticals, Inc. 
PPD
PPD
PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 6  
Protocol 
 
25 January 2016 
 
4 
Protocol Signature Page 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Efficacy, Safety, Tolerability, and Pharmacokinetics 
of Multiple Doses of Nusinersen (ISIS 396443) Delivered Intrathecally to 
Patients with Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
Amendment 6 
Date: 
25 January 2016 
 
I hereby acknowledge that I have read and understand the attached clinical protocol, entitled “A 
Study to Assess the Efficacy, Safety, Tolerability, and Pharmacokinetics of Multiple Doses of 
Nusinersen (ISIS 396443) Delivered Intrathecally to Patients with Infantile-Onset Spinal 
Muscular Atrophy”, dated 25 January 2016, and agree to conduct the study as described herein. 
I agree to comply with the International Conference on Harmonization Tripartite Guideline on 
Good Clinical Practice (E6). 
I agree to ensure that the confidential information contained in this document will not be used for 
any purpose other than the evaluation or conduct of the clinical investigation without the prior 
written consent of Ionis Pharmaceuticals, Inc. 
 
 
 
 
 
 
Investigator’s Signature 
 
 
 
Investigator’s Name (please print) 
 
Date (DD Month YYYY) 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 6  
Protocol 
 
25 January 2016 
 
5 
TABLE OF CONTENTS 
Page 
PROTOCOL AMENDMENT.......................................................................................................9 
PROTOCOL SYNOPSIS ............................................................................................................10 
STUDY GLOSSARY ...................................................................................................................14 
1. 
OBJECTIVES .....................................................................................................................16 
1.1 
Primary Objective .....................................................................................................16 
1.2 
Secondary Objectives................................................................................................16 
2. 
BACKGROUND AND RATIONALE ..............................................................................16 
2.1 
Spinal Muscular Atrophy ..........................................................................................16 
2.2 
Therapeutic Rationale ...............................................................................................17 
2.3 
ISIS 396443 ..............................................................................................................18 
2.3.1 
Mechanism of Action ...............................................................................18 
2.3.2 
Chemistry .................................................................................................18 
2.3.3 
Preclinical Experience ..............................................................................18 
2.3.4 
Clinical Experience ..................................................................................19 
2.4 
Rationale for Dose and Schedule of Administration ................................................20 
3. 
EXPERIMENTAL PLAN ..................................................................................................21 
3.1 
Study Design .............................................................................................................21 
3.2 
Number of Study Centers..........................................................................................21 
3.3 
Number of Subjects...................................................................................................21 
3.4 
Overall Study Duration and Follow-up.....................................................................21 
3.4.1 
Screening ..................................................................................................22 
3.4.2 
Treatment ..................................................................................................22 
3.4.3 
Post-Treatment Follow-up ........................................................................22 
3.5 
End of Study .............................................................................................................22 
3.6 
Safety Monitoring and Dose Escalation ...................................................................22 
3.7 
Dose Limiting Toxicity .............................................................................................23 
4. 
SUBJECT ENROLLMENT ...............................................................................................23 
4.1 
Screening...................................................................................................................23 
4.2 
Registration ...............................................................................................................23 
4.3 
Replacement of Subjects ...........................................................................................24 
5. 
SUBJECT ELIGIBILITY ..................................................................................................24 
5.1 
Inclusion Criteria ......................................................................................................24 
5.2 
Exclusion Criteria .....................................................................................................25 
6. 
STUDY PROCEDURES ....................................................................................................25 
6.1 
Study Schedule..........................................................................................................25 
6.2 
Study Assessments ....................................................................................................25 
6.2.1 
Collection of CSF .....................................................................................25 


ISIS 396443-CS3A 
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6.2.2 
Laboratory Analytes .................................................................................26 
6.2.3 
Neurological Examinations as Assessed by the Hammersmith Infant 
Neurological Examination (HINE) ..........................................................26 
6.2.4 
Pharmacokinetics Specimen Collection ...................................................26 
6.2.5 
Children’s Hospital of Philadelphia Infant Test for Neuromuscular 
Disease (CHOP INTEND) .......................................................................26 
6.2.6 
Hammersmith Functional Motor Scale – Expanded ................................27 
6.2.7 
Compound Muscle Action Potential ........................................................27 
6.2.8 
Growth Parameters ...................................................................................27 
6.2.9 
Ventilation Use .........................................................................................27 
6.2.10 
Phone Assessments ...................................................................................27 
6.2.11 
Safety Evaluations ....................................................................................27 
7. 
STUDY DRUG ....................................................................................................................28 
7.1 
Study Drug ................................................................................................................28 
7.1.1 
Two-Vial Configuration (ISIS 396443 and Diluent) Description ............28 
7.1.2 
Ready-To-Use Vial Configuration Description .......................................29 
7.2 
Packaging and Labeling ............................................................................................29 
7.3 
Study Drug Accountability .......................................................................................29 
8. 
TREATMENT OF SUBJECTS .........................................................................................29 
8.1 
Study Drug Administration .......................................................................................29 
8.2 
Other Protocol-Required Drugs ................................................................................31 
8.3 
Other Protocol-Required Procedures ........................................................................31 
8.4 
Treatment Precautions ..............................................................................................31 
8.5 
Safety Monitoring Rules ...........................................................................................31 
8.6 
Stopping Rules ..........................................................................................................31 
8.7 
Adjustment of Dose and/or Treatment Schedule ......................................................31 
8.8 
Discontinuation of Study Treatment .........................................................................31 
8.9 
Withdrawal of Subjects from the Study ....................................................................31 
8.10 
Concomitant Therapy and Procedures ......................................................................32 
8.10.1 
Concomitant Therapy ...............................................................................32 
8.10.2 
Concomitant Procedures ...........................................................................33 
9. 
SERIOUS AND NON-SERIOUS ADVERSE EVENT REPORTING ..........................33 
9.1 
Sponsor Review of Safety Information.....................................................................33 
9.2 
Regulatory Requirements..........................................................................................33 
9.3 
Definitions.................................................................................................................33 
9.3.1 
Adverse Event ..........................................................................................33 
9.3.2 
Adverse Reaction and Suspected Adverse Reaction ................................33 
9.3.3 
Serious Adverse Event (SAE) ..................................................................33 
9.4 
Monitoring and Recording Adverse Events ..............................................................34 
9.4.1 
Serious Adverse Events ............................................................................34 


ISIS 396443-CS3A 
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25 January 2016 
 
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9.4.2 
Non-Serious Adverse Events ....................................................................35 
9.4.3 
Evaluation of Adverse Events (Serious and Non-Serious) ......................35 
9.4.3.1 
Relationship to the Study Drug ...................................................35 
9.4.3.2 
Severity .......................................................................................35 
9.4.3.3 
Action Taken with Study Drug ...................................................36 
9.4.3.4 
Treatment Given for Adverse Event ...........................................36 
9.4.3.5 
Outcome of the Adverse Event ...................................................36 
9.5 
Procedures for Handling Special Situations .............................................................36 
9.5.1 
Abnormalities of Laboratory Tests ...........................................................36 
9.5.2 
Prescheduled or Elective Procedures or Routinely Scheduled Treatments37 
9.5.3 
Dosing Errors ...........................................................................................37 
10. STATISTICAL CONSIDERATIONS ..............................................................................37 
10.1 
Study Endpoints ........................................................................................................37 
10.1.1 
Primary Endpoint .....................................................................................37 
10.1.2 
Secondary Endpoints ................................................................................38 
10.1.3 
Exploratory Endpoints ..............................................................................38 
10.2 
Sample Size Considerations ......................................................................................38 
10.3 
Populations ................................................................................................................39 
10.4 
Interim Analysis ........................................................................................................39 
10.5 
Planned Methods of Analysis ...................................................................................39 
10.5.1 
Demographic and Baseline Characteristics ..............................................39 
10.5.2 
Efficacy Analysis .....................................................................................39 
10.5.3 
Safety and Pharmacokinetic Analysis ......................................................39 
11. INVESTIGATOR’S REGULATORY OBLIGATIONS .................................................40 
11.1 
Informed Consent......................................................................................................40 
11.2 
Ethical Conduct of the Study ....................................................................................40 
11.3 
Institutional Review Board/Research Ethics Board ..................................................41 
11.4 
Subject Confidentiality .............................................................................................41 
12. ADMINISTRATIVE AND LEGAL OBLIGATIONS ....................................................41 
12.1 
Protocol Amendments ...............................................................................................41 
12.2 
Study Termination ....................................................................................................42 
12.3 
Study Documentation and Storage ............................................................................42 
12.4 
Study Monitoring ......................................................................................................42 
12.5 
Language ...................................................................................................................43 
12.6 
Compensation for Injury ...........................................................................................43 
13. REFERENCES ....................................................................................................................44 
14. APPENDICES .....................................................................................................................46 
Appendix A 
Schedule of Procedures .....................................................................47 
Appendix B 
Laboratory Analytes .........................................................................52 
Appendix C 
Pharmacokinetic Sampling Schedule................................................54 


ISIS 396443-CS3A 
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Amendment 6  
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TABLE OF TABLES 
Page 
Table 1 
Study Drug Characteristics ................................................................................... 28 
Table 2 
Study Drug Characteristics:  Ready-To-Use Vial Configuration ......................... 29 
Table 3 
ISIS 396443 Dose Equivalents and Concentration During the ‘Loading’ 
Dosing Treatment Period ...................................................................................... 30 
Table 4 
ISIS 396443 Dose Equivalent and Concentration During the 
‘Maintenance’ Dosing Treatment Period .............................................................. 30 
Table 5 
ISIS 396443 Dose Volume to be Injected ............................................................. 30 
 
 
TABLE OF FIGURES 
Page 
Figure 1 
ASO Therapeutic Approach for Treatment of SMA ............................................. 17 
 
 
 


ISIS 396443-CS3A 
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Amendment 6
Protocol 
25 January 2016
 
9 
PROTOCOL AMENDMENT 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Efficacy, Safety, Tolerability, and Pharmacokinetics 
of Multiple Doses of Nusinersen (ISIS 396443) Delivered Intrathecally to 
Patients with Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
6 
Amendment Date: 
25 January 2016 
 
The following modifications to Protocol ISIS 396443-CS3A, have been made. 
Minor changes (not included in the list of changes below) have been made throughout the 
protocol to correct errors and/or to improve the overall clarity of the original protocol but these 
changes do not impact subject safety, exposure, or the overall study design. 
If there are more than 1 or 2 changes, insert the following table. 
The following table provides a summary list of major changes to the protocol: 
Protocol Section 
Description of Change 
Rationale 
Study Title 
Added “Efficacy” and “Nusinersen” 
Study title was updated to appropriately 
reflect the study objectives.  Nusinersen 
is an approved USAN name for 
ISIS 396443. 
Protocol Synopsis: Objectives, 
Section 1: Objectives 
Amended Study Objectives to 
reflect efficacy assessment as the 
primary study objective and safety 
and tolerability as the secondary 
study objectives. 
Study is of sufficient duration now to 
assess the efficacy of nusinersen in this 
patient population. 
Protocol Synopsis: Criteria for 
Evaluation 
Section 10.1: Study Endpoints 
Efficacy and Safety endpoints are 
listed according to the amended 
study objectives 
Efficacy endpoints related to motor 
function and event-free survival 
represent the most meaningful endpoints 
in this patient population. 
Section 10.5: Planned Methods 
of Analysis 
Inserted the methods for analysis of 
efficacy endpoints of the study 
Description of the methodology for 
analysis of the efficacy endpoints related 
to motor function and event-free survival 
was included to clarify how these data 
will be analyzed and presented. 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 6  
Protocol 
 
25 January 2016 
 
10 
PROTOCOL SYNOPSIS 
Protocol Title  
A Study to Assess the Efficacy, Safety, Tolerability, and Pharmacokinetics of Multiple 
Doses of Nusinersen (ISIS 396443) Delivered Intrathecally to Patients with Infantile-
Onset Spinal Muscular Atrophy 
Study Phase 
2 
Indication 
Spinal Muscular Atrophy (SMA) 
Objectives 
Primary Objective: 
To examine the clinical efficacy of multiple doses of ISIS 396443 administered 
intrathecally to patients with Infantile-Onset SMA. 
Secondary Objectives: 
To examine the safety and tolerability of multiple doses of ISIS 396443 administered 
intrathecally to patients with Infantile-Onset SMA. 
To examine the CSF and plasma pharmacokinetics of multiple doses of ISIS 396443 
administered intrathecally to patients with Infantile-Onset SMA. 
Number of Subjects 
Approximately 8-20 subjects may be enrolled into this multiple-dose study of 
ISIS 396443.  The number of subjects may be higher if some subjects must be replaced 
and/or if the sizes of cohorts are expanded in order to obtain further experience with 
one or both dose levels.  
Study Design and 
Methodology 
This study will test the clinical efficacy, safety, tolerability, and pharmacokinetics of 
multiple doses of ISIS 396443 administered as intrathecal (IT) injections by lumbar 
puncture (LP).  All subjects will receive ISIS 396443.  Two ‘loading’ dose levels (scaled 
by infant age to be equivalent to 2 year old doses of 6 mg or 12 mg, based on CSF 
volume) will be evaluated sequentially.  The initial dose level of 6 mg will be studied in a 
cohort of 4 subjects.  The 12 mg dose level will be studied in 4 to approximately 
16 subjects.  Following this, all subjects will receive ‘maintenance’ doses of 12 mg 
equivalent ISIS 396443 on Days 253, 379, 505, 631, 757, 883, 1009, 1135, and 1261. 
Cohort 1 (n = 4):  ‘Loading’ dosing of 6 mg equivalent ISIS 396443 on Days 1, 15, 85, 
IT injection; followed by ‘maintenance’ doses of 12 mg equivalent ISIS 396443 on 
Days 253, 379, 505, 631, 757, 883, 1009, 1135, and 1261 IT injection. 
Cohort 2 (n = 4-16):  ‘Loading’ dosing of 12 mg equivalent ISIS 396443 on Days 1, 15, 
85, IT injection; followed by ‘maintenance’ doses of 12 mg equivalent ISIS 396443 on 
Days 253, 379, 505, 631, 757, 883, 1009, 1135, and 1261 IT injection. 
After Informed consent is obtained, subjects will undergo a Screening evaluation no 
greater than 21 days prior to first dose administration at which their eligibility for the 
study will be examined.  Subjects who meet the eligibility criteria will be admitted to the 
study center on Study Day 1, undergo pre-dose evaluations, and then receive an LP 
injection of Study Drug (ISIS 396443).  Subjects will return to the study center on 
Days 15, 85, 253, 379, 505, 631, 757, 883, 1009, 1135, and 1261 for follow-up 
evaluations of clinical efficacy and safety and subsequent injections.  Following the LP 
injection on Day 1, subjects will remain at the study center for at least 24 hours post-
injection for safety monitoring.  Following LP injections on Days 15, 85, 253, 379, 505, 
631, 757, 883, 1009, 1135, and 1261, subjects will remain at the study center for at 
least 6 hours post-injection for safety monitoring.  Safety monitoring visits will occur on 
Days 16, 29, 86, 92, 169, 254, 337, 442, 568, 694, 820, 946, 1072, 1198, and 1352 
(through 13 weeks after the last dose of ISIS 396443).  In addition, the study center will 
monitor the subject’s condition through telephone contact on Study Days 8, 43, 57, 71, 
106, 127, 134, 148, 189, 197, 218, 239, 274, 295, 316, 358, 380, 400, 421, 463, 484, 
506, 526, 547, 589, 610, 632, 652, 673, 715, 736, 758, 778, 799, 841, 862, 884, 904, 
925, 967, 988, 1010, 1030, 1051, 1093, 1114, 1136, 1156, 1177, 1219, 1240, 1262, 
1282, 1303, and 1324 (i.e., approximately every 3 weeks). 
A CSF sample will be taken pre-dose on each injection day for safety laboratory 
evaluation and PK analyses. 
If a subject terminates early from the study, they will be encouraged to complete 
assessments per the Day 1352 visit. 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 6  
Protocol 
 
25 January 2016 
 
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Protocol Synopsis Continued 
Study Population and 
Main Criteria for 
Inclusion/ Exclusion 
Inclusion Criteria: 
Subjects must meet all of the following criteria at screening to be eligible: 
1. 
Signed informed consent of parent(s) or guardian(s) 
2. 
Genetic documentation of 5q SMA homozygous gene deletion or mutation 
3. 
Onset of clinical signs and symptoms consistent with SMA at ≥ 21 days and 
≤ 6 months (180 days) of age 
4. 
Males and females between ≥ 21 days and ≤ 7 months (210 days) of age at 
Screening 
5. 
At study entry, receiving adequate nutrition and hydration (with or without 
gastrostomy), in the opinion of the Site Investigator 
6. 
Body weight > 5th percentile for age using CDC guidelines  
7. 
Medical care meets and is expected to continue to meet guidelines set out in the 
Consensus Statement for Standard of Care in SMA (Wang et al. 2007), in the 
opinion of the Site Investigator 
8. 
Gestational age of 35 to 42 weeks and gestation body weight ≥ 2 kg 
9. 
Reside within approximately 9 hours ground-travel distance from a participating 
study center for the duration of the study. Residence > 2 hours ground-travel 
distance from a study center must obtain clearance from the Site Investigator and 
the study Medical Monitor 
10. Able to complete all study procedures, measurements and visits and parent or 
guardian/subject has adequately supportive psychosocial circumstances, in the 
opinion of the Site Investigator 
Exclusion Criteria: 
Subjects meeting any of the following criteria are not eligible for the study: 
1. 
Hypoxemia (O2 saturation awake < 96% or O2 saturation asleep < 96%, without 
ventilation support) 
2. 
Presence of an untreated or inadequately treated active infection requiring 
systemic antiviral or antimicrobial therapy at any time during the screening period 
3. 
History of brain or spinal cord disease that would interfere with the LP procedures, 
CSF circulation, or safety assessments 
4. 
Presence of an implanted shunt for the drainage of CSF or an implanted CNS 
catheter 
5. 
History of bacterial meningitis 
6. 
Clinically significant abnormalities in hematology or clinical chemistry parameters, 
as assessed by the Site Investigator, at Screening that would render the subject 
unsuitable for inclusion 
7. 
Treatment with another investigational drug (e.g., albuterol, riluzole, carnitine, 
creatine, sodium phenylbutyrate, salbutamol, valproate, hydroxyurea etc.), 
biological agent, or device within 90 days prior to enrollment or anytime during the 
study.  Any history of gene therapy or cell transplantation 
8. 
The subject’s parent(s) or legal guardian(s) is unable to understand the nature, 
scope, and possible consequences of the study, or does not agree to comply with 
the protocol defined schedule of assessments 
9. 
Ongoing medical condition that according to the Site Investigator would interfere 
with the conduct and assessments of the study.  Examples are medical disability 
other than SMA that would interfere with the assessment of safety or would 
compromise the ability of the subject to undergo study procedures 


ISIS 396443-CS3A 
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Amendment 6
Protocol 
25 January 2016
 
12 
Protocol Synopsis Continued 
Study Drug and 
Administration 
ISIS 396443 (20 mg/mL, 2.5 mL, diluted to specified concentration with artificial CSF 
diluent OR 2.4 mg/mL as ‘Ready-to-Use Vial’; see Section 7.1.2. for more details) will 
be administered as an IT LP injection.  The volume of the injection and thus the dose 
will be adjusted for the subject’s age, based on scaling to CSF volume.  Thus, younger 
subjects will be given a lower dose of drug, achieved by injecting a smaller volume that 
is proportional to estimated CSF volume for age, such that dose volume will be 
equivalent to 5 mL for age 2 to adult. Dosing instructions and details regarding 
administration will be provided in the Study Drug Manual. 
Criteria for Evaluation 
Efficacy Evaluations: 
 
Achievement of motor milestones as evaluated by the Hammersmith Infant 
Neurological Examination (Module 2) 
 
Event-free survival determined by the proportion of subjects who are alive and do 
not require permanent ventilatory support (defined as ≥ 16 hours ventilation/day 
continuously for at least 14 days in the absence of an acute reversible illness) 
 
Improvement in muscle strength as measured by the Children's Hospital of 
Philadelphia Infant Test for Neuromuscular Disease (CHOP INTEND) 
 
Improvement in neuromuscular electrophysiology measured by the Compound 
Muscle Action Potential (CMAP) of the ulnar and peroneal nerves 
 
Measures of respiratory status (number of respiratory events, respiratory-related 
hospitalizations, ventilator use) 
 
Growth parameters (weight for age/length, head circumference, chest 
circumference, head to chest circumference ratio, arm circumference) 
Safety/Tolerability Endpoints: 
 
Adverse events 
 
Vital signs (HR, BP, respiratory rate, pulse oximetry awake) 
 
Neurological examinations 
 
Physical examinations and weight 
 
Clinical laboratory tests  
 
CSF laboratory tests 
 
ECGs 
 
Use of concomitant medications 
Pharmacokinetic Measures: 
 
Plasma levels of ISIS 396443 
 
CSF levels of ISIS 396443 
Additional Measures: 
 
SMN2 copy number (if no Athena result existing) 
 
SMN2 gene sequencing (if no Athena result existing) 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 6  
Protocol 
 
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Protocol Synopsis Continued 
Safety Monitoring and 
Dose Escalation 
Safety data will be reviewed on an ongoing basis by the Medical Monitor and by the 
DSMB. 
At the onset of the study, a Dose-Limiting-Toxicity (DLT) is defined as an adverse event 
that in the judgment of a Site Investigator is of sufficient significance to be dose limiting,  
is possibly or definitely related to Study Drug (i.e., the adverse event is substantially 
less likely to occur in patients not treated with Study Drug), and that is not a known: 
1) sign or symptom of SMA disease, or 2) effect of the LP injection procedure, or 
3) effect of anesthesia/sedation in SMA patients, if utilized. 
The progression of the study from the initial dose level (Cohort 1) to Cohort 2 will be 
determined by the Sponsor and the DSMB and generally be based on the number of 
DLTs observed in patients treated with ISIS 396443.  In general, the occurrence of 
DLTs in 2 patients in Cohort 1 may result in the dose tested being considered as dose 
limiting.  The occurrence of 1 DLT in 4 subjects in Cohort 1 may result in the cohort 
being expanded to 6 subjects (2 additional subjects) to further assess the safety within 
that dose level.  After the last patient within Cohort 1 completes the Day 29 visit, safety 
results for the cohort will be reviewed by the Medical Monitor and the DSMB and a 
recommendation regarding further enrollment and escalation to Cohort 2 will be made. 
The Sponsor and DSMB will determine if enrollment of additional patients is required to 
confirm there is an acceptable toxicity profile prior to its designation as MTD. 
Statistical 
Considerations 
The sample size selection is based on prior experience to ensure that the safety, 
tolerability, and pharmacokinetics will be adequately assessed while minimizing 
unnecessary patient exposure rather than statistical considerations for the efficacy 
endpoints. 
Interim efficacy and safety analyses will be performed to provide content for regulatory 
submissions and to support ISIS 396443 drug development planning and business 
activities.  Details of the analyses are contained in the Statistical Analysis Plan. 
Sponsor 
Ionis Pharmaceuticals, Inc. 
 
 


ISIS 396443-CS3A 
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Amendment 6  
Protocol 
 
25 January 2016 
 
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STUDY GLOSSARY 
Abbreviation/Acronym 
Definition 
AE 
Adverse event/experience 
ALT 
Alanine aminotransferase  
aPTT 
Activated partial thromboplastin time 
ASO 
Antisense oligonucleotide 
AST 
Aspartate aminotransferase 
BUN 
Blood urea nitrogen 
CMAP 
Compound muscle action potential 
Cmax 
Maximum concentration 
CNS 
Central nervous system 
CPK 
Creatinine phosphokinase 
CRF 
Case report form 
CSF 
Cerebrospinal fluid 
CT 
Computerized tomography 
DLT 
Dose limiting toxicity 
DSMB 
Data and Safety Monitoring Board 
eCRF 
Electronic case report form 
ECG 
Electrocardiogram 
FL 
Full-length  
GCP 
Good Clinical Practice 
HFMSE 
Hammersmith Functional Motor Scale Expanded 
hnRNP 
Heterogeneous nuclear ribonucleoproteins 
ICH 
International Conference on Harmonisation  
INR 
International normalized ratio 
IRB 
Institutional Review Board 
IT 
Intrathecal 
LP 
Lumbar puncture 
MAD 
Multiple ascending-dose 
MedDRA 
Medical Dictionary for Regulatory Activities 
miRNA 
Micro ribonucleic acid (RNA) 
MOE 
2′-O-(2-methoxyethyl)  


ISIS 396443-CS3A 
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Amendment 6  
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MRI 
Magnetic resonance imaging 
mRNA 
Messenger ribonucleic acid 
MTD 
Maximum tolerated dose 
NCS 
Not clinically significant 
NOAEL 
No observed adverse effect level 
PK 
Pharmacokinetic(s) 
PT 
Prothrombin Time 
SAE 
Serious adverse event 
SMA 
Spinal muscular atrophy 
SMN 
Survival motor neuron 
snRNA 
Small nuclear ribonucleic acid 
SUSARs 
Suspected unexpected serious adverse reactions 
t½ 
Terminal elimination half-life 
TIMPSI 
Test for Infant Motor Performance Screening Items 
Tmax 
Time to maximal concentration 
WMA 
World Medical Association 
 


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1. 
OBJECTIVES 
The objectives of this study are to evaulate the clinical efficacy, safety, tolerability, and 
pharmacokinetics (PK) of multiple doses of ISIS 396443 administered intrathecally to patients 
with infantile-onset Spinal Muscular Atrophy (SMA). 
1.1 
Primary Objective 
To examine the clinical efficacy of multiple doses of ISIS 396443 administered intrathecally to 
patients with Infantile-Onset SMA. 
1.2 
Secondary Objectives 
To examine the safety and tolerability of multiple doses of ISIS 396443 administered 
intrathecally to patients with infantile-onset SMA. 
To examine the cerebral spinal fluid (CSF) and plasma PK of multiple doses of ISIS 396443 
administered intrathecally to patients with infantile-onset SMA. 
2. 
BACKGROUND AND RATIONALE 
2.1 
Spinal Muscular Atrophy 
SMA is an autosomal recessive neuromuscular disease characterized by degeneration of the 
motor neurons in the anterior horn of the spinal cord, resulting in atrophy of the voluntary 
muscles of the limbs and trunk.  With an incidence of 1:6000 to 1:10,000 live births, it is the 
most common genetic cause of infant mortality, and a major cause of childhood morbidity due to 
weakness, in the U.S.  The natural history of SMA includes four major phenotypes that are 
recognized dependent on age of onset and achieved motor abilities.  The most severe form, 
Type I SMA, has a disease onset within the first few months of life; these children are never able 
to sit or walk and usually die from respiratory failure by the age of 2 years.  Type II SMA 
patients are able to sit but never walk unaided, with symptoms presenting between 6-18 months 
of age.  Type III SMA patients are able to sit and walk but individuals with this form may 
become severely and increasingly disabled.  Type IV or adult-onset SMA patients have an age of 
onset over 18 years of age and have normal life expectancies. 
In 95% of SMA patients, a deletion in the SMN1 gene on Chromosome 5q11-q13 is found; with 
the remaining 5% attributable to small mutations in the same gene (Lefebvre et al. 1995; 
Helmken et al. 2003).  SMN1 lies in the telomeric portion of an inverted duplication of a region 
of Chromosome 5.  The centromeric half of the duplication contains a homologous gene, named 
SMN2 that differs from SMN1 by 11 nucleotides.  The open reading frames for both genes 
encode for proteins with identical amino acid sequences.  Survival motor neuron (SMN) gene 
transcripts, similar to most mammalian transcripts, undergo alternative splicing in which certain 
exons are either included or excluded from the mature protein coding transcripts 
(Keren et al. 2010).  In particular, Exon 7 of the SMN1 gene is alternatively spliced with 90 to 
95% of the mature messenger ribonucleic acid (mRNA) transcripts derived from the SMN1 gene 
containing Exon 7, and 5 to 10% of transcripts missing Exon 7.  The transcripts missing Exon 7 
(often referred to as Δ7) produce a truncated protein which is defective and unstable 
(Cho et al. 2010).  One (1) of the 11 nucleotide differences between SMN1 and SMN2, a C to T 
substitution occurs in Exon 7 of the SMN2 gene resulting in an alternative splicing pattern that 


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favors skipping of Exon 7.  The result is that as much as 90% of the transcripts produced from 
SMN2 are missing Exon 7.  The remainder, SMN2 transcripts containing Exon 7, produces a 
full-length (FL) protein product identical to the SMN1 protein, since the C to T substitution is 
silent. Humans have a variable copy number of the SMN2 gene (0-8 copies).  The number of 
SMN2 copies and the resulting amount of FL-SMN protein expressed in SMA patients (10-40% 
of normal SMN protein levels) correlates with SMA disease severity and thus SMN2 is a key 
modifier of disease phenotype (Coovert et al. 1997; Lefebvre et al. 1997; Feldkotter et al. 2002; 
Prior et al. 2004). 
2.2 
Therapeutic Rationale 
Since the number of SMN2 gene copies and resulting amount of SMN protein is correlated with 
disease onset and severity, a therapeutic approach predicted to benefit SMA patients is to 
increase the levels of full length SMN2 pre-mRNA by restoring the splicing pattern that gives 
rise to full length SMN2 mRNA.  Increasing inclusion of Exon 7 in the SMN2 transcript will 
increase FL-SMN protein levels and SMN protein activity.  A therapeutic strategy for promoting 
Exon 7 inclusion is through the use of antisense oligonucleotides (ASOs) (see Figure 1). 
 
Figure 1 
ASO Therapeutic Approach for Treatment of SMA 
 


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The known potential risks associated with ISIS 396443 are elaborated on in the Guidance to 
Investigator section of the Investigator’s Brochure.  Additional study associated risks relate to 
the lumbar puncture (LP) procedure are also elaborated on in the Guidance to Investigator 
section of the Investigator’s Brochure. 
2.3 
ISIS 396443 
2.3.1 
Mechanism of Action 
ISIS 396443 is a fully modified, 2′-O-2-methoxyethyl (MOE), ASO drug designed to bind to a 
specific sequence in the intron downstream of Exon 7 of the SMN2 transcript.  The region of the 
pre-mRNA targeted by ISIS 396443 is normally occupied by heterogeneous nuclear 
ribonucleoproteins (hnRNP) A1/2 proteins, masking the U1 small nuclear ribonucleic acid 
(snRNA) binding site at the 5′-exon-intron junction of Exon 7, and is referred to as ISS-N1.  
U1 snRNA base pairs to the sequences that define the 5′-splice site, which is thought to be one of 
the first steps that initiates splicing of an intron.  ISIS 396443 displaces the hnRNP A1/2 proteins 
from the pre-mRNA binding site, allowing U1 snRNA to bind to the exon-intron junction and 
promote assembly of the spliceosomal complex, thus promoting inclusion of Exon 7 into the 
mRNA which results in production of FL-SMN protein. 
2.3.2 
Chemistry 
Chemically, ISIS 396443 is a synthetic oligomer of 18 nucleotides (i.e., an 18-mer) that are 
connected sequentially by phosphorothioate linkages.  Each of the 17 internucleotide linkages is 
a 3′-O to 5′-O phosphorothioate diester.  The 18 sugar residues are uniformly modified with 
2′-O-(2-methoxyethyl) (MOE).  These MOE-modified nucleotides confer (1) increased affinity 
to the target mRNA (McKay et al. 1999), (2) increased resistance to exonucleases and 
endonucleases (thereby increasing stability in tissue) (Geary et al. 2003), and (3) amelioration of 
some of the high dose toxicities associated with ASO containing only the phosphorothioate 
linkages (Henry et al. 2000). 
The sequence of ISIS 396443 is written as follows: 
5′ -MeUMeCAMeCMeUMeUMeUMeCAMeUAAMeUGMeCMeUGG- 3′ 
Where A and G are 2′-O-(2-methoxyethyl)nucleosides, MeC is 5-methyl-2′-O-(2-
methoxyethyl)cytidine and MeU designates 5-methyl-2′-O-(2-methoxyethyl)uridine. 
2.3.3 
Preclinical Experience 
Detailed information concerning the preclinical studies conducted with ISIS 396443 can be 
found in the Investigator’s Brochure.  A summary is included below. 
ISIS 396443 was identified after an extensive screen of greater than 500 2′-MOE 
oligonucleotides in in vitro splicing assays, reporter gene assays and in SMA patient fibroblasts 
(Hua et al. 2007; Hua et al. 2008).  Data have shown that ISIS 396443 promotes a 
concentration-dependent increase in full-length transcripts (including Exon 7) in patient 
fibroblast cells, achieving greater than 90% full length SMN2 transcripts and forms nuclear 
structures, called gems, known to contain SMN protein.  In a mild mouse model of SMA, 
ISIS 396443 promoted inclusion of Exon 7 in the SMN2 transgene in a variety of peripheral 


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tissues when dosed systemically (Hua et al. 2008) and in central nervous system (CNS) tissue, 
including spinal cord, when injected into the lateral ventricle.  ISIS 396443 produced greater 
than 90% Exon 7 inclusion in the transgenic mice and increased SMN protein production in 
motor neurons, resulting in the appearance of gems in motor neurons.  These studies were 
extended to a more severe mouse model of SMA (SMA Δ7) (Le et al. 2005), where the CNS 
delivery of drug produced a dose-dependent effect on SMN2 Exon 7 inclusion, SMN protein 
production, and survival.  These mice treated with ISIS 396443 demonstrated improved weight 
gain, improvements in muscle morphology, muscle strength, and motor coordination and 
improved morphology of the motor neuron junctions (Passini et al. 2011).  Further, ISIS 396443 
was shown to distribute widely in the CNS following intrathecal (IT) administration in monkey 
(Passini et al. 2011). 
The pharmacokinetics and toxicity of ISIS 396443 were assessed following:  1) single intrathecal 
(IT) lumbar bolus injections (1 to 7 mg) in adult monkeys 2) following 14 weeks (with a 4-week 
interim sacrifice) of repeated IT lumbar bolus injections (0.3 to 3 mg/wk or every other week) in 
juvenile monkeys and 3) following 1-year of repeated IT lumbar bolus injections in juvenile 
monkeys.  Detailed results from these preclinical studies conducted with ISIS 396443 can be 
found in the Investigator’s Brochure. 
2.3.4 
Clinical Experience 
Detailed information concerning the clinical studies conducted with ISIS 396443 can be found in 
the Investigator’s Brochure.  A summary is included below. 
ISIS 396443 has been evaluated in a completed open-label, single ascending-dose (SAD) Phase 1 
study designed to assess the safety, tolerability and pharmacokinetics of ISIS 396443 in patients 
with SMA (ISIS 396443-CS1).  A single-dose of ISIS 396443 was administered by IT injection 
to SMA patients aged 2 to 14 years of age.  Four (4) dose levels (1, 3, 6, and 9 mg) were 
evaluated sequentially.  Each dose level was studied in a cohort of 6 or 10 patients, where all 
patients received drug.  In this study all subjects completed dosing and the follow-up visits per 
protocol.  Overall, ISIS 396443 was well-tolerated and no safety concerns were identified up to 
the 9.0 mg dose level, given as a single IT injection.  No serious adverse events (SAEs) or dose-
limiting toxicities (DLTs) were reported in ISIS 396443-CS1.  Adverse events (AEs) reported 
were mild or moderate in severity and there was no relationship with ISIS 396443 dose level. In 
addition, no ISIS 396443 related adverse changes in neurological exams were reported, despite 
intensive monitoring during the immediate post-dosing period.  CSF and plasma drug 
concentrations observed were generally consistent with predictions made from nonclinical 
monkey studies. 
ISIS 396443 is also being evaluated in seven ongoing studies: ISIS 396443-CS2, 
ISIS 396443-CS10, ISIS 396443-CS12, ISIS 396443-CS3B (ENDEAR), ISIS 396443-CS4 
(CHERISH), 232SM201 (NURTURE) and 232SM202 (EMBRACE). 
ISIS 396443-CS10 is an open-label, single dose, re-dosing study for SMA patients who 
previously participated in Cohorts 2, 3 and 4 in ISIS 396443-CS1. 
ISIS 396443-CS2 is an open-label, multiple ascending-dose (MAD) Phase 1/2a study designed to 
assess the safety, tolerability and pharmacokinetics of ISIS 396443 in patients with SMA.  


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Multiple doses of ISIS 396443, ranging from 3 mg to 12 mg, are being administered by 
intrathecal injection to SMA patients aged 2 to 15 years of age. 
ISIS 396443-CS12 is an open-label, multiple-dose study to assess the safety and tolerability of a 
12 mg dose level of ISIS 396443 administered intrathecally in patients with spinal muscular 
atrophy who previously participated in ISIS 396443-CS2 or ISIS 396443-CS10. 
ISIS 396443-CS3B (ENDEAR) is a randomized, double-blind, sham-procedure controlled study 
designed to assess clinical efficacy and safety of ISIS 396443 in patients with infantile-onset 
SMA.  A 12 mg dose equivalent scaled by CSF volume is being evaluated in symptomatic SMA 
infants ≤ 7 months at the time of consent of age utilizing a loading regimen of 4 intrathecal doses 
on Days 1, 15, 29 and 64, followed by a q4 months maintenance period. 
ISIS 396443-CS4 (CHERISH) is a randomized, double-blind, sham-procedure controlled study 
designed to assess clinical efficacy and safety of ISIS 396443 in patients with later-onset SMA 
aged 2-12 years.  A 12 mg dose is being evaluated utilizing a loading regimen of 3 intrathecal 
doses on Days 1, 29 and 85, followed by a q6 months maintenance period. 
232SM201 (NURTURE) is an open-label study to assess the efficacy, safety, tolerability and 
pharmacokinetics of multiple doses of ISIS 396443 delivered intrathecally to subjects with 
genetically diagnosed and pre-symptomatic SMA. 
232SM202 (EMBRACE) is a randomized, double-blind, sham-procedure controlled study to 
assess safety and tolerability and to explore efficacy of ISIS 396443 administered intrathecally in 
subjects with SMA who are not eligible to participate in the clinical studies ISIS 396443-CS3B 
or ISIS 396443-CS4. 
2.4 
Rationale for Dose and Schedule of Administration 
The proposed study will test the clinical efficacy, safety, tolerability, and PK of multiple doses of 
ISIS 396443 administered as IT injections.  Two (2) ‘loading’ dose levels (scaled by infant age 
to be equivalent to 2 year old doses of 6 mg or 12 mg, based on CSF volume) will be evaluated 
sequentially, given during a‘loading’ dosing period on Study Days 1, 15, and 85.  Following that, 
all subjects will be given ‘maintenance’ doses of 12 mg equivalent ISIS 396443 on 
Study Days 253, 379, 505, 631, 757, 883, 1009, 1135, and 1261. 
ISIS 396443 dose levels and dose interval for ISIS 396443-CS3A were selected based on 
preclinical toxicology and pharmacokinetic observations from monkey studies utilizing single-
dose and repeat dosing (for 14 weeks) IT administration, consideration of the target tissue 
concentration anticipated for drug pharmacology, and safety data in the completed and ongoing 
clinical studies of ISIS 396443 to date.  Based upon pharmacology and pharmacokinetic results 
in SMA transgenic mice, we estimate that the target tissue concentration to produce 50 to 90% 
SMN2 Exon 7 inclusion is between 1 and 10 µg/g spinal cord tissue.  Nonclinical studies in 
juvenile monkeys receiving IT doses of ISIS 396443 showed a resulting gradient of distribution 
of ISIS 396443 along the spinal cord, with mean lumbar spinal cord concentrations ranging from 
1.6-2.3 fold and 2.0-3.5 fold higher than thoracic and cervical spinal cord levels, respectively.  
The lowest dose selected for this multiple-dose clinical study (6 mg ISIS 396443) is predicted to 
achieve spinal cord tissue concentrations at the low end of this range (approximately 6 µg/g 
lumbar and 2 μg/g cervical concentrations), while the highest dose (12 mg ISIS 396443) is 


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predicted to achieve levels at the high end of this range (approximately 10 µg/g lumbar and 
3 μg/g cervical spinal cord tissue concentrations), following the first dose. 
The loading dose interval was selected based on the nonclinical pharmacokinetic and 
pharmacology data as the dose interval to achieve and maintain ISIS 396443 spinal cord tissue 
levels that are predicted to be within the upper end of the pharmacologically active range by 
Day 85 (predicted to be approximately 30 µg/g lumbar and 10 µg/g cervical tissue 
concentration), while at the same time considering subject safety and convenience for repeated 
LP intrathecal injections.  The maintenance dose interval (once every 4 months) was selected 
based on the estimated spinal tissue and CSF drug half-life (4-6 months) and was selected to 
maintain spinal cord tissue levels of ISIS 396443 at a steady-state level within the estimated 
pharmacologically active range. 
Additional details on dose scaling and expected CSF and tissue concentrations are summarized 
in the Investigator’s Brochure. 
3. 
EXPERIMENTAL PLAN 
3.1 
Study Design 
This study will test the clinical efficacy, safety, tolerability, and PK of multiple doses of 
ISIS 396443 administered as IT injections by LP.  Two (2) ‘loading’ dose levels (scaled by 
infant age to be equivalent to 2 year old doses of 6 mg or 12 mg, based on CSF volume) will be 
evaluated sequentially.  The initial dose level of 6 mg will be studied in a cohort of 4 subjects.  
The 12 mg dose level will be studied in 4 to approximately 16 subjects.  Following this, all 
subjects will receive ‘maintenance’ doses of 12 mg equivalent ISIS 396443 on Days 253, 379, 
505, 631, 757, 883, 1009, 1135, and 1261.  
Cohort 1 (n = 4):  ‘Loading’ dosing of 6 mg equivalent ISIS 396443 on Days 1, 15, 85, 
IT injection; ‘maintenance’ doses of 12 mg equivalent on Days 253, 379, 505, 631, 757, 883, 
1009, 1135, and 1261, IT injection. 
Cohort 2 (n = 4-16):  ‘Loading’ dosing of 12 mg equivalent ISIS 396443 on Days 1, 15, 85, 
IT injection, ‘maintenance’ doses of 12 mg equivalent on Days 253, 379, 505, 631, 757, 883, 
1009, 1135, 1261, IT injection. 
3.2 
Number of Study Centers 
This study will be conducted at multiple centers in the United States and Canada. 
3.3 
Number of Subjects 
Approximately 8-20 subjects will be enrolled into this multiple-dose study of ISIS 396443.  The 
number of subjects may be higher if some subjects must be replaced and/or if the sizes of some 
cohorts are expanded in order to obtain further experience with some dose levels. 
3.4 
Overall Study Duration and Follow-up 
The Study will consist of screening, treatment period including ‘loading’ dosing and 
‘maintenance’ dosing, and post-treatment follow-up period.  The total duration of participation in 
the study is approximately 3.7 years.  Please refer to the Schedule of Procedures in Appendix A. 


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3.4.1 
Screening 
After informed consent/assent is obtained, subjects will undergo a Screening evaluation no 
greater than 21 days prior to first dose administration at which their eligibility for the study will 
be examined. 
3.4.2 
Treatment 
Subjects who meet the eligibility criteria will be admitted to the study center on Study Day 1, 
undergo pre-dose evaluations, and then receive an LP injection of Study Drug (ISIS 396443).  
The subject will remain at the study center for at least 24 hours post-injection for safety 
monitoring and laboratory specimen collection.  Subjects will return to the tudy center on 
Days 15, 85, 253, 379, 505, 631, 757, 883, 1009, 1135, and 1261 for follow-up evaluations and 
subsequent injections.  Following LP injections on Study Days 15, 85, 253, 379, 505, 631, 757, 
883, 1009, 1135, and 1261 subjects will not stay overnight in the hospital but will be monitored 
for at least 6 hours post-injection before leaving the study facility.  A CSF sample will be taken 
pre-dose on each injection day for safety and Pharmacokinetic (PK) analyses.  During the 
treatment period, the study center will monitor the subject’s condition through safety monitoring 
visits on Study Days 16, 29, 86, 92, 169, 254, 337, 442, 568, 694, 820, 946, 1072, and 1198, and 
by telephone contact on Study Days 8, 43, 57, 71, 106, 127, 134, 148, 189, 197, 218, 239, 274, 
295, 316, 358, 380, 400, 421, 463, 484, 506, 526, 547, 589, 610, 632, 652, 673, 715, 736, 758, 
778, 799, 841, 862, 884, 904, 925, 967, 988, 1010, 1030, 1051, 1093, 1114, 1136, 1156, 1177, 
1219, 1240, 1262, 1282, 1303, and 1324 (i.e., approximately every 3 weeks). 
3.4.3 
Post-Treatment Follow-up 
During the post-treatment follow-up period, the study center will monitor the subject’s condition 
through telephone contact on Study Days 1282, 1303, and 1324 (i.e., every 3 weeks through 
13 weeks after the last dose of ISIS 396443).  A safety monitoring visit will occur on Day 1352 
(through 13 weeks after the last dose of ISIS 396443). 
If a subject terminates early from the study, they will be encouraged to complete all assessments 
per the Day 1352 visit. 
3.5 
End of Study 
The end of study is last subject, last visit (either in-person visit or telephone contact). 
3.6 
Safety Monitoring and Dose Escalation 
Safety data will be reviewed on an ongoing basis by the Sponsor and the Ionis Medical Monitor. 
Safety data will also be reviewed on an ongoing basis by an independent Data and Safety 
Monitoring Board (DSMB). 
The progression of the study from the initial dose level (Cohort 1) to Cohort 2 will be determined 
by the Sponsor and the DSMB and generally be based on the number of DLTs observed in 
patients treated with ISIS 396443.  In general, the occurrence of DLTs in 2 patients in Cohort 1 
may result in the dose tested being considered as dose limiting.  The occurrence of 1 DLT in 
4 subjects in Cohort 1 may result in the cohort being expanded to 6 subjects (2 additional 
subjects) to further assess the safety within that dose level. 


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After the last patient within Cohort 1 completes the Day 29 visit, safety results for the cohort will 
be reviewed by the Sponsor and the DSMB and a recommendation regarding further enrollment 
and escalation to Cohort 2 will be made.  The Sponsor and DSMB will determine if enrollment 
of additional patients is required to confirm there is an acceptable toxicity profile prior to its 
designation as MTD. 
3.7 
Dose Limiting Toxicity 
At the onset of the study, a DLT is defined as an adverse event that in the judgment of a 
Site Investigator is of sufficient significance to be dose limiting, is possibly or definitely related 
to Study Drug (i.e., the AE is substantially less likely to occur in subjects not treated with 
Study Drug), and that is not a known:  1) sign or symptom of SMA disease, or 2) effect of the LP 
injection procedure. 
If a suspected DLT occurs during injection of Study Drug, treatment of the subject with Study 
Drug should be stopped (i.e., the injection should be immediately discontinued).  The Site 
Investigator should contact the Ionis Medical Monitor as soon as possible following any dosing 
discontinuations to discuss the case.  The DSMB will also be informed and will determine if any 
relevant findings have been observed with other subjects in the study. 
Subjects that experience a DLT will discontinue study treatment but will be monitored by 
follow-up safety visits. 
4. 
SUBJECT ENROLLMENT 
4.1 
Screening 
Before subjects may be enrolled into the study, the Sponsor requires a copy of the Study Center’s 
written Institutional Review Board (IRB) or Research Ethics Board (REB) approval of the 
protocol, informed consent form, informed assent form (if applicable) and all other subject 
information and/or recruitment material. 
Before a subject’s participation in the trial, the Investigator is responsible for obtaining written 
informed consent from the parent(s) or legal guardian(s).  At the time of consent, the subject will 
be considered enrolled into the study and will be assigned a unique screening number before any 
study procedures, including screening procedures, are performed.  At the time of registration, 
subjects will be assigned a unique subject identification number.  This number will be used to 
identify the subject throughout the trial and must be used on all study documentation related to 
that subject.  The screening number and subject identification number must remain constant 
throughout the entire trial.  In the event the subject is re-consented and re-screened the subject 
must be given a new screening number.  Screening numbers and subject identification numbers, 
once assigned, will not be re-used. 
4.2 
Registration 
Subjects will be registered after all screening assessments have been completed and after the 
Investigator and the Ionis Medical Monitor have verified that they are eligible per criteria in 
Sections 5.1 and 5.2. 


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No subject may begin treatment prior to registration and assignment of a unique subject 
identification number. 
4.3 
Replacement of Subjects 
If a subject withdraws from the trial prior to the administration of Study Drug, the subject will be 
replaced.  If a subject withdraws for reasons other than safety after receiving any amount of 
Study Drug and prior to the Day 85 visit, the Site Investigator and the Ionis Medical Monitor will 
discuss the reason(s) for the discontinuation and the appropriateness of enrolling a replacement 
subject.  If a subject withdraws for safety reasons that are potentially drug related after receiving 
any amount of Study Drug, the DSMB will review the case and provide recommendations 
regarding replacement of the subject. 
5. 
SUBJECT ELIGIBILITY 
5.1 
Inclusion Criteria 
To be eligible to participate in this study candidates must meet the following eligibility criteria at 
screening. 
1. Signed informed consent of parent(s) or guardian(s) 
2. Genetic documentation of 5q SMA homozygous gene deletion or mutation 
3. Onset of clinical signs and symptoms consistent with SMA at ≥ 21 days and ≤ 6 months 
(180 days) of age 
4. Males and females between ≥ 21 days and ≤ 7 months (210 days) of age at Screening 
5. At study entry, receiving adequate nutrition and hydration (with or without gastrostomy), 
in the opinion of the Site Investigator 
6. Body weight > 5th percentile for age using CDC guidelines 
7. Medical care meets and is expected to continue to meet guidelines set out in the 
Consensus Statement for Standard of Care in SMA (Wang et al. 2007), in the opinion of 
the Site Investigator 
8. Gestational age of 35 to 42 weeks and gestation body weight ≥ 2 kg 
9. Reside within approximately 9 hours ground-travel distance from a participating study 
center for the duration of the study.  Residence > 2 hours ground-travel distance from a 
study center must obtain clearance from the Site Investigator and the study Medical 
Monitor 
10. Able to complete all study procedures, measurements and visits and parent or 
guardian/subject has adequately supportive psychosocial circumstances, in the opinion of 
the Site Investigator 


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5.2 
Exclusion Criteria 
Subjects meeting any of the following criteria are not eligible for the study: 
1. Hypoxemia (O2 saturation awake < 96% or O2 saturation asleep < 96%, without 
ventilation support) 
2. Presence of an untreated or inadequately treated active infection requiring systemic 
antiviral or antimicrobial therapy at any time during the screening period 
3. History of brain or spinal cord disease that would interfere with the LP procedures, CSF 
circulation, or safety assessments 
4. Presence of an implanted shunt for the drainage of CSF or an implanted CNS catheter 
5. History of bacterial meningitis 
6. Clinically significant abnormalities in hematology or clinical chemistry parameters, as 
assessed by the Site Investigator, at screening that would render the subject unsuitable for 
inclusion 
7. Treatment with another investigational drug (e.g., albuterol, riluzole, carnitine, creatine, 
sodium phenylbutyrate, salbutamol, valproate, hydroxyurea, etc.), biological agent, or 
device within 90 days prior to enrollment or anytime during the study.  Any history of 
gene therapy or cell transplantation 
8. The subject’s parent(s) or legal guardian(s) is unable to understand the nature, scope, and 
possible consequences of the study, or does not agree to comply with the protocol defined 
schedule of assessments 
9. Ongoing medical condition that according to the Site Investigator would interfere with 
the conduct and assessments of the study.  Examples are medical disability other than 
SMA that would interfere with the assessment of safety or would compromise the ability 
of the subject to undergo study procedures 
6. 
STUDY PROCEDURES 
6.1 
Study Schedule 
All required study procedures are outlined in Appendices A, B and C. 
6.2 
Study Assessments 
6.2.1 
Collection of CSF 
Subjects will have CSF collected pre-dose during the LP procedure on Days 1, 15, 85, 253, 379, 
505, 631, 757, 883, 1009, 1135, and 1261 for safety and PK analyses.  Approximately 5 mL of 
CSF will be collected by lumbar puncture, using a ‘spinal anesthesia’ needle and a standard LP 
collection kit.  A 22G anesthesia needle is recommended.  Depending on institutional guidelines, 
anesthesia or sedation may be used for the procedure.  Spinal ultrasound may be used for the LP 
procedure, if deemed necessary, but is not required.  CSF will be used for standard laboratory 


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measurement of cells, glucose, and protein, and ISIS 396443 pharmacokinetic analyses.  Extra 
CSF may be stored for investigation of possible biomarkers of SMA disease or the 
pharmacodynamic effects of ISIS 396443 (e.g., CSF mRNA, CSF protein, CSF miRNA panel) or 
for profiling of drug binding proteins, bioanalytical method validation purposes, stability 
assessments, metabolite assessments, immunogenicity assessments (including assay development 
and validation purposes), or to assess other actions of ISIS 396443 with CSF constituents. 
6.2.2 
Laboratory Analytes 
Laboratory measurements of serum chemistry, hematology, urinalysis, coagulation parameters, 
and plasma antibodies to ISIS 396443 will be performed at the times shown in the Schedule of 
Procedures (Appendix A).  The analytes to be measured are shown in Appendix B. 
6.2.3 
Neurological Examinations as Assessed by the Hammersmith Infant Neurological 
Examination (HINE) 
Neurological examinations will be performed using the Hammersmith Infant Neurological 
Examination (HINE).  These examinations will include assessment of mental status, level of 
consciousness, sensory function, motor function, cranial nerve function, and reflexes.  Motor 
milestones will be accessed using Section 2 of the HINE, which is comprised of 8 independent 
milestone categories:  voluntary grasp, ability to kick in supine position, head control, rolling, 
sitting, crawling, standing, and walking.  HINE will be performed at the times/dates according to 
the schedule as shown in Appendix A (Schedule of Procedures). 
6.2.4 
Pharmacokinetics Specimen Collection 
Plasma and CSF specimens will be collected as shown in Appendix A (Schedule of Procedures) 
and Appendix C (Pharmacokinetic Sampling Schedule).  The following ISIS 396443 plasma PK 
parameters (though not necessarily limited to) will be derived when appropriate from the 
individual subject concentration vs. time profiles using noncompartmental-based methods and 
based on actual sampling times: 
 The maximal observed plasma drug concentration (Cmax) 
 The time to reach Cmax in plasma (Tmax) 
 The area under the plasma concentrations time curve from the time of the IT dose to the 
last collected sample 
 The apparent terminal elimination half-life (t½), if possible 
6.2.5 
Children’s Hospital of Philadelphia Infant Test for Neuromuscular Disease 
(CHOP INTEND) 
Subjects will be evaluated using the CHOP INTEND test according to the schedule as shown in 
Appendix A (Schedule of Procedures).  The CHOP INTEND test was specifically designed to 
evaluate the motor skills of infants with significant motor weakness, including infants with SMA 
(Glanzman et al. 2010).  The CHOP INTEND test captures neck, trunk, proximal and distal limb 
strength in 14 elicited and 2 observational items.  The CHOP INTEND has been established as a 
safe and reliable infant motor measure in infantile-onset SMA. 


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6.2.6 
Hammersmith Functional Motor Scale – Expanded  
All subjects who have maintained a CHOP INTEND total score of ≥ 50 for 2 consecutive study 
visits will be evaluated using the Hammersmith Functional Motor Scale Expanded (HFMSE) at 
the times shown in Appendix A (Schedule of Procedures).  This evaluation will be performed in 
addition to the CHOP INTEND.  The HFMSE is a reliable and validated tool to assess motor 
function in children with SMA.  The scale was originally developed with 20 scored activities and 
was devised for use in children with SMA type 2 and type 3 with limited ambulation to give 
objective information on motor ability and clinical progression (Main et al. 2003).  The expanded 
scale includes an additional module of 13 items developed to allow for evaluation of ambulatory 
SMA patients (O’Hagen et al. 2007).  The HFMSE has been shown to be highly correlated with 
other clinical assessments and shows good test-retest reliability.  The HFMSE is easy to use and 
quickly administered. 
6.2.7 
Compound Muscle Action Potential  
Measurements of Compound Muscle Action Potential (CMAP) will be assessed (ulnar and 
peroneal nerves) according to the schedule as shown in Appendix A (Schedule of Procedures).  
CMAP is an electrophysiological technique that can be used to determine the approximate 
number of motor neurons in a muscle or group of muscles.  CMAP is a well validated method for 
tracking disease progression in neuromuscular disorders such as spinal muscular atrophy (SMA) 
(Swoboda et al. 2005; Lewalt et al. 2010) and amyotrophic lateral sclerosis (Shefner et al. 2011) 
and has been proposed as a potential biomarker of a therapeutic effect in SMA. 
6.2.8 
Growth Parameters 
Subjects will be assessed by growth measurements according to the schedule as shown in 
Appendix A (Schedule of Procedures).  Growth parameters of body length, head circumference, 
chest circumference, and arm circumference will be measured.  Additional parameters of weight 
for age, weight for length, and head to chest circumference ratio will be calculated. 
6.2.9 
Ventilation Use 
The subject’s ventilator use will be recorded at the times/dates according to the schedule as 
shown in Appendix A (Schedule of Procedures). 
6.2.10 
Phone Assessments 
Monitoring telephone calls will occur on Study Days 8, 43, 57, 71, 106, 127, 134, 148, 189, 197, 
218, 239, 274, 295, 316, 358, 380, 400, 421, 463, 484, 506, 526, 547, 589, 610, 632, 652, 673, 
715, 736, 758, 778, 799, 841, 862, 884, 904, 925, 967, 988, 1010, 1030, 1051, 1093, 1114, 1136, 
1156, 1177, 1219, 1240, 1262, 1282, 1303, and 1324 (i.e., approximately every 3 weeks between 
study visits).  Changes in concomitant medications and adverse events will be recorded.  In 
addition, information on the subject’s ventilator use and SMA disease status will be collected. 
6.2.11 
Safety Evaluations 
Safety will be evaluated by assessment of AEs including SAEs as described in Section 9.  
Additional safety evaluations include the following parameters: 
 Neurological examinations (HINE) 


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 Vital signs 
 Physical examinations and weight 
 Clinical laboratory tests (serum chemistry, hematology, urinalysis) 
 CSF laboratory tests (cell count, protein, glucose) 
 Electrocardiogram (ECG) 
 Use of concomitant medications 
7. 
STUDY DRUG 
7.1 
Study Drug 
There are 2 types of Study Drug that may be provided for this study: 
a. Study Drug at a concentration of 20 mg/mL which is diluted prior to use; this is described 
as the ‘Two-Vial’ configuration.  See Section 7.1.1 for more details. 
b. Study Drug at a concentration of 2.4 mg/mL; this is described as ‘Ready-to-Use Vial’ 
configuration.  See Section 7.1.2 for more details. 
7.1.1 
Two-Vial Configuration (ISIS 396443 and Diluent) Description 
ISIS 396443 active drug product and artificial CSF diluent are manufactured by Ionis 
Pharmaceuticals, Inc., Carlsbad, CA, USA.  The active drug product is supplied at a 
concentration of 20 mg/mL as a 2.5 mL fill volume in a 5 mL clear glass vial.  The diluent 
(artificial CSF) is supplied as a 20 mL fill volume in a 30 mL clear glass vial.  These 
configurations allow for various clinical doses by using different dilution procedures between the 
active drug product and diluent vials.  Study Drug characteristics are described in Table 1.  More 
details are provided in the Investigators Brochure. 
Table 1 
Study Drug Characteristics 
Study Drug 
ISIS 396443 
Diluent 
(artificial CSF) 
Strength 
20 mg/mL 
NA 
Volume/vial 
2.5 mL solution per vial 
20 mL solution per vial 
Route of Administration 
IT injection 
IT injection 
 
ISIS 396443 and the diluent (artificial CSF) must be stored securely at 2° C to 8° C.  
ISIS 396443 must be protected from light. 


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7.1.2 
Ready-To-Use Vial Configuration Description 
In the ‘Ready-to-use’ vial configuration, the Study Drug is supplied in 6 mL clear glass vials at a 
concentration of 2.4 mg/mL.  The Study Drug and its storage and preparation instructions will be 
provided by the Sponsor or designee.  Study Drug characteristics are listed under Table 2. 
Table 2 
Study Drug Characteristics:  Ready-To-Use Vial Configuration 
Study Drug 
ISIS 396443 Drug Product 
Strength 
2.4 mg/mL  
Volume/vial 
5.0 mL solution per vial 
Route of Administration 
IT injection 
 
ISIS 396443 must be stored securely at 2° to 8° C and protected from light. 
7.2 
Packaging and Labeling 
Ionis Pharmaceuticals, Inc. will provide the Site Pharmacists with packaged Study Drug labeled 
in accordance with regulatory requirements. 
7.3 
Study Drug Accountability 
The study staff is required to document receipt, dispensing and return of Study Drug supplies 
provided by the Sponsor.  Drug accountability documentation and all used and unused Study 
Drug vials (ISIS 396443 and diluent) must be returned to Ionis Pharmaceuticals, Inc. or 
designee. 
8. 
TREATMENT OF SUBJECTS 
8.1 
Study Drug Administration 
Each subject will receive a single IT bolus (1-3 minute) LP injection of Study Drug using a 
‘spinal anesthesia’ needle and 5 mL syringe.  A 22G spinal anesthesia needle is recommended.  
The target site for needle insertion is the L3/L4 space, but may be 1 segment above or 
1-2 segments below this level, if needed.  Prior to the injection 5-6 mL of CSF fluid is to be 
collected for analyses.  Depending on institutional guidelines, anesthesia or sedation may be used 
for the LP procedure, following institutional procedures.  Spinal ultrasound may be used for the 
LP procedure, if deemed necessary, but is not required. 
Table 3 outlines the dose equivalent and ISIS 396443 concentration for ISIS 396443 dose 
cohorts to be delivered during the ‘loading’ dosing treatment period (i.e., dosing on Days 1, 15 
and 85).  Study Drug dosing during the ’loading’ dosing treatment period will be done using the 
two-vial configuration. 
Table 4 outlines the dose equivalent and ISIS 396443 concentration for ISIS 396443 to be 
delivered during the ‘maintenance’ dosing treatment period (i.e., dosing on Days 253, 379, 505, 
631, 757, 883, 1009, 1135, and 1261).  Study Drug dosing during the ‘maintenance’ dosing 


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treatment period will be done using the two-vial configuration and will transition to the  
ready-to-use vial configuration once the current protocol amendment has been approved. 
Table 3 
ISIS 396443 Dose Equivalents and Concentration During the ‘Loading’ 
Dosing Treatment Period 
Cohort 
Dose Equivalent 
(mg) 
Concentration 
(mg/mL) 
Injection Volume 
(mL) 
1 
6 
1.2 
See Table 5 
2 
12 
2.4 
See Table 5 
 
Table 4 
ISIS 396443 Dose Equivalent and Concentration During the ‘Maintenance’ 
Dosing Treatment Period 
Cohorts 
Dose Equivalent 
(mg) 
Concentration 
(mg/mL) 
Injection Volume 
(mL) 
1 and 2 
12 
2.4 
See Table 5 
 
The volume of the injection will be adjusted based on the subject’s age.  Subjects who are older 
than 24 months (730 Days) on the day of dosing will receive the full 12 mg dose of the 
Study Drug (5 mL).  For subjects younger than 24 months of age, the volume of the injection 
will be adjusted based on the subject’s age on the day of dosing per Table 5, such that each 
subject will receive a 12 mg equivalent dose based on CSF volume scaling. 
Table 5 
ISIS 396443 Dose Volume to be Injected 
Age 
Estimated 
CSF Volume* 
Injection Volume 
(mL) 
0-3 months  
(0-90 days) 
120 mL 
4 mL 
3-6 months  
(91-182 days) 
130 mL 
4.3 mL 
6-12 months  
(183 – 365 days 
135 mL 
4.5 mL 
12-24 months  
(366 – 730 days) 
140 mL 
4.7 mL 
> 24 months 
(> 730 days) 
150 mL 
5.0 mL 
* 
Matsuzawa et al. 2001 
 
Please refer to the Study Drug Manual provided by the Sponsor for detailed instructions for 
Study Drug preparation and administration. 


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8.2 
Other Protocol-Required Drugs 
There are no other protocol required drugs, but depending on institutional guidelines anesthesia 
or sedation may be used for the LP procedure, following institutional procedures.  
8.3 
Other Protocol-Required Procedures 
There are no other protocol-required treatment procedures. 
8.4 
Treatment Precautions 
Subjects will be encouraged to lie flat for 1 hour following dosing, if possible. 
8.5 
Safety Monitoring Rules 
Please refer to the Guidance to Investigator section of the Investigator Brochure. 
8.6 
Stopping Rules 
Please refer to Section 8.8.  There are no additional specific stopping rules for this study but the 
Investigator should discuss significant concerns relating to individual subjects with the 
Ionis Medical Monitor to ensure that it is appropriate for the subject to continue Study Drug. 
8.7 
Adjustment of Dose and/or Treatment Schedule 
No adjustment of dose is permitted.  In the event of a concurrent illness that would prevent the 
dosing procedure from being performed safely, an adjustment in the dose schedule may be 
permitted, but must be approved by the Sponsor Medical Monitor.  In this case, the scheduled 
Day 15 dosing visit may be delayed until up to Day 70 and the scheduled Day 85 dosing visit 
may be delayed up to Day 120.  The scheduled dosing visits during the ‘maintenance’ period 
(i.e., Days 253, 379, 505, 631, 757, 883, 1009, 1135, and 1261) may be delayed by up to 
8 weeks. 
8.8 
Discontinuation of Study Treatment 
A subject must permanently discontinue study treatment for any of the following: 
 The subject withdraws consent 
 The subject experiences an adverse event that necessitates permanent discontinuation of 
study treatment 
 The subject experiences a DLT as defined in Section 3.7 
The reason for discontinuation of study treatment must be recorded in the Case Report Form 
(CRF) and source documentation.  
Subjects that discontinue treatment will enter the post-treatment follow-up period unless consent 
is withdrawn (see Appendix A). 
8.9 
Withdrawal of Subjects from the Study 
Subjects must be withdrawn from the study for any of the following: 
 Withdrawal of consent 


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 The subject is unwilling or unable to comply with the protocol 
Other reasons for withdrawal of subjects from the study might include: 
 At the discretion of the Investigator for medical reasons 
 At the discretion of the Investigator or Sponsor for noncompliance 
 Significant protocol deviation 
 Administrative decision by the Investigator or Sponsor 
All efforts will be made to complete and report the observations as thoroughly as possible up to 
the date of withdrawal.  All information, including the reason for withdrawal from Study, must 
be recorded in the CRF. 
Any subject who withdraws consent to participate in the study will be removed from further 
treatment and study observation immediately upon the date of request.  These subjects should be 
encouraged to complete the early termination study procedures and observations at the time of 
withdrawal (Appendix A). 
For subjects withdrawn for reasons other than withdrawal of consent every effort should be made 
to complete the early termination study procedures and observations at the time of withdrawal 
(see Appendix A). 
8.10 
Concomitant Therapy and Procedures 
The use of concomitant therapies or procedures defined below must be recorded on the subject’s 
CRF.  AEs related to administration of these therapies or procedures must also be documented on 
the appropriate CRF. 
8.10.1 
Concomitant Therapy 
A concomitant therapy is any non-protocol specified drug or substance (including over-the-
counter medications, herbal medications and vitamin supplements) administered between 
screening and last telephone contact. 
Subjects should consult with the Site Investigator or qualified designee prior to initiating any 
new medication, including non-prescription compounds or any other non-drug therapy. 
Allowed Concomitant Therapy 
Throughout the study, Site Investigators or designated licensed physicians involved in the study 
may prescribe concomitant medications or treatments deemed necessary for adverse events or to 
provide adequate supportive care. 
Disallowed Concomitant Therapy 
Study subjects are prohibited from receiving other experimental agents during the study.  This 
includes marketed agents being used off-label and/or at experimental doses that are being tested 


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for the treatment of SMA.  The following agents are specifically prohibited:  valproate, riluzole, 
carnitine, creatine, sodium phenylbutyrate, hydroxyurea, salbutamol. 
8.10.2 
Concomitant Procedures 
A concomitant procedure is any therapeutic intervention (e.g., surgery/biopsy) or diagnostic 
assessment (e.g., blood gas measurement, bacterial cultures) performed between screening and 
last visit. 
9. 
SERIOUS AND NON-SERIOUS ADVERSE EVENT REPORTING  
9.1 
Sponsor Review of Safety Information 
Safety information will be collected, reviewed, and evaluated by the Sponsor in accordance with 
the Safety Management Plan throughout the conduct of the clinical trial. 
9.2 
Regulatory Requirements 
The Sponsor is responsible for regulatory submissions and reporting to the Investigators of SAEs 
including suspected unexpected serious adverse reactions (SUSARs) per the International 
Conference on Harmonization (ICH) guidelines E2A and ICH E6.  Country-specific regulatory 
requirements will be followed in accordance with local country regulations and guidelines. 
IRBs/REBs will be notified of any serious adverse event (SAE) according to applicable 
regulations.  The DSMB will be notified of any SAE as specified in the DSMB charter. 
The Sponsor will evaluate the available information and decide if there is a reasonable possibility 
that the Study Drug caused the AE and, therefore, meets the definition of a SUSAR. 
For the purpose of regulatory reporting of SUSARs, there are no “expected” AEs in this study 
population.  For Study Drug “expected” AEs, refer to the Investigator Brochure. 
9.3 
Definitions 
9.3.1 
Adverse Event 
An adverse event is any unfavorable and unintended sign (including a clinically significant 
abnormal laboratory finding, for example), symptom, or disease temporally associated with the 
study or use of investigational drug product, whether or not the AE is considered related to the 
investigational drug product. 
9.3.2 
Adverse Reaction and Suspected Adverse Reaction 
An adverse reaction is any adverse event caused by the Study Drug. 
A suspected adverse reaction is any adverse event for which there is a reasonable possibility that 
the drug caused the adverse event.  A suspected adverse reaction implies a lesser degree of 
certainty about causality than an adverse reaction. 
9.3.3 
Serious Adverse Event (SAE) 
A serious adverse event is any adverse event that in the view of either the Investigator or 
Sponsor, meets any of the following criteria: 


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 Results in death 
 Is life threatening: that is, poses an immediate risk of death at the time of the event 
An AE or suspected adverse reaction is considered “life-threatening” if, in the view of 
either the Investigator or Sponsor, its occurrence places the subject at immediate risk of 
death.  It does not include an AE or suspected adverse reaction that, had it occurred in a 
more severe form, might have caused death. 
 Requires inpatient hospitalization or prolongation of existing hospitalization 
Hospitalization is defined as an admission of greater than 24 hours to a medical facility 
and does not always qualify as an AE 
 Results in a persistent or significant incapacity or substantial disruption of the ability to 
conduct normal life functions 
 Results in congenital anomaly or birth defect in the offspring of the subject (whether the 
subject is male or female) 
 Important medical events that may not result in death, are not life-threatening, or do not 
require hospitalization may also be considered serious when, based upon appropriate 
medical judgment, they may jeopardize the subject and may require medical or surgical 
intervention to prevent one of the outcomes listed in this definition.  Examples of such 
medical events include allergic bronchospasm requiring intensive treatment in an 
emergency room or at home, blood dyscrasias or convulsions that do not result in 
inpatient hospitalization, or the development of drug dependency or drug abuse. 
9.4 
Monitoring and Recording Adverse Events 
Any pre-existing conditions or signs and/or symptoms present in a subject prior to the start of the 
study (i.e., before informed consent) should be recorded as Medical History and not recorded as 
AEs unless the pre-existing condition worsened.  The Investigator should always group signs and 
symptoms into a single term that constitutes a single unifying diagnosis if possible. 
9.4.1 
Serious Adverse Events 
In the interest of subject safety, and in order to fulfill regulatory requirements, all SAEs 
(regardless of their relationship to Study Drug) should be reported to the Sponsor within 
24 hours of the Study Center’s first knowledge of the event.  The collection of SAEs will begin 
after the subject signs the informed consent form and stop at the end of the subject’s follow-up 
period which is defined as the subject’s last visit.  When the Investigator is reporting by 
telephone, it is important to speak to someone in person versus leaving a message.  An Initial 
Serious Adverse Event Form should be completed and a copy should be faxed to the Sponsor. 
Detailed information should be actively sought and included on Follow-Up Serious Adverse 
Event Forms as soon as additional information becomes available.  All SAEs will be followed 
until resolution.  SAEs that remain ongoing past the subject’s last protocol-specified follow-up 
visit will be evaluated by the Investigator and Sponsor.  If the Investigator and Sponsor agree the 
subject’s condition is unlikely to resolve, the Investigator and Sponsor will determine the follow-
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9.4.2 
Non-Serious Adverse Events 
The recording of non-serious AEs will begin after the subject signs the informed consent form 
and will stop at the end of the subject’s follow-up period, which is defined as subject’s last visit.  
The Investigator will monitor each subject closely and record all observed or volunteered AEs on 
the Adverse Event Case Report Form. 
9.4.3 
Evaluation of Adverse Events (Serious and Non-Serious) 
The Investigator’s opinion of the following should be documented on the Adverse Event Case 
Report Form: 
9.4.3.1 
Relationship to the Study Drug 
The event’s relationship to the Study Drug is characterized by one of the following: 
 Related:  There is clear evidence that the event is related to the use of Study Drug 
e.g., confirmation by positive re-challenge test 
 Possible:  The event cannot be explained by the subject’s medical condition, concomitant 
therapy, or other causes, and there is a plausible temporal relationship between the event 
and Study Drug administration 
 Unlikely/Remote:  An event for which an alternative explanation is more likely 
(e.g., concomitant medications or ongoing medical conditions) or the temporal 
relationship to Study Drug administration and/or exposure suggests that a causal 
relationship is unlikely (For reporting purposes, Unlikely/Remote will be grouped 
together with Not Related) 
 Not Related:  The event can be readily explained by the subject’s underlying medical 
condition, concomitant therapy, or other causes, and therefore, the Investigator believes 
no relationship exists between the event and Study Drug 
9.4.3.2 
Severity 
The event’s severity is characterized by one of the following: 
 Mild:  The event is easily tolerated by the subject and does not affect the subject’s usual 
daily activities 
 Moderate:  The event causes the subject more discomfort and interrupts the subject’s 
usual daily activities 
 Severe:  The event is incapacitating and causes considerable interference with the 
subject’s usual daily activities 
If the event is an SAE, then all applicable seriousness criteria must be indicated (criteria listed in 
Section 9.3.3). 


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9.4.3.3 
Action Taken with Study Drug 
Action taken with Study Drug due to the event is characterized by one of the following. 
 None:  No changes were made to Study Drug administration and dose 
 Permanently Discontinued:  Study drug was discontinued and not restarted 
 Temporarily Interrupted, restarted – same dose:  Dosing was temporarily interrupted 
or delayed due to the AE and restarted at the same dose 
9.4.3.4 
Treatment Given for Adverse Event 
Any treatment (e.g., medications or procedures) given for the AE should be recorded on the 
Adverse Event Case Report Form (treatment should also be recorded on the concomitant 
treatment or ancillary procedures CRF as appropriate). 
9.4.3.5 
Outcome of the Adverse Event 
If the event is a non-serious AE then the event’s outcome is characterized by one of the 
following: 
 AE Persists:  Subject terminates from the trial and the AE continues 
 Recovered:  Subject recovered completely from the AE 
 Became Serious:  The event became serious (the date that the event became serious 
should be recorded as the Resolution Date of that AE and the Onset Date of the 
corresponding SAE) 
 Change in Severity (if applicable):  AE severity changed 
If the event is a SAE then the event’s outcome is characterized by one of the following: 
 Ongoing:  SAE continuing 
 Persists (as non-serious AE):  Subject has not fully recovered but the event no longer 
meets serious criteria and should be captured as an AE on the non-serious AE electronic 
case report form (eCRF) (the SAE resolution date should be entered as the date of onset 
of that AE) 
 Recovered:  Subject recovered completely from the SAE (the date of recovery should be 
entered as the SAE resolution date) 
 Fatal:  Subject died (the date of death should be entered as the SAE resolution date) 
9.5 
Procedures for Handling Special Situations 
9.5.1 
Abnormalities of Laboratory Tests 
Clinically significant abnormal laboratory test results may, in the opinion of the Investigator, 
constitute or be associated with an AE.  Examples of these include abnormal laboratory results 


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that are associated with symptoms, or require treatment, e.g., bleeding due to thrombocytopenia, 
tetany due to hypocalcemia, or cardiac arrhythmias due to hyperkalemia.  Whenever possible, the 
underlying diagnosis should be listed in preference to abnormal laboratory values as AEs.  
Clinically significant abnormalities will be monitored by the Investigator until the parameter 
returns to its baseline value or until agreement is reached between the Investigator and 
Ionis Medical Monitor.  Laboratory abnormalities deemed not clinically significant (NCS) by the 
Investigator should not be reported as AEs.  Similarly, laboratory abnormalities reported as AEs 
by the Investigator should not be deemed NCS on the laboratory sheet. 
The Investigator is responsible for reviewing and signing all laboratory reports.  The signed 
clinical laboratory reports will serve as source documents. 
9.5.2 
Prescheduled or Elective Procedures or Routinely Scheduled Treatments 
A prescheduled or elective procedure or a routinely scheduled treatment will not be considered 
an SAE, even if the subject is hospitalized; the Study Center must document all of the following: 
 The prescheduled or elective procedure or routinely scheduled treatment was scheduled 
(or was on a waiting list to be scheduled) prior to obtaining the subject’s consent to 
participate in the study 
 The condition that required the prescheduled or elective procedure or routinely scheduled 
treatment was present before and did not worsen or progress in the opinion of the 
Investigator between the subject’s consent to participate in the study and the timing of the 
procedure or treatment 
 The prescheduled or elective procedure or routinely scheduled treatment is the sole 
reason for the intervention or hospital admission 
9.5.3 
Dosing Errors 
Study Drug dosing errors should be documented as Protocol Deviations.  A brief description 
should be provided in the deviation, including whether the subject was symptomatic (list 
symptoms) or asymptomatic, and the event accidental or intentional. 
Dosing details should be captured on the Dosing CRF.  If the subject takes a dose of Study Drug 
that exceeds protocol specifications and the subject is symptomatic, then the symptom(s) should 
be documented as an AE and be reported per Section 9.4. 
Should an overdose occur, the Investigator or designee should refer to the Guidance to 
Investigator’s section of the Investigator’s Brochure and contact the Sponsor within 24 hours. 
10. 
STATISTICAL CONSIDERATIONS 
10.1 
Study Endpoints 
10.1.1 
Primary Endpoint 
 Achievement of motor milestones as evaluated by Module 2 of the Hammersmith Infant 
Neurological Examination  


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10.1.2 
Secondary Endpoints 
The efficacy of multiple doses of ISIS 396443 as assessed by: 
 Event-free survival determined by the proportion of subjects who are alive and do not 
require permanent ventilatory support (defined as tracheostomy or the need for 
≥ 16 hours ventilation/day continuously for at least 14 days in the absence of an acute 
reversible illness) 
 Improvement in muscle strength as measured by the Children’s Hospital of Philadelphia 
Infant Test for Neuromuscular Disease (CHOP INTEND) 
 Improvement in neuromuscular electrophysiology measured by the Compound Muscle 
Action Potential (CMAP) of the ulnar and peroneal nerves 
The safety and tolerability of multiple doses of ISIS 396443 as assessed by: 
 Adverse events 
 Neurological examinations 
 Vital signs 
 Physical examinations and weight 
 Clinical laboratory tests (serum chemistry, hematology, urinalysis) 
 CSF laboratory tests (cell count, protein, glucose) 
 ECGs 
 Use of concomitant medications 
CSF and plasma PK of ISIS 396443. 
10.1.3 
Exploratory Endpoints 
The following efficacy evaluations will be performed as exploratory endpoints: 
 Measures of respiratory status (number of respiratory events, respiratory infections, 
respiratory-related hospitalizations, ventilator use, O2 saturation awake) 
 Growth parameters (weight for age/length, head circumference, chest circumference, 
head to chest circumference ratio, arm circumference) 
10.2 
Sample Size Considerations 
There is no statistical rationale for the selected sample size of 4 treated subjects in Cohort 1 and 
4 to approximately 16 subjects in Cohort 2.  The sample size was selected to ensure that the 
safety and tolerability of ISIS 396443 will be adequately assessed while minimizing unnecessary 
subject exposure rather than statistical considerations for the efficacy endpoints. 


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10.3 
Populations 
Safety Population:  All subjects who are registered and receive at least one dose of Study Drug. 
Pharmacokinetic Population:  All subjects who are registered and for which there is at least one 
evaluable post-dose pharmacokinetic sample. 
Efficacy Population: all subjects who are registered, receive all scheduled loading doses of Study 
Drug (i.e., Days 1, 15 and 85), and complete the Day 92 visit. 
10.4 
Interim Analysis 
Interim efficacy and safety analyses will be performed to provide content for regulatory 
submissions and to support ISIS 396443 drug development planning and business activities. 
Details of the analyses are contained in the Statistical Analysis Plan. 
10.5 
Planned Methods of Analysis 
10.5.1 
Demographic and Baseline Characteristics 
Demographic and baseline characteristics will be summarized descriptively for each dose cohort. 
10.5.2 
Efficacy Analysis 
The primary analysis will include the proportion of subjects who achieved improvement in motor 
milestones at their last visit.  It will be tabulated by cohort and overall. 
Motor milestones will be accessed using Section 2 of the Hammersmith Infant Neurological 
Exam (HINE) which is comprised of 8 independent milestone categories:  voluntary grasp, 
ability to kick in supine position, head control, rolling, sitting, crawling, standing, and walking.  
Within each of these categories, subjects can progress from complete inability to perform a 
motor task, for example no grasp, no kicking, inability to maintain head upright, all the way to 
complete mastery of each category, for example achievement of pincer grasp, touching toes 
while supine, or maintaining their head upright. 
Event-free survival will be evaluated by determining a proportion of subjects who are alive and 
do not require permanent ventilatory support.  Permanent ventilation is defined as tracheostomy 
or the need for ≥ 16 hours ventilation/day continuously for at least 14 days in the absence of an 
acute reversible illness. 
The CHOP INTEND infant motor function scale total score, change, and percent change from 
baseline will be summarized by cohort and overall, and by visit. 
The absolute values for CMAP amplitude and CMAP area, as well as absolute and percent 
changes from baseline, will be summarized by cohort and overall, and by visit. 
Full details of the methodology for analyses described above, as well as all additional analyses, 
are contained in the Statistical Analysis Plan. 
10.5.3 
Safety and Pharmacokinetic Analysis 
All subjects exposed to ISIS 396443 will be included in the safety analyses. 


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Treatment duration and amount of Study Drug received will be summarized by dose cohort.  
Subject incidence rates of all adverse events will be tabulated by Medical Dictionary for 
Regulatory Activities (MedDRATM) system organ class, and by MedDRATM term.  Tables and/or 
narratives of “on-study” deaths, serious and significant adverse events, including early 
withdrawals due to adverse events, will also be provided. 
All treatment emergent AEs, all treatment emergent AEs potentially related to Study Drug, all 
treatment emergent serious AEs, and all treatment emergent serious AEs potentially related to 
Study Drug will be summarized. 
Laboratory tests including chemistry panel, complete blood count with differential, urinalysis, 
and CSF components will be summarized by study visit for each dose cohort.  These safety 
variables will also be presented over time after Study Drug administration, as appropriate, using 
results prior to Day 1 dosing as Baseline.  Vital sign results will be presented similarly. 
Physical and neurological examination findings and results from ECG will be listed for review.  
As appropriate, results will also be summarized descriptively for each dose cohort.  Concomitant 
medication usage for each subject will be listed for review.  Plasma pharmacokinetic parameters 
and ISIS 396443 concentrations in plasma and CSF for the Pharmacokinetic population will be 
summarized by dose cohort using descriptive statistics and, where warranted, presented 
graphically. 
11. 
INVESTIGATOR’S REGULATORY OBLIGATIONS 
11.1 
Informed Consent 
The written informed consent documents should be prepared in the language(s) of the potential 
subject population, based on an English version provided by the Sponsor and should be easy to 
understand. 
Before a subject’s participation in the trial, the Investigator is responsible for obtaining written 
informed consent from the parent or legal guardian after adequate explanation of the aims, 
methods, anticipated benefits, and potential hazards of the study and before any protocol-specific 
screening procedures or any Study Drugs are administered.  Sufficient time must be given to 
consider whether to participate in the study. 
The acquisition of informed consent and the parent/legal guardian’s agreement or refusal of 
his/her notification of the primary care physician should be documented in the subject’s medical 
records, and the informed consent form(s) should be signed and personally dated by the 
parent/legal guardian and by the study person who conducted the informed consent discussion.  
The original signed informed consent form(s) should be retained in the Study Master File and in 
any other locations required by institutional policy, and a copy of the signed consent form(s) 
should be provided to the parent or guardian. 
11.2 
Ethical Conduct of the Study 
The Guidelines of the World Medical Association (WMA) Declaration of Helsinki dated 
October 2002 the applicable regulations and guidelines of current Good Clinical Practice (GCP) 
as well as the demands of national drug and data protection laws and other applicable regulatory 
requirements will be strictly followed. 


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11.3 
Institutional Review Board/Research Ethics Board 
A copy of the protocol, proposed informed consent form, other written subject information, and 
any proposed advertising material must be submitted to the IRB/REB for written approval.  A 
copy of the written approval of the protocol and informed consent form must be received by the 
Sponsor before recruitment of subjects into the study and shipment of Study Drug.  A copy of the 
written approval of any other items/materials that must be approved by the Study Center or 
IRB/REB must also be received by the Sponsor before recruitment of subjects into the study and 
shipment of Study Drug.  The Investigator’s Brochure must be submitted to the IRB/REB for 
acknowledgement. 
The Investigator must submit to and, where necessary, obtain approval from the IRB/REB for all 
subsequent protocol amendments and changes to the informed consent documents.  The 
Investigator should notify the IRB/REB of deviations from the protocol in accordance with ICH 
GCP Section 4.5.2.  The Investigator should also notify the IRB/REB of serious adverse events 
occurring at the Study Center and other adverse event reports received from the Sponsor, in 
accordance with local procedures. 
The Investigator will be responsible for obtaining annual IRB/REB approval/renewal throughout 
the duration of the study.  Copies of the Investigator’s reports, all IRB/REB submissions and the 
IRB/REB continuance of approval must be sent to the Sponsor. 
11.4 
Subject Confidentiality 
The Investigator must ensure that the subject’s confidentiality is maintained.  On the CRFs or 
other documents submitted to the Sponsor, subjects should be identified by unique initials and a 
subject study number only.  Documents that are not for submission to the Sponsor (e.g., signed 
informed consent/assent forms) should be kept in strict confidence by the Investigator. 
In compliance with federal and local regulations/ICH GCP Guidelines, it is required that the 
Investigator and institution permit authorized representatives of the company, of the regulatory 
agency(s), and the IRB/REB direct access to review the subject’s original medical records for 
verification of study-related procedures and data.  Direct access includes examining, analyzing, 
verifying, and reproducing any records and reports that are important to the evaluation of the 
study.  The Investigator is obligated to inform and obtain the consent of the subject’s parent or 
guardian to permit named representatives to have access to his/her study-related records without 
violating the confidentiality of the subject. 
12. 
ADMINISTRATIVE AND LEGAL OBLIGATIONS 
12.1 
Protocol Amendments 
Protocol amendments must be made only with the prior approval of the Sponsor.  Agreement 
from the Investigator must be obtained for all protocol amendments and amendments to the 
informed consent document.  The regulatory authority and IRB/REB must be informed of all 
amendments and give approval for any amendments likely to affect the safety of the subjects or 
the conduct of the trial.  The Investigator must send a copy of the approval letter from the 
IRB/REB to the Sponsor. 


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12.2 
Study Termination 
Both the Sponsor and the Investigator reserve the right to terminate the study, according to the 
terms of the study contract.  The Investigator/Sponsor should notify the IRB/REB in writing of 
the trial’s completion or early termination. 
12.3 
Study Documentation and Storage 
The Investigator should ensure that all appropriately qualified persons to whom he/she has 
delegated trial duties are recorded on a Sponsor-approved Delegation of Site Responsibilities 
Form. 
Source documents are original documents, data, and records from which the subject’s CRF data 
are obtained.  These include but are not limited to hospital records, clinical and office charts, 
laboratory and pharmacy records, imaging, and correspondence.  In this study, CRFs may not be 
used as source documents. 
The Investigator and Study Center staff is responsible for maintaining a comprehensive and 
centralized filing system of all study-related (essential) documentation in accordance with 
Section 8 of the ICH Guidelines (E6), suitable for inspection at any time by representatives from 
the Sponsor and/or applicable regulatory authorities.  Elements should include: 
 Subject files containing completed CRFs, informed consents, and supporting copies of 
source documentation 
 Study files containing the protocol with all amendments, Investigator’s Brochure, copies 
of pre-study documentation and all correspondence to and from the IRB/REB and the 
Sponsor 
 If drug supplies are maintained at the Study Center, proof of receipt, Study Drug Product 
Accountability Record, Return of Study Drug Product for Destruction, and all drug-
related correspondence 
In addition, all original source documents supporting entries in the CRFs must be maintained and 
be readily available. 
No study document should be destroyed without prior written agreement between the Sponsor 
and the Investigator.  Should the Investigator wish to assign the study records to another party or 
move them to another location, he/she must notify the Sponsor. 
12.4 
Study Monitoring 
The Sponsor representative and regulatory authority inspectors are responsible for contacting and 
visiting the Investigator for the purpose of inspecting the facilities and, upon request, inspecting 
the various records of the trial (e.g., CRFs and other pertinent data) provided that subject 
confidentiality is respected. 
The Sponsor monitor is responsible for inspecting the CRFs at regular intervals throughout the 
study to verify adherence to the protocol; completeness, accuracy, and consistency of the data; 
and adherence to local regulations on the conduct of clinical research.  The monitor should have 


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access to subject medical records and other study-related records needed to verify the entries on 
the CRFs. 
The Investigator agrees to cooperate with the monitor to ensure that any problems detected in the 
course of these monitoring visits, including delays in completing CRFs, are resolved. 
In accordance with ICH GCP and the Sponsor’s audit plans, this study may be selected for audit 
by representatives from the Sponsor’s Clinical Quality Assurance Department. Inspection of 
Study Center facilities (e.g., pharmacy, drug storage areas, laboratories) and review of study-
related records will occur to evaluate the trial conduct and compliance with the protocol, ICH 
GCP, and applicable regulatory requirements. 
To ensure the quality of clinical data a clinical data management review will be performed on 
subject data received by the Sponsor.  During this review, subject data will be checked for 
consistency, omissions, and any apparent discrepancies.  In addition, the data will be reviewed 
for adherence to the protocol and GCP.  To resolve any questions arising from the clinical data 
management review process, data queries and/or Study Center notifications will be sent to the 
Study Center for completion and return to Sponsor. 
The Principal Investigator will sign and date the indicated places on the CRF.  These signatures 
will indicate that the principal Investigator inspected or reviewed the data on the CRF, the data 
queries, and the Study Center notifications, and agrees with the content. 
12.5 
Language 
CRFs must be completed in English.  Generic names for concomitant medications should be 
recorded in English if possible, unless it is a combination drug, then record the trade name in 
English. 
All written information and other material to be used by subjects and investigative staff must use 
vocabulary and language that are clearly understood. 
12.6 
Compensation for Injury 
The Sponsor maintains appropriate insurance coverage for clinical trials and will follow 
applicable local compensation laws.  Subjects will be treated and/or compensated for any 
study-related illness/injury in accordance with the information provided in the Compensation for 
Injury section of the Informed Consent document. 
 
 


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13. 
REFERENCES 
Campbell SK, Swanlund A, Smith E, et al. Validity of the TIMPSI for estimating concurrent performance on the test 
of infant motor performance. Pediatric Physical Therapy 2008 Spring; 20: 3-10. 
Cho S, Dreyfuss G. A degron created by SMN2 Exon 7 skipping is a principal contributor to spinal muscular 
atrophy severity. Genes Dev 2010; 24: 438-442. 
Coovert DD, Le TT, McAndrew PE, et al. The survival motor neuron protein in spinal muscular atrophy. Hum Mol 
Genet 1997; 6: 1205-1214. 
Feldkotter M, Schwarzer V, Wirth R, et al. Quantitative analyses of SMN1 and SMN2 based on real-time light 
cycler PCR: fast and highly reliable carrier testing and prediction of severity of spinal muscular atrophy. Am J Hum 
Genet 2002; 70: 358-368. 
Finkel RS, Hynan LS, Glanzman AM, et al; AmSMART Group. The test of infant motor performance: reliability in 
spinal muscular atrophy type I. Pediatric Physical Therapy 2008 Fall; 20: 242-246. 
Geary RS, Yu RZ, Watanabe T, et al. Pharmacokinetics of a tumor necrosis factor-α phosphorothioate 2′-O-(2-
Methoxyethyl) modified antisense oligonucleotide: comparison across species. Drug Metab Disp 2003; 31: 
1419-1428. 
Glanzman AM, Mazzone E, Main M, et al. The Children’s Hospital of Philadelphia Infant Test of Neuromuscular 
Disorders (CHOP INTEND): Test development and reliability. Neuromuscular Disorders 2010; 20: 155-161. 
Helmken C, Hofmann Y, Schoenen F, et al. Evidence for a modifying pathway in SMA discordant families: reduced 
SMN level decreases the amount of its interacting partner and Htra2-beta1. Hum Genet 2003; 114: 11-21. 
Henry S, Stecker K, Brooks D, et al. Chemically modified oligonucleotides exhibit decreased immune stimulation in 
mice. J Pharmacol Exp Ther 2000; 292: 468-479. 
Henry SP, Johnson M, Zanardi TA, et al. Renal uptake and tolerability of a 2′-O-methoxyethyl modified antisense 
oligonucleotide (ISIS 113715) in monkey. Toxicology 2012; 301: 13-20. 
Hua Y, Vickers TA, Baker BF, et al. Enhancement of SMN2 Exon 7 inclusion by antisense oligonucleotides 
targeting the exon. PLoS Biol 2007; 5: e73. 
Hua Y, Vickers TA, Okunola HL, et al. Antisense masking of an HnRNP A1/A2 intronic splicing silencer corrects 
SMN2 splicing in transgenic mice. Am J Hum Genet 2008; 82: 834-848. 
Keren H, Lev-Maor G, Ast G. Alternative splicing and evolution: diversification, exon definition and function. 
Nat.Rev.Genet 2010; 11: 345-355. 
Le TT, Pham LT, Butchbach ME, et al. SMNΔ7, the major product of the centromeric survival motor neuron 
(SMN2) gene, extends survival in mice with spinal muscular atrophy and associates with full-length SMN. Hum 
Mol Genet 2005; 14: 845-857. 
Lee LCY, Sennett M, Erickson JM. Prevention and management of post-lumbar puncture headache in pediatric 
oncology patients. J Ped Onc Nurs 2007; 24: 200-207. 
Lefebvre S, Bürglen L, Reboullet S, et al. Identification and characterization of a spinal muscular atrophy-
determining gene. Cell 1995; 80: 155-165. 
Lefebvre S, Burlet P, Liu Q, et al. Correlation between severity and SMN protein level in spinal muscular atrophy. 
Nat Genet 1997; 16: 265-269. 
Lewalt AJ, Krosschell KJ, Scott C, et al. Compound muscle action potential and motor function in children with 
spinal muscular atrophy. Muscle Nerve 2010; 42(5); 703-708. 
Main M, Kairon H, Mercuri E, Muntoni F. The Hammersmith Functional Motor Scale for children with spinal 
muscular atrophy: a scale to test ability and monitor progress in children with limited ambulation. Eur J Paediatr 
Neurol 2003; 7(4): 155-159. 


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Matsuzawa J, Matsui M, Konishi T, et al. Age-related volumetric changes of brain gray and white matter in healthy 
infants and children. Cereb Cortex 2001 Apr; 11(4):335-342. 
McKay RA, Miraglia LJ, Cummins LL, et al. Characterization of a potent and specific class of antisense 
oligonucleotide inhibitor of human protein kinase C-α expression. J Bio Chem 1999; 274: 1715-1722. 
O’Hagen JM, Glanzman AM, McDermott MP, et al. An expanded version of the Hammersmith Functional Motor 
Scale for SMA II and III patients. Neuromuscular Disorders 2007; 17: 693-697. 
Passini MA, Bu J, Richards AM, et al. Antisense oligonucleotides delivered to the mouse CNS ameliorate symptoms 
of severe spinal muscular atrophy. Science Trans Med 2011; 3: 1-12. 
Prior TW, Swoboda KJ, Scott HD, Hejmanowski AQ. Homozygous SMN1 deletions in unaffected family members 
and modification of the phenotype by SMN2 gene. Am J Med Genet 2004; 130A: 307-310. 
Shefner JM, Watson ML, Simionescu L, et al. Multipoint incremental motor unit number estimation as an outcome 
measure in ALS. Neurology 2011; 77: 235-241. 
Swoboda KJ, Prior TW, Scott CB, et al. Natural history of denervation in SMA: relation to age, SMN2 copy 
number, and function. Annals of Neurology 2005; 57: 704-712. 
Wang CH, Finkel RS, Bertini ES, et al; Participants of the International Conference on SMA Standard of Care. 
Consensus statement for standard of care in spinal muscular atrophy. J Child Neurol 2007; 22:1027-1049. 
 


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14. 
APPENDICES 


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Appendix A 
Schedule of Procedures 
 


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Appendix A – Schedule of Procedures for ‘Loading’ Dosing Period, Visits Only* 
* 
Note that monitoring telephone calls will also occur on Study Days 8 (±1), 43 (±2), 57 (±2), 71 (±2), 106 (±3), 
127 (±3), 134 (±3), 148 (±3), 189 (±3), 197 (±3), 218 (±3), 239 (±3) (i.e., every 2-3 weeks throughout the study)13 
Study Period 
Screen 
Dosing 
Study Day 
D -21 
to 
D -1 
D1 
Injection 
D2 
D15 (±1D) 
Injection 
D16 D29 
(±1D) 
D85 (±2D) 
Injection 
D86 
 
D92 
(±1D) 
D169 
(±5D) 
Pre- 
dose LP Post- 
dose 
Pre- 
dose LP Post- 
dose 
Pre- 
dose LP 
Post-
dose 
Study Drug:  LP 
Injection10 
 
 
X 
 
 
 
X9 
 
 
 
 
X9 
 
 
 
 
In-Patient Stay 
(24 hours) 
 
 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
Informed Consent 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Inclusion/Exclusion 
Criteria 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Medical History 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
SMN Genetics1 
 
 
 
 
 
 
 
 
 
 
X 
 
 
 
 
 
Vital Signs2 
X 
X 
 
4X3 
X5 
X 
 
4X3 
X 
X 
X 
 
4X3 
X 
X 
X 
Weight 
X 
X 
 
 
X 
 
 
 
X 
X 
X 
 
 
X 
X 
X 
Growth Parameters4 
X 
 
 
 
 
 
 
 
 
X 
 
 
 
 
X 
X 
Physical Examination 
X 
X 
 
 
 
X 
 
 
 
X 
X 
 
 
 
X 
X 
Ventilator Use 
X 
X 
 
 
 
X 
 
 
 
X 
X 
 
 
 
X 
X 
Neurological 
Examination 
X 
X 
 
2X6 
X5 
X 
 
2X6 
X 
X 
X 
 
2X6 
X 
X 
X 
ECG 
X 
 
 
 
 
 
 
 
 
X 
 
 
 
 
X 
 
Safety Labs7, 12 
X 
 
 
 
 
 
 
 
 
X 
 
 
 
 
X 
X 
Coagulation Labs 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Immunogenicity  
 
X 
 
 
 
 
 
 
 
 
X 
 
 
 
 
X 
CSF Safety12 
 
X 
 
 
 
X 
 
 
 
 
X 
 
 
 
 
 
CSF PK11 
 
X 
 
 
 
X 
 
 
 
 
X 
 
 
 
 
 
Plasma PK11 
 
X 
 
3X 
X 
 
 
 
 
 
X 
 
X 
 
X 
 
CHOP INTEND 
X8 
 
 
 
 
 
 
 
 
X 
 
 
 
 
X 
X 
CMAP 
X8 
 
 
 
 
 
 
 
 
X 
 
 
 
 
X 
X 
Con Med Recording 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Adverse Event 
Collection 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
1 
Only for those patients who do not have documented evidence of SMN genetics from Athena Diagnostics 
2 
Resting blood pressure, pulse, respiratory rate, temperature, and pulse oximetry awake.  Pulse oximetry asleep will 
also be assessed at screening only 
3 
Vital signs performed 1, 2, 4, 6 hours after dosing 
4 
Length, weight for age/length, head circumference, chest circumference, head to chest circumference ratio, arm 
circumference 
5 
Conducted within 20-24 hours after dosing 
6 
Neurological exams at 3 and 6 hours after dosing 
7 
Serum chemistry, hematology, urinalysis panels (see Appendix B for analytes).  Cystatin C will not be analyzed at 
Day 29 due to blood volume limitations 
 


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Appendix A – Schedule of Procedures for ‘Loading’ Dosing Period, Visits Only* Continued 
Legend Text Continued 
8 
CMAP and CHOP INTEND assessments do not form part of the screening assessment or inclusion/exclusion criteria 
but are baseline measurements taken during the screening phase of the study 
9 
Overnight stay is optional on Day 15 and Day 85, if needed due to complications of the LP procedure 
10 Injections may not be scheduled within 72 hours after an immunization 
11 Refer to Appendix C for PK sampling schedule 
12 Refer to Appendix B for analytes 
13 At telephone contact, changes in concomitant medications and adverse events will be recorded as well as information 
on the subject’s ventilator use and SMA disease status 


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Appendix A – Schedule of Procedures for ‘Maintenance’ Dosing Period, Visits Only* 
* 
Note that monitoring telephone calls will also occur on Study Days 274 (±3), 295 (±3), 316 (±3), 358 (±3), 
400 (±3), 421 (±3), 463 (±3), 484 (±3), 526 (±3), 547 (±3), 589 (±3), 610 (±3), 652(±3), 673 (±3), 715 (±3), 
736 (±3), 778 (±3), 799 (±3), 841 (±3), 862 (±3), 904 (±3), 925 (±3), 967 (±3), 988 (±3), 1030 (±3), 1051 (±3), 
1093 (±3), 1114 (±3), 1156 (±3), 1177 (±3), 1219 (±3), 1240 (±3), 1282 (±3), 1303 (±3), and 1324 (±3) (i.e., 
approximately every 3 weeks)10 
Study Period 
Dosing/Followup 
Study Day 
D253 
Injection 
(±7D) 
D254 D337 
(±7D) 
D379, 505, 631, 757, 
883, 1009, 1135, and 
1261 Injection 
(±7D) 
D380, 
506, 
632,7
58, 
884, 
1010, 
1136, 
1262 
D442, 
568, 
694, 
820, 
946, 
1072, 
1198 
(±7D) 
D1352 
(±7D) 
or 
Early 
Termination 
(ET) 
Pre- 
dose 
LP 
Post- 
dose 
Pre-
dose 
LP 
Post-
dose 
Study Drug: LP Injection7 
 
X6 
 
 
 
 
X6 
 
 
 
 
Vital Signs1 
X 
 
4X2 
X 
X 
X 
 
4X2 
 
X 
X 
Weight 
X 
 
 
 
X 
X 
 
 
 
X 
X 
Growth Parameters3 
X11 
 
 
 
X 
X11 
 
 
 
X 
X 
Physical Examination 
X 
 
 
 
X 
X 
 
 
 
X 
X 
Ventilator Use 
X 
 
 
 
X 
X 
 
 
 
X 
X 
Neurological Examination 
X 
 
2X4 
X 
X 
X 
 
2X4 
 
X 
X 
ECG 
 
 
 
 
X 
 
 
 
 
X12 
X 
Safety Labs5, 9 
X11 
 
 
 
X 
 
 
 
 
X 
X 
Immunogenicity13 
X11 
 
 
 
X 
 
 
 
 
X 
X 
CSF Safety9 
X 
 
 
 
 
X 
 
 
 
 
 
CSF PK8 
X 
 
 
 
 
X 
 
 
 
 
 
Plasma PK8 
X11 
 
 
 
 
X11 
 
 
 
 
X 
CHOP INTEND 
X11 
 
 
 
X 
X11 
 
 
 
X 
X 
HFMSE14 
 
 
 
 
 
X11 
 
 
 
X 
X 
CMAP 
X11 
 
 
 
X 
 
 
 
 
X 
X 
Phone Contact 
 
 
 
 
 
 
 
 
X 
 
 
Con Med Recording 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Adverse Event Collection 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
1 
Resting blood pressure, pulse, respiratory rate, temperature, and pulse oximetry awake 
2 
Vital signs performed 1, 2, 4, 6 hours after dosing 
3 
Length, weight for age/length, head circumference, chest circumference, head to chest circumference ratio, arm 
circumference 
4 
Neurological exams at 3 and 6 hours after dosing 
5 
Serum chemistry, hematology, urinalysis panels (see Appendix B for analytes) 
6 
Overnight stay is optional on Days 253, 379, 505, 631, 757, 883, 1009, 1135, and 1261 if needed due to 
complications of the LP procedure 
7 
Injections may not be scheduled within 72 hours after an immunization 
8 
Refer to Appendix C for PK sampling schedule 
9 
Refer to Appendix B for analytes 
10 At telephone contact, changes in concomitant medications and adverse events will be recorded as well as 
information on the subject’s ventilator use and SMA disease status 
11 These assessments can be performed up to 7 days prior to dosing, if necessary 


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Appendix A – Schedule of Procedures for ‘Maintenance’ Dosing Period, Visits Only* 
Continued 
Legend Text Continued 
12 ECG should be performed on Day 568 only 
13 Immunogenicity assessments will be performed as deemed necessary on samples collected on other days 
14 All subjects who have maintained a CHOP INTEND total score of ≥ 50 for 2 consecutive study visits will be 
evaluated using the Hammersmith Functional Motor Scale Expanded (HFMSE) in addition to CHOP INTEND 
 


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Appendix B 
Laboratory Analytes 
 
 
 


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Appendix B Laboratory Analytes 
CLINICAL SAFETY ASSESSMENTS 
(minimum requirements) 
OTHER ASSESSMENTS 
Clinical Chemistry 
Sodium 
Potassium 
Chloride 
Total protein 
Albumin 
Calcium 
Phosphorus 
Bicarbonate 
Glucose 
BUN 
Creatinine 
Cystatin C 
Total serum Bilirubin 
Alkaline phosphatase 
AST (SGOT) 
ALT (SGPT) 
CPK 
 
Coagulation 
aPTT 
PT 
INR 
 
Immunogenicity Evaluation 
ISIS 396443 plasma Abs 
 
CSF Safety (Minimum 
Requirements) 
Red blood cells 
White blood cells 
Glucose 
Protein 
 
Urinalysis  
Specific gravity 
pH 
Protein 
Glucose 
Ketones 
Bilirubin 
Blood 
Red blood cells 
White blood cells 
Epithelial cells 
Bacteria 
Casts 
Crystals 
 
Hematology 
Red blood cells 
Hemoglobin 
Hematocrit 
Platelets 
White blood cells 
WBC Differential 
 
(% and absolute) 
 
Neutrophils 
 
Eosinophils 
 
Basophils 
 
Lymphocytes 
 
Monocytes 
*PK 
Plasma ISIS 396443 levels 
CSF ISIS 396443 levels 
 
SMN Genetics 
SMN2 Copy Number 
SMN Gene Sequencing 
 
 
 
* 
Any of the collected PK plasma and CSF samples from the study patients may also be used for investigation of 
possible biomarkers of SMA disease or the pharmacodynamic effects of ISIS 396443 (e.g., CSF mRNA, CSF 
protein, CSF miRNA panel) or for profiling of drug binding proteins, bioanalytical method validation purposes, 
stability assessments, metabolite assessments, immunogenicity assessments (including assay development and 
validation purposes), or to assess other actions of ISIS 396443 with plasma and CSF constituents 
 
 
 


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Appendix C 
Pharmacokinetic Sampling Schedule 
 
 


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Appendix C Pharmacokinetic Sampling Schedule 
Treatment Period 
Study Day 
Timepoints 
Blood 
Collection  
CSF Collection  
Multiple Dose: 
LP Injection 
D1 
Predose 
0.35 mL 
0.5 mL 
1 hr 
0.35 mL 
NA 
2 hr 
0.35 mL 
NA 
4 hr 
0.35 mL 
NA 
D2 
24 hr 
0.35 mL 
NA 
D15 
Predose 
NA 
0.5 mL 
D85 
Predose 
0.35 mL 
0.5 mL 
4 hr 
0.35 mL 
NA 
D92 
Anytime 
0.35 mL 
NA 
D253 
Predose 
0.35 mL 
0.5 mL 
D379 
Predose 
0.35 mL 
0.5 mL 
D505 
Predose 
0.35 mL 
0.5 mL 
D631 
Predose 
0.35 mL 
0.5 mL 
D757 
Predose 
0.35 mL 
0.5 mL 
D883 
Predose 
0.35 mL 
0.5 mL 
D1009 
Predose 
0.35 mL 
0.5 mL 
D1135 
Predose 
0.35 mL 
0.5 mL 
D1261 
Predose 
0.35 mL 
0.5 mL 
D1352/ET 
Anytime 
0.35 mL 
NA 
NA 
Not applicable (No Collection Scheduled) 
Details on sampling, preparation, and shipment are included in the study laboratory manual 
Any of the collected PK plasma and CSF samples from the study patients may also be used for investigation of 
possible biomarkers of SMA disease or the pharmacodynamic effects of ISIS 396443 (e.g., CSF mRNA, CSF protein, 
CSF miRNA panel) or for profiling of drug binding proteins, bioanalytical method validation purposes, stability 
assessments, metabolite assessments, immunogenicity assessments (including assay development and validation 
purposes), or to assess other actions of ISIS 396443 with plasma and CSF constituents 


 
 
 
 
ISIS PHARMACEUTICALS, INC. 
 
 
ISIS 396443-CS3A 
 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Amendment 5 – 29 May 2015 
 
 
 
 
 
Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 


PPD


ISIS 396443-CS3A 
CONFIDENTIAL
Amendment 5
Protocol 
29 May 2015
 
3 
 
ISIS 396443 
Isis Protocol Number ISIS 396443-CS3A 
Protocol Amendment 5 
 
Clinical Phase:  2 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Trial Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 
Phone: + 01 760 931 9200 
Fax: 
+ 01 760 603 2700 
Key Sponsor Contact: 
 
 
 
Phone: +
 
Fax: 
+ 
 
Date: 
29 May 2015 
 
 
Confidentiality Statement 
This document contains confidential information of Isis Pharmaceuticals, Inc. that must not be 
disclosed to anyone other than the recipient study staff and members of the institutional review 
board or authorized regulatory agencies.  This information cannot be used for any purpose other 
than the evaluation or conduct of the clinical investigation without the prior written consent of 
Isis Pharmaceuticals, Inc. 
PPD
PPD
PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 5  
Protocol 
 
29 May 2015 
 
4 
Protocol Signature Page 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
Amendment 5 
Date: 
29 May 2015 
 
I hereby acknowledge that I have read and understand the attached clinical protocol, entitled “A 
Study to Assess the Safety, Tolerability, and Pharmacokinetics of Multiple Doses of ISIS 396443 
Delivered Intrathecally to Patients with Infantile-Onset Spinal Muscular Atrophy”, dated 
29 May 2015, and agree to conduct the study as described herein. 
I agree to comply with the International Conference on Harmonization Tripartite Guideline on 
Good Clinical Practice (E6). 
I agree to ensure that the confidential information contained in this document will not be used for 
any purpose other than the evaluation or conduct of the clinical investigation without the prior 
written consent of Isis Pharmaceuticals, Inc. 
 
 
 
 
 
 
Investigator’s Signature 
 
 
 
Investigator’s Name (please print) 
 
Date (DD Month YYYY) 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 5  
Protocol 
 
29 May 2015 
 
5 
TABLE OF CONTENTS 
Page 
PROTOCOL AMENDMENT.......................................................................................................9 
PROTOCOL SYNOPSIS ............................................................................................................10 
STUDY GLOSSARY ...................................................................................................................14 
1. 
OBJECTIVES .....................................................................................................................16 
1.1 
Primary Objective .....................................................................................................16 
1.2 
Secondary Objective .................................................................................................16 
1.3 
Exploratory Objective ...............................................................................................16 
2. 
BACKGROUND AND RATIONALE ..............................................................................16 
2.1 
Spinal Muscular Atrophy ..........................................................................................16 
2.2 
Therapeutic Rationale ...............................................................................................17 
2.3 
ISIS 396443 ..............................................................................................................18 
2.3.1 
Mechanism of Action ...............................................................................18 
2.3.2 
Chemistry .................................................................................................18 
2.3.3 
Preclinical Experience ..............................................................................18 
2.3.4 
Clinical Experience ..................................................................................19 
2.4 
Rationale for Dose and Schedule of Administration ................................................20 
3. 
EXPERIMENTAL PLAN ..................................................................................................21 
3.1 
Study Design .............................................................................................................21 
3.2 
Number of Study Centers..........................................................................................21 
3.3 
Number of Subjects...................................................................................................21 
3.4 
Overall Study Duration and Follow-up.....................................................................21 
3.4.1 
Screening ..................................................................................................22 
3.4.2 
Treatment ..................................................................................................22 
3.4.3 
Post-Treatment Follow-up ........................................................................22 
3.5 
End of Study .............................................................................................................22 
3.6 
Safety Monitoring and Dose Escalation ...................................................................22 
3.7 
Dose Limiting Toxicity .............................................................................................23 
4. 
SUBJECT ENROLLMENT ...............................................................................................23 
4.1 
Screening...................................................................................................................23 
4.2 
Registration ...............................................................................................................23 
4.3 
Replacement of Subjects ...........................................................................................24 
5. 
SUBJECT ELIGIBILITY ..................................................................................................24 
5.1 
Inclusion Criteria ......................................................................................................24 
5.2 
Exclusion Criteria .....................................................................................................25 
6. 
STUDY PROCEDURES ....................................................................................................25 
6.1 
Study Schedule..........................................................................................................25 
6.2 
Study Assessments ....................................................................................................25 
6.2.1 
Collection of CSF .....................................................................................25 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 5  
Protocol 
 
29 May 2015 
 
6 
6.2.2 
Laboratory Analytes .................................................................................26 
6.2.3 
Neurological Examinations ......................................................................26 
6.2.4 
Pharmacokinetics Specimen Collection ...................................................26 
6.2.5 
Children’s Hospital of Philadelphia Infant Test for Neuromuscular 
Disease (CHOP INTEND) .......................................................................26 
6.2.6  
Hammersmith Functional Motor Scale – Expanded ................................27 
6.2.7 
Compound Muscle Action Potential ........................................................27 
6.2.8 
Growth Parameters ...................................................................................27 
6.2.9 
Ventilation Use .........................................................................................27 
6.2.10 
Phone Assessments ...................................................................................27 
6.2.11 
Safety Evaluations ....................................................................................27 
7. 
STUDY DRUG ....................................................................................................................28 
7.1 
Study Drug ................................................................................................................28 
7.1.1 
Two-Vial Configuration (ISIS 396443 and Diluent) Description ............28 
7.1.2 
Ready-To-Use Vial Configuration Description .......................................29 
7.2 
Packaging and Labeling ............................................................................................29 
7.3 
Study Drug Accountability .......................................................................................29 
8. 
TREATMENT OF SUBJECTS .........................................................................................29 
8.1 
Study Drug Administration .......................................................................................29 
8.2 
Other Protocol-Required Drugs ................................................................................30 
8.3 
Other Protocol-Required Procedures ........................................................................31 
8.4 
Treatment Precautions ..............................................................................................31 
8.5 
Safety Monitoring Rules ...........................................................................................31 
8.6 
Stopping Rules ..........................................................................................................31 
8.7 
Adjustment of Dose and/or Treatment Schedule ......................................................31 
8.8 
Discontinuation of Study Treatment .........................................................................31 
8.9 
Withdrawal of Subjects from the Study ....................................................................31 
8.10 
Concomitant Therapy and Procedures ......................................................................32 
8.10.1 
Concomitant Therapy ...............................................................................32 
8.10.2 
Concomitant Procedures ...........................................................................33 
9. 
SERIOUS AND NON-SERIOUS ADVERSE EVENT REPORTING ..........................33 
9.1 
Sponsor Review of Safety Information.....................................................................33 
9.2 
Regulatory Requirements..........................................................................................33 
9.3 
Definitions.................................................................................................................33 
9.3.1 
Adverse Event ..........................................................................................33 
9.3.2 
Adverse Reaction and Suspected Adverse Reaction ................................33 
9.3.3 
Serious Adverse Event (SAE) ..................................................................33 
9.4 
Monitoring and Recording Adverse Events ..............................................................34 
9.4.1 
Serious Adverse Events ............................................................................34 
9.4.2 
Non-Serious Adverse Events ....................................................................35 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 5  
Protocol 
 
29 May 2015 
 
7 
9.4.3 
Evaluation of Adverse Events (Serious and Non-Serious) ......................35 
9.4.3.1 
Relationship to the Study Drug ...................................................35 
9.4.3.2 
Severity .......................................................................................35 
9.4.3.3 
Action Taken with Study Drug ...................................................36 
9.4.3.4 
Treatment Given for Adverse Event ...........................................36 
9.4.3.5 
Outcome of the Adverse Event ...................................................36 
9.5 
Procedures for Handling Special Situations .............................................................36 
9.5.1 
Abnormalities of Laboratory Tests ...........................................................36 
9.5.2 
Prescheduled or Elective Procedures or Routinely Scheduled Treatments37 
9.5.3 
Dosing Errors ...........................................................................................37 
10. STATISTICAL CONSIDERATIONS ..............................................................................37 
10.1 
Study Endpoints ........................................................................................................37 
10.1.1 
Primary Endpoint .....................................................................................37 
10.1.2 
Secondary Endpoint .................................................................................38 
10.1.3 
Exploratory Endpoints ..............................................................................38 
10.2 
Sample Size Considerations ......................................................................................38 
10.3 
Populations ................................................................................................................38 
10.4 
Interim Analysis ........................................................................................................39 
10.5 
Planned Methods of Analysis ...................................................................................39 
10.5.1 
Demographic and Baseline Characteristics ..............................................39 
10.5.2 
Safety and Tolerability .............................................................................39 
10.5.3 
Pharmacokinetic Analysis ........................................................................39 
10.5.4 
Exploratory Biomarker and Efficacy Analysis ........................................39 
11. INVESTIGATOR’S REGULATORY OBLIGATIONS .................................................40 
11.1 
Informed Consent......................................................................................................40 
11.2 
Ethical Conduct of the Study ....................................................................................40 
11.3 
Institutional Review Board/Research Ethics Board ..................................................40 
11.4 
Subject Confidentiality .............................................................................................41 
12. ADMINISTRATIVE AND LEGAL OBLIGATIONS ....................................................41 
12.1 
Protocol Amendments ...............................................................................................41 
12.2 
Study Termination ....................................................................................................41 
12.3 
Study Documentation and Storage ............................................................................41 
12.4 
Study Monitoring ......................................................................................................42 
12.5 
Language ...................................................................................................................43 
12.6 
Compensation for Injury ...........................................................................................43 
13. REFERENCES ....................................................................................................................44 
14. APPENDICES .....................................................................................................................46 
Appendix A 
Schedule of Procedures .....................................................................47 
Appendix B 
Laboratory Analytes .........................................................................52 
Appendix C 
Pharmacokinetic Sampling Schedule................................................54 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 5  
Protocol 
 
29 May 2015 
 
8 
TABLE OF TABLES 
Page 
Table 1 
Study Drug Characteristics ................................................................................... 28 
Table 2 
Study Drug Characteristics:  Ready-To-Use Vial Configuration ......................... 29 
Table 3 
ISIS 396443 Dose Equivalents and Concentration During the ‘Loading’ 
Dosing Treatment Period ...................................................................................... 30 
Table 4 
ISIS 396443 Dose Equivalent and Concentration During the 
‘Maintenance’ Dosing Treatment Period .............................................................. 30 
Table 5 
ISIS 396443 Dose Volume to be Injected ............................................................. 30 
 
 
TABLE OF FIGURES 
Page 
Figure 1 
ASO Therapeutic Approach for Treatment of SMA ............................................. 17 
 
 
 


ISIS 396443-CS3A 
CONFIDENTIAL
Amendment 5
Protocol 
29 May 2015
 
9 
PROTOCOL AMENDMENT 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
5 
Amendment Date: 
29 May 2015 
 
The purpose of this amendment is to (1) extend the ‘maintenance’ dosing treatment period by 
adding 4 doses of 12 mg equivalent of ISIS 396443 given every 4 months (i.e., on 
Study Days 883, 1009, 1135, and 1261); (2) add the Hammersmith Functional Motor 
Scale-Expanded as an outcomes measure for all subjects who have maintained a CHOP INTEND 
total score of ≥ 50 for two consecutive study visits; (3) add a ready-to-use vial configuration of 
the Study Drug; (4) add language on the possibility to perform an interim analysis to support 
potential discussions with regulatory agencies.  The following table provides a summary list of 
changes to the protocol: 
Section 
Description of Change 
Justification 
Protocol Synopsis 
2.4 Rationale for Dose and 
Schedule of Administration 
3. Experimental Plan 
6. Study Procedures 
Appendix A 
Appendix C 
Maintenance dosing extended to add 
4 additional doses of 12 mg equivalent 
of ISIS 396443 given once every 
4 months.  Additional telephone 
contact assessments (approximately 
once every 3 weeks) have been added 
to accommodate the extension of the 
treatment period. 
Due to the severe, life-threatening 
nature of infantile-onset SMA, and 
the encouraging risk-benefit profile 
of ISIS 396443 observed in this 
patient population in early parts of 
the study, the treatment duration is 
being extended (including 
additional study visit and 
telephone contact assessments). 
2.3.4 Clinical Experience 
Update clinical section with respect to 
Phase 2 study status. 
This section was updated to list all 
ongoing studies. 
Protocol Synopsis 
6.2 Study Assessments 
10.1.3 Exploratory Endpoints 
13. References 
Appendix A 
Hammersmith Functional Motor Scale 
Expanded (HFMSE) was added as an 
exploratory endpoint for subjects who 
have maintained a CHOP INTEND 
total score of ≥ 50 for two consecutive 
study visits.  Two (2) references on the 
HFMSE were added. 
The HFMSE is added to allow 
capturing functional improvements 
seen in stronger infants who have 
started to reach the ceiling of the 
CHOP INTEND. 
Protocol Synopsis 
7. Study Drug 
8.1 Study Drug Administration 
Added information on the two types of 
Study Drug supply:  (1) a two-vial 
configuration in which Study Drug is at 
a concentration of 20 mg/mL which is 
diluted prior to use, and (2) a ready-to-
use vial configuration in which the 
Study Drug is at a concentration of 
2.4 mg/mL. 
As the study progresses, the 
transition will be made from the 
two-vial configuration to the ready-
to-use vial configuration for ease 
of administration. 
Protocol Synopsis 
10.4 Interim Analysis 
Added language to allow for an interim 
analysis. 
The possibility to perform an 
interim analysis was added to 
support potential discussions with 
regulatory agencies. 


 
 
 
 
ISIS PHARMACEUTICALS, INC. 
 
 
ISIS 396443-CS3A 
 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Amendment 4 – 19 May 2014 
 
 
 
 
 
Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 


PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 4  
Protocol 
 
19 May 2014 
 
3 
 
ISIS 396443 
Isis Protocol Number ISIS 396443-CS3A 
Protocol Amendment 4 
 
Clinical Phase:  2 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Trial Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 
Phone: + 01 760 931 9200 
Fax: 
+ 01 760 603 2700 
Key Sponsor Contact: 
 
 
Phone: + 
 
Fax: 
+ 
 
 
 
Date: 
19 May 2014 
 
 
Confidentiality Statement 
This document contains confidential information of Isis Pharmaceuticals, Inc. that must not be 
disclosed to anyone other than the recipient study staff and members of the institutional review 
board or authorized regulatory agencies.  This information cannot be used for any purpose other 
than the evaluation or conduct of the clinical investigation without the prior written consent of 
Isis Pharmaceuticals, Inc. 
PPD
PPD
PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 4  
Protocol 
 
19 May 2014 
 
4 
Protocol Signature Page 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
Amendment 4 
Date: 
19 May 2014 
 
I hereby acknowledge that I have read and understand the attached clinical protocol, entitled “A 
Study to Assess the Safety, Tolerability, and Pharmacokinetics of Multiple Doses of ISIS 396443 
Delivered Intrathecally to Patients with Infantile-Onset Spinal Muscular Atrophy”, dated 
19 May 2014, and agree to conduct the study as described herein. 
I agree to comply with the International Conference on Harmonization Tripartite Guideline on 
Good Clinical Practice (E6). 
I agree to ensure that the confidential information contained in this document will not be used for 
any purpose other than the evaluation or conduct of the clinical investigation without the prior 
written consent of Isis Pharmaceuticals, Inc.  
 
 
 
 
 
 
Investigator’s Signature 
 
 
 
Investigator’s Name (please print) 
 
Date (DD Month YYYY) 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 4  
Protocol 
 
19 May 2014 
 
5 
TABLE OF CONTENTS 
Page 
PROTOCOL AMENDMENT.......................................................................................................9 
PROTOCOL SYNOPSIS ............................................................................................................10 
STUDY GLOSSARY ...................................................................................................................14 
1. 
OBJECTIVES .....................................................................................................................16 
1.1 
Primary Objective .....................................................................................................16 
1.2 
Secondary Objective .................................................................................................16 
1.3 
Exploratory Objective ...............................................................................................16 
2. 
BACKGROUND AND RATIONALE ..............................................................................16 
2.1 
Spinal Muscular Atrophy ..........................................................................................16 
2.2 
Therapeutic Rationale ...............................................................................................17 
2.3 
ISIS 396443 ..............................................................................................................18 
2.3.1 
Mechanism of Action ...............................................................................18 
2.3.2 
Chemistry .................................................................................................18 
2.3.3 
Preclinical Experience ..............................................................................18 
2.3.4 
Clinical Experience ..................................................................................19 
2.4 
Rationale for Dose and Schedule of Administration ................................................20 
3. 
EXPERIMENTAL PLAN ..................................................................................................20 
3.1 
Study Design .............................................................................................................20 
3.2 
Number of Study Centers..........................................................................................21 
3.3 
Number of Subjects...................................................................................................21 
3.4 
Overall Study Duration and Follow-up.....................................................................21 
3.4.1 
Screening ..................................................................................................21 
3.4.2 
Treatment ..................................................................................................21 
3.4.3 
Post-Treatment Follow-up ........................................................................22 
3.5 
End of Study .............................................................................................................22 
3.6 
Safety Monitoring and Dose Escalation ...................................................................22 
3.7 
Dose Limiting Toxicity .............................................................................................22 
4. 
SUBJECT ENROLLMENT ...............................................................................................23 
4.1 
Screening...................................................................................................................23 
4.2 
Registration ...............................................................................................................23 
4.3 
Replacement of Subjects ...........................................................................................23 
5. 
SUBJECT ELIGIBILITY ..................................................................................................23 
5.1 
Inclusion Criteria ......................................................................................................23 
5.2 
Exclusion Criteria .....................................................................................................24 
6. 
STUDY PROCEDURES ....................................................................................................25 
6.1 
Study Schedule..........................................................................................................25 
6.2 
Study Assessments ....................................................................................................25 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 4  
Protocol 
 
19 May 2014 
 
6 
6.2.1 
Collection of CSF .....................................................................................25 
6.2.2 
Laboratory Analytes .................................................................................25 
6.2.3 
Neurological Examinations ......................................................................25 
6.2.4 
Pharmacokinetics Specimen Collection ...................................................25 
6.2.5 
Children’s Hospital of Philadelphia Infant Test for Neuromuscular 
Disease (CHOP INTEND) .......................................................................26 
6.2.6 
Compound Muscle Action Potential ........................................................26 
6.2.7 
Growth Parameters ...................................................................................26 
6.2.8 
Ventilation Use .........................................................................................26 
6.2.9 
Phone Assessments ...................................................................................26 
6.2.10 
Safety Evaluations ....................................................................................27 
7. 
STUDY DRUG ....................................................................................................................27 
7.1 
Study Drug and Diluent Description.........................................................................27 
7.2 
Packaging and Labeling ............................................................................................27 
7.3 
Study Drug Accountability .......................................................................................28 
8. 
TREATMENT OF SUBJECTS .........................................................................................28 
8.1 
Study Drug Administration .......................................................................................28 
8.2 
Other Protocol-Required Drugs ................................................................................29 
8.3 
Other Protocol-Required Procedures ........................................................................29 
8.4 
Treatment Precautions ..............................................................................................29 
8.5 
Safety Monitoring Rules ...........................................................................................29 
8.6 
Stopping Rules ..........................................................................................................29 
8.7 
Adjustment of Dose and/or Treatment Schedule ......................................................29 
8.8 
Discontinuation of Study Treatment .........................................................................30 
8.9 
Withdrawal of Subjects from the Study ....................................................................30 
8.10 
Concomitant Therapy and Procedures ......................................................................31 
8.10.1 
Concomitant Therapy ...............................................................................31 
8.10.2 
Concomitant Procedures ...........................................................................31 
9. 
SERIOUS AND NON-SERIOUS ADVERSE EVENT REPORTING ..........................31 
9.1 
Sponsor Review of Safety Information.....................................................................31 
9.2 
Regulatory Requirements..........................................................................................31 
9.3 
Definitions.................................................................................................................32 
9.3.1 
Adverse Event ..........................................................................................32 
9.3.2 
Adverse Reaction and Suspected Adverse Reaction ................................32 
9.3.3 
Serious Adverse Event (SAE) ..................................................................32 
9.4 
Monitoring and Recording Adverse Events ..............................................................33 
9.4.1 
Serious Adverse Events ............................................................................33 
9.4.2 
Non-Serious Adverse Events ....................................................................33 
9.4.3 
Evaluation of Adverse Events (Serious and Non-Serious) ......................33 
9.4.3.1 
Relationship to the Study Drug ...................................................33 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 4  
Protocol 
 
19 May 2014 
 
7 
9.4.3.2 
Severity .......................................................................................34 
9.4.3.3 
Action Taken with Study Drug ...................................................34 
9.4.3.4 
Treatment Given for Adverse Event ...........................................34 
9.4.3.5 
Outcome of the Adverse Event ...................................................34 
9.5 
Procedures for Handling Special Situations .............................................................35 
9.5.1 
Abnormalities of Laboratory Tests ...........................................................35 
9.5.2 
Prescheduled or Elective Procedures or Routinely Scheduled 
Treatments ................................................................................................35 
9.5.3 
Dosing Errors ...........................................................................................36 
10. STATISTICAL CONSIDERATIONS ..............................................................................36 
10.1 
Study Endpoints ........................................................................................................36 
10.1.1 
Primary Endpoint .....................................................................................36 
10.1.2 
Secondary Endpoint .................................................................................36 
10.1.3 
Exploratory Endpoints ..............................................................................37 
10.2 
Sample Size Considerations ......................................................................................37 
10.3 
Populations ................................................................................................................37 
10.4 
Interim Analysis ........................................................................................................37 
10.5 
Planned Methods of Analysis ...................................................................................37 
10.5.1 
Demographic and Baseline Characteristics ..............................................37 
10.5.2 
Safety and Tolerability .............................................................................37 
10.5.3 
Pharmacokinetic Analysis ........................................................................38 
10.5.4 
Exploratory Biomarker and Efficacy Analysis ........................................38 
11. INVESTIGATOR’S REGULATORY OBLIGATIONS .................................................38 
11.1 
Informed Consent......................................................................................................38 
11.2 
Ethical Conduct of the Study ....................................................................................39 
11.3 
Institutional Review Board/Research Ethics Board ..................................................39 
11.4 
Subject Confidentiality .............................................................................................39 
12. ADMINISTRATIVE AND LEGAL OBLIGATIONS ....................................................40 
12.1 
Protocol Amendments ...............................................................................................40 
12.2 
Study Termination ....................................................................................................40 
12.3 
Study Documentation and Storage ............................................................................40 
12.4 
Study Monitoring ......................................................................................................41 
12.5 
Language ...................................................................................................................41 
12.6 
Compensation for Injury ...........................................................................................41 
13. REFERENCES ....................................................................................................................42 
14. APPENDICES .....................................................................................................................44 
Appendix A 
Schedule of Procedures ..............................................................................45 
Appendix B 
Laboratory Analytes...................................................................................49 
Appendix C 
Pharmacokinetic Sampling Schedule .........................................................51 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 4  
Protocol 
 
19 May 2014 
 
8 
TABLE OF TABLES 
Page 
Table 1 
Study Drug Characteristics ................................................................................... 27 
Table 2 
ISIS 396443 Dose Equivalents and Concentration During the ‘Loading’ 
Dosing Treatment Period ...................................................................................... 28 
Table 3 
ISIS 396443 Dose Equivalent and Concentration During the 
‘Maintenance’ Dosing Treatment Period .............................................................. 28 
Table 4 
ISIS 396443 Dose Volume to be Injected ............................................................. 29 
 
 
TABLE OF FIGURES 
Page 
Figure 1 
ASO Therapeutic Approach for Treatment of SMA ............................................. 17 
 
 
 


ISIS 396443-CS3A 
CONFIDENTIAL
Amendment 4
Protocol 
19 May 2014
 
9 
PROTOCOL AMENDMENT 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
4 
Amendment Date: 
19 May 2014 
 
The purpose of this amendment is to extend maintenance dosing with ISIS 396443 by adding 
4 additional doses of 12 mg equivalent given every 4 months (i.e., on Study Days 379, 505, 631, 
and 757).   
The following table provides a summary list of changes to the protocol: 
Section 
Description of Change 
Justification 
Protocol Synopsis,  
3. 
Experimental Plan,  
6.2 
Study Procedures,  
8. 
Treatment of Subjects 
Appendix A,  
Appendix C 
Maintenance dosing extended to add 
4 additional doses of 12 mg equivalent 
of ISIS 396443 given once every 
4 months 
Due to the severe, life-threatening 
nature of infantile-onset SMA, and 
the encouraging risk-benefit profile 
of ISIS 396443 observed in this 
patient population in the early part 
of the study, the treatment 
duration is being extended 
2.3.3 Preclinical Experience  
Detailed results of preclinical studies 
replaced with a summary statement 
and the relevant sections of the 
Investigator Brochure are referenced 
Investigator Brochure has been 
updated with results of the long-
term toxicology study in juvenile 
monkeys 
2.4 
Rationale for Dose and 
Schedule of 
Administration  
Rationale for loading and maintenance 
dose and regimen has been updated  
Dose and dosing regimen 
selection have been justified on 
the basis of available clinical and 
preclinical PK data 
8.1 
Study Drug Administration  Row added for study drug injection 
volume in subjects >24 months of age 
Necessary to accommodate 
extension of the treatment period 
 
 
 


 
 
 
 
ISIS PHARMACEUTICALS, INC. 
 
 
ISIS 396443-CS3A 
 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Amendment 3 – 14 March 2014 
 
 
 
 
 
Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 


PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 3  
Protocol 
 
14 March 2014 
 
3 
 
ISIS 396443 
Isis Protocol Number ISIS 396443-CS3A 
Protocol Amendment 3 
 
Clinical Phase:  2 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Trial Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 
Phone: + 01 760 931 9200 
Fax: 
+ 01 760 603 2700 
Key Sponsor Contact: 
 
 
Phone: + 
 
Fax: 
+ 
 
 
 
Date: 
14 March 2014 
 
 
Confidentiality Statement 
This document contains confidential information of Isis Pharmaceuticals, Inc. that must not be 
disclosed to anyone other than the recipient study staff and members of the institutional review 
board or authorized regulatory agencies.  This information cannot be used for any purpose other 
than the evaluation or conduct of the clinical investigation without the prior written consent of 
Isis Pharmaceuticals, Inc. 
PPD
PPD
PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 3  
Protocol 
 
14 March 2014 
 
4 
Protocol Signature Page 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
Amendment 3 
Date: 
14 March 2014 
 
I hereby acknowledge that I have read and understand the attached clinical protocol, entitled “A 
Study to Assess the Safety, Tolerability, and Pharmacokinetics of Multiple Doses of ISIS 396443 
Delivered Intrathecally to Patients with Infantile-Onset Spinal Muscular Atrophy”, dated 
14March 2014, and agree to conduct the study as described herein. 
I agree to comply with the International Conference on Harmonization Tripartite Guideline on 
Good Clinical Practice (E6). 
I agree to ensure that the confidential information contained in this document will not be used for 
any purpose other than the evaluation or conduct of the clinical investigation without the prior 
written consent of Isis Pharmaceuticals, Inc.  
 
 
 
 
 
 
Investigator’s Signature 
 
 
 
Investigator’s Name (please print) 
 
Date (DD Month YYYY) 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 3  
Protocol 
 
14 March 2014 
 
5 
TABLE OF CONTENTS 
Page 
PROTOCOL AMENDMENT.......................................................................................................9 
PROTOCOL SYNOPSIS ............................................................................................................10 
STUDY GLOSSARY ...................................................................................................................14 
1. 
OBJECTIVES .....................................................................................................................16 
1.1 
Primary Objective .....................................................................................................16 
1.2 
Secondary Objective .................................................................................................16 
1.3 
Exploratory Objective ...............................................................................................16 
2. 
BACKGROUND AND RATIONALE ..............................................................................16 
2.1 
Spinal Muscular Atrophy ..........................................................................................16 
2.2 
Therapeutic Rationale ...............................................................................................17 
2.3 
ISIS 396443 ..............................................................................................................18 
2.3.1 
Mechanism of Action ...............................................................................18 
2.3.2 
Chemistry .................................................................................................18 
2.3.3 
Preclinical Experience ..............................................................................18 
2.3.4 
Clinical Experience ..................................................................................19 
2.4 
Rationale for Dose and Schedule of Administration ................................................20 
3. 
EXPERIMENTAL PLAN ..................................................................................................21 
3.1 
Study Design .............................................................................................................21 
3.2 
Number of Study Centers..........................................................................................22 
3.3 
Number of Subjects...................................................................................................22 
3.4 
Overall Study Duration and Follow-up.....................................................................22 
3.4.1 
Screening ..................................................................................................22 
3.4.2 
Treatment ..................................................................................................22 
3.4.3 
Post-Treatment Follow-up ........................................................................22 
3.5 
End of Study .............................................................................................................22 
3.6 
Safety Monitoring and Dose Escalation ...................................................................23 
3.7 
Dose Limiting Toxicity .............................................................................................23 
4. 
SUBJECT ENROLLMENT ...............................................................................................23 
4.1 
Screening...................................................................................................................23 
4.2 
Registration ...............................................................................................................24 
4.3 
Replacement of Subjects ...........................................................................................24 
5. 
SUBJECT ELIGIBILITY ..................................................................................................24 
5.1 
Inclusion Criteria ......................................................................................................24 
5.2 
Exclusion Criteria .....................................................................................................25 
6. 
STUDY PROCEDURES ....................................................................................................25 
6.1 
Study Schedule..........................................................................................................25 
6.2 
Study Assessments ....................................................................................................26 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 3  
Protocol 
 
14 March 2014 
 
6 
6.2.1 
Collection of CSF .....................................................................................26 
6.2.2 
Laboratory Analytes .................................................................................26 
6.2.3 
Neurological Examinations ......................................................................26 
6.2.4 
Pharmacokinetics Specimen Collection ...................................................26 
6.2.5 
Children’s Hospital of Philadelphia Infant Test for Neuromuscular 
Disease (CHOP INTEND) .......................................................................27 
6.2.6 
Compound Muscle Action Potential ........................................................27 
6.2.7 
Growth Parameters ...................................................................................27 
6.2.8 
Ventilation Use .........................................................................................27 
6.2.9 
Phone Assessments ...................................................................................27 
6.2.10 
Safety Evaluations ....................................................................................27 
7. 
STUDY DRUG ....................................................................................................................28 
7.1 
Study Drug and Diluent Description.........................................................................28 
7.2 
Packaging and Labeling ............................................................................................28 
7.3 
Study Drug Accountability .......................................................................................28 
8. 
TREATMENT OF SUBJECTS .........................................................................................28 
8.1 
Study Drug Administration .......................................................................................28 
8.2 
Other Protocol-Required Drugs ................................................................................30 
8.3 
Other Protocol-Required Procedures ........................................................................30 
8.4 
Treatment Precautions ..............................................................................................30 
8.5 
Safety Monitoring Rules ...........................................................................................30 
8.6 
Stopping Rules ..........................................................................................................30 
8.7 
Adjustment of Dose and/or Treatment Schedule ......................................................30 
8.8 
Discontinuation of Study Treatment .........................................................................30 
8.9 
Withdrawal of Subjects from the Study ....................................................................30 
8.10 
Concomitant Therapy and Procedures ......................................................................31 
8.10.1 
Concomitant Therapy ...............................................................................31 
8.10.2 
Concomitant Procedures ...........................................................................32 
9. 
SERIOUS AND NON-SERIOUS ADVERSE EVENT REPORTING ..........................32 
9.1 
Sponsor Review of Safety Information.....................................................................32 
9.2 
Regulatory Requirements..........................................................................................32 
9.3 
Definitions.................................................................................................................32 
9.3.1 
Adverse Event ..........................................................................................32 
9.3.2 
Adverse Reaction and Suspected Adverse Reaction ................................32 
9.3.3 
Serious Adverse Event (SAE) ..................................................................32 
9.4 
Monitoring and Recording Adverse Events ..............................................................33 
9.4.1 
Serious Adverse Events ............................................................................33 
9.4.2 
Non-Serious Adverse Events ....................................................................34 
9.4.3 
Evaluation of Adverse Events (Serious and Non-Serious) ......................34 
9.4.3.1 
Relationship to the Study Drug ...................................................34 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 3  
Protocol 
 
14 March 2014 
 
7 
9.4.3.2 
Severity .......................................................................................34 
9.4.3.3 
Action Taken with Study Drug ...................................................35 
9.4.3.4 
Treatment Given for Adverse Event ...........................................35 
9.4.3.5 
Outcome of the Adverse Event ...................................................35 
9.5 
Procedures for Handling Special Situations .............................................................35 
9.5.1 
Abnormalities of Laboratory Tests ...........................................................35 
9.5.2 
Prescheduled or Elective Procedures or Routinely Scheduled 
Treatments ................................................................................................36 
9.5.3 
Dosing Errors ...........................................................................................36 
10. STATISTICAL CONSIDERATIONS ..............................................................................36 
10.1 
Study Endpoints ........................................................................................................36 
10.1.1 
Primary Endpoint .....................................................................................36 
10.1.2 
Secondary Endpoint .................................................................................37 
10.1.3 
Exploratory Endpoints ..............................................................................37 
10.2 
Sample Size Considerations ......................................................................................37 
10.3 
Populations ................................................................................................................37 
10.4 
Interim Analysis ........................................................................................................38 
10.5 
Planned Methods of Analysis ...................................................................................38 
10.5.1 
Demographic and Baseline Characteristics ..............................................38 
10.5.2 
Safety and Tolerability .............................................................................38 
10.5.3 
Pharmacokinetic Analysis ........................................................................38 
10.5.4 
Exploratory Biomarker and Efficacy Analysis ........................................38 
11. INVESTIGATOR’S REGULATORY OBLIGATIONS .................................................38 
11.1 
Informed Consent......................................................................................................38 
11.2 
Ethical Conduct of the Study ....................................................................................39 
11.3 
Institutional Review Board/Research Ethics Board ..................................................39 
11.4 
Subject Confidentiality .............................................................................................39 
12. ADMINISTRATIVE AND LEGAL OBLIGATIONS ....................................................40 
12.1 
Protocol Amendments ...............................................................................................40 
12.2 
Study Termination ....................................................................................................40 
12.3 
Study Documentation and Storage ............................................................................40 
12.4 
Study Monitoring ......................................................................................................41 
12.5 
Language ...................................................................................................................41 
12.6 
Compensation for Injury ...........................................................................................42 
13. REFERENCES ....................................................................................................................43 
14. APPENDICES .....................................................................................................................45 
Appendix A 
Schedule of Procedures ...........................................................................46 
Appendix B 
Laboratory Analytes................................................................................50 
Appendix C 
Pharmacokinetic Sampling Schedule ......................................................52 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 3  
Protocol 
 
14 March 2014 
 
8 
 
TABLE OF TABLES 
Page 
Table 1 
Study Drug Characteristics ................................................................................... 28 
Table 2 
ISIS 396443 Dose Equivalents and Concentration During the ‘Loading’ 
Dosing Treatment Period ...................................................................................... 29 
Table 3 
ISIS 396443 Dose Equivalent and Concentration During the 
‘Maintenance’ Dosing Treatment Period .............................................................. 29 
Table 4 
ISIS 396443 Dose Volume to be Injected ............................................................. 29 
 
 
TABLE OF FIGURES 
Page 
Figure 1 
ASO Therapeutic Approach for Treatment of SMA ............................................. 17 
 
 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 3  
Protocol 
 
14 March 2014 
 
9 
PROTOCOL AMENDMENT 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
3 
Amendment Date: 
14 March 2014 
 
The purpose of this amendment is to add permanent ventilation to the combined efficacy 
endpoint.  The definition of permanent ventilation is ≥16 hours ventilation/day continuously for 
>2 weeks in the absence of an acute reversible illness.  This definition is consistent with what 
was done in Natural History Study data in the infantile SMA patient population (PNCR study, 
Finkel et al., unpublished data). 
The following table provides a summary list of changes to the protocol: 
Section 
Description of Change 
Synopsis;  
10.1.3 Exploratory Endpoints 
Changed efficacy endpoint to: Age at death or 
permanent ventilation (defined as ≥16 hours 
ventilation/day continuously for >2 weeks in the 
absence of an acute reversible illness), to be 
consistent with natural history study data in the 
infantile SMA population (PNCR study, Finkel et al.). 
 
 


 
 
 
 
ISIS PHARMACEUTICALS, INC. 
 
 
ISIS 396443-CS3A 
 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Amendment 2 – 18 January 2014 
 
 
 
 
 
Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 


PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 2  
Protocol 
 
18 January 2014 
 
3 
 
ISIS 396443 
Isis Protocol Number ISIS 396443-CS3A 
Protocol Amendment 2 
 
Clinical Phase:  2 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Trial Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 
Phone: + 01 760 931 9200 
Fax: 
+ 01 760 603 2700 
Key Sponsor Contact: 
 
 
Phone: + 
 
Fax: 
+ 
 
 
 
Date: 
18 January 2014 
 
 
Confidentiality Statement 
This document contains confidential information of Isis Pharmaceuticals, Inc. that must not be 
disclosed to anyone other than the recipient study staff and members of the institutional review 
board or authorized regulatory agencies.  This information cannot be used for any purpose other 
than the evaluation or conduct of the clinical investigation without the prior written consent of 
Isis Pharmaceuticals, Inc. 
PPD
PPD
PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 2  
Protocol 
 
18 January 2014 
 
4 
Protocol Signature Page 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
Amendment 2 
Date: 
18 January 2014 
 
I hereby acknowledge that I have read and understand the attached clinical protocol, entitled “A 
Study to Assess the Safety, Tolerability, and Pharmacokinetics of Multiple Doses of ISIS 396443 
Delivered Intrathecally to Patients with Infantile-Onset Spinal Muscular Atrophy”, dated 18 
January 2014, and agree to conduct the study as described herein. 
I agree to comply with the International Conference on Harmonization Tripartite Guideline on 
Good Clinical Practice (E6). 
I agree to ensure that the confidential information contained in this document will not be used for 
any purpose other than the evaluation or conduct of the clinical investigation without the prior 
written consent of Isis Pharmaceuticals, Inc.  
 
 
 
 
 
 
Investigator’s Signature 
 
 
 
Investigator’s Name (please print) 
 
Date (DD Month YYYY) 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 2  
Protocol 
 
18 January 2014 
 
5 
TABLE OF CONTENTS 
Page 
PROTOCOL AMENDMENT.......................................................................................................9 
PROTOCOL SYNOPSIS ............................................................................................................10 
STUDY GLOSSARY ...................................................................................................................14 
1. 
OBJECTIVES .....................................................................................................................16 
1.1 
Primary Objective .....................................................................................................16 
1.2 
Secondary Objective .................................................................................................16 
1.3 
Exploratory Objective ...............................................................................................16 
2. 
BACKGROUND AND RATIONALE ..............................................................................16 
2.1 
Spinal Muscular Atrophy ..........................................................................................16 
2.2 
Therapeutic Rationale ...............................................................................................17 
2.3 
ISIS 396443 ..............................................................................................................18 
2.3.1 
Mechanism of Action ...............................................................................18 
2.3.2 
Chemistry .................................................................................................18 
2.3.3 
Preclinical Experience ..............................................................................18 
2.3.4 
Clinical Experience ..................................................................................19 
2.4 
Rationale for Dose and Schedule of Administration ................................................20 
3. 
EXPERIMENTAL PLAN ..................................................................................................21 
3.1 
Study Design .............................................................................................................21 
3.2 
Number of Study Centers..........................................................................................22 
3.3 
Number of Subjects...................................................................................................22 
3.4 
Overall Study Duration and Follow-up.....................................................................22 
3.4.1 
Screening ..................................................................................................22 
3.4.2 
Treatment ..................................................................................................22 
3.4.3 
Post-Treatment Follow-up ........................................................................22 
3.5 
End of Study .............................................................................................................22 
3.6 
Safety Monitoring and Dose Escalation ...................................................................23 
3.7 
Dose Limiting Toxicity .............................................................................................23 
4. 
SUBJECT ENROLLMENT ...............................................................................................23 
4.1 
Screening...................................................................................................................23 
4.2 
Registration ...............................................................................................................24 
4.3 
Replacement of Subjects ...........................................................................................24 
5. 
SUBJECT ELIGIBILITY ..................................................................................................24 
5.1 
Inclusion Criteria ......................................................................................................24 
5.2 
Exclusion Criteria .....................................................................................................25 
6. 
STUDY PROCEDURES ....................................................................................................25 
6.1 
Study Schedule..........................................................................................................25 
6.2 
Study Assessments ....................................................................................................26 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 2  
Protocol 
 
18 January 2014 
 
6 
6.2.1 
Collection of CSF .....................................................................................26 
6.2.2 
Laboratory Analytes .................................................................................26 
6.2.3 
Neurological Examinations ......................................................................26 
6.2.4 
Pharmacokinetics Specimen Collection ...................................................26 
6.2.5 
Children’s Hospital of Philadelphia Infant Test for Neuromuscular 
Disease (CHOP INTEND) .......................................................................27 
6.2.6 
Compound Muscle Action Potential ........................................................27 
6.2.7 
Growth Parameters ...................................................................................27 
6.2.8 
Ventilation Use .........................................................................................27 
6.2.9 
Phone Assessments ...................................................................................27 
6.2.10 
Safety Evaluations ....................................................................................27 
7. 
STUDY DRUG ....................................................................................................................28 
7.1 
Study Drug and Diluent Description.........................................................................28 
7.2 
Packaging and Labeling ............................................................................................28 
7.3 
Study Drug Accountability .......................................................................................28 
8. 
TREATMENT OF SUBJECTS .........................................................................................28 
8.1 
Study Drug Administration .......................................................................................28 
8.2 
Other Protocol-Required Drugs ................................................................................30 
8.3 
Other Protocol-Required Procedures ........................................................................30 
8.4 
Treatment Precautions ..............................................................................................30 
8.5 
Safety Monitoring Rules ...........................................................................................30 
8.6 
Stopping Rules ..........................................................................................................30 
8.7 
Adjustment of Dose and/or Treatment Schedule ......................................................30 
8.8 
Discontinuation of Study Treatment .........................................................................30 
8.9 
Withdrawal of Subjects from the Study ....................................................................30 
8.10 
Concomitant Therapy and Procedures ......................................................................31 
8.10.1 
Concomitant Therapy ...............................................................................31 
8.10.2 
Concomitant Procedures ...........................................................................32 
9. 
SERIOUS AND NON-SERIOUS ADVERSE EVENT REPORTING ..........................32 
9.1 
Sponsor Review of Safety Information.....................................................................32 
9.2 
Regulatory Requirements..........................................................................................32 
9.3 
Definitions.................................................................................................................32 
9.3.1 
Adverse Event ..........................................................................................32 
9.3.2 
Adverse Reaction and Suspected Adverse Reaction ................................32 
9.3.3 
Serious Adverse Event (SAE) ..................................................................32 
9.4 
Monitoring and Recording Adverse Events ..............................................................33 
9.4.1 
Serious Adverse Events ............................................................................33 
9.4.2 
Non-Serious Adverse Events ....................................................................34 
9.4.3 
Evaluation of Adverse Events (Serious and Non-Serious) ......................34 
9.4.3.1 
Relationship to the Study Drug ...................................................34 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 2  
Protocol 
 
18 January 2014 
 
7 
9.4.3.2 
Severity .......................................................................................34 
9.4.3.3 
Action Taken with Study Drug ...................................................35 
9.4.3.4 
Treatment Given for Adverse Event ...........................................35 
9.4.3.5 
Outcome of the Adverse Event ...................................................35 
9.5 
Procedures for Handling Special Situations .............................................................35 
9.5.1 
Abnormalities of Laboratory Tests ...........................................................35 
9.5.2 
Prescheduled or Elective Procedures or Routinely Scheduled 
Treatments ................................................................................................36 
9.5.3 
Dosing Errors ...........................................................................................36 
10. STATISTICAL CONSIDERATIONS ..............................................................................36 
10.1 
Study Endpoints ........................................................................................................36 
10.1.1 
Primary Endpoint .....................................................................................36 
10.1.2 
Secondary Endpoint .................................................................................37 
10.1.3 
Exploratory Endpoints ..............................................................................37 
10.2 
Sample Size Considerations ......................................................................................37 
10.3 
Populations ................................................................................................................37 
10.4 
Interim Analysis ........................................................................................................38 
10.5 
Planned Methods of Analysis ...................................................................................38 
10.5.1 
Demographic and Baseline Characteristics ..............................................38 
10.5.2 
Safety and Tolerability .............................................................................38 
10.5.3 
Pharmacokinetic Analysis ........................................................................38 
10.5.4 
Exploratory Biomarker and Efficacy Analysis ........................................38 
11. INVESTIGATOR’S REGULATORY OBLIGATIONS .................................................38 
11.1 
Informed Consent......................................................................................................38 
11.2 
Ethical Conduct of the Study ....................................................................................39 
11.3 
Institutional Review Board/Research Ethics Board ..................................................39 
11.4 
Subject Confidentiality .............................................................................................39 
12. ADMINISTRATIVE AND LEGAL OBLIGATIONS ....................................................40 
12.1 
Protocol Amendments ...............................................................................................40 
12.2 
Study Termination ....................................................................................................40 
12.3 
Study Documentation and Storage ............................................................................40 
12.4 
Study Monitoring ......................................................................................................41 
12.5 
Language ...................................................................................................................41 
12.6 
Compensation for Injury ...........................................................................................42 
13. REFERENCES ....................................................................................................................43 
14. APPENDICES .....................................................................................................................45 
Appendix A 
Schedule of Procedures ...........................................................................46 
Appendix B 
Laboratory Analytes................................................................................50 
Appendix C 
Pharmacokinetic Sampling Schedule ......................................................52 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 2  
Protocol 
 
18 January 2014 
 
8 
 
TABLE OF TABLES 
Page 
Table 1 
Study Drug Characteristics ................................................................................... 28 
Table 2 
ISIS 396443 Dose Equivalents and Concentration During the ‘Loading’ 
Dosing Treatment Period ...................................................................................... 29 
Table 3 
ISIS 396443 Dose Equivalent and Concentration During the 
‘Maintenance’ Dosing Treatment Period .............................................................. 29 
Table 4 
ISIS 396443 Dose Volume to be Injected ............................................................. 29 
 
 
TABLE OF FIGURES 
Page 
Figure 1 
ASO Therapeutic Approach for Treatment of SMA ............................................. 17 
 
 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 2  
Protocol 
 
18 January 2014 
 
9 
PROTOCOL AMENDMENT 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
2 
Amendment Date: 
18 January 2014 
 
The purpose of this amendment is to extend the study to add one additional ‘maintenance’ dose 
of 12 mg equivalent ISIS 396443 on Study Day 253, approximately 6 months after the subjects 
have received their Day 85 dose.  A few typographical errors were also corrected for 
consistency. 
The following table provides a summary list of changes to the protocol: 
Section 
Description of Change 
PROTOCOL SYNOPSIS, Study Design and Methodology 
Added ‘maintenance’ dose treatment and follow-up 
to the study design 
2.4 Rationale for Dose and Schedule of Administration 
Added dose rationale for the single ‘maintenance’ 
dose 
3. EXPERIMENTAL PLAN 
Added ‘maintenance’ dose treatment and follow-up 
to the study design 
6.2 Study Assessments 
Added ‘maintenance’ dose treatment and follow-up 
to the study assessments 
8.1 Study Drug Administration 
Added study drug administration instructions for the 
‘maintenance’ dose 
8.7 Adjustment of Dose and/or Treatment Schedule 
Added instructions for the ‘maintenance’ dose 
APPENDIX A  Schedule of Procedures 
Added ‘maintenance’ dose treatment and follow-up 
to the schedule of procedures 
APPENDIX C  Pharmacokinetic Schedule 
Added pharmacokinetic sample schedule for the 
‘maintenance’ dose period 
 
 


 
 
 
 
ISIS PHARMACEUTICALS, INC. 
 
 
ISIS 396443-CS3A 
 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Amendment 1 – 14 August 2013 
 
 
 
 
 
Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 


PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 1  
Protocol 
 
14 August 2013 
 
3 
 
ISIS 396443 
Isis Protocol Number ISIS 396443-CS3A 
Protocol Amendment 1 
 
Clinical Phase:  2 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients 
with Infantile-Onset Spinal Muscular Atrophy 
 
Trial Sponsor: 
Isis Pharmaceuticals, Inc. 
2855 Gazelle Court 
Carlsbad, CA  92010 
Phone: + 01 760 931 9200 
Fax: 
+ 01 760 603 2700 
Key Sponsor Contact: 
 
 
Phone: +
 
Fax: 
+
 
 
 
Date: 
14 August 2013 
 
 
Confidentiality Statement 
This document contains confidential information of Isis Pharmaceuticals, Inc. that must not be 
disclosed to anyone other than the recipient study staff and members of the institutional review 
board or authorized regulatory agencies.  This information cannot be used for any purpose other 
than the evaluation or conduct of the clinical investigation without the prior written consent of 
Isis Pharmaceuticals, Inc. 
PPD
PPD
PPD


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 1  
Protocol 
 
14 August 2013 
 
4 
Protocol Signature Page 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
Amendment 1 
Date: 
14 August 2013 
 
I hereby acknowledge that I have read and understand the attached clinical protocol, entitled “A 
Study to Assess the Safety, Tolerability, and Pharmacokinetics of Multiple Doses of ISIS 396443 
Delivered Intrathecally to Patients with Infantile-Onset Spinal Muscular Atrophy”, dated 
14 August 2013, and agree to conduct the study as described herein. 
I agree to comply with the International Conference on Harmonization Tripartite Guideline on 
Good Clinical Practice (E6). 
I agree to ensure that the confidential information contained in this document will not be used for 
any purpose other than the evaluation or conduct of the clinical investigation without the prior 
written consent of Isis Pharmaceuticals, Inc.  
 
 
 
 
 
 
Investigator’s Signature 
 
 
 
Investigator’s Name (please print) 
 
Date (DD Month YYYY) 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 1  
Protocol 
 
14 August 2013 
 
5 
TABLE OF CONTENTS 
Page 
PROTOCOL AMENDMENT.......................................................................................................9 
PROTOCOL SYNOPSIS ............................................................................................................10 
STUDY GLOSSARY ...................................................................................................................14 
1. 
OBJECTIVES .....................................................................................................................16 
1.1 
Primary Objective .....................................................................................................16 
1.2 
Secondary Objective .................................................................................................16 
1.3 
Exploratory Objective ...............................................................................................16 
2. 
BACKGROUND AND RATIONALE ..............................................................................16 
2.1 
Spinal Muscular Atrophy ..........................................................................................16 
2.2 
Therapeutic Rationale ...............................................................................................17 
2.3 
ISIS 396443 ..............................................................................................................18 
2.3.1 
Mechanism of Action ...............................................................................18 
2.3.2 
Chemistry .................................................................................................18 
2.3.3 
Preclinical Experience ..............................................................................18 
2.3.4 
Clinical Experience ..................................................................................19 
2.4 
Rationale for Dose and Schedule of Administration ................................................20 
3. 
EXPERIMENTAL PLAN ..................................................................................................21 
3.1 
Study Design .............................................................................................................21 
3.2 
Number of Study Centers..........................................................................................22 
3.3 
Number of Subjects...................................................................................................22 
3.4 
Overall Study Duration and Follow-up.....................................................................22 
3.4.1 
Screening ..................................................................................................22 
3.4.2 
Treatment ..................................................................................................22 
3.4.3 
Post-Treatment Follow-up ........................................................................22 
3.5 
End of Study .............................................................................................................22 
3.6 
Safety Monitoring and Dose Escalation ...................................................................22 
3.7 
Dose Limiting Toxicity .............................................................................................23 
4. 
SUBJECT ENROLLMENT ...............................................................................................23 
4.1 
Screening...................................................................................................................23 
4.2 
Registration ...............................................................................................................24 
4.3 
Replacement of Subjects ...........................................................................................24 
5. 
SUBJECT ELIGIBILITY ..................................................................................................24 
5.1 
Inclusion Criteria ......................................................................................................24 
5.2 
Exclusion Criteria .....................................................................................................25 
6. 
STUDY PROCEDURES ....................................................................................................25 
6.1 
Study Schedule..........................................................................................................25 
6.2 
Study Assessments ....................................................................................................25 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 1  
Protocol 
 
14 August 2013 
 
6 
6.2.1 
Collection of CSF .....................................................................................25 
6.2.2 
Laboratory Analytes .................................................................................26 
6.2.3 
Neurological Examinations ......................................................................26 
6.2.4 
Pharmacokinetics Specimen Collection ...................................................26 
6.2.5 
Children’s Hospital of Philadelphia Infant Test for Neuromuscular 
Disease (CHOP INTEND) ................................................................26 
6.2.6 
Compound Muscle Action Potential ........................................................27 
6.2.7 
Growth Parameters ...................................................................................27 
6.2.8 
Ventilation Use .........................................................................................27 
6.2.9 
Phone Assessments ...................................................................................27 
6.2.10 
Safety Evaluations ....................................................................................27 
7. 
STUDY DRUG ....................................................................................................................28 
7.1 
Study Drug and Diluent Description.........................................................................28 
7.2 
Packaging and Labeling ............................................................................................28 
7.3 
Study Drug Accountability .......................................................................................28 
8. 
TREATMENT OF SUBJECTS .........................................................................................28 
8.1 
Study Drug Administration .......................................................................................28 
8.2 
Other Protocol-Required Drugs ................................................................................29 
8.3 
Other Protocol-Required Procedures ........................................................................29 
8.4 
Treatment Precautions ..............................................................................................29 
8.5 
Safety Monitoring Rules ...........................................................................................29 
8.6 
Stopping Rules ..........................................................................................................29 
8.7 
Adjustment of Dose and/or Treatment Schedule ......................................................29 
8.8 
Discontinuation of Study Treatment .........................................................................30 
8.9 
Withdrawal of Subjects from the Study ....................................................................30 
8.10 
Concomitant Therapy and Procedures ......................................................................31 
8.10.1 
Concomitant Therapy ...............................................................................31 
8.10.2 
Concomitant Procedures ...........................................................................31 
9. 
SERIOUS AND NON-SERIOUS ADVERSE EVENT REPORTING ..........................31 
9.1 
Sponsor Review of Safety Information.....................................................................31 
9.2 
Regulatory Requirements..........................................................................................31 
9.3 
Definitions.................................................................................................................32 
9.3.1 
Adverse Event ..........................................................................................32 
9.3.2 
Adverse Reaction and Suspected Adverse Reaction ................................32 
9.3.3 
Serious Adverse Event (SAE) ..................................................................32 
9.4 
Monitoring and Recording Adverse Events ..............................................................33 
9.4.1 
Serious Adverse Events ............................................................................33 
9.4.2 
Non-Serious Adverse Events ....................................................................33 
9.4.3 
Evaluation of Adverse Events (Serious and Non-Serious) ......................33 
9.4.3.1 
Relationship to the Study Drug ...................................................33 


ISIS 396443-CS3A 
CONFIDENTIAL
Amendment 1 
Protocol 
14 August 2013
 
7 
9.4.3.2 
Severity .......................................................................................34 
9.4.3.3 
Action Taken with Study Drug ...................................................34 
9.4.3.4 
Treatment Given for Adverse Event ...........................................34 
9.4.3.5 
Outcome of the Adverse Event ...................................................34 
9.5 
Procedures for Handling Special Situations .............................................................35 
9.5.1 
Abnormalities of Laboratory Tests ...........................................................35 
9.5.2 
Prescheduled or Elective Procedures or Routinely Scheduled  
Treatments ................................................................................................35 
9.5.3 
Dosing Errors ...........................................................................................36 
10.  STATISTICAL CONSIDERATIONS ..............................................................................36 
10.1 
Study Endpoints ........................................................................................................36 
10.1.1 
Primary Endpoint .....................................................................................36 
10.1.2 
Secondary Endpoint .................................................................................36 
10.1.3 
Exploratory Endpoints ..............................................................................36 
10.2 
Sample Size Considerations ......................................................................................37 
10.3 
Populations ................................................................................................................37 
10.4 
Interim Analysis ........................................................................................................37 
10.5 
Planned Methods of Analysis ...................................................................................37 
10.5.1 
Demographic and Baseline Characteristics ..............................................37 
10.5.2 
Safety and Tolerability .............................................................................37 
10.5.3 
Pharmacokinetic Analysis ........................................................................38 
10.5.4 
Exploratory Biomarker and Efficacy Analysis ........................................38 
11.  INVESTIGATOR’S REGULATORY OBLIGATIONS .................................................38 
11.1 
Informed Consent......................................................................................................38 
11.2 
Ethical Conduct of the Study ....................................................................................38 
11.3 
Institutional Review Board/Research Ethics Board ..................................................38 
11.4 
Subject Confidentiality .............................................................................................39 
12.  ADMINISTRATIVE AND LEGAL OBLIGATIONS ....................................................39 
12.1 
Protocol Amendments ...............................................................................................39 
12.2 
Study Termination ....................................................................................................39 
12.3 
Study Documentation and Storage............................................................................40 
12.4 
Study Monitoring ......................................................................................................40 
12.5 
Language ...................................................................................................................41 
12.6 
Compensation for Injury ...........................................................................................41 
13.  REFERENCES ....................................................................................................................42 
14.  APPENDICES .....................................................................................................................44 
Appendix A 
Schedule of Procedures ..............................................................................45 
Appendix B 
Laboratory Analytes ...................................................................................48 
Appendix C 
Pharmacokinetic Sampling Schedule .........................................................50 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 1  
Protocol 
 
14 August 2013 
 
8 
TABLE OF TABLES 
Page 
Table 1 
Study Drug Characteristics ................................................................................... 28 
Table 2 
ISIS 396443 Dose Equivalents and Concentration ............................................... 29 
Table 3 
ISIS 396443 Dose Volume to be Injected ............................................................. 29 
 
 
TABLE OF FIGURES 
Page 
Figure 1 
ASO Therapeutic Approach for Treatment of SMA ............................................. 17 
 
 
 


ISIS 396443-CS3A 
CONFIDENTIAL 
Amendment 1  
Protocol 
 
14 August 2013 
 
9 
PROTOCOL AMENDMENT 
 
Protocol Number: 
ISIS 396443-CS3A 
Protocol Title: 
A Study to Assess the Safety, Tolerability, and Pharmacokinetics of 
Multiple Doses of ISIS 396443 Delivered Intrathecally to Patients with 
Infantile-Onset Spinal Muscular Atrophy 
Amendment: 
1 
Amendment Date: 
14 August 2013 
 
The purpose of this amendment is to: 1) change the dose from 9 to 12 mg ISIS 396443; 2) add 
additional subjects to the study, such that Cohort 2 now may enroll between 4 to approximately 
16 subjects; 3) add age at death and age at death or ≥16 hours ventilation/day for >2 weeks as 
exploratory endpoints; and 4) update the clinical experience section, specifically regarding the 
ISIS 396443-CS2 and ISIS 396443-CS12 studies. 
The following table provides a summary list of changes to the protocol: 
Section 
Description of Change 
2.3.4 Clinical Experience 
Updated information regarding ISIS 396443-CS2 and 
ISIS 396443-CS12 studies 
2.4 Rationale for Dose and Schedule of 
Administration 
Changed “9 mg ISIS 396443” to “12 mg ISIS 396443” 
throughout the section and added rationale for 12 mg 
dose  
3.3 Number of Subjects 
For Cohort 2 changed “4” to “4 to approximately 16” 
subjects to allow additional subjects to enroll in this study 
prior to when the anticipated Phase 3 study in Infantile-
onset SMA commences 
8.1 Study Drug Administration 
Updated dose and concentration in Table 2 and Table 3 
for 12 mg dosing 
10.2 Sample Size Considerations 
Changed “sample size of 8” to “sample size of 
approximately 20” 
10.1.3 
Exploratory Endpoints 
Added age at death and age at death or ≥16 hours 
ventilation/day for >2 weeks as exploratory endpoints 
 
 

SAP_001.pdf: 
Ionis Pharmaceuticals, Inc.
Statistical Analysis Plan
CONFIDENTIAL
February 10,2016
ISIS 396443-CS3A
loNls-
PHARMACEUTICALS
Statistical Analysis Plan
rsts 396443-GS3A
A Study to Assess the Efficacy, Safetyo Tolerability, and
Pharmacokinetics of Muttiple Doses of Nusinersen (ISIS 396443)
Delivered Intrathecally to Patients with Infantile-Onset Spinal
Muscular Atrophy
Date: 2110120',6
Version: 4.0
CoNFIDENTIAL
ConfidentiallTrade Secret information subject to 18-USC-1905 and to which all claims of privilege and
confidentiality are asserted in both statutory and common law. Further dissemination may be made
only with fhe express written permission of /sis Pharmaceuticals, lnc.


Statistical Analysis Plan Signature Page
Ionis Pharmaceuticals, Inc.
2855 Gazelle Court
Carlsbad, CA 92010
Compound Name: 396443
Date:
acology
Date:
Protocol
Study Title:
Issue Date:
CS3A
A Study to Assess the Efficacy, Safety, Tolerability, and
Pharmacokinetics of Multiple Doses of Nusinersen (ISIS 396443)
Delivered Intrathecally to Patients with Infantile-Onset Spinal
Muscular Atrophy
25 JAN 2016 (Amendment 6)
Date:
(D
7'0
a
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to7eltu b
Zftc> ,/z- / 6
2 (o h/ó
1
Date:
PPD


5
5
6
6
6
6
6
6
7
1.
1.1
1.2
1.3
Table of Contents
I ntro duction ...............
Study Overview
Objectives
1.2.2 SecondaryObjectivse
Endpoints
1.3.1 Primary Endpoint
1.3.2 SecondaryEndpoints..
1.3.3 ExploratoryEndpoints
2 Procedures
2.1 
GeneralOverviewofProcedures..............
2.2 
Randomization & Treatment Allocation.
2.3 
Conduct....
2.4 
AE Reporting.............
2.5 
Data Management......
2.5.1 Case Report Form Data
2.5.2 Laboratory Data
1.2.1 PrimaryObjective
2.5.3 Pharmacokinetics Data.....
7
7
7
7
8
I
8
8
8
3 
Analytìcøl P1an...,....,.
3.1 
General Overview of Analyses
3.1.1 StatisticalMethods.......
3.1.2 Subject Population Analyzed
3.1.3 Sample Size Consideration...
9
9
9
9
3.1 .4 
Safety Review and Planned Interim Analysis
3.1.5 Incomplete or Missing Data
3.1.5.1 MotorMilestones.
3.1.5.2 CHOP-INTEND Total Score
3.1.5.3 Ventilator Use
3.2 
Primary Efficacy Analysis.... 
................12
3.3 
Secondary Efficacy Analysis 
................15
3.3.1 Event-freeSurvival
3.3.2 CHOP-INTEND Infant Motor Function Scale.......
16
3.3.3 CMAP Ulnar and Peroneal Nerves
16
3.4 
Exploratory Analysis 
.........17
3.4.1 Measures of Respiratory Status
t7
18
t9
t9
t9
20
20
20
2l
3.4.2 GrowthParameters
3.4.3 Hammersmith Functional Motor Scale - Expanded..
3.5 
Pharmacokinetic Analyses..............
10
10
11
11
11
t2
3.5.1 CSF Concentration Data..
3.5.2 PlasmaPharmacokinetics
3.5.2.1 PlasmaConcentrationData.
3.5.2.2 PlasmaPharmacokineticParameters
3.5.2.3 ImmunogenicityAnalyses
2


3.6
Safety Analyses...
3.6.1 AdverseEvents..........
3.6.1.1 Adverse Events over Time....
3.6.1.2 Adverse events following Dosing...
3.6.2 LaboratoryMeasurements
3.6.5 LeadElectrocardiograms(ECG)
3.6.6 ConcomitantMedications
3.6.7 Hammersmith Infant Neurological Examinations - Sections 1 and 3
22
22
24
24
24
25
26
26
26
26
3


Primarv Rationale for Amendment:
The following major modifïcations have been made to 396443 CS3A SAP, version 3.0, dated
September 25,2015
List of Modifications
Section
Title
Ghange/Rationale
1.2
Objectives
Updated to reflect the changes in protocol amendment 6
1.3
Endpoints
Updated to reflect the changes in protocol amendment 6
3.L.2
Subject
Analyzed
Population
Added 2 SMN2 Copy Set/To provide additional analysis for the
subset of subjects who have 2 copies of the SMN2 gene
3.1.5
lncomplete or Missing
Data
Added missing value imputation method for motor milestone
Changed missing value imputation method for CHOP-INTEND
assessment/To apply the same imputation method to both
primarv endpoint and CHOP-INTEND assessment
3.2
Primary Efficacy Ana lysis
New section/To provide details of the primary efficacy analysis
3.3
Secondary
Analysis
Efficacy
New section/To provide details ofthe secondary efficacy
analyses
3.6.1
Adverse Events
Added definition for lumber puncture related treatment
emergent adverse events and added information about the
linked ae records/Clarification
3.6.1.1
Adverse Events over Time
New section
3.6.!.2
Adverse events following
Dosing
New section
3.6.5
Lead Electrocardiograms
(EcG)
Added outlier analysis
4


L. [t\tRoDUCTIoN
This document provides a description of the study organization, study procedures, and the
plan for the statistical analysis of the study data. Section I discusses study design, objectives,
and endpoints; Section 2 provides the study procedures; Section 3 provides the detailed plan
for the statistical analyses.
As with any statistical analysis plan (SAP), the proposed methods and approaches to the data
analysis should be viewed as flexible. The statistical analysis to some degree is iterative since
so much of the planning is based on statistical and other assumptions, which require
verification.
1.1 Study Overview
This is a Phase 2, open label study conducted at multiple centers in the United States and
Canada. Approximately 8-20 subjects will be enrolled into this multiple-dose study of ISIS
396443. The number of subjects may be higher if some subjects must be replaced and/or if the
sizes of some cohorts are expanded in order to obtain further experience with some dose levels.
This study will test the clinical efficacy, safety, tolerability, and pharmacokinetics of multiple
doses of ISIS 396443 administered as intrathecal (IT) injections by lumbar puncture (LP).
Two 'loading' dose levels (scaled by infant age to be equivalent to 2 year old doses of 6 mg
or 12 mg, based on CSF volume) will be evaluated sequentially. The initial dose level of 6
mg will be studied in a cohort of 4 subjects. The 12 mg dose level will be studied in 4 to
approximately 16 subjects. Following this, all subjects will receive 'maintenance' dosing of
12mgequivalent ISIS 396443 on Days 253,379,505,631,757,883, 1009, ll35,andl26L
Cohort 1 (n:4): 'Loading' dosing of 6 mg equivalent ISIS 396443 on Days l, 15,85, IT
injection; 'maintenance' dosing of 12 mg equivalent on Days 253,379,505, 631, 151,883,
1009, 1135, and 126I,lT injection
Cohort 2(n:4-16):'Loading'dosing of 12mgequivalentlSlS 396443 onDays 1, 15,85, IT
injection, 'maintenance' dosing of 12 mg equivalent on Days 253, 379, 505, 63I, l5l, 883,
1009, 1135, and 1261.,1T injection
After parental informed consent is obtained, subjects will undergo a Screening evaluation no
greater than 2l days prior to first dose administration at which their eligibility for the study
will be examined. Subjects who meet the eligibility criteria will be admitted to the study
center on Study Day l, undergo pre-dose evaluations, and then receive an LP injection of
study drug (ISIS 396443). Following the LP injection on Day 1, subjects will remain at the
study center for at least 24 hours post injection for safety monitoring. Subjects will return to
the study center on Days 15, 85, 253,379,505, 63I,757,883, 1009, 1135, and 126l for
follow-up evaluations of clinical efficacy and safety and subsequent injections. Following LP
injections on Study Days 15, 85,253,379, 505, 631,757,883, 1009, 1 135, and 1261 subjects
5


will not stay overnight in the hospital but will be monitored for at least 6 hours post-injection
before leaving the study facility. A CSF sample will be taken pre-dose on each injection day
for safety and Pharmacokinetic (PK) analyses. During the treatment period, the study center
will monitor the subject's condition through safety monitoring visits on Study Days 16,29,
86, 92, 169, 254, 337, 442, 568, 694, 820, 946, 1072, and I 198, and by telephone contact on
Study Days 8, 43, 57, 71, 106, l2l, I34, 148, 189, I97, 218, 239, 274, 295,316, 358, 380,
400, 421, 463, 484, 506, 526, 541, 589, 610, 632,652, 673,715,736,758,778,799,841,862,
984,904,925,967,988, 1010, 1030, 1051, 1093, lll4,1136,1156, 1177,1219, 1240, 1262
(i.e., approximately every 3 weeks). During the post-treatment follow-up period, the study
center will monitor the subject's condition through telephone contact on Study Days 1282,
1303, and 1324 (i.e., every 3 weeks through 13 weeks after the last dose of ISIS 396443).
Safety monitoring visit will occur on Day 1352 (through 13 weeks after the last dose of ISIS
396443).If a subject terminates early from the study, they will be encouraged to complete all
assessments per the Day 1352 visit.
1.2 Objectives
1.2.1 Primøry Objective
To examine the clinical efficacy of multiple doses of ISIS 396443 administered intrathecally
to patients with tnfantile-Onset SMA.
1.2.2 Secondøry Objectivse
To examine the safety and tolerability of multiple doses of ISIS 396443 administered
intrathecally to patients with Infantile-Onset SMA'
To examine the cerebral spinal fluid (CSF) and plasma PK of multiple doses of ISIS 396443
administered intrathecally to patients with infantile-onset SMA.
1.3 Endpoints
1.3.1 Prímøry Endpoint
The primary endpoint is the proportion of subjects who achieve improvement in motor
milestones as evaluated by Module 2 of the Hammersmith Infant Neurological Examination.
Improvement is def,rned as achievement of at least one of the following:
(Ð 
at least a2-pointincrease from baseline or achievement of pincer grasp in the
category ofvoluntary grasp, or
(iÐ 
at least a2-pointincrease in the ability to kick or achievement of touching
toes, or
(iiÐ a l-point increase in any of the remaining 6 categories: head control, rolling,
sitting, crawling, standing, or walking.
1.3.2 SecondaryEndpoints
Secondary efficacy endpoints include :
o Event-free survival determined by the proportion of subjects who are alive and do not
require permanent ventilatory support (defined as tracheostomy or the need for >16
6


hours ventilation/day continuously for at least 2 weeks in the absence of an acute
reversible illness)
. Improvement in muscle strength as measured by the Children's Hospital of
Philadelphia tnfant Test for Neuromuscular Disease (CHOP INTEND)
o Improvement in neuromuscular electrophysiology measured by the Compound
Muscle Action Potential (CMAP) of the ulnar and peroneal nerves
The safety and tolerability of multiple doses of ISIS 396443 as assessed by:
. Adverse events
o Neurologicalexaminations
o Vital signs
o Physical examinations and weight
, 
. Clinical laboratory tests (serum chemistry, hematology, urinalysis)
o CSF laboratory tests (cell count, protein, glucose)
o ECGs
o Use of concomitant medications
Pharmacokinetic Measures
o CSF and Plasma pharmacokinetics of ISIS 396443
1.3.3 Explorøtory Endpoìnts
The following efficacy evaluations will be performed as exploratory endpoints
Measures of respiratory status (number of respiratory events,, respiratory
infections, respiratory-related hospitalizations, ventilator use and 02 saturation
awake)
Growth parameters (weight for agellength, head circumference, chest
circumference, head to chest circumference ratio, arm circumference)
2 PnoCnDURES
2.1 General Overview of Procedures
Ionis will review all study data including source documents, CRFs, and laboratory reports.
Study site will enter subject source data into the case report form.
2.2 Randomization & Treatment Allocation
This is an open label study. Subjects won't be randomized. Subjects will be registered for the
trial before they begin dosing.
2.3 Conduct
The study will be conducted in accordance with current Good Clinical Practice (GCP) and
International Conference on Harmonization (ICH) guidelines, the V/orld Medical Association
Declaration of Helsinki guidelines, the Food and Drug Administration (FDA) Code of
Federal Regulations, and all other local regulatory requirements.
o
l


2.4 AE Reporting
Ionis Pharmaceuticals, Inc. is responsible for processing all reported AEs. All serious
adverse events, reported to lonis Pharmaceuticals, Inc. (or designee), will be reviewed
according to Ionis standard operating procedures. The Sponsor Medical Monitor and Drug
Safety Physician will review all AEs and SAEs on an ongoing basis throughout the study.
Ionis Pharmaceuticals, Inc. (or designee) will prepare and submit safety reports to the health
authorities. If it becomes necessary to communicate new safety information, Ionis
Pharmaceuticals, Inc. (or designee) will also prepare a safety notification letter and transmit
it to study sites.
2,5 Data Management
2.5J Case Report Form Datø
BioClinica (or designee) is responsible for creating the Electronic Data Capture (EDC) data
entry screens, database and edit checks using definitions developed by Ionis Pharmaceuticals,
Inc. Ionis Pharmaceuticals, Inc. is responsible for the review, data management querying and
locking of the database.
Data are single-entered into the EDC system by the investigator site staff. Programmed edit
checks (computer logic that checks the validity of the data entered and also prompts for
missing datathat is expected to be entered) are run and automatic queries are generated. Ionis
Pharmaceuticals, Inc. reviews all data for accuracy and validity and generates additional
queries in the EDC system when necessary. The data are corrected or an explanation
conceming the query is provided in the EDC system. After all data are entered, source data
verified, reviewed (by Data Management and Clinical Development) and queried, and all
queries resolved, the database is locked
2.5.2 Løborøtory Døta
The safety laboratory analysis including chemistry, hematology, urinalysis, coagulation, and
CSF safety lab data (CSF V/BC, CSF RBC, CSF Protein, CSF Glucose) will be performed at
each site's local lab and will be entered on a CRF. These data will be electronically
transferred to Ionis Pharmaceuticals, Inc. and stored as SAS data files along with the SAS
data files for all other CRFs.
2.5.3 Pharmacokinetícs Døtø
Ionis Pharmaceuticals, Inc. is responsible for the management and review of the plasma and
CSF drug concentration data. Final QA'd data will be stored in Documentum.
8


3 Ax¿,r-yrICAL Pr,¡.x
3.1 General Overview of Analyses
3.1.1 Støtístical Methods
Simple descriptive summary statistics, such as n, mean, standard deviation, standard elror,
median, interquartile range (25th percentile, 75th percentile), minimum, and maximum for
continuous variables, and counts and percentages for categorical variables will be used to
summarize most data. Where appropriate, p-values will be reported. Null hypotheses will be
tested using two-sided tests whose Type I error rates are controlled at alpha: 0.05 unless
otherwise stated. Since the primary analysis and secondary analyses are not comparative
analysis, adjustments for multiplicity of testing will not be used.
Baseline is defined as the last non-missing value prior to the first dose for ISIS 396443 CS3A
Age (months) at death or permanent ventilation will be calculated using the following SAS
code:
yrdif(date 1 date2,' ACT I ACT')* 12
Subject demographic (e.g., age, gendeÍ, race, etc.) and baseline characteristics (e.g., age at
onset of symptoms, age at SMA diagnosis, SMN2 copy number, subtype of SMA fType IB if
onset of symptoms {2 weeks of age, Type IC if onset of symptoms >12 weeks of age],
number of hours of BiPAP support, presence/absence of supplemental G{ube or NG tube
feeding within 30 days prior to the first dose, etc.) obtained before the first study drug
administration will be summarizedby cohort and overall using descriptive statistics.
Subject enrollment and disposition will be summarized. Protocol deviations will be listed.
For by-visit data summary, multiple records with the same visit label will be averaged.
Unscheduled results will not be included in the summary tables except for determining
baseline, but will be presented in data listings.
The presence/absence of supplemental G-tube or NG tube feeding within 30 days prior to the
first dose will be manually reviewed by Ionis physicians.
3.1.2 Subject Populatíon Anølyzed
Safety Population will include all subjects who are registered and receive at least one dose of
study drug.
Pharmacokinetic (PK) Population will include all subjects who are registered and for which
there is at least one evaluable post-dose pharmacokinetic sample. The Efficacy Evaluable
(Evaluable) Population will include all subjects who are registered, receive all scheduled
9


loading doses of study drug (i.e., Days 1, 15 and 85), and complete visits through at least Day
92. Two SMN2 Copy Set will include subset of Evaluable Population who have 2 copies of
lhe SMN2 gene. Safety analyses will be conducted on the Safety Population, PK analyses will
be conducted in PK Population, efficacy and biomarker analyses will be conducted on the
Evaluable Population and 2 SMN2 Copy Set. In addition to the Safety Population, PK
Population and Evaluable Population, it is recognized that some data displays will be
provided for "All Screened", "All Enrolled", and "screening Failures" subjects but no data
analysis will be executed in these populations.
3.1.3 Sample Size Considerstíon
There is no statistical rationale for the selected sample size for the assessment of efficacy,
since no data on the rate of motor milestone response, the primary efficacy endpoint of the
study, existed at the time of the study design. The sample size was selected to ensure that the
safety, tolerability, and pharmacokinetics of ISIS 396443 will be adequately assessed while
minimizing unnecessary subject exposure. According to Table 1, the sample size also allows
for sufficient power (-90%) to detect a response rate of at least30Yo.
Table I below provides the power analysis for 20 patients using the proportion of patients
who achieved improvement based on the mofor milestone endpoint (as defined in section
1.3.1). A two-sided exact test based on the binomial probability distribution with an alpha of
0.05 is used.
Table 1. Power analysis for 20 patients using the proportion of patients who achieved
improvement based on the motor milestone
Null hypothesis, H¡
Alternative hypothesis, Hu
Power
5% (1 responder)
20% (4 responders)
58%
5%
25% (5 responders)
77%
5%
30% (6 responders)
89%
5%
35% (7 responders)
95%
3.1.4 Safety Review and Plønned Interim Anølysís
To ensure subject safety, safety data will be reviewed on an ongoing basis by the Ionis Safety
Physician. Safety data will also be reviewed on an ongoing basis by the Data and Safety
Monitoring Board (DSMB). After the last patient within Cohort 1 completes the Day 29 visit,
safety results for the cohort will be reviewed by the Medical Monitor and the DSMB and a
recommendation regarding further enrollment and escalation to Cohort 2 will be made. The
Sponsor and DSMB will determine if enrollment of additional patients is required to confirm
there is an acceptable toxicity profile prior to its designation as MTD.
10


Interim efficacy and safety analyses will be performed to provide content for regulatory
submissions and to support ISIS 396443 drug development planning and business activities.
Specifically, analyses of primary and secondary efficacy endpoints (Sections 3.2 and 3.3), as well
as PK (Section 3.5) and safety analyses (Section 3.6), will be conducted at time points that allow
for reasonable interpretation of the data, such as all continuing subjects passing a specific age
threshold (e.g., 18, 24 or 36 months).
3.1.5 Incomplete or Missing Døta
3.1.5.1 Motor Mílestones
If not all tests were performed at a visit, the approach described below will be used for the
handling of missing data. Missing data will be imputed on an individual item level. If a test is
missing at screening, then the missing test will be imputed as the median of the non-missing
values of the group to which the subject belongs: age at s¡rmptom onset ({2 weeks, >12
weeks) by disease duration ( 12 weeks, >I2 weeks). If for the subject with the missing test,
the corresponding visit is flanked by visits with non-missing tests, the missing test will be
imputed using linear interpolation with the result rounded to the nearest integer score.
Otherwise, the missing test will be imputed as the lowest score of the visit in the group (age
at symptom onset by disease duration) to which the subject belongs. If the group is empty,
the missing test will be imputed as the lowest score of all available subjects of the visit.
Disease duration (week) will be calculated as (screening date-birth date)|7- age at symptom
onset in week.
3.1.5.2 CHOP-INTEND Total Score
The CHOP-INTEND assessment contains 16 items, 13 of them are tested on both right side
and left side. The item score will be equal to the maximum score of both sides for those 13
items. If the score for one side is missing the item score will be equal to the score of the non-
missing side.
If not all tests were performed at a visit, the approach described below will be used for the
handling of missing data. Missing data will be imputed on an individual item level. If a test is
missing at screening, then the missing test will be imputed as the median of the non-missing
values of the group to which the subject belongs: age at. symptom onset (12 weeks, >12
weeks) by disease duration ( {2 weeks, )12 weeks). If for the subject with the missing test,
the corresponding visit is flanked by visits with non-missing tests, the missing test will be
imputed using linear interpolation with the result rounded to the nearest integer score.
Otherwise, the missing test will be imputed as the lowest score of the visit in the group (age
at symptom onset by disease duration) to which the subject belongs. If the group is empty,
the missing test will be imputed as the lowest score of all available subjects of the visit.
11


3.1.5.3 Ventilutor Use
If data indicates a patient requires BiPAP or CPAP daily, but the number of hours ventilator use
is missing then the missing value will be imputed using the greater requirement of the visits
that flank the missing visit(s).
3,2 Primary Efficacy Analysis
Motor milestones will be assessed using Section 2 of the Hammersmith Infant Neurological
Exam (HINE) which is comprised of eight independent milestone categories: voluntary grasp,
ability to kick in supine position, head control, rolling, sitting, crawling, standing, and walking.
Within each of these categories, subjects can progress from complete inability to perform a motor
task, for example no grasp, no kicking, unable to maintain head upright, all the way to complete
mastery of each category, for example achievement of pincer grasp, touching toes while supine,
maintaining their head upright for a maximum of 26 motor milestones (see Table 2 below).
Table 2. Motor Milestone Achievement (Module 2, HINE)
Milestone progression
Motor
milestone
Voluntary
grasp
No grasp
Ability to kick No kicking
(in supine)
Head control
Rolling
Standing
Walking
Uses whole hand
Finger and thumb; Pincer grasP
immature grasp
Kicks horizontal; Upward (vertically) Touches leg 
Touches toes
legs do not lift
All the time upright
Unable 
to Wobbles
maintain upright
No rolling
Rolling to side 
Prone to supine 
Supine to prone
Sitting
Cannot sit
Crawling
Does not lift head On elbow
Stable sit
outstretched Crawling flat on
abdomen
Sit with support at ProPs
hips
hand
On
Pivots (rotates)
On hands and
knees
Does not support SupPorts weight
weight
No walking
Bouncing
with Stands unaided
(holding Walking
independently
Stands
support
Cruising
on)
For each dosing visit, in addition to the scheduled pre-dose assessment, the HINE including
motormilestonesarealsoassessed at3,6 hoursandI dayafter dosingtomonitorpatient
safety. Since the patient's motor function could be temporarily impacted by the lumbar
puncture procedure, only the motor milestone results from the pre-dose assessment will be
t2


used for the efficacy analysis and the results at 3, 6 hours and I day after dosing will be
excluded.
The improvement in motor milestones is defined as achievement of any of the following :
(Ð 
at least a 2-point increase from baseline or achievement of pincer grasp in the
category ofvoluntary grasp, or
(ii) 
at least a2-point increase in the ability to kick or achievement of touching toes, or
(iiÐ a l-point increase in any of the remaining 6 categories: head control, rolling,
sitting, crawling, standing, or walking.
Subjects who die or withdraw from the study will be counted as non-responders and will be
included in the denominator for the calculation of the proportion. As a result, mortality will
be accounted for in the motor milestone analysis. For subjects on permanent ventilation,
because motor milestone assessment continues, functional scores after permanent ventilation
will be used to assess improvement in motor milestones.
The primary analysis will be the proportion of subjects who achieve improvement in motor
milestones at their last available visit. It will be tabulated by cohort and overall. The primary
analysis will be conducted using the Evaluable Population.
As the sensitivity analyses, the proportion of subjects who achieve improvement in motor
milestones at their last available visit will be tabulated
1).by cohort and overall using the 2 SMN2 Copy Set
2).by cohort and overall using the Safety Population
3).by SMA subtype using the Safety Population
In addition, the following analyses will also be conducted to assess effects of treatment with
ISIS 396443 on the motor milestone improvement over time:
o The proportion of subjects who achieve improvement in motor milestones will be
tabulated by visit.
Subjects who die or withdraw from the study prior to a visit will be counted as non-
responders for the visit and all subsequent visits and will be included in the
denominator for the calculation of the proportion. As a result, mortality will be
accounted for in the motor milestone analysis. For subjects on peñnanent ventilation,
because motor milestone assessment continues, functional scores after permanent
ventilation will be used to assess improvement in motor milestones
If a subject misses a visit and has visit(s) beyond the visit, then the subject will be
counted as a non-responder and included in the denominator for the calculation of the
proportion for the visit.
13


a
a
o
o
Subjects who are ongoing in the study and haven't reached a visit will not be included
in the denominator for the calculation of the proportion for the visit and all
subsequent visits.
The proportion of subjects who achieve full head control (defined as All the Time
Upright), independent sitting (defined as either Prop, Stable Sit or Pivot), rolling
(Prone to Supine or Supine to Prone), crawling (defined as crawling On outstretched
hand, flat on abdomen, or On hands and knees), standing (defined as Stands with
support or Stands unaided), or walking (Cruising (holding on) or V/alking
independently) at their last available visit. Subjects who die or withdraw from the
study will be counted as non-responders and will be included in the denominator for
the calculation of the proportion. For subjects on permanent ventilation, because
motor milestone assessment continues, functional scores after permanent ventilation
will be used to assess improvement in motor milestones.
The proportion of subjects who achieve full head control (defined as All the Time
Upright), independent sitting (defined as either Prop, Stable Sit or Pivot), rolling
(Prone to Supine or Supine to Prone), crawling (defined as crawling On outstretched
hand, flat on abdomen, or On hands and knees), standing (defined as Stands with
support or Stands unaided), or walking (Cruising (holding on) or Walking
independently) will be tabulated by visit. Subjects who die or withdraw from the
study prior to a visit will be counted as non-responders for the visit and all subsequent
visits and will be included in the denominator for the calculation of the proportion.
For subjects on permanent ventilation, because motor milestone assessment
continues, functional scores after permanent ventilation will be used to assess
improvement in motor milestones.
If a subject misses a visit and has visit(s) beyond the visit, then the subject will be
counted as a non-responder and included in the denominator for the calculation of the
proportion for the visit.Subjects who are ongoing in the study and haven't reached a
visit will not be included in the denominator for the calculation of the proportion for
the visit and all subsequent visits.
Summary of change from baseline in the motor milestone total score at their last
available visit. The total motor milestone score equals to the sum of individual scores.
For example, if the subject cannot maintain head control this is scored as 0, head
wobbles as 1, and all the time upright as 2. Complete inability to perform any motor
task will therefore be as zero. The ability to master all motor milestones will be
scored as26.
Summary of change from baseline in the motor milestone total score by visit.
The above analyses will be tabulated
1).by cohort and overall using the Evaluable Population
2).by cohort and overall using the 2 SMN2 Copy Set
14


3). by cohort and overall using the Safety Population
4).by SMA subtype using the Safety Population
3.3 Secondary Effïcacy Analysis
3.3.1 Event-free Survívøl
Event-free survival will be estimated using the Kaplan-Meier (K-M) methodology. The
proportion of subjects who are alive and do not require permanent ventilatory support at age
of 3,6, 9,12,15, 18, 21,24,27,and 30 months will be estimated from the Kaplan-Meier
curve.
The medians and95%o CIs will be estimated if possible. The data will be tabulated
1).by cohort and overall using the Evaluable Population
2).by cohort and overall using the 2 SMN2 Copy Set
3).by cohort and overall using the Safety Population
4).by SMA subtype using the Safety Population
Event-free survival will be calculated from a patient's birth date to death date or permanent
ventilation date. Permanent ventilation is defined as tracheostomy OR >16 hours of
ventilation support/day continuously for ã4 days in the absence of an acute reversible
illness. The endpoint was determined by the investigator using chart review of all available
patient data. The date of the permanent ventilation event will be the date of reaching 14 days
of continuous BiPAP support for at least 16 hours; i.e., at the end of the l4-day period.
If a patient is not known to have died or have had permanent ventilation, the data will be
censored at the latest visit date with non-missing ventilation use assessment, either as an in-
person visit or by telephone contact, whether or not the patient had received the full course of
treatment, and whether the patient completed the study or withdrew prematurely.
The calculation will be as follows:
If a patient is not on permanent ventilation but died, event-free survival : number of months
between birth date and death date, censor:0;
If a patient is on permanent ventilation, event-free survival : number of months between
birth date and the date when the patient is first time on permanent ventilation, censor:O;
If a patient is alive and not on permanent ventilation, event-free survival : number of months
between birth date and last ventilation assessment date, censor:I. Where last ventilation
assessment date equals the latest date from the last visit date from Ventilation CRFs and last
phone contact date from Phone Contact CRFs where BiPAP or CPAP assessment is recorded.
15


3.3.2 CHOP-INTEND Infant Motor Function Scule
The CHOP-INTEND assessment contains 16 items, the total score ranges from worst
possible score of 0 to a best possible score of 64
The CHOP-INTEND infant motor function scale total scors, change, and percent change from
baseline will be summarized by visit.
In addition, the CHOP-INTEND infant motor function scale total score, change, and percent
change from baseline to the last available visit for each patient will also be summarized.
The proportion of subjects who achieve improvement in CHOP INTEND total scores will be
tabulated by visit. Improvement is defined as at least a four-point increase from baseline.
Subjects who die or withdraw from the study prior to a visit will be counted as non-
responders for the visit and all subsequent visits and will be included in the denominator for
the calculation of the proportion. For subjects on permanent ventilation, because CHOP
INTEND assessment continues, functional scores after permanent ventilation will be used to
assess improvement in CHOP-INTEND. If a subject misses a visit and has visit(s) beyond the
visit, then the subject will be counted as a non-responder and included in the denominator for
the calculation of the proportion for the visit.
Subjects who are ongoing in the study and haven't reached a visit will not be included in the
denominator for the calculation of the proportion for the visit and all subsequent visits.
The proportion of subjects who achieve improvement in CHOP INTEND total scores at their
last available visit will be tabulated. Subjects who die or withdraw from the study will be
counted as non-responders and will be included in the denominator for the calculation of the
proportion. As a result, mortality will be accounted for in the CHOP INTEND analysis. For
subjects on permanent ventilation, because CHOP INTEND assessment continues, functional
scores after permanent ventilation will be used to assess improvement in CHOP INTEND.
The above analyses will be tabulated
1). by cohort and overall using the Evaluable Population
2).by cohort and overall using the 2 SMN2 Copy Set
3). by cohort and overall using the Safety Population
a).by SMA subtype using the Safety Population
3.3.3 CMAP Ulnør and Peroneal Nerves
CMAP is an electrophysiological measure used to determine the approximate number of
motor neurons in a muscle or group of muscles. CMAP amplitude and CMAP area will be
evaluated at two sites, right side ulnar nefve and right side peroneal nerve.
T6


The absolute values for CMAP amplitude and CMAP area, as well as changes and percent
changes from baseline will be summarized by visit.
In addition, the absolute values for CMAP amplitude and CMAP area, as well as changes and
percent change from baseline to the last available visit for each patient will also be
summarized.
The proportion of subjects with a change from baseline CMAP amplitude at the peroneal
nerve greater than 0.5 mV will be tabulated by visit. Subjects who die or withdraw from the
study prior to a visit will be counted as non-responders for the visit and all subsequent visits
and will be included in the denominator for the calculation of the proportion. As a result,
mortality will be accounted for in the CMAP analysis. For subjects on permanent ventilation,
because CMAP assessment continues, the scores after permanent ventilation will be used to
assess improvement in CMAP. If a subject misses a visit and has visit(s) beyond the visit,
then the subject will be counted as a non-responder and included in the denominator for the
calculation of the proportion for the visit.Subjects who are ongoing in the study and haven't
reached a visit will not be included in the denominator for the calculation of the proportion
for the visit and all subsequent visits.
The proportion of subjects with a change from baseline CMAP amplitude at the peroneal
nerve greater than 0.5 mV at their last available visit will be tabulated. Subjects who die or
withdraw from the study will be counted as non-responders and will be included in the
denominator for the calculation of the proportion. As a result, mortality will be accounted
for in the CMAP analysis. For subjects on permanent ventilation, because CMAP assessment
continues, the assessments after permanent ventilation will be used to assess improvement in
CMAP.
The above analyses will be tabulated
1). by cohort and overall using the Evaluable Population
2).by cohort and overall using the 2 SMN2 Copy Set
3).by cohort and overall using the Safety Population
a). by SMA subtype using the Safety Population
3.4 ExploratoryAnalysis
3.4.1 Meøsures of Respiratory Status
Measures of respiratory status include number of respiratory events, respiratory infections,
respiratory-related hospitalizations, as well as number of hours per day ventilator use and 02
saturation awake.
t7


All serious adverse events that are coded into the System Organ Class (SOC) Respiratory,
thoracic, and mediastinal disorders, will be considered respiratory events.
Respiratory infections will include those serious adverse events that are coded under the
System Organ Class (SOC) of infections and infestations as well as those in the System
Organ Class (SOC) of investigations. For the sake of completeness, respiratory infections
will be identified via a two-step process: first, all serious adverse events that code to the High
Level Group Term of Respiratory tract infections under the SOC Respiratory, thoracic, and
mediastinal disorders will be extracted progtammatically; secondly, all remaining SAEs of
infections will be manually reviewed by a Safety Physician who will identify those that are
respiratory in nature.
Respiratory-related hospitalizations will combine the above two categories of respiratory
events and respiratory infections where the SAE had resulted in an inpatient hospitalization.
The number of respiratory events, respiratory infections, and respiratory-related
hospitalizations during the study will be analyzed using the rate at which they occur. For
descriptive purposes, the aggregate event rate will be calculated by dividing the total number
of respiratory events that occurred in the group by the total number of subject-years on study.
The aggregate rates for respiratory infections and respiratory-related hospitalizations will be
calculated similarly. The aggregate rates will be tabulated.
If a patient doesn't require BiPAP or CPAP daily, then the number of hours per day ventilator
use will be equal to 0. If ventilattor use is reported as a range, e.g, 6 to 12 hours, the maximum
will be used. The handling of missing values is described in Section 3.1.5.2.
The absolute values for number of hours per day ventilator use and 02 saturation awake, as
well as changes and percent changes from baseline will be summarized by visit.
The above analyses will be tabulated
1). by cohort and overall using the Evaluable Population
2).by cohort and overall using the 2 SMN2 Copy Set
3).by cohort and overall using the Safety Population
4). by SMA subtype using the Safety Population.
3.4.2 GrowthParameters
The growth parameters will include weight to age ratio, weight to length ratio, head
circumference, chest circumference, head to chest circumference ratio, arm circumference,
and weight for age percentile, height for age percentile, and head circumference percentile
using the V/HO child growth standards (V/HO Child Growth Standards, 2006). The absolute
values as well as changes, and percent changes from baseline will be summarized by visit.
18


The above analyses will be tabulated
1). by cohort and overall using the Evaluable Population
2).by cohort and overall using the 2 SMN2 Copy Set
3).by cohort and overall using the Safety Population
4).by SMA subtype using the Safety Population.
3.4.3 Hsmmersmith Functionøl Motor Scale - Expønded
All subjects who have maintained a CHOP INTEND total score of >50 for two consecutive
study visits will be evaluated using the Hammersmith Functional Motor Scale Expanded
(HFMSE). This evaluation will be performed in addition to the CHOP INTEND. Since the
data is only collected for a subset of patients, the data will be provided in a data listing.
3.5 PharmacokineticAnalyses
CSF and plasma samples will be collected at protocol designated times for ISIS 396443
pharmacokinetic assessments from all treatment groups (cohorts).
3.5,1 CSF Concentrøtion Døta
CSF concentrations of ISIS 396443, along with the scheduled (nominal) and actual samples
times (i.e., time from IT dosing) will be listed (when applicable) for each patient, treatment
group (cohort), nominal dose, and day. Differences between scheduled and actual sampling
days will also be listed for all patients. Percent differences between actual administered dose
and nominal dose will also be listed.
CSF concentrations below the lower limit of quantification (LLOQ) will be indicated by
ooBLQ". For the pu{pose of calculating typical descriptive statistics (n, mean, SD, o/oCV,
geometric mean, geometric o/oCV, median, minimum, and maximum) for CSF
concentrations, all BLQ values will be set to zero. Mean CSF concentrations that are BLQ
will be presented as BLQ, and the SD and %CV will be reported as not applicable. Summary
statistics of the ISIS 396443 CSF concentrations will be tabulated by treatment group
(cohort), nominal dose, duy, and scheduled time point. At the discretion of the
pharmacokineticist and/or biostatistician, samples may be excluded from descriptive statistics
if there are large deviations between scheduled and actual sampling days or times, or large
deviations between actual dose and nominal dose.
ISIS 396443 CSF concentration versus time (actual) profiles from Day 1 to Day 673, for each
patient, as well as the mean (+SE) CSF concentration versus time (scheduled) profiles for
each treatment cohort, will be presented graphically. Samples may be excluded from the
mean plots if there are large deviations between scheduled and actual sampling times, or
large deviations between actual dose and nominal dose.
r9


Due to the limited CSF samples collected no CSF pharmacokinetic parameters will be
calculated.
3.5.2 Plasma Pharmacokinetics
3.5.2.1 Plasma Concentrøtion Døta
Plasma concentrations of ISIS 396443, along with the scheduled (nominal) and actual
samples times (i.e., time from IT dosing) will be listed (when applicable) for each patient,
treatment group (cohort), nominal dose, and day. Percent differences between scheduled and
actual sampling times will also be listed for all patients. Percent differences between actual
administered dose and nominal dose will also be listed.
Plasma concentrations below the lower limit of quantification (LLOQ) will be indicated by
"BLQ". For the purpose of calculating typical descriptive statistics (n, mean, SD, o¿CV,
geometric mean, geometric yoCV, median, minimum, and maximum) for plasma
concentrations, all BLQ values will be set to zero. Mean plasma concentrations that are BLQ
will be presented as BLQ, and the SD and %CY will be reported as not applicable. Summary
statistics of the ISIS 396443 plasma concentrations will be tabulated by treatment group
(cohort), nominal dose, duy, and scheduled time point. At the discretion of the
pharmacokineticist andlor biostatistician, samples may be excluded from descriptive statistics
if there are large deviations between scheduled and actual sampling times, or large deviations
between actual dose and nominal dose.
ISIS 396443 plasma concentration versus time (actual) profiles from Day 1 to Day 15
(Cohorts I and 2), for each patient, as well as the mean (+SE) plasma concentration versus
time (scheduled) profiles for each treatment cohort, will be presented graphically on linear
and semilogarithmic scales. Additionally, ISIS 396443 plasma concentration versus time
(actual) profiles (Day 1) from 0 to 24 hours for all patients , as well as the mean (+SE)
plasma concentration versus time (scheduled) profiles (0 to 2+ hours) for each applicable
treatment cohort will be presented graphically on linear and semilogarithmic scales. Samples
may be excluded from the mean plots if there are large deviations between scheduled and
actual sampling times, or large deviations between actual dose and nominal dose.
ISIS 396443 plasma trough þredose) concentration versus time (actual) profiles from Day 1
to Day 673, for each patient, as well as the mean (+SE) plasma concentration versus time
(scheduled) profiles for each treatment cohort, will be presented graphically. Samples may
be excluded from the mean plots if there are large deviations between scheduled and actual
sampling times, or large deviations between actual dose and nominal dose.
3.5.2.2 Pløsma Pharmøcokinetic Parameters
Non-compartmental pharmacokinetic analysis of ISIS 396443 will be carried out on each
individual subject data set using Phoenix WinNonlin version 6.0 or higher (Pharsight
20


Corporation, Mountain View, CA). Plasma pharmacokinetic parameters in each patient
(when applicable) will be determined. For calculation of PK parameters, all BLQ values will
be set to zero. The following plasma PK parameters will be calculated (when applicable) and
based on actual sampling times:
o C-u*t the maximum observed drug concentration in plasma.
o T,nu*i the time at which C*u" occurs.
The following PK parameters may be calculated using actual sampling times where
appropriate data exists at the discretion of the pharmacokinetic scientist:
o AUCo-+¡t Ítroâs under the plasma concentration-time curve from zero time (pre-dose)
to 4 hours after IT administration will be calculated using the linear trapezoidal rule.
o Additional partial AUC values (e.g., AUCo-z¿n) may be calculated depending upon the
samples collected during the study and the number of patients for which additional
samples are collected via an indwelling catheter, as well as when warranted and at the
discretion of the pharmacokinetic scientist.
. Though outlined in the study protocol, apparent terminal elimination half-life (tz)
values will not be determined as the limited post-treatment sampling schedule precludes
this evaluation.
Plasma pharmacokinetic parameters (if applicable) will be summarized using descriptive
statistics (n, mean, SD, o/oCV, geometric mean, geometric yoCV, median, minimum, and
maximum) by treatment cohort, nominal dose, and day.
3.5.2.3 fmmunogenicity Analyses
Immunogenicity (IM) testing (anti-ISIS 396443 antibody positivity), using designated plasma
samples collected from each study subject, is planned to be conducted and reported. Initially,
designated available immunogenicity plasma samples collected only on Days I (predose
only), 85 (predose only), 169,253,337, 42I,505, 589, and 673 are planned to undergo
immunogenicity testing. Plasma samples collected at other time points for ISIS 396443
concentration determinations may also be potentially evaluated for IM testing if of further
interest and deemed warranted by the pharmacokinetic scientist. An individual sample result
will be designated 'antibody positive' based on both positive screening and confirmation
assay results (i.e., confirmed positive result), and otherwise will be deemed 'antibody
negative'. A study subject will be given'antibodypositive' status if they have at least one
conf,rrmed positive sample result at any time during the treatment or post-treatment
evaluation periods. A study subject will be given 'antibody negative' status if all evaluated
IM sample results are antibody negative and they have at least one evaluable IM result from
Day 85 or later during the treatment and post-treatment evaluation periods. Otherwise, a
study subject will be given 'antibody inconclusive' status.
2t


The IM incidence and IM incidence rate at each evaluated study timepoint, and for the
overall treatment and post-treatment evaluation period, will be determined and appropriately
summarized, by treatment, as the number of and percent (%) of evaluated subjects with
antibody negative, antibody positive, and antibody inconclusive status. In addition, in
antibody positive study subjects, antibody titers of any antibody positive samples will be
reported (listed) and also appropriately summarized across subjects and by treatment (e.g., at
each evaluated timepoint, or by observed peak titer values, etc.) at the discretion of the
de si gnated study pharmacokineticist and/ or statistician.
At this time, antibody positivity in any antibody positive study subjects from this study is not
yet planned to be designated as being either 'persistent' or 'transient'. However, the IM
results from this study may be evaluated at alater date (and possibly in combination with IM
results from other studies potentially involving these patients) to determine these
designations. At that time, criteria for persistent versus transient positive antibody responses
will also be defined.
V/hen and where warranted, PK and selected safety and eff,tcacy results may be further
summarized (stratified) by antibody status (antibody negative vs. positive subjects) and
treatment.
3.6 Safety Analyses
Safety will be assessed by summarizing the incidence and type of AEs and SAEs.
Laboratory parameters will be provided in the data listings.
Days of exposure, number of doses and total amount of drug received for each subject will be
summarized by cohort and overall.
Time on study will be categorized into intervals and summarized by cohort and overall.
The time on stuáy will be defined as the total number of days a subject is known to be
followed on study calculated as follows:
Time on study: Last date on study -Date of first dose+ 1.
Where the last date on study is defined as the date of the latest visit or evaluation or
telephone contact, or time of death from all available data for a given subject.
3,6.1 Adverse Events
The incidence of AEs will be summarizedby Medical Dictionary for Regulatory Activities
(MedDRA) preferred term and system organ class for:
22


. Any treatment emergent adverse events
o Related treatment emergent adverse events. Related adverse event is defined as those
events with related or possible related relationship to study drug.
. Any treatment emergent adverse events by severity
o Related treatment emergent adverse events by severity
o Serious treatment emergent adverse events
o Serious and related treatment emergent adverse events.
o Lumber puncture related treatment emergent adverse events. Lumber puncture related
adverse event is defined as those events with related or possible related relationship to
study procedure/other study drug.
AEs that lead to treatment discontinuation will be listed separately. Early dosing termination
and early study termination will be summarized by termination reason.
A treatment emergent adverse event is defined as an event occurring after the initiation of
study treatment and before the end of the follow-up period. Non-treatment emergent adverse
event will be flagged in the data listing.
If there is no "Formlink" link, and the AE (start date/time) occurs after the subject's first
dosing dateltime, then the AE is treatment-emergent. Otherwise, if the AE (start date/time)
occurs prior to the subject's first dosing date/time, then the AE is not treatment-emergent.
If there is a "Formlink" link between two AE records, then we compare them pairwise, and
consider two cases, where we compare the AE severity (mild/moderate/severe) between the
two records in the pair. We chronologically order the 2 records (by AE start date) and refer
to the o'first" and "second" AEs.
Case 1: The fîrst AE record in the pair occurs before first dosing, and the second record
occurs after dosing.
If the AE severity on the second record is worse than the severity on the first record, then
only count the second AE as treatment-emergent. But, if the severity improves (second
record sev.erity is less severe than the first record severity), then neither record is counted as
treatment-emergent.
Case 2: Both AE records in the pair occur after first dosing.
If the AE severity on the second record is worse than the severity on the first record, then
count both records as treatment-emergent. But, if the severity improves, then only count the
f,rrst record as treatment-emergent.
23


When counting the total number of treatment-emergent events, events linked together
through change in severity will still be counted as separate events.
The most conservative approach will be used to determine if the event occurs during the
treatment or follow-up periods. For example,
AEs that have onset dates or resolution dates prior to the first study treatment dates will
be considered to have occurred prior to the study period. If the onset or resolution date of
an AE is a partial date with only month or year available or completely missing, then the
event is assumed to be within the study period unless the year is prior to the year of the
first study treatment date, or if in the same year, the month is prior to the month of the
first study treatment date.
o
o AEs that have onset dates after study termination date will be assumed to have occurred
after the study period. If the onset of an AE is a partial date with only month or year
available or complete missing, then similar approach as above will be used.
3.6.1.1 Adverse Events over Time
The incidence of AEs will be evaluated by time of onset by 90-day time intervals. For a
given time interval, the number of subjects who were followed for adverse events during that
time interval will be presented along with the incidence of adverse events during that time
interval. Therefore, for a given System Organ Class (SOC) or preferred term (PT), subjects
will be counted only once for a given time interval but may be counted more than once
across time intervals.
3.6.1.2 Adverse events following Dosing
To examine the onset of any adverse events following dosing, the incidence of events that
occur during loading and maintenance dosing periods will be presented by treatment group
and overall. In addition, presentations of events that occurred in the first 24 hours and the
first72 hours following dosing will be provided.
3.6.2 Løboratory Meøsurements
The following is the list of lab analytes that will be collected throughout the study:
o Chemistry: Sodium, Potassium, Chloride, Total protein, Albumin, Calcium, Phosphorus,
Bicarbonate, Glucose, BlIlI, Creatinine, Cystatin C, Total serum bilirubin, ALT, AST,
Alkaline phosphatase, CPK
o Hematology: Hematocrit, Hemoglobin, Platelets, RBC, 'WBC, and WBC differential
(Basophils, Eosinophils, Lymphocytes, Monocytes, and Neutrophils)
. 
Coagulation: PT, aPTT, INR
o Urinalysis: Specific gravity, pH, Protein, Glucose, Ketones, Bilirubin, Blood, Red blood
cells, White blood cells, Epithelial cell, Bacteria, Casts, Crystals
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. 
CSF safety: Red blood cells, White blood cells, Glucose, Protein.
o SMN Genetics: SMN2 copy number and SMN gene sequencing
If V/BC differential absolute counts are missing, and percentages are available, then the
absolute counts will be calculated by multiplying the percentage by total WBC count.
Conversely, if absolute counts are available, percentages are missing, percentages will be
calculated by dividing absolute count by the total WBC count. If neutrophils counts and
percentages are missing, and segmented neutrophils and bands neutrophils results are
available, then neutrophils will be calculated by adding segmented neutrophils and bands
neutrophils.
Each subject's laboratory values will be classified according to whether the test result is
oolo\ry'" (i.e., below the lower limit of normal [LLN]), 'onormal" (within the normal range), or
"high" (i.e., above the upper limit of normal [ULN]). If a subject is missing a baseline value
but had a post-baseline value, then the baseline assessment is labeled as "unknown".
Likewise, if a subject had a baseline value but had no post-baseline values, then the
minimum and maximum are labeled as "unknown". Post-baseline laboratory results are
defined as any assessment taken after the first dose. The shifts (relative to the normal range)
from baseline to the minimum and maximum post-baseline values will be presented. Should
a treatment affect a laboratory parameter, that parameter could be affected at different times
for different subjects. Therefore, these analyses present the most extreme values for each
subject. For many laboratory parameters, the effect could be in either direction, (i'e., an
increase or a decrease), so both the maximum and minimum values have been analyzed.
From these, the shifts (relative to the normal range) from baseline to low and high will be
calculated. If a subject's value shifts, it can change from normal to either low or high, from
low to normal or high, from high to normal or low, or from unknown to low, normal, or high.
For each parameter, the incidence of shift to low will be summarized using the minimum
post-baseline values. Shift to low includes subjects with a normal, high, or unknown
baseline value and at least one post-baseline value of the given test. Similarly, the incidence
of shift to high will be summarized using the maximum post-baseline values. Shift to high
includes subjects with a low, normal, or unknown baseline value and at least one post-
baseline value.
3.6.3 Vital Sígns
Vital signs will include body temperature, heart rate, respiratory rate, and systolic and
diastolic blood pressure, pulse oximetry awake and pulse oximetry asleep. The vital sign
data will be provided in a data listing.
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3.6.4 Physícal Examinatìons
Adverse changes in physical examinations that are deemed clinically significant by the
investigator will be classified as adverse events. All physical examination data will be
provided in a data listing.
3.6,5 Leød Electrocørdiograms (ECG)
The ECG data will include ventricular rate (VR), PR interval, QRS duration, QT/QTC, and
Overall interpretation.
Outlier analyses will be performed for the corrected Fridericia QT interval (QTcF). This
includes summaries of the number and percentage of subjects with a post-baseline QToF
greater than 450 msec, 480 msec, and 500 msec) and the number and percentage of subjects
with an increase of 30 msec and 60 msec from baseline in corrected QT interval (QTc).
QTçF will be calculated based on the subject's reportable ECG data at each visit using the
formula described below :
QTcF: QT / (RR) t", where RR:60/VR
The ECG data will also be provided in a data listing'
3.6.6 Concomitant Medicøtions
Concomitant medications will be coded using V/HO Drug dictionary and summarized by
ATC class, generic name and treatment group.
3.6.7 Hammersmith Infønt Neurologìcal Examinatíons - Sections I ønd 3
Adverse changes in neurological examinations that are deemed clinically significant by the
investigator will be classified as adverse events. Data collected for HINE sections 1 and 3
will be provided in a data listing.
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