Conditions: Hypertension
Keywords: Hypertension Hypertensive AGT Angiotensinogen Blood Pressure
Allocation: RANDOMIZED | Intervention model: PARALLEL
Enrollment: 160
Healthy volunteers: no

Brief summary:
The purpose of this study was to evaluate the effect of IONIS-AGT-LRx compared to placebo on seated automated office systolic blood pressure (SBP) from baseline to Study Day 85 in uncontrolled hypertensive participants on ≥ 3 antihypertensive medications and to evaluate the effect of IONIS-AGT-LRx on ambulatory blood pressure, seated automated office SBP, seated automated office diastolic blood pressure (DBP), and plasma angiotensinogen (AGT) at each scheduled visit in uncontrolled hypertensive participants on ≥ 3 antihypertensive medications.

Detailed description:
This was a phase 2, double-blind, randomized, placebo-controlled study in up to 160 participants. Participants were randomized in a 2:1 ratio and received a once-weekly subcutaneous (SC) treatment with either IONIS-AGT-LRx or matching placebo. The length of participation in the study was approximately 31 weeks, which included an up to 6-week screening period, a 12-week treatment period, and a 13-week post-treatment period.

Arms:
arm 1: Participants received ISIS 757456 matching placebo subcutaneously once weekly for 12 weeks. arm 2: Participants received ISIS 757456 80 milligrams (mg), subcutaneous (SC) injection, once weekly for 12 weeks. arm 3: Participants received ISIS 757456 120 mg, SC injection, once weekly for 12 weeks.

Interventions: intervention 1: Placebo intervention 2: IONIS-AGT-LRx
Intervention descriptions: intervention 1: ISIS 757456 matching placebo administered by SC injection. intervention 2: IONIS-AGT-LRx administered by SC injection.

Protocol document:
Prot_SAP_001.pdf: 
 
 
A Double-Blind, Placebo-Controlled, Phase 2 Study to Assess the Safety, Tolerability and Efficacy 
of IONIS-AGT-LRx, an Antisense Inhibitor of Angiotensinogen Production Administered 
Subcutaneously for 12 Weeks to Hypertensive Patients With Uncontrolled Blood Pressure   
 
NCT04714320 
 
15 Aug 2022 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Ionis.com 


 
 
IONIS PHARMACEUTICALS, INC. 
 
ISIS 757456-CS4 
 
A Double-Blind, Placebo-Controlled, Phase 2 Study to Assess the 
Safety, Tolerability and Efficacy of IONIS-AGT-LRX, an Antisense 
Inhibitor of Angiotensinogen Production Administered 
Subcutaneously for 12 Weeks to Hypertensive Patients with 
Uncontrolled Blood Pressure 
 
Amendment 4 – 4 November 2021 
 
 
ASO Targeting RAAS  
 
 
Trial Sponsor 
Ionis Pharmaceuticals 
2855 Gazelle Court 
Carlsbad, CA  92010 
Phone: 760-931-9200 
Fax: 760-603-2588
 


ISIS 757456-CS4 (ASTRAAS) 
CONFIDENTIAL 
Amendment 4
Protocol 
4 November 2021
 
2 
 
ISIS 757456-CS4 
Amendment 4 
 
Clinical Phase:  2 
 
A Double-Blind, Placebo-Controlled, Phase 2 Study to Assess the 
Safety, Tolerability and Efficacy of IONIS-AGT-LRX, an Antisense 
Inhibitor of Angiotensinogen Production Administered 
Subcutaneously for 12 Weeks to Hypertensive Patients with 
Uncontrolled Blood Pressure 
 
 
ASO Targeting RAAS  
 
Protocol History 
Original Protocol: 
8 September 2020 
Amendment 1: 
5 November 2020 
Amendment 2:  
12 April 2021 
Amendment 3: 
15 June 2021 
 
Sponsor 
Ionis Pharmaceuticals, Inc. 
Carlsbad, CA  92010 
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 independent ethics committee 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.   


ISIS 757456-CS4 (ASTRAAS) 
CONFIDENTIAL 
Amendment 4
Protocol 
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Protocol Signature Page 
 
Protocol Number: 
ISIS 757456-CS4 
Protocol Title: 
A Double-Blind, Placebo-Controlled, Phase 2 Study to Assess the Safety, 
Tolerability and Efficacy of IONIS-AGT-LRX, an Antisense Inhibitor of 
Angiotensinogen Production Administered Subcutaneously for 12 Weeks 
to Hypertensive Patients with Uncontrolled Blood Pressure  
Amendment:  
4 
Date:  
 
4 November 2021 
 
I hereby acknowledge that I have read and understand the attached clinical protocol, entitled 
“A Double-Blind, Placebo-Controlled, Phase 2 Study to Assess the Safety, Tolerability and 
Efficacy of IONIS-AGT-LRX, an Antisense Inhibitor of Angiotensinogen Production 
Administered Subcutaneously for 12 Weeks to Hypertensive Patients with Uncontrolled Blood 
Pressure,” dated 4 November 2021, 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 and the Declaration of Helsinki. 
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 757456-CS4 (ASTRAAS) 
CONFIDENTIAL 
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Protocol 
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TABLE OF CONTENTS 
PROTOCOL AMENDMENT .......................................................................................................10 
PROTOCOL SYNOPSIS ..............................................................................................................18 
STUDY DESIGN AND TREATMENT SCHEMA ......................................................................26 
STUDY GLOSSARY ....................................................................................................................27 
1. 
OBJECTIVES AND ENDPOINTS ............................................................................29 
1.1. 
Objectives ...................................................................................................................29 
1.1.1. 
Primary Objective .......................................................................................................29 
1.1.2. 
Secondary Objectives .................................................................................................29 
1.1.3. 
Safety Objectives ........................................................................................................29 
1.1.4. 
Exploratory Objectives ...............................................................................................29 
1.2. 
Study Endpoints ..........................................................................................................30 
1.2.1. 
Primary Endpoint ........................................................................................................30 
1.2.2. 
Secondary Endpoints ..................................................................................................30 
1.2.3. 
Safety Endpoint ..........................................................................................................30 
1.2.4. 
Exploratory Endpoints ................................................................................................30 
2. 
BACKGROUND AND RATIONALE .......................................................................31 
2.1. 
Overview of Disease ...................................................................................................31 
2.2. 
Therapeutic Rationale .................................................................................................31 
2.3. 
ISIS 757456 ................................................................................................................33 
 
 
 
 
2.3.3. 
Preclinical Experience ................................................................................................34 
2.3.4. 
Clinical Experience .....................................................................................................34 
 
 
2.5. 
Benefit-Risk Assessment ............................................................................................35 
2.5.1. 
Benefit Assessment .....................................................................................................35 
2.5.2. 
Risk Assessment .........................................................................................................36 
3. 
EXPERIMENTAL PLAN ..........................................................................................37 
3.1. 
Study Design ...............................................................................................................37 
3.2. 
Number of Study Centers ...........................................................................................37 
3.3. 
Number of Patients .....................................................................................................37 


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3.4. 
Overall Study Duration and Follow-up ......................................................................37 
3.4.1. 
Screening ....................................................................................................................37 
3.4.2. 
Treatment ....................................................................................................................37 
3.4.3. 
Post-Treatment ............................................................................................................37 
3.5. 
End-of-Study ...............................................................................................................38 
4. 
PATIENT ENROLLMENT ........................................................................................38 
4.1. 
Screening ....................................................................................................................38 
4.2. 
Randomization ............................................................................................................38 
4.3. 
Replacement of Patients .............................................................................................38 
4.4. 
Unblinding of Treatment Assignment ........................................................................38 
5. 
PATIENT ELIGIBILITY ...........................................................................................39 
5.1. 
Inclusion Criteria ........................................................................................................39 
5.2. 
Exclusion Criteria .......................................................................................................40 
6. 
STUDY PROCEDURES ............................................................................................43 
6.1. 
Study Schedule ...........................................................................................................43 
6.1.1. 
Screening ....................................................................................................................43 
6.1.1.1. 
24-Hour Ambulatory Blood Pressure Monitoring ......................................................43 
6.1.1.2. 
Seated Automated Office Blood Pressure Measuring Instructions at Each 
Visit .............................................................................................................................44 
6.1.1.3. 
Orthostatic Blood Pressure Assessment .....................................................................45 
6.1.2. 
Treatment Period ........................................................................................................45 
6.1.3. 
Post-Treatment Period ................................................................................................46 
6.2. 
Laboratory Assessments .............................................................................................46 
6.3. 
Patient Reported Outcomes ........................................................................................47 
6.3.1. 
Patient Global Impression Items .................................................................................47 
6.3.2. 
EuroQol-5 Dimension 5 Level (EQ-5D-5L)...............................................................47 
6.3.3. 
12-Item Short Form Survey (SF-12)...........................................................................47 
6.3.4. 
Power Over Pressure- Hypertension Specific Questionnaire (POP-HSQ) .................47 
6.4. 
Restriction on the Lifestyle of Patients .......................................................................47 
6.4.1. 
Contraception Requirements ......................................................................................47 
7. 
STUDY DRUG ...........................................................................................................48 
 
8 
7.2. 
Packaging and Labeling ..............................................................................................49 


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7.3. 
Study Drug Accountability .........................................................................................49 
8. 
TREATMENT OF PATIENTS ..................................................................................49 
8.1. 
Study Drug Administration.........................................................................................49 
8.2. 
Other Protocol-Required Drugs ..................................................................................49 
8.3. 
Other Protocol-Required Treatment Procedures ........................................................49 
8.4. 
Treatment Precautions ................................................................................................49 
8.5. 
Safety Monitoring Rules .............................................................................................49 
8.5.1. 
Safety Monitoring Rules for Liver Chemistry Tests ..................................................50 
8.5.2. 
Safety Monitoring Rules for Platelet Count Results...................................................51 
8.5.3. 
Safety Monitoring for Minor Bleeding Events ...........................................................53 
8.5.4. 
Safety Monitoring for Potassium ................................................................................53 
8.5.5. 
Safety Monitoring for Blood Pressure ........................................................................53 
8.6. 
Stopping Rules ............................................................................................................54 
8.6.1. 
Stopping Rules for Liver Chemistry Elevations .........................................................54 
8.6.2. 
Stopping Rules for Renal Function Test Results ........................................................54 
8.6.3. 
Stopping Rule for Platelet Count Results ...................................................................54 
8.6.4. 
Bleeding Events ..........................................................................................................56 
8.6.4.1. 
Definition of Major Bleeding Events (Schulman and Kearon 2005) .........................56 
8.6.4.2. 
Definition of Clinically Relevant Non-Major Bleeding Events .................................56 
8.6.4.3. 
Definition of Minor Bleeding Events .........................................................................56 
8.6.5. 
Stopping Rule for Potassium ......................................................................................57 
8.6.6. 
Stopping Rule for Blood Pressure ..............................................................................57 
8.7. 
Adjustment of Dose and/or Treatment Schedule ........................................................59 
8.8. 
Discontinuation of Study Drug ...................................................................................59 
8.9. 
Withdrawal of Patients from the Study Procedures ....................................................59 
8.10. 
Concomitant Therapy and Procedures ........................................................................60 
8.10.1. 
Concomitant Therapy .................................................................................................60 
8.10.2. 
Concomitant Procedures .............................................................................................61 
8.11. 
Treatment Compliance ................................................................................................61 
9. 
SERIOUS AND NON-SERIOUS ADVERSE EVENT REPORTING .....................61 
9.1. 
Sponsor Review of Safety Information ......................................................................61 
9.2. 
Regulatory Reporting Requirements ..........................................................................61 


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9.3. 
Definitions ..................................................................................................................62 
9.3.1. 
Adverse Event .............................................................................................................62 
9.3.2. 
Adverse Drug Reaction and Suspected Unexpected Adverse Drug Reaction ............62 
9.3.3. 
Serious Adverse Event ................................................................................................63 
9.3.4. 
Adverse Event of Special Interest ...............................................................................63 
9.4. 
Monitoring and Recording Adverse Events ...............................................................64 
9.4.1. 
Serious Adverse Events ..............................................................................................64 
9.4.2. 
Non-Serious Adverse Events ......................................................................................64 
9.4.3. 
Evaluation of Adverse Events (Serious and Non-Serious) .........................................64 
9.4.3.1. 
Relationship to the Study Drug...................................................................................65 
9.4.3.2. 
Severity .......................................................................................................................65 
9.4.3.3. 
Action Taken with Study Drug ...................................................................................65 
9.4.3.4. 
Treatment Given for Adverse Event ...........................................................................66 
9.4.3.5. 
Outcome of the Adverse Event ...................................................................................66 
9.4.3.6. 
Follow-up of Adverse Event .......................................................................................66 
9.5. 
Procedures for Handling Special Situations ...............................................................67 
9.5.1. 
Abnormalities of Laboratory Tests .............................................................................67 
9.5.2. 
Prescheduled or Elective Procedures or Routinely Scheduled Treatments ................67 
9.5.3. 
Medication Errors .......................................................................................................68 
9.5.4. 
Contraception and Pregnancy .....................................................................................68 
10. 
STATISTICAL CONSIDERATIONS .......................................................................69 
10.1. 
Stratification, Subsets, and Covariates .......................................................................69 
10.2. 
Sample Size Considerations .......................................................................................69 
10.3. 
Populations .................................................................................................................69 
10.4. 
Definition of Baseline .................................................................................................69 
10.5. 
Interim Analysis and Multiplicity ...............................................................................69 
10.6. 
Planned Methods of Analysis .....................................................................................70 
10.6.1. 
Demographic and Baseline Characteristics ................................................................70 
10.6.2. 
Safety Analysis ...........................................................................................................70 
10.6.3. 
Efficacy Analysis ........................................................................................................71 
10.6.4. 
Pharmacokinetic Analysis ..........................................................................................71 
10.6.5. 
Exploratory Analyses ..................................................................................................72 


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11. 
INVESTIGATOR’S REGULATORY OBLIGATIONS ...........................................72 
11.1. 
Informed Consent .......................................................................................................72 
11.2. 
Ethical Conduct of the Study ......................................................................................73 
11.3. 
Independent Ethics Committee/Institutional Review Board ......................................73 
11.4. 
Patient Confidentiality ................................................................................................73 
12. 
ADMINISTRATIVE AND LEGAL OBLIGATIONS ..............................................74 
12.1. 
Protocol Amendments ................................................................................................74 
12.2. 
Study Termination ......................................................................................................74 
12.3. 
Study Documentation and Storage .............................................................................74 
12.4. 
Study Monitoring ........................................................................................................75 
12.5. 
Language .....................................................................................................................75 
12.6. 
Compensation for Injury .............................................................................................75 
13. 
REFERENCES ...........................................................................................................76 
APPENDIX A. SCHEDULE OF PROCEDURES .....................................................................78 
APPENDIX B. LIST OF LABORATORY ANALYTES ..........................................................82 
APPENDIX C. PK SAMPLING SCHEDULE ...........................................................................84 
APPENDIX D. GRADING SCALE FOR ADVERSE EVENTS RELATING TO 
LABORATORY ABNORMALITIES ..............................................................86 
 
 
 


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LIST OF TABLES 
Table 1: 
Known Risks Associated with RAAS Blockade ........................................................36 
Table 2: 
Orthostatic Hypotension Assessment .........................................................................45 
Table 3: 
Study Drug Characteristics .........................................................................................48 
Table 4: 
Study Drug Dosing Information .................................................................................49 
Table 5: 
Monitoring Schedule and Stopping Rules for Platelet Counts ...................................52 
Table 6: 
Recommended Laboratory Analyses ..........................................................................56 
Table 7: 
Blood Pressure Stopping and Monitoring Rules ........................................................58 
 
LIST OF FIGURES 
Figure 1: 
The Renin-Angiotensin-Aldosterone (RAAS) System and Site of Action for 
Principle Antagonists ..................................................................................................32 
 
 
 


ISIS 757456-CS4 (ASTRAAS) 
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Amendment 4
Protocol 
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PROTOCOL AMENDMENT 
 
Protocol Number: 
ISIS 757456-CS4 
Protocol Title: 
A Double-Blind, Placebo-Controlled, Phase 2 Study to Assess the Safety, 
Tolerability and Efficacy of IONIS-AGT-LRX, an Antisense Inhibitor of 
Angiotensinogen Production Administered Subcutaneously for 12 Weeks 
to Hypertensive Patients with Uncontrolled Blood Pressure 
Amendment:  
4 
Date:  
 
4 November 2021 
 
The main purpose of this amendment is to update some of the inclusion and exclusion criteria 
related to baseline requirements for ECG abnormalities, renal function, diuretic use and the use 
of medications that may cause hyperkalemia.  Based on the collective safety and laboratory data 
currently available across ISIS 757456 clinical studies and the lack of safety signals related to 
hyperkalemia, ECG, and renal function, the criteria below have been revised to allow patients 
with these baseline abnormalities:  Other changes are outlined below as well as the rationale for 
the updates: 
i) 
update the eGFR exclusion criterion to < 30 mL/min/1.73 m2  
ii) 
update the UPCR exclusion criterion to > 0.5 mg/mg and remove the UACR exclusion 
criterion 
iii) 
revise the renal stopping rules to align with the modified eligibility criteria  
iv) 
allow patients with ECG abnormalities, aside from those who have clinically relevant 
prolonged QTc per Investigator judgement  
v) 
allow patients on potassium sparing diuretics while excluding patients with a history of 
hyperkalemia per Investigator judgement 
vi) 
revise the definition of baseline for renal stopping rules in order to more accurately 
determine if additional monitoring or stopping of Study Drug is necessary 
Further, to clarify that in the event Day 1 lab results are uninterpretable (e.g., due to hemolysis 
or missing sample), Day 8 dosing is allowed but the investigator is instructed to obtain an 
interpretable result before study drug dosing on Day 15.  And finally, to clarify the testing 
needed for HBV at screening 
The table below provides a list of modifications to the protocol; added text in bold, deleted text 
in strikethrough.  


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Protocol Section 
Description of Change 
Rationale 
Protocol Synopsis 
Section 5.1 Inclusion 
Criteria 
6. At Screening, the patient must have been on a 
stable, maximally tolerated regimen (per 
Investigator judgement) of 3 or more 
antihypertensive medications for at least 1 month 
prior to screening and will be required to 
maintain this regimen throughout the Study.  The 
combination of antihypertensive medications 
must be in the following categories:   
a. angiotensin-converting enzyme inhibitor 
(ACEi) or angiotensin II receptor blocker 
(ARB) 
b. beta blocker 
c. calcium channel blocker 
d. non potassium sparing diuretic 
e. alpha-1 blocker 
f. centrally acting sympatholytic agent 
g. direct acting vasodilators (e.g., hydralazine)  
Based on the collective 
safety and laboratory data 
currently available across 
ISIS 757456 clinical 
studies and lack of safety 
signals related to 
hyperkalemia, the 
inclusion criterion has 
been updated to also allow 
patients on potassium 
sparing diuretics.  This 
modification is further 
supported by the updated 
eligibility criterion that 
excludes patients with a 
known history of 
hyperkalemia per 
Investigator judgement.  
Protocol Synopsis 
Section 5.2 
Exclusion Criteria 
5. The use of the following at time of Screening and 
during the course of the study: 
a. Other medications for the treatment of HTN 
(e.g., minoxidil, diazoxide, renin inhibitors) 
b. Medications that may cause hyperkalemia 
unless on a stable dose at least 1 month prior 
to the Screening visit and no known history 
of hyperkalemia per Investigator judgement 
(e.g., cyclosporine or tacrolimus, pentamidine, 
trimethoprim sulfamethoxazole, all heparins) 
c. Use of oral anticoagulants, unless stable for 
4 weeks prior to the first dose of Study Drug 
and regular monitoring must be performed per 
clinical practice during the study unless the 
patient is receiving vitamin K agonists.  If the 
patient is receiving vitamin K antagonists 
(e.g., warfarin) international normalized ratio 
(INR) should be in therapeutic range, as 
established by the Investigator, for 4 weeks 
prior to the first dose 
d. Chronic administration (defined as > 3 days per 
week for the duration of the trial) of NSAIDs or 
Cox-2 inhibitors (except aspirin for 
cardiovascular provided the total daily dose 
does not exceed 325 mg) 
e. Use of phosphodiesterase 5 inhibitors 
(e.g., sildenafil, tadalafil, vardenafil, avanafil) 
within 7 hours prior to any scheduled visit 
f. Potassium sparing diuretics (e.g., eplerenone, 
spironolactone, amiloride, triamterene) 
Intentionally left blank 
The exclusion criterion 5b 
has been updated to allow 
patients who are on 
medications that may 
cause hyperkalemia, but 
do not have hyperkalemia 
at Screening or a history 
of hyperkalemia per 
Investigator judgement.  
This will allow patients 
who have stable, within 
normal range potassium 
levels.  Exclusion 
criterion 5f has been 
removed and added to 
inclusion criterion 6. 


ISIS 757456-CS4 (ASTRAAS) 
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Protocol 
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Protocol Section 
Description of Change 
Rationale 
Protocol Synopsis 
Section 5.2 Exclusion 
Criteria 
8. Active infection with human immunodeficiency 
virus (HIV), hepatitis C (HCV) hepatitis B 
(HBV) (as evidenced by a positive test for 
hepatitis B surface antigen) or hepatitis C 
(HCV) (diagnosed by initial serological testing 
and confirmed with RNA testing) or prior 
treatment for hepatitis C.  Patients at Screening 
who test positive by serology, but negative by 
RNA may be allowed in consultation with the 
Sponsor Medical Monitor or designee 
Correction. HBV diagnosis 
is based on a positive test 
for hepatitis B surface 
antigen. 
12. Unstable/underlying known cardiovascular 
disease defined as: 
a. Any history of congestive heart failure (New 
York Heart Association [NYHA] Class 
III-IV) 
b. Any history of previous myocardial 
infarction, coronary revascularization, 
unstable or stable angina pectoris ˂ 1 year 
prior to screening 
c. 12 lead ECG demonstrating a QT interval 
(corrected using Fridericia's formula [QTcF]) 
˃ 450 msec in males and ˃ 470 msec in 
females at Screening, or a history or 
evidence of long QT syndrome Clinically 
relevant prolonged QTc (using 
Fridericia’s formula [QTcF]) per 
Investigator judgement 
d. Any hemodynamically unstable atrial or 
ventricular arrhythmias 
e. Significant uncorrected valvular heart 
disease  
f. Any history of stroke or transient ischemic 
attack < 1 year prior to screening 
The exclusion criterion was 
updated to allow patients 
with ECG abnormalities, 
aside from those who have 
clinically relevant 
prolonged QTc per 
Investigator judgement and 
are otherwise eligible.  
 


ISIS 757456-CS4 (ASTRAAS) 
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Amendment 4
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Protocol Section 
Description of Change 
Rationale 
Protocol Synopsis 
Section 5.2 
Exclusion Criteria 
16. Screening laboratory results as follows, or any 
other clinically significant abnormalities in 
screening laboratory values that would render a 
patient unsuitable for inclusion 
a. Urine protein-creatinine ratio (UPCR) 
≥ 0.53 mg/mg  In the event of a UPCR above 
this threshold, eligibility may be confirmed 
by a quantitative total urine protein 
measurement of ˂ 300 mg/24 hr 
b. Urine albumin creatinine ratio (UACR) 
˃ 300 mg/g Intentionally left blank 
c. A positive test for blood (including trace) on 
urinalysis that is subsequently confirmed with 
urine microscopy showing ˃ 5 red blood cells 
per high power field and is related to 
glomerulopathies.  In women, this exclusion 
criterion must be assessed outside of 
menstrual period.  If, in the opinion of the 
Investigator, the haematuria is not considered 
related to glomerulopathies the patient may 
be considered eligible, pending proper 
follow-up and a discussion with the Sponsor 
Medical Monitor or designee.  Patients with 
history of bladder cancer must have been 
treated with curative intent and have not 
presented recurrence within the prior 5 years 
d. Alanine aminotransferase (ALT), aspartate 
aminotransferase (AST) > 1.5 × upper limit of 
normal (ULN) 
e. Total bilirubin ≥ 1.5 × ULN (patients with 
total bilirubin ≥ 1.5 × ULN may be allowed 
on study if only indirect bilirubin is elevated, 
ALT/AST is not greater than the ULN, and 
known to have Gilbert’s disease) 
f. Platelet count ˂ 125,000/mm3 
g. Serum potassium ˃ 5.1 mmol/L 
h. Estimated glomerular filtration rate (eGFR) of 
˂ 3045 mL/min/1.73 m2 using the Chronic 
Kidney Disease Epidemiology Collaboration 
formula 
i. Hemoglobin A1c (HbA1c) of ˃ 9% or 
diabetes mellitus type 2 not well-controlled 
based on the Investigator’s judgement   
Based on the collective 
renal safety and laboratory 
data currently available 
across ISIS 757456 clinical 
studies, the UPCR and 
eGFR criteria were 
modified to allow the 
inclusion of patients who 
have a higher UPCR and 
lower eGFR.  The UACR 
criterion has been removed 
as patients with 
macroalbuminuria will still 
be excluded per the UPCR 
threshold for eligibility.  
 


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Protocol Section 
Description of Change 
Rationale 
Section 2.5.2 Risk 
Assessment,  
Table 1 
ISIS 757456 is expected to have less effect on the 
renal RAAS system (less elevation of serum 
potassium) than ACEi, ARB or direct renin 
antagonists 
Most likely to occur in patients with high serum 
potassium.  Eligibility criteria excludes K+ 
> 5.1 mmol/L and patients on medications that 
may cause hyperkalemia unless on a stable dose 
at least 1 month prior to the Screening visit and 
no known history of hyperkalemia per 
Investigator judgement.  
Eligibility criteria exclude patients at enhanced 
risk 
Monitoring and stopping rules in place for 
hyperkalemia 
The language is updated to 
be consistent with the 
revised eligibility criteria. 
Section 3.2 
Number of Study 
Centers 
This study will be conducted at multiple centers in 
the United States and Canada. 
This study will now also 
be conducted at sites in 
Canada. 
Section 6.2 
Laboratory 
Assessments  
Laboratory analyte samples will be collected 
throughout the study.  A list of these analytes is 
contained in Appendix B.   
If the platelet value, serum creatinine eGFR, 
UPCR, potassium, or liver enzyme tests are 
uninterpretable (e.g., due to clumping, hemolysis, 
or quantity not sufficient) or missing, a repeat 
blood specimen should be re drawn as soon as 
possible (ideally within 7 days, either in clinic, by 
Home Healthcare or local laboratory).  Any 
uninterpretable platelet count, serum creatinine 
eGFR, UPCR, liver enzyme, or potassium result 
must be rechecked and determined not to have met 
a stopping rule before dosing can continue.  If 
Day 1 results are uninterpretable, Day 8 dosing 
may proceed, however, an interpretable result 
is required (and must be reviewed) before study 
drug dosing on Day 15.  If there is suspicion of 
EDTA mediated platelet clumping, a repeat 
platelet count should be collected in a sodium 
citrate tube as soon as possible.  Treatment should 
be held if there is no evaluable platelet count 
within 18 days prior to the scheduled dose 
(Section 8.5.2).   
Correction. Added eGFR 
and UPCR tests to align 
with safety monitoring 
rules and clarified how to 
manage uninterpretable 
values after Day 1. 
 


ISIS 757456-CS4 (ASTRAAS) 
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Amendment 4
Protocol 
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Protocol Section 
Description of Change 
Rationale 
Section 8.5.2 
Safety Monitoring 
Rules for Platelet 
Count Results  
Treatment should be held if there is no evaluable 
platelet count within 18 days prior to the scheduled 
dose. Any unreportable platelet count result must 
be rechecked and determined not to have met a 
stopping rule before dosing can continue.  If Day 1 
results are uninterpretable, Day 8 dosing may 
proceed, however, an interpretable result is 
required (and must be reviewed) before study 
drug dosing on Day 15.  
Provide further 
clarification on how to 
manage uninterpretable 
values after Day 1. 
Section 8.6 
Stopping Rules 
For the purposes of the stopping rules, Baseline is 
defined as:  
• Temporary stopping rules for renal function tests
- Baseline is defined as the average of the pre-
dose test closest to Day 1 and Day 1 last non
missing measurement prior to the first dose
• Stopping rules for platelets – Baseline is defined
as the last non missing measurement prior to the
first dose
The definition of baseline 
for renal stopping rules 
was modified in order to 
provide a more accurate 
pre-dose clinical status 
while taking physiological 
variability into 
consideration.  This will 
better indicate if additional 
monitoring or the stopping 
of Study Drug is 
necessary. 
Section 8.6.2 
Stopping Rules for 
Renal Function 
Test Results 
1.Confirmed 30% decline in eGFR from Baseline
eGFR values [eGFR should be calculated using
the CKD-EPI creatinine equation
(Levey et al. 2009)]
2. Confirmed proteinuria (UPCR ≥ 0.7 mg/mg):
Absolute UPCR value ≥ 1000 mg/g
The follow up schedule for any events meeting
either of these stopping criteria will be 
determined by the Investigator in consultation 
with the Sponsor Medical Monitor or designee. 
The possible dosing re-initiation or follow-up 
schedule for any events meeting either of these 
criteria will be determined by the Investigator in 
consultation with the Sponsor Medical Monitor 
or designee. 
At the discretion of the Investigator, a decision 
to hold or permanently stop study drug may be 
made based on lesser changes in these 
parameters observed in isolation or in 
association with other renal-related 
abnormalities. 
The renal function 
monitoring and stopping 
rules were updated to 
align with the modified 
eligibility criteria based on 
the collective renal safety 
and laboratory data 
currently available across 
ISIS 757456 clinical 
studies.  The updated 
language also clarifies the 
guidelines for Study Drug 
reinstitution. 


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Protocol Section 
Description of Change 
Rationale 
Section 8.10.1 
Concomitant Therapy 
The following are disallowed concomitant 
therapies: 
• Other medications for the treatment of HTN 
(e.g., minoxidil, diazoxide, renin inhibitors) 
• Medications that may cause hyperkalemia unless 
on a stable dose at least 1 month prior to the 
Screening visit (e.g., cyclosporine or tacrolimus, 
pentamidine, trimethoprim sulfamethoxazole, all 
heparins) 
• Use of oral anticoagulants unless stable for 4 
weeks prior to the first dose of Study Drug and 
regular monitoring must be performed, per 
clinical practice during the study unless the 
patient is receiving vitamin K agonists.  If the 
patient is receiving vitamin K antagonists (e.g., 
warfarin) INR should be in therapeutic range, as 
established by the Investigator, for 4 weeks prior 
to the first dose 
• Chronic administration (defined as > 3 days per 
week for the duration of the trial) of NSAIDs or 
COX-2 inhibitors (except aspirin for 
cardiovascular disease provided the total daily 
dose does not exceed 325 mg) 
• Use of phosphodiesterase 5 inhibitors (e.g., 
sildenafil, tadalafil, vardenafil, avanafil) within 
72 hours prior to any scheduled visit 
• Potassium sparing diuretics (e.g., eplerenone, 
spironolactone, amiloride, triamterene) 
The disallowed 
concomitant therapies 
were updated to reflect 
the updated exclusion 
criterion. 
Section 9.3.3 Serious 
Adverse Event 
The terms "severe" and "serious" are not 
synonymous.  Severity refers to the intensity of an 
AE (e.g., rated as mild, moderate, or severe, or 
according to National Cancer Institute Common 
Terminology Criteria for Adverse Events [NCI 
CTCAE]; OR Toxicity Grading Scale for Healthy 
Adult and Adolescent Volunteers Enrolled in 
Preventive Vaccine Clinical Trials; the event itself 
may be of relatively minor medical significance 
(such as severe headache without any further 
findings). 
Correction. Severity of 
adverse events and 
serious adverse events 
will be graded based on 
criteria from the 
Common Terminology 
Criteria for Adverse 
Events (CTCAE) and 
not the Toxicity 
Grading Scale for 
Healthy Adult and 
Adolescent Volunteers 
Enrolled in Preventive 
Vaccine Clinical Trials. 
 
 


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Protocol Section 
Description of Change 
Rationale 
Appendix A 
Schedule of Procedures 
Footnote 7 
If the platelet value, serum creatinine, eGFR, UPCR, 
potassium, or liver enzyme tests are uninterpretable 
(e.g., due to clumping, hemolysis or quantity not 
sufficient) a repeat blood specimen; should be 
re-drawn as soon as possible (ideally within 7 days 
either in clinic, by Home Healthcare, or local 
laboratory) and not meet a stopping rule prior to next 
dose.  If Day 1 results are uninterpretable, Day 8 
dosing may proceed, however, an interpretable 
result is required (and must be reviewed) before 
study drug dosing on Day 15.   
Correction. Added eGFR 
and UPCR tests to align 
with safety monitoring 
rules and clarified how 
to manage 
uninterpretable values 
after Day 1. 


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PROTOCOL SYNOPSIS 
Protocol Title 
A Double-Blind, Placebo-Controlled, Phase 2 Study to Assess the Safety, 
Tolerability and Efficacy of IONIS-AGT-LRX, an Antisense Inhibitor of 
Angiotensinogen Production Administered Subcutaneously for 12 Weeks to 
Hypertensive Patients with Uncontrolled Blood Pressure 
Study Phase 
2 
Indication 
Hypertension 
Primary 
Objective 
• 
To evaluate the effect of ISIS 757456 compared to placebo on seated automated 
office systolic blood pressure (SBP) from Baseline to Study Day 85 in 
uncontrolled hypertensive patients on ≥ 3 antihypertensive medications 
Secondary 
Objectives 
• 
To evaluate the effect of ISIS 757456 on plasma angiotensinogen (AGT) at each 
scheduled visit in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• 
To evaluate the effect of ISIS 757456 on 24-hour ambulatory blood pressure at 
Study Day 85 in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• 
To evaluate the effect of ISIS 757456 on seated automated office SBP at each 
scheduled visit in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• 
To evaluate the effect of ISIS 757456 on seated automated office diastolic blood 
pressure (DBP) at each scheduled visit in uncontrolled hypertensive patients on 
≥ 3 antihypertensive medications 
Exploratory 
Objectives 
• 
To evaluate the effects of ISIS 757456 administered subcutaneously (SC) 
(e.g., angiotensin II, renin [Plasma renin activity (PRA); direct renin], urinary 
AGT) in uncontrolled hypertensive patients on ≥ 3 antihypertensive medications 
• 
To evaluate the pharmacokinetics (PK) of ISIS 757456 (as total full length 
ASO, including fully conjugated, partially conjugated, and unconjugated 
ISIS 757456) administered SC in uncontrolled hypertensive patients on 
≥ 3 antihypertensive medications 
• 
To evaluate potential PK / pharmacodynamic (PD) correlation with relevant 
biomarkers and/or clinical endpoints 
Safety Objective • 
To evaluate the safety and tolerability of ISIS 757456 in uncontrolled 
hypertensive patients on ≥ 3 antihypertensive medications 
• 
To evaluate the effects of ISIS 757456 on patient reported quality of life 
outcomes 
Study Design 
Randomized, double blind, placebo controlled, multi-center study 
Number of 
Patients 
Approximately 150 patients will be enrolled 
 
 
 


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PROTOCOL SYNOPSIS (CONTINUED) 
Study 
Population 
Inclusion Criteria 
1. Must have given written informed consent (signed and dated) and any 
authorizations required by local law and be able to comply with all study 
requirements 
2. Males or females aged 18–80 inclusive and weighing ≥ 50 kg at the time of 
informed consent 
3. Satisfy the following: 
a. Females:  must be non-pregnant and non-lactating, and either: 
i. 
surgically sterile (e.g., bilateral tubal occlusion, hysterectomy, bilateral 
salpingectomy, bilateral oophorectomy) 
ii. 
post-menopausal (defined as 12 months of spontaneous amenorrhea in 
females ˃ 55 years of age or, in females ≤ 55 years, 12 months of 
spontaneous amenorrhea without an alternative medical cause and 
follicle stimulating hormone (FSH) levels in the postmenopausal range 
for the laboratory involved) 
b. Males must be abstinent*, surgically sterile (vasectomy with negative semen 
analysis at Follow-up) or if engaged in sexual relations with a woman of 
childbearing potential (WOCBP), a highly effective contraceptive method 
must be used (refer to Section 6.4.1) from the time of signing the informed 
consent form until at least 20 weeks after the last dose of Study Drug 
(ISIS 757456 or placebo) or abstinent* 
* Abstinence is only acceptable as true abstinence, i.e., when this is in line with 
the preferred and usual lifestyle of the patient.  Periodic abstinence 
(e.g., calendar, ovulation, symptothermal, post-ovulation methods), declaration 
of abstinence for the duration of a trial and withdrawal are not acceptable 
methods of contraception. 
4. Body mass index (BMI) ≤ 45.0 kg/m2 
5. Brachial circumference ≥ 22 and ≤ 43 cm (8.7 and 16.9 inches) 
6. At Screening, the patient must have been on a stable, maximally tolerated 
regimen (per Investigator judgement) of 3 or more antihypertensive medications 
for at least 1 month prior to screening and will be required to maintain this 
regimen throughout the Study.  The combination of antihypertensive medications 
must be in the following categories:   
a. angiotensin-converting enzyme inhibitor (ACEi) or angiotensin II receptor 
blocker (ARB) 
b. beta blocker 
c. calcium channel blocker 
d. diuretic 
e. alpha-1 blocker 
f. centrally acting sympatholytic agent 
g. direct acting vasodilators (e.g., hydralazine)  
7. At Screening Visit 1 and on Screening Visit 2 prior to the 24-hr ambulatory 
blood pressure monitoring, the average seated automated office BP (using the 
Sponsor provided BP machine) must be within > 130–≤ 170mmHg systolic and 
meet the Exclusion Criterion #4 Pulse Pressure requirement.  The average is to be 
derived from 3 assessments taken within 10 minutes.  Up to 2 additional tests at 
each time point are allowed in order to qualify 
 
 


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PROTOCOL SYNOPSIS (CONTINUED) 
Study 
Population 
(Continued) 
Inclusion Criteria (Continued) 
8. Agree to abstain from alcoholic beverages for at least 24 hours prior to clinic 
visits 
9. Agree to maintain adequate hydration and adhere to a low sodium diet 
throughout the duration of the study (from the Screening visit onward) 
10. Agree to abstain from exercise, smoking, and caffeine use 30 minutes prior to 
seated automated office BP measurements 
Exclusion Criteria  
1. Clinically significant abnormalities in medical history according to Investigator 
judgement (e.g., major surgery non-inclusive of exclusion criteria 12 within 
2 months of Screening, type 1 diabetes mellitus)  
2. History of secondary hypertension (HTN) including, but not limited to any of the 
following:  renovascular HTN (unilateral or bilateral renal artery stenosis), 
coarctation of the aorta, primary hyperaldosteronism, Cushing’s disease, 
pheochromocytoma, polycystic kidney disease, and drug-induced HTN 
3. Patient with borderline orthostatic hypotension (assessed at Screening Visit 1), 
when they assume a standing position (within >1 to ≤3 minutes of standing up), 
defined as: 
a. A decrease in SBP of ≥ 15 mmHg  
Up to 1 additional test may be done once the patient is adequately hydrated if 
orthostasis is thought to be attributed to volume depletion 
4.  Patient with a pulse pressure at Screening Visit 1 of ˃ 80 mmHg.  Patients with a 
pulse pressure >80 mmHg at Screening Visit 1 may be allowed in consultation 
with the Sponsor Medical Monitor or designee.  Pulse pressure is calculated by 
the difference between the office seated SBP and the office seated DBP (seated 
automated office blood pressure is to be derived by the average of three 
assessments) 
5. The use of the following at time of Screening and during the course of the study:   
a. Other medications for the treatment of HTN (e.g., minoxidil, diazoxide, 
renin inhibitors) 
b. Medications that may cause hyperkalemia unless on a stable dose at least 
1 month prior to the Screening visit and no history of hyperkalemia per 
Investigator judgement  
c. Use of oral anticoagulants, unless stable for 4 weeks prior to the first dose of 
Study Drug and regular monitoring must be performed per clinical practice 
during the study unless the patient is receiving vitamin K agonists.  If the 
patient is receiving vitamin K antagonists (e.g., warfarin) international 
normalized ratio (INR) should be in therapeutic range, as established by the 
Investigator, for 4 weeks prior to the first dose 
d. Chronic administration (defined as > 3 days per week for the duration of the 
trial) of NSAIDs or COX-2 inhibitors (except aspirin for cardiovascular 
disease provided the total daily dose does not exceed 325 mg) 
e. Use of phosphodiesterase 5 inhibitors (e.g., sildenafil, tadalafil, vardenafil, 
avanafil) within 72 hours prior to any scheduled visit 
f. Intentionally left blank 


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PROTOCOL SYNOPSIS (CONTINUED) 
Study 
Population 
(Continued) 
Exclusion Criteria (Continued) 
6. Treatment with another Study Drug, biological agent, or device within 1 month of 
Screening, or 5 half-lives of study agent, whichever is longer 
7. Previous treatment with an oligonucleotide or other RNA therapeutic (including 
small interfering ribonucleic acid [siRNA]) within 4 months of Screening if single 
dose received, or within 12 months of Screening if multiple doses received.  This 
exclusion criterion does not apply to the COVID-19 vaccines which are allowed.  
COVID-19 vaccination performed prior to study participation or during study 
participation will be collected in the electronic database. 
8. Active infection with human immunodeficiency virus (HIV), hepatitis B (HBV) 
(as evidenced by a positive test for hepatitis B surface antigen), or hepatitis C 
(HCV) (diagnosed by initial serological testing and confirmed with RNA testing), 
or prior treatment for hepatitis C.   
9. Malignancy within 5 years, except for basal or squamous cell carcinoma of the 
skin or carcinoma in situ of the cervix that has been successfully treated.  Patients 
with a history of other malignancies that have been treated with curative intent and 
which have no recurrence within 5 years may also be eligible if reviewed by the 
Sponsor Medical Monitor or designee 
10. History of bleeding diathesis, coagulopathy, immune thrombocytopenic purpura 
(ITP), thrombotic cytopenic purpura (TTP), or any qualitative or quantitative 
platelet defect 
11. Recent history of, or current drug or alcohol abuse 
12. Unstable/underlying known cardiovascular disease defined as: 
a. Any history of congestive heart failure (New York Heart Association [NYHA] 
Class III-IV) 
b. Any history of previous myocardial infarction, coronary revascularization, 
unstable or stable angina pectoris ˂ 1 year prior to screening 
c. Clinically relevant prolonged QTc (using Fridericia’s formula [QTcF]) per 
Investigator judgement 
d. Any hemodynamically unstable atrial or ventricular arrhythmias 
e. Significant uncorrected valvular heart disease 
f. Any history of stroke or transient ischemic attack < 1 year prior to screening 


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PROTOCOL SYNOPSIS (CONTINUED) 
Study 
Population 
(Continued) 
Exclusion Criteria (Continued) 
13. A cardiac valve repair, cardiac device implantation, and/or a hospitalization for 
heart failure within 3 months of Screening 
14. Active infection requiring systemic antiviral or antimicrobial therapy that will not 
be completed prior to Study Day 1 
15. Abnormal thyroid function tests at Screening with clinical significance, per 
Investigator judgment.  Patients receiving dose-stable thyroid hormone 
replacement therapy for at least 3 months prior to screening will be allowed to 
participate as long as thyroid tests (TSH/T3/T4) show that patient is euthyroid 
16. Screening laboratory results as follows, or any other clinically significant 
abnormalities in screening laboratory values that would render a patient unsuitable 
for inclusion 
a. Urine protein-creatinine ratio (UPCR) ≥ 0.5 mg/mg   
b. Intentionally left blank 
c. A positive test for blood (including trace) on urinalysis that is subsequently 
confirmed with urine microscopy showing ˃ 5 red blood cells per high-power 
field and is related to glomerulopathies.  In women, this exclusion criterion 
must be assessed outside of menstrual period.  If, in the opinion of the 
Investigator, the haematuria is not considered related to glomerulopathies the 
patient may be considered eligible, pending proper follow-up and a discussion 
with the Sponsor Medical Monitor or designee.  Patients with history of 
bladder cancer must have been treated with curative intent and have not 
presented recurrence within the prior 5 years 
d. Alanine aminotransferase (ALT), aspartate aminotransferase (AST) 
> 1.5 × upper limit of normal (ULN) 
e. Total bilirubin ≥ 1.5 × ULN (patients with total bilirubin ≥ 1.5 × ULN may be 
allowed on study if only indirect bilirubin is elevated, ALT/AST is not greater 
than the ULN, and known to have Gilbert’s disease) 
f. Platelet count ˂ 125,000/mm3 
g. Serum potassium ˃ 5.1 mmol/L 
h. Estimated glomerular filtration rate (eGFR) of ˂ 30 mL/min/1.73 m2 using the 
Chronic Kidney Disease Epidemiology Collaboration formula 
i. Hemoglobin A1c (HbA1c) of ˃ 9% or diabetes mellitus type 2 not 
well-controlled based on the Investigator’s judgement 
 


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PROTOCOL SYNOPSIS (CONTINUED) 
Study 
Population 
(Continued) 
Exclusion Criteria (Continued) 
17. Clinically significant abnormalities upon physical examination which in the 
Investigator’s opinion should exclude the patient from study participation 
18. Patient works nighttime shifts (e.g., 11 PM to 7 AM) 
19. Have any other conditions, which, in the opinion of the Investigator or Sponsor 
would make the patient unsuitable for inclusion, or could interfere with the patient 
participating in or completing the study 
20. Unwilling to comply with study procedures, including follow-up, as specified by 
this protocol, or unwillingness to cooperate fully with the Investigator 
Treatment 
Groups 
Approximately 150 patients will be stratified based on Screening eGFR result of 
˂ 60 or ≥ 60 mL/min/1.73 m2 and randomized to 1 of 2 dose cohorts in a 1:1 ratio 
(Cohort A or Cohort B).  Within each dose cohort, patients will be further randomized 
to receive ISIS 757456 or placebo in a 2:1 ratio. 
Study Drug 
Dosage and 
Administration 
ISIS 757456 (100 mg/mL) and placebo will be supplied in vials of 0.8 mL in a 
stoppered glass vial.  Study Drug (ISIS 757456 or placebo) injection volumes will be 
0.8 mL or 1.2 mL for Cohort A and Cohort B, respectively.  All Study Drug injections 
will be administered by qualified personnel at the clinic or by a Home Healthcare 
Professional. 
Rationale for 
Dose and 
Schedule 
Selection 
The dose levels of 80 mg and 120 mg were selected based on safety, PK, tolerability 
and PD data from the ISIS 757456 Phase 1 study in healthy volunteers and 
ISIS 757456 Phase 2 studies in uncontrolled hypertensive patients (wash out patients 
and patients on 2 to 3 HTN medications).  In the ISIS 757456 Phase 1 study plasma 
AGT decreased in a dose-dependent manner after 6 weeks of 40 mg and 80 mg 
once-weekly doses, and the mean reduction of AGT was approximately 46% and 60% 
(Study Day 43), respectively.  In 2 completed Phase 2 studies in patients with 
controlled and uncontrolled HTN, the mean reduction of AGT was observed up to 
approximately 54% after 6 weeks and 67% after 8 weeks of dosing of 80 mg 
once-weekly doses.  Although neither study was powered for statistical relevant 
reductions in BP, a trend in SBP lowering > 5 mmHg was observed in patients treated 
with ISIS 757456 compared to placebo.  Hence, the 80 mg dose level was selected for 
this study as well.  Exposure-response analysis of the Phase 1 and 2 studies suggested a 
potential for more reductions in plasma AGT with a dose of 120 mg (compared to 
80 mg).  Hence, the dose level of 120 mg was also selected to evaluate plasma AGT 
dynamics and to evaluate further, the subsequent changes in SBP reductions.  In both 
Phase 2 trials, including patients on ACEi or ARB therapy, no protocol stopping rules 
were met for renal, hepatic, platelet, hypotension or hyperkalemia.   
The proposed 120 mg per week dose is also supported by the 3.5-fold margin from the 
no observed adverse effect level (NOAEL) observed at 6 mg/kg/wk in the 9-month 
chronic toxicology study.  
Adjustment of 
Dose and/or 
Treatment 
Schedule 
An adjustment to the dose frequency from weekly to every 2 weeks or a weekly dose 
reduction may be permitted for safety or efficacy reasons in consultation with the 
Sponsor Medical Monitor or designee. 


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PROTOCOL SYNOPSIS (CONTINUED) 
Study Visit 
Schedule and 
Procedures 
In-clinic BP, blood and urine samples will be collected regularly throughout the study 
for safety, efficacy, and exploratory analyses.  Appendix B shows a list of analytes 
required for the study. 
The safety of ISIS 757456 will be monitored in an ongoing fashion throughout the 
trial.   
Screening:  Week -6 to Week -1 
Laboratory and other study procedures will be performed to assess eligibility during 
the Screening Period.  Patients will be given a hydration and low sodium diet 
education at the Screening Visit.   
Treatment:  Week 1 to Week 12 
Eligible patients will be stratified based on Screening eGFR result of ˂ 60 or ≥ 60 
mL/min/1.73 m2 and randomized 1:1:1 to 80 mg ISIS 757456:  120 mg ISIS 757456:  
placebo.  Patients will receive SC doses of Study Drug on Study Days 1, 8, 15, 22, 29, 
36, 43, 50, 57, 64, 71, and 78.  Patients will continue their previous antihypertensive 
medication regimen throughout the study.   
The study site will counsel and ask the patient at each visit if (s)he is maintaining a 
low sodium diet and adequate hydration.  The patient is expected to maintain adequate 
hydration and a low sodium diet throughout the study. 
Patients that discontinue treatment are encouraged to remain in the study for the 
Post-Treatment Evaluation Period and will conduct procedures outlined at Study Day 
85 visit upon discontinuation of Study Drug. 
Post-Treatment:  Week 13 to Week 25 
Patients are to return to the Study Center for follow-up visits on Study Days 85, 92, 
106, 120, 148, and 169.   
The study site will continue to counsel and ask the patient at each visit if (s)he is 
maintaining a low sodium diet and adequate hydration in the Post-Treatment Period.   
The final study visit will be Study Day 169. 
Primary 
Endpoint 
 Change in seated automated office SBP from Baseline to Study Day 85  
Secondary 
Endpoints 
 Absolute levels and change and percent change in plasma AGT from Baseline to 
each scheduled, post-baseline visit 
 Changes from Baseline to Study Day 85 in 24-hour mean SBP and DBP measured 
by ambulatory blood pressure monitoring (ABPM) 
 Percentage of patients reaching the goals of seated automated office seated SBP ≤ 
140 mmHg, DBP ≤ 90 mmHg, and both during the study (excluding patients with 
a baseline SBP of ≤ 140 mmHg) 
 Percentage of patients reaching the goals of automated office seated SBP 
≤ 130 mmHg, DBP ≤ 80 mmHg, and both during the study  
 Change on seated automated office SBP from Baseline to each scheduled, 
post-Baseline visit 
 Change on seated automated office DBP from Baseline to each scheduled, 
post-Baseline visit 


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PROTOCOL SYNOPSIS (CONTINUED) 
Exploratory 
Endpoints 
• Absolute levels and change and percent change of angiotensin II, renin (PRA; 
direct renin), urinary AGT from Baseline to each scheduled, post-Baseline visit 
• Summary of peak and trough ISIS 757456 plasma concentrations  
• Potential exposure-response analysis using relevant exposure parameters and 
biomarkers including, but not limited to trough concentration (Ctrough) and plasma 
AGT may be performed 
• Changes from baseline to Study Day 85 and each scheduled post Baseline 
assessment in the following patient reported outcome questionnaires: EuroQol-5 
Dimension 5 Level (EQ-5D-5L), 12-Item Short Form Survey (SF-12), Power 
Over Pressure- Hypertension Specific Questionnaire (POP-HSQ), Patient Global 
Impression- Change (PGI-C), and Patient Global Impression- Severity (PGI-S) 
Safety 
Endpoint 
• Incidence and severity of treatment-emergent adverse events (TEAE), use of 
concomitant medications, abnormal findings in laboratory assessments, ECG, 
and vital signs 
Statistical 
Considerations 
Approximately 150 patients will be stratified based on Screening eGFR result of 
˂ 60 or ≥ 60 mL/min/1.73 m2 and randomized to 1 of 2 dose cohorts in a 1:1 ratio 
(Cohort A or Cohort B).  Within each dose cohort, patients will be further 
randomized to receive ISIS 757456 or placebo in a 2:1 ratio.  The planned sample 
size will provide approximate 80% of power to demonstrate the primary efficacy 
endpoint of mean SBP reduction difference of 8 mmHg between each ISIS treated 
group and pooled placebo based on the assumption of pooled standard deviation of 
13 mmHg with the two-sided test and a significant level of 0.05 while taking into 
account an estimated 10% drop-out rate.  The overall type I error rate is at 0.1 level, 
two-sided.   
The primary efficacy analysis will be the comparison of change from Baseline to 
Study Day 85 in SBP.  The multiplicity will be controlled by using the sequential 
testing strategy.  
All planned analyses will compare ISIS 757456 to pooled placebo.   
An interim analysis may be conducted after approximately 50% of the patients have 
been enrolled.   
Sponsor 
Ionis Pharmaceuticals, Inc.   




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STUDY GLOSSARY 
Abbreviation 
Definition 
2′-MOE 
2′-O-(2-methoxyethyl) 
ABP 
ambulatory blood pressure 
ABPM 
ambulatory blood pressure monitoring 
ACEi 
angiotensin-converting enzyme inhibitor 
AE 
adverse event 
AGT 
angiotensinogen 
ALP 
alkaline phosphatase 
ALT 
alanine aminotransferase (SGPT) 
aPTT 
activated partial thromboplastin time 
ARB 
angiotensin receptor blockers  
ARF 
acute renal failure 
ASGPR 
asialoglycoprotein receptor  
ASO 
antisense oligonucleotide 
BMI 
body mass index 
BP 
blood pressure 
BUN 
blood urea nitrogen 
CKD 
chronic kidney disease 
CMV 
cytomegalovirus 
CRF 
case report form 
CRNMB 
clinically relevant non-major bleeding 
CT 
computed tomography 
CTCAE 
Common Terminology Criteria for Adverse Events  
Ctrough 
trough concentration 
DBP 
diastolic blood pressure 
ECG 
electrocardiogram 
eCRF 
electronic Case Report Form 
eGFR 
estimated glomerular filtration rate 
EQ-5D-5L 
EuroQoL-5 Dimension 5 Level 
FAS 
full analysis set 
FSH 
follicle stimulating hormone 
GalNAc 
N-acetyl galactosamine 
GCP 
Good Clinical Practice 
HAV 
hepatitis A virus 
HbA1c 
Hemoglobin A1c 
HbsAg 
hepatitis B surface antigen 
HBV 
hepatitis B virus 
HCV 
hepatitis C virus 
HIV 
human immunodeficiency virus 
hs-CRP 
CRP measured by high sensitivity assay 
HTN 
hypertension 
hr(s) 
hour(s) 
ICH 
International Conference on Harmonization 
IgM 
immunoglobulin M 
INR 
international normalized ratio 


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IRB 
Institutional Review Board 
ITP 
immune thrombocytopenic purpura  
IV 
intravenous 
MB 
major bleeding 
MCH 
mean corpuscular hemoglobin 
MCHC 
mean corpuscular hemoglobin concentration 
MCV 
mean corpuscular volume 
NCI CTCAE 
National Cancer Institute Common Terminology Criteria for Adverse 
Events 
MRI 
magnetic resonance imaging 
mRNA 
messenger ribonucleic acid 
NOAEL 
No Observed Adverse Effect Level 
NYHA 
New York Heart Association 
on study 
The patient is ‘on study’ from signing of the informed consent until their 
last study visit 
PD 
pharmacodynamic(s) 
PGI-C 
Patient Global Impression- Change 
PGI-S 
Patient Global Impression- Severity 
PK 
pharmacokinetic(s) 
POP-HSQ 
Power Over Pressure- Hypertension Specific Questionnaire 
PPS 
per protocol set  
PRA 
plasma renin activity 
PT 
prothrombin time 
RAAS 
renin-angiotensin-aldosterone system 
RHTN 
resistant hypertension 
RNase H1 
an ubiquitous endonuclease that specifically hydrolyzes the RNA strand in 
RNA/DNA hybrids 
SAE 
serious adverse event 
SAP 
statistical analysis plan 
SBP 
systolic blood pressure 
siRNA 
small interfering ribonucleic acid 
SC 
subcutaneous(ly) 
SF-12 
12-Item Short Form Survey 
Study Day 1 
defined as the first day Study Drug product is administered to the patient 
Study Drug 
ISIS 757456 or placebo 
SUSAR 
suspected unexpected serious adverse reaction 
TEAE 
treatment-emergent adverse event 
TTP 
thrombotic cytopenic purpura 
UACR 
urine albumin-creatinine ratio 
ULN 
upper limit of normal 
UPCR 
urine protein-creatinine ratio  
WBC 
white blood cell 
WOCBP 
woman of childbearing potential 
 
 


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1. 
OBJECTIVES AND ENDPOINTS 
1.1. 
Objectives 
1.1.1. 
Primary Objective 
• To evaluate the effect of ISIS 757456 compared to placebo on seated automated 
office systolic blood pressure (SBP) from Baseline to Study Day 85 in uncontrolled 
hypertensive patients on ≥ 3 antihypertensive medications 
1.1.2. 
Secondary Objectives 
• To evaluate the effect of ISIS 757456 on plasma angiotensinogen (AGT) at each 
scheduled visit in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate the effect of ISIS 757456 on 24-hour ambulatory blood pressure at 
scheduled visits in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate the effect of ISIS 757456 on seated automated office SBP at each 
scheduled visit in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate the effect of ISIS 757456 on seated automated office diastolic blood 
pressure (DBP) at each scheduled visit in uncontrolled hypertensive patients on 
≥ 3 antihypertensive medications 
1.1.3. 
Safety Objectives 
• To evaluate the safety and tolerability of ISIS 757456 in uncontrolled hypertensive 
patients on ≥ 3 antihypertensive medications 
1.1.4. 
Exploratory Objectives 
• To evaluate the effects of ISIS 757456 administered subcutaneously (SC) 
(e.g., angiotensin II, renin [Plasma renin activity (PRA); direct renin], urinary 
angiotensinogen) in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate the pharmacokinetics (PK) of ISIS 757456 (as total full-length ASO, 
including fully conjugated, partially conjugated, and unconjugated ISIS 757456) 
administered SC in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate potential PK/ pharmacodynamic (PD) correlation with relevant 
biomarkers and/or clinical endpoints 
• To evaluate the effects of ISIS 757456 on patient reported quality of life outcomes 


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1.2. 
Study Endpoints 
1.2.1. 
Primary Endpoint 
• Change in seated automated office SBP from Baseline to Study Day 85  
1.2.2. 
Secondary Endpoints 
• Absolute levels and change and percent change in plasma AGT from Baseline to each 
scheduled, post-Baseline visit 
• Changes from Baseline to Study Day 85 in 24-hour mean SBP and DBP measured by 
ambulatory blood pressure monitoring (ABPM) 
• Percentage of patients reaching the goals of seated automated office seated 
SBP ≤ 140 mmHg, DBP ≤ 90 mmHg, and both during the study (excluding patients 
with a baseline SBP of ≤ 140 mmHg) 
• Percentage of patients reaching the goals of automated office seated 
SBP ≤ 130 mmHg, DBP ≤ 80 mmHg, and both during the study 
• Change in seated automated office SBP from Baseline to each scheduled, 
post-Baseline visit 
• Change in seated automated office DBP from Baseline to each scheduled, 
post-Baseline visit 
1.2.3. 
Safety Endpoint 
• Incidence and severity of treatment-emergent adverse events (TEAE), use of 
concomitant medications, abnormal findings in laboratory assessments, 
electrocardiogram (ECG), and vital signs 
1.2.4. 
Exploratory Endpoints 
• Absolute levels and change and percent change of angiotensin II, renin (PRA; direct 
renin), urinary AGT from Baseline to each scheduled, post-Baseline visit 
• Summary of peak and trough ISIS 757456 plasma concentrations  
• Potential exposure-response analysis using relevant exposure parameters and 
biomarkers including, but not limited to trough concentration (Ctrough) and plasma 
AGT may be performed 
• Changes from Baseline to Study Day 85 and each scheduled post-Baseline 
assessment in the following patient reported outcome questionnaires:  EuroQol-5 
Dimension 5 Level (EQ-5D-5L), 12-Item Short Form Survey (SF-12), Power Over 
Pressure- Hypertension Specific Questionnaire (POP-HSQ), Patient Global 
Impression- Change (PGI-C), and Patient Global Impression- Severity (PGI-S) 


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2. 
BACKGROUND AND RATIONALE 
2.1. 
Overview of Disease 
Hypertension (HTN) is defined as failure to achieve BP goal < 140/90 mmHg.  HTN is a major 
contributor to cardiovascular disease (CVD) morbidity and mortality and chronic kidney disease 
(CKD).  Approximately 1.5 M people in the US have myocardial infarction or stroke annually, 
with ~50% of these major adverse cardiovascular events attributed to HTN (Lawes, Vander 
Hoorn, and Rodgers 2008; Korsnes et al. 2015).  Inadequate BP control can lead to increase 
cardiovascular risk.  Some patients have pseudo-resistance HTN as they are not compliant with 
their medications or they have white coat HTN.  Lowering blood pressure reduces cardiovascular 
(CV) risk including major CV events, CHD, stoke, heart failure, renal failure, all-cause mortality 
(Ettehad et al. 2016).  Providing sustained and controlled blood pressure with weekly or less 
administration could benefit patients inadequately controlled with daily administered therapies.  
Night-time SBP has consistently been shown to be an important predictor of CV risk.  
A 10 mmHg increase in night-time SBP can increase risk of total CV events, stroke and cardiac 
mortality in diabetic patients (Draman et al. 2015). 
Resistant hypertension (RHTN) is defined as failure to achieve BP goal of ˂ 130/80 mmHg in 
patients adherent to adequate doses of ≥ 3 medications (1 of which is diuretic) (Judd and 
Calhoun 2014; Sigmund et al. 2020).  In US alone, 70 million adults have HTN, of which 
12-15% have RHTN.  Among these patients 33% of them have uncontrolled RHTN (Judd and 
Calhoun 2014).  In an analysis of National Health and Nutrition Examination Survey database, 
these patients are more likely to be black, with diabetes, with CKD Stage 3, with proteinuria and 
congestive heart failure compared to patients with HTN and without resistant HTN.   
2.2. 
Therapeutic Rationale 
AGT is the source of all downstream angiotensin metabolites.  Angiotensin II (ang II) is the 
principal angiotensin of the RAAS cascade (Figure 1) that elicits a multitude of effects that 
impact the cardiovascular and renal systems.  Ang II acts via multiple mechanisms to 
contribute to hypertension and related disease states such as heart failure and CKD.  Such 
mechanisms include vasoconstriction, renal sodium reabsorption, cell proliferation and 
dedifferentiation, aldosterone secretion and activation of pro-fibrotic and reactive oxygen species 
pathways (Carey 2013; Te Riet et al. 2015).  AGT is acted upon by renin and as such is the only 
known substrate for renin.  Renin cleavage of AGT generates the N-terminal derived 
decapeptide, angiotensin I (Ang I), and the remaining protein des(AngI)-AGT.  Renin cleavage 
of AGT is the rate-limiting step in Ang II formation, as Ang I is converted to Ang II by ACE in a 
substrate-limited reaction.   




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2.3. 
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2.3.3. 
Preclinical Experience 
Detailed information concerning the preclinical studies conducted with ISIS 757456 can be 
found in the Investigator’s Brochure.   
2.3.4. 
Clinical Experience 
Detailed information concerning the clinical studies conducted with ISIS 757456 can be found in 
the Investigator’s Brochure.  A summary is included below. 
The safety and tolerability of ISIS 757456 was evaluated in a total of 62 healthy volunteer 
subjects aged 18–60 in a blinded, placebo-controlled, Phase 1 study ISIS 757456-CS1.  Of these 
62 subjects, 46 received ISIS 757456 (29 in the single-dose cohorts and 17 in the multiple-dose 
cohorts), and 16 received placebo.  ISIS 757456 was well-tolerated at doses up to 80 mg 
administered as a single dose and 480 mg total dose (80 mg administered as 6 doses over 
6 weeks).  ISIS 757456 demonstrated dose-dependent plasma reduction of AGT levels following 
multiple doses, and there were no dose-dependent clinically meaningful trends in laboratory 
assessments and no serious adverse events (SAE).  The Phase 1 study did not result in 
hypotension, hyperkalemia or renal changes.  The elimination half-life of ISIS 757456 was 
estimated to be approximately 2-4 weeks.   
ISIS 757456-CS2 was a Phase 2 multicenter, double-blind, placebo-controlled study to assess the 
safety, tolerability and efficacy of ISIS 757456 in patients with controlled BP.  The study 
enrolled 25 patients randomized 2:1 (ISIS 757456 to placebo) to receive 80 mg ISIS 757456 or 
placebo for 6 weeks.  The patients were washed out of their antihypertensive medications prior to 
Study Day 1 and the BP was confirmed to be uncontrolled after washout.  A 54% mean reduction 
of AGT was observed at an 80 mg weekly dose over 6 weeks.  A trend in blood pressure 
lowering was observed in patients treated with ISIS 757456 compared to placebo.  Mean systolic 
blood pressure was reduced 7.5 mmHg in patients treated with ISIS 757456 and 1.7 mmHg in 
placebo patients.  No rebound hypertension was observed.  ISIS 757456 was well-tolerated and 
there were no clinically meaningful trends in laboratory assessments, including hypotension and 
hyperkalemia, no adverse events of special interest (AESI), and no related SAEs (Morgan et al. 
In Press).   
ISIS 757456-CS3 was a Phase 2 multicenter, double-blind, placebo-controlled study to assess the 
safety, tolerability and efficacy of ISIS 757456 in hypertensive patients with uncontrolled BP.  
The patients continued a stable regimen of 2-3 antihypertensives, one of which was an ACEi or 
ARB, throughout the duration of the study.  The study enrolled 26 patients randomized 2:1 
(ISIS 757456 to placebo) to receive 80 mg ISIS 757456 or placebo.  The Study Drug was 
administered once weekly for 8 weeks with a loading dose on Study Day 3.  A 67% mean 
reduction of AGT was observed after the 8 weeks of dosing.  A trend in blood pressure lowering 
was observed in patients treated with ISIS 757456 compared to placebo.  Mean systolic blood 
pressure was reduced 12.8 mmHg in patients treated with ISIS 757456 and 4.6 mmHg in placebo 
patients.  No rebound hypertension was observed.  ISIS 757456 was well-tolerated in 
hypertension patients taking 2 or 3 hypertension medication including an ACEi or an ARB and 


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there were no clinically meaningful trends in laboratory assessments, including hypotension and 
hyperkalemia, no AESIs, and no SAEs  (Morgan et al. In Press).   
2.5. 
Benefit-Risk Assessment 
Detailed information concerning the benefit-risk assessment of ISIS 757456 can be found in the 
Investigator’s Brochure.   
2.5.1. 
Benefit Assessment 
This compound has been administered to healthy volunteers, hypertensive patients washed out of 
their antihypertensive medications, and hypertensive patients on antihypertensive medications.  
As a GalNAc, specifically due to the mediated uptake to the hepatocytes, ISIS 757456 will 
primarily affect circulating AGT rather than decrease renal AGT transcription.  Therefore, it has 
the potential of an improved therapeutic index due to the potential of lower occurrence of 
hyperkalemia and ARF, which are issues with aggressive RAAS blockage in some subjects that 
may be due, in part, to excessive inhibition of intrarenal Ang II signaling.  Based on its MOA of 
blocking the RAAS pathway it is expected to lower the BP in patients with HTN.  While there 
are other compounds (ACEi, ARB, aldosterone blockers, renin inhibitors) that block different 
components of this pathway, this compound blocks AGT, the most upstream factor in the 
pathway.  Finally, due to its estimated half-life of 2 to 4 weeks, ISIS 757456 may have a superior 


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duration of action compared to RAAS blockers thereby contributing to the potential of 
ISIS 757456 having an enhanced pharmacological benefit relative to the standard of care.   
2.5.2. 
Risk Assessment 
The known potential risks to study participants associated with ISIS 757456 are elaborated on in 
the Guidance for the Investigator section of the Investigator’s Brochure.  Additional study 
associated risks are: 
Table 1: 
Known Risks Associated with RAAS Blockade 
Potential Risk 
of Clinical 
Significance 
Summary of Data/Rationale 
for Risk 
Mitigation Strategy 
Hypotension 
RAAS blockade may lower BP 
Duration of effect:  Long 
half-life of the ASO 
Extent of effect:  AGT is the 
most upstream molecule in 
RAAS pathway 
Eligibility criteria exclude patients at enhanced risk 
and Weekly BP monitoring 
Monitoring and stopping rules in place for 
hypotension 
Hyperkalemia 
RAAS blockade is known to 
increase serum potassium 
ISIS 757456 is expected to have less effect on the 
renal RAAS system (less elevation of serum 
potassium) than ACEi, ARB or direct renin 
antagonists 
Most likely to occur in patients with high serum 
potassium.  Eligibility criteria excludes 
K+ > 5.1 mmol/L and patients on medications that 
may cause hyperkalemia unless on a stable dose at 
least 1 month prior to the Screening visit and no 
known history of hyperkalemia per Investigator 
judgement. 
Eligibility criteria exclude patients at enhanced risk 
Monitoring and stopping rules in place for 
hyperkalemia 
ARF 
RAAS blockade is known to 
cause ARF 
Based on liver (hepatocyte) targeting via the GalNAc 
conjugation, ISIS 757456 is expected to have less 
effect on the renal RAAS system than ACEi, ARB or 
direct renin antagonists 
Monitor renal function closely 
 


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3. 
EXPERIMENTAL PLAN 
3.1. 
Study Design 
This will be a Phase 2, double-blind, randomized, placebo-controlled study of ISIS 757456 
conducted in uncontrolled hypertensive patients on ≥ 3 antihypertensive medications (see 
Section 5.1 Inclusion Criteria).  Patients will be stratified based on Screening eGFR result of 
< 60 or ≥ 60 mL/min/1.73 m2 (see Section 10.1). 
The 80 mg and 120 mg once-weekly doses are designed to assess the safety, tolerability, and 
efficacy of Study Drug (ISIS 757456 or placebo) administered over a 12-week period.   
All patients will complete a 13-week Post-Treatment Period. 
ISIS 757456 (100 mg/mL) and placebo will be supplied in vials of 0.8 mL in a stoppered glass 
vial.  Study Drug (ISIS 757456 or placebo) injection volumes will be 0.8 mL or 1.2 mL for 
Cohort A and Cohort B, respectively.  All Study Drug injections will be administered by 
qualified personnel at the clinic or by a Home Healthcare Professional (as arranged with the site 
staff and after appropriate training).  Refer to Section 8.1 for additional details. 
3.2. 
Number of Study Centers 
This study will be conducted at multiple centers in the United States and Canada. 
3.3. 
Number of Patients 
Approximately 150 patients are planned to be enrolled in this study.   
3.4. 
Overall Study Duration and Follow-up 
The study will consist of Screening (up to 6 weeks), Treatment (12 weeks), and Post-Treatment 
Periods (13 weeks).  Please refer to the Schedule of Procedures in Appendix A.   
Patients may be required to attend additional visits for monitoring of adverse events (AEs) or 
abnormal investigation results.  The frequency of additional monitoring will be determined by 
the Study Medical Monitor in consultation with the Investigator. 
3.4.1. 
Screening 
Patient eligibility for the study will be determined within 6 weeks prior to study entry 
(Study Day -42 to -1). 
3.4.2. 
Treatment 
Patients will be stratified based on based on Screening eGFR result of ˂ 60 or 
≥ 60 mL/min/1.73 m2 (see Section 10.1).  Patients will receive a total of 12 SC doses of Study 
Drug once weekly for 12 weeks (Study Days 1, 8, 15, 22, 29, 36, 43, 50, 57, 64, 71, 78). 
3.4.3. 
Post-Treatment 
Patients are to return to the Study Center for Post-Treatment visits on Study Days 85, 92, 106, 
120, 148, and 169.  The final study visit will be Study Day 169. 


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3.5. 
End-of-Study 
The End-of-Study is defined as the date of the last visit of the last patient.   
4. 
PATIENT ENROLLMENT 
4.1. 
Screening 
Before patients may be enrolled into the study, the Sponsor requires a copy of the Study Center’s 
written institutional review board (IRB) approval of the protocol, informed consent form, and all 
other patient information and/or recruitment material. 
Patients must sign the consent form before any screening tests or assessments are performed.  At 
the time of consent, the patient 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 randomization, patients will be assigned a unique patient 
identification number.  This number will be used to identify the patient throughout the trial and 
must be used on all study documentation related to that patient.  The screening number and 
patient identification number must remain constant throughout the entire trial.  Screening 
numbers and patient identification numbers, once assigned, will not be re-used. 
4.2. 
Randomization  
Patients will be randomized at Study Day 1, after all assessments have been completed and after 
the Investigator has verified that they are eligible per criteria in Section 5.1 and Section 5.2.  No 
patient may begin treatment prior to randomization and assignment of a unique patient 
identification number. 
Patients will be stratified based on Screening eGFR result of ˂ 60 or ≥ 60 mL/min/1.73 m2 
(see Section 10.1) and then patients will be randomized to 1 of 2 dose cohorts in a 1:1 ratio 
(Cohort A or Cohort B).  Within each dose cohort, patients will be further randomized to receive 
ISIS 757456 or placebo in a 2:1 ratio. 
The Sponsor or designee will prepare the randomization list and utilize an automated IRT 
(Interactive Response Technology) system. 
4.3. 
Replacement of Patients 
Patients who withdraw from the study will not be replaced. 
4.4. 
Unblinding of Treatment Assignment 
The Sponsor and all patients, monitors, and Study Center personnel related to the study, will be 
blinded throughout the study.  However, if a patient has suffered a SAE (as defined in 
Section 9.3.3), and/or when knowledge of the treatment assignment will impact the clinical 
management of the patient, the Investigator will have the ability to unblind the treatment 
assignment for that patient.  The Sponsor or designee will be informed of the unblinding of a 
patient within 24 hours.  An unblinded randomization schema will be maintained securely at the 


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Sponsor’s designated vendor.  In addition, all SUSARs will be unblinded by the Sponsor or 
designee for the purpose of regulatory reporting (see Section 9.2). 
Every reasonable attempt should be made to complete the early termination study procedures and 
observations (see Appendix A and Appendix B) prior to unblinding, as knowledge of the 
treatment arm could influence patient assessment. 
An unblinded interim analysis may be performed and the results summarized by cohort after 
approximately 50% of the patients have been enrolled.   
5. 
PATIENT ELIGIBILITY 
To be eligible to participate in this study candidates must meet the following eligibility criteria at 
the time point specified in the individual eligibility criterion listed. 
The Sponsor/Designee may be consulted if any questions arise regarding the inclusion or 
exclusion criteria.   
5.1. 
Inclusion Criteria 
1. Must have given written informed consent (signed and dated) and any authorizations 
required by local law and be able to comply with all study requirements 
2. Males or females aged 18–80 inclusive and weighing ≥ 50 kg at the time of informed 
consent 
3. Satisfy the following: 
a. Females:  must be non-pregnant and non-lactating, and either: 
i. surgically sterile (e.g., bilateral tubal occlusion, hysterectomy, bilateral 
salpingectomy, bilateral oophorectomy) or 
ii. post-menopausal (defined as 12 months of spontaneous amenorrhea in females 
> 55 years of age or, in females ≤ 55 years, 12 months of spontaneous amenorrhea 
without an alternative medical cause and follicle stimulating hormone (FSH) 
levels in the postmenopausal range for the laboratory involved) 
b. Males must be abstinent*, surgically sterile (vasectomy with negative semen analysis 
at follow-up) or if engaged in sexual relations with a woman of childbearing potential 
(WOCBP), a highly effective contraceptive method must be used (refer to 
Section 6.4.1) from the time of signing the informed consent form until at least 
20 weeks after the last dose of Study Drug (ISIS 757456 or placebo) or abstinent* 
* Abstinence is only acceptable as true abstinence, i.e., when this is in line with the 
preferred and usual lifestyle of the patient.  Periodic abstinence (e.g., calendar, 
ovulation, symptothermal, post-ovulation methods), declaration of abstinence for the 
duration of a trial and withdrawal are not acceptable methods of contraception. 
4. body mass index (BMI) ≤ 45.0 kg/m2 
5. Brachial circumference ≥ 22 and ≤ 43 cm (8.7 and 16.9 inches) 


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6. At Screening, the patient must have been on a stable, maximally tolerated regimen 
(per Investigator judgement) of 3 or more antihypertensive medications for at least 
1 month prior to screening and will be required to maintain this regimen throughout the 
Study.  The combination of antihypertensive medications must be in the following 
categories: 
a. angiotensin-converting enzyme inhibitor (ACEi) or angiotensin II receptor blocker 
(ARB) 
b. beta blocker 
c. calcium channel blocker 
d. diuretic 
e. alpha-1 blocker 
f. centrally acting sympatholytic agent 
g. direct acting vasodilators (e.g., hydralazine) 
7. At Screening Visit 1 and on Screening Visit 2 prior to the 24-hr ambulatory blood 
pressure monitoring, the average seated automated office BP (using the Sponsor provided 
BP machine) must be within ˃ 130 – ≤ 170 mmHg systolic and meet the Exclusion 
Criterion 4 Pulse Pressure requirement.  The average is to be derived from 3 assessments 
taken within 10 minutes.  Up to 2 additional tests at each time point are allowed in order 
to qualify 
8. Agree to abstain from alcoholic beverages for at least 24 hours prior to clinic visits 
9. Agree to maintain adequate hydration and adhere to a low sodium diet throughout the 
duration of the study (from the Screening visit onward) 
10. Agree to abstain from exercise, smoking, and caffeine use 30 minutes prior to seated 
automated office BP measurements 
5.2. 
Exclusion Criteria 
1. Clinically significant abnormalities in medical history according to Investigator 
judgement (e.g., major surgery non inclusive of exclusion criteria 12 within 2 months of 
Screening, type 1 diabetes mellitus)  
2. History of secondary HTN including, but not limited to any of the following: 
renovascular HTN (unilateral or bilateral renal artery stenosis), coarctation of the aorta, 
primary hyperaldosteronism, Cushing's disease, pheochromocytoma, polycystic kidney 
disease, and drug-induced HTN 
3. Patient with borderline orthostatic hypotension (assessed at Screening Visit 1), when they 
assume a standing position (within > 1 to ≤ 3 minutes of standing up), defined as: 
a. A decrease in SBP of ≥ 15 mmHg  
Up to 1 additional test may be done once the patient is adequately hydrated if orthostasis 
is thought to be attributed to volume depletion  


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4. Patient with a pulse pressure at Screening Visit 1 of ˃ 80mmHg.  Patients with a pulse 
pressure >80 mmHg at Screening Visit 1 may be allowed in consultation with the 
Sponsor Medical Monitor or designee.  Pulse pressure is calculated by the difference 
between the office seated SBP and the office seated DBP (seated automated office blood 
pressure is to be derived by the average of 3 assessments) 
5. The use of the following at time of Screening and during the course of the study: 
a. Other medications for the treatment of HTN (e.g., minoxidil, diazoxide, renin 
inhibitors) 
b. Medications that may cause hyperkalemia unless on a stable dose at least 1 month 
prior to the Screening visit and no history of hyperkalemia per Investigator judgement 
c. Use of oral anticoagulants, unless stable for 4 weeks prior to the first dose of Study 
Drug and regular monitoring must be performed per clinical practice during the study 
unless the patient is receiving vitamin K agonists.  If the patient is receiving vitamin 
K antagonists (e.g., warfarin) international normalized ratio (INR) should be in 
therapeutic range, as established by the Investigator, for 4 weeks prior to the first dose 
d. Chronic administration (defined as > 3 days per week for the duration of the trial) of 
NSAIDs or Cox-2 inhibitors (except aspirin for cardiovascular provided the total 
daily dose does not exceed 325 mg) 
e. Use of phosphodiesterase 5 inhibitors (e.g., sildenafil, tadalafil, vardenafil, avanafil) 
within 72 hours prior to any scheduled visit 
f. Intentionally left blank 
6. Treatment with another Study Drug, biological agent, or device within 1 month of 
Screening, or 5 half-lives of study agent, whichever is longer 
7. Previous treatment with an oligonucleotide or other RNA therapeutic (including small 
interfering ribonucleic acid [siRNA]) within 4 months of Screening if single dose 
received, or within 12 months of Screening if multiple doses received.  This exclusion 
criteria does not apply to the COVID-19 vaccines which are allowed.  COVID-19 
vaccination performed prior to study participation or during study participation will be 
collected in the electronic database 
8. Active infection with human immunodeficiency virus (HIV), hepatitis B virus (HBV) (as 
evidenced by a positive test for hepatitis B surface antigen), or hepatitis C (HCV) 
(diagnosed by initial serological testing  and confirmed with RNA testing) or prior 
treatment for hepatitis C.   
9. Malignancy within 5 years, except for basal or squamous cell carcinoma of the skin or 
carcinoma in situ of the cervix that has been successfully treated.  Patients with a history 
of other malignancies that have been treated with curative intent and which have no 
recurrence within 5 years may also be eligible if reviewed by the Sponsor Medical 
Monitor or designee 
10. History of bleeding diathesis, coagulopathy, immune thrombocytopenic purpura (ITP), 
thrombotic cytopenic purpura (TTP), or any qualitative or quantitative platelet defect 


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11. Recent history of, or current drug or alcohol abuse 
12. Unstable/underlying known cardiovascular disease defined as: 
a. Any history of congestive heart failure (New York Heart Association [NYHA] 
Class III-IV) 
b. Any history of previous myocardial infarction, coronary revascularization, unstable or 
stable angina pectoris ˂ 1 year prior to screening 
c. Clinically relevant prolonged QTc (using Fridericia’s formula [QTcF]) per 
Investigator judgement 
d. Any hemodynamically unstable atrial or ventricular arrhythmias 
e. Significant uncorrected valvular heart disease  
f. Any history of stroke or transient ischemic attack < 1 year prior to screening  
13. A cardiac valve repair, cardiac device implantation, and/or a hospitalization for heart 
failure within 3 months of Screening 
14. Active infection requiring systemic antiviral or antimicrobial therapy that will not be 
completed prior to Study Day 1 
15. Abnormal thyroid function tests at Screening with clinical significance, per Investigator 
judgement.  Patients receiving dose-stable thyroid hormone replacement therapy for at 
least 3 months prior to screening will be allowed to participate as long as thyroid tests 
(TSH/T3/T4) show that patient is euthyroid 
16. Screening laboratory results as follows, or any other clinically significant abnormalities 
in screening laboratory values that would render a patient unsuitable for inclusion 
a. Urine protein-creatinine ratio (UPCR) ≥ 0.5 mg/mg   
b. Intentionally left blank 
c. A positive test for blood (including trace) on urinalysis that is subsequently confirmed 
with urine microscopy showing ˃ 5 red blood cells per high power field and is related 
to glomerulopathies.  In women, this exclusion criterion must be assessed outside of 
menstrual period.  If, in the opinion of the Investigator, the haematuria is not 
considered related to glomerulopathies the patient may be considered eligible, 
pending proper follow-up and a discussion with the Sponsor Medical Monitor or 
designee.  Patients with history of bladder cancer must have been treated with 
curative intent and have not presented recurrence within the prior 5 years 
d. Alanine aminotransferase (ALT), aspartate aminotransferase (AST) > 1.5 × upper 
limit of normal (ULN) 
e. Total bilirubin ≥ 1.5 × ULN (patients with total bilirubin ≥ 1.5 × ULN may be 
allowed on study if only indirect bilirubin is elevated, ALT/AST is not greater than 
the ULN, and known to have Gilbert’s disease) 
f. Platelet count ˂ 125,000/mm3 
g. Serum potassium ˃ 5.1 mmol/L 


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h. Estimated glomerular filtration rate (eGFR) of ˂ 30 mL/min/1.73 m2 using the 
Chronic Kidney Disease Epidemiology Collaboration formula 
i. Hemoglobin A1c (HbA1c) of ˃ 9% or diabetes mellitus type 2 not well-controlled 
based on the Investigator’s judgement   
17. Clinically significant abnormalities upon physical examination which in the 
Investigator’s opinion should exclude the patient from study participation 
18. Patient works nighttime shifts (e.g., 11 PM to 7 AM) 
19. Have any other conditions, which, in the opinion of the Investigator or Sponsor would 
make the patient unsuitable for inclusion, or could interfere with the patient participating 
in or completing the study 
20. Unwilling to comply with study procedures, including follow-up, as specified by this 
protocol, or unwillingness to cooperate fully with the Investigator 
6. 
STUDY PROCEDURES 
6.1. 
Study Schedule 
All required study procedures are listed by visit in Appendix A, Appendix B and Appendix C. 
6.1.1. 
Screening 
Written informed consent for the study will be obtained prior to the performance of any 
study-related procedures including screening procedures.  A 6-week period is provided for 
completing screening assessments and determining patient eligibility for the study.  Safety labs 
may be re-tested up to 2 additional times for determination of patient eligibility. 
During the Screening Period, patients will undergo a medical history (including cardiovascular 
disease risk factors) and physical examination including vital signs, orthostatic hypotension 
assessment, 12-lead ECG, and have blood and urine samples taken for clinical laboratory testing.  
Patients will be screened for HIV, HepB, and HCV. 
During the Screening Period the study site will also educate patients regarding adequate 
hydration and low sodium diet.   
A 24-hour blood pressure (BP) monitoring will be performed during the Screening Period. 
The study site will record basic personal details, including name, contact details, gender, height, 
weight, date of birth, age, ethnicity, and racial origin (to be used only for clinical purposes), as 
well as information on medical history, and clinical data collected about the patient’s 
participation in the study. 
6.1.1.1. 
24-Hour Ambulatory Blood Pressure Monitoring 
A 24-hour ABPM, conducted with a Sponsor provided machine, will be assessed during the 
Screening Period and at one additional time point for patients who have received 8 or more doses 
as detailed in the Appendix A Schedule of Procedures.   


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The assessment should take place after the Screening Visit 1 blood pressure, pulse pressure, and 
labs have been resulted, the results are deemed not to be exclusionary, and blood pressure is 
confirmed not to be exclusionary at Screening Visit 2.  Ensure sufficient time during the 
Screening Period to allow for a repeat (if the first 24-hr monitoring is not successful) prior to the 
Study Day 1 visit.   
For patients who have been administered 8 or more doses of Study Drug,  24-hr ABPM should 
ideally be initiated in-clinic after all other study procedures have been completed either on Study 
Day 78 or Day 85 (completed 12 doses) or Treatment Early Termination/Study Day 85 
(completed 8-11 doses).  If a repeat is necessary, the 24-hr ABPM should be initiated no later 
than the Study Day 92 visit.  Ensure the 24-hr ABPM is not done concurrently with other study 
procedures.   
Please refer to the Study Manual for additional details. 
6.1.1.2. 
Seated Automated Office Blood Pressure Measuring Instructions at Each Visit 
The seated automated office blood pressure monitoring is to be conducted with Sponsor provided 
blood pressure machines throughout the study.  
Patient Instructions 
• Patient should have abstained from smoking, exercise and caffeine use at least 
30 minutes prior to BP measurements 
• Patient should be wearing a loose, short-sleeved top.  If patient is wearing a 
long-sleeved or tight garment around the arm, provide a gown or remove the arm 
from the sleeve 
• Patient should be comfortably seated (e.g., with his/her back against the chair, feet 
flat on the floor) 
• Patient’s arm should be bent at the elbow and supported by a table 
Observer Instructions 
• Confirm with that the patient has abstained from smoking, exercise and caffeine use 
at least 30 minutes prior to measurement 
• Place the seated automated office blood pressure cuff on the patient’s upper arm at 
the level of the heart and centered at the midpoint of the humerus 
• Leave patient alone for 5 minutes in a quiet setting 
• After 5 minutes, return and take 3 readings, with approximately 2-3 minutes between 
readings 
• Document the average of the 3 readings 
Screening Visit 1 BP Results 
At the Screening Visit 1, measure the BP per instructions above with both the right and left arm.  
The arm with the highest SBP (that meets inclusion/exclusion criteria) and meets the Pulse 
Pressure criteria should be used at each subsequent visit. 




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All safety data including AEs, BP, and concomitant medications will be reviewed by the 
Sponsor’s Medical Monitor or designee on an ongoing basis throughout the trial  
6.1.3. 
Post-Treatment Period 
After the last dose (Study Day 78) or last dose for early termination patients, patients will return 
to the clinic once weekly for the first 2 weeks (Study Days 85, 92) and then on Study Days 106, 
120, 148, and 169 for safety assessments.   
The study site will continue to counsel and ask the patient at each visit if (s)he is maintaining 
adequate hydration and a low sodium diet in the Post-Treatment Period.   
For patients who have been administered 8 or more doses of Study Drug, the second 24-hr 
ABPM will be conducted.  This 24-hr APBM should ideally be initiated in-clinic after all other 
study procedures have been completed either on Study Day 78 or Treatment Early 
Termination/Study Day 85.  If a repeat is necessary, the 24-hr ABPM should be initiated no later 
than the Study Day 92 visit.  Ensure the 24-hr ABPM is not done concurrently with other study 
procedures.   
All safety data including AEs, in-clinic BP, and concomitant medications will be reviewed by the 
Sponsor’s Medical Monitor or designee on an ongoing basis throughout the trial. 
Of the 6 required study visits during the Post-Treatment Period, 4 selected study visits have the 
option to be performed outside the clinic by a Sponsor selected Home Health Care professional.  
Home Healthcare assessments and procedures may be conducted by a Home Healthcare 
professional at specified visits per the Schedule of Procedures (Appendix A) and as arranged by 
site staff.  If 3 consecutive Home Healthcare visits have been conducted, the Sponsor must be 
consulted prior to the scheduling of a 4th in-home visit.  If a visit cannot be performed in the 
clinic or by Home Health Care, patient contact (e.g., video call, text, email, etc.) is required by 
site personnel to assess any AEs, changes in concomitant medications, adequate hydration and 
low sodium diet counseling. 
6.2. 
Laboratory Assessments  
Laboratory analyte samples will be collected throughout the study.  A list of these analytes is 
contained in Appendix B.   
If the platelet value, eGFR, UPCR, potassium, or liver enzyme tests are uninterpretable (e.g., due 
to clumping, hemolysis, or quantity not sufficient) or missing a repeat blood specimen should be 
re-drawn as soon as possible (ideally within 7 days, either in clinic, by Home Healthcare or local 
laboratory).  Any uninterpretable platelet count, eGFR, UPCR, liver enzyme, or potassium result 
must be rechecked and determined not to have met a stopping rule before dosing can continue.  
If Day 1 results are uninterpretable, Day 8 dosing may proceed, however, an interpretable result 
is required (and must be reviewed) before study drug dosing on Day 15.  If there is suspicion of 
EDTA mediated platelet clumping, a repeat platelet count should be collected in a sodium citrate 
tube as soon as possible.  Treatment should be held if there is no evaluable platelet count within 
18 days prior to the scheduled dose (Section 8.5.2).   


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6.3. 
Patient Reported Outcomes 
The assessments listed in the sections below should be, ideally, performed prior to any 
interventional study procedures, such as blood draws and vital signs.  Appropriate instructions 
will be provided to the patient by site staff.  
6.3.1. 
Patient Global Impression Items  
The Patient Global Impression- Change (PGI-C) is a questionnaire which asks the patient to rate 
the perceived level of improvement (or worsening) in the patient’s hypertension since the start of 
study treatment, using categorical response options.  The Patient Global Severity- Change 
(PGI-S) is a questionnaire which asks the patient to rate his or her current hypertension disease 
severity, using categorical response options.  The assessments may take approximately 3 to 
5 minutes each to complete.  
6.3.2. 
EuroQol-5 Dimension 5 Level (EQ-5D-5L) 
The EQ-5D-5L is a two-part, self-reported instrument that assesses the patient’s health state.  
The patient must self-rate their level of severity for five dimensions that include mobility, 
self-care, usual activities, pain/discomfort, and anxiety/depression.  In a second part of the 
assessment, the patient must report their perceived health on a visual analog scale ranging from 
0 to 100.  The questionnaire may take approximately 3 to 5 minutes to complete.  
6.3.3. 
12-Item Short Form Survey (SF-12) 
Patients will be asked to complete the SF-12, which is a set of generic, coherent, and easily 
administered quality of life measures.  The SF-12 consists of 12 questions that measure 8 health 
domains:  general health, physical functioning, role physical, bodily pain, vitality, social 
functioning, role emotional, and mental health.  The assessment should take approximately 3 to 
5 minutes to complete.  
6.3.4. 
Power Over Pressure- Hypertension Specific Questionnaire (POP-HSQ) 
The POP-HSQ is a disease specific patient reported questionnaire which assesses the patient’s 
severity and frequency of nonspecific symptoms associated with hypertension and mental health 
as impacted by hypertension.  The assessment may take approximately 15 minutes to complete.   
6.4. 
Restriction on the Lifestyle of Patients 
6.4.1. 
Contraception Requirements 
All male and female patients must refrain from sperm/egg donation from the time of signing the 
informed consent until at least 20 weeks after the patient’s last dose of study treatment.   
Males must be abstinent†, or if engaged in sexual relations with a woman of childbearing 
potential, highly effective contraception must be used from the time of signing the informed 
consent until at least 20 weeks after the patient’s last dose of study treatment. 
For the purposes of this study, WOCBP are defined as any female who has experienced 
menarche, and who does not meet one of the following conditions: 


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• Postmenopausal:  12 months of spontaneous amenorrhea in females ˃ 55 years of age 
or, in females ≤ 55 years, 12 months of spontaneous amenorrhea without an 
alternative medical cause and FSH levels in the postmenopausal range for the 
laboratory involved 
• 6 weeks after surgical bilateral oophorectomy with or without hysterectomy 
• Post-hysterectomy 
For the purposes of the study, highly effective contraception is defined as follows: 
• For males:  surgical sterilization (vasectomy with negative semen analysis at 
follow-up), a surgically sterile non-pregnant female partner, or the non-pregnant 
female partner of WOCBP uses a highly effective contraceptive method (defined 
below) 
• For female partners of male subjects:  surgical sterilization (e.g., bilateral tubal 
occlusion, hysterectomy, bilateral salpingectomy, bilateral oophorectomy), hormonal 
contraception associated with inhibition of ovulation (combined estrogen and 
progestogen containing, or progestogen-only), intrauterine contraception device or 
intrauterine hormone-releasing system (IUS). 
Male patients with partners that are pregnant must use condoms as barrier method to ensure that 
the fetus is not exposed to the Study Drug. 
†Note: Abstinence (i.e., refraining from heterosexual intercourse throughout the duration of 
study participation) is only acceptable as true abstinence, i.e., when this is in line with 
the preferred and usual lifestyle of the patient.  Periodic abstinence (e.g., calendar, 
ovulation, symptothermal, post-ovulation methods), declaration of abstinence for the 
duration of a trial and withdrawal are not acceptable methods of contraception. 
7. 
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7.2. 
Packaging and Labeling 
The Sponsor will provide the Investigator with packaged Study Drug (ISIS 757456 or placebo) 
labeled in accordance with specific country regulatory requirements.   
7.3. 
Study Drug Accountability 
The study staff is required to document the receipt, dispensing, and return/destruction of Study 
Drug (ISIS 757456 or placebo) supplies provided by the Sponsor according to Sponsor 
instruction and in accordance with institutional policy. 
8. 
TREATMENT OF PATIENTS 
8.1. 
Study Drug Administration 
Please refer to the Study Drug Manual provided by the Sponsor for more detailed instructions for 
Study Drug (ISIS 757456 or placebo) preparation and administration. 
Table 4: 
Study Drug Dosing Information 
Cohort 
Total Dose 
Cohort A 
80 mg ISIS 757456 or placebo SC 
Cohort B 
120 mg ISIS 757456 or placebo SC 
* The administration of Study Drug for doses of 120 mg may be delivered as a single injection or 2 non-contiguous 
injections.  
   
 
8.2. 
Other Protocol-Required Drugs 
Per eligibility criteria, patients must be on a stable regimen of 3 antihypertensives for at least 
1 month prior to screening evaluations and will be required to continue the stable regimen 
throughout the duration of the study (Refer to Inclusion Criteria Section 5.1).   
8.3. 
Other Protocol-Required Treatment Procedures 
There are no other protocol-required treatment procedures.   
8.4. 
Treatment Precautions 
There are no treatment precautions required. 
8.5. 
Safety Monitoring Rules 
Please refer also to the ‘Guidance for Investigator’ section of the Investigator’s Brochure. 
For the purposes of safety monitoring Baseline is defined as:  


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• The last non-missing measurement prior to the first dose 
In addition to the standard monitoring of clinical safety parameters, the following guidelines are 
provided for the monitoring of selected parameters chosen based on preclinical and clinical 
observations. 
Confirmation Guidance:  At any time during the study (Treatment or Post-Treatment Periods), 
the initial clinical laboratory results meeting the safety monitoring criteria presented below must 
be confirmed by performing measurements (ideally in the same laboratory that performed the 
initial measurement) on new specimens.  All new specimen collections should take place as soon 
as possible (ideally within 3 days of the initial collection).  For stopping rules, if the initial 
laboratory result is observed during the Treatment Period, the results from the retest must be 
available prior to administering the next dose of Study Drug (ISIS 757456 or placebo). 
Re-dosing Guidance:  Patients with initial laboratory test values that reach a stopping rule must 
not be re-dosed until the re-test results are available.  In general, patients who do not meet the 
stopping rules based upon retest may continue dosing.  However, the Investigator and the 
Sponsor Medical Monitor (or appropriately qualified designee) should confer as to whether 
additional close monitoring of the patient is appropriate.  If any of the stopping criteria described 
below (refer to Sections 8.6.1 to 8.6.6) are met, the patient will be permanently discontinued 
from further treatment with Study Drug (ISIS 757456 or placebo), evaluated fully as outlined 
below and in consultation with the Sponsor Medical Monitor or appropriately qualified designee, 
and will be followed up in accordance with Section 8.8 of the protocol.   
8.5.1. 
Safety Monitoring Rules for Liver Chemistry Tests 
The following rules are adapted from the draft guidance for industry, “Drug-Induced Liver 
Injury: Premarketing Clinical Evaluation,” issued by the U.S.  Department of Health and Human 
Services, Food and Drug Administration, July 2009.  For a definition of Baseline, please refer to 
guidance in Section 8.5 above. 
In the event of an ALT or AST measurement that is ˃ 3 × ULN at any time during the study 
(Treatment or Post-Treatment Period), the initial measurement(s) should be confirmed as 
described above.  Additional, confirmatory measurements should also be performed if ALT or 
AST levels increase to 5 × ULN. 
Frequency of Repeat Measurements:  Patients with confirmed ALT or AST levels ˃ 3 × ULN 
should have their liver chemistry tests (ALT, AST, ALP, INR and total bilirubin) retested at least 
once-weekly until ALT and AST levels become ≤ 1.2 × ULN or 1.2 × baseline value if the 
baseline value was ˃ ULN. 
Further Investigation into Liver Chemistry Elevations:  For patients with confirmed ALT or AST 
levels ˃ 3 × ULN, the following evaluations should be performed: 
• Obtain a more detailed history of symptoms and prior and concurrent diseases 
• Obtain further history for concomitant drug use (including nonprescription 
medications, herbal and dietary supplement preparations), alcohol use, recreational 
drug use, and special diets 
• Obtain a history for exposure to environmental chemical agents and travel 


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• Serology for viral hepatitis (hepatitis A virus [HAV] immunoglobulin M [IgM], 
HBsAg, HCV antibody, cytomegalovirus [CMV] IgM, and EBV antibody panel) 
• Serology for autoimmune hepatitis (e.g., antinuclear antibody [ANA], smooth muscle 
antibody IgG, Liver-Kidney Microsome-1 Antibody, IgG) 
Additional liver evaluations, including gastroenterology/hepatology consultations, hepatic 
computed tomography (CT), or magnetic resonance imaging (MRI) scans, may be performed at 
the discretion of the Investigator, in consultation with the Sponsor Medical Monitor.  Repetition 
of the above evaluations should be considered if a patient’s ALT and/or AST levels reach 
5 × ULN. 
For a definition of Baseline, please refer to guidance in Section 8.5. 
8.5.2. 
Safety Monitoring Rules for Platelet Count Results 
Please refer also to Table 5. 
Platelet count will be monitored every 2 weeks during the Treatment Period and at the following 
visits in the Post-Treatment Period:  Study Days 85, 92, 106, 120, 148, and 169.  The 
Investigator should review all platelet count results within 48 hours of receipt.  If a patient’s 
platelet count falls to 100,000/mm3 or less, then the patient’s platelet counts should be monitored 
as described in Table 5.  In case of platelet reduction to ˂ 50,000/mm3, the platelet monitoring 
rule defined in Stopping rules (Section 8.6.3) should be followed. 
Treatment should be held if there is no evaluable platelet count within 18 days prior to the 
scheduled dose.  Any uninterpretable platelet count result must be rechecked and determined not 
to have met a stopping rule before dosing can continue.  If Day 1 results are uninterpretable, 
Day 8 dosing may proceed, however, an interpretable result is required (and must be reviewed) 
before study drug dosing on Day 15.   
In the event of a platelet count ˂ 75,000/mm3, additional laboratory investigations may be 
conducted after discussion between the Investigator and the Sponsor Medical Monitor including: 
fibrinogen or D-dimer (local lab), complement (total C3, total C4, C5a), peripheral smear, 
citrated sample for platelets, CBC with reticulocytes and mean platelet volume, coagulation 
panel (prothrombin time/international normalized ratio [PT/INR], activated partial 
thromboplastin time [aPTT]), total globulins [IgG and IgM]), serology for hepatitis B virus, 
HCV, HIV (if not done at Screening), anti-platelet antibodies (specialty lab), and anti-drug 
antibodies (specialty lab).   
Any case of a platelet count ˂ 50,000/mm3 should be reported to the Sponsor in an expedited 
fashion (see Section 9.3.4).  The Study Medical Monitor will discuss the assessment and 
treatment of such subjects with the Investigator. 


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Table 5: 
Monitoring Schedule and Stopping Rules for Platelet Counts 
Platelet Count on 
Rx 
Drug Dose 
Monitoring 
> 100,000/mm3 
No action 
Monitor per Schedule of Procedures 
(Appendix A) 
≥ 75,000–
≤ 100,000/mm3 
No action 
Monitor every week until 3 successive values 
> 100,000/mm3 
≥ 50,000– 
< 75,000/mm3 
Pause dosing 
When platelet count returns to 
> 100,000/mm3 restart dosing only if 
approved by Sponsor Medical 
Monitor 
Monitor at least twice weekly until 3 successive 
values > 75,000/mm3 then weekly until 3 values 
> 100,000/mm3 
If not redosing then subsequent monitoring 
should be per the Schedule of Procedures 
(Appendix A) 
If redosing then continue to monitor weekly for 
the remainder of the Treatment Period 
Consider discontinuation of antiplatelet agents / 
non-steroidal anti-inflammatory drug (NSAIDS) 
/ anticoagulant medication while platelet count 
˂ 75,000/mm3 
≥ 25,000– 
< 50,000/mm3 
Permanently discontinue Study Drug 
Report to the Sponsor in an expedited fashion 
Monitor at least twice weekly until 3 successive 
values > 75,000/mm3 then weekly until 3 values 
> 100,000/mm3.  Subsequent monitoring should 
be per the Schedule of Procedures (Appendix A) 
Discontinue antiplatelet agents / NSAIDS / 
anticoagulant medication while platelet count 
˂ 75,000/mm3 if possible 
˂ 25,000/mm3 
Permanently discontinue Study Drug 
Report to the Sponsor in an expedited fashion 
Monitor daily until 3 successive values 
> 25,000/mm3 then monitor at least twice 
weekly until 3 successive values  
> 75,000/mm3 then weekly until 3 values 
> 100,000/mm3.  Subsequent monitoring should 
be per the Schedule of Procedures (Appendix A) 
Corticosteroids recommended* 
Consider need for hospitalization and referral to 
hematologist 
Discontinue antiplatelet agents / NSAIDS / 
anticoagulant medication while platelet count 
˂ 75,000/mm3 if possible 
* Recovery in platelet count may be accelerated by administration of high-dose glucocorticoids.  Treatment as recommended by 
the American Society of Hematology (ASH) (2019) guidelines for immune thrombocytopenia (Blood Advances, 
10 DECEMBER 2019, Volume 3, Number 23) includes initial therapy with either dexamethasone 40 mg per day for 4 days, or 
prednisone 0.5 to 2.0 mg/kg per day.  Prednisolone or prednisone may be administered for up to 2 to 4 weeks with taper; 
alternatively, intravenous immunoglobulin (IVIG) may be administered at 0.4 g/kg/d for 5 days, or infusions of 1 g/kg/d for 
1-2 days (Provan et al. 2010; Provan et al. 2019).   


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8.5.3. 
Safety Monitoring for Minor Bleeding Events 
Minor bleeding events are those that do not fulfill the criteria for major bleeding (MB) or 
clinically relevant, non-major bleeding (CRNMB) events (which are defined in Section 8.6.4), 
for example excess bruising, petechiae, gingival bleeding on brushing teeth.  Additional testing 
of platelet counts and coagulation parameters (PT/INR/aPTT) should be considered after 
consultation with the Sponsor Medical Monitor. 
8.5.4. 
Safety Monitoring for Potassium 
In the event of a potassium measurement ˃ 5.3 mmol/L at any time during the study (Treatment 
or Post-Treatment Period), the initial measurement(s) should be confirmed.  If the initial 
potassium measurement is confirmed to be ˃ 5.3 mmol/L, weekly monitoring will be initiated 
until levels return to ˂ 5.3 mmol/L.   
In the event of a potassium measurement ≥ 5.5 mmol/L, but less than 6.0 mmol/L, study drug is 
to be held.  In the event of a potassium measurement ≥ 6.0 mmol/L refer to Section 8.6.5 
Stopping Rule for Potassium.  In the event of a potassium measurement ≥ 5.5 mmol/L, the 
Investigator or designee should assess for symptoms consistent with hyperkalemia (e.g., muscle 
weakness or paralysis).  An ECG may be considered per Investigator judgement to assess 
electrocardiographic signs of hyperkalemia (delayed conduction, heart block, arrhythmias) 
which, if conducted, should be discussed with the Sponsor Medical Monitor or designee to 
determine continued dosing with Study Drug.  The investigator or designee should consider 
treating the patients with therapies that reduce the serum potassium, such as a low-potassium 
diet, diuretics, or a reduction or cessation of medications that can increase the serum potassium 
(ACEi/ARB).  Serum potassium should be measured frequently until documented to be 
˂ 5.5 mmol/L.  Dosing with Study Drug may continue after a confirmed potassium measurement 
˂ 5.5 mmol/L with Investigator and Sponsor Medical Monitor or designee consultation.   
8.5.5. 
Safety Monitoring for Blood Pressure 
If the patient is symptomatic (i.e., dizziness, light-headedness, clammy skin, fatigue, blurry 
vision), at any point in the study, the patient should contact the Study Center immediately or 
should a patient experience a BP measurement of ≤ 90 mmHg (systolic) 2 additional 
measurements must be done within a 30-minute period.  If the confirmed average of the 
2 additional SBP measurements is ≤ 90 mmHg and/or the patient remains symptomatic, dosing 
of a patient with Study Drug will be paused.  Oral and/or IV hydration should be instituted.  The 
suitability of the patient for continued dosing will be determined by the Investigator in 
consultation with the Sponsor Medical Monitor after the SBP has normalized and/or patient is no 
longer symptomatic and will be based on the reversibility of initiating factors (e.g., decreased 
oral intake, cold/flu, other illness).  If the patient remains symptomatic or if the SBP does not 
normalize Study Drug should be permanently discontinued (refer to Section 8.6.6).   


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8.6. 
Stopping Rules 
For the purposes of the stopping rules, Baseline is defined as:  
• Temporary stopping rules for renal function tests – Baseline is defined as the average 
of the pre-dose test closest to Day 1 and Day 1  
• Stopping rules for platelets – Baseline is defined as the last non-missing measurement 
prior to the first dose 
8.6.1. 
Stopping Rules for Liver Chemistry Elevations 
In the event of laboratory results meeting the following criteria, and the event is without an 
alternative explanation as discussed with the Sponsor Medical Monitor, dosing of a patient 
with Study Drug (ISIS 757456 or placebo) will be stopped permanently; values that are not 
confirmed due to failure to retest or missing lab values will be presumed confirmed: 
1. ALT or AST ˃ 8 × ULN, which is confirmed 
2. ALT or AST ˃ 5 × ULN, which is confirmed and persists for ≥ 2 weeks 
3. ALT or AST ˃ 3 × ULN, which is confirmed and total bilirubin ˃ 2 × ULN or INR ˃ 1.5 
4. ALT or AST ˃ 3 × ULN, which is confirmed, and the new appearance (i.e., onset 
coincides with the changes in hepatic enzymes) of fatigue, nausea, vomiting, right upper 
quadrant pain or tenderness, fever, rash, and/or concomitant eosinophilia (˃ ULN) 
8.6.2. 
Stopping Rules for Renal Function Test Results 
1. Confirmed 30% decline in eGFR from Baseline eGFR values [eGFR should be calculated 
using the CKD-EPI creatinine equation (Levey et al. 2009)] 
2. Confirmed proteinuria:  Absolute UPCR value ≥ 1000 mg/g 
The possible dosing re-initiation or follow-up schedule for any events meeting either of these 
criteria will be determined by the Investigator in consultation with the Sponsor Medical 
Monitor or designee. 
At the discretion of the Investigator, a decision to hold or permanently stop study drug may be 
made based on lesser changes in these parameters observed in isolation or in association with 
other renal-related abnormalities. 
8.6.3. 
Stopping Rule for Platelet Count Results 
See Table 5 for an overview of monitoring and stopping rules for platelet counts.   
• In the event of a confirmed platelet count less than 50,000/mm3, dosing of a patient 
with Study Drug (ISIS 757456 or placebo) will be stopped permanently 
− The platelet count should be tested at least twice weekly until 3 successive values 
above 75,000/mm3 
− Then tested weekly until 3 values above 100,000/mm3 
− Subsequent monitoring should follow the Schedule of Procedures (Appendix A) 


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• In the event of any platelet count less than 25,000/mm3, dosing of the patient with 
Study Drug will be stopped permanently 
− Platelet count should be monitored daily until 3 successive values above 
25,000/mm3 
− Then monitor at least twice weekly until 3 successive values above 75,000/mm3 
− Then monitor weekly until 3 successive values above 100,000/mm3 
− Then monitor per the Schedule of Procedures (Appendix A) 
− Administration of corticosteroids is recommended for patients whose platelet 
count is less than 25,000/mm3, with referral to a hematologist. 
In the event of a platelet count ˂ 75,000/mm3 and ≥ 50,000/mm3, and in the absence of MB or 
CRNMB (defined below), dosing with Study Drug should be suspended temporarily until the 
platelet count has recovered to ˃ 100,000/mm3.  The suitability of the patient for continued 
dosing will be determined by the Investigator in consultation with the Study Medical Monitor 
and will be based on factors such as the original rate of decline in the patient’s platelet count, 
whether any bleeding events were experienced, and the speed of recovery of platelet count after 
interruption of dosing.  If dosing is reinitiated, platelet count must be measured twice weekly 
(consider more frequent monitoring if risk factors for bleeding are present) until 3 successive 
values above 75,000/mm3 then weekly until the end of study.  Treatment should be held if there 
is no evaluable platelet count within 18 days prior to the scheduled dose.  Any uninterpretable 
platelet count result must be rechecked and determined not to have met a stopping rule before 
dosing can continue.   
Labs to Be Performed in the Event of a Platelet Count Less than 50,000/mm3 
The following is a list of recommended lab analyses for patients who have any occurrence of 
platelet count less than 50,000/mm3.  Archived samples prior to Day 1 of investigational 
treatment may also be analyzed in order to determine the patient’s baseline conditions. 


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Table 6: 
Recommended Laboratory Analyses 
To Be Performed at Local Lab 
Peripheral smear (should be performed locally, fixed, and sent to central lab for review) 
Fibrinogen split products or D-dimer on fresh blood 
To Be Performed at Central Lab 
Citrated sample for platelets 
Coagulation panel (PT/INR, aPTT) 
CBC with reticulocytes 
Total globulins:  IgG and IgM 
Complement:  total C3, total C4, C5a 
Serology for: 
HBV, HCV, HIV (if not done recently for screening) 
To Be Performed at Specialty Lab(s) 
Antiplatelet antibodies 
Anti-ASO antibody 
Note:  These labs may be performed in the event of a platelet count less than 75,000/mm3 after discussion with the 
Study Medical Monitor.  Additional lab tests may be considered in order to ascertain potential causative conditions 
for significant platelet count decreases / thrombocytopenia.   
 
8.6.4. 
Bleeding Events 
8.6.4.1. 
Definition of Major Bleeding Events (Schulman and Kearon 2005) 
1. Fatal bleeding, and/or 
2. Symptomatic bleeding in a critical area or organ, such as intracranial, intraspinal, 
intraocular, retroperitoneal, intraarterial or pericardial, or intramuscular with 
compartment syndrome 
3. Clinically overt bleeding leading to transfusion of ≥ 2 units of packed red blood cells or 
whole blood or a fall in hemoglobin of 2.0 mg/dL (1.24 mmol/L) or more within 24 hours 
8.6.4.2. 
Definition of Clinically Relevant Non-Major Bleeding Events 
Clinically relevant non-major bleeding is defined as overt bleeding not meeting the criteria for 
MB but that resulted, for example, in medical examination, intervention, or had clinical 
consequences for a patient. 
8.6.4.3. 
Definition of Minor Bleeding Events 
Minor bleeding events are those that do not fulfill the criteria for MB or CRNMB (defined 
above), for example excess bruising, petechiae, gingival bleeding on brushing teeth. 


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8.6.5. 
Stopping Rule for Potassium 
In the event of confirmed potassium of ≥ 6.0 mmol/L dosing of a patient with Study Drug will be 
stopped permanently.  Investigator or designee should assess for symptoms (e.g., muscle 
weakness or paralysis) and electrocardiographic signs of hyperkalemia (delayed conduction, 
heart block, arrhythmias).  The Investigator or designee should consider treating the patients with 
therapies that reduce the serum potassium, such as a low-potassium diet, diuretics, or a reduction 
or cessation of medicines that can increase the serum potassium (ACEi/ARB).  Additional 
treatments for hyperkalemia (e.g., intravenous [IV] calcium, insulin and glucose, gastrointestinal 
cation exchangers) can be considered per investigator judgment.  The Investigator may consider 
obtaining a nephrology consultation with consideration for dialysis to treat hyperkalemia.  
Additional consultation of the Investigator with the Sponsor Medical Monitor may be considered 
to individualize treatment plan. 
8.6.6. 
Stopping Rule for Blood Pressure 
1. If the patient remains symptomatic (i.e., dizziness, light-headedness, clammy skin, 
fatigue, blurry vision) and/or in the event of the average of 2 additional BP measurements 
≤ 90 mmHg (systolic) after instituting oral or IV hydration, Study Drug should be 
permanently discontinued, and the patient should be referred to urgent care or the 
emergency room. 
Any other antihypertensives or other medications that could affect BP should be stopped, 
if feasible.  The use of IV angiotensin II or vasopressors should be considered in patients 
that remain symptomatic and are hemodynamically unstable.  The patient will continue to 
be followed per protocol in the study.  Follow-up for patients that screen fail due to BP 
inclusion criteria (see Section 5.1)11T will not be necessary. 
2. Should a patient experience a BP measurement ˃ 180 mmHg (systolic) or ˃ 115 mmHg 
(diastolic) 2 additional measurements must be done within a 30-minute period and the 
average must be calculated.  If the average of the 2 additional measurements is 
˃ 180 mmHg (systolic) or ˃ 115 mmHg (diastolic), they will then be withdrawn from 
further blinded, randomized treatment.  Study Drug is to be permanently discontinued 
unless the hypertensive event is transient and dependent on reversible initiating factors 
(e.g., stressful event).  The suitability of the patient for continued dosing will be 
determined by the Investigator in consultation with the Sponsor Medical Monitor after 
the patient's BP has normalized.  Additional antihypertensive medication(s) may be 
added per Investigator and Sponsor Medical Monitor judgement if the patient has 
discontinued Study Drug.  If the patient has not discontinued Study Drug, changes to the 
stable regimen of antihypertensives may result in withdrawal of the patient from the 
Treatment Period.  The patient will continue to be followed per protocol in the study.  
Follow-up for patients that screen fail due to BP inclusion criteria (see Section 5.1) will 
not be necessary. 


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Table 7: 
Blood Pressure Stopping and Monitoring Rules 
Confirmed Blood 
Pressure 
Dosing 
Additional Actions 
SBP ≤ 90 mmHg* and/or 
patient is symptomatic 
(e.g., decreased oral 
intake, cold/flu, other 
illness) 
• Pause 
• Institute oral and/or IV hydration.  If hypotension 
or symptoms of hypotension persist proceed to 
stopping rules below 
• The suitability of the patient for continued dosing 
will be determined by the Investigator in 
consultation with the Sponsor Medical Monitor 
after the SBP has normalized and/or patient is no 
longer symptomatic and will be based on the 
reversibility of initiating factors (e.g., decreased 
oral intake, cold/flu, other illness)  
SBP ≤ 90 mmHg* and/or 
the patient remains 
symptomatic after 
instituting oral and/or IV 
hydration and there is no 
other reason contributing 
to hypotensive event 
• Permanently 
Discontinue 
• The patient should be referred to urgent care or the 
emergency room 
• Any other antihypertensives or other medications 
that could affect BP should be stopped, if feasible 
• The use of IV angiotensin II or vasopressors 
should be considered in patients that remain 
symptomatic and are hemodynamically unstable 
• The patient will continue to be followed per 
protocol in the study.  Follow-up for patients that 
screen fail due to BP inclusion criteria (see 
Section 5.1) will not be necessary 
SBP ˃ 180 mmHg* or  
DBP ˃ 115 mmHg* 
• Pause if BP 
elevation is 
thought to be 
transient, 
reversible and 
related to external 
initiating factors 
(e.g., stressful 
event) 
• Permanently 
discontinue if no 
other reason 
contributing to 
hypertensive 
event  
• The suitability of the patient for continued dosing 
will be determined by the Investigator in 
consultation with the Sponsor Medical Monitor 
after the patient's BP has normalized 
• Additional antihypertensive medication(s) may be 
added per Investigator and Sponsor Medical 
Monitor judgement if the patient has discontinued 
Study Drug.  If the patient has not discontinued 
Study Drug, changes to the stable regimen of 
antihypertensives may result in withdrawal of the 
patient from the Treatment Period 
• The patient will continue to be followed per 
protocol in the study.  Follow-up for patients that 
screen fail due to BP inclusion criteria (see 
Section 5.1) will not be necessary 
* BP measurements must be confirmed by the average of 2 additional measurements done within a 30-minute period  
 


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8.7. 
Adjustment of Dose and/or Treatment Schedule 
An adjustment to the dose frequency from weekly to every 2 weeks or a weekly dose reduction 
may be permitted for safety or efficacy reasons in consultation with the Sponsor Medical 
Monitor or designee.   
8.8. 
Discontinuation of Study Drug 
A patient must permanently discontinue study treatment for any of the following: 
• The patient becomes pregnant.  Report the pregnancy according to instructions in 
Section 9.5.4 
• The patient withdraws consent 
• The patient experiences an AE that necessitates permanent discontinuation of Study 
Drug  
• The patient develops laboratory test abnormalities that meet any of the stopping rules 
listed in Sections 8.6.1 to 8.6.3 
• The patient experiences an AE that necessitates unblinding of the Investigator  
The reason for discontinuation of Study Drug must be recorded in the electronic Case Report 
Form (eCRF) and source documentation. 
Patients who early terminate from treatment will complete the Post-Treatment study procedures 
per the Appendix A.   
For patients withdrawn for reasons other than withdrawal of consent every effort should be made 
to complete the early termination of study procedures and observations at the time of withdrawal 
(see Appendix A) and ideally within 2 weeks from the last dose of Study Drug.   
8.9. 
Withdrawal of Patients from the Study Procedures 
Patients must be withdrawn from study procedures for any of the following: 
• Withdrawal of consent 
• The patient is unwilling or unable to comply with the protocol 
Other reasons for withdrawal of patients from study procedures might include: 
• At the discretion of the Investigator for medical reasons 
• At the discretion of the Investigator or Sponsor for noncompliance 
• Significant protocol deviation 
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 eCRF. 
Any patient who withdraws consent to participate in the study will be removed from further 
treatment and study observation immediately upon the date of request.  These patients should be 


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encouraged to complete the early termination study procedures and observations at the time of 
withdrawal (Appendix A). 
For patients withdrawn for reasons other than withdrawal of consent every effort should be made 
to complete the early termination of study procedures and observations at the time of withdrawal 
(see Appendix A) and ideally within 2 weeks from the last dose of Study Drug.   
8.10. 
Concomitant Therapy and Procedures 
The use of concomitant therapies or procedures defined below must be recorded on the patient’s 
eCRF.  Adverse events related to administration of these therapies or procedures must also be 
documented on the appropriate eCRF. 
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 the Post-Treatment Evaluation Period. 
Allowed Concomitant Therapy 
Any other medications deemed necessary by the Investigator are allowed except those listed in 
the disallowed concomitant therapy. 
COVID-19 vaccines are allowed during study participation.  However, because the vaccines are 
associated with local injection site reactions and constitutional symptoms, the vaccination should 
NOT be dosed on the same day of Study Drug injection and avoid the same site of any Study 
Drug injection.  If a patient has a reaction from a COVID-19 vaccination, ensure patient recovers 
prior to administering the next Study Drug injection (even if it incurs a delay). 
Disallowed Concomitant Therapy 
The following are disallowed concomitant therapies: 
• Other medications for the treatment of HTN (e.g., minoxidil, diazoxide, renin 
inhibitors) 
• Medications that may cause hyperkalemia unless on a stable dose at least 1 month 
prior to the Screening visit  
• Use of oral anticoagulants unless stable for 4 weeks prior to the first dose of Study 
Drug and regular monitoring must be performed, per clinical practice during the study 
unless the patient is receiving vitamin K agonists.  If the patient is receiving vitamin 
K antagonists (e.g., warfarin) INR should be in therapeutic range, as established by 
the Investigator, for 4 weeks prior to the first dose 
• Chronic administration (defined as > 3 days per week for the duration of the trial) of 
NSAIDs or COX-2 inhibitors (except aspirin for cardiovascular disease provided the 
total daily dose does not exceed 325 mg) 
• Use of phosphodiesterase 5 inhibitors (e.g., sildenafil, tadalafil, vardenafil, avanafil) 
within 72 hours prior to any scheduled visit 


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Changes to the stable regimen of antihypertensives allowed at Screening may result in 
withdrawal of the patient from the Treatment Period. 
8.10.2. 
Concomitant Procedures 
A concomitant procedure is any therapeutic intervention (e.g., surgery/biopsy, physical therapy) 
or diagnostic assessment (e.g., blood gas measurement, bacterial cultures) performed between 
Screening and the Post-Treatment Evaluation Period. 
8.11. 
Treatment Compliance 
Compliance with treatment dosing is to be monitored and recorded in the eCRF by Study Center 
staff.   
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 or designee in 
accordance with the applicable Ionis and/or designee SOPs throughout the conduct of the clinical 
trial. 
9.2. 
Regulatory Reporting Requirements 
The Sponsor or designee 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 GCP.  
Country-specific regulatory requirements will be followed in accordance with local country 
regulations and guidelines. 
Institutional Review Boards will be notified of any SAE according to applicable regulations. 
In addition to the Investigator’s assessment of relatedness, the Sponsor or designee will evaluate 
the available information and perform an independent assessment of all reported SAEs and 
determine if there is a reasonable possibility that the Study Drug (ISIS 757456 or placebo) is 
causally related to a reported SAE and, therefore, meets the definition of an SUSAR.  While the 
Sponsor’s independent causality assessment may differ from the Investigator’s assessment, the 
country-specific regulatory requirements must be followed for expedited reporting of SUSAR to 
local regulatory authorities. 
Appropriate personnel at the Sponsor or designee will unblind SUSARs for the purpose of 
regulatory reporting.  The Sponsor or designee will submit SUSARs to Regulatory Agencies in 
blinded or unblinded fashion according to local law.  The Sponsor or designee will submit 
SUSARs to Investigators in a blinded fashion. 
For the purpose of regulatory reporting of SUSARs, there are no “expected” AEs in this study 
population.  Investigator’s Brochure (provided separately) for expected AEs. 


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9.3. 
Definitions 
9.3.1. 
Adverse Event 
An AE can be any unfavorable and unintended sign (including an abnormal laboratory finding, 
for example), symptom, or disease temporally associated with the use of medicinal 
(investigational) product, whether or not the AE is considered related to the medicinal 
(investigational) product. 
An AE can therefore be any of the following:   
• Any unfavorable and unintended sign (including an abnormal laboratory finding), 
symptom, or disease temporally associated with the use of a medicinal product, 
whether or not considered related to the medicinal product 
• Any new disease or exacerbation of an existing disease (a worsening in the character, 
frequency, or severity of a known condition) 
• Recurrence of an intermittent medical condition (e.g., headache) not present at 
Baseline 
• Any deterioration in a laboratory value or other clinical test (e.g., ECG, X-ray) that is 
associated with symptoms or leads to a change in study treatment or concomitant 
treatment or discontinuation from Study Drug 
• Adverse events that are related to a protocol-mandated intervention, including those 
that occur prior to assignment of study treatment (e.g., screening invasive procedures 
such as biopsies). 
9.3.2. 
Adverse Drug Reaction and Suspected Unexpected Adverse Drug Reaction 
Adverse Drug Reaction (ADR) 
In the pre-approval clinical experience with a new medicinal product or its new usages, 
particularly as the therapeutic dose(s) may not have been established, ADR is defined as follows: 
All noxious and unintended responses to a medicinal product related to any dose should 
be considered adverse drug reactions. 
The phrase "responses to a medicinal product" means that a causal relationship between the 
medicinal product and the AE has been determined by the Sponsor as at least a reasonable 
possibility, i.e., the relationship cannot be ruled out. 
Suspected Unexpected Adverse Drug Reaction  
A suspected unexpected ADR is any ADR, the nature or severity of which is not consistent with 
the applicable product information, e.g., Investigator's Brochure for an unapproved medicinal 
(investigational) product. 
A suspected adverse reaction implies a lesser degree of certainty about causality than an adverse 
reaction. 


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9.3.3. 
Serious Adverse Event 
A SAE is any AE that in the view of either the Investigator or Sponsor, meets any of the 
following criteria: 
• 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 patient 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 a congenital anomaly or birth defect in the offspring of the patient (whether 
the patient 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 patient 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 
The terms "severe" and "serious" are not synonymous.  Severity refers to the intensity of an AE 
(e.g., rated as mild, moderate, or severe, or according to National Cancer Institute Common 
Terminology Criteria for Adverse Events [NCI CTCAE]; the event itself may be of relatively 
minor medical significance (such as severe headache without any further findings). 
Severity and seriousness need to be independently assessed for each AE recorded on the eCRF. 
9.3.4. 
Adverse Event of Special Interest  
Adverse events of special interest (AESI), including both serious or non-serious events, is one of 
scientific and medical concern specific to the Sponsor’s product or program, for which ongoing 
monitoring and rapid communication by the Investigator to the Sponsor could be appropriate. 
For the purpose of this study, severe reductions in platelet count ˂ 50,000/mm3 accompanied by 
an MB event or CRNMB event, or platelet count of ˂ 25,000/mm3 independent of an MB or 
CRNMB event are considered as AEs of special interest.  AESI must be reported to the Sponsor 
within 24 hours to ensure timely submission as the 15-day expedited report by the Sponsor to 
regulatory agencies. 


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Adverse events of special interest are required to be reported by the Investigator to the Sponsor 
immediately, no more than 24 hours of the Investigator’s first knowledge of the event for 
expedited reporting to Regulatory Authorities. 
9.4. 
Monitoring and Recording Adverse Events 
Any pre-existing conditions or signs and/or symptoms present in a patient 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.  Before a 
diagnosis is confirmed, all symptoms should be reported as separate AEs. 
9.4.1. 
Serious Adverse Events 
In the interest of patient safety, and in order to fulfill regulatory requirements, all SAEs 
(regardless of their relationship to Study Drug) should be reported to the Sponsor or designee 
within 24 hours of the Study Center’s first knowledge of the event.  The collection of SAEs will 
begin after the patient signs the informed consent form and stop at the end of the patient’s 
Follow-up Period which is defined as Study Day 169.  When the Investigator is reporting by 
telephone, it is important to speak to someone in person versus leaving a message.   
Once the patient is randomized to study drug, SAEs should be reported using electronic SAE 
electronic data capture system (EDC) whenever possible.  In situations where electronic SAE 
submission is unavailable or of an SAE occurs prior to the patient’s randomization, a paper 
Serious Adverse Event Form should be completed, and a copy should be faxed or emailed to the 
Sponsor or designee.  For SAEs reported after randomization, information submitted on paper 
form should be entered into EDC as soon as the system becomes available.  The SAE reporting 
instruction, including the fax number and email address can be found in the Investigator site file 
for the study. 
Detailed follow-up information should be actively sought and promptly submitted electronically 
or on a Serious Adverse Event Form if reporting by paper before randomization. 
All SAEs will be followed until resolution.  Serious Adverse Events that remain ongoing past the 
patient’s last protocol-specified follow-up visit will be evaluated by the Investigator and 
Sponsor.  If the Investigator and Sponsor agree the patient’s condition is unlikely to resolve, the 
Investigator and Sponsor will determine the follow-up requirement. 
9.4.2. 
Non-Serious Adverse Events 
The recording of non-serious AEs will begin after the patient signs the informed consent form 
and will stop at the end of the patient’s Follow-up Period, which is defined as Study Day 169.  
The Investigator will monitor each patient 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. 


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9.4.3.1. 
Relationship to the Study Drug 
The event’s relationship to the Study Drug (ISIS 757456 or placebo) 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 patient’s medical condition, 
concomitant therapy, or other causes, and there is a plausible temporal relationship 
between the event and Study Drug (ISIS 757456 or placebo) 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 (ISIS 757456 or placebo) 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 patient’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 severity of AEs and SAEs relating to laboratory tests and AEs at the injection site will be 
graded based on criteria from the Common Terminology Criteria for Adverse Events (CTCAE) 
Version 5.0, November 2017 (refer to Appendix D).  Any AE not listed in Appendix D will be 
graded as follows: 
• Mild:  The event is easily tolerated by the patient and does not affect the patient’s 
usual daily activities 
• Moderate:  The event causes the patient more discomfort and interrupts the patient’s 
usual daily activities 
• Severe:  The event is incapacitating and causes considerable interference with the 
patient’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). 
9.4.3.3. 
Action Taken with Study Drug 
Action taken with Study Drug (ISIS 757456 or placebo) due to the event is characterized by one 
of the following. 
• None:  No changes were made to Study Drug (ISIS 757456 or placebo) 
administration and dose 
• Not Applicable:  SAE/AE was reported during Screening Period prior to Study Drug 
administration 
• Permanently Discontinued:  Study Drug was discontinued and not restarted 


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• Temporarily Interrupted, Restarted – Same Dose:  Dosing and/or dosing 
frequency was temporarily interrupted/changed or delayed due to the AE and 
restarted at the same dose 
• Reduced Dose:  Dosing was reduced, temporarily interrupted, or delayed due to the 
AE and restarted at the next lower dose or reduced dosing frequency  
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 eCRF, 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:  Patient terminates from the trial and the AE continues 
• Recovered:  Patient 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 an SAE, then the event’s outcome is characterized by one of the following: 
• Ongoing:  SAE continuing 
• Persists (as non-serious AE):  Patient has not fully recovered but the event no longer 
meets serious criteria and should be captured as an AE on the non-serious AE eCRF 
(the SAE resolution date should be entered as the date of onset of that AE) 
• Recovered:  Patient recovered completely from the SAE (the date of recovery should 
be entered as the SAE resolution date) 
• Recovered with Sequelae:  The signs/symptoms of the reported SAE have improved 
but not completely resolved, and a new baseline for the patient is established since 
full recovery is not expected 
• Fatal:  Patient died (the date of death should be entered as the SAE resolution date) 
• Unknown:  The outcome of the reported SAE is not available, e.g., patient is lost to 
follow-up 
9.4.3.6. 
Follow-up of Adverse Event 
Investigator Follow-Up 
During the Study Period, the Investigator should follow each AE until the event has resolved to 
baseline grade or better, the event is assessed as stable, the patient is lost to follow-up, or the 


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patient withdraws consent.  Every effort should be made to follow all SAEs considered to be 
related to Study Drug or related to study procedures until a final outcome can be reported. 
Resolution of AE (with dates) should be documented on the Adverse Event eCRF and in the 
patient’s medical record to facilitate source data verification.   
Investigator should follow-up or support the Sponsor’s effort to follow up with all pregnancies 
reported during the study from either the study patient or the female partner of male study patient 
until pregnancy outcome is available.   
Sponsor Follow-Up 
For SAEs, AESI , and pregnancy cases in patients who has completed or terminated study, the 
Sponsor or a designee should follow-up by telephone, fax, email, and/or a monitoring visit to 
obtain additional case details and outcome information (e.g., from hospital discharge summaries, 
consultant reports, autopsy reports) in order to perform an independent medical assessment of 
these case. 
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 
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 Sponsor 
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 and should include the Investigator’s 
assessment of clinical significance of out of range/abnormal laboratory values. 
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 patient 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 patient’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 patient’s consent to participate in the study and the 
timing of the procedure or treatment 


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• The prescheduled or elective procedure or routinely scheduled treatment is the sole 
reason for the intervention or hospital admission 
9.5.3. 
Medication Errors 
Study Drug (ISIS 757456 or placebo) errors (for example, overdose, underdose, and 
administration error) should be reported as a medication error on the Protocol Deviation CRF.  A 
brief description should be provided when reporting the event, including whether the patient was 
symptomatic or asymptomatic, and whether the event was accidental or intentional. 
Dosing details should also be captured on the Dosing CRF.  If the patient takes a dose of Study 
Drug that exceeds protocol specifications and the patient is symptomatic, then an AE of 
medication error should be reported per Section 9.4, and all symptoms should be included in the 
event description.   
An overdose is the accidental or intentional use of a drug in an amount higher than the dose 
being studied.  An overdose or incorrect administration of study treatment should be reported as 
an AE of medication error.  All symptoms associated with an overdose or incorrect 
administration of Study Drug should be recorded on the Adverse Event eCRF.  The event should 
be reported to the Sponsor immediately (i.e., no more than 24 hours after learning of the event). 
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 or designee within 
24 hours. 
9.5.4. 
Contraception and Pregnancy 
Male patients must continue to use appropriate contraception with their partners, or refrain from 
sexual activity, as described in Section 6.4.1. 
If a male patient makes or believes that he has made someone pregnant during the study, then the 
Study Center staff must be informed immediately.  An Initial Pregnancy Form should be 
submitted to the Sponsor or designee within 24 hours of first learning of the occurrence of 
pregnancy.  Follow-up information including delivery or termination should be reported by 
designating as ‘Follow-up’ on the Pregnancy Forms and reported within 24 hours. 
Payment for all aspects of obstetrical care, child or related care will be the patient’s 
responsibility.   
Male patients:  The progress of the pregnancy of a male patient’s partner should be followed 
until the outcome of the pregnancy is known (i.e., delivery, elective termination, or spontaneous 
abortion).  If the pregnancy results in the birth of a child, the Study Center and Sponsor may 
follow-up with the mother and may request access to the mother and infant’s medical records to 
obtain additional information relevant to the pregnancy progress and outcome.  A longer 
follow-up may be required if a newborn child experiences a medical condition.  Follow-up will 
be performed to the extent permitted by the applicable regulations and privacy considerations, 
e.g., partner ICF may be required. 


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10. 
STATISTICAL CONSIDERATIONS 
10.1. 
Stratification, Subsets, and Covariates 
Patients will be stratified on Screening eGFR result of ˂ 60 or ≥ 60 mL/min/1.73 m2.  In general, 
the covariates will include stratification factors and baseline measurement.  
10.2. 
Sample Size Considerations 
Approximately 150 patients will be stratified based on Screening eGFR result of ˂ 60 or 
≥ 60 mL/min/1.73 m2 and randomized to 1 of 2 dose cohorts in a 1:1 ratio (Cohort A or 
Cohort B).  Within each dose cohort, patients will be further randomized to receive ISIS 757456 
or placebo in a 2:1 ratio.  The planned sample size will provide approximate 80% of power to 
demonstrate the mean SBP reduction difference of 8 mmHg between each ISIS treated group and 
pooled placebo based on the assumption of pooled standard deviation of 13 mmHg with the 
two-sided test and a significant level of 0.05 while taking into account an estimated 10% 
drop-out rate.  The overall type I error rate is at 0.1 level, two-sided.   
10.3. 
Populations 
UFull Analysis Set (FAS)U:  All randomized patients who have received at least 1 injection of 
Study Drug (ISIS 757456 or placebo) and who have at least 1 post-Baseline efficacy 
measurements. 
Per Protocol Set (PPS):  All FAS patients who received at least 10 of the 12 doses of Study Drug, 
did not alter Screening antihypertensive medications during the Treatment Period and prior to 
Study Day 85, and have no significant protocol deviations that would be expected to affect 
efficacy assessments. 
USafety SetU:  All patients who are randomized and receive at least 1 dose of Study Drug. 
UPK SetU:  All patients who are randomized and receive at least 1 dose of Study Drug and have at 
least 1 evaluable PK sample. 
10.4. 
Definition of Baseline 
The baseline for plasma AGT will be define as the average of all values prior to the first dose of 
Study Drug.   
The baseline for all other assessments will be defined as the last non-missing measurement prior 
to the first dose.   
10.5. 
Interim Analysis and Multiplicity 
An interim analysis may be conducted after approximately 50% of the patients have been 
enrolled. 
For the final analysis, the multiplicity of primary and key secondary endpoints comparisons will 
be controlled by using sequential testing strategy in the following testing sequence: 
• Comparison of change from baseline to Study Day 85 in SBP between each ISIS 757456 
treated group and pooled placebo in the PPS 


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• Comparison of percent change from baseline to Study Day 85 in AGT between each 
ISIS 757456 treated group and pooled placebo in the PPS 
• Comparison of change from baseline to Study Day 85 in SBP from 24 hr mean ABPM 
between each ISIS 757456 treated group and pooled placebo in the PPS 
• Comparison of portion of patients reaching the goals of seated automated SBP 
≤ 140 mmHg and DBP ≤ 90 mmHg at Day 85 between each ISIS 757456 treated group 
and pooled placebo in the PPS 
The statistical comparison will be conducted with 2-sided alpha level of 0.05 for each 
ISIS 757456 treated group verse pooled placebo.  If an efficacy endpoint comparison in the 
sequence of an ISIS treated group is not statistically significant, then the subsequent endpoints 
analyses will be considered as exploratory for that ISIS 757456 treated group.  Details will be 
provided in the Statistical Analysis Plan (SAP). 
10.6. 
Planned Methods of Analysis 
All case report form (CRF) data, lab data transfers, and any outcomes derived from the data may 
be provided in the patient data listings.  Patient data listings will be presented for all patients 
randomized in the study.  Descriptive summary statistics including n, mean, median, standard 
deviation, standard error, interquartile range (25th percentile, 75th percentile), and range 
(minimum, maximum) for continuous variables, and counts and percentages for categorical 
variables will be used to summarize most data by treatment group.  Where appropriate, p-values 
will be reported.  The placebo patients from both cohorts will be pooled.  All primary and 
secondary endpoints will be assessed on the Full Analysis Set and Per-Protocol Set, with the 
latter being the basis for the primary efficacy analysis.  All safety assessments will be performed 
on the Safety Set.   
10.6.1. 
Demographic and Baseline Characteristics 
Demographic and Baseline characteristics will be summarized using descriptive statistics by 
treatment group.  Patient randomization will be summarized by cohort and treatment group.  The 
patient disposition will be summarized.  All patients enrolled will be included in a summary of 
patient disposition. 
10.6.2. 
Safety Analysis 
The safety analysis will be conducted on the Safety Population. 
Treatment duration and amount of Study Drug (ISIS 757456 or placebo) received will be 
summarized by treatment group.  Patient incidence rates of all AEs will be tabulated by 
MedDRA™ system organ class, and by Preferred Term.  Narratives of treatment emergent deaths, 
SAEs and AEs of special interest, including early withdrawals due to AEs, will also be provided. 
All treatment-emergent AEs and SAEs as well as all treatment-emergent AEs and SAEs 
potentially related to Study Drug (ISIS 757456 or placebo) will be summarized. 
Laboratory tests to ensure patient safety including chemistry panel, complete blood count with 
differential, coagulation panel, complement etc., will be summarized by study visits for each 
treatment group.  These safety variables will also be presented as change and percent change 


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from Baseline over time after Study Drug (ISIS 757456 or placebo) administration, as 
appropriate.   
Vital sign and ECG measures will be tabulated by treatment group.  In addition, the number of 
patients who experience abnormalities in clinical laboratory evaluations will be summarized by 
treatment group. 
10.6.3. 
Efficacy Analysis 
The primary analysis will be the comparison of change from Baseline to Study Day 85 in SBP 
between each ISIS 757456 treated group and pooled placebo group in the PPS.  The data will be 
analyzed using analysis of covariance (ANCOVA) with treatment, and randomization 
stratification factor (Screening eGFR result of ˂ 60 or ≥ 60 mL/min/1.73 m2), and baseline 
measure (SBP) as independent variables.  In the case data departs substantially from normality, 
the nonparametric test will be employed instead.   
For the dichotomous endpoints, logistic regression model will be used.   
The secondary efficacy analyses will be performed in a similar way to the primary analysis, 
which include: 
• Comparison of change from Baseline to Study Day 85 in SBP and DBP from 24 hr 
mean ABPM between each ISIS 757456 treated group and pooled placebo group in 
the PPS and FAS 
• Comparison of change and percentage change from Baseline to each schedule post 
Baseline visit in plasma AGT between each ISIS 757456 treated group and pooled 
placebo group in PPS and FAS 
• Comparison of proportion of patients reached the goal of SBP ≤ 140 mmHg, DBP 
≤ 90 mmHg, and both for each scheduled visit between each ISIS 757456 treated 
group and pooled placebo group in PPS and FAS, excluding patients with baseline 
SBP ≤ 140 mmHg 
• Comparison of proportion of patients reached the goal of SBP ≤ 130 mmHg, DBP 
≤ 80 mmHg, and both for each scheduled visit between each ISIS 757456 treated 
group and pooled placebo group in PPS and FAS 
• Comparison of change from Baseline to each schedule post-Baseline visit in SBP 
between each ISIS 757456 treated group and pooled placebo group in PPS and FAS 
• Comparison of change from Baseline to each schedule post Baseline visit in DBP 
between each ISIS 757456 treated group and pooled placebo group in PPS and FAS 
10.6.4. 
Pharmacokinetic Analysis 
Plasma ISIS 757456 concentrations at trough during the Treatment Period and concentrations 
observed during the Post-Treatment Evaluation Period will be listed by dose, study day, time 
point, and summarized using descriptive statistics. 
Other plasma PK parameters, as appropriate, may be determined or calculated at the discretion of 
the PK scientist. 


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Metabolite identification and profiling may be conducted on select plasma samples. 
Additional details regarding the PK analysis will be described in the Statistical Analysis Plan 
(SAP). 
Analysis of potential exposure-response relationship between plasma AGT and ISIS 757456 
exposure (such as CRtrough
R) will be conducted.  Relationships between other relevant biomarkers 
and clinical endpoints with PK measures may also be explored, if deemed appropriate.   
Population PK and PK/PD analysis may be performed using PK data from this Study, and/or 
combined with other ISIS 757456 clinical PK/PD data later in the development timeline. 
10.6.5. 
Exploratory Analyses 
The change and percent change from Baseline to each schedule post-Baseline visit in 
angiotensin II and renin (PRA; direct renin) will be compared between each ISIS 757456 treated 
group and pooled placebo group in PPS and FAS.  The data will be analyzed in a similar way to 
the primary analysis. 
Change from Baseline to each schedule post-Baseline visit in SF-12 scores (domain, physical 
component summary, and mental component summary) and EQ-5D-5L (dimension, visual acuity 
score, index score) will be compared between each ISIS 757456 treated group and pooled 
placebo group in PPS and FAS.   
Proportion of improvement from baseline to each scheduled post Baseline in PGI-S and PGI-C 
will be compared between each ISIS 757456 treated group and pooled placebo group in PPS and 
FAS. 
Proportion of each questionnaire over time in POP-HSQ will be compared between each 
ISIS 757456 treated group and pooled placebo group in PPS and FAS. 
The analyses will be detailed in the SAP.   
11. 
INVESTIGATOR’S REGULATORY OBLIGATIONS 
11.1. 
Informed Consent 
The written informed consent document should be prepared in the language(s) of the potential 
patient population, based on an English version provided by the Sponsor. 
Before a patient’s participation in the trial, the Investigator is responsible for obtaining written 
informed consent from the patient 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 Drug (ISIS 757456 or placebo) is administered.  The patient must be 
given sufficient time to consider whether to participate in the study.   
The acquisition of informed consent and the patient’s agreement or refusal to notify his/her 
primary care physician should be documented in the patient’s medical records and the informed 
consent form should be signed and personally dated by the patient and by the person who 
conducted the informed consent discussion (not necessarily an Investigator).  The original signed 
informed consent form should be retained in the Study Master File and in any other locations 


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required by institutional policy, and a copy of the signed consent form should be provided to the 
patient. 
11.2. 
Ethical Conduct of the Study 
All applicable regulations and guidelines of current Good Clinical Practice (GCP), the 
Declaration of Helsinki as well as the demands of national drug and data protection laws and 
other applicable regulatory requirements must be followed.   
11.3. 
Independent Ethics Committee/Institutional Review Board 
A copy of the protocol; proposed informed consent form, other written patient information, and 
any proposed advertising material must be submitted to the IRB 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 patients 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 must 
also be received by the Sponsor before recruitment of patients into the study and shipment of 
Study Drug.  The Investigator’s Brochure must be submitted to the IRB for acknowledgement. 
The Investigator must submit to and, where necessary, obtain approval from the IRB for all 
subsequent protocol amendments and changes to the informed consent document.  The 
Investigator should notify the IRB of deviations from the protocol in accordance with ICH GCP.  
The Investigator should also notify the IRB of SAEs occurring at the Study Center and other AE 
reports received from the Sponsor, in accordance with local procedures. 
The Investigator will be responsible for obtaining annual IRB approval/renewal throughout the 
duration of the study.  Copies of the Investigator’s reports, all IRB submissions and the IRB 
continuance of approval must be sent to the Sponsor. 
11.4. 
Patient Confidentiality 
The Investigator must ensure that the patient’s confidentiality is maintained.  On the CRFs or 
other documents submitted to the Sponsor, patients should be identif 
ied by initials (if permitted by local law) and a patient identification number only.  Documents 
that are not for submission to the Sponsor (e.g., signed informed consent 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 direct access to review the patient’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 patient to permit 
named representatives to have access to his/her study-related records without violating the 
confidentiality of the patient. 


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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 must be informed of all 
amendments and give approval for any amendments likely to affect the safety of the patients or 
the conduct of the trial.  The Investigator must send a copy of the approval letter from the IRB to 
the Sponsor. 
12.2. 
Study Termination 
The Sponsor reserves the right to terminate the study.  The Investigator reserves the right to 
terminate their participation in the study, according to the terms of the site contract.  The 
Investigator should notify the IRB in writing of the trial’s completion or early termination and 
send a copy of the notification to the Sponsor or designee. 
12.3. 
Study Documentation and Storage 
An eCRF utilizing an Electronic Data Capture (EDC) application will be used for this study. 
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 patient’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, eCRF may not be 
used as source documents. 
The Investigator and Study Center staff are responsible for maintaining a comprehensive and 
centralized filing system of all study-related (essential) documentation in accordance with 
ICH GCP, suitable for inspection at any time by representatives from the Sponsor and/or 
applicable regulatory authorities.  Elements should include: 
• Patient 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 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, final 
Study Drug product reconciliation, 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. 


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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 patient 
confidentiality is respected. 
The Sponsor monitor or designee 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 access to patient 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 
patient data received by the Sponsor.  During this review, patient 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 
Case report forms must be completed in English.  Generic names and trade names are acceptable 
for concomitant medications.  Combination medications should be recorded using their trade 
name. 
All written information and other material to be used by patients 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.  Patients 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 
Altmann, K-H, Dean NM, Fabbro D, et al. Second generation of antisense oligonucleotides: 
From nuclease resistance to biological efficacy in animals. CHIMIA Int J Chem 1996; 50: 
168‑176. 
Carey, RM. Newly discovered components and actions of the renin-angiotensin system. 
Hypertension 2013; 62: 818-822. 
Crooke, ST, and Bennett CF. Progress in antisense oligonucleotide therapeutics. Annu Rev 
Pharmacol Toxicol 1996; 36: 107-129. 
Draman, MS, Dolan E, van der Poel L, et al. The importance of night-time systolic blood 
pressure in diabetic patients: Dublin Outcome Study. J Hypertens 2015; 33: 1373-1377. 
Ettehad, D, Emdin CA, Kiran A, et al. Blood pressure lowering for prevention of cardiovascular 
disease and death: a systematic review and meta-analysis. The Lancet 2016; 387: 957-967. 
Geary, RS, Yu RZ, Watanabe T, et al. Pharmacokinetics of a tumor necrosis factor-alpha 
phosphorothioate 2'-O-(2-methoxyethyl) modified antisense oligonucleotide: comparison across 
species. Drug Metab Dispos 2003; 31: 1419-1428. 
Henry, SP, Kim T-W, Kramer-Strickland K, et al. Toxicologic Properties of 2'-O-Methoxyethyl 
Chimeric Antisense Inhibitors in Animals and Man. In In Antisense Drug Technology, 
Principles, Strategies and Applications,2008. 327-364. Boca Ratan, FL: Taylor & Francis 
Group.2. 
Inoue, H, Hayase Y, Iwai S, et al. Sequence-dependent hydrolysis of RNA using modified 
oligonucleotide splints and RNase H. FEBS Lett 1987; 215: 327-330. 
Judd, E, and Calhoun DA. Apparent and true resistant hypertension: definition, prevalence and 
outcomes. J Hum Hypertens 2014; 28: 463-468. 
Korsnes, JS, Davis KL, Ariely R, et al. Health care resource utilization and costs associated with 
nonfatal major adverse cardiovascular events. J Manag Care Spec Pharm 2015; 21: 443-450. 
Lawes, CM, Vander Hoorn S, and Rodgers A. Global burden of blood-pressure-related disease, 
2001. Lancet 2008; 371: 1513-1518. 
Levey, AS, Stevens LA, Schmid CH, et al. A new equation to estimate glomerular filtration rate. 
Ann Intern Med 2009; 150: 604-612. 
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 Biol Chem 1999; 
274: 1715-1722. 
Monia, BP, Lesnik EA, Gonzalez C, et al. Evaluation of 2'-modified oligonucleotides containing 
2'-deoxy gaps as antisense inhibitors of gene expression. J Biol Chem 1993; 268: 14514-14522. 
Morgan, E, Tami  Y, Hu K, et al. Antisense Inhibition of Angiotensinogen With IONIS-AGT-
LRx : Results of Phase 1 and Phase 2 Studies. J Am Coll Cardiol Basic Trans Science In Press. 
Neunert, C, Terrell DR, Arnold DM, et al. American Society of Hematology 2019 guidelines for 
immune thrombocytopenia. Blood Adv 2019; 3: 3829-3866. 
Prakash, TP, Graham MJ, Yu J, et al. Targeted delivery of antisense oligonucleotides to 
hepatocytes using triantennary N-acetyl galactosamine improves potency 10-fold in mice. 
Nucleic Acids Res 2014; 42: 8796-8807. 


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Provan, D, Arnold DM, Bussel JB, et al. Updated international consensus report on the 
investigation and management of primary immune thrombocytopenia. Blood Adv 2019; 3: 3780-
3817. 
Provan, D, Stasi R, Newland AC, et al. International consensus report on the investigation and 
management of primary immune thrombocytopenia. Blood 2010; 115: 168-186. 
Schulman, S, and Kearon C. Definition of major bleeding in clinical investigations of 
antihemostatic medicinal products in non-surgical patients. J Thromb Haemost 2005; 3: 692-694. 
Sigmund, CD, Carey RM, Appel LJ, et al. Report of the National Heart, Lung, and Blood 
Institute Working Group on Hypertension: Barriers to Translation. Hypertension 2020; 75: 902-
917. 
Stockert, RJ. The asialoglycoprotein receptor: relationships between structure, function, and 
expression. Physiol Rev 1995; 75: 591-609. 
Te Riet, L, van Esch JH, Roks AJ, et al. Hypertension: renin-angiotensin-aldosterone system 
alterations. Circ Res 2015; 116: 960-975. 
Zhang, H, Lowenberg EC, Crosby JR, et al. Inhibition of the intrinsic coagulation pathway factor 
XI by antisense oligonucleotides: a novel antithrombotic strategy with lowered bleeding risk. 
Blood 2010; 116: 4684-4692. 
 


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APPENDIX A. SCHEDULE OF PROCEDURES 
 
 






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Appendix A Schedule of Procedures (Continued) 
8 Height at Screening only 
9 Patients who terminate treatment or post-treatment early from the study should be encouraged to participate in an early termination visit, at which time the Study D85 or Study D169 
assessments should be conducted, respectively 
10 Home Healthcare assessments and procedures may be conducted by a Home Healthcare professional at specified visits and as arranged by the site staff.  If 3 consecutive Home 
Healthcare visits have been conducted, the Sponsor must be consulted prior to the scheduling of a 4th in-home visit 
11 24-hr ABPM should take place after the visit 1 Screening labs have been resulted and the results are deemed not to be exclusionary.  Ensure sufficient time during the Screening 
Period to allow for a repeat (if the first 24-hr monitoring is not successful) prior to the Study Day 1 visit  
12 Study Drug may be administered by a trained Home Healthcare Professional  
13 For patients who have been administered 8 or more doses of Study Drug, a 24-hr ABPM should ideally be initiated in-clinic after all study procedures have been completed either on 
Study D78 or Treatment Early Termination/Study D85.  If a repeat is necessary, the 24-hr ABPM should be initiated no later than the Study D92 visit.  Ensure the 24-hr APBM is not 
done concurrently with other study procedures 
14 The study visit must occur in-clinic if the 24-hour ambulatory blood pressure is initiated 
15 Ideally should be completed prior to any interventional study procedures (e.g., blood draws, vital signs) 
16 Urine angiotensinogen is not required at Screening 
 
Time (time is in reference to Study Drug (ISIS 757456 or placebo) administration) 
a Pre-dose 
b Pre-dose, optional 2-hr post-dose   
 
 


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APPENDIX B. LIST OF LABORATORY ANALYTES 
 
 


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Appendix B List of Laboratory Analytes 
Based on emerging data from this or future studies, additional tests not listed below may be 
performed on stored samples to better characterize the profile of ISIS 757456 or other similar 
oligonucleotides. 
Clinical Chemistry 
Panel 
• Sodium 
• Potassium 
• Chloride 
• Bicarbonate 
• Total protein 
• Albumin 
• Calcium 
• Magnesium 
• Phosphorus 
• Glucose 
• BUN 
• Creatinine 
• Creatinine Clearance  
• Uric Acid 
• Total bilirubin 
• Direct (conjugated) 
bilirubin 
• Indirect 
(unconjugated) 
bilirubin 
• ALT 
• AST 
• ALP 
• Creatinine kinase 
• GGT 
 
Thyroid Panel 
• TSH 
• Free T4 
• Free T3 
Screening Tests 
• Hepatitis B surface 
antigen 
• Hepatitis C antibody 
• HIV antibody 
• FSH 
(women only, if 
applicable) 
• Serum βhCG  
(women only, if 
applicable) 
• Hemoglobin A1c 
 
Coagulation 
• aPTT (sec) 
• PT (sec) 
• INR 
 
Efficacy 
• Plasma AGT 
 
Pharmacokinetics1 
• ISIS 757456 
concentrations in 
plasma 
 
ADA 
• Anti-ISIS 757456 
antibodies3 
 
Hematology 
• Red blood cells 
• Hemoglobin 
• Hematocrit 
• MCV, MCH, MCHC 
• Platelets 
• White blood cells 
(WBC) 
• WBC Differential 
(% and absolute) 
• Neutrophils 
• Eosinophils 
• Basophils 
• Lymphocytes 
• Monocytes 
 
Exploratory 
• Angiotensin II 
• Renin (plasma renin 
activity) 
• Direct renin  
• Plasma aldosterone3 
• NT- pro B-type 
natriuretic peptide 
• NT-pro Atrial 
natriuretic peptide3  
• Endothelin I3 
Inflammatory 
• hs-CRP  
 
Urinalysis 
• Color 
• Appearance 
• Specific gravity 
• pH 
• Protein/Creatinine 
Ratio (UPCR) 
• Protein 
• Blood 
• Ketones 
• Urobilinogen 
• Glucose 
• Bilirubin 
• Leukocyte esterase 
• Nitrate 
• Microscopic 
examination2 
• Albumin 
• Creatinine 
• Protein  
• A/C Ratio (UACR) 
• Angiotensinogen 
1 Plasma PK samples may also be used for profiling of drug binding proteins, bioanalytical method validation 
purposes, stability assessments, metabolite assessments, or to assess other actions of ISIS 757456 with plasma 
constituents 
2 Will be performed on abnormal findings unless otherwise specified 
3 May be analyzed 


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APPENDIX C. PK SAMPLING SCHEDULE  
 
 




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APPENDIX D. GRADING SCALE FOR ADVERSE EVENTS RELATING 
TO LABORATORY ABNORMALITIES 
 
 


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Appendix D Grading Scale for Adverse Events Relating to Laboratory Abnormalities 
The following grading recommendations for adverse events relating to lab test abnormalities are 
based upon the Common Terminology Criteria for Adverse Events (CTCAE) Version 5.0, 
November 2017. 
 
 
 


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Version: 1
Version Date: 05 Nov 2021
Title: 757456-CS4 Amendment 4: A DB, Placebo-Controlled, Phase 2 Study to Assess the
Safety, Tolerability & Efficacy of IONIS-AGT-LRX, an Antisense Inhibitor of
Angiotensinogen Production Administered SC for 12 Wks to Hypertensive Pts with
Protocol


 
 
Statistical Analysis Plan 
 
ISIS 757456-CS4 
 
A Double-Blind, Placebo-controlled, Phase 2 Study to Assess the 
Safety, Tolerability and Efficacy of IONIS-AGT-LRX, an Antisense 
Inhibitor of Angiotensinogen Production Administered 
Subcutaneously for 12 Weeks to Hypertensive Patients with 
Uncontrolled Blood Pressure 
 
Date:  15 August 2022 
 
Version:  2.0 
 
 
Confidential 
Confidential/Trade 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 the 
express written permission of Ionis Pharmaceuticals, Inc 




Ionis Pharmaceuticals 
CONFIDENTIAL 
15 August 2022
Statistical Analysis Plan v2 
ISIS 757456-CS4
 
3 
TABLE OF CONTENTS 
SPONSOR SIGNATURE PAGE ....................................................................................................2 
REVISION HISTORY.....................................................................................................................5 
ABBREVIATIONS .........................................................................................................................6 
1. 
INTRODUCTION ........................................................................................................7 
1.1. 
Study Overview ............................................................................................................7 
1.2. 
Objectives .....................................................................................................................9 
1.2.1. 
Primary Objective .........................................................................................................9 
1.2.2. 
Secondary Objectives ...................................................................................................9 
1.2.3. 
Safety Objectives ..........................................................................................................9 
1.2.4. 
Exploratory Objectives .................................................................................................9 
1.3. 
Endpoints ....................................................................................................................10 
1.3.1. 
Primary Endpoint ........................................................................................................10 
1.3.2. 
Secondary Endpoints ..................................................................................................10 
1.3.3. 
Safety Endpoints .........................................................................................................10 
1.3.4. 
Exploratory Endpoints ................................................................................................10 
2. 
PROCEDURES ..........................................................................................................11 
2.1. 
General Overview of Procedures ................................................................................11 
2.2. 
Randomization & Treatment Allocation ....................................................................11 
2.3. 
Conduct .......................................................................................................................11 
2.4. 
Data Monitoring ..........................................................................................................12 
2.4.1. 
Safety Data Monitoring ..............................................................................................12 
2.5. 
Data Management .......................................................................................................12 
2.5.1. 
Case Report Form (CRF) Data ...................................................................................12 
2.5.2. 
Laboratory Data ..........................................................................................................12 
2.5.3. 
Pharmacokinetics (PK) and Immunogenicity (IM) Data ............................................13 
3. 
ANALYSIS PLAN .....................................................................................................13 
3.1. 
Statistical Design Summary ........................................................................................13 
3.2. 
General Overview of Analyses ...................................................................................14 
3.2.1. 
Statistical Methods ......................................................................................................14 
3.2.2. 
Subject Population Analyzed ......................................................................................15 
3.2.3. 
Handling of Missing Data ...........................................................................................16 


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3.2.4. 
Disposition of Subjects ...............................................................................................16 
3.2.5. 
Demographic and Baseline Characteristics ................................................................16 
3.2.6. 
Protocol Deviations ....................................................................................................17 
3.2.7. 
Stratification, Subsets, and Covariates .......................................................................17 
3.2.8. 
Scoring of Questionnaires ...........................................................................................18 
3.3. 
Primary Analysis ........................................................................................................19 
3.3.1. 
Primary Efficacy Analysis ..........................................................................................19 
3.3.2. 
Sample Size consideration ..........................................................................................19 
3.4. 
Secondary Efficacy Analyses .....................................................................................19 
3.4.1. 
Secondary Efficacy Endpoints ....................................................................................19 
3.4.2. 
Multiplicity .................................................................................................................20 
3.4.3. 
Interim Analysis ..........................................................................................................21 
3.5. 
Exploratory Analyses ..................................................................................................21 
3.6. 
Pharmacokinetic (PK) and Immunogenicity (IM) Analysis .......................................22 
3.6.1. 
PK Analysis ................................................................................................................22 
3.6.2. 
Pharmacokinetic/Pharmacodynamic Exposure-Response Analysis ...........................23 
3.6.3. 
IM Analysis ................................................................................................................24 
3.7. 
Safety Analyses ..........................................................................................................26 
3.7.1. 
Exposure .....................................................................................................................26 
3.7.2. 
Adverse Events ...........................................................................................................26 
3.7.3. 
Laboratory Measurements ..........................................................................................28 
3.7.4. 
2.12.4 Vital Signs .......................................................................................................29 
3.7.5. 
12-Lead Electrocardiograms (ECG) ...........................................................................30 
3.7.6. 
Prior and Concomitant Medications ...........................................................................30 
4. 
REFERENCES ...........................................................................................................30 
APPENDIX 1. EQ-5D HEALTH STATE INDEX SCORE DERIVATION 
ALGORITHM ......................................................................................................31 
LIST OF TABLES 
Table 1: 
Weight of EQ-5D-5L Response by Region ................................................................31 
LIST OF FIGURES 
Figure 1: 
Testing Strategy illustration ........................................................................................21 
 


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REVISION HISTORY 
Version No. 
Effective Date 
Summary of Changes 
1.0 
28 March 2022 
New Document 
2.0 
1 July 2022 
Updated language for Section 3.5, Exploratory Analyses endpoints 
for PGI-C, PGI-S and POP-HSQ.  
• Section 3.2.1, for the continuous endpoints, Van Elteren 
test with treatment and randomization stratification factor 
will be conducted  if the data departs substantially from 
normality.  Changed from rank ANCOVA to Van Elteren 
test. 
• Section 3.7.3, modified potassium (K) categories to 
“Absolute levels ≥ 5.5–< 6, confirmed”; “Absolute levels 
≥ 6, confirmed”.  Removed Platelet count first category 
“Confirmed Platelet count > 100,000/mm3”.  Clarified 
local lab data will be included when derive confirmed 
abnormal results. 
 
 
 


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ABBREVIATIONS 
In addition to the study glossary provided in the protocol, here are additional abbreviations 
provided for the study SAP. 
Abbreviation  
 
Definition 
ABP   
 
 
ambulatory blood pressure 
ABPM  
 
 
ambulatory blood pressure monitoring 
AE  
 
 
 
adverse event 
AGT   
 
 
angiotensinogen 
ALP   
 
 
alkaline phosphatase 
ALT   
 
 
alanine aminotransferase (SGPT)  
ANCOVA 
 
 
Analysis of Covariance 
BMI   
 
 
body mass index 
BP  
 
 
 
blood pressure 
CI 
 
 
 
confidence interval 
CRF   
 
 
case report form  
CVD  
 
 
cardiovascular disease 
CSR   
 
 
clinical study report 
DBP   
 
 
diastolic blood pressure 
ECG   
 
 
electrocardiogram 
eCRF   
 
 
electronic Case Report Form 
eGFR   
 
 
estimated glomerular filtration rate 
EQ-5D-5L  
 
 
EuroQoL-5 Dimension 5 Level 
FAS   
 
 
full analysis set 
HTN   
 
 
hypertension 
LCRIS  
 
 
Local cutaneous reaction at injection site  
LLQ   
 
 
lower limit of quantitation  
MMRM 
 
 
mixed models with repeated measures 
PD 
 
 
 
pharmacodynamic(s) 
PGI-C   
 
 
Patient Global Impression- Change 
PGI-S   
 
 
Patient Global Impression- Severity 
PK  
 
 
 
pharmacokinetic(s) 
POP-HSQ  
 
 
Power Over Pressure- Hypertension Specific Questionnaire 
PPS  
 
 
 
per protocol set 
SAE   
 
 
serious adverse event 
SAP   
 
 
statistical analysis plan 
SBP   
 
 
systolic blood pressure 
SF-12   
 
 
12-Item Short Form Survey 
TEAE   
 
 
treatment-emergent adverse event 
UACR  
 
 
urinary albumin/creatinine ratio 
ULN   
 
 
upper limit of normal 


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1. 
INTRODUCTION 
This document provides a description of the study organization, study procedures, and the plan 
for the statistical analysis of the study data.  Section 1 discusses study design, objectives, and 
endpoints; Section 2 provides the study procedures; Section 3 provides the detailed plan for the 
statistical analyses. 
The purpose of this plan is to provide specific guidelines from which the analysis will proceed.  
Any deviations from these guidelines will be documented in the clinical study report (CSR). 
Within this document, the terms ‘patient’ and ‘subject’ are both used to describe the individual 
who enrolls in this study. 
1.1. 
Study Overview 
This is a Phase 2, double-blind, randomized, placebo-controlled study of ISIS 757456 conducted 
in uncontrolled hypertensive patients on ≥ 3 antihypertensive medications.  Eligible patients will 
be stratified based on a screening eGFR result (<60 vs. ≥ 60 mL/min/1.73 m2) and randomized 
to 1 of 2 dose cohorts in a 1:1 ratio (Cohort A (80 mg ISIS 757456 or placebo) or Cohort B 
(120mg ISIS 757456 or placebo)).  Within each dose cohort, patients will be further randomized 
to receive ISIS 757456 or placebo in a 2:1 ratio.  Patients will receive SC doses of Study Drug on 
Study Days 1, 8, 15, 22, 29, 36, 43, 50, 57, 64, 71, and 78 during treatment period.  Patients will 
continue their antihypertensive medication regimen throughout the study.  The patient is 
expected to maintain adequate hydration and a low sodium diet throughout the study. After the 
Treatment Period, all patients will then complete a 13-week Post-Treatment Period.  
This study will be conducted at multiple centers in the United States.  Approximately 
150 patients are planned to be enrolled in this study.  Study Drug (ISIS 757456 or placebo) 
injection volumes will be 0.8 mL or 1.2 mL for cohort A or Cohort B, respectively.  All Study 
Drug injections will be administered by qualified personnel at the clinic or by a home healthcare 
professional (as arranged with the site staff and after appropriate training). 
The study will consist of Screening, Treatment, and Post-treatment.  The overall length of a 
subject’s participation will be approximately 31 weeks (up to 6 weeks for Screening, a 12-week 
Treatment Period, and a 13-week Post-Treatment Evaluation Period). 


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The study design and treatment schema are depicted as follows: 
 
TREATMENT PERIOD 
Study Days 1, 8, 15, 22, 29, 36, 43, 
50, 57, 64, 71, 78 
POST-TREATMENT 
EVALUATION PERIOD 
Study Days 85-169 
EARLY 
TREATMENT 
TERMINATION 
POST-TREATMENT 
EVALUATION 
PERIOD 
13 Wks 
(ET/ Study Days 85, 92, 
106, 120, 148,169) 
Patient Withdrawn 
from Study or 
Consent Withdrawn 
WITHDRAWAL 
FROM STUDY 
Early Term Visit 
(Study Day 85) 
SCREEN 
Study Day -42 to -1 
END-OF-STUDY 
PROCEDURES 
Post-Treatment Early Termination/ 
Study Day 169 
Screen Failure 
Treatment Terminated; 
Consent Not Withdrawn 


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1.2. 
Objectives 
1.2.1. 
Primary Objective 
To evaluate the effect of ISIS 757456 compared to placebo on seated automated office systolic 
blood pressure (SBP) from Baseline to Study Day 85 in uncontrolled hypertensive patients on 
≥ 3 antihypertensive medications 
1.2.2. 
Secondary Objectives 
• To evaluate the effect of ISIS 757456 on plasma angiotensinogen (AGT) 
concentration at each scheduled visit in uncontrolled hypertensive patients on 
≥ 3 antihypertensive medications 
• To evaluate the effect of ISIS 757456 on 24-hour ambulatory blood pressure at 
scheduled visits in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate the effect of ISIS 757456 on seated automated office SBP at each 
scheduled visit in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate the effect of ISIS 757456 on seated automated office diastolic blood 
pressure (DBP) at each scheduled visit in uncontrolled hypertensive patients on 
≥ 3 antihypertensive medications 
1.2.3. 
Safety Objectives 
To evaluate the safety and tolerability of ISIS 757456 in uncontrolled hypertensive patients on 
≥ 3 antihypertensive medications 
1.2.4. 
Exploratory Objectives 
• To evaluate the effects of ISIS 757456 administered subcutaneously (SC) 
(e.g., angiotensin II, renin [Plasma renin activity (PRA); direct renin], urinary 
angiotensinogen) in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate the pharmacokinetics (PK) of ISIS 757456 (as total full-length ASO, 
including fully conjugated, partially conjugated, and unconjugated ISIS 757456) 
administered SC in uncontrolled hypertensive patients on ≥ 3 antihypertensive 
medications 
• To evaluate potential PK/ pharmacodynamic (PD) correlation with relevant 
biomarkers and/or clinical endpoints 
• To evaluate the effects of ISIS 757456 on patients reported quality of life outcomes 


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1.3. 
Endpoints 
1.3.1. 
Primary Endpoint 
Change in seated automated office SBP from baseline to Study Day 85  
1.3.2. 
Secondary Endpoints 
• Absolute levels and change and percent change in plasma AGT concentration from 
baseline to each scheduled, post-baseline visit  
• Changes from Baseline to Study Day 85 in 24-hour mean SBP and DBP measured by 
ambulatory blood pressure monitoring (ABPM) 
• Percentage of patients reaching the goals of seated automated office 
SBP ≤ 140 mmHg, DBP ≤ 90 mmHg, and both for each scheduled, post-Baseline 
visit (excluding patients with a baseline SBP of ≤ 140 mmHg) 
• Percentage of patients reaching the goals of automated office seated 
SBP ≤ 130 mmHg, DBP ≤ 80 mmHg, and both for each scheduled, post-Baseline 
visit 
• Change on seated automated office SBP from Baseline to each scheduled, 
post-Baseline visit 
• Change on seated automated office DBP from Baseline to each scheduled, 
post-Baseline visit 
1.3.3. 
Safety Endpoints 
Incidence and severity of treatment-emergent adverse events (TEAE), use of concomitant 
medications, abnormal findings in laboratory assessments, electrocardiogram (ECG), and vital 
signs 
1.3.4. 
Exploratory Endpoints 
• Absolute levels and change and percent change of angiotensin II, renin (PRA; direct 
renin), urinary AGT concentration from Baseline to each scheduled, post-Baseline 
visit 
• Summary of peak and trough ISIS 757456 plasma concentrations 
• Potential exposure-response analysis using relevant exposure parameters and 
biomarkers including, but not limited to trough concentration (Ctrough) and plasma 
AGT may be performed 
• Changes from Baseline to Study Day 85 and each scheduled post-Baseline 
assessment in the following patient reported outcome questionnaires:  EuroQol-5 
Dimension 5 Level (EQ-5D-5L), 12-Item Short Form Survey (SF-12), Power Over 
Pressure- Hypertension Specific Questionnaire (POP-HSQ), Patient Global 
Impression- Change (PGI-C), and Patient Global Impression-Severity (PGI-S) 


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o Change from Baseline to Study Day 85 in daytime SBP and DBP from 24 hr 
ABPM  
o Change from Baseline to Study Day 85 in nighttime SBP and DBP from 24 hr 
ABPM  
o Change from Baseline to Day 85 in eGFR  
o Change from Baseline to Day 85 in urinary albumin/creatinine ratio (UACR)  
o Primary endpoint, selective secondary and exploratory endpoints will be also 
analyzed using a sub-population of patients with baseline 24 hr ABPM SBP > 130  
2. 
PROCEDURES 
2.1. 
General Overview of Procedures 
Ionis Pharmaceuticals, Inc. (or designee) will review all study data including source documents, 
case report forms, and laboratory reports.  The study site will enter subject source data into the 
case report form.  Central laboratory data will be transferred electronically from Medpace 
Reference Laboratories (MRL) Inc. for those analytes noted in the MRL Laboratory Services 
Agreement, PK Plasma concentration data will be from Pharmaceutical Product Development 
(PPD) Inc., and immunogenicity data will be from Charles River Laboratories in Montreal 
(CR-SEN) to Ionis Pharmaceuticals, Inc.  Additionally, plasma AGT,  Ang II, aldosterone and 
PRA data analyzed by Attoquant Diagnostics GmbH will be transferred to Ionis via separate 
Data Transfer Agreement (DTA). 
2.2. 
Randomization & Treatment Allocation 
Patients will be randomized after all screening assessments have been completed and after the 
Investigator has verified that they are eligible per criteria in Clinical Study Protocol Sections 5.1 
and 5.2.  No subject may begin treatment prior to randomization and assignment of a unique 
subject identification number.   
Eligible patients will be stratified based on a screening eGFR result (< 60 vs. 
≥ 60 mL/min/1.73 m2) and then patients will be randomized to 1 of 2 dose cohorts in a 1:1 ratio 
(Cohort A or Cohort B).  Within each dose cohort, patients will be further randomized in a 2:1 to 
receive ISIS 757456 or placebo as outlined in Clinical Study Protocol Section 4.2.  The Sponsor 
or designee will prepare the randomization list and utilize an automated IRT (Interactive 
Response Technology) system for randomization assignment. 
2.3. 
Conduct 
The study will be conducted in accordance with current Good Clinical Practice (GCP) and 
International Conference on Harmonization (ICH) guidelines, the World Medical Association 
Declaration of Helsinki guidelines, the Food and Drug Administration (FDA) Code of Federal 
Regulations, and all other local regulatory requirements. 


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2.4. 
Data Monitoring 
2.4.1. 
Safety Data Monitoring 
Ionis Pharmaceuticals, Inc. (or designee) is responsible for processing all reported adverse events 
(AEs).  All serious adverse events (SAEs), reported to Ionis Pharmaceuticals, Inc. (or designee), 
are reviewed according to standard operating procedures.  The medical monitor 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 worldwide in 
accordance with local requirements.  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 site. 
2.5. 
Data Management 
An electronic case report form (eCRF) utilizing an Electronic Data Capture (EDC) application 
will be used for this Study. 
2.5.1. 
Case Report Form (CRF) Data 
Clario (Formerly BioClinica) (or designee) is responsible for creating the 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 
data that 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 is corrected or an explanation concerning the 
query is provided in the EDC system.  After all data is entered, source data verified, reviewed 
and queried the database is closed, the data is then reviewed by Ionis Pharmaceuticals, Inc. and 
additional queries may be generated if necessary.  After all queries are resolved and PI signature 
are received, the database is locked. 
2.5.2. 
Laboratory Data  
Ionis Pharmaceuticals, Inc. is responsible for the format of the laboratory electronic data 
transfers, the transfer schedule, and the review of the clinical laboratory data.  Investigator sites 
have access to safety data via lab reports sent directly from the laboratory and online using the 
MRL ClinTrack® database. The laboratory data will be stored as SAS datasets. 
Plasma AGT, Ang II, aldosterone, and PRA by Attoquant Diagnostics GmbH will be transferred 
to Ionis via separate Data Transfer Agreement (DTA). 


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2.5.3. 
Pharmacokinetics (PK) and Immunogenicity (IM) Data 
Ionis Pharmaceuticals, Inc. is responsible for the management and review of the plasma drug 
ISIS 757456 concentration data and immunogenicity sample analysis.  This review process 
involves reconciling the patient and visit identifiers (e.g. patient demographics) with the clinical 
data collected in the EDC system.  The final PK data will be stored as CSV and/or EXCEL files 
per the Analysis Plan. 
3. 
ANALYSIS PLAN 
3.1. 
Statistical Design Summary 
This is a Phase 2, randomized, placebo-controlled, multi-center study.  Approximated 
150 patients will be stratified based on a screening eGFR result (< 60 vs. ≥ 60 mL/min/1.73 m2) 
and randomized to 1 of 2 dose cohorts in a 1:1 ratio (Cohort A or Cohort B).  Within each dose 
cohort, patients will be further randomized to receive ISIS 757456 or placebo in a 2:1 ratio.  
All planned analyses will compare each ISIS 757456 treatment group to pooled Placebo.  The 
overall study type I error is at 0.1 level, two-sided.  The multiplicity of study treatment groups 
comparison and study endpoints testing sequence will be further described in Section 3.4.2.  
The planned sample size will provide approximate 80% of power to demonstrate the mean SBP 
reduction difference of 8 mmHg between each ISIS 757456 treated group and pooled placebo 
based on the assumption of pooled standard deviation of 13 mmHg with the two-sided test and a 
significant level of 0.05.  
An interim analysis may be conducted after approximately 50% of patients have been enrolled.  
The unblinding information dissemination during this interim analysis will be controlled and 
described in the request to unblind data for study analysis document to ensure the trial integrity.  
Please refer to Section 3.3.2 for further details. 
The primary analysis will take place after all patients complete Treatment Period (Day 85) 
assessments and the database has been locked.  The final analysis will be performed after all 
patients have completed or have had the opportunity to complete the study Day 169 assessments 
(such as patients who have terminated early) and the database has been locked.   
The primary endpoint is the change from baseline to Study Day 85 in seated automated office 
SBP and will be analyzed by analysis of covariance (ANCOVA) with treatment, randomization 
stratification factor and baseline measure (SBP) as independent variables.  The statistical 
inferences will be summarized by treatment group.  Analyses will assess the hypothesis there are 
difference in the SBP change from baseline at Day 85 between each ISIS 757456 treatment 
group and pooled placebo.  The estimated treatment effect with 95% CI and p-values will be 
provided.   
Plasma AGT concentration from Attoquant will be used for AGT analysis. 


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3.2. 
General Overview of Analyses  
3.2.1. 
Statistical Methods 
The study outcomes will be summarized by treatment group.  Descriptive summary statistics 
including number of patients, mean, median, standard deviation, standard error of mean, 25th 
percentile, 75th percentile, minimum, and maximum for continuous variables, and counts and 
percentages for categorical variables will be provided to summarize most data.  The efficacy 
endpoints will be assessed on the full analysis set (FAS) and per-protocol set (PPS) with the 
latter being the basis for the primary efficacy analysis.  The safety analyses will be performed on 
the Safety Set.  Pharmacokinetic analysis will be conducted in the PK Set.   
All planned analyses will compare each ISIS 757456 treatment group to pooled placebo. 
All statistical tests will be conducted using 2-sided tests with 5% Type I error rate unless 
otherwise stated.  P-values will be provided to assess the null hypothesis that there is no 
statistical difference between an ISIS 757456 treatment group and pooled placebo in study 
endpoints.  The stratification factor (screening eGFR status [<60 vs. ≥ 60 mL/min/1.73 m2]), 
treatment received, and baseline measure are included in the Analysis of Covariate (ANCOVA) 
model as independent variables.  For the continuous endpoints, the ANCOVA model will be 
applied or Van Elteren test with treatment and randomization stratification factor  if the data 
departs substantially from normality.  The normality assumption will be assessed by the Shapiro-
Wilk test based on the residuals from ANCOVA modeling.  Least Square mean (LSM) 
difference with 95% CI and p-values between treatment groups in study endpoints will be 
provided based on ANCOVA model.  For the dichotomous endpoints, logistic regression model 
with treatment, randomization stratification factor and baseline measure (SBP) as independent 
variables will be used instead.   
To evaluate treatment difference across scheduled post-baseline visits, an additional mixed 
model with repeated measures (MMRM) analysis may be utilized.  Besides stratification factor, 
visit, treatment received, baseline measure, and treatment-by-visit interaction will be included in 
the model, which visit as an ordinally categorical variable.  An unstructured covariance will be 
used initially, with correlation coefficients being greater for visits that are closer.  Other 
covariance structures, such as first-order autoregressive covariance structure based on the 
assumption of the measurements over time are expected to be correlated, may be considered 
based on model fitting consideration.  The adjusted mean with 95% CI for each treatment group 
and 95% CIs for the mean difference between pooled placebo and each ISIS 757456 treatment 
groups at each scheduled time point will be provided.  
Additional covariates, such as baseline characteristics, may be considered as an additional 
covariate in the comparison statistical modeling and for subgroup summary for the exploratory 
analysis purpose.  
Laboratory outcomes with lower limit of quantitation (LLQ) or less will be imputed as LLQ in 
the summary tables and figures.  
PK parameters will be summarized using include number of patients, mean, standard deviation, 
coefficient of variation (CV), geometric mean, median, minimum, and maximum. 


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Additional subject listings including case report form (CRF) data and derived outcomes from the 
data may be presented.  
Baseline Definition 
The baseline for plasma AGT concentration from Attoquant will be defined as the average of all 
values prior to the first dose of Study Drug.  The baseline for all other assessments will be 
defined as the last non-missing measurement prior to the first dose.   
Analytical Visits 
In general, all post-baseline data will be summarized using the visit labels provided in the data.  
Multiple results with the same visit label will be averaged for continuous variables, and the worst 
result will be used for categorical variables.  Results with visit labels as “Unscheduled” will not 
be included in by-visit summary tables and figures except for determining baseline and the 
incidence of abnormality lab summary including, but not limited to, AST, ALT, potassium, 
eGFR, urine protein/creatinine ratio, and platelet specified in Section 3.7.4, but will be presented 
in data listings.  Other situations, such as required additional confirmation, will be described 
separately in the related endpoint analysis approach. 
3.2.2. 
Subject Population Analyzed 
The following analysis populations are defined for this study: 
• Full Analysis Set (FAS):  All randomized patients who have received at least 
1 injection of Study Drug (ISIS 757456 or Placebo) and who have at least 
1 post-Baseline primary efficacy measurements. 
• Per Protocol Set (PPS):  All FAS patients who received at least 10 of the 12 doses of 
Study Drug, did not alter antihypertensive medications during the Treatment Period 
and prior to Study Day 85, and have no significant protocol deviations that would be 
expected to affect efficacy assessment.  Significant protocol deviations are defined as 
those deviations from the protocol likely to impact the perceived efficacy and/or 
safety of study treatments, for example, other antihypertensive medication usage 
compliance during the study.  After all data are entered, reviewed, and queried, the 
database is closed and sent to clinical group for review and identification of 
significant protocol deviations before unblinding. 
• Safety Set:  All patients who are randomized and receive at least 1 dose of 
Study Drug. 
• Pharmacokinetics (PK) Set:  All patients who are randomized and receive at least 
1 dose of ISIS 757456 and have at least 1 evaluable PK sample.   
ln addition to the above analysis sets, it is recognized that some data displays will be provided 
for the number of patients in "All Screened", "Screening Failures" and "All Randomized"  but no 
data analysis will be executed in these populations.  A by-patient listing based on all randomized 
will be provided. 


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3.2.3. 
Handling of Missing Data 
Unless otherwise specified, missing values will not be imputed. 
Patients with missing data for a scheduled assessment time point will be excluded from the 
summary for that time point.  
Number of patients with missing data and missing data related to COVID-19 public health 
emergency may be provided.   
3.2.4. 
Disposition of Subjects 
The number of patients screened, screen failures, the number and percentage of patients 
randomized, dosed, completed the study treatment, discontinued from treatment early and 
reason, completed post-treatment follow-up, and discontinued from post-treatment follow-up and 
reason will be tabulated by treatment group.   
A by-patient listing will also be provided. 
Additional by-patient listing will be provided to describe screen failures, treatment/post-
treatment follow-up termination information related COVID-19 public health emergency.   
3.2.5. 
Demographic and Baseline Characteristics  
Demographic and Baseline characteristics (e.g., age, gender, ethnicity, race, cardiovascular 
disease (CVD) risk factors, tobacco/nicotine use, number of antihypertensive medications 
received, diuretic use (Y/N), CVD history, diabetic status, weight, height, BMI) will be 
summarized using descriptive statistics by treatment group.    
Additionally, the following additional baseline characteristics will be summarized: 
• Age (yrs): < 65, ≥ 65 
• BMI (Kg/m2): < 30, ≥ 30 
• eGFR (mL/min/1,73 m2): ≥ 60, < 60 
• heart rate 
• Antihypertensive medications received by category (RAS inhibitor, beta-blocker, 
calcium channel blocker, diuretic, alpha-1 blocker, centrally acting sympatholytic 
agent, vasodilators, aldosterone antagonists)  
• ≥ 3 anti-HTN med with diuretic  
• 
Number anti-HTN medications 3, 4, ≥ 5 
• SBP and DBP (mmHg) at baseline 
• 24 hr ABPM SBP at baseline >130 SBP  
• Daytime ABPM SBP at baseline > 135 
• Plasma AGT 
• HbA1c 


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• eGFR 
• urinary albumin/creatinine ratio (UACR): normal:  < 30 mg/g was considered normal; 
micro-albuminuria: 30-300 mg/g; macro-albuminuria: > 300 mg/g  
• CV Risk 
For race summary, if multiple races are recorded in the database, then ‘Multiple Race’ will be 
used in the summary table.  The listing will display the specific race values. 
BMI will be computed using the formula:  
BMI = (weight in kilograms) / [screening height in cm / 100]2 
Listing of medical history will be provided.  
3.2.6. 
Protocol Deviations 
Protocol deviations will be classified to major or minor based on the study protocol deviation 
process plan.  Protocol deviations will be provided in the data listings.  Additional tables may be 
provided to summarize the protocol deviations related to COVID-19 public health emergency. 
A listing of all patients affected by the COVID-19 public health emergency related study 
disruption by subject number identifier and by investigational site, and a description of how the 
individual’s participation was altered will be provided.   
3.2.7. 
Stratification, Subsets, and Covariates 
Eligible Patients will be stratified on Screening eGFR result of ˂ 60 or ≥ 60 mL/min/1.73 m2.  
In the case of errors in stratification, the analyses should be based on the actual stratification, 
regardless of IRT entry.   
The following patient demographic and baseline characteristics may be considered as additional 
covariates in the comparison statistical modeling for exploratory analysis purposes:   
• Age group (< 65 years vs. ≥ 65 years) 
• Race (Caucasian vs non-Caucasian) 
• Gender 
• Obesity status 
• Number of antihypertensive medications received 
• Diabetic status 
• UACR categories 
• Two or more CV risk factors 
• 24 hr ABPM SBP > 130 mmHg 
• Chronic kidney disease status defined by eGFR (> 90 mL/min, 60-90 mL/min, and 
45-59 mL/min)   


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3.2.8. 
Scoring of Questionnaires 
EuroQoL-5 Dimensional 5-level Questionnaire (EQ-5D-5L) 
EQ-5D-5L is a standardized measure of health status developed by the EuroQoL Group to 
provide a simple, generic measure of health for clinical and economic appraisal.  The EQ-5D-5L 
consists of the EQ-5D-5L descriptive system and the EQ visual analog scale (VAS). 
The EQ-5D-5L descriptive system comprises five dimensions (mobility, self-care, usual 
activities, pain/discomfort and anxiety/depression), and the response to each dimension is scored 
as:  1 = no problems/pain/anxious, 2 = slight problems/pain/anxious, 3 = moderate 
problems/pain/anxious, 4 = severe problems/pain/anxious, 5 = unable/extreme.  Lower score 
indicates worsening of QoL. 
The EQ VAS is used as a quantitative measure of health outcome as judged by the individual 
respondent.  The EQ VAS records a self-rating of health status on a vertical VAS anchored at 
100 (best imaginable health state) at the top and 0 (worst imaginable health state) at the bottom. 
Additionally, the EQ-5D health state index score reflects how good or bad a health state is 
according to the preferences of the general population of a country/region will be derived and 
summarized.  If any of five dimensions is missing, then the index value will be set as missing.  
The details of EQ-5D health state index score derivation algorithm is described in Appendix 1. 
Patient Global Impression 
Patient Global Impression of Severity (PGI-S) is a single-item scale instrument asking patients to 
rate the severity of their health condition on a scale of (1) none, (2) mild, (3) moderate, 
(4) severe, (5) very severe. 
Patient Global Impression of Change (PGI-C) is a single-item scale, which aims to evaluate all 
aspects of patients’ health and determine if there has been an improvement or not in the patients’ 
overall change in health condition. Patients rate their change as (1) much better, (2) a little better, 
(3) no change, (4) a little worse, (5) much worse. 
Power Over Pressure - Hypertension Specific Questionnaire (POP-HSQ)  
The POP-HSQ (Schmider, et. al 2017) comprising questionnaire A and questionnaire B (what 
symptoms have you experienced in the last 4 weeks).  In questionnaire A, patients rate their 
health state in part 1 to part 4 and their blood pressure management and impact in part 5 to 
part 7.  In questionnaire B, patients provide the experienced symptoms’ frequency and intensity.    
Short Form (SF) -12  
The SF-12 comprising 12 items that yield 8 healthy domains profile of physical and mental 
well-being.  These 8 domains are: health domains: physical functioning (PF), role physical (RP), 
bodily pain (BP), general health (GH), vitality (VT), social functioning (SF), role emotional 
(RE), and mental health (MH).  Each health domain score contributes to the physical component 
summery (PCS, including PF, RP, GH and BP) and a mental component summary (MCS, 
including MH, RE, SF, and VT).  Other measures can be derived from SF-12 data, including 
SF-6D preference-based utility index and the medical expenditure prediction to help understand 
the economic impact of a patient’s condition and treatment.  The norm-based score for each 
domain and components will be calculated using Optum’s SF36v2/SF-12v2 scoring software 


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operated by Ionis data management function personal.  Higher scores are associated with a better 
quality of life (QOL).   
3.3. 
Primary Analysis 
3.3.1. 
Primary Efficacy Analysis 
The primary analysis will be the comparison of change from Baseline to Study Day 85 
(Study Week 13) in seated automated office SBP between each ISIS 757456 treatment dose 
group and pooled placebo group in the PPS.  The data will be analyzed as described in 
Section 3.2.1.  Least square mean (LSM) estimated treatment difference with 95% CI and 
p value will be provided.  Additional MMRM analysis will be conducted to assess the treatment 
effect.  
Additional covariates, such as baseline characteristics, may be considered, in the comparison 
statistical modeling for exploratory analyses. 
Additional analysis summary based on the FAS will also be provided.  
3.3.2. 
Sample Size consideration 
Approximately 150 patients will be stratified and randomized to 1 of 2 dose cohorts in 1:1 ratio 
(Cohort A or Cohort B).  Within each dose cohort, patients will be further randomized to receive 
ISIS 757456 or placebo in a 2:1 ratio.   
With the assumptions of mean seated automated office SBP reduction difference of 8 mmHg 
between an ISIS treated group and pooled placebo with the common standard deviation of 
13 mmHg, there are at least 86 eligible patients (n = 43 for an ISIS 757456 treated group and 
n = 43 for pooled placebo) are needed to provide approximate 80% of power to demonstrate the 
mean difference between an ISIS treated group and pooled placebo by ANCOVA with the 
two-sided significant level of 0.05.  Additional patients are included while considering an 
estimated approximate 10% drop-out rate.   
The overall type I error rate is controlled at 0.1 level, two-sided. 
3.4. 
Secondary Efficacy Analyses 
3.4.1. 
Secondary Efficacy Endpoints 
The secondary efficacy analyses will be performed using statistical methods described in 
Section 3.2.1.  ANCOVA model will be used for continuous endpoints, logistic regression model 
will be used for dichotomous endpoints.  Secondary endpoints include: 
1. Comparison of percent change from baseline to Study Day 85 in AGT between an 
ISIS 757456-treated group and pooled-placebo in the PPS 
2. Comparison of change from baseline to Study Day 85 in SBP from 24-hrs mean ABPM 
between an ISIS 757456-treated group and pooled-placebo in the PPS  
3. Comparison of proportion of patients reaching the goals of seated automated office SBP 
≤ 140 mmHg and DBP ≤ 90 mmHg at Study Day 85 between an ISIS 757456-treated 
group and pooled-placebo in the PPS, excluding patients with baseline SBP ≤ 140 mmHg 


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4. Comparison of absolute value, change and percent change from Baseline to each 
scheduled post-Baseline visit in plasma AGT between each ISIS 757456-treated group 
and pooled-placebo group in PPS and FAS  
5. Comparison of change from Baseline to Study Day 85 in SBP and DBP from 24 hr mean 
ABPM between each ISIS 757456-treated group and pooled-placebo group in the PPS 
and FAS 
6. Comparison of patient incidence in reaching the goals of seated automated office 
SBP ≤ 140 mmHg, DBP ≤ 90 mmHg, and both for each scheduled, post-Baseline visit 
between each ISIS 757456-treated group and pooled-placebo group in PPS and FAS, 
excluding patients with Baseline SBP ≤ 140 mmHg 
7. Comparison of patient incidence in reaching the goals of seated automated office 
SBP ≤ 130 mmHg, DBP ≤ 80 mmHg, and both for each scheduled, post-Baseline visit 
between each ISIS 757456-treated group and pooled-placebo group in PPS and FAS 
8. Comparison of change from Baseline to each scheduled post-Baseline visit in seated 
automated office SBP between each ISIS 757456-treated group and pooled-placebo 
group in PPS and FAS 
9. Comparison of change from Baseline to each scheduled post-Baseline visit in seated 
automated office DBP between each ISIS 757456-treated group and pooled-placebo 
group in PPS and FAS 
Figures contain the summary (mean with standard error of mean) by scheduled visit overtime 
based on PPS will be provided.  Spaghetti plots demonstrating each individual’s efficacy 
assessment over time by treatment group will be provided.  
3.4.2. 
Multiplicity 
The multiplicity of primary and the first three secondary endpoints will be controlled by using 
sequential (closed) testing strategy in the following testing sequence for interim analysis (if 
applicable) and final analysis, which is also illustrated in Figure 1 below. 
• Comparison of change from baseline to Study Day 85 in in seated automated office 
SBP between an ISIS 757456-treated group and pooled-placebo in the PPS (primary) 
• Comparison of percent change from baseline to Study Day 85 in AGT between an 
ISIS 757456-treated group and pooled-placebo in the PPS (secondary 1) 
• Comparison of change from baseline to Study Day 85 in SBP from 24-hrs mean 
ABPM between an ISIS 757456-treated group and pooled-placebo in the PPS 
(secondary 2)  
• Comparison of portion of patients reaching the goals of seated automated office SBP 
≤ 140 mmHg and DBP ≤ 90 mmHg at Study Day 85 between an ISIS 757456-treated 
group and pooled-placebo in the PPS, excluding patients with baseline SBP ≤ 140 
mmHg (secondary 3) 
 




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• Proportion of non-worsening from baseline to each scheduled post-Baseline PGI-C 
and PGI-S will be compared between each ISIS 757456-treated group and 
pooled-placebo group.  PGI-C will be categorized by non-worsening and worsening, 
where non-worsening includes categories of “Much better”, “A little better” and 
“No change”.  PGI-S will be categorized by non-worsening and worsening, where 
non-worsening defined as ≥ 0 point improvement in the change from baseline score.  
The adjusted odds ratio (each ISIS 757456-treated dose group vs. pooled-placebo 
group) with 95% CI from logistic regression model will be provided. 
• Comparison of change from Baseline to Study Day 85 in daytime SBP and DBP from 
24 hr ABPM will be conducted between each ISIS 757456-treated group and 
pooled-placebo group  
• Comparison of change from Baseline to Study Day 85 in nighttime SBP and DBP 
from 24 hr ABPM will be conducted between each ISIS 757456-treated group and 
pooled-placebo group  
• Comparison of change from Baseline to Day 85 in eGFR will be conducted between 
each ISIS 757456-treated group and pooled-placebo group 
• Comparison of change from Baseline to Day 85 in UACR will be conducted between 
each ISIS 757456-treated group and pooled-placebo group 
• Primary endpoint, selective secondary and exploratory endpoints will be also 
analyzed using a sub-population of patients with Baseline 24 hr ABPM SBP > 130  
• POP-HSQ scores will be listed. 
Additional POP-HSQ summaries and analysis will be conducted, details are described in 
POP-HSQ SAP.  Other exploratory PK endpoints will be further described in the Section 3.6.   
3.6. 
Pharmacokinetic (PK) and Immunogenicity (IM) Analysis 
3.6.1. 
PK Analysis 
The plasma pharmacokinetics of ISIS 757456 (as total full-length oligonucleotides or 
ISIS 757456-equivalent, ISIS 757456-eq.) will be assessed following SC administration(s).  
PK analysis summary will be based on PK set.   
Metabolite identification and profiling may be determined in some of the collected plasma 
samples and will be reported separately.   
Plasma Concentration Data of Total Full-Length Oligonucleotides 
Plasma concentrations of ISIS 757456 (ISIS 757456eq.), along with the scheduled (nominal) and 
actual sample times (i.e., time from SC dosing) will be listed (when applicable) for each patient, 
by treatment group, actual dose, subject IM status and study day.  In addition, percent differences 
between scheduled and actual sampling times will also be listed for all patients, as well as 
percent differences between actual administered dose and nominal dose.  
ISIS 757456 concentrations will be summarized using descriptive statistics by treatment, dose, 
cohort, study day, and scheduled time point, without and with stratification by IM status.  Plasma 


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concentrations below the lower limit of quantification (LLOQ) will be indicated by “BLQ”.  For 
the purpose of calculating typical descriptive statistics (n, mean, SD, SE, %CV, geometric mean, 
geometric %CV, 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 %CV will be reported as not applicable.  Other stratifications may also be performed 
if deemed warranted to properly interpret the pharmacokinetic analysis.  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 times, or large deviations 
between actual dose and nominal dose (> 30%). 
Plasma Pharmacokinetic Parameters 
The plasma PK of ISIS 757456 (as total full-length oligonucleotides) will be assessed following 
the SC administration.  Non-compartmental PK analysis of ISIS 757456 (total full-length 
oligonucleotides) will be carried out on each individual subject data set using Phoenix 
WinNonlin version 8.0 or higher (Pharsight Corporation, Mountain View, CA).  For calculation 
of PK parameters, all BLQ values will be set to zero.  The plasma PK parameters for 
ISIS 757456eq. will be calculated based on actual sampling times.  Since only trough and post-
treatment follow up PK samples (and optional samples 2 hours post-dose on Day 1 and Day 78) 
are to be collected, only the following PK parameters will be calculated: 
• Ctrough  
• t1/2λz 
The terminal half-life shall be determined over a time interval equal to at least 1.5 × t1/2λz 
(Span > 1.5).  In addition, a minimum of three data points in the elimination phase will be used 
to define λz, and the coefficient of determination (r2_adjusted) shall be greater than or equal to 
0.8 for the estimate to be accepted.  Other plasma PK parameters, as appropriate, may be 
determined or calculated at the discretion of the PK scientist. 
Plasma PK parameters will be listed by treatment, dose, subject ID, subject IM status, and 
study day; and appropriately summarized using descriptive statistics (n, mean, SD, SE, %CV, 
geometric mean, geometric %CV, median, minimum, and maximum) by treatment and 
study day.  Additionally, subject IM status stratified plasma PK parameters will be similarly 
summarized. Other stratifications may also be performed if deemed warranted at the discretion of 
the pharmacokineticist and/or biostatistician.  
3.6.2. 
Pharmacokinetic/Pharmacodynamic Exposure-Response Analysis 
Exposure-response correlations may be explored graphically between plasma exposure 
(e.g., Ctrough,), and selected PD measures (e.g., plasma AGT level), other relevant biomarkers 
(such as plasma renin activity, angiotensin II, etc.) and relevant clinical endpoints (such as 
systolic blood pressure).  In addition, the relationship between serum AGT level with plasma 
concentrations (Ctrough) of ISIS 757456 eq. may be further evaluated with an inhibitory effect 
Emax model.  The analysis may include stratification by IM status as well. 
Population PK and PKPD analysis may be performed using the PK and PD data from this Study, 
and/or combined with other ISIS 757456 clinical PK/PD data from any previous and future 
studies in the development timeline. 


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3.6.3. 
IM Analysis 
Samples collected at pre-dose on Study Days 1 (baseline), 15, 29, 57 and anytime on Days 85 
and 169 in the post-treatment period will be analyzed for anti-ISIS 757456 antibodies.  
Sample Level ADA data 
An evaluable sample will be designated ‘IM positive’ based on both positive screening and 
confirmation assay results (i.e., confirmed positive result), and otherwise will be deemed 
‘IM negative’. Sample IM results (screen positive/negative, confirmed positive/negative or 
unevaluable, and when applicable, titer of anti-ISIS 757456 antibodies) before, during, and after 
treatment with study drug (sample IM status) will be listed by treatment, dose, and day of 
collection.   
The sample ADA incidence (number) and incidence rate (percent) at each evaluated study time 
point will be determined and appropriately summarized by treatment and dose as the total 
number of and percentage of evaluated subjects with sample ADA negative, positive, and 
unknown status.  Furthermore, titer over time will be also appropriately summarized (using 
descriptive statistics) as median, quartiles (25% and 75%), and range by treatment and dose. 
Subject Level ADA Data 
Subject’s overall ADA status will be defined as ‘Positive’ if they have at least one confirmed 
positive sample result at any time during the treatment or post-treatment evaluation periods; 
‘Negative’ if all evaluated ADA sample results during the treatment and post-treatment 
evaluation periods are ADA negative and they have at least one evaluable ADA result collected 
post study drug treatment.  Otherwise, a study subject will be assigned ‘Unknown’ ADA status.   
Furthermore, subjects with overall positive ADA status will be further classified into different 
ADA types based on their baseline ADA status and change in ADA titer post treatment as 
described below (Shankar et al. 2014): 
• Treatment-Emergent ADA:  sum of treatment-induced ADA and treatment-boosted 
ADA as described below: 
o Treatment-Induced ADA:  ADA developed de novo (seroconversion) following 
biologic drug administration (i.e., formation of ADA any time after the initial 
drug administration in a subject without pre-existing ADA, i.e., baseline negative 
ADA) 
o Treatment-Boosted ADA:  pre-existing ADA that were boosted to a higher level 
following biologic drug administration (i.e., any time after the initial drug 
administration the ADA titer is greater than the baseline titer by a factor of 8-fold 
or more) 
• Treatment-Unaffected ADA:  pre-existing ADA that were not affected (boosted) 
following biologic drug administration (i.e., any time after the initial drug 
administration the ADA titer is 4-fold or less) 
• ADA type would be not applicable (NA) if the subject’s overall ADA status is 
negative. 


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Other subject level IM parameters to be calculated/defined may include but not limited to: 
• Subject ADA Status at Baseline: "Positive" if the subject has Week 1 Day 1 pre-dose 
sample (baseline) tested as confirmed positive; "Negative" if the subject has Week 1 
Day 1 pre-dose sample (baseline) tested as confirmed negative; "Unknown" if the 
subject has Week 1 Day 1 pre-dose sample (baseline) unevaluable. 
• Onset of ADA: i.e., the first day ADA positive sample observed, will be calculated 
by: the date of first sample has “positive” sample IM status - first dose date +1 
• Last Positive ADA Study Day: defined as the last positive ADA sample observed 
from the start of study drug treatment and will be calculated by: the date of last 
sample has “positive” sample IM status - first dose date +1 
• Last IM Sampling Study Day: defined as the last ADA sample collected from the start 
of study drug treatment and will be calculated by:  the date of last sample collected - 
first dose date +1 
• Peak titer: the highest titer observed for the subject  
• Time to peak titer: the time to reach peak titer will be calculated by:  the date of first 
peak titer observed- first dose date +1 
• Total number of ADA Positive Samples:  the total number of ADA samples being 
confirmed positive for the subject  
• Total number of ADA Samples evaluated:  the total number of ADA samples being 
collected and analyzed successfully with reportable results for the subject 
Lastly, subjects with positive ADA status may further be classified as being transient or 
persistent ADA response, if there are sufficient number of subjects with transient ADA status.  
Transient and persistent ADA definitions are defined below and based on (Shankar et al. 2014): 
Transient ADA response:  
• Treatment-induced ADA detected only at one sampling time point during the 
treatment or follow-up observation period (excluding the last sampling time point, 
which will be considered persistent unless shown to be undetectable at a later time) or 
• Treatment-induced ADA detected at two or more sampling time points during the 
treatment (including follow-up period if any), where the ﬁrst and last ADA-positive 
samples (irrespective of any negative samples in between) are separated by a period 
less than 16 weeks, and the subject’s last sampling time point is ADA-negative. 
Persistent ADA response: 
• Treatment-induced ADA detected at two or more sampling time points during the 
treatment (including follow-up period if any), where the ﬁrst and last ADA-positive 
samples (irrespective of any negative samples in between) are separated by a period 
of 16 weeks or longer or 


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• Treatment-induced ADA detected only at the last sampling time point of the study 
treatment period or at a sampling time point with less than 16 weeks before an 
ADA-negative last sample. 
The subject level ADA prevalence, incidence, and positive ADA response being transient or 
persistent (if applicable) will be calculated as the number and the proportion (percent) of the 
study population during the study period by treatment and dose.  Subject level IM parameters (as 
described above) will be listed by treatment and dose for all evaluable subjects, and also 
appropriately summarized (using descriptive statistics) as median, quartiles (25% and 75%) and 
range, by treatment and dose. 
Evaluation of IM Impact on PK, PD, Efficacy and Safety 
The impact of IM on PK and PD will be evaluated by stratifying plasma PK parameters, plasma 
trough and post-treatment ISIS 757456 concentrations and PD biomarker levels by subject ADA 
status, summarized using typical descriptive statistics, and presented graphically and/or in tables.  
The impact of IM may also be evaluated on selected clinical efficacy end points and safety 
measures.  
Lastly, other stratifications (e.g., based on antibody titer, onset of ADA, etc.) of selected PK, 
efficacy and safety assessments may also be performed if deemed warranted at the discretion of 
the pharmacokineticist, medical monitor, and/or biostatistician. 
3.7. 
Safety Analyses 
The safety analysis will be conducted on the Safety Set. 
3.7.1. 
Exposure 
Total amount of study drug (Placebo, ISIS 757456 80 mg or ISIS 757456 120 mg) received 
(mg), total number of doses administered, and treatment duration will be summarized by the 
treatment group.    
The treatment duration will be defined as the total number of days a subject is known to be 
followed on study calculated as follows:  
Treatment duration = Last dose date  – Date of first dose + 1 
3.7.2. 
Adverse Events 
The number and incidence of adverse events (AEs) will be summarized by Medical Dictionary 
for Regulatory Activities (MedDRA) version 24.0 preferred term (PT) and system organ class 
(SOC) for: 
Any treatment emergent adverse events (TEAEs). 
Serious TEAEs (Treatment-emergent SAEs). 
Related TEAEs and SAEs.  Related is defined as “Related”, “Possible”, or missing relationship 
to study drug.  


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Any TEAEs by severity.  At each severity level of subject summarization, a subject is classified 
according to the highest severity if the subject reported one or more events.  AEs with missing 
severity will be categorized as “Missing” for this summary. 
TEAEs leading to permanent study drug discontinuation. 
Serious and non-serious AEs that lead to study discontinuation or investigational drug 
discontinuation will be listed separately.  Non-treatment emergent adverse event will be included 
and be noted in the subject AE listing. 
TEAEs will be defined as those adverse events that either start or worsen in severity on or after 
the date/time of first dose of study treatment.  
In addition, if severity of an AE changes during the study, a separate AE will be recorded for 
each severity on the AE CRF. “first” and “second” AE will be identify based on AE start date. 
AE start date of the second record is AE stop date of first record.  These linked events will be 
identified based on “Formlink” dataset and compared pairwise, and consider two cases, where 
the AE severity (mild/moderate/severe) are compared between the two records in the pair.    
Case 1:  The first AE record in the pair occurs before first dosing, and the second AE record 
occurs after first dosing.   
If the AE severity or seriousness of the second record is worse than that of the first 
record, then only the second AE is deemed as a TEAE.  Otherwise, neither record is 
considered as TEAE. 
Case 2:  Both AE records in the pair occur after first dosing.   
Only the worst AE will be deemed as one TEAE. 
All TEAEs identified based on the rules above will be summarized in the event number analysis.    
The most conservative approach will be used to determine if the event occurs after the treatment.  
For example, if the onset date or resolution date of an AE is prior to the first study treatment 
date, it 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 complete 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.   
Addition summary of hypotension and hyperkalemia incidence will be provided.     
Local Cutaneous Reactions at the Injection Site 
Local cutaneous reaction at injection site (LCRIS) is defined as (A) moderate or severe adverse 
events with the PTs Injection site erythema, Injection site swelling, Injection site pruritus, or 
Injection site pain that started on the day of injection, persisted for at least two days or ongoing; 
or (B) any AE at the Study Drug injection site, regardless of severity, that leads to 
discontinuation of study drug, where AE at the Study Drug injection site is the principal reason 
for discontinuation. 


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Percentage of injections leading to LCRIS will be calculated as follows for each subject: 
(A/B)*100, where A = number of injections with an LCRIS, and B = total number of injections. 
Doses that are split across multiple injections are counted as a single injection 
LCRIS will be summarized using the MedDRA coding system, by SOC/PT.  Percentage of the 
injections leading to LCRIS at injection site will also be summarized.  
LCRIS will be listed by preferred term. 
Flu-like Reactions 
Flu-like reactions are defined as adverse events with PTs including either (A) Influenza like 
illness or (B) Pyrexia or Feeling hot or Body temperature increased, plus at least two of the 
following symptoms with the PTs:  Chills, Myalgia, or Arthralgia, starting on day of injection or 
the next day.  
Percentage of injections leading to flu-like reactions will be summarized using the descriptive 
statistics. 
Percentage of injections leading to flu-like reactions will be calculated as follows for each 
subject:  (A/B)*100, where A = number of injections leading to flu-like reactions, and B = total 
number of injections. 
FLRs will be summarized using the MedDRA coding system by SOC/PT.  Percentage of the 
injections leading to FLRs at injection site will also be summarized.  
FLRs will be listed by preferred term. 
AE of Special Interest (AESI):  Platelet reduction 
Per protocol, severe reductions in platelet count < 50,000/mm3 accompanied by a clinically 
relevant bleeding event or platelet count of < 25,000/mm3 independent of a clinically relevant 
bleeding event are considered as AESI.  The AEs meeting the AESI criteria will be captured in 
the AE CRF page with a check box to indicate.  
AESI will be summarized using the MedDRA coding system, by SOC/PT.  A listing of AESI 
will also be generated.  
3.7.3. 
Laboratory Measurements 
Chemistry including thyroid panel and inflammatory, hematology, coagulation, and urinalysis 
(result, change and percent change from baseline) will be summarized by each treatment group 
and each post-baseline visit.  Listing of laboratory assessments in Chemistry, Hematology and 
urinalysis will be provided.  Local laboratory data will also be provided in the listings separately. 
For alanine aminotransferase (ALT) and aspartate aminotransferase (AST), the number and 
percent of subjects with post-Baseline results falling in each of the following categories (using 
available central and local laboratory assessments) will be tabulated:  
ALT or AST > 3 × ULN, confirmed 
ALT or AST > 5 × ULN, confirmed 
ALT or AST > 3 × ULN and serum total bilirubin > 2 × ULN, confirmed or INR > 1.5 




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3.7.5. 
12-Lead Electrocardiograms (ECG) 
Safety 12 lead ECG will be performed at the visits indicated in the protocol Schedule of 
Procedures.   
The ECG data will include ventricular rate (VR), PR interval, QRS duration, QT, and corrected 
QT intervals (QT corrected using the Fridericia’s formula), and overall interpretation.  
For the continuous variables above, descriptive statistics (n, mean, standard error, standard 
deviation, median, 25th percentile, 75th percentile, minimum and maximum) of results at study 
screening and study Week 13 visit, as well as the change and percent change from Baseline, will 
be presented in summary tables; for the categorical responses to overall interpretation.  
All the ECG data collected will be listed.  
3.7.6. 
Prior and Concomitant Medications 
Prior and Concomitant medications will be coded using WHO Drug dictionary (Global B3 
March 2021) and summarized by ATC class, generic name and treatment group.  
Prior medications include medications that started prior to the first dose of study medication 
regardless of whether they continued while on treatment or not.  Concomitant medications 
include medications that patients are exposed to on or after the first dose of study medication.  
Additional summary of HTN medications uses at baseline and new initiation of HTN medication 
during the study will be summarized by ATC class and treatment group.  
Listings will be provided. 
4. 
REFERENCES 
Levey, A. S., L. A. Stevens, C. H. Schmid, Y. L. Zhang, 3rd Castro F. A., H. I. Feldman, J. W. 
Kusek, et al. 2009. “A New Equation to Estimate Glomerular Filtration Rate.” Ann Intern Med 
150: 604–12. https://doi.org/10.7326/0003-4819-150-9-200905050-00006. 
Shankar, G., S. Arkin, L. Cocea, V. Devanarayan, S. Kirshner, A. Kromminga, V. Quarmby, et 
al. 2014. “Assessment and Reporting of the Clinical Immunogenicity of Therapeutic Proteins and 
Peptides-Harmonized Terminology and Tactical Recommendations.” Aaps j 16: 658–73. 
https://doi.org/10.1208/s12248-014-9599-2. 
  
 
 


Ionis Pharmaceuticals 
CONFIDENTIAL 
15 August 2022
Statistical Analysis Plan v2 
ISIS 757456-CS4
 
31 
APPENDIX 1. EQ-5D HEALTH STATE INDEX SCORE DERIVATION 
ALGORITHM 
To derive this index value, the response of each dimension will be first translated into a weight 
based on the value sets issued by EuroQoL group (see table x).  The index value then can be 
calculated as 1- sum of weights from these 5 dimensions’ response.  The possible best value is 
1 for both regions and the possible worst value is -0.285 for UK region and -0.573 for US region.  
If there is a missing response from a dimension, then the index value will not be calculated and 
be treated as missing.  For this study, all the other non-US counties will utilize UK county 
weight instead.  The display of the index value is with 3 digits of decimal points.   
Table 1: 
Weight of EQ-5D-5L Response by Region 
  
  
  
  
EQ-5D-5L Dimension 
  
Region 
Response 
Mobility 
Self-
Care 
Usual 
Activities 
Pain/ 
Discomfort 
Anxiety/ 
Depression 
UK 
1) No problem 
0 
0 
0 
0 
0 
(16/11/2020) 
2) Slight problem 
0.058 
0.05 
0.05 
0.063 
0.078 
  
3) Moderate problems 
0.076 
0.08 
0.063 
0.084 
0.104 
  
4) Severe problems 
0.207 
0.164 
0.162 
0.276 
0.285 
  
5) Unable to  
0.274 
0.203 
0.184 
0.335 
0.289 
US 
1) No problem 
0 
0 
0 
0 
0 
(16/11/2020) 
2) Slight problem 
0.096 
0.089 
0.068 
0.06 
0.057 
  
3) Moderate problems 
0.122 
0.107 
0.101 
0.098 
0.123 
  
4) Severe problems 
0.237 
0.22 
0.255 
0.318 
0.299 
  
5) Unable to  
0.322 
0.261 
0.255 
0.414 
0.321 
Sources:  https://euroqol.org/support/tools/analysis-tools/index-value-set-calculators/ 
 


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