Phase I Placebo-Controlled Study of the Infectivity, Safety and Immunogenicity of a Single Dose of a Recombinant Live-Attenuated Respiratory Syncytial Virus Vaccine, RSV 6120/∆NS2/1030s, Lot RSV#012A, Delivered as Nose Drops to RSV-Seropositive Children 12 to 59 Months of Age and RSV-Seronegative Infants and Children 6 to 24 Months of Age
Phase: PHASE1
Conditions: RSV Infection
Sponsor: National Institute of Allergy and Infectious Diseases (NIAID)

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RSV 6120/∆NS2/1030s, Lot#012A  
           
Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
Phase I Placebo-Controlled Study of the Infectivity, Safety and Immunogenicity of a 
Single Dose of a Recombinant Live-attenuated Respiratory Syncytial Virus Vaccine, 
RSV 6120/∆NS2/1030s, Lot RSV#012A, Delivered as Nose Drops to  
RSV-Seropositive Children 12 to 59 Months of Age and  
RSV-Seronegative Infants and Children 6 to 24 Months of Age 
 
Sponsored by:  
National Institute of Allergy and Infectious Diseases (NIAID) 
 
Center for Immunization Research (CIR) Protocol Number: CIR 322 
Western IRB Protocol Number: WIRB # 20172138 
FDA Investigational New Drug (IND) Number: 17681 
Project Assurance: FWA #00000287 
 
IND Sponsor:  
Office of Clinical Research Policy and Regulatory Operations (OCRPRO) 
Division of Clinical Research (DCR) 
National Institute of Allergy and Infectious Diseases (NIAID) 
National Institutes of Health (NIH) 
5601 Fishers Lane 
Room 4B11, MSC 9820 
Bethesda, MD 20892 
 
 
Respiratory Syncytial Virus (RSV) 6120/∆NS2/1030s 
Provided by: 
Laboratory of Infectious Diseases (LID), NIAID, NIH 
 
Medical Monitor:  
Shirley Jankelevich, MD 
 
Principal Investigator: 
Ruth Karron, MD 
 
Version 3.0 
April 6, 2020 
 
 
 
 
 
 
 
The information contained herein is the property of the NIAID and may not be reproduced, published 
or disclosed to others without the written authorization of the NIAID. 

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RSV 6120/∆NS2/1030s, Lot#012A  
           
Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
STUDY STAFF ROSTER 
 
PRINCIPAL INVESTIGATOR 
Ruth A. Karron, MD 
CIR 
Johns Hopkins Bloomberg School of Public Health (JHSPH) 
624 N. Broadway, Room 217 
Baltimore, MD 21205 
Phone: 410-614-0319 
Administrative Coordinator: 410-955-1624  
Email: rkarron@jhu.edu 
 
CLINICAL INVESTIGATORS  
Kawsar Talaat, MD 
CIR, JHSPH 
624 N. Broadway, Room 249 
Baltimore, MD 21205 
Phone: 410-502-9627 
Email: ktalaat1@jhu.edu 
 
 
Kristi Herbert, CRNP-P 
 
CIR, JHSPH 
624 N. Broadway, Room 205 
Baltimore, MD 21205 
Phone: 410-955-4362  
Email: kherber1@jhu.edu 
 
Elizabeth Schappell, RN, MSN, CCRP 
CIR, JHSPH 
624 N. Broadway, Room 205 
Baltimore, MD 21205 
Phone: 410-614-9114  
Email: eschappe@jhu.edu 
 
Jocelyn San Mateo, CRNP-F, CCRP 
CIR, JHSPH 
624 N. Broadway, Room 201 
Baltimore, MD 21205 
Phone: 410-614-4306 
Email: jsanmat1@jhu.edu  
Beulah Sabundayo, PharmD, MPH 
CIR, JHSPH 
624 N. Broadway, Room 200 
Baltimore, MD 21205 
Phone: 410-502-7451 
Email: bsabund1@jhu.edu 
 
Suzanne Woods, CRNP-P, CCRP 
CIR, JHSPH 
624 N. Broadway, Room 212 
Baltimore, MD 21205 
Phone: 410-614-1880  
Cell: 443-813-0697 
Email: swoods12@jhu.edu 
 
SCIENTIFIC INVESTIGATORS  
 
Ursula Buchholz, PhD 
LID, NIAID, NIH 
Building 50, Room 6503 
50 South Drive, MSC 8007 
Bethesda, MD 20892 
Phone: 301-594-1533 
Email: ubuchholz@niaid.nih.gov 
 
 
Peter Collins, PhD  
LID, NIAID, NIH  
Building 50, Room 6517 
50 South Drive, MSC 8007 
Bethesda, MD 20892 
Phone: 301-594-1590 
Email: pcollins@niaid.nih.gov 
 

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RSV 6120/∆NS2/1030s, Lot#012A  
           
Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
CLINICAL TRIAL SITES 
Center for Immunization Research 
Johns Hopkins Bloomberg School of Public 
Health  
624 N. Broadway, Room 117 
Baltimore, MD 21205 
Phone: 410-502-3333 
 
CLINICAL LABORATORIES 
 
Johns Hopkins Hospital 
Laboratories of Pathology 
600 N. Wolfe Street 
Baltimore, MD 21287 
Phone: 410-955-3142 
 
Center for Immunization Research East  
Johns Hopkins Bayview Medical Center  
Campus 
301 Mason Lord Drive 
Baltimore, MD 21224 
Phone: 410-550-2725 
 
Center for Immunization Research South 
9811 Mallard Drive, Suite 210 
Laurel, MD 20708 
 
Quest Diagnostics  
1901 Sulphur Spring Road 
Baltimore, MD 21227 
Phone: 410-247-0001 
VIROLOGY/IMMUNOLOGY LABORATORIES 
CIR Laboratories 
615 N. Wolfe Street, Room E5402 
Baltimore, MD 21205 
Phone: 410-955-7230 
INSTITUTIONAL REVIEW BOARD 
 
Western Institutional Review Board® (WIRB) 
1019 39th Avenue SE Suite 120 
Puyallup, WA 98374-2115 
Phone: 1-800-562-4789 or 360-252-2500 
 
INSTITUTIONAL BIOSAFETY OFFICE (IBC) 
2024 E. Monument Street, Room B-200 
Baltimore, MD 21287 
 
 
 
 
SPONSOR REPRESENTATIVE 
John Tierney, RN, MPM 
OCRPRO, DCR, NIAID, NIH 
5601 Fishers Lane 
Room 4B11, MSC 9820 
Bethesda, MD 20892 
Phone: 301-451-5136  
Email: jtierney@niaid.nih.gov 
 
SPONSOR MEDICAL MONITOR 
Shirley Jankelevich, MD 
Medical Monitor 
Clinical Monitoring Research Program 
Leidos Biomedical Research, Inc. 
5705 Industry Lane, Room 248  
Frederick, Maryland  21704 
Phone 301-846-7322 
Email: shirley.jankelevich@nih.gov 
 
 
 

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RSV 6120/∆NS2/1030s, Lot#012A  
           
Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
TABLE OF CONTENTS 
 
STUDY STAFF ROSTER ............................................................................................................................................. 2 
TABLE OF CONTENTS .............................................................................................................................................. 4 
LIST OF TABLES......................................................................................................................................................... 8 
LIST OF FIGURES ....................................................................................................................................................... 9 
LIST OF ABBREVIATIONS ...................................................................................................................................... 10 
PROTOCOL SUMMARY........................................................................................................................................... 13 
 
BACKGROUND AND SCIENTIFIC RATIONALE ....................................................................................... 15 
 
BACKGROUND............................................................................................................................................. 15 
1.1.1 
Epidemiology, Disease Burden, and the Need for a Vaccine ............................................................... 15 
 
PRIOR RESEARCH ........................................................................................................................................ 16 
1.2.1 
Experimental Vaccines against RSV .................................................................................................... 16 
1.2.1.1 
RSV Vaccine Candidates with an NS2 Deletion ........................................................................................... 17 
1.2.2 
Preclinical Studies ................................................................................................................................ 18 
1.2.2.1 
Temperature sensitivity of RSV 6120/ΔNS2/1030s ...................................................................................... 18 
1.2.2.2 
Replication of an experimental lot of RSV 6120/ΔNS2/1030s in an in-vitro model of mucociliary normal 
human tracheal/bronchial epithelial cells ......................................................................................................................... 18 
1.2.2.3 
Replication of experimental lot and CTM of RSV 6120/ΔNS2/1030s in African green monkeys (AGMs) . 20 
1.2.3 
Previous Clinical Experience ............................................................................................................... 20 
 
RATIONALE ................................................................................................................................................. 21 
 
CLINICAL DEVELOPMENT PLAN .................................................................................................................. 21 
 
HYPOTHESES ............................................................................................................................................... 21 
 
OBJECTIVES ................................................................................................................................................... 21 
2.1. 
PRIMARY OBJECTIVES................................................................................................................................. 21 
2.2. 
SECONDARY OBJECTIVES ............................................................................................................................ 22 
2.3. 
EXPLORATORY OBJECTIVE ......................................................................................................................... 22 
 
STUDY DESIGN ............................................................................................................................................. 22 
 
STUDY POPULATION ................................................................................................................................... 23 
 
INCLUSION CRITERIA FOR RSV-SEROPOSITIVE CHILDREN ......................................................................... 24 
 
EXCLUSION CRITERIA FOR RSV-SEROPOSITIVE CHILDREN ........................................................................ 24 
 
INCLUSION CRITERIA FOR RSV-SERONEGATIVE INFANTS & CHILDREN ..................................................... 26 
 
EXCLUSION CRITERIA FOR RSV-SERONEGATIVE INFANTS & CHILDREN .................................................... 26 
 
INCLUSION CRITERIA FOR RSV-SERONEGATIVE CHILDREN PARTICIPATING IN A SECOND SEASON OF RSV 
SURVEILLANCE ..................................................................................................................................... 28 
 
EXCLUSION CRITERIA FOR RSV-SERONEGATIVE CHILDREN PARTICIPATING IN A SECOND SEASON OF RSV 
SURVEILLANCE ..................................................................................................................................... 28 
 
CO-ENROLLMENT CONSIDERATIONS .......................................................................................................... 28 
 
RE-ENROLLMENT CONSIDERATIONS ........................................................................................................... 28 
 
RECRUITMENT PROCESS ............................................................................................................................. 28 
 
PARTICIPANT RETENTION ........................................................................................................................... 29 
 
PARTICIPANT WITHDRAWAL OR TERMINATION FROM THE STUDY .............................................................. 29 
 
STUDY PRODUCT ......................................................................................................................................... 29 
 
STUDY PRODUCT REGIMENS ....................................................................................................................... 30 
 
STUDY PRODUCT FORMULATION ................................................................................................................ 30 
5.2.1 
RSV 6120/∆NS2/1030s ........................................................................................................................ 30 

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Principal Investigator: Ruth A. Karron, MD 
        
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5.2.2 
Diluent for RSV 6120/∆NS2/1030s ..................................................................................................... 30 
5.2.3 
Placebo for RSV 6120/∆NS2/1030s ..................................................................................................... 30 
 
STUDY PRODUCT STORAGE......................................................................................................................... 30 
 
STUDY PRODUCT PREPARATION ................................................................................................................. 31 
5.4.1 
Diluent .................................................................................................................................................. 31 
5.4.2 
Placebo ................................................................................................................................................. 31 
5.4.3 
Live RSV 6120/∆NS2/1030s ................................................................................................................ 31 
 
STUDY PRODUCT INOCULATION PROCEDURE ............................................................................................. 32 
 
STUDY PRODUCT ACQUISITION ................................................................................................................... 32 
 
STUDY PRODUCT ACCOUNTABILITY ........................................................................................................... 32 
 
DISPOSITION OF USED/UNUSED STUDY PRODUCT ...................................................................................... 33 
 
FINAL DISPOSITION OF STUDY PRODUCTS .................................................................................................. 33 
 
CONCOMITANT MEDICATIONS .................................................................................................................... 33 
 
STUDY VISITS AND PROCEDURES ........................................................................................................... 33 
 
CONSENTING PROCESS ................................................................................................................................ 35 
 
SCREENING VISIT ........................................................................................................................................ 35 
6.2.1 
Participants Enrolled in Surveillance for a Second RSV Season ......................................................... 36 
 
RANDOMIZATION ........................................................................................................................................ 36 
 
ENROLLMENT ............................................................................................................................................. 37 
6.4.1 
Enrollment - Inoculation (Day 0) ......................................................................................................... 37 
6.4.2 
Enrollment in the Second Season of RSV Surveillance (Group 2 Only).............................................. 39 
 
STUDY PHASES ........................................................................................................................................... 39 
6.5.1 
RSV-Seropositive Children (Group 1) ................................................................................................. 39 
6.5.1.1 
Acute Phase: In-person Study Visit Days ..................................................................................................... 39 
6.5.1.2 
Acute Phase: Non-Visit Study Day Contacts ................................................................................................ 40 
6.5.1.3 
Study Day 11 ................................................................................................................................................ 40 
6.5.1.4 
Post-Acute Phase .......................................................................................................................................... 41 
6.5.1.5 
Study Day 28 Visit ........................................................................................................................................ 41 
6.5.2 
RSV-Seronegative Infants and Children .............................................................................................. 41 
6.5.2.1 
Acute Phase: In-person Study Visit Days ..................................................................................................... 41 
6.5.2.2 
Acute Phase: Non-Visit Study Day Contacts ................................................................................................ 42 
6.5.2.3 
Study Day 29 ................................................................................................................................................ 43 
6.5.2.4 
Post-Acute Phase .......................................................................................................................................... 43 
6.5.2.5 
Study Day 56 Visit ........................................................................................................................................ 43 
6.5.2.6 
Period after Day 56 through October 31st ..................................................................................................... 44 
 
RSV SURVEILLANCE: FIRST SEASON .......................................................................................................... 44 
6.6.1 
Pre-RSV Season Surveillance Study Visit ........................................................................................... 44 
6.6.2 
Weekly Contact for Surveillance during the RSV Season ................................................................... 45 
6.6.3 
Post-RSV Season Surveillance Study Visit .......................................................................................... 46 
 
RSV SURVEILLANCE: SECOND SEASON ...................................................................................................... 46 
 
ILLNESS VISIT ............................................................................................................................................. 46 
 
EARLY DISCONTINUATION STUDY VISIT .................................................................................................... 47 
 
LABORATORY PROCEDURES ....................................................................................................................... 48 
6.10.1 Specimen Collection............................................................................................................................. 48 
6.10.2 Virus Detection .................................................................................................................................... 48 
6.10.3 Immunologic Assays ............................................................................................................................ 49 
6.10.4 Specimen Preparation, Testing, Storage, and Shipping ........................................................................ 49 
6.10.5 Research Laboratories .......................................................................................................................... 49 
6.10.6 Plan for Use and Storage of Biological Samples .................................................................................. 50 
6.10.7 Biohazard Containment ........................................................................................................................ 50 
 
SAFETY ASSESSMENT, MONITORING, AND REPORTING ................................................................... 50 
 
SAFETY-RELATED ROLES AND RESPONSIBILITIES....................................................................................... 50 

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7.1.1 
Principal Investigator ........................................................................................................................... 50 
7.1.2 
Safety Review and Communications Plan (SRCP) and Transfer of Regulatory Obligations (TORO) 51 
7.1.3 
Sponsor Medical Monitor ..................................................................................................................... 51 
7.1.4 
Data Safety Monitoring Board ............................................................................................................. 51 
7.1.5 
Sponsor Reporting ................................................................................................................................ 51 
 
SAFETY-RELATED RECORDING ON CRFS ................................................................................................... 51 
 
SERIOUS ADVERSE EVENT REPORTING ....................................................................................................... 56 
 
REPORTING OF UNANTICIPATED PROBLEMS TO OCRPRO .......................................................................... 56 
 
PARTICIPANT MANAGEMENT ................................................................................................................... 57 
 
MANAGEMENT OF ADVERSE EVENTS.......................................................................................................... 57 
8.1.1 
Solicited Adverse Events ...................................................................................................................... 58 
8.1.2 
Serious Adverse Events ........................................................................................................................ 59 
8.1.3 
Unexpected Adverse Event .................................................................................................................. 59 
 
GRADING SEVERITY OF ADVERSE EVENTS ................................................................................................. 59 
8.2.1 
AE Grading .......................................................................................................................................... 59 
8.2.2 
Fever Grading ....................................................................................................................................... 60 
 
PAUSING AND STOPPING RULES .................................................................................................................. 60 
 
STATISTICAL CONSIDERATIONS .............................................................................................................. 61 
 
GENERAL DESIGN ISSUES ........................................................................................................................... 61 
9.1.1 
General Design ..................................................................................................................................... 61 
9.1.2 
Description of the Statistical Methods to be Employed ....................................................................... 61 
 
OUTCOME MEASURES ................................................................................................................................. 61 
9.2.1 
Primary Outcome Measures ................................................................................................................. 61 
9.2.2 
Secondary Outcome Measures ............................................................................................................. 62 
 
SAMPLE SIZE AND ACCRUAL ...................................................................................................................... 62 
9.3.1 
Sample Size and Randomization .......................................................................................................... 62 
 
MONITORING .............................................................................................................................................. 65 
9.4.1 
Site Monitoring Plan ............................................................................................................................ 65 
9.4.2 
Monitoring by the NIAID Intramural Data and Safety Monitoring Board ........................................... 65 
 
ANALYSES .................................................................................................................................................. 66 
9.5.1 
Assessment of Primary Objectives ....................................................................................................... 66 
9.5.2 
Assessment of Secondary Objectives ................................................................................................... 67 
 
DATA HANDLING AND RECORD KEEPING ............................................................................................. 67 
 
DATA MANAGEMENT RESPONSIBILITIES .................................................................................................... 67 
 
ESSENTIAL AND SOURCE DOCUMENTS AND ACCESS TO SOURCE DATA ..................................................... 68 
 
CLINICAL INVESTIGATOR’S BROCHURE ...................................................................................................... 68 
 
QUALITY CONTROL AND QUALITY ASSURANCE ......................................................................................... 68 
 
CLINICAL SITE MONITORING .................................................................................................................... 68 
 
HUMAN SUBJECTS PROTECTIONS ........................................................................................................... 69 
 
INSTITUTIONAL REVIEW BOARD/ETHICS COMMITTEE REVIEW AND APPROVAL ......................................... 69 
 
VULNERABLE PARTICIPANTS ...................................................................................................................... 69 
 
INFORMED CONSENT ................................................................................................................................... 69 
 
POTENTIAL BENEFITS ................................................................................................................................. 70 
 
POTENTIAL RISKS ....................................................................................................................................... 70 
12.5.1 Venipuncture ........................................................................................................................................ 70 
12.5.2 Nasal Wash ........................................................................................................................................... 70 
12.5.3 Nasosorption SAM Strips ..................................................................................................................... 70 
12.5.4 Topical Anesthetic Cream .................................................................................................................... 70 
12.5.5 Receipt of Study Product ...................................................................................................................... 70 

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Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
 
REIMBURSEMENT/COMPENSATION ............................................................................................................. 71 
12.6.1 Study Compensation: Acute Phase and First Season Surveillance ....................................................... 71 
12.6.2 Study Compensation: Second Season Surveillance .............................................................................. 71 
 
PRIVACY AND CONFIDENTIALITY................................................................................................................ 71 
 
MANAGEMENT OF INCIDENTAL FINDINGS ................................................................................................... 71 
 
ADMINISTRATIVE PROCEDURES.............................................................................................................. 71 
 
REGULATORY OVERSIGHT .......................................................................................................................... 71 
 
PROTOCOL REGISTRATION .......................................................................................................................... 72 
 
STUDY IMPLEMENTATION ........................................................................................................................... 72 
 
PROTOCOL DEVIATION REPORTING ............................................................................................................ 72 
 
CLINICALTRIALS.GOV ................................................................................................................................. 72 
 
PUBLICATIONS .............................................................................................................................................. 72 
 
REFERENCES ................................................................................................................................................. 73 
APPENDICES ............................................................................................................................................................. 77 
APPENDIX I: TABLES REFERENCED IN THE BACKGROUND SECTION ....................................................... 77 
APPENDIX II: SCHEDULE OF EVENTS: SCREENING, ACUTE PHASE, AND POST-ACUTE PHASE .......... 80 
APPENDIX III: SCHEDULE OF EVENTS: RSV SEASONAL SURVEILLANCE ................................................. 82 
APPENDIX IV: DEFINITIONS OF SOLICITED ADVERSE EVENTS .................................................................. 83 
APPENDIX V: RSV SEASONALITY IN BALTIMORE .......................................................................................... 84 
 
 
 
 

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LIST OF TABLES 
 
Table 1: Inoculation Schedule .......................................................................................................13 
Table 2: Temperature Sensitivity of Experimental Lots of RSV 6120/ΔNS2/1030s and RSV 
ΔNS2/Δ1313/I1314L ......................................................................................................18 
Table 3: Screening Visit Procedures ..............................................................................................36 
Table 4: Enrollment Visit Procedures ............................................................................................38 
Table 5: Acute Phase In-Person Visit Procedures – RSV-Seropositive Children .........................39 
Table 6: Acute Phase Non-Visit Contact Procedures – RSV-Seropositive Children ....................40 
Table 7: Day 11 Non-Visit Procedures – RSV-Seropositive Children ..........................................40 
Table 8: Day 28 Visit Procedures – RSV-Seropositive Children ..................................................41 
Table 9: Acute Phase In-Person Visit Procedures – RSV-Seronegative Children ........................42 
Table 10: Acute Phase Non-Visit Contact Procedures – RSV-Seronegative Children .................42 
Table 11: Day 29 Non-Visit Procedures – RSV-Seronegative Children .......................................43 
Table 12: Day 56 Visit Procedures – RSV-Seronegative Children ...............................................44 
Table 13: Pre-RSV Season Surveillance Study Visit Procedures ..................................................45 
Table 14: RSV Season Surveillance Procedures ...........................................................................45 
Table 15: Post-RSV Seasonal Surveillance Study Visit Procedures .............................................46 
Table 16: Illness Visit Timeframe .................................................................................................47 
Table 17: Illness Visit Procedures .................................................................................................47 
Table 18: Early Discontinuation Procedures .................................................................................48 
Table 19: AE CRF Recording Requirements for RSV-Seropositive Subjects ..............................53 
Table 20: AE CRF Recording Requirements for RSV-Seronegative Subjects .............................54 
Table 21: Grading for Adverse Events ..........................................................................................59 
Table 22: Grading for Fever ..........................................................................................................60 
Table 23: Pausing and Stopping Rules ..........................................................................................60 
Table 24: The Probability of Observing LRI Events in Seronegative Vaccine Recipients ...........63 
Table 25: Percent of Participants Experiencing LRI or AEs with Exact 90% CI..........................63 
Table 26: Magnitude of Difference in Responses Detectable with 80% Power ............................64 
Table 27: Viral Titers of Nasopharyngeal Swab Samples from AGMs Inoculated with the 
Experimental Lot or CTM of RSV 6120/ΔNS2/1030s, with RSV ΔNS2/Δ1313/I1314L, 
or with Recombinant wt RSV rA2 ..................................................................................77 
Table 28: Viral Titer of Tracheal Lavage Samples from AGMs Inoculated with the Experimental 
Lot or CTM of RSV 6120/ΔNS2/1030s, with RSV ΔNS2/Δ1313/I1314L, or with 
Recombinant wt RSV rA2 ..............................................................................................78 
Table 29: Serum PRNT60 Titers in AGMs Inoculated with the Experimental Lot or with CTM of 
RSV 6120/ΔNS2/1030s or RSV ΔNS2/Δ1313/I1314L, or Recombinant wt RSV rA2 .79 
Table 30: Schedule of Events: Group 1: RSV-Seropositive Participants ......................................80 
Table 31: Schedule of Events: Group 2: RSV-Seronegative Participants .....................................81 
Table 32: Schedule of Events: RSV Seasonal Surveillance ..........................................................82 
Table 33: Definitions of Solicited Adverse Events........................................................................83 
 
 
 

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RSV 6120/∆NS2/1030s, Lot#012A  
           
Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
LIST OF FIGURES 
 
Figure 1: Group 1 – Study Overview .............................................................................................14 
Figure 2: Group 2 – Study Overview .............................................................................................14 
Figure 3: Multicycle Replication of Experimental Lots in an in-vitro Model of Normal Human 
Bronchial/Tracheal Epithelial Cells ................................................................................19 
Figure 4: Investigational Product Label Sample ............................................................................30 
Figure 5: Power Curves for Assessment of Vaccine Virus Shedding in RSV seronegative 
Vaccinees ........................................................................................................................64 
Figure 6: RSV Seasonality in Baltimore ........................................................................................84 
 
 
 
 
 
 
 
 

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RSV 6120/∆NS2/1030s, Lot#012A  
           
Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
 
LIST OF ABBREVIATIONS 
 
ACIP 
AE 
Advisory Committee on Immunization Practices (CDC) 
adverse event 
AGM 
African green monkey 
AIDS 
acquired immunodeficiency syndrome 
BSC 
biologic safety cabinet 
CACI 
compounding aseptic containment isolator 
CDC 
Centers for Disease Control and Prevention 
cDNA 
complementary deoxyribonucleic acid 
CFR 
Code of Federal Regulations 
CGMP 
Current Good Manufacturing Practice 
CI 
confidence interval 
CIR 
Center for Immunization Research 
CL 
clinical lot 
cp 
cold-passaged 
CRADA 
Cooperative Research and Development Agreement 
CRF 
case report form 
CRL 
Charles River Laboratories 
CSO 
Clinical Safety Office 
CTM 
clinical trial material 
DAERS 
DAIDS Adverse Event Reporting System 
DAIDS 
Division of AIDS 
DAIDS PRO 
Division of AIDS Protocol Registration Office 
DC 
discontinuation 
DCR 
Division of Clinical Research 
DHHS 
Department of Health and Human Services 
DMC 
Data Management Center 
DMEM 
Dulbecco’s Modified Eagle Medium 
DNA 
deoxyribonucleic acid 
DSMB 
Data and Safety Monitoring Board 
EAE 
expedited adverse event 
EENT 
ears, eyes, nose, throat 
ELISA 
enzyme-linked immunosorbent assay 
F protein 
fusion protein  
FDA 
Food and Drug Administration 
FDAAA 
Food and Drug Administration Amendments Act of 2007 
FSTRF 
Frontier Science & Technology Research Foundation, Inc. 
GCP 
Good Clinical Practice  
GLP 
Good Laboratory Practice 
GMP 
Good Manufacturing Practice 
GMT 
geometric mean titer 
HIPAA 
Health Insurance Portability and Accountability Act 
HIV 
human immunodeficiency virus 
HJF 
Henry M. Jackson Foundation for the Advancement of Military Medicine 
IB 
investigator’s brochure 
ICD 
informed consent document 
ICH 
International Conference on Harmonisation 
IgA, IgG, IgE 
immunoglobulin A, G, E 
IMPAACT 
International Maternal Pediatric Adolescent AIDS Clinical Trials Network 
i.n. 
intranasally 
IND 
investigational new drug 

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IoR 
Investigator of Record 
IRB 
institutional review board 
i.t. 
intratracheally 
JHSPH 
Johns Hopkins Bloomberg School of Public Health 
JHU 
Johns Hopkins University 
LDMS 
Laboratory Data Management System 
LID 
Laboratory of Infectious Diseases 
LPC 
laboratory processing chart 
LRI 
lower respiratory illness 
LRT 
lower respiratory tract 
MA-LRI 
medically attended lower respiratory illness 
MOP 
manual of procedures 
mRNA 
messenger ribonucleic acid 
NHP 
nonhuman primate 
NIAID 
National Institute of Allergy and Infectious Diseases 
NIAID CRMS 
NIAID Clinical Research Management System 
NICHD 
National Institute of Child Health and Development 
NIH 
National Institutes of Health 
nt 
nucleotide 
OCRPRO 
Office of Clinical Research Policy & Regulatory Operations 
OHRP 
Office for Human Research Protections 
ORF 
open reading frame 
OTC 
over-the-counter 
PBMC 
peripheral blood mononuclear cell 
PCR 
polymerase chain reaction 
PE 
physical examination 
PFU 
plaque-forming units 
PI 
principal investigator 
PID 
participant identification number 
PRNT 
plaque reduction neutralization 
PSRT 
Protocol Safety Review Team 
r 
recombinant 
RE 
regulatory entity 
RNA 
ribonucleic acid 
rRT-PCR 
reverse transcription polymerase chain reaction 
RSC 
Regulatory Support Center 
RSV 
respiratory syncytial virus 
RT-qPCR 
quantitative reverse transcription polymerase chain reaction 
SAE 
serious adverse event 
SAM 
synthetic absorptive matrix 
SDAC 
Statistical & Data Analysis Center, Harvard School of Public Health 
SDMC 
Statistical and Data Management Center 
SERF 
Safety Expedited Report Form 
SES 
subject enrollment system 
SID 
study identification number 
SMM 
sponsor medical monitor 
SOP 
standard operating procedure 
SPG 
sucrose-phosphate-glutamate buffer 
SRCP 
Safety Review and Communication Plan 
SUSAR 
serious, unexpected, suspected adverse reaction 
TL 
tracheal lavage 
ts 
temperature sensitivity 
TSH 
shut-off temperature 
TSP 
small plaque temperature 

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UP 
unanticipated problem  
URI 
upper respiratory tract illness 
URT 
upper respiratory tract 
US 
United States 
VAR 
vaccine administration record 
WIRB 
Western Institutional Review Board 
wt 
wild-type 
∆M2-2 
deleted M2-2 
 
 
 
 

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PROTOCOL SUMMARY 
 
Phase I Placebo-Controlled Study of the Infectivity, Safety and Immunogenicity of a 
Single Dose of a Recombinant Live-attenuated Respiratory Syncytial Virus Vaccine, 
RSV 6120/∆NS2/1030s, Lot RSV#012A, Delivered as Nose Drops to  
RSV-Seropositive Children 12 to 59 Months of Age and RSV-Seronegative Infants and 
Children 6 to 24 Months of Age 
 
SHORT TITLE  
RSV 6120/∆NS2/1030s 
 
PURPOSE 
To determine whether study vaccine is attenuated and immunogenic in this age 
group 
 
DESIGN 
A double blind, randomized, placebo-controlled study design will be used to 
evaluate the safety and immunogenicity of the study product in 
RSV-seropositive and RSV-seronegative participants.  
 
STUDY POPULATION Group 1: Healthy RSV-seropositive* children ≥ 12 months to < 60 months of age 
 
Group 2: Healthy RSV-seronegative^ infants and children ≥ 6 months to < 25 months of age 
 
*RSV-seropositive: RSV serum neutralizing antibody titer ≥ 1:40;  
  determined prior to inoculation and within the calendar year of inoculation  
  
^RSV-seronegative: RSV serum neutralizing antibody titer < 1:40;  
  determined ≤ 42 days prior to inoculation  
 
SAMPLE SIZE 
Group 1: Approximately 15 RSV-seropositive children 
Group 2: Approximately 21-30 RSV-seronegative infants and children  
 
STUDY PRODUCT 
Single dose of intranasal RSV 6120/∆NS2/1030s vaccine or placebo 
 
Table 1: Inoculation Schedule 
 
 
 
 
 
 
  
*plaque-forming units (PFU) 
 
The study will start with group 1 and proceed to group 2 if there are not more than 2 vaccine-associated 
solicited adverse events (AEs) (Days 0 through 10) and no vaccine-associated lower respiratory illnesses 
(LRIs) (Days 0 through 28) in RSV-seropositive children. If 3 or more RSV-seropositive children (group 
1) have respiratory or febrile illness that coincides with shedding of > 102.5 PFU of vaccine virus as 
detected by culture, then we will evaluate the safety, infectivity, and immunogenicity of the vaccine virus 
in an additional 15 RSV-seropositive children before determining whether to continue with group 2.   
 
Group # 
N 
Study Product 
Dose 
1 
10 
RSV 6120/∆NS2/1030s  
105.7 PFU* 
5 
Placebo 
0 
2 
14-20 
RSV 6120/∆NS2/1030s  
105.0 PFU* 
7-10 
Placebo 
0 

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STUDY DURATION 
 
Participants will be enrolled in the protocol between April 1 and October 31, outside of the RSV season. 
RSV-seropositive participants will be followed for 28 days after inoculation, and RSV-seronegative 
participants will remain on the study until they complete the post–RSV season visit. 
 
After the first season of RSV surveillance is completed, research staff may invite the families of 
participants in group 2 to participate in an optional second season of RSV surveillance during November-
March of the second year following enrollment.  
 
Figure 1: Group 1 – Study Overview 
 
 
Figure 2: Group 2 – Study Overview 
 
*Post-RSV Season specimens to be collected beginning April 1st 
 

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BACKGROUND AND SCIENTIFIC RATIONALE  
 
 
BACKGROUND 
 
1.1.1 Epidemiology, Disease Burden, and the Need for a Vaccine 
 
In the United States alone, RSV is responsible for 75,000 to 125,000 hospitalizations of infants 
yearly (1), and worldwide, RSV infects at least 34 million children under 5 years old, resulting in 
an estimated 3.4 million RSV LRI hospitalizations and 66,000 to 199,000 RSV-attributable 
deaths each year (2). In temperate climates, annual RSV epidemics occur in late winter and early 
spring, and nearly all children are infected within the first 2 years of life. RSV illness can range 
from mild upper respiratory tract illness (URI), including rhinitis, pharyngitis, and coryza, to 
severe LRI, including bronchiolitis and pneumonia. Beyond the acute burden of disease caused 
by RSV, severe RSV disease in infancy may predispose to reactive airway disease during 
childhood (3, 4). 
 
RSV is an enveloped RNA virus that is a member of the newly organized Pneumoviridae family, 
genus Orthopneumovirus (5). RSV has a single negative-sense strand RNA genome of 15.2 
kilobases encoding 10 mRNAs. Each mRNA encodes a single protein, with the exception of the 
M2 mRNA, which contains 2 overlapping ORFs. The 11 RSV proteins are the viral RNA-binding 
nucleoprotein N, the phosphoprotein P, the large polymerase protein L, the attachment 
glycoprotein G, the fusion glycoprotein F, the small hydrophobic surface glycoprotein SH, the 
internal matrix protein M, the 2 nonstructural proteins NS1 and NS2, and the M2-1 and M2-2 
proteins encoded by the M2 mRNA. The gene order is: 3'-NS1-NS2-N-P-M-SH-G-F-M2-L-5'. 
RSV transcription and genome replication take place exclusively in the cytoplasm, and virions 
form by budding from the apical plasma membrane of respiratory epithelial cells.  
 
Currently, no licensed vaccine against RSV is available, although there is broad consensus that 
such a vaccine is urgently needed and should be a global health priority. Although passive 
immunoprophylaxis with the monoclonal RSV-neutralizing antibody palivizumab (Synagis®; 
MedImmune) is available for high-risk infants, this approach is not feasible for general use. A 
formalin-inactivated vaccine against RSV was evaluated clinically in the 1960s and did not 
confer protection; instead, disease enhancement occurred at a high rate following natural 
infection of vaccinees with wild-type (wt) RSV (6). Studies in experimental animals established 
that disease enhancement was specific to non-replicating RSV vaccines and not seen with 
infectious RSV or replicating vaccine vectors (7, 8).   
 
Following the failure of the formalin-inactivated RSV vaccine, attempts at developing RSV 
vaccines at NIAID have focused largely on live-attenuated approaches (9). Importantly, 
throughout a period of over 20 years, a number of live-attenuated investigational RSV vaccines 
have been evaluated in RSV-naïve infants and children, and enhanced disease following wt RSV 
infection of vaccinees has not been observed (10). Apart from the absence of enhanced disease, 
live-attenuated RSV vaccines have a number of known advantages over non-replicating RSV 
vaccines. They can be administered intranasally, induce protective mucosal immunity in the 
respiratory tract (as well as systemic immunity), infect in the presence of maternally-derived RSV 
serum antibody, and have been well tolerated and immunogenic when administered to infants as 
young as four weeks of age (11).  
 
Human RSV has a single serotype with two antigenic subgroups, A and B. The two subgroups 
exhibit a 3- to 4-fold reciprocal difference in neutralization by polyclonal convalescent serum. 
Analysis of glycoprotein-specific responses in infants by enzyme-linked immunosorbent assay 

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(ELISA) with purified F and G glycoproteins showed that the fusion proteins (F proteins) were 
50% related antigenically, and the G proteins were 7% related (12). Consistent with this level of 
antigenic relatedness, F protein expressed by a recombinant vaccinia virus was equally protective 
in cotton rats against challenge with either subgroup A or B, whereas the G protein was 13-fold 
less effective against the heterologous subgroup (13). Thus, the F protein is responsible for most 
of the observed cross-subgroup neutralization and protection, and a subgroup A vaccine virus is 
likely to induce a broad immune response against wt RSV of either subgroup. Antibodies to the F 
protein are one of the endpoints evaluated in this study. 
 
The RSV vaccines to be evaluated in this study were derived using a recombinant 
deoxyribonucleic acid (DNA)–based technique called reverse genetics (14). The technique of 
reverse genetics has been used to produce a number of licensed vaccines; among them is 
FluMist® (MedImmune). This technique allows de novo recovery of infectious virus entirely 
from complementary DNA (cDNA) in a qualified cell substrate under defined conditions. 
Reverse genetics provides a means to introduce predetermined mutations into the RSV genome 
via the cDNA intermediate. Derivation of vaccine virus from cDNA minimizes the risk of 
contamination with adventitious agents and helps to keep the passage history brief and well 
documented. Once recovered, the vaccine virus is propagated in the same manner as a 
biologically derived virus. As a result of repeated passage and amplification, the drug substance 
(clinical trial material [CTM]) does not contain any recombinant DNA. The RSV vaccine 
candidate to be tested under this protocol is a derivative of strain A2, subgroup A. 
 
The drug substance RSV 6120/∆NS2/1030s is a recombinant live-attenuated RSV vaccine 
candidate that contains a deletion of the NS2 gene (ΔNS2) and a genetically stabilized version of 
the attenuating temperature sensitivity mutation at codon 1321 of the L gene termed “1030s” 
(S1313[TCA]/Y1321K[AAA]). The RSV NS2 protein functions as an interferon antagonist, and 
deletion of NS2 is attenuating (15-21). The 1030s mutation is also attenuating and confers a 
moderate temperature-sensitive phenotype to RSV (22). RSV 6120/∆NS2/1030s has a shutoff 
temperature for replication of 40°C, and it produces only small plaques at 38°C or above. 
Compared to recombinant wt RSV, the RSV 6120/∆NS2/1030s candidate vaccine was 
significantly restricted in replication in a primary human airway epithelial cell model, and 
moderately attenuated and immunogenic in African green monkeys (AGM). 
 
 
PRIOR RESEARCH 
 
1.2.1 Experimental Vaccines against RSV 
 
Efforts have been directed toward the development of a live-attenuated RSV vaccine because of 
the advantages of live-attenuated vaccines over inactivated or subunit vaccines. These advantages 
include the ability to (i) induce the full spectrum of protective immune responses including serum 
and local antibodies as well as CD4+ and CD8+ T cells and innate immunity; (ii) infect and 
replicate in the presence of maternal antibody, permitting immunization of young infants; and (iii) 
produce an acute, self-limited, attenuated infection that is well tolerated and readily eliminated 
from the respiratory tract. Another important advantage is the absence of vaccine-related 
enhanced disease, as has been confirmed in clinical studies (10). 
 
Several live-attenuated RSV vaccines have been evaluated in clinical trials in adult and pediatric 
populations as part of NIAID's ongoing RSV vaccine development program (11, 23-26). Four 
vaccine viruses with the NS2 deletion have been evaluated in clinical studies: rA2cpΔNS2, 
rA2cp248/404ΔNS2, rA2cp530/1009ΔNS2 (27), and RSV ΔNS2/Δ1313/I1314L. A brief 
summary of clinical studies involving these vaccine candidates is provided below.  

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RSV Vaccine Candidates with an NS2 Deletion 
 
rA2cpΔNS2 contains the NS2 deletion as well as a set of 5 amino acid substitutions in the N, F, 
and L proteins that were identified in a cold-passaged (cp) RSV (V267I in N; E218A and T523I 
in F; and C319Y and H1690Y in L) (28, 29). rA2cp248/404ΔNS2 contains all of the mutations in 
rA2cpΔNS2 as well as two additional point mutations: "248," an amino acid substitution in the L 
protein (Q831L) (28, 30, 31), and "404," a nucleotide substitution in the M2 gene start signal (30, 
32). rA2cp530/1009ΔNS2 also contains all of the mutations in rA2cpΔNS2 as well as two 
additional point mutations in L: “530” (F521L) and “1009” (M1169V) (33, 34).  
 
rA2cpΔNS2, which had the fewest attenuating mutations, was evaluated in a dose-escalating 
study (105.0 PFU, 106.0 PFU, or 107.0 PFU) administered to a total of 45 adults, and was also 
evaluated at a dose of 105.0 PFU in eight 15- to 59-month-old RSV-seropositive children.  
Shedding of rA2cpΔNS2 was not detected in any of the adults, and only 2 of 16 adults who 
received 107.0 PFU had an antibody response. In contrast, at the dose of 105.0 PFU, rA2cpΔNS2 
was shed by 3 of 8 vaccinated seropositive children with mean peak titers of 103.4 PFU/mL of 
nasal wash. Two additional children had detectable virus by polymerase chain reaction (PCR). 
Based on these results, rA2cpΔNS2 was deemed under-attenuated, and was not further evaluated 
in seronegative children.  
 
rA2cp248/404ΔNS2 was evaluated in adults (n = 16 vaccinees), 15- to 59-month-old 
RSV-seropositive children (n = 16 vaccinees), and 6- to 24-month-old RSV-seronegative children 
(n = 10 vaccinees). rA2cp248/404ΔNS2 was not shed by adults. Only one seropositive child shed 
vaccine virus (peak titer of 100.4 PFU/mL of nasal wash), and four seronegative children shed 
vaccine virus (mean peak titer of 102.3 PFU/mL of nasal wash). 
  
rA2cp530/1009ΔNS2 was also evaluated in adults (n = 19 vaccinees), 15- to 59-month-old 
RSV-seropositive children (n = 14 vaccinees), and 6- to 24-month-old RSV-seronegative children 
(n = 15 vaccinees). rA2cp530/1009ΔNS2 was shed by one adult (peak titer of 100.4 PFU/mL of 
nasal wash), none of the seropositive children, and three seronegative children (mean peak titer of 
101.3 PFU/mL of nasal wash). In summary, deletion of the NS2 gene attenuates RSV. rA2cpΔNS2 
was over-attenuated for adults but under-attenuated for young children, whereas both 
rA2cp248/404ΔNS2 and rA2cp530/1009ΔNS2 were over-attenuated and insufficiently 
immunogenic for seronegative children (27). 
 
Based on these results, RSV∆NS2/∆1313/I1314L was developed. In a recent phase I trial 
(NCT01893554), this candidate vaccine was evaluated in RSV-seropositive children 12-59 
months of age at a dose of 106 PFU (10V, 5P), and no shedding or immune response was 
detected, indicative of attenuation. Next, it was evaluated in seronegative children 6-24 months of 
age at two sequential dose levels. At the lower dose of 105 PFU (15V, 7P), the vaccine was poorly 
infectious and immunogenic:  7% of recipients shed virus detected by culture, and  53% 
developed a RSV neutralizing antibody responses. At the higher dose of 106 PFU (20V, 10P), 
80% of recipients shed virus detected by culture, and 80% developed a RSV neutralizing 
antibody response. 
 
To generate a vaccine candidate with a slightly increased level of replication compared to RSV 
ΔNS2/Δ1313/I1314L, the current candidate vaccine RSV 6120/∆NS2/1030s was designed. RSV 
6120/∆NS2/1030s is expected to be slightly less restricted in replication than 
RSV∆NS2/∆1313/I1314L, and more immunogenic in RSV-seronegative infants and children. 
 

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1.2.2 Preclinical Studies 
1.2.2.1 Temperature sensitivity of RSV 6120/ΔNS2/1030s 
 
The shutoff temperature (TSH) was evaluated by efficiency of plaque formation at temperatures 
indicated in Table 2. RSV 6120/ΔNS2/1030s is moderately temperature sensitive, with a TSH of 
40°C. The TSH is defined as the lowest restrictive temperature at which the reduction in plaque 
number compared to that at the permissive temperature of 32ºC is 100-fold or greater than that 
observed for wt RSV at the 2 temperatures. Small plaques were observed at 38°C, and plaques 
formed at 39°C and above were characterized as micro-plaques. Temperature sensitivity results of 
the vaccine candidate RSV ∆NS2/∆1313/I1314L were included for comparison (Table 2). As 
expected, the temperature sensitivity of RSV ∆NS2/∆1313/I1314L was slightly greater than that 
of RSV 6120/∆NS2/1030s (TSH of 39°C, small plaque temperature [TSP] of 37°C for RSV 
∆NS2/∆1313/I1314L; TSH of 40°C, TSP of 38°C for RSV 6120/∆NS2/1030s). 
 
Table 2: Temperature Sensitivity of Experimental Lots of RSV 6120/ΔNS2/1030s 
and RSV ΔNS2/Δ1313/I1314L 
Virus 
Virus titer (log10 PFU per mL) at indicated temperature (°C) 
a 
Repl b 
32 
35 
36 
37 
38 
39 
40 
TSH c 
TSP d 
wt RSV 
1 
8.0 
8.0 
7.9 
7.9 7.8 
7.6 
7.5 
> 40 
> 40   
wt RSV 
2 
6.7 
6.9 
6.8 
6.8 6.9 
6.8 
6.6 
> 40 
> 40   
RSV ∆NS2 ∆1313 I1314L 
1 
7.5 
7.3 
7.0 
6.7 5.7 
1.7 
< 1 
39 
37 
RSV 6120/∆NS2/1030s 
2 
7.8 
7.9 
7.9 
7.7 7.8 7.5e 5.5e 
40 
38 
 
a   The temperature sensitivity (ts) phenotype for each virus was evaluated by plaque assay on Vero cells at the 
indicated temperatures. For viruses with ts phenotype, titers at shutoff temperatures are marked (bold, underlined). 
Vero cells were used for the ∆NS2 viruses because loss of expression of NS2 renders RSV more sensitive to type I 
interferon, which is not made by Vero cells. See footnote c for the definition of shutoff temperature. 
b 
wt RSV rows 1 and 2 represent duplicate dilution series of the same virus that were examined in parallel in the same 
experiment. Differences between these replicas illustrate the variability that is inherent in these biological 
experiments.  
c 
Shutoff temperature (TSH, bold, underlined) is defined as the lowest restrictive temperature at which the reduction 
compared to 32ºC is 100-fold or greater than that observed for wt RSV at the two temperatures. The ts phenotype is 
defined as having a shutoff temperature of 40ºC or less. 
d 
TSP, small plaque temperature, is defined as the lowest restrictive temperature at which the small-plaque phenotype 
is observed.  
e 
At 39 and 40°C, RSV 6120/∆NS2/1030s formed micro-plaques. 
1.2.2.2 Replication of an experimental lot of RSV 6120/ΔNS2/1030s in an in-vitro model of 
mucociliary normal human tracheal/bronchial epithelial cells 
 
Multicycle growth of an experimental lot of RSV 6120/ΔNS2/1030s was evaluated in a 3D 
mucociliary human tissue model consisting of normal differentiated primary human 
tracheal/bronchial airway epithelial cells (EpiAirway, Mattek, Inc.). Virus replication was 
evaluated at two different temperatures (32°C and 37°C) to model the temperatures of the upper 
and lower airways. Pseudostratified differentiated airway cells were cultured in an air-liquid 
interface in 12-mm transwell inserts with 0.4 µM pore sizes in Dulbecco’s Modified Eagle 
Medium (DMEM), supplemented with a proprietary mix of epidermal growth factors and other 
proprietary factors. Prior to infection, the apical surfaces of the cultures were washed with 500 µL 

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PBS to remove any accumulated mucus. Triplicate wells were infected at a multiplicity of 
infection of 0.1 with 100 µl per well of RSV 6120/ΔNS2/1030s. To evaluate replication at 32°C, 
recombinant wt RSV A2 and RSV ΔNS2/Δ1313/I1314L were included for comparison. At 37°C, 
only recombinant wt RSV was included for comparison. After 2 hours of adsorption, the 
inoculum was gently removed, and the apical surfaces of the cultures were washed 3 times with 
300 µL PBS. Apical washes were collected daily on days 1-7 post-infection to assay viral 
replication, then snap-frozen, and virus titers were determined by immunoplaque assay on Vero 
cells.  
 
Recombinant wt RSV of strain A2 replicated efficiently in differentiated primary human 
tracheal/bronchial epithelial cells at 32°C and at 37°C (Figure 3). At 32°C, peak titers of wt RSV 
were detectable on day 6, whereas at 37°C, peak titers were reached by day 5 post-infection. The 
temperature-sensitive virus RSV ΔNS2/Δ1313/I1314L, which differs from RSV 
6120/ΔNS2/1030s only by the attenuating mutation in the L ORF, was included for comparison at 
the lower temperature, 32°C. Peak titers of this virus were about 200-fold lower than those of wt 
RSV at 32°C. RSV 6120/ΔNS2/1030s replicated to slightly higher titers than RSV 
ΔNS2/∆1313/I1314L, showing that indeed, RSV ΔNS2 with the 1030s mutation is slightly less 
restricted for replication than RSV ΔNS2 with the genetically stabilized deletion of codon 1313 
of the L ORF. At 37°C, the difference in peak titers between RSV 6120/ΔNS2/1030s and wt RSV 
was statistically significant.  
 
Figure 3: Multicycle Replication of Experimental Lots in an in-vitro Model of 
Normal Human Bronchial/Tracheal Epithelial Cells 
 
 
 
Cultures were infected at a multiplicity of infection of 0.1, and supernatants were harvested daily for 7 
days, snap-frozen, and titered later on Vero cells. A: replication at 32°C. B: replication at 37°C. 
 
 

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1.2.2.3 Replication of experimental lot and CTM of RSV 6120/ΔNS2/1030s in African 
green monkeys (AGMs) 
 
Replication and immunogenicity of RSV 6120/ΔNS2/1030s were evaluated in nonhuman 
primates (NHPs), specifically African green monkeys (AGM). AGMs are semi-permissive for 
RSV. The first NHP study was done in AGMs to evaluate an experimental lot of RSV 
6120/ΔNS2/1030s, and a second study was done to evaluate the CTM (Lot RSV#012A). AGMs 
that were seronegative for RSV were inoculated intranasally (i.n.) and intratracheally (i.t.) with 
RSV 6120/ΔNS2/1030s; a dose of 1 x 106 PFU of vaccine in a 1-mL volume per site was 
administered to sedated juvenile male and female AGMs (total dose per animal: 2 x 106 PFU). 
Nasopharyngeal swabs were collected daily on days 0 through 10 and on days 12 and 14; tracheal 
lavage (TL) samples were collected every other day from days 2 through 14; and virus shedding 
was analyzed by immunoplaque assay. Serum RSV neutralizing antibody titers were determined 
by a complement-enhanced 60% plaque reduction neutralization assay. Results from studies 
following the same protocol, performed in animals from the same group and origin, inoculated 
with recombinant wtRSV A2 at the same dose, were included for comparison. Studies were 
approved by the Animal Care and Use Committee of NIAID, NIH.  
 
In both studies, shedding of the RSV 6120/ΔNS2/1030s virus from the upper respiratory tract 
(URT) and lower respiratory tract (LRT) was detectable over several days. In both studies, the 
mean peak titers of RSV 6120/ΔNS2/1030s in the nasopharyngeal samples was 3.8 log10 PFU per 
mL. These mean peak titers were slightly lower than that of wt RSV A2 (4.2 log10 PFU per mL), 
but higher than those of the previously tested RSV ΔNS2/Δ1313/I1314L virus (3.2 log10 PFU per 
mL). The mean peak titers in the LRT, detectable in TL samples, were 4.6 log10 PFU per mL for 
the Experimental Lot of RSV 6120/ΔNS2/1030s, and 2.9 PFU per mL for the CTM of RSV 
6120/ΔNS2/1030s. Shedding was observed in all AGMs in both URT and LRT. Compared to the 
RSV ΔNS2/Δ1313/I1314L virus, which showed much lower mean peak titers (1.4  log10 PFU per 
mL) with only a 75% infection rate (virus replication detected in 3 out of 4 animals), the study 
virus RSV 6120/ΔNS2/1030s was less attenuated in the AGMs (Appendix I, Table 27, Table 28).  
 
In both studies, the RSV 6120/ΔNS2/1030s virus induced serum neutralizing antibodies. In the 
first study of the experimental lot, the neutralizing antibody titers were comparable to or slightly 
higher than wt RSV A2 and the previously tested RSV vaccine virus (RSV ΔNS2/Δ1313/I1314L) 
(Appendix I, Table 29). The CTM induced titers that were slighly lower than those of wt RSV, 
and comparable to those of RSV ΔNS2/Δ1313/I1314L. These results show that at a total dose of 
2 x 106 PFU, administered i.n. and i.t., RSV 6120/ΔNS2/1030s is immunogenic in AGMs. 
 
1.2.3 Previous Clinical Experience 
 
The live-attenuated recombinant RSV 6120/∆NS2/1030s vaccine virus is being evaluated for 
the first time in humans. Based on preclinical data, RSV 6120/∆NS2/1030s is expected to be 
more attenuated than the previously tested vaccine candidate rA2cpΔNS2, but less attenuated 
than the vaccine candidates rA2cp248/404ΔNS2 and rA2cp530/1009ΔNS2, which were 
deemed over-attenuated in seronegative infants and children, and slightly less attenuated and 
more immunogenic than RSV∆NS2/∆1313/I1314L. As noted above, this vaccine is predicted to 
be attenuated and well tolerated in seropositive and seronegative children. 
 
 
 

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RATIONALE 
 
 
CLINICAL DEVELOPMENT PLAN 
 
The investigational RSV vaccine RSV 6120/ΔNS2/1030s will be evaluated sequentially in 
RSV-seropositive children followed by evaluation in RSV-seronegative infants and children. The 
main purpose of this study is to determine whether RSV 6120/ΔNS2/1030s is well tolerated and 
restricted in replication in RSV-seropositive children, and well tolerated, infectious, 
immunogenic, and genetically stable in RSV-seronegative infants and children. If the study 
product is found to be well tolerated, infectious, and immunogenic in RSV-seronegative infants 
and children, then a safety and immunogenicity study in infants < 6 months of age will be 
initiated. Plans will be made in consultation with industry collaborators as a lead RSV vaccine 
candidate is identified.  
 
The primary immunogenicity endpoints to be evaluated are RSV neutralizing antibody titer, and 
RSV F protein antibody (by ELISA). Neutralizing antibody is a well-established and important 
surrogate marker of effective immunity to RSV disease. Antibodies to the F protein are also 
associated with cross-subgroup neutralization and protection (13). These assays will be performed 
at the Johns Hopkins University (JHU) CIR laboratory. 
 
 
HYPOTHESES 
 
RSV 6120/∆NS2/1030s will be well-tolerated in RSV-seropositive and RSV-naïve participants. 
In RSV-seronegative participants, RSV 6120/ΔNS2/1030s is expected to be safe and result in 
infection, limited vaccine replication, and the induction of a neutralizing antibody response to 
RSV.  
 
 
OBJECTIVES 
 
2.1. 
PRIMARY OBJECTIVES 
 
Safety 
1. To assess the frequency and severity of study product–related solicited and unsolicited AEs in 
RSV-seropositive participants (group 1) from day 0 through the 10th day following inoculation  
2. To assess the frequency and severity of study product–related solicited and unsolicited AEs in 
RSV-seronegative participants (group 2) from day 0 through the 28th day following inoculation 
3. To assess study product–related serious adverse events (SAEs) in RSV-seropositive participants 
(group 1) from day 0 through the 28th day following inoculation  
4. To assess study product–related SAEs in RSV-seronegative participants (group 2) from day 0 
through the 56th day following inoculation  
  
Infectivity  
1. To determine the peak titer of vaccine virus shed and duration of virus shedding by each 
participant  
2. To assess the proportion of vaccinated participants infected with study vaccine (If < 70% of 
RSV-seronegative vaccinees are infected with vaccine virus, the study product will be considered 
over-attenuated.) 
 
Immunogenicity 
1. To characterize serum antibody responses to the study product 28 days after inoculation of RSV-
seropositive vaccine recipients (group 1)  

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2. To characterize serum antibody responses to the study product 56 days after inoculation of RSV-
seronegative vaccine recipients (group 2)   
2.2. 
SECONDARY OBJECTIVES 
 
1. To characterize the frequency and severity of symptomatic medically attended respiratory and 
febrile illness in RSV-seronegative vaccine and placebo recipients (group 2) who experience 
natural infection with wt RSV during the subsequent RSV season(s)  
2. To characterize serum antibody responses in RSV-seronegative vaccine and placebo recipients 
(group 2) who experience natural infection with wt RSV during the subsequent RSV season(s) 
3. To characterize mucosal antibody responses to vaccine in RSV-seropositive (group 1) and RSV-
seronegative (group 2) participants  
 
2.3. 
EXPLORATORY OBJECTIVE 
 
Study samples may be used in comparative assays with samples from other RSV vaccine studies 
initiated by the LID, NIAID, NIH. 
 
 
STUDY DESIGN 
 
The study will be double-blind, randomized, and placebo-controlled. Participants in groups 1 and 
2 will be block-randomized in groups of 3 at a ratio of 2:1 to receive vaccine or placebo, 
respectively. The vaccine will be evaluated in a stepwise fashion beginning with 
RSV-seropositive children (Group 1) and proceeding sequentially in RSV-seronegative children 
(Group 2).  For the purpose of this study, RSV-seropositive is defined as having a serum 
neutralizing antibody titer of ≥ 1:40, and RSV-seronegative is defined as having a serum 
neutralizing antibody titer of < 1:40. This definition has been used in previous evaluations of live-
attenuated RSV vaccines (11, 24, 35). In these previous studies, live-attenuated RSV vaccines 
were highly restricted in replication and poorly immunogenic in children with titers ≥ 1:40 but 
were far less restricted in replication and highly immunogenic in children with titers < 1:40. 
These data suggest that this neutralizing antibody cutoff can distinguish effectively between 
RSV-experienced and RSV-naïve children. 
 
The study will proceed to group 2 if there are not more than 2 vaccine-associated solicited AEs 
and no vaccine-associated LRIs in RSV-seropositive children. If 3 or more RSV-seropositive 
children have respiratory or febrile illness that coincides with shedding of > 102.5 PFU of vaccine 
virus as detected by culture, then we will enroll an additional 15 RSV-seropositive children to 
continue evaluating the safety, infectivity, and immunogenicity of the vaccine virus.   
 
Enrollment occurs at the time of inoculation with study product, Day 0, and will occur between 
April 1 and October 31 to avoid the time during which wt RSV typically circulates in the 
community (Appendix V, Figure 6). Accrual will stop prior to November 1. 
  
For group 1, RSV-seropositive participants, the duration of participation in the initial phase of the 
study is 28 days. The initial phase consists of an Acute Phase from inoculation through the 10th 
day after inoculation, and a Post-Acute Phase from the 11th day through the 28th day after 
inoculation. For group 2, RSV-seronegative participants, the duration of participation in the initial 
phase of the study is 56 days. The initial phase consists of an Acute Phase from inoculation 
through the 28th day after inoculation, and a Post-Acute Phase from the 29th day through the 56th 
day after inoculation.  
 

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During the Acute Phase, the participants’ parents/guardians will be contacted daily. These 
contacts will consist either of an in-person evaluation of interim medical history, clinical 
assessment, and nasal rinse, or an interim medical history conducted by a mutually agreed upon 
communication method. During the Acute Phase, the participants will be evaluated for AEs or 
SAEs. Participants who have a febrile or respiratory illness or otitis media will have a study visit 
to perform a nasal rinse. The nasal rinse will be tested for RSV and for other respiratory 
pathogens that are considered adventitious agents. During the Post-Acute Phase, study 
participants’ parents/guardians will be instructed to contact the study staff if any SAEs occur. The 
RSV-seropositive participants will have a scheduled follow-up visit on the 28th day after 
inoculation, and the RSV-seronegative participants will have a scheduled follow-up visit on the 
56th day after inoculation. The schedules of evaluations during the Acute Phase and Post-Acute 
Phase are shown in Appendix II, Table 30. 
 
For the RSV-seronegative participants (group 2), the study has a third phase that assesses the 
incidence and severity of illness suggestive of RSV occurring during the RSV season following 
inoculation. During the RSV Season Surveillance Period, encompassing November 1 to March 
31, study staff will make weekly contact with the participants’ parents/guardians to identify 
medically attended episodes of fever, URI, LRI, or otitis media. Participants who have such an 
episode will have an in-person evaluation of interim medical history, clinical assessment, and 
nasal rinse to evaluate for RSV and other respiratory pathogens that are considered adventitious 
agents (Appendix III).  
 
The RSV-seronegative participants will also have a study visit during the pre-RSV season 
between October 1 and 31 to collect a blood sample and nasal rinse sample and/or nasosorption 
synthetic absorptive matrix (SAM) strip for immunological assays. The samples will be used to 
assess the durability of the vaccine response. Participants will have a post-RSV season visit to 
collect blood and nasal rinse and/or nasosorption SAM strip specimens to further assess the 
durability of the vaccine response and to assess the immune response to naturally occurring wt 
RSV infection.. The RSV-seronegative participants may have an overlap between the initial 
phases and the RSV surveillance phase of the study, depending upon the date of inoculation.  
 
 
STUDY POPULATION 
 
Approximately 15 RSV-seropositive children ≥ 12 months to < 60 months of age will be enrolled; 
10 will receive a single dose of 105.7 PFU of vaccine and 5 will receive a single dose of placebo. 
Subsequently, 21 to 30 RSV-seronegative infants and children ≥ 6 months to < 25 months of age 
will be enrolled in the study and will receive a single dose of either 105.0 PFU of vaccine or 
placebo in a 2:1 ratio. These numbers were chosen based upon experience with phase I evaluation 
of other live-attenuated respiratory virus candidate vaccines (11,23,25) and statistical 
considerations (Section 9).  
 
Placebo recipients are needed in pediatric studies to distinguish the background respiratory and 
febrile illnesses that occur in infants and children from those attributable to study vaccine. These 
numbers were chosen based upon experience with phase I evaluation of other live-attenuated 
respiratory virus candidate vaccines (11,23,25) and statistical considerations (Section 9). 
 
Infants and children will be selected for participation according to the described study-specific 
co-enrollment considerations; the recruitment, screening, and enrollment process; and participant 
retention and withdrawal or termination. 
 

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INCLUSION CRITERIA FOR RSV-SEROPOSITIVE CHILDREN  
 
Potential RSV-seropositive participants must meet all of the following inclusion criteria to be 
enrolled in this study: 
 
4.1.1. ≥ 12 months of age and < 60 months of age at the time of inoculation 
4.1.2. Screening serum specimen for RSV-neutralizing antibody is obtained within the calendar 
year of inoculation  
4.1.3. Seropositive for RSV antibody, defined as serum RSV-neutralizing antibody titer ≥ 1:40 
4.1.4. Pre-inoculation serum sample for RSV-neutralizing antibody specimen is obtained no 
more than 56 days prior to inoculation 
4.1.5. In good health based on review of the medical record, history, and physical examination at 
the time of inoculation 
4.1.6. Received routine immunizations appropriate for age based on the Advisory Committee on 
Immunization Practices (ACIP) Recommended Immunization Schedule for Children and 
Adolescents Aged 18 Years or Younger 
4.1.7. Growing normally for age as demonstrated on a standard growth chart and has a current 
height and weight above the 3rd percentile for age  
4.1.8. Expected to be available for the duration of the study 
4.1.9. Parent/guardian is willing and able to provide written informed consent  
 
 
EXCLUSION CRITERIA FOR RSV-SEROPOSITIVE CHILDREN 
 
Potential RSV-seropositive participants who meet any of the following exclusion criteria will be 
excluded from participation in this study: 
 
4.2.1. 
Born at less than 34 weeks gestation 
4.2.2. Maternal history of positive HIV test  
4.2.3. Evidence of chronic disease 
4.2.4. Known or suspected impairment of immune function 
4.2.5. Bone marrow/solid organ transplant recipient 
4.2.6. Major congenital malformations, including congenital cleft palate or cytogenetic 
abnormalities 
4.2.7. Suspected or documented developmental disorder, delay, or other developmental problem  
4.2.8. Cardiac abnormality requiring treatment 
4.2.9. Lung disease or reactive airway disease 
4.2.10. More than one episode of wheezing in the first year of life 
4.2.11. Wheezing episode or received bronchodilator therapy within the past 12 months 
4.2.12. Wheezing episode or received bronchodilator therapy after the age of 12 months 
4.2.13. Previous receipt of supplemental oxygen therapy in a home setting 

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4.2.14. Previous receipt of an investigational RSV vaccine  
4.2.15. Previous receipt or planned administration of anti-RSV antibody product including 
ribavirin, RSV Ig or RSV mAb 
4.2.16. Previous receipt of immunoglobulin or any antibody products within the past 6 months 
4.2.17. Previous receipt of any other blood products within the past 6 months 
4.2.18. Previous anaphylactic reaction 
4.2.19. Previous vaccine-associated adverse reaction that was Grade 3 or above 
4.2.20. Known hypersensitivity to any vaccine component 
4.2.21. Member of a household that contains an infant who is less than 12 months of age at the 
date of inoculation through the 10th day after inoculation  
4.2.22. Member of a household that, at the date of inoculation through the 10th day after 
inoculation, contains an immunocompromised individual including but not limited to: 
• a person who is HIV-infected 
• a person who has received chemotherapy within the 12 months prior to enrollment 
• a person receiving immunosuppressant agents 
• a person living with a solid organ or bone marrow transplant 
4.2.23. Will attend a daycare facility that does not separate children by age and contains an infant 
who is < 12 months of age at the date of inoculation through the 10th day after inoculation 
4.2.24. Receipt of any of the following prior to enrollment: 
• any inactivated vaccine or live-attenuated rotavirus vaccine within the 14 days prior, or  
• any live vaccine, other than rotavirus vaccine, within the 28 days prior, or 
• another investigational vaccine or investigational drug within 28 days prior, or 
• salicylate (aspirin) or salicylate-containing products within the past 28 days 
4.2.25. Scheduled administration of any of the following after planned inoculation: 
• inactivated vaccine or live-attenuated rotavirus vaccine within the 14 days after, or 
• any live vaccine other than rotavirus in the 28 days after, or 
• another investigational vaccine or investigational drug in the 28 days after 
4.2.26. Receipt of any of the following medications within 3 days of study enrollment: 
• systemic antibacterial, antiviral, antifungal, anti-parasitic, or antituberculous agents, 
whether for treatment or prophylaxis, or 
• intranasal medications, or 
• other prescription medications except the permitted concomitant medications listed 
below 
Permitted concomitant medications (prescription or non-prescription) include nutritional 
supplements, medications for gastroesophageal reflux, eye drops, and topical medications, 
including (but not limited to) cutaneous (topical) steroids, topical antibiotics, and topical 
antifungal agents. 
4.2.27. Any of the following events at the time of enrollment: 
• fever (temporal or rectal temperature of ≥100.4°F), or  
• upper respiratory signs or symptoms (rhinorrhea, cough, or pharyngitis) or 
• nasal congestion significant enough to interfere with successful inoculation, or  
• otitis media 

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INCLUSION CRITERIA FOR RSV-SERONEGATIVE INFANTS & CHILDREN 
 
Potential RSV-seronegative participants must meet all of the following inclusion criteria to be 
enrolled in this study: 
 
4.3.1. ≥ 6 months of age and < 25 months of age at the time of inoculation 
4.3.2. Screening and pre-inoculation serum specimens for RSV-neutralizing antibody are 
obtained no more than 42 days prior to inoculation  
4.3.3. Seronegative for RSV antibody, defined as serum RSV-neutralizing antibody titer < 1:40 
4.3.4. In good health based on review of the medical record, history, and physical examination at 
the time of inoculation 
4.3.5. Received routine immunizations appropriate for age based on the ACIP Recommended 
Immunization Schedule for Children and Adolescents Aged 18 Years or Younger 
4.3.6. Growing normally for age as demonstrated on a standard growth chart, AND 
• If < 1 year of age: has a current height and weight above the 5th percentile for age  
• If ≥ 1 year of age: has a current height and weight above the 3rd percentile for age  
4.3.7. Expected to be available for the duration of the study 
4.3.8. Parent/guardian is willing and able to provide written informed consent  
 
 
EXCLUSION CRITERIA FOR RSV-SERONEGATIVE INFANTS & CHILDREN 
 
Potential RSV-seronegative participants who meet any of the following criteria will be excluded 
from this study: 
 
4.4.1. Born at less than 34 weeks gestation 
4.4.2. Born at less than 37 weeks gestation, and at the date of innoculation less than 1 year of age 
4.4.3. Maternal history of a positive HIV test 
4.4.4. Evidence of chronic disease 
4.4.5. Known or suspected infection or impairment of immunological functions 
4.4.6. Bone marrow/solid organ transplant recipient 
4.4.7. Major congenital malformations, including congenital cleft palate or cytogenetic 
abnormalities 
4.4.8. Suspected or documented developmental disorder, delay, or other developmental problem 
4.4.9. Cardiac abnormality requiring treatment 
4.4.10. Lung disease or reactive airway disease 
4.4.11. More than one episode of wheezing in the first year of life 
4.4.12. Wheezing episode or received bronchodilator therapy within the past 12 months 
4.4.13. Wheezing episode or received bronchodilator therapy after the age of 12 months 
4.4.14. Previous receipt of supplemental oxygen therapy in a home setting 

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4.4.15. Previous receipt of an investigational RSV vaccine   
4.4.16. Previous receipt or planned administration of anti-RSV antibody product including 
ribavirin, RSV Ig, or RSV mAb 
4.4.17. Previous receipt of immunoglobulin or any antibody products within the past 6 months 
4.4.18. Previous receipt of any blood products within the past 6 months  
4.4.19. Previous anaphylactic reaction 
4.4.20. Previous vaccine-associated adverse reaction that was Grade 3 or above 
4.4.21. Known hypersensitivity to any study product component 
4.4.22. Member of a household that contains an infant who is less than 6 months of age at the date 
of inoculation through the 28th day after inoculation 
4.4.23. Member of a household that, at the date of inoculation through the 28th day after 
inoculation, contains an immunocompromised individual including but not limited to: 
• 
a person who is HIV-infected 
• 
a person who has cancer and has received chemotherapy within the 12 months prior to 
enrollment 
• 
a person living with a solid organ or bone marrow transplant 
4.4.24. Attends a daycare facility that does not separate children by age and contains an infant < 6 
months of age at the date of inoculation through the 28th day after inoculation 
4.4.25. Receipt of any of the following prior to enrollment: 
• 
any inactivated vaccine or live-attenuated rotavirus vaccine within the 14 days prior, or  
• 
any live vaccine, other than rotavirus vaccine, within the 28 days prior, or 
• 
another investigational vaccine or investigational drug within 28 days prior, or 
• 
salicylate (aspirin) or salicylate-containing products within the past 28 days 
4.4.26. Scheduled administration of any of the following after planned inoculation 
• 
inactivated vaccine or live-attenuated rotavirus vaccine within the 14 days after, or 
• 
any live vaccine other than rotavirus in the 28 days after, or 
• 
another investigational vaccine or investigational drug in the 56 days after 
4.4.27. Receipt of any of the following medications within 3 days of study enrollment: 
• systemic antibacterial, antiviral, antifungal, anti-parasitic, or antituberculous agents, 
whether for treatment or prophylaxis, or 
• intranasal medications, or 
• other prescription medications except the permitted concomitant medications listed 
below 
Permitted concomitant medications (prescription or non-prescription) include nutritional 
supplements, medications for gastroesophageal reflux, eye drops, and topical medications, 
including (but not limited to) cutaneous (topical) steroids, topical antibiotics, and topical 
antifungal agents. 
4.4.28. Any of the following events at the time of enrollment: 
• 
fever (temporal or rectal temperature of ≥100.4°F), or  
• 
upper respiratory signs or symptoms (rhinorrhea, cough, or pharyngitis) or 
• 
nasal congestion significant enough to interfere with successful inoculation, or  
• 
otitis media 

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INCLUSION CRITERIA FOR RSV-SERONEGATIVE CHILDREN PARTICIPATING IN A SECOND 
SEASON OF RSV SURVEILLANCE  
 
4.5.1. RSV-seronegative participants who have completed the initial RSV season surveillance 
portion of the study and whose parent/guardian is willing and able to provide written 
informed consent  
4.5.2. Expected to be available during the second year of RSV surveillance 
 
 
EXCLUSION CRITERIA FOR RSV-SERONEGATIVE CHILDREN PARTICIPATING IN A 
SECOND SEASON OF RSV SURVEILLANCE  
 
4.6.1.  Currently enrolled in another RSV study 
 
 
CO-ENROLLMENT CONSIDERATIONS 
 
Co-enrollment to an investigational vaccine or investigational drug study is not allowed during 
the Acute Phase or Post-Acute Phase of this study. After the Post-Acute Phase, co-enrollment 
may be considered at the discretion of the principal investigator (PI). 
 
 
RE-ENROLLMENT CONSIDERATIONS 
 
Participants who receive placebo may re-enroll in the initial study following completion of the 
initial RSV surveillance period if they continue to meet all eligibility requirements. 
 
 
RECRUITMENT PROCESS 
 
CIR research staff will recruit participants from pediatric practices and clinics in the greater 
Baltimore/Washington area. Upon referral by the participants’ primary care provider or the 
provider’s staff, CIR staff will approach parents/guardians, present them with IRB-approved 
informational brochures, and obtain contact information. Study staff will contact interested 
parents/guardians to elaborate upon the details and requirements of the study.   
 
CIR research staff may also recruit participants through mailing IRB-approved documents to 
children of local pediatric practices and clinics, and to households in local zip codes containing age-
appropriate children. In addition, CIR staff may recruit participants through group gatherings such 
as health fairs and by posting IRB-approved recruitment materials.  
 
If parents/guardians are interested in having their child participate and the child meets the minimum 
inclusion and exclusion criteria, then the study staff will schedule a screening visit to determine the 
child’s eligibility.   
 
After the first year of RSV surveillance is completed, the parents/guardians of children enrolled in 
group 2 will be invited to have their children participate in a second year of RSV surveillance.  
Participation is optional, and separate consent will be obtained for participation in this portion of 
the study. 
 

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 PARTICIPANT RETENTION 
 
Study staff will make every effort to retain participants in the study, thereby minimizing potential 
biases associated with loss to follow-up.   
 
 PARTICIPANT WITHDRAWAL OR TERMINATION FROM THE STUDY 
 
Participants in this study may voluntarily withdraw from the study at any time. Any participant 
who has received study product will be encouraged to remain in the safety evaluation for the 
duration of the study even if sample collection is refused.  
 
A participant may withdraw or terminate participation in the study early for any of the following 
reasons: 
• 
Withdrawal of consent – applies to a parent/guardian who verbally or in writing withdraws 
consent for the participant to continue in the study for any reason. 
• 
Noncompliant with protocol – applies to a parent/guardian who does not comply with 
protocol-specific visits or evaluations on a consistent basis, such that adequate follow-up is 
not possible and the participant’s safety would be compromised by continuing in the study. 
• 
Investigator discretion – participant withdrawal may occur if the investigator believes that it 
is in the best interest of the participant. 
• 
Other – a category used when previous categories do not apply; requires an explanation. 
 
The study may be ended for the following reasons: 
• 
Research is terminated by sponsor or investigator – applies to the situation where the entire 
study is terminated by the sponsor or investigator for any reason. 
• 
The study sponsor, CIR, the institutional review board (IRB), the Office for Human Research 
Protections (OHRP), NIAID, or the US Food and Drug Administration (FDA) may decide to 
end the study. 
 
For any participant who withdraws or who is terminated from the study prior to completion of 
follow-up, study staff will document the reason for the withdrawal or termination. In the event 
that the circumstances that led to a participant’s withdrawal or termination change, the study staff 
will contact the PI to discuss options for resumption of follow-up.  Withdrawn subjects will not 
be replaced. 
 
Participants enrolled in the optional second year of RSV season surveillance may voluntarily 
withdraw before the conclusion of the second year RSV surveillance season. 
 
 
STUDY PRODUCT 
 
The unblinded dispenser should consult standard operating procedures (SOPs) and the study 
manual of operations (MOP). Refer to Figure 1 for an overview of the study design and to the 
investigator’s brochure (IB) for further information about the study product. 
 
• 
Group 1: Live Recombinant Respiratory Syncytial Virus RSV 6120/∆NS2/1030s, approximately 
105.7 PFU per 1.0 mL vaccine 
• 
Group 2: Live Recombinant Respiratory Syncytial Virus RSV 6120/∆NS2/1030s, approximately 
105.0 PFU per 0.5 mL vaccine 
• 
Group 1: Placebo for the RSV vaccine will be Lactated Ringer’s Solution for Injection, USP 1.0 
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• 
Group 2: Placebo for the RSV vaccine will be Lactated Ringer’s Solution for Injection, USP 0.5 
mL 
 
 
STUDY PRODUCT REGIMENS 
 
Enrolled study participants will receive a single dose of RSV 6120/∆NS2/1030s vaccine or 
placebo, administered as nose drops.  
 
 
STUDY PRODUCT FORMULATION  
 
5.2.1 RSV 6120/∆NS2/1030s  
 
The RSV 6120/∆NS2/1030s vaccine is provided in a sterile 2.0-mL cryovial, each containing 0.6 
mL of vaccine (Lot RSV#012A). The vaccine virus concentrate is diluted by trained research 
personnel to a dose of approximately 105.7 PFU (group 1) in 1.0 mL or 105.0 PFU (group 2) in a 
0.5-mL volume. The vaccine vial is labeled as shown in Figure 4.  
 
Figure 4: Investigational Product Label Sample 
Live recombinant Respiratory Syncytial Virus   
RSV 6120/ΔNS2/1030s 
VERO GROWN VIRUS VACCINE 
Date: 29AUG2016 
Vial#:XXXX 
Lot: RSV#012A 
CAUTION:NEW DRUG LIMITED 
 BY FEDERAL (USA) LAW 
TO INVESTIGATIONAL USE 
Store at –80°C ± 15°C 
Charles River Laboratories, Malvern, PA 
(Enlarged Sample) 
 
5.2.2 Diluent for RSV 6120/∆NS2/1030s  
 
The diluent for RSV 6120/∆NS2/1030s is Lactated Ringer’s Solution for Injection, USP.  
 
5.2.3 Placebo for RSV 6120/∆NS2/1030s   
 
The placebo for RSV 6120/∆NS2/1030s is Lactated Ringer’s Solution for Injection, USP. 
 
 
 
STUDY PRODUCT STORAGE 
 
Vaccine will be stored in a secure freezer at -80°C ± 15°C. It must remain frozen until the time of 
use. Once the vaccine is thawed, it should never be refrozen for reuse. Vaccine will be prepared 
from new, unopened containers for each use.  
 

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Lactated Ringer’s Solution for Injection, USP should be stored at room temperature as 
recommended by the supplier until the day before study product preparation. Vaccine 
diluent/placebo must be transferred to a secure 2°C to 8°C refrigerator at least 24 hours before 
use.  
 
Procedures for managing the vaccine and diluent/placebo shipment are in the MOP. 
 
 
STUDY PRODUCT PREPARATION 
 
The diluent for the vaccine, the placebo for the vaccine, and the RSV vaccine must be prepared 
by following the detailed instruction in the MOP. 
 
The unblinded dispenser will prepare the correct dose of study product for each participant in a 
biological safety cabinet (BSC) or compounding aseptic containment isolator (CACI) using 
aseptic technique. If necessary to preserve blinding, all prepared syringes will be as described in 
the MOP. 
  
5.4.1 Diluent  
 
The diluent is Lactated Ringer’s Solution for Injection, USP. 
 
 
5.4.2 Placebo 
 
Placebo is Lactated Ringer’s Solution for Injection, USP.  
 
Placebo will be drawn up in a sterile syringe to a volume of 1.0 mL for group 1 and to a volume 
of 0.5 mL for group 2 and labeled per instructions in the MOP. If necessary to preserve blinding, 
all prepared syringes will be as described in the MOP. 
 
5.4.3 Live RSV 6120/∆NS2/1030s  
 
Diluent will be prepared prior to removal of vaccine from the freezer. The MOP will be followed 
for proper handling of the study product.  
 
Approximately three vials per dose of undiluted vaccine will be used to prepare the 
administration dose. When manipulating the undiluted study product, the smallest gauge needle 
possible will be used to avoid loss of study product in the needle and syringe hub. Concentration 
of the undiluted study product is approximately 106 PFU per mL. The frozen study product will 
be thawed and diluted with Lactated Ringer’s Solution for Injection, USP to a dose of 
approximately 105.7 PFU in 1.0 mL prior to administration for group 1. For subjects in group 2, 
the frozen study product will be thawed and diluted with Lactated Ringer’s Solution for Injection, 
USP to a dose of approximately 105.0 PFU in 0.5mL prior to administration.  
 
The diluted study product will be drawn up to a volume of 1.0 mL (group 1) or 0.5 mL (group 2) 
in a sterile syringe and labeled per instructions in the MOP. The labeled filled syringes will be 
transported in a cooler at 2ºC to 8ºC with ice or cold packs to the clinical site for administration. 
Vaccine must be administered within 4 hours of being removed from the freezer.  Placebo must 
be administered within 4 hours of being removed from the refrigerator. 
 

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Samples of undiluted (if available) and diluted study product will be aliquoted from the remaining 
vaccine that has been prepared. The samples will be snap-frozen as per the MOP and stored at  
-80°C ± 15°C separate from the concentrated study product. Titration of vaccine will be 
completed to confirm the titer of the vaccine administered to the participants.  
 
The MOP provides detailed instructions on vaccine storage, handling, preparation, labeling and 
transport to the clinic. 
 
 
STUDY PRODUCT INOCULATION PROCEDURE 
 
All study participants will receive a single dose of study product, administered as nose drops. 
There is no nasal preparation prior to administration. While the participant is supine, a volume of 
1.0 mL (group 1) or 0.5 mL (group 2) of study product will be delivered as nose drops 
(approximately 0.5 mL [group 1] or 0.25 mL [group 2] per nostril) using a sterile, needle-less, 
masked syringe. Participant will remain supine for approximately 60 seconds following 
inoculation. 
 
 
STUDY PRODUCT ACQUISITION 
 
The clinical lot of RSV 6120/∆NS2/1030s was generated by Charles River Laboratories (CRL) 
using the seed virus provided by the NIH.  
 
Lactated Ringer’s Solution for Injection, USP will be used  as diluent and placebo. 
 
Vaccine virus will be stored at a NIAID-designated commercial repository until formally 
requested by the PI/designee. Prior to IRB approval and FDA safe to proceed determination, the 
PI/designee at JHU CIR may request that vials of vaccine be transferred from the sponsor to the 
CIR laboratories at the JHSPH for confirmation of the vaccine titer. However, only after receipt 
of IRB approval and FDA safe to proceed determination, will vials of vaccine be used for 
administration to study subjects. Such an initial shipment may contain the full number of vials 
needed for implementation of the protocol, but no vaccine will be administered to participants 
until all necessary approvals have been received. Procedures for ordering and shipping of the 
study product are in the MOP.  
 
 
STUDY PRODUCT ACCOUNTABILITY 
 
The unblinded dispenser is responsible for maintaining an accurate inventory and accountability 
record of study-product and Lactated Ringer’s Solution for Injection, USP for this study. A copy 
of the randomization code will be retained by the unblinded dispenser as outlined in the MOP. 
Without written request to unblind, the randomization code may not be released. The unblinded 
dispenser will be responsible for maintaining the blind.  
  

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DISPOSITION OF USED/UNUSED STUDY PRODUCT 
 
After the unblinded dispenser dilutes the vaccine and draws up the vaccine into a syringe for 
administration, the label will be removed from the vaccine vials used for preparation and placed 
in the accountability log. In this manner, monitoring personnel will be able to verify the 
accountability of all vaccine vials used for the study. If there is any vaccine left after the syringes 
have been drawn up and aliquots have been removed for titering, it will be destroyed by research 
personnel as per the MOP.  
 
 
FINAL DISPOSITION OF STUDY PRODUCTS 
 
After study completion or termination, all unused study product will be disposed of per the 
sponsor’s instructions.  
  
 CONCOMITANT MEDICATIONS  
 
Permitted concomitant medications at enrollment (prescription or non-prescription) include 
nutritional supplements, medications for gastroesophageal reflux, eye drops, and topical 
medications, including (but not limited to) cutaneous (topical) steroids, topical antibiotics, and 
topical antifungal agents. 
 
The use of prophylactic antipyretics, decongestants, or antihistamines is discouraged during the 
Acute Phase: day of inoculation day through the 10th day after inoculation for group 1; and day of 
inoculation through the 28th day following inoculation for group 2. However, use of these 
medications for treatment of symptoms is allowed.  
 
Due to the potentially confounding effect on study immunogenicity results, the following 
concomitant medications should be avoided after inoculation unless deemed clinically necessary.  
 
• 
Licensed inactivated vaccine or live-attenuated rotavirus vaccine within 14 days of 
inoculation 
• 
Licensed live virus vaccine, other than rotavirus vaccine, within 28 days of inoculation 
• 
Systemic corticosteroids for more than 14 days at a dosage equivalent to prednisone at 
> 2 mg/kg or 20 mg daily, or other immune-modifying drugs within 28 days of inoculation 
• 
Immunoglobulins and/or any blood products within 28 days of inoculation 
• 
Investigational drug or investigational vaccine within 28 days of inoculation for group 1 and 
within 56 days of inoculation for group 2 
 
 
STUDY VISITS AND PROCEDURES 
 
An overview of the study visits, evaluation schedule, and specimen collection is provided in 
Appendix II and Appendix III. This section contains additional information on visit-specific study 
procedures.  
 

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Study visits, except inoculation, may be performed at one of the clinical sites or at a mutually 
agreed-upon location. Inoculation must be performed at one of the clinical sites. Unless otherwise 
specified, visits may be split, with required procedures performed on more than one day within 
the allowable visit window. All clinical tasks will be performed by a medical professional. The 
physical examination will include temperature, heart rate, respiratory rate, assessment of ears, 
eyes, nose, and throat (EENT), lungs, heart, and skin, as well as abdominal, musculoskeletal, and, 
as appropriate, neurological exams. A focused clinical examination will include temperature, 
heart rate, respiratory rate, EENT, lung, heart and lymph nodes assessment. 
 
All specimens will be obtained, processed, stored and transferred per the MOP.  
 
In addition to the protocol-specified procedures listed in this section, study staff may complete 
other tasks consistent with SOPs, including but not limited to collecting, reviewing, and updating 
demographic and locator information; reviewing elements of informed consent; scheduling 
telephone contacts and visits; providing instructions for contacting study staff between visits; 
providing visit reminders; and following up on missed visits. All visit procedures will be 
documented in accordance with the MOP. Refer to Section 10 for more information on 
documentation requirements and completion of case report forms (CRFs).  
 
The study’s medical professional will inform parent/guardian of any significant abnormal 
physical findings and, after obtaining parental release of information, will make appropriate 
referrals back to the child’s primary caregiver, if necessary. 
 

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CONSENTING PROCESS          
 
The screening process may be completed under a separate screening protocol and consent or 
under the study protocol and consent. The parent/guardian must complete the informed consent 
process and sign the informed consent document (ICD) before screening or study procedures are 
performed. The consenting process for the study may take place during the screening visit or may 
be conducted at a separate visit. During the consenting process, the child’s parent will review the 
ICD, be encouraged to ask questions, and complete a comprehension assessment to evaluate 
consent understanding. Study staff will use incorrect answers from the comprehension assessment 
to identify areas of the ICD that need further review with the parent/guardian. This will help 
ensure that the parent/guardian has sufficient understanding of the process before the ICD is 
signed. Parent/guardian will be offered a copy of the ICD. 
 
For group 2, enrollment into the second RSV surveillance period will be initiated under a separate 
consent from the ICD used for the primary study. 
 
During the consenting process, the parent will be given the opportunity to provide permission for 
use of their child’s picture in advertisement flyers, articles, or presentations. In addition, the 
parent will be given the opportunity to provide permission for storing their child’s specimens for 
future respiratory virus vaccine studies and for research purposes. 
 
As needed, a parent/guardian will complete a Health Insurance Portability and Accountability Act 
(HIPAA) medical record release to allow study staff to review medical history and immunization 
records, and to allow review of AEs during the study. Only those portions of the medical record 
that are pertinent to the study will be maintained in the study chart. 
 
 
SCREENING VISIT 
 
The screening process will include reviewing the medical record, obtaining a medical history 
from parent/guardian, and conducting a physical examination. The medical record review should 
include review of history related to the study eligibility criteria, the immunization record, and 
growth chart data. The medical history from the parent should include demographics, prior 
diagnoses, current medications, signs and symptoms, developmental status, age of household 
members, day care attendance, and use of medications prior to inoculation. 
 
The screening visit will also include obtaining a serum sample to test for the presence of RSV 
antibodies. Approximately 5 mL of blood is sufficient quantity for the screening and pre-
inoculation assays. The procedure will be performed and documented per the SOP. As in previous 
phase I trials of other live-attenuated RSV vaccines (11, 23-25), other screening laboratory tests 
will not be performed on the participant. Such tests are not routinely performed as part of well-
child care, given that the risk of undiagnosed hepatic, metabolic, and renal diseases is much lower 
in children than in adults (36).  
 
For the RSV-seropositive participants, the screening serum sample must be obtained in the same 
calendar year as study inoculation. The pre-inoculation serum antibody sample must be obtained 
no more than 56 days prior to inoculation.  
 
For the RSV-seronegative participants, the screening serum sample must be obtained no more 
than 42 days prior to inoculation. Study staff should consider potential randomization and 
inoculation dates when scheduling the participant screening visits. 
 

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Table 3: Screening Visit Procedures 
Screening Visit Procedures  
Administrative Tasks 
 
• Conduct consenting process 
• Confirm parent/guardian’s informed consent comprehension 
• Obtain release of medical records as required per HIPAA 
• Obtain available medical records 
• Assess eligibility 
•  
Clinical Tasks 
• Obtain, review, and document medical history 
• Perform complete physical examination, or 
physical examination may be deferred until study day 0 
• Address any concerns 
• Document findings 
• Obtain and review participant’s immunization record  
• Review participant’s medical records to determine age-appropriate 
developmental assessment, and participant’s weight and length 
Laboratory 
Tasks 
Blood 
Collect blood for: 
• Screening1 and pre-inoculation2 sample for RSV serum antibody testing  
Follow-up Preparation 
• Schedule enrollment visit  
1. Obtained within the same calendar year as inoculation for RSV-seropositive participants and no more than 
42 days prior to inoculation for RSV-seronegative participants 
 2. Obtained no more than 56 days prior to inoculation for RSV-seropositive participants and no more than 
42 days prior to inoculation for RSV-seronegative participants 
 
6.2.1 Participants Enrolled in Surveillance for a Second RSV Season  
 
No screening is needed prior to the second RSV surveillance period. 
 
 
RANDOMIZATION 
 
Randomization numbers will be assigned by the unblinded laboratory vaccine dispenser and will 
be forwarded to the Data Safety Monitoring Board (DSMB) Executive Secretary. Unblinded 
laboratory personnel will label the vaccine syringes with the study virus vaccine name, PI initials, 
study number, subject’s randomization number, preparer and vaccine verifier’s initials and 
expiration date and time. The vaccine administrator and verifier will initial the label and place the 
label in the subject’s study chart. The vaccination time will be documented on the vaccine 
administration record (VAR). All syringes will be disposed of by the study staff following the 
enrollment visit. A copy of the VAR will be sent to the laboratory. 
  
A copy of the randomization code will be retained by the unblinded dispenser in the laboratory 
where the vaccine is prepared. Without the PI's written request to unblind, the randomization code 
will not be released to the clinical staff until the acute monitoring phase is complete for each 
subject within the randomization group. Pediatric subjects will be block-randomized in groups of 3 
(2 vaccinees and 1 placebo recipient per block); this allows for maintenance of the blind during 
each subject’s acute monitoring phase but also provides early information regarding safety, which 
is appropriate and common practice for phase 1 respiratory virus vaccine studies (11,23,25). 
Detailed procedures for randomization and unblinding will be placed in the laboratory study 
binder. 
 
 

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ENROLLMENT  
 
6.4.1 Enrollment - Inoculation (Day 0) 
Enrollment will occur between April 1 and October 31 to avoid the RSV season. For the purpose 
of this protocol, enrollment will correspond to inoculation with investigational product, which 
occurs on study day 0, and must be completed at a CIR office site or pediatric practice where 
emergency supplies are available. Prior to inoculation, an authorized prescriber will provide a 
request for study product to the unblinded dispenser. The request must include the information 
outlined in the MOP.  
 
The ICD will be obtained from the parent/guardian of each child who participates in this study 
prior to the performance of any study procedures or inoculation. If not previously obtained, then 
the ICD will be completed on the day of enrollment. The child’s parent/guardian will be 
encouraged to ask questions and complete a comprehension assessment to evaluate understanding 
of the study. Study staff will use incorrect answers from the comprehension assessment to identify 
those areas of the ICD that need further review with the parent/guardian. This will help ensure that 
the parent/guardian has sufficient understanding of the study process before the ICD is signed.  
 
If the participant is noted to have any of the following on enrollment day, inoculation must be 
deferred: 
• 
fever (temporal or rectal temperature of ≥ 100.4°F), or  
• 
URI or LRI symptoms or signs (including but not limited to rhinorrhea, cough, or 
pharyngitis), or 
• 
nasal congestion significant enough to interfere with successful inoculation, or  
• 
otitis media. 
 
If the inoculation for an RSV-seropositive participant has to be deferred to the following calendar 
year, or if the 42-day window from screening to inoculation is exceeded for an RSV-seronegative 
participant, then the infant or child must be rescreened.  

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Table 4: Enrollment Visit Procedures 
Enrollment Visit Procedures (Day 0)*   
Administrative Tasks 
 
• Confirm study ICD is completed and signed and dated by both 
parent/guardian and study staff who completed the consenting process 
• Complete eligibility determination and confirmation 
• Complete paper-based eligibility checklist 
• Obtain release of medical records as required per HIPAA 
Clinical Tasks 
• Obtain interim history from parent/guardian 
• Perform physical exam: 
o 
Complete physical exam if deferred at screening, OR 
o 
Focused physical exam if physical completed at 
screening 
• Address any concerns  
• Document findings  
• Confirm eligibility 
Laboratory 
Tasks 
Blood  
If insufficient volume obtained at screening, collect blood for: 
• Pre-inoculation RSV antibody titer  
Nasosorption 
SAM Strip 
and/or Nasal 
Wash  
Collect nasosorption strip for: 
• RSV antibody assays 
Note: The nasosorption must be performed prior to the nasal wash. 
The nasal wash must be obtained prior to administering the study 
product. 
Collect nasal wash for: 
• RSV antibody assays 
• RSV viral detection and quantification 
Study Product Administration 
• Administer study product and maintain participant in a supine position 
for 1 minute 
• Observe for approximately 30 minutes after inoculation to evaluate for 
immediate hypersensitivity reactions 
Follow-up Preparation 
• Provide the following: temperature card with explanation, temporal and 
rectal thermometers with instructions for use, illness criteria 
explanation, and study personnel contact information  
• Schedule non-visit day contact and schedule next in-person visit 
• Offer parent/guardian safety seat education materials, and a safety seat 
educator appointment 
• Offer parent lactation services by a certified lactation counselor, if 
appropriate 
*Within same calendar year as screening for RSV-seropositive participants, and no more than 42 days from 
screening for RSV-seronegative participants 
 
Following the inoculation day visit, the parent/guardian will record the infant/child’s 
temperatures and signs of illness on the temperature card and provide these to study personnel 
during an in-person visit or non-visit day contact. New rectal thermometers will be given and 
temporal artery thermometers will be provided to parent/guardian for use during the study. For 
temperature measurements, parent/guardian will be instructed to use the study-provided temporal 
artery thermometer to screen for elevated temporal artery temperatures. This device is used to 
minimize the number of rectal temperature measurements and has been shown to be an effective 
screening tool for rectal fever (37). The parent/guardian will measure temporal artery 
temperatures following the manufacturer’s directions. If any temporal artery temperature is 
≥ 100.0°F, parent/guardian will be asked to measure a rectal temperature within 20 minutes (37). 
For study-specific management and grading of temperatures, see Section 8, Table 22. 
 

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6.4.2 Enrollment in the Second Season of RSV Surveillance (Group 2 Only) 
 
Participation in RSV surveillance for a second season will require completion of an additional 
consent document by the parent or guardian. The consent document for this optional portion of the 
study may be done at the pre–RSV second seasonal surveillance visit or at a separate visit prior to 
the pre–RSV second seasonal surveillance visit. 
 
 
STUDY PHASES  
 
Refer to Figures 1 and 2 for timelines of study visits. The Acute Phase begins with inoculation 
and ends at midnight on the 10th day after inoculation for RSV-seropositive participants, and at 
midnight on the 28th day after inoculation for RSV-seronegative participants. During the Acute 
Phase of the study, a study healthcare professional will be available by telephone 24 hours a day 
for consultation with parent/guardian regarding any illnesses that may occur. 
 
Study personnel will have daily contact with RSV-seropositive participants’ parents/guardians for 
the first 10 days after inoculation, and for the RSV-seronegative participants during the first 28 
days after inoculation. This 28-day period is consistent with the duration of shedding of live-
attenuated respiratory virus vaccines in RSV-seronegative participants (28, 37-39).  
 
 
On non-visit days, study staff will contact the parent/guardian and will record the 
parent/guardian-provided temperatures and signs of illness. Participants with illness may have 
additional visits to assess the illness (Section 6.8, Illness Visit). 
 
6.5.1 RSV-Seropositive Children (Group 1) 
6.5.1.1 Acute Phase: In-person Study Visit Days  
 
An in-person study visit and clinical assessment performed by a healthcare professional will be 
completed during visits on days 3, 4, 5, 6, 7, and 10 after inoculation, with a visit window of ± 1 
day. If an in-person visit is moved by ± 1 day, then the non-visit day contact will be completed in 
place of the original interim visit date. 
 
Table 5: Acute Phase In-Person Visit Procedures – RSV-Seropositive Children 
Days 3, 4, 5, 6, 7 and 10 In-person Visit Procedures (each visit ± 1 days) 
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s 
previous days’ interim history including:  
 
medications and/or immunizations 
 
signs and symptoms of illness 
 
highest temperature reading, indicate temperature method    
• Perform focused clinical examination 
• Address any concerns 
• Review safety data 
• Document findings  
Laboratory 
Tasks 
Nasal wash  
Collect nasal wash for: 
• RSV viral detection and quantification 
Follow-up Preparation 
• Schedule non-visit day contact and follow-up, in-person visits 
Day 10 only:  
• Review SAE criteria with participants and how to contact study 
personnel during Post-Acute Phase  

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During an Acute Phase study visit, if the participant is diagnosed with or suspected of having 
URI, LRI, otitis media or fever (as defined in Appendix IV), testing of the nasal wash specimen 
for adventitious agents will be performed as described in the MOP.  
6.5.1.2 Acute Phase: Non-Visit Study Day Contacts 
 
In the 10 days following inoculation, parental contact will be made on days that an in-person visit 
is not completed: days 1, 2, 8, and 9 (each visit ± 1 day). On non-visit days, study staff will 
contact the parent/guardian and will record the parent/guardian-provided temperatures and signs 
of illness. Participants with illness may have additional visits to assess the severity of the illness 
(Section 6.8). 
 
Table 6: Acute Phase Non-Visit Contact Procedures – RSV-Seropositive Children 
Days 1, 2, 8, and 9 Contact Procedures (each visit ± 1 day) 
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s 
interim history from previous days, including:  
 
medications and/or immunizations 
 
signs and symptoms of illness 
 
highest temperature reading, indicate temperature method 
• Address any concerns 
• Review safety data 
• Document findings 
Follow-up Preparation 
• Schedule an illness appointment if necessary 
 
6.5.1.3 Study Day 11  
 
There will be a non-visit contact on Day 11 to obtain interim history through midnight on the 10th 
day following inoculation. If the Day 10 Visit takes place on Day 11, it is not necessary to have 
an additional contact with the family on Day 11. 
 
Table 7: Day 11 Non-Visit Procedures – RSV-Seropositive Children 
Day 11 Non-Visit Procedures  
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s 
previous days’ interim history including:  
 
medications and/or immunizations 
 
signs and symptoms of illness 
 
highest temperature reading, indicate temperature method 
• Address any concerns 
• Review safety data 
• Document findings 
Follow-up Preparation 
• Review SAE criteria with participants and how to contact study 
personnel during Post-Acute Phase  
 
 
 

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6.5.1.4 Post-Acute Phase  
 
The Post-Acute Phase begins on the 11th day after inoculation and ends on the 28th day after 
inoculation. During the Post-Acute Phase, parent/guardian will be instructed to monitor for and 
contact the study staff if their child has symptoms that are suggestive of a SAE. If the parent 
reports an SAE that may meet the study pause or stop criteria (Section 8.3), then an Illness Visit 
will be scheduled (Section 6.8). 
 
6.5.1.5 Study Day 28 Visit 
 
The Day 28 Visit should be conducted between 28 and 35 days following inoculation. Because 
the Post-Acute Phase ends as the 28th day following inoculation, only events through that time 
should be evaluated as having occurred during the Post-Acute Phase.  
 
Table 8: Day 28 Visit Procedures – RSV-Seropositive Children 
Day 28 Visit (+7 Days) 
Administrative Tasks 
• Provide study compensation 
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s interim 
history from midnight of the 10th day through the  28th day 
following inoculation, including:  
 
visits to medical provider/hospitalizations 
 
medications and/or immunizations 
 
signs and symptoms of illness meeting SAE criteria 
• Address any concerns 
• Review safety data 
• Document findings 
Laboratory 
Tasks 
Blood 
Collect blood for: 
• Serum antibodies to RSV 
Nasosorption 
SAM Strip 
and/or Nasal 
Wash  
Collect nasosorption strip for: 
• RSV antibody assays 
Note: The nasosorption must be performed prior to the nasal wash. 
The nasal wash must be obtained prior to administering the study 
product. 
Collect nasal wash for: 
• RSV antibody assays 
 
6.5.2 RSV-Seronegative Infants and Children 
6.5.2.1 Acute Phase: In-person Study Visit Days  
 
An in-person study visit and clinical assessment performed by a healthcare professional will be 
completed during visits on Study Days 3, 5, 7, 10, 12, 14, 17 and 28 after inoculation with a visit 
window of ± 1 day. If an in-person visit is moved by ± 1 day, then the non-visit day contact is 
completed in place of the original interim visit date. 
 
 

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Table 9: Acute Phase In-Person Visit Procedures – RSV-Seronegative Children 
Days 3, 5, 7, 10, 12, 14, 17, and 28 In-person Visit Procedures (each visit ± 1 days) 
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s 
previous days’ interim history including:  
 
medications and/or immunizations 
 
signs and symptoms of illness 
 
highest temperature reading, indicate temperature method    
• Perform focused clinical examination 
• Address any concerns 
• Review safety data 
• Document findings  
Laboratory 
Tasks 
Nasal Wash 
and/or 
Nasosorption 
SAM Strip 
Collect nasal wash for: 
• RSV viral detection and quantification 
Day 28 only: Collect nasal wash for: 
• RSV antibody assays 
• RSV viral detection and quantification 
Day 28 only: Collect nasosorption strip for:  
• RSV antibody assays 
Note: The nasosorption must be performed prior to the nasal wash. 
Follow-up Preparation 
• Schedule non-visit day contact and follow-up, in-person visits 
Day 28 only:  
• Review SAE criteria with participants and how to contact study 
personnel during Post-Acute Phase  
6.5.2.2 Acute Phase: Non-Visit Study Day Contacts 
 
In the 28 days following inoculation, parental contact will be made on days that an in-person visit 
is not completed: days 1, 2, 4, 6, 8, 9, 11, 13, 15, 16, and 18–27 (each visit ± 1 day). On non-visit 
days, study staff will contact the parent/guardian and will record the parent/guardian-provided 
temperatures and signs of illness. Participants with illness may have additional visits to assess the 
severity of the illness (Section 6.8). 
 
Table 10: Acute Phase Non-Visit Contact Procedures – RSV-Seronegative Children 
Days 1, 2, 4, 6, 8, 9, 11, 13, 15, 16, 18 , 19, 20, 21, 22, 23, 24, 25, 26 and 27 Contact Procedures 
(each visit ± 1 day) 
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s 
previous days’ interim history including:  
 
medications and/or immunizations 
 
signs and symptoms of illness 
 
highest temperature reading, indicate temperature method 
• Address any concerns 
• Review safety data 
• Document findings 
Follow-up Preparation 
• Schedule an illness appointment if necessary 
 
 
 

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6.5.2.3 Study Day 29  
 
There will be a non-visit contact on Day 29 to obtain interim history through midnight on the 28th 
day following inoculation. If the Day 28 Visit takes place on Day 29, it is not necessary to have 
an additional contact with the family on Day 29. 
 
Table 11: Day 29 Non-Visit Procedures – RSV-Seronegative Children 
Day 29 Non-Visit Procedures  
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s 
previous days’ interim history including:  
 
medications and/or immunizations 
 
signs and symptoms of illness 
 
highest temperature reading, indicate temperature method 
• Address any concerns 
• Review safety data 
• Document findings 
Follow-up Preparation 
• Review SAE criteria with participants and how to contact study 
personnel during Post-Acute Phase  
 
6.5.2.4 Post-Acute Phase  
 
The Post-Acute Phase begins on the 29th day after inoculation and ends on the 56th day after 
inoculation. During the Post-Acute Phase, parent/guardian will be instructed to monitor for and 
contact the study staff if their infant or child has symptoms that are suggestive of a SAE. If the 
parent reports an SAE that may meet the study pause or stop criteria (Section 8.3) then schedule 
an Illness Visit (Section 6.8). 
 
6.5.2.5 Study Day 56 Visit 
 
The Day 56 Visit should be conducted between 56 and 63 days following inoculation. Because 
the Post-Acute Phase ends on the 56th day following inoculation, only events through that time 
should be evaluated as having occurred during the Post-Acute Phase.  
 
 

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Table 12: Day 56 Visit Procedures – RSV-Seronegative Children 
Day 56 Visit (+7 Days) 
Administrative Tasks 
• Provide first study compensation payment 
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s 
interim history from midnight of the 28th day through the 56th day 
following inoculation, including:  
 
visits to medical provider/hospitalizations 
 
medications and/or immunizations 
 
signs and symptoms of illness meeting SAE criteria 
• Address any concerns 
• Review safety data 
• Document findings 
Laboratory 
Tasks 
 
Blood 
Collect blood for: 
• Serum antibodies to RSV 
Nasosorption 
SAM Strip 
and/or Nasal 
Wash  
Collect nasosorption strip for: 
• RSV antibody assays 
• Note: The nasosorption must be performed prior to the nasal wash. 
The nasal wash must be obtained prior to administering the study 
product. 
Collect nasal wash for: 
• RSV antibody assays 
Follow-up Preparation 
Follow-up depends on when, during the calendar year, the Day 56 
Visit is conducted. If Day 56 Visit is conducted: 
• Prior to October 1 
 
schedule Pre-RSV Season Visit (Section 6.6.1) 
• On or after October 1 
 
Day 56 Visit will also be the Pre-RSV Season Visit 
 
Review plans for weekly contact during the RSV Season 
Surveillance Period (Section 6.6.2) 
 
6.5.2.6 Period after Day 56 through October 31st 
 
During this period, contact with the participant is not required except for the Pre-RSV Season 
Study Visit described in Section 6.6.1. No clinical data will be recorded on CRFs or reported 
under this protocol except for data as outlined in Section 7.2. 
 
 
RSV SURVEILLANCE: FIRST SEASON 
 
6.6.1 Pre-RSV Season Surveillance Study Visit  
 
The Pre-RSV Season Study Visit is not required if the Day 56 Visit is conducted on or after 
October 1; the samples collected at the Day 56 Visit are sufficient for the Pre-RSV Season Study 
Visit. Otherwise, an in-person visit is expected of participants during the Pre-RSV Season for 
collection of a blood sample, nasal wash sample and/or nasosorption strip for RSV antibody 
assays.  
 

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Table 13: Pre-RSV Season Surveillance Study Visit Procedures 
Pre-RSV Season Study Visit (October 1 through October 31 of enrollment year) 
Clinical Tasks 
• Document findings related to study procedure 
Laboratory 
Tasks 
 
Blood 
 
Collect blood for: 
• Serum antibodies to RSV  
Nasosorption 
SAM Strip 
and/or Nasal 
Wash  
Collect nasosorption strip for: 
• RSV antibody assays 
• Note: The nasosorption must be performed prior to the nasal wash. 
The nasal wash must be obtained prior to administering the study 
product. 
Collect nasal wash for: 
RSV antibody assays 
Follow-up Preparation 
• Review plans for weekly contact during the RSV Season 
Surveillance Period (see Section 6.6.2) 
 
6.6.2 Weekly Contact for Surveillance during the RSV Season  
 
Based on previous data regarding the seasonality of RSV in the Baltimore, MD, area (Appendix 
IV, Figure 6), surveillance for RSV-associated disease will be conducted between November 1 
and March 31 during the first RSV season following receipt of study product. For some RSV-
seronegative participants, surveillance during the first RSV season may overlap with the Acute 
and/or Post-Acute study phases. In this case, all evaluations required for each of the relevant 
phases of the study will be conducted.  
 
During the first RSV season following receipt of study product, participants enrolled in this study 
will be monitored for symptomatic, medically attended, RSV-like illnesses listed below via 
weekly telephone or email communication or an in-person visit. (Rhinorrhea and cough need not 
meet the Appendix IV criteria if they are documented by the health care provider): 
• 
Medically attended fever 
• 
Medically attended URI  
• 
Medically attended otitis media 
• 
Medically attended LRI  
 
An Illness Visit will be scheduled within 3 days of study staff notification of any of these events 
(Section 6.8). 
 
Table 14: RSV Season Surveillance Procedures 
RSV Season Surveillance (November 1 through March 31 following inoculation) 
Clinical Tasks 
• Obtain interim history 
• Review safety data 
• Document findings 
Follow-up Preparation 
• Continue with weekly contacts through March 31 
• Schedule an Illness Visit if warranted  
• Schedule the Post-RSV Season Study Visit (targeted April 1st - 
April 30th with allowable window through September 30th) 
 

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6.6.3 Post-RSV Season Surveillance Study Visit 
 
The Post-RSV Season Surveillance Visit will occur in the calendar year following receipt of 
study product, ideally between April 1st and April 30th through September 30th. 
 
Table 15: Post-RSV Seasonal Surveillance Study Visit Procedures 
Post-RSV Season Study Visit  
Administrative Tasks 
• Provide second study compensation 
Clinical Tasks 
• Document findings related to study procedures 
Laboratory 
Tasks 
 
Blood 
 
Collect blood for: 
• Serum antibodies to RSV  
Nasosorption 
SAM Strip  
Collect nasosorption strip for: 
• RSV antibody assays 
 
 
 
 
RSV SURVEILLANCE: SECOND SEASON  
 
The second RSV seasonal surveillance is November 1 through March 31 in the second season 
following inoculation and will be optional for RSV-seronegative children (group 2) who have 
participated in the first surveillance season. An ICD for the second season of RSV surveillance 
must be signed before any procedures are performed. If a participant’s parent/guardian agrees to 
have their child participate in a second season of RSV surveillance, the procedures for the second 
season will be identical to those outlined for the first season in Section 6.6, except that a Pre-
Season 2 Visit with blood and nasal sample will always be required, and the Post-Season 2 Visit 
will occur in the second calendar year following enrollment in the study.  
 
 
ILLNESS VISIT 
 
The timeframe after staff notification in which the Illness Visit must occur depends on the study 
phase and the grading of the fever and respiratory symptoms per Section 8.2 and If the Illness 
Visit occurs on a day concurrent with an in-person study visit, a single nasal wash collection is 
required and adventitious agent testing will be requested. All symptoms will be followed until 
resolution or deemed stable or chronic by appropriate medical personnel (e.g., medical doctor or 
nurse practitioner). Illness Visits may occur during any of the study phases. 
 
If the acute or post-acute phases overlap the surveillance period (between November 1 and March 
31), then the timelines for the acute and post-acute phases will be used. 

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Table 16: Illness Visit Timeframe 
Illness Visit Timeframe 
Phase 
Symptoms 
Grade 
Visit Timeframe 
Acute 
Fever, otitis media or URI 
1 
Within 3 days  
Acute 
Fever, otitis media or URI 
≥ 2 
Within 2 days  
Acute 
LRI 
Any 
Within 1 day 
Post-Acute 
SAE that meets study pause 
or stop criteria (Section 8.3) 
≥ 2 
Within 3 days 
RSV Season 
Surveillance 
Medically attended fever, 
otitis media, URI or LRI 
≥ 2 
Within 3 days 
 
Table 17: Illness Visit Procedures 
Illness Visit Procedures 
Administrative Tasks 
• Complete Adventitious Agent Assay Request for rRT-PCR on 
nasal wash for adventitious agents 
Clinical Tasks 
• Obtain and document from parent/guardian the participant’s 
interim history including:  
 medications and/or immunizations 
 signs and symptoms of illness 
 highest temperature reading, indicate temperature method    
• Perform focused clinical examination 
• Address any concerns 
• Review safety data 
• Document findings  
Laboratory 
Tasks 
Nasal Wash 
Collect nasal wash for: 
• Viral detection and quantification 
Follow-up Preparation 
• Schedule follow-up as appropriate 
 
 
EARLY DISCONTINUATION STUDY VISIT 
 
In the event that a child is unable to continue participation in the study, parent/guardian will be 
encouraged to allow the participant to complete safety monitoring through Day 28 for RSV-
seropositive participants and through Day 56 for RSV-seronegative participants.  Every effort 
should be made to schedule a final Early Discontinuation Visit. 

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Table 18: Early Discontinuation Procedures 
Early Discontinuation 
Administrative Tasks 
• Record data on CRF  
Clinical Tasks 
• Obtain interim history 
• Address any concerns 
• Review safety data 
• Encourage parent/guardian to allow the participant to complete 
safety monitoring through Day 28 for RSV-seropositive 
participants and through Day 56 for RSV-seronegative participants 
• Document findings 
Laboratory 
Tasks 
 
Blood 
Collect blood for: 
• Serum antibodies to RSV 
Nasal Wash 
and/or 
Nasosorption 
SAM Strip 
If Early Discontinuation Visit is within 28 days of inoculation for 
RSV-seropositive participants or 56 days for RSV-seronegative 
participants (Appendix II), collect nasal wash for: 
• Viral detection and quantification 
• RSV antibody assays 
Collect nasosorption strip for: 
• RSV antibody assays 
Note: The nasosorption must be performed prior to the nasal wash. 
 
 LABORATORY PROCEDURES 
 
6.10.1 Specimen Collection 
 
Specimens will be collected for this study as indicated in the Schedule of Events. Further 
information on collection of blood and nasal wash specimens will also be provided in the MOP.  
 
6.10.2 Virus Detection 
 
Specimens for viral culture and quantification of vaccine virus shedding will be obtained by nasal 
wash with approximately 20 mL of Ringer’s lactate solution once before and approximately 6 
times after inoculation for RSV-seropositive participants and approximately 8 times after 
inoculation RSV-seronegative participants as shown in Appendix II. These specimens may also 
be tested for adventitious respiratory viruses if needed. Additional nasal wash specimens for 
detection of RSV and adventitious respiratory viruses by culture and rRT-PCR will also be 
obtained from participants who meet illness criteria during the initial phase (Day 0 through Day 
28 for RSV-seropositive participants; Day 0 through Day 56 for RSV-seronegative participants) 
as well as during the RSV First Seasonal Surveillance Period (November 1 – March 31) for all 
RSV-seronegative participants and the RSV Second Season Surveillance for RSV-seronegative 
participants whose parents/guardians elect to participate. If a participant becomes ill during the 
Acute or Post-Acute Phase, then up to 10 additional nasal washes over a 28-day period for group 
1 participants and over a 56-day period for group 2 participants may be obtained to exclude 
infection with an adventitious virus. Laboratory testing will be performed by personnel who are 
not involved with clinical assessment to maintain the blinding status. 
 

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6.10.3 Immunologic Assays 
 
For measurement of RSV serum antibodies, serum specimens will be obtained in 
RSV-seropositive children not more than 56 days prior to inoculation and on Day 28 (+7) 
postinoculation; and in RSV-seronegative infants and children not more than 42 days prior to 
inoculation and on Day 56 (+7) postinoculation. In addition, pre-first RSV season and post-first 
RSV season serum specimens will be collected in all RSV-seronegative infants and children. 
These samples will be used to determine whether a four-fold or greater rise in antibody titer has 
occurred during the first RSV season, which would signify infection with wt RSV. This will 
allow comparison of the rate and severity of significant RSV illness following infection with wt 
virus, as well as comparison of the antibody responses, in vaccine and placebo recipients.  
 
For subjects in group 2 enrolled in surveillance during the second RSV season, RSV 
neutralizing antibodies will also be measured in serum specimens collected before and after the 
second RSV season: between October 1 and October 31 (pre-season 2), and with a target 
window of  April 1 to April 30 which can be extended through September 30 (post-season 2). 
This will allow assessment of the duration of antibody responses to vaccination, and 
comparison of the rate and severity of significant RSV illness following infection with wt virus 
in vaccine and placebo recipients during the second season. 
 
Specimens will be obtained by venipuncture, finger-stick, or heel-stick. No more than 5 mL of 
blood will be drawn at each blood draw visit. A maximum of 10 mL of blood will be taken from 
RSV-seropositive subjects for study purposes. A maximum of 20 mL of blood will be taken from 
RSV-seronegative subjects for study purposes from screening through season 1 surveillance. A 
maximum of an additional 10 mL of blood will be taken from RSV-seronegative subjects who 
participate in season 2 surveillance. 
 
For RSV-seropositive participants, nasal wash specimens and/or nasosorption strips for 
measurement of secretory immunity will be obtained before inoculation and 28 days after 
inoculation. For RSV-seronegative participants, nasal wash specimens and/or nasosorption strips 
for measurement of secretory immunity will be obtained before inoculation, 28 and 56 days after 
inoculation, and at the pre- and post-RSV season 1 surveillance visits. These specimens may be 
generated from the same nasal wash obtained for viral culture, but specimen processing is 
different as described in the MOP. If the parent/guardian opts to participate in the second year of 
RSV surveillance, a nasal wash specimen and/or nasosorption strip will also be obtained at the 
second year pre- and post-RSV surveillance visits to measure secretory immunity.    
 
6.10.4 Specimen Preparation, Testing, Storage, and Shipping 
 
All specimens collected for this study will be labeled, transported, processed, tested, stored and/or 
shipped in accordance with SOPs. The frequency of specimen collection and testing will be 
directed by the Schedule of Evaluations (Appendix II and Appendix III).  
 
6.10.5 Research Laboratories 
 
Quantitation of the amount of vaccine virus shed, assays to measure immune responses before 
and after inoculation, and assessment of nasal washes for adventitious viral agents will be 
performed at the CIR. Nasosorption assays will be performed at the LID, NIAID. 
Cytokine/chemokine assays may also be performed on nasal washes from participants infected 
with vaccine virus if sufficient material is available. Selected specimens may be sent to LID, 
NIAID for confirmatory testing.  

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6.10.6 Plan for Use and Storage of Biological Samples 
 
All specimens collected as part of this study may, with the parent/guardian’s permission, be 
stored for future research as part of CIR’s approved biospecimen repository for vaccine research. 
These samples and data may be used for future screening for respiratory virus vaccine studies and 
to learn more about RSV infection and other diseases. The parent/guardian or child will not own 
the blood specimens, nasal fluid, or data after it is given to the study. No financial benefit will be 
provided to the parent/guardian or child from any product or idea created by the investigators 
using the data or materials. These samples will not be sold or used to make commercial products. 
Genetic tests will not be performed on these samples unless separate informed consent is 
obtained.  
 
Samples stored in the repository will be labeled with the study identification number of the 
participants that, by themselves, cannot identify study participants but are linkable to the study 
databases generated by the main study. The repository database will contain only the study 
participants’ numbers. A master log linking the study participants’ identification numbers to their 
names is maintained by the study’s clinical staff in a password-protected computer system with 
limited access to authorized research team members and will not be shared with the laboratory. 
Study participants, or their parents/guardians, may withdraw consent for future testing of their 
specimens at any time. 
 
6.10.7 Biohazard Containment 
 
As the transmission of blood-borne pathogens can occur through contact with contaminated 
needles, blood, and blood products, appropriate blood and secretion precautions will be employed 
by all personnel. The procedures for obtaining, shipping, and handling of all specimens for this 
study will follow the current recommendation of the Centers for Disease Control and Prevention 
(CDC) and the NIH. All infectious specimens will be transported using packaging mandated in 
Title 42 of the Code of Federal Regulations (CFR), Part 72 (42 CFR 72) and in accordance with 
individual carrier guidelines (e.g., Federal Express). 
 
 
SAFETY ASSESSMENT, MONITORING, AND REPORTING 
 
Participant safety will be carefully assessed, monitored, and reported at multiple levels 
throughout this study. Sections 7.1-7.4 describe safety-related roles, responsibilities, and 
procedures. The safety monitoring roles of the NIAID Intramural Data Safety and Monitoring 
Board (DSMB) are briefly referenced in Section 7.1.2 and described in detail in Section 9.4.2. 
 
 
SAFETY-RELATED ROLES AND RESPONSIBILITIES 
 
7.1.1 Principal Investigator 
 
The PI is responsible for continuous monitoring of the CIR’s study participants and for alerting 
the protocol team if unexpected concerns arise. Trained study staff will record safety-related data 
on CRFs as indicated in Section 7.2. The PI is also responsible for prompt reporting to the IRBs 
and other applicable review bodies of any unanticipated problems (UPs) involving risks to 
participants or others.  
 

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7.1.2 Safety Review and Communications Plan (SRCP)  
A Safety Review and Communication Plan (SRCP) was developed for the protocol. The SRCP is 
an internal communications document between the PI and the IND sponsor (OCRPRO) Clinical 
Safety Office, which delineates the safety oversight responsibilities of the PI, the CSO, and other 
stakeholders. The SRCP also includes the overall plan for conducting periodic safety surveillance 
assessments.  
 
7.1.3 Sponsor Medical Monitor 
 
A medical monitor representing the IND sponsor (OCRPRO) has been appointed for oversight of 
safety in this clinical study. The sponsor medical monitor will be responsible for performing 
safety assessments as outlined in a Safety Review and Communications Plan (SRCP). 
 
7.1.4 Data Safety Monitoring Board 
 
An independent DSMB will monitor participant safety through routine and as-needed reviews of 
study data. Refer to Section 9.4.2 for more information on the composition and role of the DSMB 
in the monitoring of this study.   
 
7.1.5 Sponsor Reporting 
 
A SUSAR is a suspected adverse reaction that is both serious and unexpected, as defined in 21 
CFR 312.32. SUSARs will be reported to the FDA and all participating investigators as IND 
Safety Reports. The Regulatory Sponsor will also submit a brief report of the progress of the 
investigation to the FDA on an annual basis as defined in 21 CFR 312.33. 
Following notification from the PI, OCRPRO, as the IND Sponsor, will report all SAEs to the 
FDA within the required timelines. Fatal and life-threatening events will be reported within 
7 calendar days of the sponsor’s awareness and all other SAEs will be reported within 15 calendar 
days of the sponsor’s awareness.  
 
 
SAFETY-RELATED RECORDING ON CRFS 
 
AEs that occur during protocol-specified AE reporting periods following inoculation of study 
product should be considered AEs. The current section outlines which events should be collected 
on source documents and which should be recorded on CRFs for inclusion in the database. 
 
AEs may be observed by the study investigator or designee, elicited or volunteered from the 
parent/guardian or participant, or captured on participant’s temperature cards. Assessment of 
safety will include clinical observation and monitoring of laboratory parameters as necessary. 
Follow-up measures such as history, physical examination, and laboratory testing and/or 
treatment may be necessary if a participant experiences an AE. Details of AEs will be properly 
documented on the source documents, recorded on CRFs, and reported to LID investigators and 
the DSMB in separate semi-annual and annual reports. AEs will be provided to the IRB as 
defined by the IRB policy. 
 
This study has several periods of AE observation that have different AE CRF recording 
requirements. In addition, for RSV-seronegative participants, there may be a period when no AEs 
are recorded on CRFs if the Day 56 Visit occurs in advance of the start of the RSV Season 
Surveillance Period (November 1). The AEs (solicited and unsolicited; and SAEs) to be recorded 
on CRFs and the study phase and the calendar dates during which they are to be reported are 
defined in Table 19 and Table 20.  

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Concomitant medications and AEs identified in this study will be recorded on CRFs. AEs 
will be recorded as signs, symptoms, laboratory test results, and diagnoses, as shown in 
Table 19 and Table 20.  
 
 

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Table 19: AE CRF Recording Requirements for RSV-Seropositive Subjects 
Study Phase at the Time of Event 
Onset 
Calendar 
Date 
AEs to Record on CRFs 
Concomitant Medications to 
Record on CRFs 
Day 0 through midnight of 10th day 
following inoculation 
(Acute Phase) 
ANY 
• All SAEs 
• All solicited AEs that meet 
Appendix IV, criteria  
• All unsolicited AEs (Grades 1 to 
4), with the exception of the 
following conditions if not 
treated with prescription 
medication or non-prescription 
medications with antipyretic 
properties: diaper rashes, teething 
pain, and spitting up 
Record these medications on the 
CRFs regardless of whether the 
related event is recorded on the 
CRFs: 
 
• All cough and cold remedies 
including decongestants, cough 
suppressants, expectorants 
• All nasal sprays (except saline 
spray) 
• All antihistamines 
• All antipyretics 
• All prescription medications 
 
For SAEs and LRIs: 
• All medications related to the 
recorded event 
From the 11th day after inoculation 
to the 28th day after inoculation 
(Post-Acute Phase) 
ANY 
• All LRIs and SAEs 
 
 
• All medications related to the 
recorded event 
Throughout study 
ANY 
• Unresolved AEs or SAEs with 
onset from Day 0 to midnight on 
the 10th day after inoculation 
• Unresolved LRIs and SAEs with 
onset prior to the 28th day 
following inoculation 
•  
• All medications related to the 
recorded event 
 
 
 

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Table 20: AE CRF Recording Requirements for RSV-Seronegative Subjects 
Study Phase at the Time of Event 
Onset 
Calendar 
Date 
AEs to Record on CRFs 
Concomitant Medications to 
Record on CRFs 
Day 0 through midnight of 28th day 
following inoculation 
(Acute Phase) 
ANY 
• All SAEs 
• All solicited AEs that meet 
Appendix IVcriteria  
• All unsolicited AEs (Grades 1 to 
4), with the exception of the 
following conditions if not 
treated with prescription 
medication or non-prescription 
medications with antipyretic 
properties: diaper rashes, teething 
pain, and spitting up 
 
 
Record these medications on the 
CRFs regardless of whether the 
related event is recorded on the 
CRFs: 
 
• All cough and cold remedies 
including decongestants, cough 
suppressants, expectorants 
• All nasal sprays (except saline 
spray) 
• All antihistamines 
• All antipyretics 
• All prescription medications 
 
For SAEs and LRIs: 
• All medications related to the 
recorded event 
From the 29th day after inoculation 
to the 56th day after inoculation 
(Post-Acute Phase) 
ANY 
• All SAEs 
 
 
• All medications related to the 
recorded event 
From Day 56 Visit through start of 
RSV Season Surveillance Period 
 
Up to 
October 31 
in year of 
inoculation 
• Grade ≥ 3 AEs or SAEs that is 
deemed related to Pre-RSV 
Season Study Visit procedures 
• All medications related to the 
recorded event 
RSV First and Second Season 
Surveillance Periods 
November 
1 to March 
31  
• Fevers, LRIs, URIs, and/or otitis 
media that are medically 
attended  
• All SAEs  
 
Note: These events do not need to 
meet the Appendix IV criteria 
For SAEs and LRIs (all grades): 
• All medications related to the 
recorded event 
 
Medications related to recorded 
medically attended illness should 
be documented in source notes 
Post-RSV Season 
Ideally 
April 1 to 
April 30; 
allowable 
through 
Sept 30th 
in the year 
after the 
inoculation 
• Grade ≥ 3 AEs or SAEs that are 
deemed related to Post-RSV 
Season Study Visit procedures 
• All medications related to the 
recorded event 
Throughout study 
ANY 
• Unresolved AEs or SAEs with 
onset from Day 0 to midnight on 
the 28th day after inoculation 
• Unresolved SAEs with onset prior 
to the 56th day following 
inoculation 
• Unresolved SAEs with onset 
during RSV Surveillance Period 
or related to the Pre- or Post-RSV 
Season Study Visit 
• All medications related to the 
recorded event 
 

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SERIOUS ADVERSE EVENT REPORTING 
 
All SAEs will be reviewed by a study physician, recorded on the Safety Expedited Report Form (SERF) 
and followed through to resolution. SAEs that have not resolved by the end of the follow-up period are 
followed until final outcome is known. If it is not possible to obtain a final outcome for an SAE (e.g., the 
subject is lost to follow-up), the reason a final outcome could not be obtained will be recorded by the 
investigator on the AE CRF and the SERF. In addition to the SAE Reporting Category identified below, 
other AEs that must be reported in an expedited manner are LRIs as defined in Appendix IV if they occur 
during study days 0 to 10 in groups 1 and study days 0 to 28 in group 2. 
 
 
Deaths and immediately life-threatening SAEs will be reported within 1 business day after the site 
becomes aware of the event. All other SAEs will be reported within 3 business days of site awareness. 
SAEs will be reported either by fax or e-mail to all of the following: 
 
• 
Sponsor: Office of Clinical Research Policy and Regulatory Operations, NIAID, NIH: Clinical 
Safety Office (CSO): Phone: 301-846-5301, Fax: 301-846-6224, rchspsafety@mail.nih.gov 
• 
DSMB Executive Secretary: Phone 301-846-5301, Fax: 301-846-6224, 
niaiddsmbia@niaid.nih.gov 
• 
LID, NIAID: Ursula Buchholz, PhD, NIAID/NIH, 301-594-1533, Fax: 301-480-1268, 
Email: ubuchholz@niaid.nih.gov 
  
SAEs will also be reported to WIRB (contact information below) based on its reporting requirements: 
 
• 
Western Institutional Review Board (WIRB), 1019 39th Ave SE, Puyallup, WA 98374, Phone: 1-
800-562-4789 
 
 
REPORTING OF UNANTICIPATED PROBLEMS TO OCRPRO 
 
An UP is defined as any event, incident, experience, or outcome that is: 
1. Unexpected in terms of nature, severity, or frequency in relation to 
a. The research procedures that are described in the IRB-approved research protocol and 
informed consent or other study documents; and 
b. The characteristics of the participant population being studied; and  
2. Possibly, probably, or definitely related to participation in the research; and 
3. Places participants or others at a greater risk of harm (including physical, psychological, 
economic, or social harm) than was previously known or recognized. Per the IND sponsor, an 
AE with a serious outcome will be considered increased risk. 
 
UPs must be reported to the local IRB per their requirements. Non-Serious AEs that are UPs must 
also be reported to the sponsor CSO. Submit the local IRB UP report form to the CSO at the 
following address no later than 7 calendar days of the PI awareness of the event.   
 
SPONSOR CLINICAL SAFETY OFFICE CONTACT INFORMATION:  
 
Clinical Safety Office 
5705 Industry Lane 
Frederick, MD 21704 
Phone 301-846-5301 
Fax 301-846-6224 

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E-mail: rchspsafety@mail.nih.gov 
 
UPs that are not AEs (UPnonAE) are not routinely reported to the CSO. However, an UPnonAE 
that may, in the opinion of the investigator, involve risk to the participant, affect others in the 
research study, or significantly impact the integrity of research data would be considered a non-
serious UP and would be reported to the CSO. For example, we will report occurrences of 
breaches of confidentiality, accidental destruction of study records, or unaccounted-for study 
drug. 
 
 
 
PARTICIPANT MANAGEMENT 
 
 
MANAGEMENT OF ADVERSE EVENTS 
 
An AE is any untoward medical occurrence in a participant administered a pharmaceutical 
product that does not necessarily have a causal relationship with this treatment. An AE can 
therefore be any unfavorable and unintended sign (including an abnormal laboratory finding), 
symptom, or disease temporally associated with the subject’s participation in the research, 
whether or not related to the subject’s participation in the research. This includes exacerbation of 
pre-existing conditions and intercurrent illnesses.  
 
Causality assessment is based on available information at the time of the assessment of the AE. 
The investigator may revise the causality assessment as additional information becomes 
available. 
 
All AEs identified in this study will be source documented, consistent with the policies and 
procedures referenced in Section 10. Among other details, source documentation will include the 
severity of each event (graded as described in Sections 8.2.1 and 8.2.2 and its relationship to 
study product, assessed by the study clinician according to the following categories and 
definitions: 
 
Related 
There is a reasonable possibility that the adverse event may be 
related to the study drug. 
 
Not related 
There is not a reasonable possibility that the adverse event may be 
related to the study drug.  
 
There are two categories of AEs specific to CIR 322: solicited and unsolicited. Solicited AEs are 
described in Section 8.1.1. Unsolicited AEs are all other AEs. However, for infants and children, 
the following common events will not be recorded as AEs unless a prescribed concomitant 
medication is used to treat them: non-infectious rashes, teething pain, and spitting up. The use of 
medications will only be captured in the event of an AE or pre-existing condition. 
 
Serious Adverse Events (SAEs) are described in Section 8.1.2. 
 

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8.1.1 Solicited Adverse Events 
 
Solicited AEs are predefined AEs that can occur after study product administration, may be 
expected to occur if the study product is insufficiently attenuated, and have protocol-specific 
criteria for reporting.  
 
Solicited AEs, whether identified by a parent/guardian or clinician, are only recorded on CRFs if 
they meet the definitions per Appendix IV Individual symptoms listed in the “events” column that 
fail to meet the criteria in the “definition” column in Appendix IV are recorded in source 
documents but are not recorded on the CRFs. During the Acute Phase of this study, days 0 
through 10 for RSV-seropositive participants and days 0 through 28 for RSV-seronegative 
participants, solicited AEs meeting the criteria for reporting will be recorded on CRFs, assigned a 
severity grade (Section 8.2), and assessed for relationship to study product (see Section 8.1). 
Solicited AEs are defined in Appendix IV and include the following:  
 
1. Fever 
2. URI 
a. Rhinorrhea, 
b. Pharyngitis,  
c. Cough without LRI, or 
d. Hoarseness 
3. Otitis Media 
4. LRI 
a. Wheezing, 
b. Pneumonia, 
c. Laryngotracheobronchitis (croup), 
d. Rhonchi, or 
e. Rales 
 
Solicited AEs Elicited by History Unconfirmed by Clinical Assessment 
 
With the exception of fever, solicited AEs reported by parents/guardians are NOT recorded on 
CRFs if a clinical assessment done on the day of the event(s) does/did not confirm their presence. 
For example, if a parent/guardian reports rhinorrhea on the day of visit, and there is/was no 
rhinorrhea upon exam, then the participant is considered to not have rhinorrhea that day. 
 
If the parent/guardian report of a fever meets the “definition” column criteria in Appendix IV on a 
day on which there was a clinical assessment, the fever will be recorded on CRFs regardless of 
whether the clinical assessment confirmed its presence. 
 
Events elicited by parent/guardian history for days on which there was no clinical exam will be: 
 
1) Recorded on the CRFs as AEs if the parent/guardian description meets the “definition” 
column criteria in Appendix IV. 
2) Recorded only on the source document, and NOT on the CRF, if the parent/guardian 
description fails to meet the “definition” column criteria in Appendix IV. For example, both 
rhinorrhea and cough must each occur on 2 consecutive days to meet the definition required 
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8.1.2 Serious Adverse Events 
 
A SAE is an AE, whether considered related to the study product or not, that meets one or more 
of the following criteria: 
1. Results in death during the period of protocol-defined surveillance. 
2. Is life threatening: defined as an event in which the participant was at immediate risk of death 
at the time of the event; it does not refer to an event that hypothetically might have caused 
death were it more severe. 
3. Requires inpatient hospitalization (or prolongation of existing hospitalization): defined as at 
least an overnight stay in the hospital or emergency ward for treatment that would have been 
inappropriate if administered in the outpatient setting. 
4. Results in a persistent or significant disability/incapacity. 
5. Is a congenital anomaly or birth defect. 
6. Is an important medical event that may not be immediately life threatening or result in death 
or hospitalization but may jeopardize the patient or may require intervention to prevent one of 
the outcomes listed above. 
 
8.1.3 Unexpected Adverse Event 
 
An AE is unexpected if it is not listed in the Investigator’s Brochure or Package Insert (for 
marketed products) or is not listed at the specificity or severity that has been observed. 
It is the responsibility of the IND Sponsor to make this determination. 
 
 
GRADING THE SEVERITY OF ADVERSE EVENTS 
 
The Investigator will assess all AEs with respect to Seriousness (criteria listed above), Severity 
(intensity or grade), and Causality (relationship to study agent and relationship to research).  All 
AEs and fever will be graded using the protocol-defined grading system.  
 
8.2.1 AE Grading  
 
Table 21: Grading for Adverse Events 
Severity 
Defined 
Grade 1 Mild 
No medical intervention required; may include over-the-counter medications 
managed by the participant or caregiver for treatment of symptoms 
Grade 2 Moderate 
Outpatient medical intervention by a health care provider required; may include 
use of over-the-counter and/or prescription medications 
Grade 3 Severe 
Prolonged medical intervention and/or hospitalization required 
Grade 4 Life threatening Illness requiring hospitalization with intensive care  
Grade 5 Death 
Event resulting in fatal outcome to the participant 
 

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8.2.2 Fever Grading 
 
 
 
 
Table 22: Grading for Fever 
Severity 
Defined 
Grade 1 
≥100.4ºF but ≤101.4ºF 
Grade 2 
≥101.5ºF but ≤102.4ºF 
Grade 3 
≥102.5ºF but ≤104.8ºF 
Grade 4 
≥104.9ºF 
Applies to any modality of temperature measurement  
 
 
PAUSING AND STOPPING RULES 
 
If any of the following occur in a participant during the specified period after receiving the study 
product, additional enrollment/inoculations will be temporarily suspended at all sites (Table 23). 
 
Table 23: Pausing and Stopping Rules 
Specified Phase 
Event 
Reporting 
Acute and Post-Acute 
Days 0 through 28 
following inoculation 
for RSV-seropositive 
subjects and days 0 
through 56 following 
inoculation for RSV-
seronegative subjects 
An SAE that cannot be attributed to 
an etiology or cannot be attributed 
to a cause unrelated to the study 
product. 
A description of the vaccine-associated 
AE(s) or safety issue must be reported by 
the PI or study staff, within one business 
day of the PI's awareness, to the CSO and 
the DSMB by fax or email. 
 
Acute Only 
Days 0 through 10 
following inoculation 
for RSV-seropositive 
subjects and days 0 
through 28 following 
inoculation for RSV-
seronegative subjects 
An LRI per Appendix IV, OR 
A fever of Grade 4 OR 
Any Grade 3 or above solicited AE 
(other than fever) 
 
The regulatory sponsor (OCRPRO) will determine if the FDA needs to be notified. The DSMB 
and regulatory sponsor will be informed and receive pertinent data of the event by the PI. Follow-
up visits for participants already inoculated will continue as outlined in Appendix II (Table 30).  
 
If the event is determined to have occurred in a participant who received active agent (vaccine), 
and the event meets one of the following stopping rule criteria, then the event will be reviewed by 
the DSMB prior to resuming enrollment.  

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1. One or more participants experience an SAE that cannot be attributed to an etiology or cannot 
be attributed to a cause unrelated to the study vaccine, OR   
2. One or more participants develop LRI associated with shedding of vaccine virus at the time 
of the LRI (even if another pathogen is identified, unless the RSV is confirmed to be wt 
RSV), OR 
3. One or more participants develop LRI that is not explained by a diagnosis unrelated to the 
vaccine virus, OR  
4. One or more participants experiences a Grade 4 fever or any Grade 3 or Grade 4 solicited AE 
other than fever associated with shedding of vaccine virus, OR 
5. Any pattern of research laboratory values or clinical symptoms is observed that the protocol 
team considers a significant safety issue for participants. 
 
The DSMB will notify the PI (via the study sponsor) of their recommendations as to whether 
enrollment can resume, or if the study needs to be stopped. The regulatory sponsor (OCRPRO) 
will be informed about this decision. In the event of an SAE, the study may be resumed if it can 
be demonstrated to the DSMB that there is no proven causal relationship with the vaccine. 
 
 
STATISTICAL CONSIDERATIONS 
 
 
GENERAL DESIGN ISSUES 
 
9.1.1 General Design 
 
The goal of this phase I, blinded, randomized, placebo-controlled vaccine trial is to assess the 
safety, infectivity, and immunogenicity of the RSV 6120/∆NS2/1030s vaccine candidate in 
RSV-seropositive and RSV-seronegative pediatric participants. Fifteen RSV-seropositive 
participants will be randomized in a 2:1 ratio to receive either the candidate vaccine at a dose of 
105.7 PFU or placebo. Twenty-one to thirty RSV-seronegative participants will be randomized in a 
2:1 ratio to receive either the candidate vaccine at a dose of 105.0 PFU or placebo.   
 
9.1.2 Description of the Statistical Methods to be Employed 
 
This study, like other phase I studies, is exploratory, rather than confirmatory; its purpose is to 
assess frequencies of AEs and patterns of immune responses. Descriptive approaches will be used 
to meet the protocol objectives as stated in Section 2 of this protocol, as well as formal statistical 
tests as outlined in Section 9.5.  
 
 
OUTCOME MEASURES 
 
9.2.1 Primary Outcome Measures 
 
Safety: Types and grades of study product-related: 
• 
Solicited AEs as defined in Appendix IV from Study Days 0-10 for RSV-seropositive and 
Study Days 0-28 for RSV-seronegative participants 
• 
Unsolicited AEs from Study Days 0-10 for RSV-seropositive and Study Days 0-28 for RSV-
seronegative participants 
• 
SAE (Section 8.1.2) from Study Days 0-28 for RSV-seropositive and Study Days 0-56 for 
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Infectivity: 
• 
Infection with RSV as defined as  
1) vaccine virus identified in a nasal wash from Study Days 0-10 for RSV-
seropositive and Study Days 0-28 for RSV-seronegative (a binary outcome based 
on nasal washes done throughout the study period; Day 0 nasal wash will be 
counted as baseline); or  
2) ≥4-fold rise in RSV neutralizing antibody titer from Study Days 0-28 for RSV-
seropositive and Study Days 0-56 for RSV-seronegative subjects 
• 
Peak titer of vaccine virus shed from Study Days 0-10 for RSV-seropositive and Study Days 
0-28 for RSV-seronegative subjects 
• 
Duration of virus shedding in nasal washes as determined by a) culture and b) RT-PCR from 
Study Days 0-10 for RSV-seropositive and Study Days 0-28 for RSV-seronegative subjects 
 
Immunogenicity: 
• 
≥4-fold rise in RSV-neutralizing antibody titer from study entry to Study Day 28 for 
RSV-seropositive and from study entry to Study Day 56 for RSV-seronegative subjects 
• 
≥4-fold rise in IgG antibody responses to RSV F glycoprotein (ELISA) from study entry to 
Study Day 28 for RSV-seropositive and from study entry and Study Day 56 for RSV-
seronegative subjects 
 
9.2.2 Secondary Outcome Measures 
 
• 
The frequency and severity of symptomatic, medically attended respiratory and febrile illness 
in the RSV-seronegative (group 2) vaccine and placebo recipients who experience natural 
infection with wt RSV during the first RSV season and through the second RSV season for 
group 2 subjects who participate in a second RSV season surveillance. 
• 
The antibody responses in the RSV-seronegative vaccine and placebo recipients who 
experience natural infection with wt RSV during the first RSV season and during a second 
RSV season for those who choose to participate. 
• 
The quality and epitope specificity of RSF F-specific antibody 
 
 
SAMPLE SIZE AND ACCRUAL 
 
 
 
9.3.1 Sample Size and Randomization 
 
In group 1, approximately 15 RSV-seropositive children will be enrolled in the study and will 
receive either 105.7 PFU of vaccine or placebo, at a ratio of 2:1.  In group 2, approximately 21-30 
RSV-seronegative infants and children will be enrolled in the study and will receive either 105.0 
PFU of vaccine or placebo at a ratio of 2:1. If 30 seronegative participants are enrolled, we 
anticipate that 20 seronegative vaccinees and 10 seronegative placebo recipients will provide data 
for the primary objectives. The sample size was chosen based upon past experience with phase I 
evaluation of other live-attenuated respiratory virus candidate vaccines (24-26). The 2:1 
randomization ratio will be used to maximize the information obtained regarding the response of 
children to the RSV 6120/∆NS2/1030s vaccine. In the event that a participant is discontinued 
early from the study and the team decides that additional data would be needed to answer the 
study objectives, an additional participant may be enrolled in the same treatment arm as the 
discontinued participant. 
 

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Eligible subjects whose parents/guardians decline participation in the second RSV season 
surveillance will not be replaced. 
 
Given the small sample size, the study will have limitations with respect to detecting AEs and 
estimating the rates of such events in the population represented by the study sample. 
 
The following calculations focus on the assessment of the tolerability of the vaccine and, in 
particular, occurrence of LRI, which occurs very infrequently in children who participate in these 
types of studies but would be considered a sentinel safety event if observed in children infected 
with vaccine virus.   
 
Table 24 shows the probability of observing 0 events of LRI within the sample of 20 seronegative 
vaccinees, as well as the probability of observing 1 or more events or 2 or more events, under a 
range of assumptions concerning the true rate of such events in the participant population 
represented by this sample. From this table, it can be seen that if the true proportion of LRI (or 
other AE) is at least 10%, there is a 61% chance of observing 2 or more events in a group of size 
20, and an 88% chance of observing at least a single event. 
 
Table 24: The Probability of Observing LRI Events in Seronegative Vaccine 
Recipients 
 
N = 20 
True underlying probability 
of LRI or AEs 
Pr (0 
events) 
Pr (1+ 
events) 
Pr (2+ events) 
.01 
0.82 
0.18 
0.02 
.03 
0.54 
0.46 
0.12 
.05 
0.36 
0.64 
0.26 
.1 
0.12 
0.88 
0.61 
.15 
0.04 
0.96 
0.82 
 
Table 25 presents 90% confidence intervals (CIs) around potential rates of LRI or AEs that might 
be observed in the sample of 20 vaccine recipients. The CIs around similar rates in a sample of 10 
placebo recipients are also presented. Note that if no LRI or AEs are detected among the 20 
vaccine recipients, we are 90% confident that the true probability of AEs in the population from 
which the sample is drawn is between 0 and 14%. 
 
Table 25: Percent of Participants Experiencing LRI or AEs with Exact 90% CI 
N 
% LRI or AEs 
90% CI 
10 
0% 
0% -- 26% 
20 
0% 
0% -- 14% 
10 
10% 
1% -- 39% 
20 
10% 
2% -- 28% 
10 
20% 
4% -- 51% 
20 
20% 
7% -- 40% 
10 
30% 
9% -- 61% 
20 
30% 
14% -- 51% 
 

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Group sample sizes of 20 in the vaccinated group and 10 in the placebo group would achieve 
80% power to detect a difference between the group proportions of about 0.45. The test statistic 
used is the one-sided Fisher's exact test. The alpha level of the test was targeted at 0.05.   Table 
26 presents examples of true group differences which can be detected with 80% power, and 
Figure 5 graphically shows power curves for 90% power, 80% power, and 70% power given the 
sample sizes.  
 
Table 26: Magnitude of Difference in Responses Detectable with 80% Power 
Response Proportion in the 
Placebo Group 
Response Proportion in 
the Vaccinated Group 
Difference 
0.05 
0.52 
0.47 
0.1 
0.59 
0.49 
0.15 
0.65 
0.50 
0.2 
0.71 
0.51 
0.5 
0.95 
0.45 
 
 
 
Figure 5: Power Curves for Assessment of Vaccine Virus Shedding in RSV Seronegative 
Vaccinees 
 
 
   (Alpha=.05; N Vaccine=20; N Placebo=10; 1-Sided Exact Test) 
 
With a sample size of 20 seronegative vaccine recipients, the 90% CI around a sample mean peak 
titer of 2.5 log10, with a SD of 1.5 (from IMPAACT 2000) is (1.92, 3.08). This ensures with 90% 
Response proportion in the Vaccine Group 

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confidence that the true population mean peak titer is between 1.92 and 3.08 log10, and with 95% 
confidence that the true population mean is not lower than 1.92 log10. 
 
With the same sample size of 20, the 90% CI around a proportion of 18/20 (90%) vaccine 
recipients who shed vaccine virus is (72%-98%). For a proportion of 19/20 (95%) the 90% CI is 
(78%-99.7%), and for 20/20 (100%) the 90% CI is (86%-100%). For the target proportion of 
95%, this ensures with 95% confidence that the true proportion of vaccine recipients who shed 
vaccine virus is not lower than 78%. 
 
 
MONITORING 
 
9.4.1 Site Monitoring Plan 
 
As per International Conference on Harmonisation Good Clinical Practice (ICH-GCP) 5.18 and 
FDA 21 CFR 312.50, clinical protocols are required to be adequately monitored by the study 
sponsor. This study monitoring will be conducted according to the “NIAID Intramural Clinical 
Monitoring Guidelines.” Monitors under contract to the NIAID/OCRPRO will visit the clinical 
research site to monitor aspects of the study in accordance with the appropriate regulations and 
the approved protocol. The objectives of a monitoring visit will be: 1) to verify the existence of 
signed informed consent documents and documentation of the ICF process for each monitored 
subject; 2) to verify the prompt and accurate recording of all monitored data points, and prompt 
reporting of all SAEs; 3) to compare CRIMSON data abstracts with individual subjects’ records 
and source documents (subjects’ charts, laboratory analyses and test results, physicians’ progress 
notes, nurses’ notes, and any other relevant original subject information); and 4) to help ensure 
investigators are in compliance with the protocol. The monitors also will inspect the clinical site 
regulatory files to ensure that regulatory requirements (Office for Human Research Protections, 
OHRP), FDA, and applicable guidelines (ICH-GCP) are being followed. During the monitoring 
visits, the investigator (and/or designee) and other study personnel will be available to discuss the 
study progress and monitoring visit. 
 
The investigator (and/or designee) will make study documents (e.g., consent forms, CRIMSON 
data abstracts) and pertinent hospital or clinical records readily available for inspection by the 
local IRB, the FDA, the site monitors, and the NIAID staff for confirmation of the study data.  
 
A specific protocol monitoring plan will be discussed with the PI and study staff prior to 
enrollment. The plan will outline the frequency of monitoring visits based on such factors as 
study enrollment, data collection status and regulatory obligations. 
 
9.4.2 Monitoring by the NIAID Intramural Data and Safety Monitoring Board 
 
The NIAID Intramural DSMB is constituted to review the safety data of Intramural NIAID 
clinical studies that require DSMB oversight. The NIAID Intramural DSMB includes 
independent experts in infectious diseases, biostatistics, and clinical research that do not have 
direct involvement in the conduct of the study and have no significant conflicts of interests as 
defined by NIAID policy. The DSMB will review the protocol prior to opening the study to 
enrollment. The DSMB will meet at least twice a year or on a schedule specified by the DSMB to 
review the completeness of the study data, the adherence to the protocol, and AE data. 
Cumulative safety data (pooled across arms, with the vaccine and placebo arms presented 
together) will be submitted to the DSMB Executive Secretary for DSMB review. The DSMB 
Executive Secretary will provide the PI with DSMB recommendations promptly, and the official 
DSMB Report will then be provided in a timely fashion through the office of the NIAID Clinical 

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Director. The PI will submit the written DSMB recommendations to the IRB upon receipt. All 
SAEs, LRIs, UPs and IND safety reports will be reported by the PI to the DSMB at the same 
time that they are submitted to the IRB and/or regulatory sponsor (OCRPRO). The PI will notify 
the DSMB of any cases of intentional or unintentional unblinding as soon as possible. The PI will 
notify the Board at the time a pausing or halting criteria are met and obtain a recommendation 
concerning continuation, modification, or termination of the study.   
 
 
ANALYSES 
 
9.5.1 Assessment of Primary Objectives  
 
Safety data from all participants in CIR 322 who have been inoculated will be summarized 
together, including data from participants who discontinue study early or have some missed 
visits. In the immunogenicity analyses, those who do not provide data for the Day 28 Visit 
follow-up for RSV-seropositive participants and for the Day 56 Visit follow-up for 
RSV-seronegative participants (due to early discontinuation or missed visit) will be treated as 
“failures.” Sensitivity analyses will be performed to check if the results are consistent with those 
when these participants are excluded. Participants who receive any of the disallowed treatments 
listed in Section 5.10 during the first 28 days after inoculation for RSV-seropositive participants 
and during the first 56 days after inoculation for RSV-seronegative participants may be excluded 
from the immunogenicity evaluations after the time of the treatment. These participants, however, 
will be included in the safety evaluations for the duration of the study. These participants will not 
be replaced. Details of the analyses listed below will be included in the statistical analysis plan.  
 
The frequency of solicited AEs and unsolicited AEs, along with 90% confidence intervals, during 
Study Days 0 to 10 in RSV-seropositive subjects and during Study Days 0 to 28 in RSV-
seronegative subjects and of vaccine-related SAE during Study Days 0 to 28 in RSV-seropositive 
subjects and during Study Days 0 to 56 in RSV-seronegative subjects will be summarized. In 
addition, line listing of individual clinical solicited AEs and unsolicited AEs, graded by severity, 
will be prepared for events occuring during Study Days 0 to 10 in RSV-seropositive subjects and 
during Study Days 0 to 28 in RSV-seronegative subjects and vaccine-related SAE during Study 
Days 0 to 28 in RSV-seropositive subjects and during Study Days 0 to 56 in RSV-seronegative 
subjects. 
 
The proportion of participants with infection defined as recovery of vaccine virus from a nasal 
wash as determined by culture and RT-PCR, and/or a >4-fold rise in neutralizing antibody titer to 
RSV, will be summarized. A line listing of the individual peak titer of vaccine virus shed and 
duration of virus shedding in nasal washes by each individual will be prepared. In addition, the 
geometric mean peak titer and mean duration of virus shed will be provided for each treatment 
group.  
 
The proportion of participants that develop 4-fold or greater rises in RSV-neutralizing antibody 
titer following vaccination will be summarized. A line listing of the individual RSV antibody titer 
pre- and post-vaccination will be prepared. In addition, the geometric mean and median antibody 
titers will be provided for each treatment group. Line listings of individual RSV-neutralizing 
antibody responses as well as of antibody responses to the RSV F glycoprotein will be prepared 
as well.  
 
Where appropriate, a 1-tailed Wilcoxon rank sum test will be used to test the hypothesis that the 
vaccinated group will exhibit greater peak viral titers and antibody titers following vaccination 
compared to the placebo group. A 1-tailed Fisher’s exact test will be used to test the hypothesis 

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that the vaccinated group will exhibit a greater proportion of participants who develop fourfold or 
greater rises in RSV-neutralizing antibody titer following vaccination compared to the placebo 
group.  
 
These will be the only formal statistical comparisons between the vaccinated and placebo groups. 
These tests will be carried out at a 5% significance level. 
 
The study results will be compared with the criteria listed in Section 1.1 to determine if this 
vaccine is a promising candidate for further evaluation in expanded phase I studies or phase II 
studies. 
 
9.5.2 Assessment of Secondary Objectives  
 
The summary of the frequency and severity of symptomatic, medically attended respiratory and 
febrile illness in the RSV-seronegative vaccine and placebo recipients who experience natural 
infection with wt RSV during the first RSV season will be presented. A summary will also be 
presented of the frequency and severity of symptomatic, medically attended respiratory and 
febrile illness in the RSV-seronegative vaccine and placebo recipients who participate in a second 
RSV season surveillance and who experience natural infection with wt RSV during the second 
RSV season. In addition, a line listing of the individual RSV antibody titer pre- and post-First 
RSV Season Surveillance Period will be prepared, as well as a listing of the RSV antibody titer 
pre- and post-Second RSV Season Surveillance Period for those seronegative subjects who 
participate in the second RSV season surveillance. In addition, the geometric mean and median 
antibody titers will be provided, for each treatment group. The adaptive immune responses to 
vaccine will be summarized for each treatment group. A line listing of the mucosal antibody 
response detected in nasal wash specimens will be prepared. 
 
 
DATA HANDLING AND RECORD KEEPING 
 
 DATA MANAGEMENT RESPONSIBILITIES 
 
The CIR will maintain adequate and accurate research records containing all information 
pertinent to the study for all screened and enrolled participants, including CRFs and supporting 
source data per the SOP.  
 
Data from source documentation for subjects enrolled in the study will be entered into the study 
data system. The data entry is to be completed on an ongoing basis during the study. Data entry 
shall be performed by authorized individuals who have a unique log-on and password. 
Corrections to the data system shall be tracked electronically (password protected) with time, 
date, individual making the correction, and what was changed.  
 
Source documents include all recordings of observations or notations of clinical activities, and all 
reports and records necessary for the evaluation and reconstruction of the clinical trial. Source 
documents may include the subject’s medical records and laboratory reports. Data will be 
collected directly from subjects during study visits and contacts.   
 

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 ESSENTIAL AND SOURCE DOCUMENTS AND ACCESS TO SOURCE DATA 
 
Study-related documentation will be completed as required by the IRB, the sponsor, and 
regulatory authorities. Continuing review documentation will be submitted to the IRB as 
specified by the IRB. An annual report will be submitted by the sponsor to the FDA based on the 
anniversary date that the IND for the RSV 6120/∆NS2/1030s vaccine went into effect. These 
reports will provide a brief description of the progress of the investigation as outlined in 21 CFR 
312.33 and will include any revisions of the protocol not previously submitted to the FDA. 
 
All study records must be accessible for inspection, monitoring, and/or auditing during and after 
the conduct of the study by authorized representatives of the study sponsors and their contracted 
monitors, the FDA, regulatory authorities, IRB, OHRP, and other applicable regulatory entities. 
Records must be kept on-site throughout the period of study implementation; thereafter, 
instructions for off-site storage may be provided by NIH. No study records may be removed to an 
off-site location or destroyed prior to receiving approval from NIH.  
 
Study-related documents will be maintained for a period of at least 2 years after final marketing 
approval of the vaccine, or at least 2 years after the formal discontinuation of clinical 
development of the product (or longer based upon local laws). The sponsor is required to inform 
the PI as to when such documents need no longer be retained. No study document should be 
destroyed without prior written agreement between the sponsor and the PI. Storage of all study-
related documents will be such that confidentiality will be strictly maintained. These records are 
also to be maintained in compliance with IRB, state, and federal medical records retention 
requirements, whichever are longest. Should the PI wish to assign the study records to another 
party and/or move them to another location, the PI must provide written notification of such 
intent to the sponsor with the name of the person who will accept responsibility for the transferred 
records and/or their new location. The sponsor must be notified in writing, and written permission 
must be received from the sponsor prior to destruction or relocation of research records. 
 
 CLINICAL INVESTIGATOR’S BROCHURE 
 
The current version of the IB comprehensively describes all the available preclinical experience 
with the experimental vaccine. If relevant new information becomes available during the course 
of the trial, the PI will receive a revised IB or an amendment to the current version. 
 
 QUALITY CONTROL AND QUALITY ASSURANCE 
 
Essential documents and participant research records are subject to continuous quality control and 
quality assurance procedures consistent with the MOP. 
 
 
CLINICAL SITE MONITORING 
 
Site monitors under contract to NIAID will inspect study facilities and review participant study 
records including ICD, CRFs, medical records, laboratory records, and study product records, to 
ensure protection of study participants, compliance with the IRB approved protocol, and accuracy 
and completeness of records. The monitors also will review essential document files to ensure 
compliance with all applicable regulatory requirements. Study staff will make study facilities and 
documents available for inspection by the monitors.   
 
The sponsor will retain originals of the Form FDA 1572 and copies of other study documents as 
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Statement of Compliance  
 
The trial will be conducted in compliance with this protocol, ICH-GCP guidelines, FDA 
guidelines and applicable regulatory requirements. The CIR monitoring will be conducted 
according to the OCRPRO Clinical Trial Management Group’s Monitoring Plan. 
 
 
HUMAN SUBJECTS PROTECTIONS 
 
 INSTITUTIONAL REVIEW BOARD/ETHICS COMMITTEE REVIEW AND APPROVAL  
 
Prior to study initiation, the PI will obtain IRB review and approval of this protocol and ICDs in 
accordance with 45 CRF 46.   In addition to the initial review and approval, the IRB must review 
the study at least annually. The PI must also promptly report to the IRB any changes in the study 
and any UPs involving risks to participants or others. 
 
All IRB policies and procedures must be followed, and complete documentation of all 
correspondence to and from the IRBs must be maintained in the essential document files. 
 
A copy of the study approval (including approval of the ICD) is to be maintained in the 
investigator’s study document binder, and a copy will be supplied to the sponsor.  
 
During the study, the PI is responsible for providing the IRB with all documents subject to review 
(i.e., protocol amendments, ICD updates, advertisements, and any written information that may 
be provided to the participant’s parents/guardians). Study progress reports will be made to the 
IRB by the investigator in accordance with IRB guidelines and government regulations. 
 
 VULNERABLE PARTICIPANTS 
 
The NIH is mandated by law to ensure that children be included in clinical research when 
appropriate (40, 41). This study responds to that mandate and will provide clinical research data 
to inform RSV vaccine infectivity, safety and immunogenicity in children. Nonetheless, the 
infants who take part in this study are considered vulnerable participants per the US CFR, and site 
IRBs/IBCs must consider the potential risks and benefits to child participants as described in 45 
CFR 46 Subpart D (for children). 
 
 INFORMED CONSENT 
 
In obtaining and documenting informed consent, the PI must comply with the applicable 
regulatory requirements, ICH-GCP guidelines, and ethical principles. The written ICD must be 
approved by the IRB prior to its use. 
 
Written informed consent for study participation will be obtained before any study-specific 
procedures are performed. The informed consent process will include information exchange, 
detailed discussion, and assessment of understanding of all required elements of informed 
consent, including the potential risks, benefits, and alternatives to study participation. 
  
As part of the informed consent process, parents/guardians will also be asked whether they agree 
to storage and future research testing of the biological specimens that remaining after all protocol-
specified testing has been completed. Future research testing of residual specimens may be 
declined with no impact on other aspects of study participation.   

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 POTENTIAL BENEFITS 
 
Participants may not receive any direct vaccine-related benefit from enrollment in this study. Some 
children who receive vaccine may be protected against infections with wt RSV that circulates in 
the community. It is hoped that information gained in this study will contribute to the development 
of a safe and effective vaccine for the prevention of illness associated with RSV infection.  
 
Parents/guardian may be offered child safety seat educational material and referral to community 
inspection stations by study staff, and may be offered certified lactation counseling services, if 
appropriate. 
 
 POTENTIAL RISKS  
 
12.5.1 Venipuncture 
 
Risks occasionally associated with venipuncture include pain and bruising at the site of 
venipuncture, lightheadedness, infection, and rarely syncope. Before each blood draw, we will 
offer to use anesthetic skin cream to decrease the pain. 
 
12.5.2 Nasal Wash 
 
Risks occasionally associated with nasal wash include pain or discomfort, and occasionally 
epistaxis. Nasal washes are not standard care in well children and are not usually performed on ill 
children, although many parents are advised to use over-the-counter saline solutions and/or nasal 
bulb suction to clear a young child’s congested nostrils during an URI. The nasal bulb suction and 
saline-like solutions are 2 components of our nasal wash procedure. 
 
12.5.3 Nasosorption SAM Strips 
 
The risk associated with the Nasosorption SAM Strips is a nasolacrimal reflex which involves 
slight watering of the eyes. 
 
12.5.4 Topical Anesthetic Cream 
 
Risks occasionally associated with the use of topical anesthetic cream include temporary skin 
discoloration, skin irritation, rash, hives, and rarely, dizziness or drowsiness. 
 
12.5.5 Receipt of Study Product 
 
If the study vaccine is insufficiently attenuated, participants could experience rhinorrhea, cough, 
fever, otitis media, or LRI. Immediate hypersensitivity reactions including urticaria, anaphylaxis, 
or other immunoglobulin E (IgE)-mediated responses are possible, as with any vaccine. With any 
investigational vaccine, there is a theoretical possibility of risks about which we have no present 
knowledge. Parents/guardians will be informed of any such risks should further data become 
available. 
 
The study participant’s family does not pay for the study product or research visits including 
examinations and laboratory tests that are part of this study, including evaluation of illness, if any. 
NIAID agrees that the funding for appropriate acute care will be provided through the NIAID 

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contract (HHS272200900010C) for any side effects that are determined to be related to the 
administration of vaccine.  
 
 REIMBURSEMENT/COMPENSATION 
 
12.6.1 Study Compensation: Acute Phase and First Season Surveillance 
 
Compensation will be in the form of either check or gift card. Participants’ parent/guardian will 
receive compensation at the rate of $50 for the inoculation visit, $30 per each scheduled or 
unscheduled study visit, $5 per each scheduled non-visit day report and weekly surveillance 
report, and a $50 bonus for study completion. In addition, the parent/guardian will receive bus 
tokens, taxi fare, or parking passes as needed for study visits. Parent/guardian will be compensated 
only for those portions of the study that are completed. Compensation will be in accordance with 
IRB policies and procedures and will be subject to IRB approval. 
 
Participants may receive age-appropriate books or small toys. The total value of the books or toys 
will not exceed $10 per participant.  
 
12.6.2 Study Compensation: Second Season Surveillance 
 
Subjects’ parents/guardians will receive compensation at the rate of $50 for the enrollment visit, 
$30 per scheduled or unscheduled study visit, $5 per weekly surveillance report and a $50 bonus 
for study completion. 
 
 PRIVACY AND CONFIDENTIALITY 
 
All study procedures will be conducted in private and every effort will be made to protect 
participant privacy and confidentiality to the extent possible. Participant information will not be 
released without written permission to do so except as necessary for review, monitoring, and/or 
auditing as described in Section 9.4 and Section 10.2. 
All study-related information will be stored securely. Participant research records will be stored 
in locked areas with access limited to study staff. All laboratory specimens, CRFs, and other 
documents that may be transmitted off-site will be identified by coded number only.  
 
All local databases must be secured with password protected access systems. Lists, logbooks, 
appointment books, and any other documents that link subject identification numbers to personal 
identifying information should be stored in a separate, locked location in an area with limited 
access.   
 
 MANAGEMENT OF INCIDENTAL FINDINGS 
 
Study clinicians will inform parents/guardians of all clinically meaningful physical exam findings 
and laboratory tests. When applicable, study clinicians will provide referrals to non-study sources 
of medical care for further evaluation and/or treatment of these findings.  
 
 
ADMINISTRATIVE PROCEDURES 
 
 REGULATORY OVERSIGHT 
 
CIR 322 is sponsored by the OCRPRO, Division of Clinical Research, NIAID, NIH. 

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OCRPRO is responsible for regulatory oversight of this study. Safety-related information 
pertaining to the study product will be distributed prior to and during the conduct of the study, in 
accordance with its sponsor obligations.  
 
NIAID provides funding to the clinical research sites at which this study will be conducted. Each 
institute contracts with independent clinical site monitors who will perform monitoring visits as 
described in Section 9.4. As part of these visits, monitors will inspect study-related 
documentation to ensure compliance with all applicable local and US regulatory requirements. 
 
 PROTOCOL REGISTRATION 
 
Prior to implementation of this protocol, and any subsequent full version amendments, the CIR 
must have the protocol and the protocol ICDs approved, as appropriate, by their local IRB/IBC, 
local IBC, and any other applicable regulatory entity.  
 
For any future protocol amendments, upon receiving final IRB/IBC and any other applicable 
regulatory entity approvals, CIR should implement the amendment immediately.  
 
 STUDY IMPLEMENTATION 
 
This study will be conducted in accordance with the protocol, ICH guidelines, and all applicable 
local and US regulations. Study implementation will also be guided by the study-specific MOP 
and other study implementation materials. 
 
 PROTOCOL DEVIATION REPORTING 
 
Protocol deviations must be documented in participant research records. Reasons for the 
deviations and corrective and preventive actions taken in response to the deviations should also 
be documented. See MOP for further instructions. 
 
Deviations will be reported to the IRB and other applicable review bodies in accordance with the 
policies and procedures of these review bodies. Serious deviations that are associated with 
increased risk to one or more study participants and/or significant impacts on the integrity of 
study data must also be reported following procedures specified in the MOP.  
 
 CLINICALTRIALS.GOV 
 
This protocol is subject to the US Food and Drug Administration Amendments Act of 2007 
(FDAAA), including registration in ClinicalTrials.gov. 
 
 
PUBLICATIONS 
 
Publication of the results of this trial will be governed by NIAID policies. Any presentation, 
abstract, or manuscript will be made available for review by the pharmaceutical and NIAID 
sponsors prior to submission. Publication or presentation approval will conform to any 
Cooperative Research and Development Agreement (CRADA) or other collaborative agreement 
in place. 
 
 

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6 April 2020 
 
 
REFERENCES 
 
1. 
Hall CB, Weinberg GA, Iwane MK, Blumkin AK, Edwards KM, Staat MA, 
Auinger P, Griffin MR, Poehling KA, Erdman D, Grijalva CG, Zhu Y, Szilagyi 
P. 2009. The burden of respiratory syncytial virus infection in young children. N 
Engl J Med 360:588-598. 
2. 
Nair H, Nokes DJ, Gessner BD, Dherani M, Madhi SA, Singleton RJ, O'Brien 
KL, Roca A, Wright PF, Bruce N, Chandran A, Theodoratou E, Sutanto A, 
Sedyaningsih ER, Ngama M, Munywoki PK, Kartasasmita C, Simoes EA, 
Rudan I, Weber MW, Campbell H. 2010. Global burden of acute lower 
respiratory infections due to respiratory syncytial virus in young children: a 
systematic review and meta-analysis. Lancet 375:1545-1555. 
3. 
Hall CB. 2001. Respiratory syncytial virus and parainfluenza virus. N Engl J Med 
344:1917-1928. 
4. 
Martinez FD. 2003. Respiratory syncytial virus bronchiolitis and the 
pathogenesis of childhood asthma. PediatrInfect DisJ 22:S76-S82. 
5. 
Collins PL, Karron RA. 2013. Respiratory syncytial virus and metapneumovirus, 
p 1086-1123. In Knipe DM, Howley PM, Cohen JI, Griffin DE, Lamb RA, Martin 
MA, Racaniello VR, Roizman B (ed), Fields Virology, Sixth Edition, Sixth ed, vol 
1. Lippincott Williams & Wilkins, Philadelphia. 
6. 
Kim HW, Canchola JG, Brandt CD, Pyles G, Chanock RM, Jensen K, Parrott 
RH. 1969. Respiratory syncytial virus disease in infants despite prior 
administration of antigenic inactivated vaccine. Am J Epidemiol 89:422-434. 
7. 
Collins PL, Murphy BR. 2007. Vaccines against Human Respiratory Syncytial 
Virus. Respiratory Syncytial Virus 14:233-278. 
8. 
Acosta PL, Caballero MT, Polack FP. 2015. Brief history and characterization 
of enhanced respiratory syncytial virus disease. Clinical and Vaccine 
Immunology 23:189-195. 
9. 
Collins PL, Murphy BR. 2005. New generation live vaccines against human 
respiratory syncytial virus designed by reverse genetics. ProcAm ThoracSoc 
2:166-173. 
10. 
Wright PF, Karron RA, Belshe RB, Shi JR, Randolph VB, Collins PL, O'Shea 
AF, Gruber WC, Murphy BR. 2007. The absence of enhanced disease with wild 
type respiratory syncytial virus infection occurring after receipt of live, attenuated, 
respiratory syncytial virus vaccines. Vaccine 25:7372-7378. 
11. 
Karron RA, Wright PF, Belshe RB, Thumar B, Casey R, Newman F, Polack 
FP, Randolph VB, Deatly A, Hackell J, Gruber W, Murphy BR, Collins PL. 
2005. Identification of a recombinant live attenuated respiratory syncytial virus 
vaccine candidate that is highly attenuated in infants. J Infect Dis 191:1093-1104. 
12. 
Hendry RM, Burns JC, Walsh EE, Graham BS, Wright PF, Hemming VG, 
Rodriguez WJ, Kim HW, Prince GA, McIntosh K. 1988. Strain-specific serum 
antibody responses in infants undergoing primary infection with respiratory 
syncytial virus. J Infect Dis 157:640-647. 
13. 
Johnson PR, Jr., Olmsted RA, Prince GA, Murphy BR, Alling DW, Walsh EE, 
Collins PL. 1987. Antigenic relatedness between glycoproteins of human 
respiratory syncytial virus subgroups A and B: evaluation of the contributions of F 
and G glycoproteins to immunity. J Virol 61:3163-3166. 

74 
RSV 6120/∆NS2/1030s, Lot#012A  
           
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14. 
Collins PL, Hill MG, Camargo E, Grosfeld H, Chanock RM, Murphy BR. 1995. 
Production of infectious human respiratory syncytial virus from cloned cDNA 
confirms an essential role for the transcription elongation factor from the 5' 
proximal open reading frame of the M2 mRNA in gene expression and provides a 
capability for vaccine development. ProcNatlAcadSciUSA 92:11563-11567. 
15. 
Lo MS, Brazas RM, Holtzman MJ. 2005. Respiratory syncytial virus 
nonstructural proteins NS1 and NS2 mediate inhibition of Stat2 expression and 
alpha/beta interferon responsiveness. J Virol 79:9315-9319. 
16. 
Ramaswamy M, Shi L, Varga SM, Barik S, Behlke MA, Look DC. 2006. 
Respiratory syncytial virus nonstructural protein 2 specifically inhibits type I 
interferon signal transduction. Virology 344:328-339. 
17. 
Spann KM, Tran KC, Chi B, Rabin RL, Collins PL. 2004. Suppression of the 
induction of alpha, beta, and lambda interferons by the NS1 and NS2 proteins of 
human respiratory syncytial virus in human epithelial cells and macrophages 
[corrected]. J Virol 78:4363-4369. 
18. 
Spann KM, Tran KC, Collins PL. 2005. Effects of nonstructural proteins NS1 
and NS2 of human respiratory syncytial virus on interferon regulatory factor 3, 
NF-kappaB, and proinflammatory cytokines. J Virol 79:5353-5362. 
19. 
Swedan S, Andrews J, Majumdar T, Musiyenko A, Barik S. 2011. Multiple 
functional domains and complexes of the two nonstructural proteins of human 
respiratory syncytial virus contribute to interferon suppression and cellular 
location. J Virol 85:10090-10100. 
20. 
Swedan S, Musiyenko A, Barik S. 2009. Respiratory syncytial virus 
nonstructural proteins decrease levels of multiple members of the cellular 
interferon pathways. J Virol 83:9682-9693. 
21. 
Teng MN, Collins PL. 1999. Altered growth characteristics of recombinant 
respiratory syncytial viruses which do not produce NS2 protein. J Virol 73:466-
473. 
22. 
Luongo C, Winter CC, Collins PL, Buchholz UJ. 2012. Increased genetic and 
phenotypic stability of a promising live-attenuated respiratory syncytial virus 
vaccine candidate by reverse genetics. J Virol 86:10792-10804. 
23. 
Karron RA, Wright PF, Crowe JE, Jr., Clements-Mann ML, Thompson J, 
Makhene M, Casey R, Murphy BR. 1997. Evaluation of two live, cold-passaged, 
temperature-sensitive respiratory syncytial virus vaccines in chimpanzees and in 
human adults, infants, and children. J Infect Dis 176:1428-1436. 
24. 
Wright PF, Karron RA, Belshe RB, Thompson J, Crowe JE, Jr., Boyce TG, 
Halburnt LL, Reed GW, Whitehead SS, Anderson EL, Wittek AE, Casey R, 
Eichelberger M, Thumar B, Randolph VB, Udem SA, Chanock RM, Murphy 
BR. 2000. Evaluation of a live, cold-passaged, temperature-sensitive, respiratory 
syncytial virus vaccine candidate in infancy. J Infect Dis 182:1331-1342. 
25. 
Wright PF, Karron RA, Madhi SA, Treanor JJ, King JC, O'Shea A, Ikizler MR, 
Zhu Y, Collins PL, Cutland C, Randolph VB, Deatly AM, Hackell JG, Gruber 
WC, Murphy BR. 2006. The interferon antagonist NS2 protein of respiratory 
syncytial virus is an important virulence determinant for humans. J Infect Dis 
193:573-581. 
26. 
Karron RA, Luongo C, Thumar B, Loehr KM, Englund JA, Collins PL, 
Buchholz UJ. 2015. A gene deletion that up-regulates viral gene expression 

75 
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6 April 2020 
 
yields an attenuated RSV vaccine with improved antibody responses in children. 
Sci Transl Med 7:312ra175. 
27. 
Wright PF, Karron RA, Madhi SA, Treanor JJ, King JC, O'Shea A, Ikizler MR, 
Zhu Y, Collins PL, Cutland C, Randolph VB, Deatly AM, Hackell JG, Gruber 
WC, Murphy BR. 2006. The interferon antagonist NS2 protein of respiratory 
syncytial virus is an important virulence determinant for humans. J Infect Dis 
193:573-581. 
28. 
Crowe JE, Jr., Bui PT, London WT, Davis AR, Hung PP, Chanock RM, 
Murphy BR. 1994. Satisfactorily attenuated and protective mutants derived from 
a partially attenuated cold-passaged respiratory syncytial virus mutant by 
introduction of additional attenuating mutations during chemical mutagenesis. 
Vaccine 12:691-699. 
29. 
Whitehead SS, Juhasz K, Firestone CY, Collins PL, Murphy BR. 1998. 
Recombinant respiratory syncytial virus (RSV) bearing a set of mutations from 
cold-passaged RSV is attenuated in chimpanzees. J Virol 72:4467-4471. 
30. 
Crowe JE, Jr., Bui PT, Davis AR, Chanock RM, Murphy BR. 1994. A further 
attenuated derivative of a cold-passaged temperature-sensitive mutant of human 
respiratory syncytial virus retains immunogenicity and protective efficacy against 
wild-type challenge in seronegative chimpanzees. Vaccine 12:783-790. 
31. 
Crowe JE, Jr., Firestone CY, Whitehead SS, Collins PL, Murphy BR. 1996. 
Acquisition of the ts phenotype by a chemically mutagenized cold-passaged 
human respiratory syncytial virus vaccine candidate results from the acquisition 
of a single mutation in the polymerase (L) gene. Virus Genes 13:269-273. 
32. 
Whitehead SS, Firestone CY, Collins PL, Murphy BR. 1998. A single 
nucleotide substitution in the transcription start signal of the M2 gene of 
respiratory syncytial virus vaccine candidate cpts248/404 is the major 
determinant of the temperature-sensitive and attenuation phenotypes. Virology 
247:232-239. 
33. 
Juhasz K, Whitehead SS, Boulanger CA, Firestone CY, Collins PL, Murphy 
BR. 1999. The two amino acid substitutions in the L protein of cpts530/1009, a 
live-attenuated respiratory syncytial virus candidate vaccine, are independent 
temperature-sensitive and attenuation mutations. Vaccine 17:1416-1424. 
34. 
Juhasz K, Whitehead SS, Bui PT, Biggs JM, Crowe JE, Boulanger CA, 
Collins PL, Murphy BR. 1997. The temperature-sensitive (ts) phenotype of a 
cold-passaged (cp) live attenuated respiratory syncytial virus vaccine candidate, 
designated cpts530, results from a single amino acid substitution in the L protein. 
J Virol 71:5814-5819. 
35. 
McFarland EJ, Buchholz UJ, Muresan P, Cunningham CK, Luongo C, 
Thumar B, Collins P, Perlowski C, Valentine M, Schappell E, 
Gnanashanmugam D, Siberry G, Rexroad V, Barr E, Harding P, DiPerna A, 
Karron R. High Infectivity of recombinant live-attenuated respiratory syncytial 
virus vaccine RSV LID ΔM2-2 in RSV-seronegativeinfants and children, 
manuscript in preparation.   
36. 
Anonymous. 2007. American Academy of Pediatrics, Committee on Practice 
and Ambulatory Medicine. Recommendations for Preventive Pediatric Health 
Care. Pediatrics 120:1376. 

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37. 
Siberry GK, ener-West M, Schappell E, Karron RA. 2002. Comparison of 
temple temperatures with rectal temperatures in children under two years of age. 
ClinPediatr(Phila) 41:405-414. 
38. 
Whitehead SS, Juhasz K, Firestone CY, Collins PL, Murphy BR. 1998. 
Recombinant respiratory syncytial virus (RSV) bearing a set of mutations from 
cold-passaged RSV is attenuated in chimpanzees. J Virol 72:4467-4471. 
39. 
Gharpure MA, Wright PF, Chanock RM. 1969. Temperature-sensitive mutants 
of respiratory syncytial virus. J Virol 3:414-421. 
40. 
National Institutes of Health. 1994. Guidelines for the Inclusion of Women and 
Minorities as Subjects in Clinical Research. Federal Register.,  
41. 
National Institutes of Health. 1998. NIH Policy and Guidelines on the Inclusion 
of Children as Participants in Research Involving Human Subjects.  
 
 

 
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APPENDICES 
 
APPENDIX I: TABLES REFERENCED IN THE BACKGROUND SECTION 
 
Table 27: Viral Titers of Nasopharyngeal Swab Samples from AGMs Inoculated with the 
Experimental Lot or CTM of RSV 6120/ΔNS2/1030s, with RSV ΔNS2/Δ1313/I1314L, or 
with Recombinant wt RSV rA2 
Virus Test 
Articlea 
and Study 
Number 
AGM  
ID 
NP virus titer (log10 PFU/mL) on indicated daysb 
Peak 
virus  
titer 
Sum 
of 
daily 
titers c 
1 
2 
3 
4 
5 
6 
7 
8 
9 
10 12/ 
14 
RSV rA2 
CTM 
RSV#004Ad 
7209 
- 
- 
3.1 
4.4 
4.4 
5.2 
3.6 
4.1 
3.4 
2.0 
- 
5.2 
31.3 
7467 
- 
0.7 
2.8 
3.8 
2.1 
3.9 
3.4 
3.2 
1.9 
2.0 
- 
3.9 
24.5 
7468 
- 
1.7 
2.6 
4.0 
3.9 
3.7 
3.6 
2.7 
3.6 
2.6 
- 
4.0 
29.1 
7492 
0.7 
1.2 
2.3 
3.9 
2.9 
3.9 
3.9 
2.4 
1.7 
1.0 
- 
3.9 
24.3 
 
Mean: 
 
 
 
 
 
 
 
 
 
 
 
4.2 
27.2 
RSV 
ΔNS2/Δ1313/ 
I1314L 
CTM 
RSV#006Ae 
7648 
- 
- 
1.7 
1.2 
1.8 
2.9 
2.2 
4.0 
3.7 
3.4 
- 
4.0 
22.0 
7692 
- 
- 
- 
- 
1.9 
- 
- 
3.3 
1.0 
1.5 
- 
3.3 
10.2 
7714 
- 
- 
0.7 
- 
1.3 
- 
- 
1.4 
2.4 
2.8 
- 
2.8 
10.7 
7764 
- 
- 
- 
0.7 
0.7 
1.0 
- 
2.3 
1.2 
2.9 
- 
2.9 
10.5 
 
Mean: 
 
 
 
 
 
 
 
 
 
 
 
3.2 
13.3 
6120/ΔNS2/ 
1030s 
Exp. Lot 
8494 
- 
- 
2.4 
2.4 
2.8 
3.7 
4.1 
4.8 >3.3 
3.0 
- 
4.8 
28.0 
8542 
- 
- 
1.4 
2.0 
2.0 
1.8 
2.9 
2.9 
3.3 
1.4 
- 
3.0 
18.5 
8554 
0.7 
- 
- 
1.5 
2.0 
2.1 
4.1 
1.3 
3.0 
3.6 
- 
4.1 
19.3 
8562 
- 
- 
- 
2.4 
3.2 
2.9 
2.3 
1.8 
3.2 
3.4 
- 
3.4 
20.7 
 
Mean: 
 
 
 
 
 
 
 
 
 
 
 
3.8 
21.6 
6120/ΔNS2/ 
1030s 
CTM 
RSV#012Af 
8928 
- 
1.7 
3.3 
3.9 
4.1 
4.9 
3.4 
3.6 
3.1 
1.0 0.7 
4.9 
30.0 
8940 
- 
- 
- 
1.7 
1.5 
3.4 
0.7 
4.3 
4.0 
3.4 2.2 
4.3 
22.3 
8983 
- 
- 
- 
0.7 
1.5 
2.9 
2.7 
0.7 
2.4 
1.3 2.7 
2.9 
15.9 
9045 
- 
0.7 
0.7 
1.5 
1.3 
1.8 
3.0 
2.6 
1.3 
- 
0.7 
3.0 
13.8 
 
Mean: 
 
 
 
 
 
 
 
 
 
 
 
3.8 
20.5 
a Monkeys were inoculated i.n. and i.t. with 106 PFU of the indicated virus in a 1 mL inoculum per site (total dose = 2x106 
PFU/AGM). 
b Virus titrations were performed on Vero cells. The lower limit of detection was 0.7 log10 PFU/mL. Samples with no detectable 
virus are represented as “-“. Peak titers for each animal are underlined. 
c The sum of daily titers is used as an estimate for the magnitude of shedding (area under the curve). A value of 0.35 was used for 
samples with no detectable virus.  
d Lot RSV#004A, vial numbers 263, 886, 1108, 1567. 
e Lot RSV#006A, vial numbers 0004, 1377, 2505. 
f Lot RSV#012A, vial numbers 0019-0021, 1267-1269, 2483-2486. 
Abbreviations: AGM- African green monkey, CTM- clinical trial material, RSV- respiratory syncytial virus, NP- nasopharyngeal, 
PFU- plaque-forming units, i.n.- intranasally, i.t.- intratracheally. 
 
 

 
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Table 28: Viral Titer of Tracheal Lavage Samples from AGMs Inoculated with the 
Experimental Lot or CTM of RSV 6120/ΔNS2/1030s, with RSV ΔNS2/Δ1313/I1314L, or 
with Recombinant wt RSV rA2 
Virus Test Articlea 
and Study Number 
AGM ID 
TL Virus Titer (log10 PFU/mL) on 
Indicated Dayb 
Peak 
Virus 
Titer 
Sum of 
Daily  
Titers c 
2 
4 
6 
8 
10 
12/14 
 
 
RSV rA2 
CTM 
RSV#004Ad 
7209 
3.0 
3.8 
4.3 
4.5 
2.6 
- 
4.5 
18.9 
7467 
3.0 
3.0 
4.3 
3.4 
2.8 
- 
4.3 
17.2 
7468 
2.5 
1.7 
2.9 
3.0 
2.9 
- 
3.0 
13.7 
7492 
1.9 
3.5 
4.7 
3.3 
- 
- 
4.7 
14.8 
Mean: 
 
 
 
 
 
 
4.1 
16.2 
RSV ΔNS2/Δ1313/ 
I1314L 
CTM RSV#006Ae 
7648 
- 
- 
- 
1.5 
- 
- 
1.5 
5.0 
7692 
- 
- 
- 
1.0 
1.7 
- 
1.7 
5.5 
7714 
- 
- 
- 
- 
- 
- 
- 
4.2 
7764 
- 
- 
- 
1.3 
- 
- 
1.3 
4.8 
Mean: 
 
 
 
 
 
 
1.4 
4.9 
6120/ΔNS2/ 
1030s 
Exp. Lot 
8494 
2.5 
3.7 
3.8 
3.5 
3.0 
- 
3.8 
17.2 
8542 
- 
5.8 
4.1 
4.5 
3.5 
- 
5.8 
19.3 
8554 
2.0 
1.5 
4.3 
4.0 
3.3 
2.1 
4.3 
17.2 
8562 
- 
2.4 
2.0 
4.5 
3.3 
1.0 
4.5 
14.0 
Mean: 
 
 
 
 
 
 
4.6 
16.9 
6120/ΔNS2/ 
1030s 
CTM RSV#012Af 
 
8928 
2.3 
1.0 
2.0 
3.2 
1.0 
1.0 
3.2 
9.9 
8940 
1.0 
3.1 
2.9 
3.4 
2.8 
1.0 
3.4 
13.6 
8983 
1.0 
1.0 
1.0 
1.0 
1.0 
2.1 
2.1 
5.6 
9045 
1.0 
2.4 
2.7 
3.1 
2.3 
1.0 
3.1 
12.1 
Mean: 
 
 
 
 
 
 
2.9 
10.5 
a Monkeys were inoculated i.n. and i.t. with 106 PFU of the indicated virus in a 1 mL inoculum per site (total dose =2x106 
PFU/AGM). 
b Virus titrations were performed on Vero cells. The lower limit of detection was 1.0 log10 PFU/mL. Samples with no detectable 
virus are represented as “-“. Underlined value indicates maximum titer for each animal. 
c The sum of daily titers is used as an estimate for the magnitude of shedding (area under the curve). Values of 0.7 are used for 
samples with no detectable virus.  
d Lot RSV#004A, vial numbers 263, 886, 1108, 1567 
e CTM vial numbers 0004, 1377, 2505 
f Lot RSV#012A, vial numbers 0019-0021, 1267-1269, 2483-2486. 
Abbreviations: AGM- African green monkey, CTM- clinical trial material, RSV- respiratory syncytial virus, TL- tracheal lavage, 
PFU- plaque-forming units, i.n.- intranasally, i.t.- intratracheally. 
 
 

 
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Table 29: Serum PRNT60 Titers in AGMs Inoculated with the Experimental Lot or with 
CTM of RSV 6120/ΔNS2/1030s or RSV ΔNS2/Δ1313/I1314L, or Recombinant wt RSV rA2 
 
Virus Test Articlea 
and Study Number 
AGM ID 
RSV Neutralization Titer      
 (Log2 of reciprocal) on Indicated Days 
0 
21 
28 
RSV rA2 
wt RSV 
CTM RSV#004Aa 
7638 
- 
7.0 
6.6 
7744 
- 
6.5 
7.9 
7781 
- 
8.0 
8.6 
7799 
- 
7.6 
7.8 
 
Mean: 
 
7.3 
7.8 
RSV ΔNS2/Δ1313/ 
I1314L 
CTM RSV#006Ab 
7648 
- 
6.4 
6.8 
7692 
- 
6.6 
6.4 
7714 
- 
4.4 
6.1 
7764 
- 
5.5 
6.0 
 
Mean: 
- 
5.8 
6.3 
6120/ΔNS2/ 
1030s 
Exp. Lot 
7638 
- 
9.6 
9.7 
7744 
- 
9.3 
9.6 
7781 
- 
7.5 
9.4 
7799 
- 
8.7 
9.4 
 
Mean: 
 
8.8 
9.5 
6120/ΔNS2/ 
1030s 
CTM RSV#012A c 
8928 
- 
6.8 
6.8 
8940 
- 
5.3 
5.7 
8983 
- 
3.3 
3.3 
9045 
- 
5.6 
5.6 
 
Mean: 
 
5.3 
5.4 
a Lot RSV#004A; vial numbers 263, 886, 1108, 1567 
b Lot RSV#006A, vial numbers 0004, 1377, 2505.  
c Lot RSV#012A, vial numbers 0019-0021, 1267-1269, 2483-2486. The lower limit of detection of the 60% Plaque Reduction 
assay is 3.3 (Log2 of the dilution reciprocal). Samples below the lower limit of detection are recorded  
as “-“.  
Abbreviations: AGM- African green monkey, CTM- clinical trial material, RSV- respiratory syncytial virus. 
 
 

 
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Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
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APPENDIX II: SCHEDULE OF EVENTS: SCREENING, ACUTE PHASE, AND POST-
ACUTE PHASE  
 
Table 30: Schedule of Events: Group 1: RSV-Seropositive Participants 
 
 
ACUTE PHASE 
POST-ACUTE PHASE 
 
Screeninga 
Day 0 
Day 1 
Day 2 
Day 3 
Day 4 
Day 5 
Day 6 
Day 7 
Day 8 
Day 9 
Day 10 
Day 11 
Day 12-27 
Day 28  
Illness 
Visit  
Early DC 
Study window  
 
± 1 day 
 
+7 days  
In person visit 
X 
X 
 
 
X 
X 
X 
X 
X 
 
 
X 
 
* 
X 
X 
X 
Non-visit contact 
 
 
X 
X 
 
 
 
 
 
X 
X 
 
X 
* 
 
 
 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
History 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Interim history 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
* 
X 
X 
X 
Physical exam (full) 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Clinical assessment 
(focused PE) 
 
X 
 
 
X 
X 
X 
X 
X 
 
 
X 
 
* 
 
X 
 
Administer study 
product 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Blood for: 
immunologic assays 
5mL 
 
 
 
 
 
 
 
 
 
 
 
 
 
5mL 
 
5mL 
Nasal wash and/or 
nasosorption SAM 
strip for: RSV 
antibody 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
X 
Nasal wash for: 
viral detection & 
quantification 
 
X 
 
 
X 
X 
X 
X 
X 
 
 
X 
 
* 
 
X 
X 
Request adventitious 
agent assay 
 
 
 
 
 
 
 
 
 
 
 
 
 
* 
 
X 
 
Total blood volume 
5mL 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
5mL 
-- 
5mL 
*If a family reports an SAE that may meet the study pause or stop criteria, complete the indicated tasks 
a Blood for RSV immunological assay is obtained not more than 56 days prior to inoculation. Screening RSV 
antibody can be obtained greater than 56 days prior to inoculation so long as it is obtained within the same calendar 
year. 

 
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Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
Table 31: Schedule of Events: Group 2: RSV-Seronegative Participants 
 
 
ACUTE PHASE 
 
 
 
POST-ACUTE 
PHASE 
 
 
Screeninga 
Day 0 
Day 1 
Day 2 
Day 3 
Day 4 
Day 5 
Day 6 
Day 7 
Day 8 
Day 9 
Day 10 
Day 11 
Day 12 
Day 13 
Day 14 
Day 15 
Day 16 
Day 17 
Day 18-27 
(contact 
each day) 
Day 28 
Day 29 
Day 30-55 
Day 56  
Illness 
Visit  
Early DC 
Study window  
 
± 1 day 
 
 
+7 days  
In person visit 
X 
X 
 
 
X 
 
X 
 
X 
 
 
X 
 
X 
 
X 
 
 
X 
 
X 
 
* 
X 
X 
X 
Non-visit contact 
 
 
X 
X 
 
X 
 
X 
 
X 
X 
 
X 
 
X 
 
X 
X 
 
X 
 
X 
* 
 
 
 
Informed consent 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
History 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Interim history 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
* 
X 
X 
X 
Physical exam  
X) 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Clinical assessment 
(focused PE) 
 
X 
 
 
X 
 
X 
 
X 
 
 
X 
 
X 
 
X 
 
 
X 
 
X 
 
* 
 
X 
 
Administer study 
product 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Blood for: 
immunologic assays 
5mL 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
5mL 
 
5mL 
Nasal wash and/or 
nasosorption SAM 
strip for: RSV 
antibody 
 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
X 
 
 
X 
 
X 
Nasal wash for: 
viral detection & 
quantification 
 
X 
 
 
X 
 
X 
 
X 
 
 
X 
 
X 
 
X 
 
 
X 
 
X 
 
* 
 
X 
X 
Request adventitious 
agent assay 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
* 
 
X 
 
Total blood volume 
5mL 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
-- 
5mL 
-- 
5mL 
*If a family reports an SAE that may meet the study pause or stop criteria, complete the indicated tasks  
a Blood for screening is obtained not more than 42 days prior to inoculation. 

 
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Version 2.0 
Principal Investigator: Ruth A. Karron, MD 
        
15 March 2019 
 
APPENDIX III: SCHEDULE OF EVENTS: RSV SEASONAL SURVEILLANCE 
 
Table 32: Schedule of Events: RSV Seasonal Surveillance 
 
Pre-RSV season 
Weekly contact 
Post-RSV season 
Illness 
Visit 
Early 
DC 
Visit period 
Oct 1st to Oct 31st  Nov 1st to Mar 31st 
Ideally Apr 1st to 
Apr 30th; 
allowable through 
Sept 30th 
 
 
Clinical assessment 
(focused PE) 
 
 
 
X 
 
Interim history 
 
X 
 
X 
X 
LABORATORY EVALUATIONS 
Blood for: 
immunologic assays 
5 mL 
 
5 mL 
 
5 mL 
Nasal wash and/or 
nasosorption SAM 
strip for 
immunologic assay 
X 
 
X 
 
 
Nasal wash for 
viral detection & 
quantification 
 
 
 
X 
 
Request adventitious 
agent assay  
 
 
 
X 
 
TOTAL BLOOD 
VOLUME 
5 mL 
-- 
5 mL 
-- 
5 mL 
 

 
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Version 2.0 
Principal Investigator: Ruth A. Karron, MD 
        
15 March 2019 
APPENDIX IV: DEFINITIONS OF SOLICITED ADVERSE EVENTS 
 
Table 33: Definitions of Solicited Adverse Events 
Event 
Defined 
Fever 
Temporal or rectal temperatures ≥100.4oF 
Acute Otitis Media1 
Loss of tympanic membrane landmarks, accompanied by erythema and loss of 
mobility. May or may not be associated with fever or other respiratory symptoms. 
Confirmed with tympanometry if possible. 
Upper Respiratory Tract Illness (URI) 
Rhinorrhea 
Two or more consecutive days of clear or purulent discharge from the nares.  
Note: Not associated with crying, change of room temperature, or eating and 
drinking. 
Pharyngitis1 
Pharyngeal erythema accompanied by exudate or pharyngeal erythema with 
enlarged, tender lymph nodes.  
Note: May be associated with sore throat, or painful or difficult swallowing. 
Cough Without LRI  
Two or more consecutive days of 3 or more episodes of cough during a 15-minute 
timed observation period, or cough awakens child from sleep.  
Note: Not associated with eating, drinking or choking. 
Hoarseness 
An unnaturally deep or rough quality of voice. 
Lower Respiratory Tract Illness (LRI) 
Wheezing2,3 
Sustained, high pitched, musical breath sounds, especially during the expiratory 
phase, which do not clear with cough. 
Pneumonia1,2,3 
Rales and crackles, originating in the lower respiratory tract, usually accompanied 
by tachypnea, which do not clear with cough. May be confirmed by x-ray showing 
areas of consolidation. 
Laryngotracheobronchitis (Croup) 
1,2,3 
Barking cough, hoarseness, and inspiratory stridor. 
Rhonchi2,3 
Coarse breath sounds which are not transmitted noises from the upper airway and 
do not clear with cough. 
Rales2,3 
Abnormal lung sound heard through a stethoscope. Rales may be sibilant 
(whistling), dry (crackling) or wet (sloshy) depending on the amount and density of 
fluid refluxing back and forth in the air passages. 
1 Diagnosis must be made by a medical professional 
2 Must be sustained over 20 minutes. 
3 Clinical assessments must be made by a medical professional and confirmed by a second medical professional, if possible. 
 
NOTE: Solicited AEs will only be recorded on CRFs according to criteria defined in Section 7.2 
 
 

 
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RSV 6120/∆NS2/1030s, Lot#012A  
           
Version 3.0 
Principal Investigator: Ruth A. Karron, MD 
        
6 April 2020 
 
APPENDIX V: RSV SEASONALITY IN BALTIMORE 
 
Figure 6: RSV Seasonality in Baltimore 
 
 
 
All specimens collected and tested at Johns Hopkins Hospital through 10 March 2016.