Perturbing the HIV Reservoir With Immune Stimulation
Phase: NA
Conditions: AIDS, HIV
Sponsor: University of California, San Diego

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1.  TITLE PAGE 
Full Title: Perturbing the HIV Reservoir with Immune Stimulation 
Title of Main Protocol and Protocol #: Perturbing the HIV Reservoir 
 
Local Protocol Number: #160089 
DAIDS-ES Document Number: (protocol number provided by DAIDS): 30122 
NIAID Funding Mechanism and Principal Investigator:  
NIH grant #1R01AI118422, David (Davey) Smith MD, MAS (PI) 
 
Other Collaborators: Susan Little MD, Sara Gianella MD 
 
 
Pharmaceutical and other Study Product Support Provided by: Not Applicable 
 
IND Number: N/A 
Protocol Chair/Co-Chair:  
Chair: Dr. Smith, Co-Chairs: Dr. Little, Dr. Gianella 
 
DAIDS Medical Officer: Dr. Lawrence Fox 
 
Version Number: 6.1 
Version Date: October 31, 2017 
 
 

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2. SIGNATURE PAGE  
I agree to conduct the study in accordance with the relevant, current protocol and will not 
make changes to the protocol without permission of DAIDS, except when necessary to 
protect the safety, rights, or welfare of study participants.  
I agree to personally conduct or supervise this study.  
I will ensure that the requirements relating to obtaining informed consent and Ethics 
Committee (EC) or Institutional Review Board (IRB) review and approval (insert relevant 
terms of assurance here, e.g. 45 CFR 46, ICH/GCP, etc.) are met.  
I agree to report to the sponsor adverse experiences as per monitoring plan that occur 
during the course of this study.  
I agree to maintain adequate and accurate study records and to make those records 
available for inspection by DAIDS, DAIDS’ authorized representatives, and/or other 
applicable regulatory entities.  
I will ensure that an EC or IRB that complies with the requirements of 45 CFR Part 46 
will complete initial and continuing review and approval of the study. I also agree to 
promptly report to the EC/IRB all changes to the study and all unanticipated problems 
involving risks to human subjects or others. Additionally, I will not make any changes to 
the study without DAIDS and EC/IRB approval, except where necessary to eliminate 
apparent immediate hazards to study participants.  
I agree to ensure that all staff members involved in the conduct of this study are 
informed about their obligations in meeting the above commitments.  
Principal Investigator:  
David Smith, MD 
 Print/Type  
Signed: ____________________________________ Date: _____________ Name/Title  
 
 
 
 
 
 

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3. TABLE OF CONTENTS 
 
1.  TITLE PAGE ................................................................................................................ 1 
2. SIGNATURE PAGE ...................................................................................................... 2 
3. TABLE OF CONTENTS ................................................................................................ 3 
4. KEY ROLES .................................................................................................................. 8 
5. LIST OF ABBREVIATIONS ........................................................................................... 9 
6. PROTOCOL SUMMARY ............................................................................................. 11 
Table 1. Schedule of Evaluations ................................................................................ 12 
Figure 1. Study Design and sampling .......................................................................... 13 
7. INTRODUCTION ......................................................................................................... 13 
7.1 Background Information ........................................................................................ 13 
7.1.1 HIV cure strategies ......................................................................................... 14 
7.1.2 Vaccines can stimulate the immune system and HIV reservoir ...................... 14 
Figure 2. Preliminary Data from Spanish Study ....................................................... 16 
7.1.3 Vaccines for HIV-infected people .................................................................... 16 
7.1.4 Incidental Immune Stimulation ........................................................................ 16 
7.2 Rationale ............................................................................................................... 17 
7.3 Major Hypotheses .................................................................................................. 18 
Primary Hypothesis: ................................................................................................. 18 
Secondary Hypotheses: ........................................................................................... 18 
8. OBJECTIVES .............................................................................................................. 19 
8.1 Primary Objective .................................................................................................. 19 
8.2 Secondary Objectives ............................................................................................ 19 
8.2.1 Aim 1a ............................................................................................................. 19 
8.2.2 Aim 1b ............................................................................................................. 19 
8.2.3 Aim 2 ............................................................................................................... 19 
8.2.4 Aim 3a ............................................................................................................. 19 
8.2.5 Aim 3b ............................................................................................................. 19 
8.2.6 Aim 3c ............................................................................................................. 20 
8.2.7 Aim 4 ............................................................................................................... 20 
8.2.8 Aim 5 ............................................................................................................... 20 
9. STUDY DESIGN ......................................................................................................... 20 

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9.1. Vaccines ............................................................................................................... 20 
9.2. Primary endpoint .................................................................................................. 20 
9.3. Study Arms ........................................................................................................... 20 
9.4. Study population ................................................................................................... 20 
9.5. Sample collection ................................................................................................. 21 
9.6. Enrollment and dropout ........................................................................................ 21 
9.7 Clinical Site ............................................................................................................ 21 
10. STUDY POPULATION .............................................................................................. 21 
10.1 Inclusion/Exclusion Criteria ................................................................................. 21 
10.1.1 Participant Inclusion Criteria ......................................................................... 21 
10.1.2 Participant Exclusion Criteria ........................................................................ 22 
10.1.3 Co-enrollment Criteria ................................................................................... 22 
10.2 Recruitment Process ........................................................................................... 22 
10.3 Participant Retention ........................................................................................... 22 
Table 2. Participant Compensation .......................................................................... 23 
11. INTERVENTIONS ................................................................................................. 23 
11.1 Biomedical Interventions ..................................................................................... 23 
11.1.1 Regimen........................................................................................................ 23 
11.1.2 Study Product Formulation and Preparation ................................................. 23 
11.1.3 Device Studies: (Not applicable) ................................................................... 23 
11.1.4 Study Product Supply and Accountability ..................................................... 23 
11.1.5 Assessment of Participant Adherence with Study Intervention ..................... 24 
11.1.6 Concomitant Medications and Procedure ..................................................... 24 
11.1.7 Permitted Medications and Procedures ........................................................ 24 
11.1.8 Prohibited Medications and Procedures ....................................................... 24 
11.1.9 Precautionary Medications and Procedures ................................................. 24 
11.1.10 Required Medications ................................................................................. 24 
11.2 Behavioral Intervention(s): (not applicable) ..................................................... 24 
12. STUDY PROCEDURES/EVALUATIONS ................................................................. 24 
12.1 Clinical Evaluations and Procedures ................................................................... 25 
12.1.1 HIV Test ........................................................................................................ 25 
12.1.2 Pregnancy Test ............................................................................................. 25 
12.1.3 Medical History and Survey .......................................................................... 25 

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12.1.4 Vaccination History ....................................................................................... 25 
12.1.5 Physical Exam .............................................................................................. 25 
12.1.6 Blood Draw ................................................................................................... 25 
12.1.7 Genital Secretion Collection .......................................................................... 25 
12.1.8 Stool Collection ............................................................................................. 26 
12.1.9 Study Vaccine Injection ................................................................................. 26 
12.2 Laboratory Evaluations ........................................................................................ 26 
Table 3. Summary of Study Assays and Schedule of Evaluations .......................... 26 
12.2.1 Specimen Preparation, Handling and Shipping ............................................ 27 
12.2.2 Biohazard Containment ................................................................................ 27 
12.2.3 Total Blood Volume ....................................................................................... 27 
12.3 Schedule of Procedures/Evaluations ................................................................... 28 
12.3.1 Screening ...................................................................................................... 28 
12.3.2 Enrollment ..................................................................................................... 28 
12.3.3 HIV Counseling and Testing ......................................................................... 28 
12.3.4 Follow-up ...................................................................................................... 29 
12.3.4.1: Baseline assessments [Day 0 of each cycle] ........................................ 29 
12.3.4.2 On study evaluations post-vaccination [Days 2, 4, 7, 14 & 30] .............. 29 
12.3.5 Early Termination Visit .................................................................................. 29 
12.3.6 Pregnancy Visit ............................................................................................. 30 
12.3.7 Final Study Visit ............................................................................................ 30 
13. ASSESSMENT OF SAFETY ..................................................................................... 30 
13.1 Safety Assessment Overview .............................................................................. 30 
13.2 Adverse Event Procedures and Reporting Requirements ................................... 30 
14. CLINICAL MANAGEMENT ....................................................................................... 30 
14.1 Clinical Management of Adverse Events ............................................................. 30 
14.1.1. Safety Monitoring Committee ....................................................................... 31 
14.1.2. Most frequent Adverse Events ..................................................................... 31 
14.2 Other Disease Events .......................................................................................... 32 
14.3 Pregnancy ........................................................................................................... 32 
14.4 Treatment Failure ................................................................................................ 32 
14.5 Criteria for Discontinuation .................................................................................. 32 
15. STATISTICAL CONSIDERATIONS .......................................................................... 33 

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15.1 Overview and General Design Issues ................................................................. 33 
15.2 Study Endpoints .................................................................................................. 34 
15.2.1 Primary Endpoint .......................................................................................... 34 
15.2.2 Secondary Endpoints: ................................................................................... 34 
15.3 Study Objectives and Hypotheses ....................................................................... 34 
15.4 Sample Size Considerations ............................................................................... 35 
Figure 3 Effect sizes with 80% power to test primary hypothesis ............................ 36 
Figure 4. Power to see a difference in HIV DNA levels following vaccination versus 
placebo .................................................................................................................... 37 
15.5 Enrollment/Randomization/Blinding Procedures ................................................. 37 
15.6 Maintenance of Trial Treatment Randomization Codes ...................................... 38 
15.7 Participant Enrollment and Follow-up .................................................................. 38 
15.8 Data and Safety Monitoring ................................................................................. 38 
15.8.1 Planned Interim Analyses and Stopping Guidelines ..................................... 38 
15.8.1.1 Interim safety review: ............................................................................. 38 
15.8.1.2 Safety Monitoring Committee ................................................................. 39 
15.8.1.3 Interim Efficacy Review: ......................................................................... 40 
15.8.2 Analysis Plan ................................................................................................ 40 
16. DATA HANDLING AND RECORDKEEPING ............................................................ 40 
16.1 Data Management Responsibilities ..................................................................... 40 
16.2 Essential Documents and Access ....................................................................... 41 
16.3 Quality Control and Quality Assurance ................................................................ 41 
17. CLINICAL SITE MONITORING ................................................................................. 41 
18. HUMAN SUBJECTS PROTECTIONS ...................................................................... 41 
18.1 Institutional Review Board/Ethics Committee ...................................................... 41 
18.2 Vulnerable Participants ........................................................................................ 41 
18.3 Informed Consent ................................................................................................ 42 
18.3.1 Informed Consent Process ........................................................................... 42 
18.3.2 Documentation of Informed Consent ............................................................ 42 
18.3.3 Stored Samples and Associated Data Considerations ................................. 42 
18.4 Risks .................................................................................................................... 44 
18.5 Social Impact Events ........................................................................................... 46 
18.6 Benefits ................................................................................................................ 46 

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18.7 Compensation ..................................................................................................... 46 
18.8 Participant Privacy and Confidentiality ................................................................ 47 
18.9 Critical Event Reporting ....................................................................................... 47 
18.10 Communicable Disease Reporting .................................................................... 47 
18.11 Incidental Findings ............................................................................................. 47 
18.12 New Findings ..................................................................................................... 47 
18.13 Study Discontinuation ........................................................................................ 48 
18.14 Post-Trial Access ............................................................................................... 48 
18.15 Community Advisory Board and Other Relevant Stakeholders ......................... 48 
19. ADMINISTRATIVE PROCEDURES .......................................................................... 48 
19.1 Protocol Registration ........................................................................................... 48 
19.2 Regulatory Oversight ........................................................................................... 49 
19.3 Study Implementation .......................................................................................... 49 
19.4 ClinicalTrials.gov ................................................................................................. 49 
20. PUBLICATION POLICY ............................................................................................ 49 
21. REFERENCES .......................................................................................................... 49 
22.1 Appendix A. Concomitant Medications CRF ....................................................... 57 
22.2 Appendix B. Medical History CRF ....................................................................... 57 
22.3 Appendix C. Physical Exam CRF ........................................................................ 57 
22.4 Appendix D. Collection of Cervical Vaginal Specimens SOP .............................. 57 
22.5 Appendix E. Laboratory SOP Transporting Specimens ...................................... 57 
22.6 Appendix F. Laboratory SOP Quality Management Plan .................................... 57 
22.7 Appendix G. Stool Specimen Collection SOP ..................................................... 57 
 
 
 

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4. KEY ROLES 
 
Principal Investigator:  
Davey Smith, M.D. 
Professor of Medicine, UCSD 
AVRC (Mail Code 8208) 
150 W. Washington Street #100, San 
Diego, CA 92103 
Telephone: (858) 642-1620 
Email: davey@ucsd.edu 
Co-investigators: 
Sara Gianella Weibel, M.D. 
Assistant 
Professor 
of 
Medicine, 
UCSD 
9500 Gilman Drive #0679  
La Jolla, CA  92093-0679 
Email: gianella@ucsd.edu 
Susan Little, M.D. 
Professor of Medicine, UCSD 
UCSD Antiviral Research Center 
200 West Arbor Drive, MC 8208 
San Diego, CA 92103 USA 
Telephone: (619) 543-8080 
Fax: (619) 543-5066 
Email: slittle@ucsd.edu 
 
Study Monitor and Data Unit: 
Susanne May 
Associate Professor of Biostatistics, 
University of Washington 
UW Biostatistics, Box 357232 
Seattle, WA 98195 
Telephone: (206) 616-0461 
Email: sjmay@uw.edu 
Statisticians: 
Christy Anderson 
Senior Biostatistician, UCSD 
UCSD Antiviral Research Center 
200 W. Arbor Drive #8208  
San Diego, CA  92103-8208 
Telephone: (619) 543-8899 
Email: czanderson@ucsd.edu 
 
Susanne May 
Associate Professor of Biostatistics, 
University of Washington 
 UW Biostatistics, Box 357232 
Seattle, WA 98195 
Telephone: (206) 616-0461 
Email: sjmay@uw.edu 
 
 

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5. LIST OF ABBREVIATIONS 
 
ACTG 
AIDS Clinical Trials Group 
AE 
 
Adverse Event 
AER 
 
Adverse Event Reports 
AIDS 
Acquired Immunodeficiency Syndrome 
ART 
 
Antiretroviral Therapy 
AVRC   
Antiviral Research Center  
CB 
Clinical Investigation and Biostatistics (CB) Core 
CD4 
 
CD4 lymphocytes 
CFAR 
Center for AIDS Research 
CMV  
Cytomegalovirus 
CRF 
 
Case Report Form 
ddPCR  
droplet digital PCR 
DSMB 
Data Safety and Monitoring Board 
EDI 
Estimated date of infection  
HBsAg  
Hepatitis B surface Antigen 
HCV 
Hepatitis C Virus 
HHV 
Human Herpesviruses 
HIV-1 
Human Immuno-deficiency Virus – 1 
HDACi 
Histone Deacetylase Inhibitors 
HPV 
 
Human Papilloma Virus  
HRPP  
Human Research Protections Program  
HRSA  
Health Resources and Services Administration 
IL 
 
Interleukin 
LPS 
Lipopolysaccharide 
MMR 
Measles, Mumps, Rubella 

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MSM  
Men who have sex with men 
NA 
not applicable  
RCT 
Randomized Controlled Trial 
RT-PCR 
Real Time-PCR  
PBMC 
Peripheral Blood Mononuclear Cells  
PCR 
Polymerase Chain Reaction 
PID  
 
Participant Identifier 
SAE 
 
Severe Adverse Reaction 
SCA 
Single Copy Assay 
SDPIRC  
San Diego Primary HIV Infection Consortium  
TDAP 
Tetanus, Diphtheria and Pertussis 
UCSD  
University of California San Diego 
VZV 
Varicella Zoster Virus 
 
 
 

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6. PROTOCOL SUMMARY 
Title: Perturbing the HIV Reservoir 
Sample Size: 56 
Study Population: HIV-infected individuals between 18 and 65 years old who started 
antiretroviral therapy (ART) during chronic infection (at least one year from estimated date of 
infection) and remained virally suppressed for at least 48 weeks before enrollment. Participants 
will have CD4 >250 cells/μl at enrollment and a CD4 nadir >100 cells/μl.  
Participating Sites: UCSD’s Antiviral Research Center (AVRC) 
Study Design: The proposed study is a randomized double-blinded controlled trial conducted 
over 28 weeks. Randomized interventions will be injections of Influenza vaccine, Pneumococcal 
vaccine, and Placebo. Each participant will receive each injection but in a randomized order.  
See Figure 1 below. 
Schedule of Evaluations: Study evaluations will be based on three 30 day cycles (Influenza 
vaccine, Pneumococcus vaccine, Placebo in random order) over 28 weeks of the RCT. Pre 
injection: one paired blood, optional stool and optional genital secretion sample will be collected 
before each injection. Post-injection: Blood will be collected on days 2, 4 and 7 while paired 
blood and optional genital secretion samples will be collected on days 14 and 30 after each 
injection. 
 
 
 
 
 
 
 
 
 
 
 
 

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a. 
 Cycle 1 to begin within 30 days of screening visit 
b. 
± 2 days  
c. 
Minimum 6 week washout period between cycles 
d. 
Paired blood and optional genital secretion sample will be collected before each injection  
e. 
Pregnancy Test will be repeated only if pregnancy is suspected.  
Table 1. Perturb Study - Schedule of Evaluations 
Activity 
Screen
ing 
Cycle 1 Treatment Period  
Fluarix /Fluzone, 
Pneumovax 23 or Placebo a 
Cycle 2 Treatment Period  
Fluarix/Fluzone, 
Pneumovax 23 or Placebo c 
Cycle 3 Treatment Period 
Fluarix/Fluzone, 
Pneumovax 23 or Placebo c 
Premature 
Discontinu
ation 
 
 
Baseline 
(Day 0)
Day 2b 
Day 4 b 
Day 7 b 
Day 14 b 
Day 30b 
Week 12 
(Day 0)
Day 2b 
Day 4 b 
Day 7 b 
Day 14 b 
Day 30 b 
 
Week 24  
(Day 0)
Day 2 b 
Day 4 b 
Day 7 b 
Day 14 b 
Day 30 b 
Informed 
Consent 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Eligibility 
Checklist 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Medical Release 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Vaccination 
History 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Randomization 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Blood collectiond 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Pregnancy teste 
X 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Physical Exam 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Medical History, 
Health Survey 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Concomitant 
Medications 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
ARV Adherence 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Post-vax 
Symptoms 
 
X 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X 
 
X 
X 
X 
X 
X 
X 
Adverse Events 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Study Drug 
Injection 
 
X 
 
 
 
 
 
X 
 
 
 
 
 
X 
 
 
 
 
 
 
Optional Stool 
Collection 
 
X 
 
 
 
 
 
X 
 
 
 
 
 
X 
 
 
 
 
 
 
Optional Genital 
Secretiond 
 
X 
 
 
 
X 
X 
X 
 
 
 
X 
X 
X 
 
 
 
X 
X 
X 
Hematology/Che
mistry/ 
Coagulation 
Panel 
 
X 
 
 
 
X 
X 
X 
 
 
 
X 
X 
X 
 
 
 
X 
X 
X 
Immune 
responses 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
HIV-1 RNA 
sequencing 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
CD4+ T-cell 
Count 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
HIV-1 DNA 
sequencing 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Archive Plasma  
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 
X 

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Study Duration: 240 weeks 
Study Regimen/Intervention:  This is a double blind RCT of two vaccines (Pneumovax®23 and 
Fluarix® or Fluzone®) plus placebo (sterile saline injection). Study participants will be followed 
for 28 weeks after enrollment. During this 28-week period, blood samples will be collected on 
day 0 and five subsequent time points after each injection (days 2, 4, 7, 14 and 30). Optional 
genital secretion samples will be collected on day zero and on days 14 and 30 after each 
injection. Optional stool will be collected at day zero. Injections (vaccine or placebo) will be 
administered 12 weeks apart and in a random order, to minimize a possible bias due to the 
order of the vaccines.  
NOTE: Fluarix® quadravalent vaccine is the flu vaccine of choice for this study. Fluzone® 
quadravalent vaccine may be substituted for Fluarix® only in the event that Fluarix® is 
unobtainable from the wholesaler/manufacturer.  
Primary Objective: To determine the impact of Pneumococcus and Influenza vaccines on the 
HIV transcriptional activity in individuals who are virologically suppressed for at least 48 weeks 
on ART. 
Primary Outcome: Average change in level of CD4+ T-cell-associated HIV RNA transcription 
from baseline to day 7 after each injection. 
Figure 1. Study Design and sampling 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
7. INTRODUCTION 
7.1 Background Information 
Figure 1. Study Design and sampling 
Legend: 56 chronically HIV+ individuals on ART will receive Influenza 
and Pneumococcus vaccines and Placebo injections in random order 
over 24 weeks (red arrows). Blood will be collected before each 
injection (blue arrow day 0) and 2, 4, 7, 14 and 30 days after each 
injection (open arrows). Optional genital secretion will be collected on 
Days 0, 14 and 30. Optional stool samples will be collected before the 
injection on Day 0. 
 
HIV+ par cipants 
N=56 
Injections 
1 
2 
3 
4 weeks 
4 weeks 
4 weeks 
12 weeks 
12 weeks 
28 weeks 

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7.1.1 HIV cure strategies  
HIV can be effectively treated, but not cured, except for very rare and special circumstances [5]. 
The recent report of the ‘Berlin Patient’ [6] has sparked new enthusiasm for such efforts. The 
first problem with curing HIV is that the virus has a latent stage that persists indefinitely, even 
during suppressive ART, resulting in recurrence of plasma viremia if ART is stopped. The 
second problem is identification of specific cells that are latently infected [5]. Most current 
curative strategies have focused on developing methods that are able to induce the virus from 
latently infected cells during ART, so that viral proteins are revealed and allow these cellular 
reservoirs to be cleared by the host immune response, while ART prevents new cells from being 
infected [7, 8]. Unfortunately, recent interventions to perturb the reservoir, like histone 
deacetylase inhibitors (HDACi), IL-7, and disulfram, have demonstrated only modest activity [7, 
9-13]. It is also becoming increasingly apparent that it is not enough to just induce HIV RNA 
expression during ART to clear latently infected cells, but activation of HIV-specific immune 
responses is also needed [7, 8]. Therefore, it seems likely that parallel efforts to augment the 
HIV-specific immune response may be required [8, 10, 14]. This proposal aims to evaluate two 
clinically indicated vaccines with longstanding safety records as immune stimulatory methods 
for perturbing the HIV reservoir and inducing release of virus and the subsequent potential for 
viral clearance by the activated host immune system. 
7.1.2 Vaccines can stimulate the immune system and HIV reservoir 
T-cell activation has been shown to effectively reactivate HIV replication in various in vitro 
models [13, 15-17]. Similar effects have been observed with in vivo immune activation 
secondary to current illness [18-26], human herpesvirus (HHV) shedding [27-30] and gut 
microbial translocation [31-34]. Prophylactic vaccination represents another means of activating 
the immune system and has been associated with mostly transient increases in levels of cell-
free HIV RNA after vaccination for influenza [21, 35-43], pneumococcus [44-46], tetanus [42, 
47], hepatitis B [48] and cholera [49]. This stimulation of the immune system and the HIV 
reservoir probably occurs because standard clinical vaccines activate not only the small subset 
of antigen specific cells within the lymphocyte population but also induce a bystander activation 
of non-specific cells. This hypothesis is supported by the measurable increases in various 
markers of systemic inflammation (e.g. pro-inflammatory cytokines [interleukin (IL)-6, IL-1, C 
reactive protein, tumor necrosis factor (TNF)], T-cell activation [CD38+HLA-DR+] and 
proliferation [Ki67+]) [50-55] after vaccination.  For inactivated vaccines, this inflammation is 
largely mediated by an adjuvant. Studies of vaccine-associated effects on HIV RNA are mixed 
however, with several studies demonstrating no significant effect on plasma viral loads (i.e., 
influenza [56-61], pneumococcus [62-64], tetanus [65], hepatitis A [66, 67], hepatitis B [68], 
measles/mumps/rubella (MMR) [69], Haemophilus influenzae type b [70] and yellow fever [71]). 
The inconsistent findings in the literature are likely secondary to [72-74]: (1) observational 
convenience studies with varying sampling times (e.g. 1 week versus 1 month), (2) 
heterogeneous study populations (e.g. different ART regimens, durations of infection, baseline 
viral loads and CD4+ T cell counts [36]), and (3) insensitive methods applied to measure how 
much HIV RNA is being induced from infected cells. The proposed study will attempt to address 
these issues by: (1) performing a randomized, double-blind, placebo-controlled trial (RCT) 
(vaccine vs. vaccine placebo) with optimized sampling times post-vaccination, (2) investigating 

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participants who have received ART for at least 48 weeks, which was started during chronic 
infection, and (3) using more sensitive methods to measure virus production from cells (single 
copy assays to measure cell-free HIV RNA and droplet digital PCR (ddPCR) for cell-associated 
HIV RNA [75]. 
While no previous study has examined the impact of vaccines on the HIV reservoir, one 
Spanish RCT of seven vaccines [1] in chronically infected individuals receiving ART did 
evaluate the hypothesis that transitory increases in HIV replication during ART would strengthen 
specific responses against HIV and induce a better control of HIV replication after ART 
interruption. Although increases in HIV-specific T-cell responses and in T-cell activation were 
observed in the vaccine group compared to the placebo group, interruption of ART was not 
associated with control of HIV replication. While this study did not show a significant increase in 
levels of cell-free HIV RNA production in the vaccine vs. placebo arms, it did suggest that 
enough HIV transcription was induced to increase HIV-specific immune responses. In contrast 
to the proposed study, this Spanish study only used standard viral load measures at wide 
intervals (30 days post-injection)[1]. To further evaluate this hypothesis, we generated 
preliminary data from the Spanish study [1] using more appropriate assays in support of our 
hypothesis that vaccination can activate proviruses during ART more than any published HDACi 
compound [7, 9, 12, 13, 76]. In these experiments, we found (see Figure 2 below):  
 
no difference in cellular HIV RNA levels between vaccine and placebo groups at 
baseline (p>0.1),  
 
significant increases in HIV transcription following each vaccine (all p<0.05),  
 
mean fold change in cell-associated HIV RNA gag transcripts following both Influenza 
and Pneumococcal vaccines was considerably higher than what has been observed in 
two clinical studies of the HDACi Vorinostat (Archin et al.[3] and Elliott et al.[4]).  
In summary, we acknowledge that the literature is not clear whether or not clinical vaccination 
can perturb the HIV reservoir, but this is because all previous studies: 
 
Were observational with a very wide range of sampling times (<1 week to >1 month).  
 
Had very heterogeneous populations with different ART regimens, ages, duration of 
infection, baseline viral loads, CD4 T-cell counts, etc. In observational studies that do 
account for these factors, such heterogeneity can confound any true relationship.  
 
Used older insensitive methods (i.e. basic standard viral load assays with lower limit of 
detections between 50 and 400 copies/ml) to measure HIV RNA transcription. 
Taken together, we feel that it would be a mistake to discard this line of investigation based 
on historical, observational studies that were not designed to address the important 
questions posed in this proposal. 

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Figure 2. Preliminary Data from Spanish Study 
 
7.1.3 Vaccines for HIV-infected people 
HIV-infected individuals are at increased risk for a variety of communicable diseases compared 
to uninfected people and several vaccines are routinely recommended. In general, vaccines 
have better efficacy in HIV-infected individuals with CD4 counts >200 cells/µL [77]. However, all 
vaccines are not created equal [78]. They can differ in immune stimulating mechanism; some 
induce mostly neutralizing antibodies while others also induce cytotoxic T-cell responses [78-
86]. They can also differ in efficacy; for example the human papilloma virus (HPV) vaccine is 
close to 100% effective while influenza is only ~65% effective [87]. Further, immune responses 
wane over time and at different rates; the tetanus vaccine requires repeat vaccination every 10 
years, while the protection from the smallpox vaccine is likely life-long [87]. Additionally, 
effective inactivated vaccines often require an adjuvant (Alum, AS04, or MF59), which primes 
the innate and adaptive immune responses quite differently [78]. This study will evaluate the two 
standard vaccines that showed the greatest mean increase in HIV RNA transcription following 
vaccination in the Spanish Trial (see Research Plan). Other vaccines could be evaluated in this 
proposal, but we chose the Influenza and Pneumococcal vaccines that are commonly used 
clinically and have a very long safety record.  
7.1.4 Incidental Immune Stimulation 
Immune stimulation following vaccination has the potential to be confounded by immune 
activation associated with other specific and non-specific stimuli [28, 88-90]. In an effort to 
Figure 2. Preliminary Data from Spanish Study [1, 2] 
A: Large increases in cell 
associated HIV RNA after 
each vaccine administration, 
but variable across subjects. 
B: Cell associated HIV RNA 
before 
and 
after 
influenza 
vaccine administration. 
C: Cell associated HIV RNA 
before and after pneumococcus 
vaccine administration. 
D: HIV transcription induced by 
influenza, pneumococcus and 
Vorinostat. 
 
Legend: We used samples shared from the Spanish study [1] to measure cell associated HIV RNA in 12 vaccinated and 11 control 
individuals. At baseline (pre-vaccine), there was no difference in cellular HIV RNA levels between those who would receive the vaccine 
and those who would receive placebo (controls) (p>0.1). Panel A: After each vaccine we found significant median increases in HIV 
transcription; like other interventions aimed to perturb the reservoir, there was considerable differences between participants. Panels B 
and C: Here, we will focus on the evaluation of the two vaccines that demonstrated the highest fold increases in cell associated HIV 
RNA (Influenza and Pneumococcus). Panel D: The mean fold change (with standard error) in cell-associated HIV RNA gag transcripts 
following both of these vaccines was considerably higher than what has been observed in two studies of in vivo administration of  
Vorinostat (Archin et al.[3] and Elliott et al.[4])  
3
4
5
6
7
12
1
2
8
9
10 11
0
500
1000
1500
2000
3000
HIV RNA 
(Gag/ 106 CD4 T cells)
Influenza
MMR
TDaP
Pneumococcus
Hep A/ VZV
Hep A/ VZV

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optimally control for possible confounding within a relatively small RCT, we propose a detailed 
analysis of the possible contribution of incidental immune stimulation resulting from HHV 
infections, microbial translocation, drug use and clinical illnesses. For example, we recently 
showed that CMV replication in seminal plasma of HIV infected subjects on suppressive ART 
was associated with increased levels of T-cell activation and higher levels of proviral HIV DNA 
and cellular HIV RNA compared to subjects with no detectable CMV (Figure 2) [30]. 
Additionally, our group has identified an association between methamphetamine use and 
increases in both immune activation and HIV DNA levels during suppressive ART [91]. 
Therefore, we will try to identify these factors and account for their potential effect. Specifically, 
we will identify and measure: 1) genital shedding of 7 HHV (by RT-PCR), 2) microbial 
translocation (as measured by lipopolysaccharide [LPS], bacterial 16s rDNA and soluble 
[s]CD14), 3) active or recent illicit substance use (using validated questionnaires [92-94] and 
urine toxicology) and 4) recent clinical illnesses (e.g. colds, skin infections, sexually transmitted 
infections) and clinical events (e.g. surgeries). 
7.2 Rationale  
Most recent HIV cure efforts have focused on strategies to induce virus from latently infected 
cells during ART, thus leaving the cellular reservoir vulnerable to the host immune system, while 
ART prevents new cells from being infected. Unfortunately, such efforts have not demonstrated 
much activity. On the other hand, it has been documented for almost two decades that routine 
clinical vaccines can induce viral production from latently infected cells, even during ART, and 
that these levels of induction are much higher than those seen by recently described 
interventions (e.g. histone deacetylase inhibitors (HDACi), IL-7, disulfram). Therefore, we 
propose a RCT with cross-over to determine how two commonly used vaccines (Influenza and 
Pneumococcus) can stimulate the immune system and induce HIV transcription. In preliminary 
data, these vaccines demonstrate a higher cell associated HIV RNA transcriptional response 
even 30 days after vaccination than the HDACi Vorinostat; see Figure 2. In this proposal, we will 
use a much safer method (vaccination with Influenza and Pneumococcus) and observe 
timepoints much closer to the time of vaccination to tease out how immune stimulation perturbs 
the HIV DNA reservoir suppressed by ART. 
Rationale for measuring total and replication competent HIV DNA 
Currently, much emphasis has been placed on the kick portion of the kick-and-kill HIV cure 
strategy; however, only modest benefit has been seen with kick interventions and almost none 
with the killing of infected cells- at least with the current measurements in circulating blood [7, 9-
13]. In this study, we will use clinically available influenza and pneumococcus vaccines to 
stimulate the HIV reservoir and stimulate the immune response. In the prior Spanish Study, 
described in the Background, it was observed that HIV-specific immune response was 
enhanced during the 'kick' of the reservoir in the blood after a clinical vaccine. This study did not 
measure HIV DNA levels or replication competent proviral levels; therefore, we do not know if 
this HIV-specific immune stimulation had an impact on the HIV reservoir in circulating blood 
cells. Therefore, in the proposed RCT, we will assess both total HIV DNA levels and replication 
competent provirus (using an “inducible RNA assay” [95]) among the five participants with the 
highest cell associated HIV RNA response to the vaccine intervention. 

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Microbiome Composition  
Infection with HIV is associated with dramatic alterations in the gut-associated lymphoid tissue, 
which occur early in the course of infection and contribute to persistent immune dysfunction and 
HIV disease progression. Mucosal HIV replication and consequent depletion of gut-associated 
CD4+ T cells is associated with epithelial barrier damage and increased translocation of 
bacterial products from the gut lumen into the systemic blood circulation (i.e. microbial 
translocation).  In turn, microbial translocation is associated with systemic immune activation 
and might play a role in immune response and disease progression in untreated and treated 
HIV infected individuals.  
As part of this study we will determine the effect of the intestinal microbiome composition on the 
viral and immunologic dynamics before and after vaccine administration.  
7.3 Major Hypotheses  
(All hypotheses are presented “under the alternative”) 
Primary Hypothesis:  
On average, participants will have a higher absolute increase in levels of cell-associated HIV 
RNA transcription in CD4+ T-cells seven days after receiving either Pneumococcal or 
Influenza vaccinations, when compared to seven days after receiving placebo. 
Secondary Hypotheses:  
 
Among the five participants with the largest increases in cell-associated HIV RNA in 
CD4+ T-cells following active vaccination (maximum across all observed time-points), 
we will deep sequence cell-associated HIV RNA and DNA populations at 0, 2, 4 7, 14 
and 30 days. We hypothesize that active vaccines will non-selectively activate cellular 
transcription of HIV, as evaluated by panmixis tests between sequences of HIV DNA and 
RNA populations at these sampled timepoints. 
 
On average, participants will have a greater absolute maximum decrease in levels of 
total CD4+ T-cell-associated HIV DNA measured 2-30 days after receiving either 
Pneumococcal or Influenza vaccinations, when compared to 2-30 days after receiving 
placebo. 
 
Among the five participants with the largest increases in cell-associated HIV RNA in 
CD4+ T-cells following active vaccination (maximum across all observed time-points), 
we will perform inducible RNA assay. We hypothesize that these five participants will 
have lower levels of replication competent proviruses 30 days after active vaccination, 
as compared to their levels of replication competent proviruses sampled before this 
vaccination. 
 
Participants will have a greater absolute maximum increase in levels of (a) inflammatory 
cytokines, (b) T-cell activation and proliferation, (c) HIV-specific immune responses, and 
(d) vaccine-specific immune responses over the 30 days of observation after receiving 
either active vaccination, when compared to maximum measures taken over the 30 days 
of observation after receiving placebo. 

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 
Levels of non-antigen specific immune stimulation (cytokines and general T-cell 
activation) and vaccine-specific immune responses will be positively associated with 
increased CD4+ T-cell-associated HIV RNA transcription and negatively associated with 
lower levels of HIV DNA, over the 30 days of observation after receiving either 
Pneumococcal or Influenza vaccinations. 
 
Maximum levels of cell-associated HIV RNA over the 30 days of observation following 
active vaccination will be positively associated with HIV-specific responses 30 days after 
receiving active vaccinations. 
 
Genital shedding of human herpesviruses, increased levels of microbial translocation 
and incident illnesses will stimulate the immune system and induce increased cell-
associated HIV RNA transcription from CD4+ T-cells in participants independent of 
receiving active vaccine or placebo. 
 
 
Presence of dysbiosis (for example increased relative frequency of Proteobacteria in 
stool samples) will be associated with increased microbioal translocation and with 
increased cell-associated HIV RNA transcription from CD4+ T-cells in participants 
independent of receiving active vaccine or placebo. 
8. OBJECTIVES 
8.1 Primary Objective 
 
Determine the impact of Pneumococcus and Influenza vaccines on the HIV transcriptional 
activity in individuals who are virologically suppressed for at least 48 weeks on ART. 
8.2 Secondary Objectives 
8.2.1 Aim 1a  
Determine the timing and duration of increased HIV transcription following active vaccination. 
8.2.2 Aim 1b 
Determine if vaccination selectively or non-selectively activates HIV transcription from HIV DNA 
populations 
8.2.3 Aim 2 
Determine if active vaccination influences levels of cell associated HIV DNA or replication 
competent provirus. 
8.2.4 Aim 3a 
Determine to what degree immune stimulation with an Influenza or Pneumococcal vaccine can 
induce HIV transcription during ART. 
8.2.5 Aim 3b 
Determine if immune stimulation associated with active vaccination is associated with reduction 
in number of HIV DNA populations. 

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8.2.6 Aim 3c 
Determine if HIV-specific, vaccine-specific, or bystander immune stimulation is associated with 
HIV transcription. 
8.2.7 Aim 4 
Determine how genital shedding of human herpesviruses, microbial translocation and incident 
illnesses stimulate the immune system and influence levels HIV transcription. 
8.2.8 Aim 5 
Determine how dysbiosis contribute to stimulate the immune system and influence HIV 
transcription. 
9. STUDY DESIGN 
The proposed study is a randomized double-blinded control trial conducted over 28 weeks. 
Randomized interventions will be injections of Influenza vaccine, Pneumococcal vaccine, and 
Placebo. Each participant will receive each injection but in a randomized order.  
9.1. Vaccines 
Influenza (Fluarix®, GSK or Fluzone®, Sanofi -Pasteur), Pneumococcal (Pneumovax®23, 
Merck), and placebo (sterile saline injection). 
NOTE: Fluarix® quadravalent vaccine is the flu vaccine of choice for this study. Fluzone® 
quadravalent vaccine may be substituted for Fluarix® only in the event that Fluarix® is 
unobtainable from the wholesaler/manufacturer.  
9.2. Primary endpoint 
average change in level of CD4+ T-cell-associated HIV RNA transcription from baseline to day 
7 after each injection. 
9.3. Study Arms 
Each subject will take Influenza (Fluarix®, GSK or Fluzone®, Sanofi -Pasteur), Pneumococcal 
(Pneumovax®23, Merck), and placebo. Randomization will determine the order in which the 
subjects receive the injections. There are six possible study arms, one for each order in which 
someone can receive the injections: 
Placebo, Flu, Pneumovax 
Placebo, Pneumovax, Flu 
Flu, Placebo, Pneumovax 
Flu, Pneumovax, Placebo  
Pneumovax, Placebo, Flu 
Pneumovax, Flu, Placebo 
9.4. Study population 

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This study will enroll 56 HIV-infected individuals between the ages of 18 and 65 who started 
ART during chronic infection. These subjects will be recruited from both the UCSD AntiViral 
Research Center and the Owen Clinic. Eligible participants will have CD4 >250 cells/μl at 
enrollment and a CD4 nadir >100 cells/μl. We chose to limit enrollment to younger individuals 
with higher CD4 counts to enhance the likelihood of having a more robust immune response to 
any given vaccine. We limited enrollment to individuals who started ART during chronic infection 
(at least one-year from estimated date of infection) to enhance the likelihood of having a large 
replication competent viral reservoir, thus increasing our chances to observe an effect of 
vaccine administration on HIV RNA transcription [96]. To reduce variability in vaccine 
stimulation (between antigen naïve versus experienced participants), we will also limit 
participation to individuals who have documented or reported vaccine history consistent with 
previous influenza and pneumococcal vaccination.   
9.5. Sample collection 
Study evaluations will be based on three 30 day cycles (Influenza vaccine, Pneumococcus 
vaccine, Placebo in random order) over 28 weeks of the RCT. Pre injection: one paired blood, 
optional stool and optional genital secretion sample will be collected before each injection. Post-
injection: Blood samples will be collected on days 2, 4, 7, 14 and 30 after each injection. 
Optional genital secretion samples will be collected on days 14 and 30 after each injection; see 
Table 1 Schedule of Evaluations. 
9.6. Enrollment and dropout 
All participants will be enrolled in the first 36 months of the study to allow for a complete 28 
weeks of observation for each subject. To decrease variability, we will only enroll individuals 
who have remained suppressed with no viral loads >50 copies/ml 48 weeks before enrollment. 
Viral loads are performed as part of recently funded ‘ART NET’ study in San Diego 
(1R01MH100974-01A1, PI Little) every 6 months and in the Owen clinic every 3 months, and 
we will have access to both measures. Anticipating a 10% dropout, we will enroll 56 subjects 
into our RCT. We thus conservatively expect to achieve an evaluable sample size of 50 at study 
end. 
9.7 Clinical Site 
All study visits will take place at the UCSD Antiviral Research Center (AVRC) located at 220 
Dickinson St, San Diego, CA 92103. 
10. STUDY POPULATION 
10.1 Inclusion/Exclusion Criteria 
10.1.1 Participant Inclusion Criteria 
10.1.1.1 
Documented HIV-1 infection more than 365 days ago (HIV antibody or  
 
            viral load positive). 
 
10.1.1.2 
Capable of signing written informed consent. 
10.1.1.3 
Documented viral suppression for at least 48 weeks (≤50 copies/mL)  

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10.1.1.4 
Men and women between 18 and 65 years of age.  
 
10.1.1.5 
Read and comprehend English.  
 
10.1.1.6 
Documented CD4 count at enrollment (>250 cells/µl) 
 
10.1.1.7 
Reported CD4 nadir >100 cells/µl. 
 
10.1.1.8 
Received seasonal flu vaccination at least 6 weeks prior to first trial  
 
                        injection (and not more recently) 
10.1.1.9 
Received vaccination for pneumococcal disease at least 12 months prior    
 
 
to first trial injection (and not more recently) 
10.1.1.10 
Started ART >6 months  from estimated date of infection) 
10.1.2 Participant Exclusion Criteria 
 
10.1.2.1 
Uncontrolled psychiatric condition. 
 
10.1.2.2 
Under the influence of drug(s) or alcohol at time of screening. 
 
10.1.2.3 
Any condition that, in the opinion of the investigator, would limit  
 
 
 
 
follow-up and adequate consent. 
 
10.1.2.4 
History of allergic reactions to any of the proposed vaccines or  
 
 
 
 
egg allergy. 
 
10.1.2.5 
History of Gullian Barre syndrome. 
 
10.1.2.6 
Receiving immunosuppressive medications. 
 
10.1.2.7 
Pregnancy or lactation.  
10.1.3 Co-enrollment Criteria 
Co-enrollment in this study and other studies (other than ART NET as discussed above) will be 
discussed with the protocol team and will be decided on a case-by-case basis. 
10.2 Recruitment Process 
Subjects will be recruited from the UCSD AVRC and the Owen Clinic. Owen Clinic is an HIV 
care program affiliated with UC San Diego Health System. It offers HIV primary care in addition 
to a number of services such as ART therapy, mental health services, medication adherence 
and substance abuse counseling. Subjects visiting either AVRC or the Owen clinic who are 
known to be HIV positive and meet the inclusion criteria will be recruited to this study and a 
screening visit will be scheduled. 
10.3 Participant Retention 

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If a subject meets the inclusion criteria and completes all activities of the screening visit, the 
study clinician will personally assist the patient in scheduling follow-up visits in order to increase 
participant retention. The subject will be compensated for each follow up visit according to the 
following table. Each participant has the opportunity to complete this 30 day cycle a total of 
three times, one for each study vaccine and placebo. The total amount each participant may be 
compensated is $570 ($190 for each cycle); see Table 2 Participant Compensation. 
Table 2. Participant Compensation 
Study Visit 
Laboratory Procedure
Compensation 
Baseline 
Blood Draw, optional Genital Secretion, optional 
stool, Intramuscular Injection 
$50
Day 2 
Blood Draw 
$20
Day 4 
Blood Draw 
$20
Day 7 
Blood Draw 
$20
Day 14 
Blood Draw, Optional Genital Secretion
$30
Day 30 
Blood Draw, Optional Genital Secretion
$50
 
11. INTERVENTIONS 
11.1 Biomedical Interventions 
11.1.1 Regimen 
Each study subject will receive one dose each of the following: Pneumovax®23, Fluarix® (or 
Fluzone®), and placebo (sterile saline solution). These will be administered at the baseline visit 
(Day 0) of each cycle and will precede 30 days of follow-up. After the 30 days of each cycle, 
there is a washout period (minimum 6 weeks) before the next cycle begins. Each study drug will 
be administered as a one time intramuscular injection. 
NOTE: Fluarix® quadravalent vaccine is the flu vaccine of choice for this study. Fluzone® 
quadravalent vaccine may be substituted for Fluarix® only in the event that Fluarix® is 
unobtainable from the wholesaler/manufacturer.  
11.1.2 Study Product Formulation and Preparation 
In order to achieve double blind administration of the vaccines, each drug or placebo will first be 
transferred to generic, non-labeled syringes by the Study Pharmacist before being delivered to 
the clinical staff who will administer the injection. This preparation (along with preparation of the 
sterile saline placebo) will take place in the AVRC pharmacy. No after-market modifications will 
be made to the formulation of Pneumovax®23 (Merck) or Fluarix® (GSK), or Fluzone® (Sanofi 
Pasteur) vaccines. 
11.1.3 Device Studies: (Not applicable) 
11.1.4 Study Product Supply and Accountability 

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All study product supply will be stored and handled by AVRC pharmacy staff. All pharmacy 
procedures will be done in accordance with existing verified Standard Operating Procedures 
(SOPs) that pertain to pharmacy inventory, storage, security and accountability. 
11.1.5 Assessment of Participant Adherence with Study Intervention 
Since the study intervention (intramuscular injection) will only be administered three times and 
always with a study clinician present, there is no need to assess participant adherence aside 
from attending the baseline visit of each study regimen cycle. 
11.1.6 Concomitant Medications and Procedure 
All concomitant medications will be recorded (including start/stop date, medication type, dose 
and indications) in a Concomitant Medications Flowsheet (see Appendix A. CRF #CON030). 
Whenever a concomitant medication or study product is initiated or the dose changed, 
investigators must review the concomitant medications’ and study products’ most recent 
package inserts, investigator's brochure, as well as updated information from DAIDS to obtain 
the most current information on drug interactions, contraindications, and precautions. 
11.1.7 Permitted Medications and Procedures 
Because it may interfere with the findings of the study, participants are asked to refrain from 
receiving routine vaccinations during the study unless otherwise instructed by their primary 
physician. However, vaccination outside the study protocol will not result in discontinuation of 
the study. Instead, these instances will be identified and documented, and investigators will 
control for this instances during analysis. Similarly, any change in any medication, including gain 
or loss, will be recorded on the Medical History CRF (See Appendix B). 
11.1.8 Prohibited Medications and Procedures 
Medications that have been suggested to perturb the HIV reservoir or the immune system will 
require early discontinuation of the study. These medications include: disulfram, histone 
deacetylase inhibitors, other cancer chemotherapies, biologic immunosuppressives, high dose 
steroids etc. Other medications or therapies that are taken will be documented during the study, 
and will be reported to the PI. 
11.1.9 Precautionary Medications and Procedures 
Other medications that have been suggested to perturb the HIV reservoir or the immune 
system, like antibiotics, non-steroidal anti-inflammatory drugs, aspirin, or low dose steroids, will 
be noted and accounted for in the final analyses. 
11.1.10 Required Medications 
Study participants must be on ART. Moreover, their HIV therapy must remain successful 
throughout the study (CD4 count >250 cells/ml and nadir >100 cells/ml). The study participants 
must also have a recent vaccination history including previous seasonal flu vaccination and 
previous pneumovax vaccination at least 12 months prior to screening visit. 
11.2 Behavioral Intervention(s): (not applicable) 
12. STUDY PROCEDURES/EVALUATIONS 

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12.1 Clinical Evaluations and Procedures 
12.1.1 HIV Test 
An HIV test will be performed to confirm HIV infection if documentation is not available. 
Acceptable HIV tests include any FDA approved test that is approved for diagnosing HIV 
infection, such as Oraquick ADVANCE, OraSure Technologies, Inc., INSTI HIV-1/HIV-2 
Antibody Test, bioLytical Laboratories Inc., ARCHITECT HIV Ag/Ab Combo, Abbott 
Laboratories, etc. Participants will receive a finger prick and a few drops of blood will be tested 
before enrollment.  
12.1.2 Pregnancy Test 
If the participant is a woman and can become pregnant, a urine pregnancy test will be 
performed at screening and will be repeated only if pregnancy is suspected.  
12.1.3 Medical History and Survey 
Participants will be asked about their medical history and will be administered a brief survey of 
incident illness, which will be recorded on Medical History CRF; See Appendix B. 
12.1.4 Vaccination History 
Participants will be asked about their vaccination history, specifically, the previous seasonal flu 
and pneumovax vaccines. A documented or reported pneumovax vaccine will be required at 
least 12 months prior to the screening visit. 
12.1.5 Physical Exam 
A brief, symptom-targeted physical exam will be performed and recorded on the Physical 
History CRF; See Appendix C. 
12.1.6 Blood Draw 
Approximately 50 mL of blood will be drawn at each visit for the purpose of CD4+ T-cell counts, 
HIV-1 RNA viral load, HIV genetic testing, and immunologic testing. At baseline, day 14 and day 
30 of each cycle, we will collect additional 10mL (60mL total) to perform routine blood chemistry 
tests, hematology and coagulation (safety tests).     
12.1.7 Genital Secretion Collection (optional) 
 
For Men: Men will be asked to provide a semen specimen. In the privacy of their home, 
or in the privacy of our clinic, they will be asked to give us a semen sample. This will 
require that they do not have sexual activity for 24 hours prior to the semen collection.  
They will need to wash their hands and then masturbate using only water or KY jelly as 
a lubricant and deposit their semen into a plastic container which will be provided to 
them. If they have collected the specimen at home, they will be asked to bring this 
container to the clinic within 1 hour after collecting their semen sample. This procedure 
is optional for participants.  
 
For Women: Women will be asked to self-collect cervical-vaginal secretion swabs. See 
Appendix D. This procedure is optional for participants.  
 
 

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12.1.8 Stool Collection (optional) 
Stool specimens will be collected from study participants (See Appendix G) for evaluation of 
the microbiome composition and its connection to immunologic and viral dynamics.  This 
procedure is optional for participants.  
12.1.9 Study Vaccine Injection 
Participant will be injected with the study drug. Administration of the intramuscular injection 
will be blinded. Subjects will complete 1 dose each of Influenza Vaccine, Pneumococcus 
vaccine and placebo injection every 12 weeks in random order.  
 
Fluarix (or Fluzone), a vaccine used to prevent infection by the Flu (influenza) virus, or 
 
Pneumovax 23, a vaccine used to prevent pneumococcal disease, or  
 
Placebo, a non-active saline solution that contains no vaccine. 
12.2 Laboratory Evaluations 
All laboratory procedures will be conducted in accordance with Good Clinical Laboratory 
Practices (GCLP) according to the DAIDS guidelines. 
A urine pregnancy test will be performed at screening in women who can become pregnant and 
will be repeated only if pregnancy is suspected. 
A rapid HIV test will be performed if no documentation of previous HIV infection is provided. 
Table 3 is a summary table of study assays, methods and time points. 
Table 3. Summary of Study Assays and Schedule of Evaluations 
 
Measurements 
Methods 
Sample 
Sampled Time-points (Days) 
Virologic 
(Aims 1+2) 
Cell-associated HIV RNA levels 
(dd)PCR 
PBMC 
(CD4+) 
0, 2, 4, 7, 14, 30 after each 
injection 
HIV DNA levels (pol and 2-LTR)  
(dd)PCR 
PBMC 
(CD4+) 
Cell-free HIV RNA levels 
SCA 
Blood 
plasma 
Cell-associated HIV RNA  env 
diversity* 
NGS  
PBMC 
(CD4+) 
  0 and time point with highest 
cell associated HIV RNA level 
after the active vaccination 
Cell-associated HIV DNA  env 
diversity* 
NGS 
PBMC 
(CD4+) 
Replication competent HIV DNA 
levels* 
Inducible 
RNA assay 
PBMC 
(CD4+) 
  0 and 30 after each active 
vaccine 
   
 
 
 
 
Immunologic 
Soluble inflammation markers  
ELISA 
Blood 
plasma 
0, 2, 4, 7, 14, 30 after each 
injection 

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(Aim 3) 
Vaccine-specific response 
ELISA 
Blood 
plasma 
Cellular inflammation markers 
Flow 
PBMC 
HIV-specific T-cell response 
Flow ICC 
PBMC 
 
 
 
 
 
 
Microbiome Cmposition 
r16s 
Sequencing 
Stool 
Day 0 
Incidental 
stimulation 
HHV quantification 
qRT-PCR 
GS 
 0, 14, 30 after each injection 
Incident Illness surveys 
Survey 
-- 
Legend: ddPCR: digital droplet PCR, SCA: single copy assay, PBMC: peripheral blood mononuclear cells, ELISA: 
enzyme linked immunoassay, Flow: flow cytometry, ICC: intracellular cytokine staining, qRT-PCR: quantitative real 
time PCR. GS: genital secretion. *The 5 participants with the highest sum levels of cell-associated HIV RNA within the 
30 days (five measures) following both active vaccinations will undergo these assessments. These same 5 
participants will be used for both of these analyses. 
12.2.1 Specimen Preparation, Handling and Shipping 
All laboratory procedures related to specimen preparation, handling and shipping will be 
conducted in accordance with Refer to DAIDS Policy Requirements for Laboratories in Clinical 
Trials, Guidelines for Good Clinical Laboratory Practice Standards and DAIDS Manual of 
Operational Procedures. The following AVRC Site SOPs are listed below and attached in the 
Appendix. 
 
Laboratory SOP “Transporting Specimens within and outside UCSD” (Appendix E) 
 
Laboratory SOP “Quality Management Plan” (Appendix F) 
12.2.2 Biohazard Containment 
Transmission of HIV and other 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 in the collection of clinical samples and the shipping and 
handling of all clinical samples and isolates for this study, as currently recommended by the 
Centers for Disease Control and Prevention in the United States, the WHO internationally and 
the National Institutes of Health.  
All protocol specimens will be shipped using packaging that meets requirements specified by 
the International Air Transport Association Dangerous Goods Regulations for UN 3373, 
Biological Substance, Category B, and Packing Instruction 650. Culture isolates, if obtained in 
this study, are to be shipped as specified for UN 2814 Category A Infectious Substances.  All 
infectious specimens will be transported using packaging mandated in the Federal Code of 
Regulations, CDC 42 CFR Part 72. Please also refer to individual carrier guidelines, e.g., 
FedEx, Airborne, for specific instructions. 
12.2.3 Total Blood Volume 
Participants will have blood drawn at each visit. Approximately 50 or 60 mL of blood will be 
drawn for the following tests and storage for future research: 

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 
CD4+/CD8+ T-cell Count (3 mL of blood).  
 
HIV-1 RNA (Viral Load) (6 mL of blood).  
 
Routine blood chemistry tests, a comprehensive metabolic panel and a complete blood 
count will be performed to determine overall health at baseline, day 14 and day 30 (we will 
collect 10 mL of additional blood). 
 
Blood will be collected for storage (6 mL of blood plasma and all PBMC). Some of the 
blood will be stored to perform future HIV testing and human genetic testing as well as to 
assess adherence to ART. Blood will be stored with the usual protectors of identity. The 
results of these tests will not be shared with the participant since these tests are not used 
for clinical management of their HIV. 
Participants will have approximately 50 or 60 mL of blood drawn at a time with an approximate 
total of 990 ml (4.5 cups) drawn for this study over the course of 8 months. 
12.3 Schedule of Procedures/Evaluations  
See Table 1 (page 12) for schedule of evaluations. 
12.3.1 Screening 
At screening a survey will establish inclusion and exclusion criteria. A screening CRF checklist 
and a screening survey will be completed by nursing staff. The screening visit will involve a 
discussion of vaccination history and medical history relevant to the inclusion and exclusion 
criteria. In addition documentation of plasma HIV-1 RNA and CD4+ T-cell counts will be 
recorded in the source documents and electronic CRF. A urine pregnancy test will be performed 
at screening in women who can become pregnant and will be repeated only if pregnancy is 
suspected. 
Written informed consent explained by trained clinical staff will occur before starting screening 
procedures. The baseline visit (study entry) and administration of the vaccine or vaccine-
placebo intervention will be performed within 30 days of the screening visit. 
12.3.2 Enrollment 
If the participant meets eligibility criteria, a participant identification number (PID) will be 
assigned to each individual screened and determined to be eligible for the study. PIDs will 
include a site code and three-digit extension. PIDs may not be reassigned even if the individual 
fails to enter the study.  The PID must be included on every CRF and blood sample. The site 
must maintain a master list of PIDs in a central location. Participant registration and 
inclusion/exclusion CRF must be completed. Study entry will occur within 30 days of screening.  
The study statistician will provide a randomization list to the pharmacist prior to study start. This 
list will be used in consecutive order to randomize participants at the time of enrolment. 
12.3.3 HIV Counseling and Testing 

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No HIV testing will occur, as participants must have documented HIV-1 infection more than 365 
days prior to enrollment (HIV antibody or viral load positive). Participants are required to be on 
ART for at least 48 weeks.  Counseling can be provided to any participant.  
12.3.4 Follow-up 
 
12.3.4.1: Baseline assessments [Day 0 of each cycle]  
(These visits will last about 1.5 hours): At study entry the subject should complete all 
assessments and clinical evaluations with blood sample for banking. Antibody levels for 
each tested vaccine, cell-associated HIV RNA levels in blood plasma and HIV DNA 
levels in PBMC should be measured. An optional genital secretion and an optional stool 
sample will also be collected at baseline assessments. Refusal to provide genital 
secretion and/or stool sample will not result in termination from the study. 
Placebo or vaccine injections: Will be performed by study nursing personnel based on 
stratification delivered by study pharmacist at baseline. 
 
12.3.4.2 On study evaluations post-vaccination [Days 2, 4, 7, 14 & 30]  
(These visits will last about 1 hour): At the time of each injection and five sampling time 
points afterwards, the following should be measured: cell-associated and cell-free HIV 
RNA levels, HIV DNA levels, vaccine-specific antibody levels, soluble and cellular 
inflammatory and proliferation markers, and HIV specific T-cell response. Genital 
secretion samples will be collection at post-vaccination evaluations 14 and 30 only. 
12.3.5 Early Termination Visit 
If a subject is unable to complete all of the study visits, they will be asked to return to the clinic 
to for a study discontinuation visit.  The following procedures will be performed: 
 
brief physical exam and concomitant medication questionnaire 
 
blood drawn for routine safety tests to check your blood count, liver and kidney function, 
CD4+cell count, viral load and storage for future testing 
 
survey about recent illnesses and symptoms 
 
optional collection of genital secretion 
CRITERIA FOR EARLY TERMINATION. 
 
Active drug or alcohol use or dependence that, in the opinion of the investigator, would 
interfere with adherence to study requirements. 
 
The subject becomes pregnant during the course of the study. 
 
Failure by the subject to attend scheduled clinic visits may result in discontinuation. 
 
Request by the subject to withdraw from the study. 
 
The participant’s HIV therapy fails. 

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 
Request of the primary care provider if s/he thinks the study is no longer in the best 
interest of the subject. 
 
Clinical reasons believed life threatening by the physician. 
 
Subject judged by the investigator to be at significant risk of failing to comply with the 
provisions of the protocol as to cause harm to self or seriously interfere with the validity of 
the study results. 
12.3.6 Pregnancy Visit 
Subjects will be screened for pregnancy at the screening visit and at subsequent follow up visits 
if they are thought to have become pregnant. If a participant becomes pregnant during the 
course of the study they will be asked to complete the early termination visit as described above 
in section 12.3.5. 
12.3.7 Final Study Visit 
The final study visit will take place as the day 30 follow up visit at the completion of the third and 
final cycle. This visit will have the same procedures and evaluations of all other follow up visits. 
13. ASSESSMENT OF SAFETY 
13.1 Safety Assessment Overview 
Although serious adverse events are not expected in this study, any serious adverse events 
(SAEs) will be reported in accordance with DAIDS recommendations and local UCSD IRB 
requirements. 
13.2 Adverse Event Procedures and Reporting Requirements 
All SAEs will be documented on the SAE Reporting Form within five working days of site 
awareness of the event and submitted to the Safety Monitoring Committee (SMC). 
14. CLINICAL MANAGEMENT 
14.1 Clinical Management of Adverse Events 
The study investigators will review all adverse events (AEs) from subjects followed as part of the 
project.  Unanticipated and anticipated toxicities will be graded according to the Division of AIDS 
Table for Grading the Severity of Adult and Pediatric Adverse Events, Corrected Version 2.1 
dated July 2017.  
Subjects with AE will be followed until the event has resolved or stabilized.  Out of range 
laboratory values will be followed closely until resolved.  In every situation, the study 
investigators will assess if it is safe for the subject to continue in the study.  A physician is on-
call at the AVRC 24/7 for evaluation and management of study-related emergencies or AEs. 
Any new, unusual, or unexpected toxicities will be reported to the Human Subjects Committee, 
other investigators participating in the trial, the FDA, the NIAID/DAIDS, and other regulatory 
agencies or entities as specified by NIAID, FDA or UCSD policies and procedures. AEs 

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assessed as related to non-study drugs (concomitant medications) will be handled according to 
the relevant package inserts and the best medical judgment of the site investigator. 
Because all proposed vaccines are FDA-approved for clinical use and clinically recommended 
for HIV-infected individuals, a low likelihood for study-related adverse events is anticipated. 
Serious adverse events (SAEs) will be defined using standard FDA guidelines. All SAEs 
occurring at the clinical site will be reported to the UCSD IRB using standard reporting time 
frames and IRB requested formats. All higher-grade toxicities will be reported immediately to Dr. 
Smith (or Dr. Little if Dr. Smith is unavailable) and follow-up monitoring or clinical intervention 
with the participant’s healthcare provider will be performed in consultation with the clinical 
investigators and nursing staff. Additionally, the clinical investigator team will review safety 
report summaries from the clinical nursing staff every three months.  
14.1.1. Safety Monitoring Committee  
Because we are using standard clinical vaccines that are clinically recommended for HIV-
infected individuals, the need for a formal DSMB is not anticipated, but the protocol team will 
convene a safety monitoring committee (SMC) of at least three independent experts including 
one statistician to perform an interim safety review at least once annually. The SMC will review 
study progress and data to ensure patient safety, as well as feasibility of completion of the study 
within the grant-funding period. After the review, the SMC will be asked to make a 
recommendation for study continuation, modification or conclusion. The SMC is composed of 
clinical trials experts and a statistician who are not investigators in the specific project and have 
no conflict of interest with the clinical study. Members of the SMC include Drs. Benson, Karris 
and Jain, who are all respected experts both in the care and treatment of HIV-infected patients 
and in the design, conduct, and analysis of clinical trials.  
All members of the SMC will receive an introduction to the study trial and will have a clear 
understanding of how study investigators will approach early study termination for safety 
reasons. The Safety Monitoring Committee will review the safety and welfare of participants and 
confidentiality of data. The committee’s responsibilities include: reviewing study protocols 
including plans for data safety monitoring, evaluating the progress of the intervention trial, 
evaluating participant risk versus benefit, performing assessments of any unexpected factors 
that can affect the safety of study participants. Following the review, the SMC will provide writing 
recommendations to the PI, who will distribute these to the study team, local IRB and the DAIDS 
Program Officer.  
14.1.2. Most frequent Adverse Events 
Participants will receive two vaccines and a vaccine placebo. This procedure may be associated 
with pain, bruising, bleeding, infection, dizziness, or fainting. This will be managed by strict 
adherence to aseptic techniques, and use of experienced clinical nurses administering the 
injections. The vaccines used in this study are generally safe for HIV infected individuals with 
CD4 > 200 cells/ul. Occasionally, subjects can develop soreness, swelling, or redness at the 
injection site. Some vaccines are associated with fever, rash, and achiness. Serious side effects 
are rare, but may include life-threatening allergic reaction or seizure. Risks associated with each 
of the specific vaccines used include: 

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 
Pneumococcal vaccine: About half of people who get Pneumovax®23 have mild side 
effects, such as redness or pain where the shot is given. Less than 1% of persons 
develop a fever, muscle aches, or more severe local reactions. Mild fever, fatigue, 
headache, chills, and muscle pain have also been reported. Life-threatening allergic 
reactions are very rare. 
 
 
Influenza vaccine: Inactivated parental vaccine that is recommended yearly to all HIV-
infected individuals. Vaccination is most effective among persons with CD4 of >100 
cells/µL and HIV RNA of <30,000 copies/mL, which is in the eligibility requirements for 
the study. In adults, the most common local and general adverse events were pain and 
redness at the injection site (20-50%), muscle aches (23%), fatigue (20%), and 
headache (19%). Vaccination should be avoided by people with known hypersensitivity 
to any ingredient of the vaccine, and persons previously diagnosed with Guillain-Barré 
syndrome. Life-threatening allergic reactions from any vaccine are very rare. 
14.2 Other Disease Events 
Any disease events or complications that occur during study visits will be handled according to 
the discretion of the study clinician and subsequently subjects will be referred to an appropriate 
primary physician. 
14.3 Pregnancy 
A urine pregnancy test will be performed at screening in women who can become pregnant and 
will be repeated only if pregnancy is suspected. If a subject becomes pregnant during the study, 
then an early termination visit will be scheduled for them and they will be taken off study. 
Reason for excluding pregnant and breastfeeding women: To date, there are still insufficient 
safety data for the use of pneumococcal and influenza vaccines in pregnant women and its 
administration is recommended only if clearly indicated for medical reasons. Additionally, as part 
of this study we will administer the vaccines outside the regular schedule to determine the effect 
of immune-stimulation on HIV transcription. It is not known if vaccine administration outside the 
clinically recommended schedule can harm unborn babies or might increase HIV transcription 
and consequently might affect the risk of vertical HIV transmission. If you are a woman and 
having sex that could lead to pregnancy, you must agree not to become pregnant 
14.4 Treatment Failure 
HIV treatment failure will be routinely tested by HIV RNA viral loads. If a participant’s HIV 
treatment fails (two consecutive viral load measurements of >500 copies/ml) or their ART 
regimen is interrupted, then an early termination visit will be scheduled and the participants will 
be taken off study. 
14.5 Criteria for Discontinuation 
 
Active drug or alcohol use or dependence that, in the opinion of the investigator, would 
interfere with adherence to study requirements. 
 
The subject becomes pregnant during the course of the study. 

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 
Failure by the subject to attend scheduled clinic visits may result in discontinuation. 
 
Request by the subject to withdraw from the study. 
 
The participant’s HIV therapy fails. 
 
Request of the primary care provider if s/he thinks the study is no longer in the best 
interest of the subject. 
 
Clinical reasons believed life threatening by the physician. 
 
Subject judged by the investigator to be at significant risk of failing to comply with the 
provisions of the protocol as to cause harm to self or seriously interfere with the validity 
of the study results. 
15. STATISTICAL CONSIDERATIONS 
15.1 Overview and General Design Issues 
The proposed study is a randomized double-blind controlled trial conducted over 28 weeks. 
Randomized interventions will be injections of Influenza vaccine, Pneumococcal vaccine, and 
Placebo. Each participant will receive each injection but in a randomized order.  
Vaccines: Influenza (Fluarix®, GSK), Pneumococcal (Pneumovax®23, Merck), and placebo 
(sterile saline injection). 
Primary endpoint: average level of CD4+ T-cell-associated HIV RNA transcription measured at 
day 7 after each injection. 
Study Arms: Each subject will take Influenza (Fluarix®, GSK), Pneumococcal (Pneumovax®23, 
Merck), and placebo. Randomization will determine the order in which the subjects receive the 
injections. There are six potential study arms, one for each possible combination in which 
someone could receive the injections.  
Study population: This study will enroll 56 HIV-infected individuals between the ages of 18 and 
65 who started ART during chronic infection. These subjects will be recruited from the UCSD 
AVRC. Eligible participants will have CD4 >250 cells/μl at enrollment and a CD4 nadir >100 
cells/μl. We chose to limit enrollment to younger individuals with higher CD4 counts to enhance 
the likelihood of having a more robust immune response to any given vaccine. We limited 
enrollment to individuals who started ART during chronic infection (>365 days from estimated 
date of infection) to enhance the likelihood of having a large replication competent viral 
reservoir, thus increasing our chances to observe an effect of vaccine administration on HIV 
RNA transcription [96]. To reduce variability in vaccine stimulation (between antigen naïve 
versus experienced participants), we will also limit participation to individuals who have 
documented or reported vaccine history consistent with previous influenza and pneumococcal 
vaccination.   

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Sample collection: Study evaluations will be based on three 30 day cycles (Influenza vaccine, 
Pneumococcus vaccine, Placebo in random order) over 28 weeks of the RCT. Pre injection: one 
paired blood, optional stool and optional genital secretion sample will be collected before each 
injection. Post-injection: Blood samples will be collected on days 2, 4, 7, 14 and 30 after each 
injection. Optional genital secretion samples will be collected on days 14 and 30 after each 
injection 
15.2 Study Endpoints 
15.2.1 Primary Endpoint 
Average change in level of CD4+ T-cell-associated HIV RNA transcription from baseline to 7 
days after each injection. 
15.2.2 Secondary Endpoints:  
1. Average change in level of CD4+ T-cell-associated HIV RNA transcription from 
baseline to 2, 4, 14 and 30 days after each injection. 
2. Average change in levels of HIV DNA from baseline to 2, 4, 7, 14 and 30 days 
after each injection. 
3. Determine selective vs. nonselective HIV RNA production from HIV DNA (by next 
generation sequencing and panmixis tests) among the five participants with the 
highest levels of cell associated HIV RNA following active vaccination. 
4. Average change in levels of immune activation (soluble and cellular markers) 
from baseline to 2, 4, 7, 14 and 30 days after each injection. 
5. Average change in levels of replication competent HIV DNA (by inducible RNA 
assay) among the five participants with the highest levels of cell associated HIV 
RNA following active vaccination. 
6. Average change in levels of HIV specific T-cell immune response from baseline 
to 30 days after each injection. 
7. Stimulation of HIV transcription by incidental illness, human herpesviruses 
replication and microbial translocation over the course of the study. 
8. Possible immunemodulatory effect of microbiome composition  
9. Vaccine administration will be safe for all included participants.  
15.3 Study Objectives and Hypotheses 
Primary Objective 
Determine the impact of Pneumococcus and Influenza vaccines on the HIV transcriptional 
activity in individuals who are virologically suppressed for at least 48 weeks on ART. 
Secondary Objectives 
Aim 1a: Determine the timing and duration of increased HIV transcription following active 
vaccination. 
Aim 1b: Determine if vaccination selectively or non-selectively activates HIV transcription 
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Aim 2: Determine if active vaccination influences levels of cell associated HIV DNA or 
replication competent provirus. 
Aim 3a: Determine to what degree immune stimulation with an Influenza or Pneumococcal 
vaccine can induce HIV transcription during ART. 
Aim 3b: Determine if immune stimulation associated with active vaccination is associated 
with reduction in number of HIV DNA populations. 
Aim 3c: Determine if HIV-specific, vaccine-specific, or bystander immune stimulation is 
associated with HIV transcription. 
Aim 4: Determine how genital shedding of human herpesviruses, microbial translocation and 
incident illnesses stimulate the immune system and influence levels HIV transcription. 
Aim 5: Determine how dysbiosis might contribute to immune stimulation and influence HIV 
transcription. 
15.4 Sample Size Considerations 
Examples of Power: 
Primary Hypothesis (stated under the alternative): On average, participants will have a higher 
absolute increase in levels of cell-associated HIV RNA in CD4+ T-cells seven days after 
receiving either Pneumococcal or Influenza vaccinations, when compared to seven days after 
receiving placebo. We have chosen to provide estimates of power on the one sampled time 
point that is likely to show the largest effect, based on our literature review. All other sampled 
time points following vaccination will be evaluated as well, but to be conservative, we focus on 
and present power for one sampled time point for our primary hypothesis. 
 
Assumption: This is a cross-over study with 56 individuals receiving Placebo, 
Pneumococcus, and Influenza injections in random order (6 possible orders) with 
sufficient wash-out period in between (minimum 6 weeks) vaccinations. We expect that 6 
individuals drop out, resulting in a fully evaluable sample size of 50 individuals. Pilot data 
were generated from the previous Spanish Study [1] to provide estimates of standard 
deviations (SD) of change in cell associated HIV RNA levels. From these data, we used a 
SD of 537 HIV RNA copies/106 CD4 T-cells for Influenza and 714 for Pneumococcus, and 
a pooled estimate of 625.  
 
Effect sizes: In Figure 3, we present the effect sizes that we have 80% power to observe 
at alpha levels of 0.05 (solid line) and 0.025 (dashed line). Since we will compare 
observations in the same individual, we will use a paired t-test that compares increases in 
cell-associated HIV RNA in CD4+ T-cells after 7 days for each of the active vaccine 
injections (Pneumococcus or Influenza) to 7 days after the placebo injection. For 
example, if the SD of the differences between active and placebo injection measures is 
625 HIV RNA copies/106 CD4 T-cells, then we will have 80% power at alpha 0.05 to 
detect an effect size of 253 HIV RNA copies/106 CD4 T-cells absolute increase or 
decrease (two-sided test) in cell associated HIV RNA levels following active vaccination 
versus placebo, which is well-within the range observed in our preliminary data. 

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Figure 3 Effect sizes with 80% power to test primary hypothesis
Legend: SD: standard deviation (x axes), effect size (y axes) 
 
Example of Secondary Hypothesis (stated under the alternative): On average, participants will 
have a greater absolute maximum decrease in levels of total CD4+ T-cell-associated HIV DNA 
measured 2-30 days after receiving either Pneumococcal or Influenza vaccinations, when 
compared to 2-30 days after receiving placebo. 
 
Assumption: This is a cross-over study of three treatments (with 6 possible orders) with a 
fully evaluable sample size of 50 individuals. We used our data from 199 subjects who had 
CD4+ T-cell associated HIV DNA levels measured after 48 weeks after virologic 
suppression on ART to provide estimates of effect sizes. In these samples, we found a 
stable average level of 41.01 of cell associated HIV DNA copies/106 PBMC with a SD of 
75.01. 
 
Effect sizes: In table 4, we present estimates of effect sizes that we have 80% power to 
observe at an alpha level of 0.05 using a paired t-test that compares maximum decreases 
(over 2-30 days) in levels of total CD4+ T-cell-associated HIV DNA for each of the active 
vaccine injections to maximum decreases (over 2-30 days) after the placebo injection, 
assuming the presented SD. For example, if the SD of the change is similar to the SD from 
our preliminary data (~75), then we have 80% power to detect an average maximum 
decrease of 30.3 HIV DNA copies/106 PBMC following active vaccination, as compared to 
after placebo. 
 
218
253
289
241
280
320
150
200
250
300
350
500
550
600
650
700
750
SD of Difference
alpha=0.025
alpha=0.05

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Figure 4. Power to see a difference in HIV DNA levels following vaccination 
versus placebo 
 
Example of Secondary Hypothesis (stated under the alternative): Participants will have a greater 
absolute maximum increase in levels of (a) inflammatory cytokines, (b) T-cell activation and 
proliferation, (c) HIV-specific immune responses, and (d) vaccine-specific immune responses 
over the 30 days of observation after receiving either active vaccination, when compared to 
maximum measures taken over the 30 days of observation after receiving placebo. 
Assumptions and effect sizes: Assuming 50 fully evaluable participants and that the SD of the 
differences in increase over days 2-30 in activated CD4+ T-cells is 4.5% (based on data from 
our CMV studies[97]), then we have 80% power for alpha levels of 0.05 using a paired t-test to 
compare maximum increases (over 2-30 days) of a 40% increase (or decrease) in levels of 
activated CD4+ T-cells for each of the active vaccine injections as compared to placebo, which 
is consistent with changes seen in CD4+ T-cell activation during genital shedding of CMV 
shedding [97]. 
15.5 Enrollment/Randomization/Blinding Procedures 
Enrollment: A screening survey will establish inclusion and exclusion criteria. A screening CRF 
checklist and a screening survey will be completed by nursing staff Written informed consent 
explained by trained clinical staff will occur before starting screening procedures. The baseline 
visit (study entry) and administration of the vaccine or vaccine-placebo intervention will be 
completed within 30 days of screening. A PID will be assigned to each individual screened for 
the study. PIDs will include a site code and three-digit extension. PIDs may not be reassigned 
even if the individual fails to enter the study.  The PID must be included on every CRF and 
blood sample. The site must maintain a master list of PIDs in a central location. Study entry 
must be completed within 30 days after participant registration and inclusion/exclusion CRF 
have been completed. 

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Randomization: The study statistician will provide a randomization list to the pharmacist prior to 
study start. Subjects will be considered randomized at the time they have completed enrollment 
procedures and have received their initial vaccination. Subjects who do not receive any 
vaccination will not be considered enrolled and will be replaced and will have no follow-up 
evaluations performed. Subjects who withdraw after receiving the vaccine or vaccine-placebo 
intervention will be asked to complete the early termination visit. 
Blinding: In order to achieve double blind administration of the vaccines, each drug or placebo 
will first be transferred to generic, non-labeled syringes before being delivered to the clinical 
staff who will administer the injection using aseptic techniques. This preparation (along with 
preparation of the sterile saline placebo) will take place in the AVRC pharmacy. Unblinding of 
the study arms for data analysis of primary and secondary endpoints will occur around 44-48 
months (after all participants have completed the entire study follow-up). 
15.6 Maintenance of Trial Treatment Randomization Codes  
Trial treatment randomization codes will be kept by Study Pharmacist (Dr. Muttera) and Study 
Biostatistician (Dr. May). Dr. Muttera is located in the AVRC Pharmacy (220 Dickinson Street, 
Suite A. San Diego, CA 92103) and her office phone number is (619) 543-2688. 
15.7 Participant Enrollment and Follow-up 
Baseline assessments [Day 0 of each cycle]  
At study entry the subject should complete all assessments and clinical evaluations with blood 
sample for banking. Antibody levels for each tested vaccine, cell-associated HIV RNA levels in 
blood plasma and HIV DNA levels in PBMC should be measured. An optional genital secretion 
and optional stool samples will also be collected at baseline assessments. 
Placebo or vaccine injections: Will be performed by study nursing personnel using the generic 
non-labelled syringe delivered by study pharmacist at baseline. 
On study evaluations post-vaccination [Days 2, 4, 7, 14 & 30]  
At the time of each injection and five sampling time points afterwards, the following will be 
measured: cell-associated and cell-free HIV RNA levels, HIV DNA levels, HIV levels, antibody 
levels, soluble and cellular inflammatory and proliferation markers, and HIV specific response. 
Optional genital secretion samples will be collected at post-vaccination evaluations 14 and 30 
only. 
15.8 Data and Safety Monitoring 
15.8.1 Planned Interim Analyses and Stopping Guidelines 
15.8.1.1 Interim safety review: 
The study investigators Drs. Smith, Little, Gianella, May and the study nurses will review the 
individual subject records every 3 months, including consent forms, case report forms 
(CRFs), supporting source data, laboratory specimen records and medical records 
(physicians’ progress notes, nurses’ notes) to ensure protection of study subjects, 
compliance with the protocol and accuracy and completeness of records.  The study 
investigators will also inspect sites’ regulatory files to ensure that regulatory requirements are 

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being followed and the site pharmacy to review product storage and management. If 
requested, the site Investigators will make study documents (e.g. consent forms, drug 
distribution forms, CRFs) readily available for inspection by the local IRB, the site monitors, 
the FDA, the NIAID, and the OHRP for confirmation of study data. 
The study investigators will review all safety events from subjects followed as part of the project.  
Adverse Events (AE) will be graded using the standard Division of AIDS Table for Grading the 
Severity of Adult and Pediatric Adverse Events, Corrected Version 2.1 dated July 2017.().  
Subjects with AE will be followed until the event has resolved or stabilized.  Out of range 
laboratory values will be followed closely until resolved.  In every situation, the study 
investigators will assess if it is safe for the subject to continue in the study.  The clinical 
investigator team (Drs. Smith and Little) will have a 24-hour call system (using the medical 
center pager system) so subjects can access a study physician at any time to answer questions 
or review potential unforeseen consequences.  
Because all proposed vaccines are standard clinical vaccines and clinically recommended for 
HIV-infected individuals, a low likelihood for study-related adverse events is anticipated. Serious 
adverse events (SAEs) will be defined using standard FDA guidelines. All SAEs occurring at the 
clinical site will be reported to the UCSD IRB using standard reporting time frames and IRB 
requested formats. All higher-grade toxicities will be reported immediately to Dr. Smith (or Dr. 
Little if Dr. Smith is unavailable) and a plan for follow-up monitoring or clinical intervention with 
the participant’s healthcare provider will be performed in consultation with the clinical 
investigators and nursing staff. Additionally, the clinical investigator team will review safety 
report summaries from the clinical nursing staff every three months.  
15.8.1.2 Safety Monitoring Committee  
Because we are using standard clinical vaccines that are clinically recommended for HIV-
infected individuals, the need for a formal DSMB is not anticipated, but the protocol team will 
convene a safety monitoring committee (SMC) of at least three independent experts including 
one statistician to perform an interim safety review once the study has reached 50% accrual or 
after approximately one year has passed, whichever occurs first. The SMC will review study 
progress and data at least annually to ensure patient safety and feasibility of completing the 
study within the grant-funding period. After the review, the SMC will be asked to make a 
recommendation for study continuation, modification or conclusion. The SMC is composed of 
clinical trials experts and statisticians who are not investigators in the specific project and have 
no conflict of interest with the clinical study. Members of the SMC include Drs. Benson, Karris 
and Jain, who are all respected experts both in the care and treatment of HIV-infected patients 
and in the design, conduct, and analysis of clinical trials.  
All members of the SMC will receive an introduction to the study trial and will have a clear 
understanding of how study investigators will approach early study termination for safety 
reasons. The Safety Monitoring Committee will review the safety and welfare of participants and 
confidentiality of data. The committee’s responsibilities include: reviewing study protocols 
including plans for data safety monitoring, evaluating the progress of the intervention trial, 
evaluating participant risk versus benefit, performing assessments of any unexpected factors 

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that can affect the safety of study participants. The SMC will provide writing recommendations 
to the PI, who will distribute these to the study team, local IRB and the DAIDS Program Officer  
15.8.1.3 Interim Efficacy Review:  
A SMC will review trial data when at least 50% of subjects have completed the trail. There will 
be no formal interim assessment of efficacy. Safety data will be evaluated to determine if the 
study should continue as planned, should be modified to ensure patient safety or should be 
stopped.   
15.8.2 Analysis Plan 
All formal analyses of the data will be performed at the end of the study with no real time clinical 
decisions based on intermediate results. We will utilize a comprehensive AVRC Data 
Management System developed through our CFAR that can handle all dynamic aspects of 
clinical research studies including electronic case report form (eCRF) creation and editing, 
protocol design interfaces, online data entry and QA workflows, real-time accrual monitoring, 
multi-study data integration, role-based user permissions, integrated specimen processing 
workflows, an auditing system to track changes to both forms and data, and data visualization 
and export tools. 
Sample and data collection from study participants will occur from study months 6-36 of the 
study. Unblinding of the study arms for data analysis of primary and secondary endpoints will 
occur around 44-48 months (after all participants have completed the entire study follow-up), 
and the final analysis of the trial will take place between 48 and 60 months. 
Analysis of the Primary Hypothesis: We will determine whether average changes in levels of 
cell-associated HIV RNA in CD4+ T-cells are associated with administration of active vaccine or 
placebo using two-sided paired t-tests with alpha level of 0.05 for each of the two tests 
(influenza versus placebo and pneumococcus versus placebo).  
Secondary analyses: Since subjects will receive multiple injections, we will determine if 
changes in cell associated HIV RNA in CD4+ T-cell levels are sustained over the weeks 
following vaccination using graphical and descriptive statistical methods. We will also explore 
the effect of the vaccines on the size of cell free and cell associated HIV RNA populations using 
linear mixed effects models and consider incorporating interactions with time to evaluate the 
change in effects of the vaccines over time. 
Additional analyses are described in the Statistical Analysis Plan (SAP).  
16. DATA HANDLING AND RECORDKEEPING 
16.1 Data Management Responsibilities 
Both archived and prospectively collected and generated data from enrolled subjects will be 
organized into our central database securely hosted at UCSD, along with all available linked 
demographic, geographic, clinical, and behavioral data. Confidentiality is maintained by linking 
data only to the unique ID, and by not storing any personal identifying information in result 
reporting systems. 

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16.2 Essential Documents and Access 
All essential documents will be maintained by the clinical data manager who will also control 
permissions and access to the sources documents and essential data. 
16.3 Quality Control and Quality Assurance 
To ensure an accurate and complete database, key forms will be monitored for discrepancies 
using automated logical checks. If problems are identified, such as missing clinical data, this will 
be communicated to study investigators. A Study Status Report including number of participants 
enrolled or off-study report (list of participants off-study; reasons off-study), SAE report (list of 
participants; diagnoses), study related AEs ≥ grade 3 will be prepared quarterly. Clinical 
evaluations and specimen collections will be conducted according to the specifications in the 
study protocol. The study protocol, including materials and informed consent forms, will undergo 
review and approval by the UCSD IRB and local Ethics Committee prior to implementation, and 
all participants will be provided with education, counseling and information, and undergo 
informed consent for all procedures. 
17. CLINICAL SITE MONITORING 
The National Institute of Allergy and Infectious Diseases (NIAID) contracted site monitors will 
visit the clinical research site (AVRC) to review participants records, including informed consent 
forms, CRFs, medical records (e.g., physicians’ progress notes, nurses’ notes, individuals’ 
hospital charts), and laboratory records to ensure protection of study participants, compliance 
with the EC/IRB approved protocol/amendments, and accuracy and completeness of records. 
The monitors will review sites’ regulatory files to ensure that local regulatory requirements, in 
addition to U.S. Federal regulations, are being followed. They will also inspect sites’ pharmacies 
and review product storage, management and accountability records 
18. HUMAN SUBJECTS PROTECTIONS 
18.1 Institutional Review Board/Ethics Committee 
This protocol and the informed consent document and any subsequent modifications will be 
reviewed and approved by the UCSD Institutional Review Board responsible for oversight of the 
study. Written informed consent will be obtained from the subject by a member of the study 
staff. At the time potential subjects contact us regarding the study, any questions they may have 
will be answered by a member of the study staff. If the potential subject is still interested in 
participating, a clinic visit will be scheduled. The subject will be informed of the time that needs 
to be allotted for their first visit in which the informed consent will be administered.  
18.2 Vulnerable Participants 
18.2.1 Pregnant women and fetuses: Women who are pregnant will be excluded from 
the study and women who become pregnant while on the study will be asked to 
complete an early termination visit and will be removed from follow up. 

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18.3 Informed Consent 
18.3.1 Informed Consent Process 
Written informed consent (previously approved by UCSD’s IRB) explained by trained 
clinical staff will occur before starting screening procedures. Study entry and 
administration of the vaccine or vaccine-placebo intervention will occur within 30 days of 
completion of screening procedures. 
The informed consent will describe the purpose of the study, the procedures to be 
followed, and the risks, and benefits of participation. This information will be explained to 
the study subject in a face-to-face setting by the individual who consents the participant. 
Participants will be encouraged to ask questions throughout the consent process and 
encouraged to discuss their participation with trusted advisors, such as family members, 
close friends, etc. Participants will be allotted sufficient time to consider whether or not to 
participate in the research study. After allowing the potential participant time to read the 
informed consent the study staff and/or investigator will answer and address any 
questions or concerns the participant may have. Once all questions and concerns have 
been addressed and the person wishes to participate, they will be asked to sign the 
informed consent.  
Also, during the consent process, the Health Insurance Portability and Accountability Act 
(HIPAA) Authorization will addressed. A copy of the consent and HIPAA Authorization 
form as well as the Notice of Privacy Practices booklet will be given to the subject. 
18.3.2 Documentation of Informed Consent 
All signed informed consent documents will be kept on file at the AVRC under the 
supervision of the AVRC regulatory office. 
18.3.3 Stored Samples and Associated Data Considerations 
18.3.3.1 Data Security: Any data collected as part of this study that is stored at the 
AVRC and/or is transferred via the internet will follow our data security process as 
outlined below. 
With the fast-developing technology, dependable and comprehensive data security 
measures are key components to defy the perceived threats of Internet hackers and 
accidental disclosure of confidential information. In the following we provide a summary 
of the key features pertinent to this project. 
 
An anonymous participant identification number is used for all data collection, 
recording, and submission to the project database.  
 
Data that contain any participant identifiers (e.g., name or contact information) other 
than the unique identifier are password protected and accessible only to staff 
members whose job requires knowledge of such data.  

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 
Laboratories are instructed not to disseminate any participant identifiers in any 
communications with, or data submissions to, any other AVRC collaborators. Any 
data transfer over the Internet uses encryption.  
 
Data transfer and all Web-based utilities use secure access (user and server 
authentication, 128-bit SSL encryption). This type of encryption is the same as is 
used for Web-based transactions that involve credit cards or Web banking.  
18.3.3.2 Research Laboratory Specimen Identification Policy: All research laboratory 
specimens leaving the AVRC to an outside laboratory will be de-identified. See the 
following procedure: 
 
The Lab Manager will create a study specific AVRC internal lab requisition. The 
requisition will be saved and accessible via the AVRC internal computer system’s 
shared drive. 
 
Each research nurse will access and print the study specific requisition(s) via the 
shared drive. 
 
Each research nurse will then complete the study specific requisition with the 
subject’s name, DOB, medical record number (MR#), PID, AVRC number and study 
week. 
 
The requisition will then be delivered by the research nurse to the AVRC lab. 
 
The AVRC laboratory staff will then complete the appropriate form for the 
corresponding laboratory to which the specimen will be sent, using two coded 
identifiers,  the subject’s PID number (in the name field) and AVRC number (in the 
medical record number ‘s field)  
 
The AVRC laboratory staff will prepare and label specimen tubes using the same two 
coded identifiers,  the subject’s PID number (in the name field) and AVRC number (in 
the medical record number’s field). No personal health identifiers will be included on 
the specimen label (i.e., no name, initials, DOB, MR#, etc).    
 
Prior to the blood draw, the phlebotomist will verbally verify the subject’s name and 
DOB. The phlebotomist will confirm the coded specimen tube(s) identifiers with the 
coded form identifiers. 
 
Coded specimens are transported to the appropriate lab either by AVRC staff or 
shipped via FedEx, under IATA regulations. 
 
All study specific completed AVRC internal lab requisitions will be retained in a 
locked and secured area for a period of six months and thereafter shredded.  
 
Samples will be kept frozen and stored for an indefinite length of time.   

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All stored samples are accessible only to the AVRC laboratory personnel and the 
appropriate study members.  Samples are stored under the coded identifiers as detailed 
above frozen and in freezers equipped with locks until they are shipped to the central 
laboratory under contract with the sponsor.  The freezers are located in the AVRC and 
CTF building behind locked doors with cypher or key pad entry. 
The stored samples are shipped as outlined above and are then secured under the 
sponsor’s SOPs for storage of human biological samples. 
18.4 Risks 
Privacy and Confidentiality: Although we will make every effort to protect the subject’s privacy 
and confidentiality it is possible that the subject’s status could become known to others. This 
could cause problems between the subject and the subject’s family and/or community and could 
cause the subject to be discriminated against.   
Personal Questions Risks: Subjects will be asked questions about personal issues during this 
study. There may be questions about their mood, about sexual functioning, about drug use, etc. 
These types of personal questions may make some subjects uncomfortable.  
Reporting Requirements: All cases of HIV, hepatitis B and hepatitis C must be reported to the 
county public health department. According to California state law, study staff is required to give 
public health department staff the participant’s name, contact information and treatment records 
for the hepatitis if requested.  
Blood Draws: The subject may experience temporary discomfort from the blood draws. The 
needlesticks may cause local pain, bleeding, bruising and swelling, as well as lightheadedness, 
dizziness and rarely, blockage of the vein, fainting and/or a local infection.  
Vaginal Secretions Collection Risks: Women may have some discomfort or feel embarrassed 
when they give vaginal fluid sample.  
Semen Collection Risks: Men may have some discomfort or feel embarrassed when they give 
semen sample.  
Stool: Subjects may have some discomfort or feel embarrassed when they give stool sample.  
Risks of Study Drugs: The drugs used in this study may have side effects, some of which are 
listed below. Please note that these lists do not include all the side effects seen with these 
drugs. These lists include the more serious or common side effects with a known or possible 
relationship. It is very important that subjects tell their study doctor of any changes in their 
medical condition while taking part in the study. At any time during the study, if subjects believe 
they are experiencing any of these side effects, they have the right to ask questions on possible 
and /or known risks.  There is a risk of serious and/or life threatening side effects when non-
study medications are taken with the study drugs. For their safety, subjects must tell the study 
doctor or nurse about all medications they are taking before they start the study and must ask 
approval for taking any new medication while they are on the study.  

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Risks of Pneumococcal (Pneumovax 23) vaccine: About half of people who get Pneumovax®23 
have mild side effects, such as redness or pain where the shot is given. Less than 1% of 
persons develop a fever, muscle aches, or more severe local reactions. Mild fever, fatigue, 
headache, chills, and muscle pain have also been reported. Life-threatening allergic reactions 
are very rare. Serious side effects include: Difficulty breathing, wheezing, rash and hives.  
These may be signs of an allergic reaction. Participants should contact the study staff 
immediately if they experience these symptoms. 
Risks of Influenza (Fluarix or Fluzone) vaccine: Common side effects include: Low fever, chills, 
redness or pain where the vaccine was injected, headache, and joint/muscle pain.  
Serious side effects include: Severe weakness or unusual feeling in arms or legs, high fever, 
seizure, convulsions, or unusual bleeding. These may be signs of an allergic reaction. Subjects 
should contact the study staff immediately if they experience these symptoms.  
Inactivated parental vaccine is recommended yearly to all HIV-infected individuals. Vaccination 
is most effective among persons with CD4 of >100 cells/µL and HIV RNA of <30,000 copies/mL, 
which is in the eligibility requirements for the study. In adults, the most common local and 
general adverse events after flu vaccine administration were pain and redness at the injection 
site (20-50%), muscle aches (23%), fatigue (20%), and headache (19%). Vaccination should be 
avoided by people with known hypersensitivity to any ingredient of the vaccine, and persons 
previously diagnosed with Guillain-Barré syndrome. Life-threatening allergic reactions from any 
vaccine are very rare. 
Reproductive Risks: It is not known if drug combinations in this study harm unborn babies. If 
participants are woman and having sex that could lead to pregnancy, they must agree not to 
become pregnant. 
If participant is a woman participating in sexual activity that could lead to pregnancy, she and/or 
her male partner must use one form of birth control that they discuss with the study staff. 
Subjects must start one method of birth control before they start taking the study drugs, while 
they are taking the study drugs, and for 30 days after stopping study drugs.  
 
Condoms (male or female) with or without a spermicidal agent. Condoms are recommended 
because their appropriate use is the only contraceptive method effective for preventing HIV 
transmission.  
 
Diaphragm or cervical cap with spermicide 
 
Intrauterine device (IUD) 
 
Hormone-based contraceptive 
Breastfeeding Risk: It is unknown whether the study drugs pass through breast milk and may 
cause harm to your infant. You must not breastfeed while you are in this study. 

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Unknown Risks: In addition to the risks listed above, there are risks that are not known or do not 
happen often when participants have these procedures performed. Participants will be informed 
in a timely manner, both verbally and in writing of any new information, findings or changes to 
the way the research will be done that might influence their willingness to continue participating 
in this study. 
18.5 Social Impact Events 
Individuals enrolled in this study may experience personal problems resulting from the study 
participation. Such problems are termed social impact events. Although we will make every 
effort to protect participant privacy and confidentiality, it is possible that participants' involvement 
in the study could become known to others, and that participants may experience stigmatization 
or discrimination as a result of being perceived as being HIV- infected or at risk for HIV infection. 
For example, participants could be treated unfairly, or could have problems being accepted by 
their families and/or communities. Problems may also occur in circumstances in which study 
participation is not disclosed, such as impact on employment related to time taken for study 
visits.  
In the event that a participant reports a social impact event, every effort will be made by study 
staff to provide appropriate assistance, and/or referrals to appropriate resources. Social impact 
events are documented and reviewed on a scheduled basis by the protocol team leadership 
with the goal of reducing their incidence and enhancing the ability of study staff to mitigate them 
when possible.  
Social impact events that are judged by the designee to be serious, unexpected, or more severe 
or frequent than anticipated, will be reported to the responsible site’s EC/IRB promptly, or 
otherwise in accordance with the EC/IRB’s requirements.  
18.6 Benefits 
Aside from compensation, the direct benefit to the participant for taking part in this study is 
provision of free Fluarix (or Fluzone) and Pneumovax vaccinations. Also, the information the 
investigators obtain from this study may help researchers design studies that could improve 
treatment for HIV in the future. 
18.7 Compensation 
Participants will be compensated for each study visit completed after their screening visit for 
their time and travel. Participants will not be compensated for their screening visit. There are a 
total of 18 visits after the screening visit (6 visits per cycle, 3 cycles total).  A breakdown of 
subject compensation is as follows: 
Screening Visit: No Compensation 
Baseline Visit (Weeks 0, 12, 24): $50 for each visit 
Cycles 1, 2 & 3 Post-vaccination days 2, 4, 7: $20 for each visit 
Cycles 1, 2 & 3 Post-vaccination days 14, 30: $30 and $50 for each visit, respectively. 

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Total: $570 
See Table 2 for summary of Participant compensation above. 
If the participant ends their participation early, he/she will be compensated for every visit they 
complete as described above. Subjects may receive up to $570 over the course of their 
approximate 28 weeks participation. 
Participants will be able to park free while visiting our clinic in the parking slots allocated for our 
patients.  Should a participant express a need for taxi or bus fare, we will provide transportation 
to the subject by way of our participant discretionary funds. 
18.8 Participant Privacy and Confidentiality 
All participant-related information including case report forms, laboratory specimens, evaluation 
forms, reports, etc., will be kept strictly confidential. All records will be kept in a secure, double 
locked location and only research staff will have access to the records. Participants will be 
identified only by means of a coded number specific to each participant. All computerized 
databases will identify participants by numeric codes only, and will be password-protected. 
Upon request, participant records will be made available to the study sponsor, the sponsor’s 
monitoring representative, and applicable regulatory entities.  
18.9 Critical Event Reporting 
All critical events occurring on study will be reported to the UCSD HRPP if they are at least 
possibly associated or of unknown association to study treatment or procedures. The team will 
use standard reporting time frames and HRPP requested formats. Critical events will be 
monitored by the study investigators and reported to the HRPP during annual reviews. 
18.10 Communicable Disease Reporting 
All cases of newly diagnosed HIV, syphilis, hepatitis B and hepatitis C must be reported to the 
county public health department. According to California state law, study staff is required to give 
public health department staff the participant’s name, contact information and treatment records 
for the hepatitis if requested. 
18.11 Incidental Findings 
Incidental findings that occur during the course of the study will be assessed among the study 
team, including investigators and NIH staff. Such findings will be reported in the literature and to 
the research community as appropriate. 
18.12 New Findings 
Publication of the results of this trial will be governed by NIH policies. Intellectual property and 
data generated under this project will be administered in accordance with both University and 
NIH policies, including the NIH Data Sharing Policy and Implementation Guidance of March 5, 
2003. Ownership of sole or joint inventions developed under the project will be owned by the 
institution(s) employing the inventor(s). Inventors shall be determined by U.S. Patent law, Title 
35 SC. University and Participating investigators/institutions will disclose any inventions 
developed under the project and such inventions will be reported and managed as provided by 
NIH policies. Sole inventions will be administered by the institution employing the inventor. Joint 

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inventions shall be administered based on mutual consultation between the parties. Similar 
procedures will be followed for copyrights. Materials generated under the project will be 
disseminated in accordance with University/Participating institutional and NIH policies. 
Depending on such policies, materials may be transferred to others under the terms of a 
material transfer agreement. Access to databases and associated software tools generated 
under the project will be available for educational, research and non-profit purposes. Such 
access will be provided using web-based applications, as appropriate. Publication of data shall 
occur during the project, if appropriate, or at the end of the project, consistent with normal 
scientific practices. Research data which documents, supports and validates research findings 
will be made available after the main findings from the final research data set have been 
accepted for publication. Such research data will be redacted to prevent the disclosure of 
personal identifiers. 
18.13 Study Discontinuation 
The study may be discontinued at any time by the EC/IRB, NIAID, or other government entities 
as part of their duties to ensure that research participants are protected.  
18.14 Post-Trial Access 
All de-identified data generated through the conduct of this trial will be made openly available to 
investigators in the field after the primary manuscript has been published. 
18.15 Community Advisory Board and Other Relevant Stakeholders 
The UCSD AVRC has a standing CAB, which will review the proposed study before its 
implementation and will receive regular updates on enrollment and any safety issues, as per 
standard AVRC protocols. 
19. ADMINISTRATIVE PROCEDURES 
19.1 Protocol Registration 
Prior to implementation of this protocol, and any subsequent full version amendments, the 
protocol and the protocol informed consent form(s) needs to be approved, as appropriate, by 
the UCSD institutional review board (IRB)/ethics committee (EC) and any other applicable 
regulatory entity (RE). Upon receiving final approval, the site will submit all required protocol 
registration documents to the DAIDS Protocol Registration Office (DAIDS PRO) at the 
Regulatory Support Center (RSC).  The DAIDS PRO will review the submitted protocol 
registration packet to ensure that all of the required documents have been received. 
Site-specific informed consent forms (ICFs) WILL be reviewed and approved by the DAIDS 
PRO and sites will receive an Initial Registration Notification from the DAIDS PRO that indicates 
successful completion of the protocol registration process. A copy of the Initial Registration 
Notification should be retained in the site's regulatory files. 
Upon receiving final IRB/EC and any other applicable RE approval(s) for an amendment, the 
site should implement the amendment immediately.  The site is required to submit an 
amendment registration packet to the DAIDS PRO at the RSC.  The DAIDS PRO will review the 

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submitted protocol registration packet to ensure that all the required documents have been 
received. Site-specific ICF(s) WILL NOT be reviewed and approved by the DAIDS PRO and 
sites will receive an Amendment Registration Notification when the DAIDS PRO receives a 
complete registration packet. A copy of the Amendment Registration Notification should be 
retained in the site's regulatory files. 
For additional information on the protocol registration process and specific documents required 
for initial and amendment registrations, refer to the current version of the DAIDS Protocol 
Registration Manual. 
19.2 Regulatory Oversight  
(pending) 
19.3 Study Implementation  
(pending) 
19.4 ClinicalTrials.gov 
This protocol is subject to the Food and Drug Administration Amendments Act of 2007 (FDAAA) 
and has been registered on ClinicalTrials.gov with the ClinicalTrials.gov unique protocol ID # 
NCT02707692. 
20. PUBLICATION POLICY 
Publication of the results of this trial will be governed by NIH policies. Intellectual property and 
data generated under this project will be administered in accordance with both University and 
NIH policies, including the NIH Data Sharing Policy and Implementation Guidance of March 5, 
2003. Ownership of sole or joint inventions developed under the project will be owned by the 
institution(s) employing the inventor(s). Inventors shall be determined by U.S. Patent law, Title 
35 SC. University and Participating investigators/institutions will disclose any inventions 
developed under the project and such inventions will be reported and managed as provided by 
NIH policies. Sole inventions will be administered by the institution employing the inventor. Joint 
inventions shall be administered based on mutual consultation between the parties. Similar 
procedures will be followed for copyrights. Materials generated under the project will be 
disseminated in accordance with University/Participating institutional and NIH policies. 
Depending on such policies, materials may be transferred to others under the terms of a 
material transfer agreement. Access to databases and associated software tools generated 
under the project will be available for educational, research and non-profit purposes. Such 
access will be provided using web-based applications, as appropriate. Publication of data shall 
occur during the project, if appropriate, or at the end of the project, consistent with normal 
scientific practices. Research data which documents, supports and validates research findings 
will be made available after the main findings from the final research data set have been 
accepted for publication. Such research data will be redacted to prevent the disclosure of 
personal identifiers. 
21. REFERENCES 

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30. Gianella S, Massanella M, Richman DD, et al. Cytomegalovirus Replication in Semen is 
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22. APPENDICES 
 
22.1 Appendix A. Concomitant Medications CRF 
 
22.2 Appendix B. Medical History CRF 
 
22.3 Appendix C. Physical Exam CRF 
 
22.4 Appendix D. Collection of Cervical Vaginal Specimens SOP 
 
22.5 Appendix E. Laboratory SOP Transporting Specimens 
 
22.6 Appendix F. Laboratory SOP Quality Management Plan 
 
22.7 Appendix G. Stool Specimen Collection SOP