A Randomized, Placebo-controlled Trial of Pre-treatment HPV Vaccination on Outcomes to LEEP Treatment of Cervical High Grade Squamous Intraepithelial Lesions in HIV-infected Women.
Phase: PHASE2
Conditions: Cervical High Grade Squamous Intraepithelial Lesion
Sponsor: University of Witwatersrand, South Africa

Page 1 of 42 
 
 
 
 
 
A randomized, placebo-controlled trial of pre-treatment HPV vaccination on outcomes to 
LEEP treatment of cervical high grade squamous intraepithelial lesions in HIV-infected 
women. 
 
 
 
 
 
Industry Support Provided by: 
VERSION 1.0 
8 November, 2014 
Includes LOA #1 
MHAVE Study 
 
Merck & Co., Inc. 
MSD # 50971 
#V501-190 
 
 

 
MERCK VACCINE STUDY VERSION 1.0 (LOA 1)  
MHAVE STUDY  
3 November 2014 
 
  
 
 
 

Page 3 of 42 
 
 
PROTOCOL TEAM ROSTER 
 
Co-Chairs 
 
Cindy Firnhaber MD, MS, DTM & H 
Cervical Cancer Screening Programme Manager | Right to Care — Treating AIDS 
Seriously  
Technical Director Clinical HIV Research Unit 
Associate Professor Department of Medicine, University of Witwatersrand 
Helen Joseph Hospital, Themba Lethu Wing, Perth Road, Westdene, Johannesburg, 
2092  |  
Tel: 011 276-8860 | Fax : 011 276-8885 | Mobile: 084 667 2348 | Email: 
cindy.firnhaber@righttocare.org 
 
Timothy J. Wilkin, MD, MPH 
Cornell Clinical Research Site 
53 W. 23rd Street, 6th Floor 
New York, NY 10010 
Phone:(212) 746-7202 
Fax: (212) 746-7203 
E-mail:tiw2001@med.cornell.edu 
 
Immunology Sub-Study Co Chairs 
 
Luis J. Montaner, D.V.M., M.Sc., D.Phil. 
Professor, HIV-1 Immunopathogenesis Laboratory 
The Wistar Institute, 3601 Spruce St., Philadelphia, PA 19104 USA 
215-898-9143 (off)/3934 (lab) Fax. 215-573-7008 (off)/9272 (lab) 
Email: Montaner@wistar.org 
 
 
Emmanouil Papasavvas, PhD, 
Emmanouil Papasavvas, PhD, Senior Staff Scientist  
The Wistar Institute,  
3601 Spruce Street, 
Philadelphia, PA, 19104 
USA 
Telephone: 215 573 7007 
email: manolisp@wistar.org 
 
 
 
 
 
 
Contracts/Finance 

 
MERCK VACCINE STUDY VERSION 1.0 (LOA 1)  
MHAVE STUDY  
3 November 2014 
 
  
Ina de Jongh 
Operations Manager  
Clinical HIV Research Unit 
Wits Health Consortium 
Department of Medicine, University of Witwatersrand 
Helen Joseph Hospital, Themba Lethu Clinic, Perth Road, Westdene 2092 
Johannesburg South Africa 
Tel: +27 11 276-8800 / Direct: +27 11 276-8803 / Mobile: 071 332-7933 / 072 345-8633 
Fax to E-mail: 086 618-0476  
E-mail: idejongh@witshealth.co.za 
 
Pharmacist 
Coreen Barker BPharm 
Rightmed Pharmacy 
Clinical HIV Research Unit 
Wits Health Consortium 
Department of Medicine, University of Witwatersrand 
Helen Joseph Hospital, Themba Lethu Clinic, Perth Road, Westdene 2092 
Johannesburg South Africa 
Tel: +27 11 276-8800 / Direct: +27 11 276-8811  
Fax to Email:  086 515 6228 / Fax: +27 11 482 5554 
E-mail: cbarker@witshealth.co.za 
 
 
 
Data Manager/Randomization/Participant registration/Computer issues 
All questions concerning this protocol should be sent a response should generally be 
received within 24 hours (Monday-Friday). 
 
Avril J. Swarts, MSc 
Clinical HIV Research Unit 
Themba Lethu Clinic, Helen Joseph Hospital 
Perth Road 
Westdene 
2092 
Phone: 
+27 11 276-8876 
Fax: 
086 5133143 
E-mail: 
aswarts@witshealth.co.za 
 

 
MERCK VACCINE STUDY VERSION 1.0 (LOA 1)  
MHAVE STUDY  
3 November 2014 
 
  
 
 
 
 
 
Protocol Document Questions- 
Cindy Firnhaber MD, MS, DTM & H 
Cervical Cancer Screening Programme Manager | Right to Care — Treating AIDS 
Seriously  
Technical Director Clinical HIV Research Unit 
Associate Professor Department of Medicine, University of Witwatersrand 
Helen Joseph Hospital, Themba Lethu Wing, Perth Road, Westdene, Johannesburg, 
2092  |  
Tel: 011 276-8860 | Fax : 011 276-8885 | Mobile: 084 667 2348 | Email: 
cindy.firnhaber@righttocare.org 
 
Timothy J. Wilkin, MD, MPH 
Cornell Clinical Research Site 
53 W. 23rd Street, 6th Floor 
New York, NY 10010 
Phone: 
(212) 746-7202 
Fax: 
(212) 746-7203 
E-mail: 
tiw2001@med.cornell.edu 
 
Product Package Inserts or Investigator Brochures 
To request copies of product package inserts or investigator brochures, contact the 
DAIDS Regulatory Support Center (RSC) at RIC@tech-res.com or call (301) 897-1708. 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

 
MERCK VACCINE STUDY VERSION 1.0 (LOA 1)  
MHAVE STUDY  
3 November 2014 
 
  
 
 
 
 
 
 
 
 
 
 
 
 
 
GLOSSARY OF PROTOCOL-SPECIFIC TERMS 
 
ASC-H 
atypical squamous cells, cannot exclude HSIL  
ASCUS 
atypical squamous cells of undetermined significance 
AIS 
adenocarcinoma in situ 
 
CIN 
cervical intraepithelial neoplasia 
HPV 
human papillomavirus 
HSIL 
high grade squamous intraepithelial lesion 
LSIL 
 
low grade squamous intraepithelial lesion 
 
LEEP/LLETZ 
 
loop electrical excision procedure/large loop electrical removal of 
transformation zone 
PPA 
per-protocol analysis 
qHPV 
quadrivalent HPV vaccine 
SUSAR 
suspected, unexpected serious adverse reactions  
VAERS 
Vaccine Adverse Event Reporting System 
VaIN 
vaginal intraepithelial neoplasia 
VIN 
vulvar intraepithelial neoplasia 
VLP 
virus-like particle 

 
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MERCK VACCINE STUDY VERSION 1.0 (LOA 1)  
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SCHEMA 
 
 
A randomized, placebo-controlled trial of pre-treatment HPV vaccination on outcomes to 
LEEP/LLETZ treatment of cervical high grade squamous intraepithelial lesions in HIV-
infected women.      
 
DESIGN 
This is a single-center, randomized, double-blinded, placebo-controlled, 
phase II trial of the quadrivalent human papillomavirus vaccine (qHPV) in 
HIV-infected women to prevent occurrence of cervical HSIL after 
LEEP/LLETZ. At entry and prior to vaccination, participants will undergo 
colposcopy with directed biopsies, cervical cytology, and stored HPV 
testing. Participants will then be randomized to the quadrivalent vaccine 
or saline placebo to be given at entry, week 4, and week 26. Women will 
have LEEP treatment at week 4.  During follow-up, participants will be 
seen for cervical cytology, colposcopy with directed biopsies at weeks 26 
and 52, and HPV specimens collection. Treatment assignment will be 
unblinded after study follow-up is completed for the last study participant.  
Women aged 45 or less randomized to placebo will be offered open label 
HPV vaccine. 
 
A subset of women (n=120) will participate in an immunology sub-study.  
These women must have a CD4 count >200 cells/mm3 and a plasma HIV-
1 RNA of <200 copies/mL.  These women will undergo cervical fluid and 
blood collection, for assessment of gene expression signatures, HPV 
antibodies, and markers for immune activation. 
 
DURATION 
Each participant will be followed for 1 year 
 
SAMPLE SIZE 
180 participants 
 
POPULATION 
HIV-1-infected women, aged 18 years or older, with cervical HSIL 
 
 
STRATIFICATION  
Participation in immunology subset 
 
REGIMEN 
 
qHPV vaccine or placebo 0.5 mL at entry, week 4, and week 26. 
 

 
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1.0 
HYPOTHESIS AND STUDY OBJECTIVES 
 
1.1 
Hypothesis 
 
HPV vaccination will reduce the occurrence of cervical HSIL post-LEEP/LLETZ. 
 
1.2 
Primary Objective 
 
To determine the effect of pre-treatment quadrivalent HPV vaccination on outcomes to 
LEEP/LLETZ treatment of cervical HSIL on persistent and/or recurrent HSIL among HIV-
infected women  
 
1.3 
Secondary Objectives 
 
1.3.1 To compare cervical cytology abnormalities, and squamous intraepithelial 
lesions on cervical biopsy between arms at week 26 and 52. 
 
1.3.2 To store cervical swabs for high-risk HPV testing to evaluate persistent, 
recurrent, and incident HPV infections. 
 
1.3.3 To explore vaccine efficacy in different subgroups:  women with cervical HSIL 
confirmed on LEEP/LLETZ specimen at week 4, and women with positive 
margins on LEEP/LLETZ specimen at week 4. 
 
1.3.4 To collect samples for immune sub-study for assessment of gene expression 
signatures, HPV antibodies, and markers for immune activation associated 
with primary objective outcomes. 
 
 
2.0 
INTRODUCTION 
 
2.1 
Background 
 
 
Cervical cancer is one of the most common cancers in women. The most recent compilation of 
global data indicates that an estimated 490,000 new cases of cervical cancer occur annually 
among women worldwide and nearly 80% of these are in developing countries, where screening 
programs are not well established. It is estimated that 270 000 women die annually from 
cervical cancer, 85% of these in low resource nations1-3.  In Africa, cervical cancer makes up 
23.3% of all cancers in women4. HIV-infected women have high rates of cervical intraepithelial 
neoplasia (CIN) especially in resource limited areas.  South Africa and Zambia report that high 
grade squamous intraepithelial lesions (HSIL) were found in approximately 30% of HIV-infected 
women 5, 6.  The Women’s Interagency HIV Study (WIHS), a US cohort, found that HIV-infected 
women had a CIN prevalence of 16%, of which 87% were low grade squamous intraepithelial 
lesions (LSIL) and 13 % (HSIL), compared to only 4 % in HIV-uninfected women7. With 

 
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improved access to ART, HIV-infected women are living longer allowing time for persistent high-
risk HPV infections and HSIL to progress to invasive cancer.    
 
Cervical HSIL (cervical intraepithelial neoplasia grade 2 or 3), the precursor lesion to cervical 
cancer, is treated by either ablative or excisional methods. A very efficient and effective 
outpatient procedure requiring only local anesthesia is LEEP/LLETZ (loop electrosurgical 
excision procedure/Large Loop Excision of the Transformation Zone).  LEEP/LLETZ uses a thin 
electrified wire to remove the lesion up to 7 mm in depth from the transitional zone. Treatment 
failure rates (as defined as incomplete ectocervical and/or endocervical margins on pathology 
specimens) are between 10-15% for the LEEP/LLETZ procedure in immunocompetent women 
8.   The LEEP/LLETZ failure rate among HIV-infected women is up to a 50% failure rate 
worldwide.  The incomplete excisional margin significantly increases the risk of persistence 
and/or recurrence of disease.   The incomplete excisional lesions are thought to be due to the 
greater size of the HSIL lesions and the multifocal nature of these lesions in HIV seropositive 
women9-12.   Additional studies suggest that a combination of cytology and high risk HPV testing 
6 months post-LEEP/LLETZ are highly predictive of persistent/recurrent HSIL13,14 . Other factors 
that may be associated with increased risk of recurrence and /or persistence of disease are 
persistent/recurrent HPV infection, CD4 count <200 cells/mm3, plasma HIV-1 RNA> 50 
copies/ml and not receiving ART 9-12, 15-17.  
 
Persistent HPV infection 
Persistent and/or recurrent HPV infection is a risk factor for refractory disease 15, 16. However 
few studies have investigated the effect of LEEP/LLETZ treatments on HPV clearance in HIV-
infected women.  Massad and colleagues reported on 170 HIV-infected women from the WIHS 
(Women’s Interagency HIV Study) and HERS (HIV Epidemiology Research Study) cohorts who 
underwent cervical treatment for CIN.  They found that HPV commonly persisted among women 
who had abnormal cervical cytology after treatment17. More data are available for HIV-
uninfected women. Elgren and colleagues reported retrospectively on 109 women treated for 
CIN with either cervical conization or cryotherapy18.  Eighty-four (81%) were positive for HPV.  
Thirty-one women were treated with cervical cryotherapy and 15 (48%) were positive for the 
same type 3 months later.  Persistence of HPV was approximately 20% 12 months after cervical 
cryotherapy. Of note, HPV clearance was greater for women treated with cervical conisation 18.  
A5282 (an ACTG international multisite test and treat study) will provide important data on 
clearance of HPV in HIV-infected women after cryotherapy and LEEP/LLETZ. 
 
HIV-infected women commonly require chronic treatment of cervical dysplasia (i.e. multiple 
LEEP/LLETZs, cone biopsies, and/or hysterectomy). These repeated procedures increase the 
morbidity for the patients in terms of increase risk of cervical incompetence and adds to the 
expense and patient load in health care systems especially in resource limited countries where 
the prevalence of high grade is so high.  New strategies are needed to improve outcomes to 
treatment of high grade CIN.    
 
HPV Vaccination in HIV-infected populations 
Gardasil (quadrivalent HPV vaccine consisting of recombinant L1 particles for types 6, 11, 16 
and 18) has been shown to prevent cervical, vaginal, anal and external genital lesions caused 

 
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by the vaccine types 19.  However, it does not hasten clearance of prevalent HPV infections20 
nor prevent or treat lesions caused by ongoing infection with the vaccine types21, 22.  
 
 
ACTG A5246/AMC052 was a single-arm, open-label trial evaluating the safety and 
immunogenicity of the quadrivalent HPV vaccine (6, 11, 16, and 18) in 109 HIV-1-infected men 
without HGAIN 23. The vaccine was found to be safe. There were no appreciable effects on 
plasma HIV-1 RNA or CD4+ counts. There were no grade 3 or 4 events that were possibly, 
probably, or definitely related to vaccination. The vaccine was highly immunogenic with 
seroconversion rates being 95% or greater for each of the four vaccine types. Among women, 
the immunogenicity, safety, and tolerability of the quadrivalent HPV recombinant vaccine 
directed against types 6, 11, 16, and 18 was assessed in the ACTG 5240 study.  This was a 
phase II, open-label, single arm study with stratification by CD4+ cell count of 319 women aged 
19 to 45 in the US, Brazil and South Africa. Among 222 women with CD4 counts over 200 and 
seronegative for each type at baseline, the vaccine was highly immunogenic with 
seroconversion rates ranging from 84 to 100% for each HPV type in the vaccine. There were no 
differences in immunogenicity rates between women with a CD4 count over 350 compared with 
counts between 200 and 350. There were no safety concerns related to the vaccine24.  
 
HPV Vaccination in the Context of Dysplasia Treatment 
Joura and colleagues examined the women (HIV negative) receiving excisional cervical 
treatments while participating in the clinical studies of the quadrivalent HPV vaccine. In the 
combined retrospective evaluation of the two phase 3 HPV vaccine trials, 587 vaccine and 763 
placebo recipients underwent treatments of the cervix25. Women who had received the 
quadrivalent HPV vaccine had a 46% lower risk of subsequent HPV disease and a 65% [95%CI: 
20-86] lower risk of high grade CIN as compared to women who had received placebo. 
Interestingly, the observed effect was not restricted to infection or dysplasia caused by vaccine 
types.  
 
Swedish and colleagues conducted a retrospective chart review of HIV-uninfected men who 
have sex with men who were diagnosed with high grade anal intraepithelial neoplasia (HGAIN).  
They showed that men who had been previously vaccinated had a 50% lower risk for recurrent 
or persistent HGAIN after undergoing ablative treatment of HGAIN 26. 
 
These data suggest that HPV vaccination may improve outcomes to treatment of dysplasia. 
HPV infects the epithelium through microabrasions in the epithelium27.  HPV antibodies induced 
by HPV vaccination, when present in high titers, prevent infection by binding the virus and 
preventing interaction with the acellular basement membrane28.   At lower antibody titers, this 
interaction is not blocked.  Lower antibody titers block the interaction of the HPV virion and the 
basal epithelial cells thereby preventing infection by a second mechanism29.  This second 
mechanism is thought to be how HPV vaccines confer long lasting protection even when 
antibody titers have fallen to low levels.  
 
Persistence of high risk HPV after LEEP/LLETZ is a major risk factor for recurrence of high 
grade CIN.  LEEP/LLETZ may clear HPV infection of the cervix, but will not clear HPV infections 
at other sites in the genital tract (i.e. vagina or vulva) that may lead to re-infection of the healing 
cervix.  Women are also at risk for re-infection from their sexual partners.  We hypothesize that 

 
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HPV antibodies induced by HPV vaccination present in the cervical area after LEEP/LLETZ will 
protect the healing epithelium from re-infection with HPV by the two mechanisms discussed 
above.  
 
There are limited data on the immunogenicity of the quadrivalent HPV vaccine 4 weeks after a 
single dose.  Among seronegative young, HIV-uninfected women, the geometric anti-HPV16 
titers 1 and 4 weeks after a single dose of the quadrivalent HPV vaccine were 30 [95% CI 18, 
50] and 113 [80, 159], mM/mL30 [unpublished data from Merck].  There are data 4 weeks after a 
single dose of Cervarix (bivalent HPV vaccine using the same viral like particles for HPV 16 and 
18, but a different adjuvant).  99% and 98% of HIV-uninfected women age 15-25 seroconverted 
to type 16 and 18 respectively31.  The Advisory Committee on Immunization Practices, which 
sets the standard for immunizations in the US, accepts a second HPV vaccination as early 28 
days after the 1st vaccine. 
 
HPV and immune regulation 
The understanding of the immunoresponse of HPV infection and to the HPV vaccine is also very 
limited. A recently presented abstract at CROI by Papasavvas et al evaluated the systemic and 
local immune responses in HIV infected women with suppressed HIV viral loads on ART with 
and without HR HPV infection.  Presence of HR HPV infection irrespective of lesion grade (CIN 
1-2-3) was associated with increased T cell activation in otherwise ART suppressed HIV 
infected women with higher CD4 counts (median CD4 count approximately 500)   Changes 
between CIN 1 and 3 did suggest greater levels of negative immunoregulatory molecules 
expression even if increased in all groups with HR HPV types as compared to women without 
HR HPV 32.  In addition, cervical biopsy analysis supports that changes in peripheral blood also 
relate to local tissue changes in gene expression as well as detectable gene expression 
changes with increasing cervical dysplasia.  Further evaluation of immune activation, 
immunoregulation, gene expression and HPV immunity in ART suppressed women is required.  
It remains unknown how quickly the biological changes associated with HR HPV cervical 
dysplasia would reverse after a LEEP/LLETZ procedure with or without vaccine immunotherapy 
if targeted to target HR types.  Note that in addition to the cervix, we will also monitor 
presence/absence of high-risk types in anal region33-37, to control for the systemic impact of 
independent high-risk infection in the anus might have on the immune response before/after 
LEEP/LLETZ.  
 
    
 
2.2 
Rationale 
Women with HIV have high rates of cervical HSIL, particularly in South Africa, and are at higher 
risk for persistent or recurrent HSIL after LEEP/LLETZ, which could increase their risk for 
progression to cervical cancer38.  Strategies are needed to improve outcomes to cervical 
treatments in HIV-infected women. This is even more critical in resource constrained settings 
where cervical cancer screening may be very infrequent; follow-up is difficult and health care 
systems overwhelmed.  
 
In South Africa, the quadrivalent HPV vaccine is indicated for the active immunisation in females 
9 to 45 years of age for the prevention of cervical, vulva, vaginal and anal cancer, precancerous 

 
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or dysplastic lesions, genital warts and in males 9 to 26 years of age for the prevention of anal 
cancer, precancerous or dysplastic lesions, external genital lesions and infection caused by 
HPV types 6, 11, 16 and 18.  The recommended age in girls is age 11-14.  HPV vaccination is 
not routinely available in South Africa, especially for those aged 18 or over.  
 
This proposal will evaluate whether these outcomes can be improved with HPV vaccination prior 
to cervical LEEP/LLETZ.  While HPV vaccination is unlikely to hasten clearance of established 
type 16 or type 18 infections, it may prevent new infections or re-infection of the cervix after 
LEEP/LLETZ treatment and subsequent recurrent disease.  If successful, this study will provide 
a proof-of-principle for this strategy. Next-generation vaccines protecting against additional 
types of high-risk HPV would lead to even greater benefits. 
 
 
3.0 
STUDY DESIGN 
 
This is a single center, randomized, double-blinded, placebo-controlled, phase II trial. The study 
population will include 180 HIV-1-infected women, aged 18 years and older with cervical HSIL 
on biopsy.  
 
Participants will be consented per SOP of the study unit. Cervical colposcopy with biopsy of 
visible lesions will be done to evaluate for cervical HSIL.  This is only done if a result is not 
available within the last 90 days. Participants will undergo blood testing for CD4 and HIV viral 
load if not available within the last 90 days. 
 
At entry, participants are randomized 1:1 to HPV vaccination or placebo. Participants will 
undergo cervical swabs for HPV DNA PCR. A pap smear is done if a result is not available 
within the last 180 days. Participants will receive vaccine or placebo at entry, week 4, and week 
26.   
 
At week 4, women will be seen for LEEP/LLETZ treatment of cervical HSIL. The LEEP/LLETZ is 
performed according to IARC or WHO recommendations at the discretion of the treating 
provider. 
 
At week 26, and week 52, participants will be seen for cervical swabs for HPV testing and 
cervical cytology. Cervical colposcopy and biopsy of visible lesions will be performed.  If no 
lesions are seen at week 26 and 52, then random biopsies will be done at 6 and 12 positions on 
the cervix.  At least two biopsies should be obtained for all participants.  Endocervical curettage 
can be performed at the discretion of the treating provider.  Participants diagnosed with cervical 
HSIL at week 26 will be treated according to South African standard of care; options include a 
repeat LEEP/LLETZ, cone biopsy or other treatment as determined necessary by the local 
provider.  These treatments are not specified by the study and do not constitute study visits. 
These women should remain in follow-up and be seen at week 52.  Women diagnosed with 
cervical HSIL at week 52 will be seen for treatment as determined by South African standard of 
care.   Other HPV related lesions, such as vaginal, vulvar or perianal HSIL, will be referred to 
appropriate specialist and treated per South African standard of care..  This is not determined 
by the study. 

 
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MERCK VACCINE STUDY VERSION 1.0 (LOA 1)  
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Participants will be told of their treatment assignment after the last participant has completed 
study follow-up.  Participants age 45 or less who received placebo will be offered open-label 
HPV vaccine.  Please note that this vaccine is approved for women up to age 45 in South 
Africa.  This will be provided by the study and dispensed by the study pharmacy.  No additional 
follow-up is planned for this open-label administration of HPV vaccine. 
 
 
In addition, 120 participants will be asked to participate in the Immunology sub-study.  
Participants are eligible if their CD4 count is >200 cells/mm3, and plasma HIV RNA level is <200 
copies /ml.  Participants in the immunology sub-study will give cervical fluid specimens using 
small sponges inserted in the cervical os for assessment of immune activation, extra blood 
specimens (i.e., PBMC for immunophenotypic characterization, plasma for HIV-1 RNA testing 
and serum for assessment of HPV antibodies and soluble markers of immune activation), at 
entry, week 4, weeks 26 and 52.  The plasma HIV-1 RNA specimens will be stored for 
evaluation when funding is available using the Abbot HIV viral load platform. The PBMC, serum, 
, and cervical fluid specimens will be stored until funding is available for testing.  This testing will 
be performed at the WISTAR Institute in Philadelphia, PA, USA. In addition, recut slides of the 
colposcopic biopsies of women participating in the immunology sub-study will be sent to Wistar 
Institute in Philadelphia, Pennsylvania for immunohistochemical evaluation of cervical cellular 
inflitrates. Anal swabs will be done tested for HR HPV typing. See Table below for description of 
specimens to be collected.   The sub-study procedures will be done before the vaccination and 
any colposcope or LLETZ/LEEP procedures if at all possible. 
 
Sample 
Purpose 
Lab 
Participan
ts to be 
notified of 
results 
Stored 
until 
funding 
available 
or real 
time 
MTA 
require
d 
Cervical 
cyto broom 
Assessment of dysplasia 
NHLS 
Yes 
Real time 
No 
Cervical 
biopsies, 
LEEP/LLET
Z 
specimens 
Assessment of dysplasia 
NHLS 
Yes 
Real time 
No 
Cervical 
Swab 
HPV testing 
UCT 
HPV 
lab 
No 
Stored 
No 
Anal swab 
To determine HPV present and type 
in anal canal 
NHLS 
No 
Stored 
No 
Cervical 
fluid 
Cytokine/Chemokine profiling 
Wistar 
No 
Stored 
Yes 
Plasma  
HIV RNA 
CLS 
Wits  
Yes 
Real time 
for 
screening
No 

 
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/ stored 
for post-
entry 
Blood 
Rapid HIV testing, CD4 cell count 
CLS 
Wits 
Yes 
Real time 
Yes 
Serum 
HPV antibodies, soluble markers of 
immune activation 
Wistar 
No 
Stored 
Yes 
PBMC 
Flow cytometry for 
immunophenotyping 
Wistar 
No 
Stored 
 Yes 
Whole blood 
lysate 
Gene expression profiling of 
pathways associated with HPV 
immune evasion 
Wistar 
No 
Stored 
Yes 
Recut 
histology 
slides from 
Colposcopic 
biopsy  
immunohistochemical evaluation of 
cervical cellular infiltrates 
Wistar 
No 
Stored 
Yes  
 
 
 
4.0 SELECTION AND ENROLLMENT OF PARTICIPANTS 
4.1 Inclusion Criteria  
4.1.1 HIV infection documented by positive HIV test by two different criteria; 
either 2 different rapid HIV tests of different manufacturers, plasma HIV-1 
RNA level >5,000 copies/mL, or Western blot). 
4.1.2 Women aged ³ 18 years. 
4.1.3 Cervical HSIL on biopsy (i.e. CIN2 and/or CIN3) 
4.1.4 For participants of reproductive potential, negative serum or urine 
pregnancy test with a sensitivity of <25 mU/mL within 48 hours prior to 
study entry.  
NOTE: Reproductive potential is defined as women who have not been 
postmenopausal for at least 24 consecutive months (i.e., who have had 
menses within the preceding 24 months) or women who have not 
undergone surgical sterilization (e.g., hysterectomy, bilateral 
oophorectomy, tubal ligation, or salpingectomy).  
4.1.5 Contraception requirements 
All study participants must agree not to participate in a conception 
process (e.g., active attempt to become pregnant or in vitro fertilization) 
during study participation (from the time of study entry until week 52). 
 
If participating in sexual activity that could lead to pregnancy, the study 
participant must agree to use at least one reliable form of contraceptive 

 
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from the time of study entry until week 52. At least one of the following 
contraceptives MUST be used appropriately: 
• 
Condoms (male or female) 
• 
Diaphragm 
• 
IUD 
• 
Hormone-based contraceptive 
 
4.2 Exclusion Criteria 
4.2.1 History or current biopsy diagnosis of invasive or microinvasive cervical, 
vaginal, vulvar or anal cancer 
4.2.2 Prior hysterectomy 
4.2.3 Cervical cryotherapy or LEEP/LEETZ within one year of entry. 
4.2.4 Cervical, vulvar, or vaginal lesions suspicious for cancer, unless biopsies 
show no invasive cancer 
4.2.5 Prior receipt of one or more doses of an HPV vaccine. 
4.2.6 Receipt of anticoagulants other than aspirin or nonsteroidal anti-
inflammatory drugs (NSAIDS) within 14 days prior to entry. 
4.2.7 Known allergy/sensitivity or any hypersensitivity to yeast or any of the 
components of the study product or its formulation (see section 5.2 for a 
list of components). 
4.2.8 Hemophilia or other bleeding diatheses. 
4.2.9 Use of any systemic antineoplastic or immunomodulatory treatment, 
systemic corticosteroids, other than inhaled corticosteroids or prednisone  
≤ 10 mg (or equivalent) , investigational vaccines, interleukins, 
interferons, growth factors, or intravenous immunoglobulin (IVIG) within 
45 days prior to study entry.  
NOTE: Routine standard-of-care vaccines (including hepatitis A, hepatitis 
B, influenza, pneumococcal, and tetanus vaccines) are not exclusionary. 
4.2.10 Breastfeeding 
4.2.11 Less than 3 months post-partum 
4.3 
Study Enrollment Procedures 
 
4.3.1 Prior to implementation of this protocol, and any subsequent full version 
amendments, the protocol and the protocol consent form approved by the local 
ethics committee (EC) and MCC. 
 
Once a candidate for study entry has been identified, details will be carefully 
discussed with the participant. The participant (or, when necessary, the legal 
guardian if the participant is under guardianship) will be asked to read and sign 
the approved protocol consent form.  

 
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4.3.2 Randomization 
 
Participants will be randomized using permuted block randomization using block 
sizes of six, stratified by participation in the immunology substudy. The 
randomization list will be kept under lock and key in the investigational pharmacy. 
 
4.4 
Co enrolment Guidelines 
 
Participants may be co-enroled in other observation clinical trials without consulting 
study chairs but will need ethics approval prior to co-enrollment.  The study chairs 
should be notified of participation in any other clinical trials 
 
 
5.0 
STUDY TREATMENT 
 
Study treatment is defined as:  
• 
Quadrivalent human papillomavirus (types 6, 11, 16, and 18) recombinant 
vaccine (qHPV) or saline placebo.  
 
• 
Loop electro excision procedure (LEEP), also known as large loop excision of the 
transformation zone (LLETZ) 
 
Antiretroviral therapy (ART) will not be supplied by the study and should be continued or 
initiated per the local standard of care. ART must be obtained outside of this study.    
 
5.1 
Regimens, Administration, and Duration 
 
5.1.1 Regimens 
 
Participants will be randomized to receive the quadrivalent HPV (types 6, 11, 16, 
and 18) recombinant vaccine, 0.5 mL, or the placebo for qHPV at entry (day 0) 
and at weeks 4 and 26.  
 
Please note:  The recommended schedule for the quadrivalent HPV vaccine is 0, 
2, and 6 months.  This study is using a modified schedule where the second 
vaccine is given earlier.  This alternative regimen is acceptable according to the 
Advisory Committee on Immunization Practices, which sets the standard for US 
immunization practices and preferred by MERCK for use in this study.  This is the 
vaccination schedule for the bivalent HPV vaccine, which is very similar to the 
quadrivalent HPV vaccine.  This minimizes the delay between diagnosis of HSIL 
and LEEP/LLETZ.  
 
All participants will undergo LEEP/LLETZ at week 4. 
 
All participants will be followed for 1 year post-randomization. 

 
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5.1.2 Study Product Administration 
 
qHPV or saline placebo will be administered as three separate 0.5 mL doses 
according to the visit schedule. The vaccine is to be used as supplied; no dilution 
or reconstitution is necessary. The vaccine will be administered intramuscularly 
in the deltoid region of the upper arm or the higher anterolateral area of the thigh 
in the clinic by study personnel using aseptic technique during preparation and 
administration. All injections should preferably be given in the deltoid region and 
in the non-dominant arm, however if this is not feasible, the dominant arm may 
be used. The participant should be seated during vaccine injection. 
 
The vaccine must not be injected intravascularly, subcutaneously, or 
intradermally.   
 
Vaccination should be deferred at the discretion of the site investigator if the 
participant is febrile or acutely ill. 
 
Following each vaccination, clinic staff will observe the participant for 15 minutes. 
 
5.2 
Study Product Formulation 
 
5.2.1 Quadrivalent HPV Virus-Like Particle (VLP) Vaccine 
Each 0.5 mL dose of qHPV (types 6, 11, 16, and 18) recombinant vaccine 
contains approximately: 
• 
HPV 6 L1 protein 
 
 
20 mcg 
• 
HPV 11 L1 protein 
 
 
40 mcg 
• 
HPV 16 L1 protein 
 
 
40 mcg 
• 
HPV 18 L1 protein 
 
 
20 mcg 
• 
Amorphous aluminum  
hydroxyphosphate sulfate adjuvant 225 mcg 
• 
Sodium chloride 
 
 
9.56 mg 
• 
L-histidine 
 
 
 
0.78 mg 
• 
Polysorbate 80 
 
 
50 mcg 
• 
Sodium borate 
 
 
35 mcg 
• 
Yeast protein  
 
 
< 7 mcg 
• 
Water  
 
 
 
q.s. (‘quantity sufficient’) 
 
5.2.3 Placebo for Quadrivalent HPV Vaccine 
 
Sodium Chloride for Injection USP, 0.9% will be used as the placebo for qHPV 
vaccine.  
 

 
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For the purpose of this study and to maintain the blind of study product, the 
Sodium Chloride for Injection USP, 0.9% must be stored refrigerated at 2°C to 
8°C (36°F to 46°F).  
 
 
5.3 
Study Product Preparation  
 
 
5.3.1 Quadrivalent HPV (VLP) Vaccine 
 
• 
One pre-filled syringe of qHPV will be used to prepare this dose. Prior to 
injection, the pharmacist will shake the vaccine syringe well before use. 
Thorough agitation immediately before administration is necessary to 
maintain suspension of the vaccine.  
 
• 
qHPV vaccine should not be diluted or mixed with other vaccines. 
 
• 
After thorough agitation, qHPV vaccine will be a white, cloudy liquid. Do not 
use the product if particulates are present or if it appears discolored. 
 
• 
Using aseptic technique, the pharmacist must inject the 0.5-mL dose of 
vaccine from the pre-filled syringe to a sterile syringe using a needle through 
the orifice of sterile syringe.  
 
• 
Once drawn into the syringe, the vaccine should be kept cold (2° to 8°C) until 
it is ready to be administered to the participant. Use promptly. 
   
• 
The pharmacist will place an overlay on the syringe. With the overlay in 
place, the syringe should be labeled as “Quadrivalent HPV Vaccine or 
Placebo” to maintain blinding. The syringe should also be labeled with the 
date/time product was withdrawn from the refrigerated vial into the syringe 
and the participant’s identifier.   
 
• 
All used vaccines should be discarded appropriately. 
 
5.4.2 Placebo for Quadrivalent HPV (VLP) Vaccine 
 
• 
One vial of normal saline (labeled as Sodium Chloride for Injection USP, 
0.9%) will be used to prepare this dose. 
 
• 
Placebo for qHPV should not be diluted or mixed with other vaccines. 
 
• 
Using aseptic technique, the pharmacist must withdraw 0.5-mL of Sodium 
Chloride for Injection, 0.9% using a sterile needle and syringe.  
 

 
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• 
Once drawn into the syringe, the placebo should be kept cold (2° to 8° C) 
until it is ready to be administered to the participant. Use promptly.   
 
• 
The pharmacist will place an overlay on the syringe. With the overlay in place 
the syringe should be labeled as “Quadrivalent HPV Vaccine or Placebo” to 
maintain blinding. The syringe should also be labeled with the date/time 
product was withdrawn from the refrigerated vial into the syringe and the 
participant’s identifier.   
 
• 
All used vaccines should be discarded appropriately. 
 
5.5 
Pharmacy: Product Supply, Distribution, and Accountability 
 
5.5.1 Study Product Acquisition/Distribution 
 
Quadrivalent HPV (types 6, 11, 16, and 18) recombinant vaccines for all 
participants are supplied by Merck & Co., Inc. The placebo for quadrivalent HPV 
vaccine (Sodium Chloride for Injection USP, 0.9%) must be obtained by the site.  
 
5.5.2 Study Product Accountability 
 
The site pharmacist is required to maintain complete records of all study vaccine 
received from Merck and subsequently dispensed. Continuous inventory is not 
required for normal saline, but all other information must be completed (including 
the lot number for the vial/ampule/IV solution used).  
 
5.6 
Concomitant Medications 
 
 
5.6.1 Required Medications 
 
None. 
 
5.6.2 Prohibited Medications 
 
• 
Receipt of licensed or experimental HPV vaccines outside of the study. 
 
5.6.3 Precautionary Medications 
 
• 
Anticoagulants other than aspirin or NSAIDS. 
• 
Systemic antineoplastic or immunomodulatory treatment, systemic 
corticosteroids, other than inhaled corticosteroids, investigational vaccines, 
interleukins, interferons, growth factors, or intravenous immunoglobulin 
(IVIG).  

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6.0 
CLINICAL AND LABORATORY EVALUATIONS 
 
6.1 
Schedule of Events for all participants 
 
Screening 
Entry 
Week 4 
(-0 days to plus 30 days) 
Week 26 
(-14 days to +45 days days) 
Week 52/Premature discontinuation 
(+/- 45 days) 
Documentation of HIV 
X 
 
 
 
 
Medical history/Clinical Assessments 
X 
X 
X 
X 
X 
Stored Cervical HPV DNA PCR 
 
X 
 
X 
X 
Cervical cytology  
 
X2 
 
X 
X 
Cervical Colposcopy and directed biopsies  
X1 
 
 
X 
X 
LEEP/LLETZ  
 
 
X 
 
 
HPV vaccine or placebo 
 
X 
X 
X 
 
Pregnancy test 
X 
X 
X 
X 
X 
Plasma HIV-1 RNA/ CD4+/CD8+ 
X1 
 
 
 
 
Immunology subset study (n=120) 
 
 
 
 
 
Cervical sponge 
 
X 
X 
X 
X 
Recut histology slides from biopsy for immunochemistry  
 
X 
X 
X 
X 
Stored PBMCs /CD4 count 
 
X 
X 
X 
                            X 
Stored serum for HPV antibodies, immune markers 
 
X 
X 
X 
X 
Anal swab  
 
X 
 
X 
X 

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Screening 
Entry 
Week 4 
(-0 days to plus 30 days) 
Week 26 
(-14 days to +45 days days) 
Week 52/Premature discontinuation 
(+/- 45 days) 
Stored plasma for HIV-1 RNA 
 
X 
X 
X 
X 
 
 
 
 
 
 
1 Only needed if results are not available from within 90 days of entry 
2 Only needed if results are not available from within 180 days of entry 

 
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6.2 
Timing of Evaluations 
 
6.2.1 Screening Evaluations  
 
Screening evaluations must occur before any study treatment or intervention. 
 
Screening evaluations to determine eligibility must be completed within 90 days 
prior to entry, unless otherwise specified. A cervical colposcopy/biopsy, 
CD4+/plasma HIV-1 RNA done as part of routine care meets this requirement if 
done within the 90 days prior to entry, and cytology meets this criteria if done 
within 180 days of entry. If patient has clinical sexual transmitted disease then 
the patient may be deferred until clinically resolved and then screening may 
continue. 
6.2.2 Entry Evaluations 
 
Screening may occur on the same day as entry if all necessary evaluations are 
available.  Entry evaluations must occur after randomization and be completed 
before initiating study vaccine. 
 
Entry evaluations may occur the same day as randomization. 
 
All entry specimens must be obtained before initiating study treatment. 
 
The participant should receive the first vaccine/placebo injection on the day of 
randomization.  The vaccine must be given within 7 days after randomization if the 
vaccine.  
6.2.3 Post-Entry Evaluations 
 
If clinically active sexually transmitted Disease (STD) then the STD should be 
treated and then the samples collected after the STD clinically resolved. This 
should occur with 30 calendar days of the study visit in which the main and the 
immunology subset study testing should have been day  
 
On-Study Evaluations 
Study visits must be scheduled on the weeks indicated in the Schedule of 
Evaluations (SOE) 
 
Week 52 Final Visit Evaluations 
These evaluations are done at the participant’s final on-study visit.  
 
6.2.4 Discontinuation Evaluations 
 
Evaluations for Registered/Randomized Participants Who Do Not Start Study 
Vaccine 
 No further evaluations should be performed.  These participants will be replaced. 
 

 
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Premature Treatment Discontinuation Evaluations 
No extra visit is required for those who initiate but do not complete the 
vaccination series. For those discontinuing the vaccine series after the first 
vaccine, the week 4 should be performed except for vaccine administration.  The 
week 26 visit should be performed and the patient continued to be followed on 
study.. For those discontinuing the vaccine after the second vaccine, the week 26 
visit should be performed. Please note that the week 26 and week 52 visits 
should be conducted even if the vaccine series is not completed. 
 
Premature Study Discontinuation Evaluations  
Participants who prematurely discontinue prior to the Week 52 window (- 45 
days) should have the Week 52 visit performed early provided it has been at 
least 12 weeks since the date of the Week 28 visit.  
 
6.3 
Instructions for Evaluations 
 
All clinical and laboratory information required by this protocol is to be present in the 
source documents.  
  
All stated evaluations are to be recorded on the CRF and keyed into the database 
unless otherwise specified. This includes events that meet the International Conference 
on Harmonisation (ICH) definitions for a serious adverse event (SAE). See section 11.2. 
 
To grade diagnoses, signs and symptoms, and laboratory results, sites must refer to the 
DAIDS AE Grading Table, Version 1.0, December 2004 (Clarification, August 2009), 
available on the RSC website at http://rsc.tech-res.com/safetyandpharmacovigilance/ 
Grading Table). 
 
Cytology and histology results, other than invasive cancer, do not need to be recorded 
as events. These results will be entered on study-specific CRFs. 
 
6.3.1 Documentation of HIV-1 
 
Refer to section 4.1.1 regarding assay requirements for HIV-1 documentation.  
 
6.3.2 Medical History 
 
The medical history must document any AIDS-defining illness or condition as 
defined by the CDC.   
 
Record the participant’s nadir CD4+ count (absolute value and date). Document 
the nadir CD4+ count when possible with a copy of the nadir CD4+ count report. 
If this documentation is not available, then subject recall will suffice. For 
participants who do not know the exact nadir value and for whom there is no 
source documentation, then recall of the categorical nadir (e.g., < 50, < 100, or < 
200 cells/mm3) will suffice. 

 
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Any prior history of treatment for cervical dysplasia or condylar should be 
documented. 
 
6.3.3 Clinical Assessments 
 
Signs and Symptoms 
At entry, record all Grade 3 or greater signs and symptoms that occurred within 
14 days before entry in source documents only. After entry, record Grade 3 or 
greater signs and symptoms regardless of their relationship to study treatment or 
study procedures in the source documents, and all signs and symptoms that led 
to a change in study treatment, regardless of grade.  Please see section 6.3.5 for 
signs and symptoms to be recorded on the CRF. 
 
Diagnoses 
After entry, sites will only record the following: 
• 
Any diagnosis of a cellulitis, abscess, or other infection at the vaccination site. 
• 
Record all admissions for LEEP/LLETZ complications 
• 
Record all cancer of the anogenital area 
 
 
Concomitant Medications 
Record the following: 
• 
For the immunology subset, any ART interruption lasting ≥ 14 days during the 
study (from enrollment visit onward).  
• 
Any precautionary or prohibited medications (see section 5.4) including 
anticoagulants other than aspirin or NSAIDS. 
• 
Any topical or surgical treatments for CIN, VIN, VaIN, or genital condyloma  
• 
Any HPV vaccination given outside of this study (this is prohibited but if given, 
needs to be recorded as a concomitant medication). 
 
6.3.5 Laboratory Evaluations and Study Procedures 
• 
After entry, record all Grade 3 or greater laboratory values if drawn to 
evaluate potential AEs related to study vaccination. 
• 
Record all signs, symptoms or toxicities that led to a change in study 
vaccination, regardless of grade.  
• 
Record all grade 3 and above AEs if deemed possibly, probably or definitely 
related to study vaccine or study procedures.  
Study visits collecting cervical specimens (i.e. swabs, lavage, fluid, biopsies 
or LEEP/LLETZ) should be deferred when the participant is menstruating.  
These visits should also be deferred if the participant exhibits visual evidence 
of bacterial sexually transmitted diseases other than bacterial vaginosis for 7 
days after appropriate treatment is initiated and the study visit should be 
completed within 30 days of the initial study visit.   
 
 
 

 
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Cervical Cytology 
Collected per the schedule of events using a cytobroom and placed on a glass 
slide (conventional cytology).  These results will be interpreted by cytologist at 
the University of the Witwatersrand/NHLS (National Health Laboratory Services), 
a laboratory accredited with the South African National Accreditation System 
(SANAS). Previous quality assurance data on the previous Pap smears with this 
team of cytologists has shown a high level of concordance with cytology readings 
from University of North Carolina (85%).  
 
 
For quality assurance, a second pathologist from South Africa will reread and the 
original Pap smear slide for diagnosis will not leave South Africa: 
• 
10% of screening/baseline cytology smears 
• 
100% of HSIL cytology specimens at weeks 26 and 52 
• 
  20% of normal cytology specimens at weeks 26 and 52 
 
Any discordant results will be adjudicated in conference. 
Cervical HPV DNA PCR 
Collected per the schedule of events. Cervical swabs should not be obtained 
during the 14 days after any cervical biopsy. 
 
Cervical colposcopy and directed biopsies 
Colposcopy will follow the cervical cancer research standard operational 
procedures. The patient is placed on the table with her feet in stirrups to position 
the pelvis for examination and the speculum is then inserted into the vagina to 
open vaginal walls and visualise the cervix. A light and low powered microscope 
(colposcope) is used to illuminate the cervix. The cervix is gently swabbed with 
5% acetic acid to remove mucus and highlight abnormal areas like abnormal 
blood vessels, acetowhite epithelium and warts.  Digital photographs and 
samples using biopsy forceps are then taken from all areas of suspicion and sent 
for histology in a container with formalin. Lugol’s Iodine may be used if further 
elucidation of the suspicious area is needed. If no lesions are visualized at the 
week 26 visits, then no biopsies are taken.  If no lesions are visualized at the 
week 52 visit, then biopsies at 6 and 12 o’clock will be taken.  At least two 
biopsies are taken for each participant at week 52.  If bleeding occurs, 
hemostasis is done with a dry swab held by forceps.  Rarely, a hemostatic 
solution called Monsel’s solution will be applied to reduce any minor bleeding. 
Inadequate colposcopy results will be repeated. 
          
If participants require anticoagulation other than aspirin or NSAIDS after study 
entry, they should not undergo cervical colposcopy and biopsies. 
 
Histology will be performed by the Department of Anatomical Pathology at the 
University of the Witwatersrand  NO HISTOLOGY BLOCKS WILL LEAVE 

 
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MERCK VACCINE STUDY VERSION 1.0 (LOA 1)  
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SOUTH AFRICA. For quality control purposes, and second pathologist will 
reread: 
• 
25% of HSIL cervical biopsies used for eligibility,  
• 
25% of LEEP/LLETZ specimens, and  
• 
100% of cervical HSIL on biopsy at weeks 26 and 52  
• 
10% of non-HSIL biopsies at weeks 26 and 52 
 
Cervical Fluid Collection (immunology subset only) 
This should be done prior to the cytobroom for cytology and cervical swab for 
stored HPV DNA PCR.  First, a speculum is inserted into the vagina exposing the 
cervix.  Using a hemostat, one ophthalmic sponge is gently inserted into the 
cervical os being careful not to touch the vaginal walls and left in place for one 
minute.  Then the sponge is removed, placed in a cryovial and frozen at -80 deg 
C.    
 
LEEP/LLETZ 
During the LLETZ procedure, the cervix is stained with 5% acetic acid and 
inspected under magnification. Abnormal areas are identified and subsequently 
stained with Lugol’s Iodine. Local anaesthetic (Xylocaine 2% with adrenalin) is 
administered with a dental syringe and a 27G needle at the usual 3, 6, 9 and 12 
o’clock positions around the cervix. A selection of a loop electrode is made 
depending on the size of the lesion that needs to be removed.  
LLETZ is performed by a trained physician (either OB/GYN or Medical Officer 
trained in LLETZ) using a Finesse II ElectroSurgical Unit (ESU) with smoke 
evacuation system. The Finesse II ESU incorporates Controlled Output Circuitry 
to produce the best histological tissue specimen for the pathologist. Controlled 
Output Circuitry is an “intelligent cut” module that maintains the power output 
within the ideal cutting range to produce a specimen with minimal thermal 
damage at the margins. The generator continuously adjusts itself to meet the 
minimum needs for optimal cutting. This eliminates any need to adjust the output 
setting during the procedure, or when changing loop sizes. The smoke 
evacuation port is connected to the non-conductive speculum and the electrode 
is introduced into the vagina without activation. Only once in front of the cervix, is 
the loop electrode activated into cutting mode before touching the tissue, thereby 
activating the smoke exhaust fan before smoke can develop. 
Under direct colposcopic vision, the loop electrode is introduced into the cervical 
tissue with gentle pressure, the operator starts cutting approximately 2 mm 
outside the identified abnormal epithelium. With a steady sweep the loop 
electrode is moved from one side of the cervix to the other, removing the 
previously identified lesion. If necessary, a second or even third sweep though 
the cervical tissue is made to remove any residual abnormal tissue. The 
specimen is immediately preserved in Formalin, properly labelled and prepared 
for transport to histopathology. hemostasis is achieved with electrocoagulation of 

 
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the crater bed using the same Finesse II ESU in coagulation mode with a ball 
electrode.  
Pregnancy Test 
A serum or urine pregnancy test is required for all women of reproductive 
potential (defined in 4.1.8) within 48 hours of entry, before each vaccination, 
before LEEP/LLETZ, and at any time pregnancy is suspected. Record the results 
of all pregnancy tests.  
 
NOTE: Refer to section 7.3 for clinical management of women who become 
pregnant on study.  
 
Plasma HIV-1 RNA 
At entry, HIV-1 RNA determinations will be obtained. Samples for plasma HIV-1 
RNA will be stored for the participants in the immunology subset at week 4, 26 
and 52. 
 
CD4+/CD8+ 
At screening, absolute CD4+/CD8+ count and percentages will be obtained. 
Samples for CD4+/CD8+ will be stored for the participants in the immunology 
subset at week 26 and 52. 
 
Stored Serum (immunology subset only) 
Serum for measures of HPV infection will be collected and stored per the SOE. 
These samples will be tested for soluble markers of inflamation and anti-HPV 
antibody concentrations once funding is identified.  This will be done on the first 
120 women who consent and qualify for the sub study 
 
Stored PBMCs (immunology subset only) 
PBMCs for future HPV and HIV immunology testing will be stored per the SOE. 
This will be done on the first 120 women who consent and qualify for the sub 
study.  
 
Stored Anal HPV swabs (immunology subset only) 
Anal HPV  swabs will be stored per the SOE. This will be done on the first 120 
women who consent and qualify for the sub study 
 
6.3.8 Vaccine/Placebo Injection 
 
Every effort should be made to administer vaccines within the protocol-defined 
visit window. Participants who do not receive the week 4 or week 26 vaccine will 
remain on study and continue the evaluations as listed in the SOE (see section 
6.2.4).  
 
The timing of the HPV vaccines is based on the Advisory Committee on 
Immunization Practices recommendations for HPV vaccination.  If the week 4 

 
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and/or week 26 vaccine is not given within the specified windows, then the 
following guidelines must be followed: 
• 
The week 4 vaccine must be given at least 28 days after the first vaccine.  If 
the week 4 is not within the protocol-specified window, then the vaccine can 
be given as late as 24 weeks post randomization.  
• 
The week 26 vaccine should be administered even if the week 4 vaccine is 
not administered because of missed visits. 
• 
The week 26 vaccine must be given at least 12 weeks after the week 4 
vaccine, and at least 24 weeks after the week 0 vaccine.  The week 26 
vaccine can be given as late 36 weeks post randomization.  
• 
The week 52 visit should occur at least 12 weeks after the week 26 visit.  
 
Vaccination should be deferred at the discretion of the site investigator if the 
participant is febrile or acutely ill. 
 
Participants will remain at the clinic for a 15-minute post-injection observation 
period following each vaccination to observe for syncope or any immediate 
adverse reactions.  
 
 
7.0 
CLINICAL MANAGEMENT ISSUES 
 
7.1 
Dose Modification 
 
No modification of the study vaccine dose is allowed. 
 
7.2 
Toxicity Management 
 
7.2.1 Management of Injection Site and Allergic Reactions 
 
Local and systemic reactions will be graded according to DAIDS AE Grading 
Table, Version 1.0, December 2004 (Clarification, August 2009). 
 
Injection Site Reactions 
• 
Swelling, induration, or redness at the injection site 
• 
Pain or tenderness, or other reactions at the injection site 
 
Systemic reactions occurring within 48 hours after vaccination 
• 
Fever or chills  
• 
Malaise or fatigue  
• 
Headache or pain, e.g., myalgia or arthralgia  
• 
Nausea or vomiting 
• 
Allergic reactions, e.g., rash, hives, dyspnea, pruritis 
• 
Syncope 
 

 
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For injection reactions judged to be severe, the protocol chairs should be notified 
within 24 hours and further vaccines should not be given to that participant prior 
to consultation with protocol co-chairs. 
 
For injection reactions judged to be life-threatening, the protocol chairs should be 
notified within 24 hours and no additional vaccinations shall be given to that 
participant. 
 
The protocol chairs should be contacted within 24 hours for any Grade 3 or 4 
reactions (e.g., injection site reactions, elevated temperatures following 
vaccination that occur at any point during study participation) thought definitely, 
possibly, or probably related to vaccination. Further vaccines should not be given 
to that participant prior to consultation with the protocol co-chairs. 
 
7.2.2 Other Adverse Events 
 
AEs related to LEEP/LLETZ or cervical biopsies should be graded using the 
DAIDS “Female Genital Grading Table for Use in Microbicide Studies,” which can 
be found on the DAIDS RSC web site: http://rsc.tech-
res.com/safetyandpharmacovigilance/.   
 
For toxicities not specifically addressed above, the following guidelines should be 
used for the management of AEs that are felt to be at least possibly related to 
vaccines: 
 
• 
Grade 1 or 2 Toxicity/AE 
 
Participants who develop a Grade 1 or 2 AE may continue study vaccine. For 
instructions regarding participants experiencing Grade 1 or 2 AEs who 
choose to permanently discontinue study treatment or study participation, see 
section 6.2.4. 
 
• 
Grade 3 or 4 Toxicity/AE 
 
Participants who develop a Grade 3 or 4 AE that is at least possibly related to 
vaccine should be reevaluated for that toxicity until the AE returns to Grade 
≤2. The study vaccine may be given at the discretion of the site investigator. 
If the same Grade 3 or 4 AE recurs and is considered by the investigator to 
be possibly, probably, or definitely related to the study vaccine, the study 
vaccine must be permanently discontinued. If, in the investigator’s opinion, 
the AE has NOT been caused by the study vaccine or if the event is an 
asymptomatic laboratory abnormality, study treatment may continue. 
 
For instructions regarding participants experiencing Grade 3 or 4 AEs that are 
at least possibly related to vaccine and that result in premature 

 
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discontinuation of study vaccine or study participation, see sections 6.2.4. 
These participants should be followed weekly until resolution of the AE. 
 
7.3 
Pregnancy and breastfeeding 
 
Notify the protocol chairs of all on-study pregnancies or occurrences of breastfeeding on 
study. 
 
Women who become pregnant on study will discontinue study treatment.  Study visits 
should be suspended until the pregnancy has been completed. In the case of an early 
termination of pregnancy, the participant may continue with evaluations and the 
vaccination schedule per the SOE.  Intrapartum complications and/or pregnancy 
outcomes will be recorded.  
 
If a woman reaches week 52 or chooses to discontinue the study before the end of 
pregnancy, site staff should request permission to contact her regarding pregnancy 
outcomes. Information obtained will be submitted on the CRF at the end of the 
pregnancy.  
 
Women who become pregnant on study through 6 weeks after the last study vaccine 
may request unblinding (see section 11). These cases of exposure to HPV vaccine will 
be reported Merck in accordance with Section 11.  
 
 
8.0 
CRITERIA FOR DISCONTINUATION 
 
8.1 
Premature Treatment Discontinuation 
• 
Request by participant to terminate treatment. 
• 
Treatment-related AEs as discussed in section 7.2.  
• 
Pregnancy or breastfeeding. 
• 
Clinical reasons believed life-threatening by the site investigator, even if not 
addressed in section 7.2. 
 
8.2 
Premature Study Discontinuation 
• 
Request by the participant to withdraw. 
• 
Request of the primary care provider if s/he thinks the study is no longer in the best 
interest of the participant. 
• 
Failure to start study vaccination within 7 days after randomization.  
• 
At the discretion of the EC, industry supporter, investigator, or other government 
agencies as part of their duties to ensure that research participants are protected.  
 
 

 
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9.0 
STATISTICAL CONSIDERATIONS  
 
9.1 
General Design Issues 
 
This is a randomized, double-blinded, placebo-controlled trial to assess the effect of pre-
treatment HPV vaccination on the occurrence of cervical HSIL post-LEEP/LLETZ. 
 
Quadrivalent HPV vaccine (6, 11, 16, and 18) will be administered in three doses with 
the second and third doses administered at weeks 4 and 26. The primary endpoint is 
defined as having cervical HSIL on cytology or biopsy at week 26 or 52.  
 
The primary analysis is modified intention-to-treat.  We will exclude randomized women 
who do not receive the first dose of study vaccine.  These women will not be followed 
and will be replaced.  Women to attend both the week 26 and week 52 visit will be 
censored from the primary analysis. 
 
The targeted accrual is 180 women. We anticipate that accrual will be completed within 
one year.   
 
9.2 
Outcome Measures 
 
9.2.1 Primary Outcome Measure  
 
 
Cervical HSIL (HSIL on cervical cytology or HSIL on cervical biopsy) at week 26 
or week 52. 
 
9.2.2 Secondary Outcome Measures 
9.2.2.1 Cervical cytology abnormalities at pre-entry, week 26, and week 52; and 
cervical biopsy abnormalities at week 26 and week 52. 
 
9.2.2.2 Cervical swabs for high-risk HPV testing at entry, week 26 and week 52. 
 
9.2.2.3 Cervical HSIL at week 26 and 52 in women stratified by whether cervical 
HSIL is found on LEEP/LLETZ biopsy, and stratified by whether LEEP/LLETZ 
margins gave HSIL detected. 
 
9.2.2.4 Cervical sponge samples,  recut slides from the cervical biopsy (samples 
from paraffin blocks collected at screening and post-entry, paraffin blocks 
from LEEP/LLETZ specimens at week 4, 26,52), PBMC, and whole blood 
lysate samples. 
 

 
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9.3 
Randomization and Stratification 
 
Participants will be randomized 1:1 to the HPV vaccination or placebo group using 
random permuted blocks.  Randomization will be stratified based on participation in the 
Immunology Sub study. 
 
9.4 
Sample Size and Accrual 
 
The primary analysis will be based on comparing proportions with cervical HSIL 
detected.  We will perform the analysis at a two-sided 0.05 significance level and target 
an 80% power to detect a 50% reduction in hazard with vaccination assuming 40% 
occurrence of cervical HSIL in the placebo group and 20% occurrence in the placebo 
group.  Alternate scenarios are shown below. 
 
Probability of endpoint in control 
group 
Probability of endpoint in vaccine 
group 
Power to detect 
difference 
50% 
25% 
92% 
50% 
30% 
74% 
40% 
20% 
80% 
40% 
25% 
53% 
30% 
15% 
63% 
 
Reimers et al33 reviewed 75 HIV-infected women who underwent LEEP/LLETZ for treatment of 
cervical HSIL.  They found that 60% developed persistent or recurrent cervical HSIL.  CD4 
count was strongly associated with recurrent/persistent disease. The women in the study by 
Reimers et al had lower CD4 counts and less use of ART than women we expect to enroll in the 
proposed study.  We expect a lower treatment failure rate for our proposed study.  Firnhaber 
and colleagues are conducting a clinical trial comparing LEEP/LLETZ and cryotherapy 
treatment for cervical HSIL in HIV-infected women. When combining participants randomized to 
LEEP/LLETZ and women with HSIL who were ineligible for randomization for having lesions 
that are no amendable to cryotherapy (i.e. extensive or endocervical lesions), we estimate that 
40% will have disease recurrence as defined by the primary endpoint. 
 
9.5 
Monitoring 
 
This study will be reviewed every 2 months by the protocol co-chairs.  They will review 
accrual, retention, pregnancies, study conduct (% of participants completing 
LEEP/LLETZ and vaccinations, % of expected visits completed and % of expected 
results received), and safety (Grade 3 or greater events at least possibly related to study 
vaccination, LEEP/LLETZ, or cervical biopsies).   
 
We will constitute a Data and Safety Monitoring Board to review study conduct and 
safety. They will conduct a review approximately 6-9 months after study accrual has 
begun, and approximately one year later.  No interim efficacy analysis is planned.  The 
study accrual should be completed within one year.  It will take one year until week 26 
visits have been completed on 50% of participants.  At this point, accrual will be 

 
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complete or nearly complete. Two or more Grade 3 or 4 adverse events or a single 
death that are at least possibly related to study vaccination will prompt an interim review 
by this advisory committee. Accrual will be halted until the Advisory Committee and the 
protocol chairs have reviewed the clinical trial data and agreed that the study should 
continue. 
 
9.6 
Analyses 
 
9.6.1 Primary Analysis 
 
The primary endpoint is the occurrence of cervical HSIL at week 26 or week 52.  
The timing of the endpoint at week 26 or 52 is not felt to be clinically relevant.  
We will not use time-to-event analyses.  Only women who have completed a 
week 26 or 52 visit will be included in the analysis.  We will compare the 
proportion of these women with cervical HSIL detected between arms.  We will 
consider the number of missing week 26 or 52 visits in both arms.  If there is an 
appreciable difference between arms, we will consider sensitivity analyses that 
adjust for missing visits.  
 
Women who do not receive the first vaccine will be excluded from study follow-up 
and the primary analysis.   
  
9.6.2 Secondary Analysis 
 
9.6.2.1 Cervical cytology abnormalities will be summarized and reported 
according to randomized group at screening, week 26 and week 52. 
 
 
Cervical biopsy results will be summarized and reported according to 
randomized group at week 26 and week 52. 
 
 
LEEP/LLETZ biopsy results will be reported. 
 
9.6.2.2 We will report on the availability on the availability of stored swabs for 
HPV testing at entry, week 4, week 26 and week 52. 
 
 
9.6.2.3 We will perform the primary analysis in various subgroups.  These 
subgroup analyses are not powered to show statistical significance.  The 
focus will be on the direction of the point estimate and to better 
understand any observed efficacy.  The subgroups include women with 
HSIL detected on LEEP/LLETZ specimen, and all other women.  Please 
note that this subgroup relies on data observed at week 4.  We do not 
expect that vaccination will affect the occurrence of HSIL on LEEP/LLETZ 
specimen based on published literature.  We will look for appreciable 
differences in the prevalence of HSIL on LEEP/LLETZ specimens 
between groups.  Any appreciable differences will limit the utility of this 

 
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analysis.   A second subgroup analysis will be women with LEEP/LLETZ 
margins positive for HSIL, and all other women.  This also involves a 
post-randomization variable and similar concerns exist for this subgroup 
analysis.   
 
9.6.2.4 We will report the availability of Cervical sponge samples, cervical biopsy 
recut slides, paraffin blocks from LEEP/LLETZ specimens on week 4, 
PBMC, and whole blood lysate samples. 
 
10.0 
PHARMACOLOGY PLAN 
Not applicable.  
 
11.0 
ADVERSE EVENT REPORTING REQUIREMENTS FOR THIS STUDY 
 
11.1 
Definitions 
 
Adverse Event (AE) shall mean any untoward medical occurrence in a Study subject who 
is administered the Study Drug regardless of whether or not a causal relationship with the 
Study Drug exists.  By way of example and without limitation, an AE can be any 
unfavorable and unintended sign (for example, an abnormal laboratory finding), symptom, 
or disease temporally associated with the use of the Study Drug,  
  
Serious Adverse Event (SAE) shall mean any untoward medical occurrence in a Study 
subject who is administered the Study Drug that results in death, a life-threatening drug 
experience, requires inpatient hospitalization or prolongation of existing hospitalization, 
results in persistent or significant disability/incapacity, or is a congenital anomaly/birth 
defect, cancer, or is a new cancer if the cancer is the condition of the study, or 
overdose.   Other important medical events that may jeopardize the patient or may require 
intervention to prevent one of the outcomes listed previously should also be considered 
“serious”. 
  
Suspected Unexpected Serious Adverse Reaction (SUSAR) shall mean any Serious 
Adverse Event, the nature, severity or frequency of which is not consistent with information 
in the most current investigator’s brochure, or with respect to a marketed product the most 
current Summary of Product Characteristics (SPC) or Package Insert. 
  
11.2 
Reporting Requirements 
 
 
11.2.1 SAE, SUSAR, pregnancy/breastfeeding, and cancer reporting:  Protocol Chair shall 
forward to MSD’s Global Safety (“MSD GS”) group, any SAE and SUSAR 
information, including, but not limited to, all initial and follow-up information involving 
any participant.  Notification shall be in the form of a completed CIOMS I/MedWatch 
within two (2) business days of learning of the SAE or SUSAR.  All SAE and 
SUSAR information shall be transmitted in the English language and contain the 

 
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reporter’s name and the participant identifier.   SUSAR information will be reported 
unblinded.  
 
Please note that all of SAE, SUSAR, and cancer diagnoses should also be reported 
to the EC and MCC within 24 business hours of site awareness. Weekends and 
public holidays are excluded.  
   
All reports of study Drug exposure during pregnancy or breastfeeding, whether 
associated with an AE or not, must be reported to MSD GS in accordance with the 
timelines and contact information for an SAE.  Study staff shall follow pregnancies 
to term to obtain the outcome of the pregnancy.  The outcome of the pregnancy 
shall be forwarded to MSD GS. 
  
All cancers should be reported as SAEs. Study drug overdoses will not be 
reported as SAEs unless there is an associated sign, symptom or diagnosis 
qualifying as an SAE. An inadvertent overdose will be recorded on the CRF. An 
overdose of vaccine/placebo is defined as administration of a 4th dose of 
vaccine/placebo during study participation. 
 
11.2.2 Reporting Procedures to Government Agencies. 
 
Adverse Reporting Department at Merck will determine the AEs reported to 
Merck that will be passed on to the US Agency, EMEA or other agencies by 
Merck.  Local reporting to the ethics committee and South African health 
authorities is the responsibility of the protocol chairs. 
 
 
11.3 Grading Severity of Events 
 
The DAIDS AE Grading Table, Version 1.0, December 2004 (Clarification, 
August 2009), will be used. It is available on the RSC website at http://rsc.tech-
res.com/safetyandpharmacovigilance/. 
 
AEs related to LEEP/LLETZ or cervical biopsies will be graded using the DAIDS 
“Female Genital Grading Table for Use in Microbicide Studies,” which can be 
found on the DAIDS RSC web site: http://rsc.tech-
res.com/safetyandpharmacovigilance/.   
 
Cervical Cytology and biopsy results do not need to be recorded as AEs unless 
invasive cancer is shown. 
 
11.4 
Unblinding Procedure 
 
Emergency unblinding is rarely allowed as the study product can be discontinued in a 
participant experiencing AEs without the need for unblinding. In the event that 
emergency disclosure of treatment assignment is thought to be required, the protocol 

 
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chairs should be contacted. The protocol chairs will discuss any unblinding request.  If it 
is still thought to be necessary by one of the protocol chairs after this discussion, then 
the unblinding request will be granted.  Should Merck have a request from an ex-US 
agency to unblind an SAE report, the request should come from Merck directly to study 
chairs and it will be handled as per the clinical trials agreement.  Unblinding is allowed 
for women who become pregnant within 6 weeks after a vaccine dose. All SUSAR’s will 
be unblinded. 
 
12.0 
HUMAN PARTICIPANTS 
 
12.1 
Ethics Committee Review and Informed Consent 
 
This protocol and the informed consent document (Appendix I) and any subsequent 
modifications will be reviewed and approved by the EC responsible for oversight of the 
study. A signed consent form will be obtained from the participant or legal 
representative. The consent form will describe the purpose of the study, the procedures 
to be followed, and the risks and benefits of participation. A copy of the consent form will 
be given to the participant or legal representative, and this fact will be documented in the 
participant’s record. 
 
12.2 
Participant Confidentiality 
 
All laboratory specimens, evaluation forms, reports, and other records that leave the site 
will be identified by coded number only to maintain participant confidentiality. All records 
will be kept locked. All computer entry and networking programs will be done with coded 
numbers only. Clinical information will be released only as necessary for monitoring by 
EC, other government agencies, or the industry supporter or designee.  
 
12.3 
Study Discontinuation 
 
The study may be discontinued at any time by the EC, the industry supporter, or other 
country-specific government agencies as part of their duties to ensure that research 
participants are protected. 
 
13.0 
PUBLICATION OF RESEARCH FINDINGS 
 
Publication of the results of this trial will be determined by the study chairs.  Submission 
of the primary manuscript is expected within 6 months of receiving the primary analysis. 
 
 
14.0 
BIOHAZARD CONTAINMENT 
 
As the 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 drawing of blood and shipping and 

 
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handling of all specimens for this study, as currently recommended by the Centers for 
Disease Control and Prevention and the National Institutes of Health. 
 
All dangerous goods materials, including diagnostic specimens and infectious 
substances, must be transported using packaging mandated by CFR 42 Part 72. Please 
refer to instructions detailed in the International Air Transport Association (IATA) 
Dangerous Goods Regulations.  
 

 
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