A Study to Assess the Reduction of Human Papillomavirus (HPV) Viral Infectivity and Transmission in HPV-Positive Women After Vaccination With 9vHPV (RIFT-HPV)

A Non-Randomized, Open-Label Study to Assess the Reduction of Human Papillomavirus (HPV) Viral Infectivity and Transmission in HPV16/18-Positive Women After Vaccination With 9vHPV, a Multivalent L1 Virus-like Particle Vaccine, Evaluated in Cervical, Anal and Oral Samples Obtained After One, Two, and Three Vaccine Doses (RIFT-HPV1/RIFT-HPV2)

This is a non-randomized, open label study to assess the reduction of Human Papillomavirus (HPV) infectivity and transmission in women positive for HPV16 and/or 18 in a cervical, oral and anal sample and vaccinated with 9vHPV/Gardasil-9™.

The primary objective of the study is to demonstrate that vaccination with a 3-dose regimen of 9vHPV will reduce viral infectivity in HPV 16/18/16+18-positive women. This objective rests upon the hypothesis that, since vaccination with 9vHPV triggers the production of type-specific HPV antibodies which are exudated to the cervical and other infected mucosae, these antibodies adhere to and neutralize newly produced HPV 16/18 viral particles also present in the mucosae, thus reducing HPV's infective capacity and transmission to sexual partners.

Secondary objectives of the study are:

• To determine HPV antibody levels before and after vaccination for each of the 9vHPV-covered HPV types (6, 11, 16, 18, 31, 33, 45, 52, and 58), to distinguish an induced antibody production due to 9vHPV vaccination from a natural response to an HPV infection (when antibody production is expected to be lower).
• To demonstrate viral infectivity reduction in HPV 16/18/16+18 after vaccination with 1-dose or 2-dose regimen of 9vHPV. Since antibody production after administration of 2 vaccine doses is not inferior to 3 doses, infectivity reduction is expected to be detected after 2 doses, and at least partially after one dose.

The main endpoint of the study is the evaluation of the HPV infective capacity in cervical, anal and oral samples from HPV 16, 18 or 16+18-positive women, using a cellular assay that models in-vitro the cervical mucosa. In brief, the specific HPV biomarker E1^E4 is measured in HaCaT keratinocytes after being cultured with study samples and thus, exposed to HPV16/18 viral particles. A reduction in E1^E4 expression is expected for keratinocytes exposed to samples taken after vaccination with 9vHPV, since the specific HPV antibodies also present in these samples would bind HPV viral particles and prevent infection of cultured keratinocytes.

Other endpoints included in the study are:

• Detection of antibodies against HPV types covered by 9vHPV (6/11/16/18/31/33/45/52/58) by specific immunoassays (ELISA, cLIA).
• HPV16/18 virion detection using ELISA and electronic microscopy.
• HPV DNA detection and genotyping, using Anyplex HPV28. These endpoints are performed in cervical, anal and oral samples from HPV 16, 18 or 16+18-positive women
• Titration of antibodies against HPV types covered by 9vHPV in serum samples from HPV 16, 18 or 16+18-positive women using ELISA or cLIA.

A minimum of 39 and 30 women will be enrolled in two different study population cohorts, respectively:

• RIFT-HPV 1 cohort: non-vaccinated adult women aged 35 years or older, positive for HPV16-, 18-, or double positive for 16 and 18, without lesion or with cervical intraepithelial neoplasia (CIN) 1/2 lesion eligible for conservative treatment.
• RIFT-HPV 2 cohort: non-vaccinated adult women aged 27 years or older, positive for HPV16-, 18-, or double positive for 16 and 18, with multiple cervical, vulvar and/or anal lesions, with cervical lesions eligible for conservative treatment.

Candidates to participate in the study are selected according to the HPV DNA test result in a cervical sample taken in their routine cervical cancer screening visit or in their routine gynaecological follow-up visit.

There is no control group in this study: all participants are expected to complete all the per-protocol procedures in a total of 4 study visits within an average of 7 months' duration: Visit 1/ Day1, Visit 2/Month 2, Visit 3/Month 6, and Visit 4/Month 7.

The study procedures are the following:

• Pregnancy test on a urine sample in Visit 1 (pregnant women are excluded from the study).
• Completion of a questionnaire about the participant's health status, use of oral contraception and sexual activity in Visits 1 and 4.
• Cervical, anal oral and blood sample collection Visits 1, 2 and 3 before receiving 9vHPV vaccination, and in Visit 4.
• Intramuscular administration of 9vHPV in a three-dose regimen in Visits 1, 2 and 3.

Regarding data analysis for primary objective assessment, differences in the infectivity rate before (Day 1/ Visit 1) and after vaccination with 3 doses of 9vHPV (Month 7/ Visit 4) will be compared in cervical, anal and oral samples using non-parametric Wilcoxon signed rank test. The same assessment will be done in 1- or 2-dose vaccination scenario.

Antibody production before and after vaccination will be summarized for each of the 9vHPV-covered HPV types.

Inclusion Criteria:

• Women.
• Aged 18 years or older for RIFT-HPV Cohort 1 and 2, attending a routine cervical cancer screening visit or gynaecological visit.
• Positive for HPV16, 18 or double-positive for 16 and 18 and negative for the rest of high-risk HPV types in a cervical sample.
• Recently diagnosed for their HPV-positivity (within the last 10 months).
• Meet one of the following criteria:

have no apparent cervical lesion (Cohort 1). have a CIN1/2 lesion which is eligible for conservative treatment (cohort 1). have HPV 16 and/or HPV 18 positive anal test with non-apparent anal lesions or with anal lesions eligible for conservative treatment (Cohort 2).

have HPV 16 and/or HPV 18 positive cervical test with vulvar premalignant lesion or condylomas, associated to HPV infection (Cohort 2).

Exclusion Criteria:

• Presence of any cervical lesion that requires clinical intervention within 7 months that could significantly affect cervical epithelia (and therefore, HPV viral production), such as cervical conization.
• History of severe allergic reaction (e.g., swelling of the mouth and throat, difficulty breathing, hypotension, or shock) that required medical intervention.
• History of allergy to any vaccine component, including aluminum, yeast, or BENZONASETM (nuclease, Nicomedia).
• History of thrombocytopenia or any coagulation disorder that would contraindicate intramuscular injection of study vaccine.
• History of splenectomy.
• History of ano-genital cancer or HPV-related head and neck cancer.
• Pregnancy at the time of signing informed consent or planning to become pregnant within the full duration of the study.