Impact of HPV Vaccination on Cervical High Grade Lesions in France
Condition: Human Papillomavirus Infection
MeSH terms: Papillomavirus Infections
Study type: Observational

Summary: 
      The objective is to estimate the prevalence of the different genotypes of human
      papillomaviruses (HPV) in histologically proven high-grade lesions of the cervix in patients
      born between 1972 and 1993. It is plan to determine if the prevalence of HPV16 and HPV18 is
      lower in the population born between 1983 and 1993 and potentially exposed to HPV vaccination
      compared to those born between 1972 and 1982 who were not exposed to vaccination. Thus the
      investigators should be able to determine whether the introduction of HPV vaccination in
      France in 2007 has had an impact on the development of high-grade lesions associated with
      HPV16 and HPV18.
    

Detailed description: 
      The prevalence of the different HPV in high-grade lesions (CIN2/3) will be determined using
      the INNOLiPA HPV extra v2 genotyping kit (Fujirebio). Only samples with a histologically
      proven high-grade cervical lesion (grade 2 or 3 intraepithelial neoplasia) are concerned. The
      patient year of birth, the year of the lesion diagnosis and the histological diagnosis will
      be recorded Participating laboratories will select eligible cervical samples (biopsies or
      surgical specimen) according to the inclusion criteria defined in the protocol. Only cervical
      samples with sufficient biological material will be selected. The selected tissue blocks will
      be sent to the Papillomavirus National Reference Center (NRC) by mail (pre-paid envelope
      provided) in accordance with the rules for the transport of biological samples. After receipt
      by the NRC, the tissue blocks will be anonymized. After HPV genotyping, the blocks will be
      returned to the laboratory. Transport costs are covered by the Papillomavirus NRC.

      Samples have to be fixed in 4% buffered formaldehyde (maximum 24 hours) and then included in
      paraffin for histopathological evaluation. On each of the selected histological pieces, the
      papillomavirus NRC will make 5μm thick sections using a microtome. The number of sections to
      be made depends on the size of the histological specimen. A minimum surface area equivalent
      to 5 sections of 5μm of a 125 mm2 piece is required for genotyping.

      Tissue sections are grouped in a labelled 2 mL screw tube. To avoid any contamination between
      samples, the microtome blade will be carefully cleaned with xylene and then 95% ethanol
      between each block.

      After DNA extraction using the QiaAmp DNA mini Kit (Qiagen), HPV genotype analysis will be
      performed using molecular techniques for the detection of human papillomavirus DNA. The
      investigators will use the INNO-LiPA HPV Genotyping Extra II (Fujirebio) kit. This is a
      Polymerase Chain Reaction (PCR) technique using degenerate primers followed by reverse
      hybridization on strips on which the specific probes of 32 alpha HPV genotypes are
      immobilized. This kit allows the detection of 13 high-risk HPV genotypes (HPV16, 18, 31, 33,
      35, 39, 45, 51, 52, 52, 56, 58, 59, 68), 6 HPVs that are likely to be at risk (HPV 26, 53,
      66, 70, 73, 82) and 14 low-risk or unclassified HPVs (HPV6, 11, 40, 42, 43, 44, 54, 61, 62,
      67, 81, 83, 89). In addition, the samples will be analyzed by PCR luminex according to the
      technique published by Schmitt et al, J Clin Microbiol 2010. In addition to alpha HPV, this
      technique allows the detection of sixty beta and gamma HPVs.

      Characteristics of the subjects to be included A description of patients included in the
      study (women born between 1972 and 1982, on the one hand, and women born between 1983 and
      1993, on the other) will include age at time of sampling, year of sampling, stage of lesion
      CIN2 or 3, and histological type.

      Comparisons between age groups will be made using Student tests for quantitative variables
      and Chi-square tests for qualitative variables, while respecting the conditions for applying
      these tests.

      Main judgment criteria Prevalences of high-risk HPV 16 or 18 genotypes will be described in
      both age groups with their 95% confidence intervals. These prevalences will be compared
      between the two age groups using the Chi-square test.

      Secondary Judgment Criterion Prevalences of high-risk HPV genotypes other than 16 or 18 will
      be described in both age groups with their 95% confidence intervals. These prevalences will
      be compared between the two age groups using the Chi-square test.

      Additional analyses The distribution of the different HPV genotypes will be described for
      both age groups.

      The number of samples required for the study is 606: 303 samples from women born between 1972
      and 1982 and 303 samples from women born between 1983 and 1993.

      The calculation of the number of samples required is based on the following assumptions: the
      prevalence of HPV16 and/or HPV18 in high-grade lesions of women is estimated at 62%. The
      minimum decrease after the introduction of HPV vaccination, interesting from a population
      health point of view, that the investigators wish to highlight is 10%, i.e. an expected
      prevalence among women born between 1983 and 1993 of 52%.

      For a power set at 80% and an alpha risk at 5%, a unilateral formulation of hypotheses (only
      a decrease in prevalence is possible) and an equal number of samples from women aged 25-34
      and 35-44, the number of samples required per group is 303.
    

Intervention name(s): biological sample analysis
Intervention type(s): Other
Intervention description(s): HPV Prevalence analysis
Observational model: Case-Control
Time perspective: Retrospective
Sampling method: Probability Sample
Study population: 
        Women born between 1972 and 1993
      
Keywords: cervical precancerous lesion; HPV; vaccination; Impact study; France