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Effectiveness of human papillomavirus vaccine against cervical precancer in Japan: Multivariate analyses adjusted for sexual activity

Abstract Japanese girls aged 12–16 years are offered free human papillomavirus (HPV) vaccination and cervical cancer screening is conducted with cytology and not HPV testing from the age of 20 years. So far, no study has analyzed the effect of HPV vaccination against cervical precancers considering HPV infection status and sexual activity. We aimed to analyze the vaccine effectiveness (VE) against HPV infection and cytological abnormalities, adjusted for sexual activity. This study comprised women aged 20–26 years who underwent cervical screening in Niigata. We obtained HPV vaccination status from municipal records and a questionnaire along with information concerning sexual activity. Of 5194 women registered for this study, final analyses included 3167 women in the vaccinated group (2821 vaccinated women prior to sexual debut) and 1386 women in the unvaccinated group. HPV 16/18 (0.2% vs 3.5%), 31/45/52 (3.4% vs 6.6%), and 31/33/45/52/58 (5.0% vs 9.3%) positive rates were significantly lower in the vaccinated group (P < 0.001). No women vaccinated before sexual debut had HPV 16/18‐related cytological abnormalities. VE for HPV 16/18 infection and high‐grade cytological abnormalities in women vaccinated prior to sexual debut were 95.8% (95% CI 81.9–99.0%; P < 0.001) and 78.3% (95% CI 11.3–94.7%; P = 0.033), respectively, in multivariate analyses adjusted for age and number of sexual partners. However, analyses of all vaccinated women did not show significant effectiveness against cytological abnormalities. Our results showed the effectiveness of HPV vaccine against high‐grade cervical cytological abnormalities and the importance of the vaccination before sexual debut.


| INTRODUC TI ON
In Japan, cervical cancer affects approximately 11,000 women per year and approximately 2900 women die of it annually, 1 with the number of patients and deaths gradually increasing. A rapid increase in the incidence of cervical cancer among young women has emerged as a social issue, particularly due to delayed marriage and childbirth. 2,3 The rate of cervical cancer screening, as a means of secondary prevention, remains low in Japan, at approximately 40%, as opposed to rates of 70%-80% in Europe and the United States. [4][5][6][7] The main method of primary prevention, namely, human papillomavirus (HPV) vaccination, began being publicly funded for 12-16-year-old girls in 2010 and was included in the national immunization program in April 2013. However, following several media reports of women experiencing various symptoms after vaccination, the Japanese Ministry of Health, Labour, and Welfare stopped actively recommending vaccination in June 2013, and the vaccination rate declined precipitously to 1%. [8][9][10] The HPV vaccine is still included as a routine childhood vaccine and, while it is provided free of charge to girls of a determined age, coverage continues to be extremely low. 11 Since the US FDA approved the HPV vaccine in 2006, national surveys globally have reported a decline in HPV infection rates, [12][13][14] a decrease in the rate of cytological abnormalities, 15 and a decline in the rate of histological abnormalities [16][17][18] as a result of the HPV vaccine. A reduction in the prevalence of invasive cervical cancer was first reported in Sweden in 2020, which has been followed by other reports from Demark and England. [19][20][21] In Japan, although a decrease in the rate of cytological and histological abnormalities due to HPV vaccination has been reported, [22][23][24][25][26] many challenges remain in terms of conducting population-based epidemiological research on the effectiveness of the HPV vaccine in Japan. First, >1700 individual municipalities individually manage vaccination records, and there is no national database. Second, reporting bias is an issue when relying on patient memory recall concerning vaccination history. 27 Third, cervical screening conducted by municipalities in Japan only involves cytology (Pap smear) and not primary HPV testing. Given these issues, large-scale epidemiological studies may not be capable of accurately evaluating the effectiveness of the HPV vaccine in Japan.
Since 2014, we have been conducting a large-scale epidemiological study in Niigata Prefecture aimed at overcoming the aforementioned challenges and obtaining an accurate assessment of HPV vaccine effectiveness in Japanese women (the Niigata study). In the Niigata study, official municipal vaccination records are checked, HPV testing is performed on registrants during cervical screening, and a questionnaire is used to obtain information on sexual activity. In an interim report of women aged 20-22 years, we reported bivalent vaccine effectiveness to be >90% in preventing HPV 16/18 infection. 28,29 The present study aimed to analyze the effectiveness of the HPV vaccine in preventing cervical cytological abnormalities in women aged 20-26 years, adjusted for sexual activity.

| Study design and target population
This population-based cross-sectional study (the Niigata study) comprised women aged 20-26 years (born in fiscal years 1992-1999) who have undergone cervical screening in six cities in Niigata Prefecture (Niigata, Nagaoka, Joetsu, Shibata, Sanjo, and Mitsuke) since April 2014. 28 Women born between April 1994 and March 2000 (fiscal years 1994-1999) were eligible to receive free HPV vaccination (50% subsidized by the central government; 50% subsidized by the municipality), starting in October 2010. Vaccination coverage in this age group is >70%. [8][9][10] In 2010, the cities of Niigata, Nagaoka, Joetsu, Shibata, and Mitsuke introduced public subsidies for girls born in or after 1994. In 2012, the city of Sanjo introduced public subsidies for girls born in or after 1996. The number of cases needed to examine the effectiveness of the HPV vaccine against the cytological abnormality of a high-grade squamous intraepithelial lesion (HSIL) or worse (HSIL+) was calculated as follows. Based on data from our previous research in Niigata Prefecture, we assumed that HSIL+ rates among unvaccinated women and women who had been vaccinated prior to sexual debut would be 0.66% and 0%, respectively, at a ratio of 1:2.2 (unvaccinated to vaccinated).
With a two-sided significance of 0.05 and a power of 0.8, we calculated that a total of 2966 participants were needed for analysis in this study.

| Vaccination status and sexual history
Japan does not have a national vaccination database, and vaccination records are managed by individual municipalities. Therefore, we obtained information on the date of vaccination, the number of doses administered, and the type of vaccine from the official vaccination records kept by the aforementioned six cities. Some registrants may have been vaccinated in a different municipality from their place of residence. Therefore, we also used a questionnaire to obtain their HPV vaccination status, along with information concerning sexual activity (age at sexual debut and number of previous sexual partners). The number of sexual partners was separated into the following five categories: 0, 1, 2-5, 6-9, and ≥ 10. We extracted "the vaccinated prior to sexual debut group" from the overall vaccinated group according to information on the date of vaccination and age at sexual debut.

| Statistical analyses
Data were analyzed using EZR (Jichi Medical University Saitama Medical Center), a graphical user interface of R (R Foundation for Data Computing, Vienna, Austria). 30 Categorical data were expressed as the overall number and percentage, and continuous data were expressed as the median or mean value. The age at sexual debut was analyzed as a continuous variable without grouping, and the number of sexual partners was analyzed in five groups: 0, 1, 2-5, 6-9, and 10 or more. Among the registrants, there was one woman who indicated that her age at sexual debut was less than 10 years old (age 8 years). To minimize the influence of outliers, this woman was excluded from the analysis. The Mann-Whitney U, Pearson's χ 2 , and Fisher's exact tests were used to compare baseline characteristics, HPV infection rates, and abnormal cytology rates in vaccinated and unvaccinated women. Univariate and multivariate logistic regression analyses were performed to evaluate vaccine effectiveness (VE) for HPV 16/18 infection and HSIL+. In the multivariate logistic regression analysis, Model 1 was adjusted for age and Model 2 was adjusted for age and total number of previous sexual partners. We set the "previous sexual partner = 0" group as the reference in Model 2. VE was calculated as 100 × [1 − odds ratio] along with the corresponding 95% confidence interval (CI). A P-value <0.05 was considered statistically significant.

| Vaccine effectiveness against human papillomavirus infection and cytological abnormality in vaccinated women prior to sexual debut
It is recommended that the HPV vaccine be administered prior to sexual debut to maximize its effectiveness. Therefore, we have added an additional analysis of the vaccine effectiveness against cytological abnormalities in vaccinated women prior to sexual debut, in addition to a comparison with all vaccinated women.

| DISCUSS ION
This study is the first in Japan to evaluate the effectiveness of HPV vaccination against cervical cytological abnormalities using accurate municipal vaccination data and taking HPV infection status and sexual activity into account. Using multivariate analyses adjusted for age and the number of sexual partners, we found a significant preventive effect against high-grade cytological abnormalities (HSIL+) for women who had received the HPV vaccine prior to sexual debut.
This study had the following three strengths. First, the analysis used accurate vaccination information obtained from municipalities (date of vaccination, type of vaccine, and number of doses administered). Second, the questionnaire survey of sexual activity allowed us to adjust for the number of sexual partners in the multivariate analysis and restrict some analyses to women who had been vaccinated prior to sexual debut. Adjusting for sexual activity, which is a confounding factor that has a considerable effect on HPV infection, has not been performed in most previous analyses from Japan and is likely to enhance the accuracy of the present study's findings on the effectiveness of the HPV vaccine. Third, HPV testing was performed at the same time as cytology examinations. Understanding HPV infection status made it possible to analyze cytological abnormalities associated with the HPV types targeted using the vaccines.
In an interim analysis as part of the Niigata study, we examined the effectiveness of the bivalent vaccine for preventing HPV  Self-reports did not match municipal records in 20.6% of cases.
Moreover, approximately 50% of women who had self-reported not having been vaccinated had actually been vaccinated but had forgotten. 27 Japan comprises many local governments, all of whom manage their own vaccination records and cervical screening results.
Therefore, due to the lack of a national registration system for vaccination and cervical screenings, and because personal information is not centrally managed, conducting large-scale epidemiological research at a national level is challenging. Therefore, previous stud- These studies obtained accurate vaccination data through checking municipal vaccination records; however, data concerning HPV infection status or sexual activity were not included in their analyses. As described above, previous analyses of vaccine effectiveness in Japan have not considered differences in sexual activity, leaving any associations with being vaccinated prior to or after sexual debut unclear.
The World Health Organization (WHO) has recommended that girls become vaccinated for HPV before they are sexually active, 32 and our findings support this recommendation for Japanese women. the WHO recommends vaccinating girls aged 9-14 years. After proactive recommendation for HPV vaccine resumes, it will be necessary to discuss the recommended age for routine HPV vaccination in Japan based on our results. As part of the Niigata study, we plan to further verify the effectiveness of the vaccines against histological abnormalities and analyze their long-term prevention effectiveness.

ACK N OWLED G M ENTS
We would like to thank Mr Kenshin Sekine and Mr Taishin Sekine for English editing, and Ms Yuka Watanabe, Ms Sachiko Ono, Ms Anna Ishida, and administrators of Niigata, Nagaoka, Joetsu, Shibata, Sanjo, Mitsuke city for their support in conducting the survey.