JNCI Cancer Spectrum (2019) 2(4): pky045 doi: 10.1093/jncics/pky045 Article ARTICLE Burden of Human Papillomavirus (HPV)-Related Cancers Attributable to HPVs 6/11/16/18/31/33/45/52 and 58 Silvia de Sanjose *,†, Beatriz Serrano*, Sara Tous, Maria Alejo, Bele n Lloveras, Beatriz Quiro  s, Omar Clavero, August Vidal, Carla Ferra  ndiz-Pulido, Miquel  Angel Pavo n, Dana Holzinger, Gordana Halec, Massimo Tommasino, Wim Quint, Michael Pawlita, Nubia Mun ~ oz, Francesc Xavier Bosch, Laia Alemany on behalf of RIS HPV TT, VVAP and Head and Neck study groups See the Notes section for the full list of authors’ affiliations. *Equally contributed.  is currently working in PATH as a Scale-up Project director, Sexual and Reproductive Health Global Program. †Silvia de Sanjose Correspondence to: Laia Alemany, PhD, Unit of Infections and Cancer, Cancer Epidemiology Research Program, Institut Catala  d’Oncologia (ICO)—Catalan Institute of Oncology, Gran Via de l’Hospitalet, 199-203, 08908 L’Hospitalet de Llobregat, Barcelona, Spain (e-mail: lalemany@iconcologia.net). Abstract Background: Many countries, mainly high- and upper-middle income, have implemented human papillomavirus (HPV) vacci- nation programs, with 47 million women receiving the full course of vaccine (three doses) in 2014. To evaluate the potential impact of HPV vaccines in the reduction of HPV-related disease, we aimed to estimate the HPV type distribution and burden of anogenital and head and neck cancers attributable to HPV types (HPVs 16/18/31/33/45/52/58/6/11) included in currently licensed HPV vaccines. Methods: In all, 18 247 formalin-fixed paraffin-embedded specimens were retrieved from 50 countries. HPV DNA detection and typing were performed with the SPF-10 PCR/DEIA/LiPA25 system. With the exception of cervical cancer, HPV DNA- positive samples were additionally subjected to HPV E6*I mRNA detection and/or p16INK4a immunohistochemistry. For cervi- cal cancer, estimates were based on HPV DNA, whereas for other sites, estimates were based on HPV DNA, E6*I mRNA, and p16INK4a biomarkers. Results: The addition of HPVs 31/33/45/52/58 to HPVs 16/18/6/11 in the nonavalent HPV vaccine could prevent almost 90% of cervical cancer cases worldwide. For other sites, the nonavalent HPV vaccine could prevent 22.8% of vulvar, 24.5% of penile, 60.7% of vaginal, 79.0% of anal cancers, 21.3% of oropharyngeal, 4.0% of oral cavity, and 2.7% of laryngeal cancer cases. Conclusions: Our estimations suggest a potential impact of the nonavalent HPV vaccine in reducing around 90% of cervical cancer cases and a global reduction of 50% of all the cases at HPV-related cancer sites. Human papillomavirus (HPV)-related cancers remain a major The global burden of cervical cancer (530 000 cases) is sub- cause of cancer both in men and women. Worldwide, HPV infec- stantially higher than that of HPV-related cancers other than tion causes up to 4.5% (640 000 cases) of all new cancer cases cervix (113 000 cases) (1,2). However, an increase in the inci- worldwide (8.6% females; 0.9% males), representing 29.5% of all dence of anal and HPV-related head and neck cancers (HNCs) infection-related cancers (1,2). has been noted in recent decades (3–6). Received: April 25, 2018; Revised: July 14, 2018; Accepted: July 27, 2018 © The Author(s) 2019. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/ licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com. 1 of 11 2 of 11 | JNCI Cancer Spectrum, 2019, Vol. 2, No. 4 HPV infection is unequivocally linked to almost all cervical that approximately 379 000 cases of cervical cancer and 156 000 cancer cases (95% of cases) and a large proportion of anal deaths could be averted in the 47 million fully vaccinated cancer cases (88% of cases attributable to HPV) (2). HPV is women by age 75 years, assuming lifelong protection (15). also causally associated with a varying percentage of cancers The evidence suggests that the combination of high HPV of the vulva, vagina, penis, and a subset of HNCs, particularly vaccination coverage in adolescents together with high cervical tonsillar cancer (7,8). Within the spectrum of HPV oncogenic screening coverage (followed by adequate treatment of the types, HPV 16 is the most prevalent in all HPV-related cancer lesions detected by screening) can eliminate cervical cancer as a sites. Based on previous analytical studies that demonstrated public health problem. As a result, it has been established that HPV to be a necessary cause of cervical cancer, HPV attribution one of the greatest priorities and challenges in global health is has been based on the detection of HPV DNA alone. However, to give access to HPV vaccination and cervical screening to the additional carcinogenic pathways other than HPV are sus- majority of women around the world. The availability of a wide pected for anal and vaginal cancers, and validated for vulvar, range of cervical screening techniques (cytology, HPV-detection penile, and HNCs. Thus, the identification of HPV DNA com- techniques, visual inspection with acetic acid) and HPV vaccina- bined with E6/E7mRNA, a marker of viral transcriptional activ- tion options (bivalent, tetravalent, and nonavalent vaccines ity, or p16INK4a, a cell surrogate marker of HPV carcinogenic with two- or three dose schedules) leads to an increase in the transformation, provides a more accurate, although conserva- complexity of decision making regarding the combination of op- tive, assessment of oncogenic activity of the virus in the spe- timal prevention strategies in each region of the world. cific sites (9–11). To evaluate the potential impact of HPV vaccines in the re- Prophylactic HPV vaccines have been introduced worldwide duction of HPV-related disease burden and to help formulate since 2006, with a high efficacy and safety record in the preven- recommendations on HPV prevention, we estimated the HPV tion of vaccine-type HPV infection and disease. Further, the HPV type distribution and burden of cancer cases attributable to the vaccines show a high effectiveness profile on prevalent infections, HPV types included in the nonavalent vaccine. For this we used precancerous lesions, and genital warts, as well as a robust herd- new data derived from a large international study that provided protection effect in nonvaccinated sexual partners with a high information on HPV DNA, mRNA, and p16INK4a from formalin- vaccine coverage rate (12,13). Currently, three HPV vaccines are fixed paraffin-embedded (FFPE) tissue of anogenital and HNC commercially available (bivalent, tetravalent, and nonavalent). All sites. three contain virus-like particles (VLPs) of HPVs 16/18. The tetra- valent vaccine also contains VLPs from HPVs 6/11. The nonava- lent vaccine additionally protects against HPVs 31/33/45/52/58. Materials and Methods The World Health Organization recognizes the importance of HPV-related diseases as global public health threats and has To estimate the prevalence and relative contribution (RC) of the reiterated the recommendation to include HPV vaccines in types included in licensed HPV vaccines in HPV-related cancers, national immunization programs (14). By October 2014, 80 coun- we used updated data from an international project designed tries (mainly high- and upper-middle income) had implemented and coordinated by the Catalan Institute of Oncology (ICO) HPV vaccination programs and 47 million women had received (Barcelona, Spain) in collaboration with DDL Diagnostic the full course of the vaccine. Even more, it has been estimated Laboratory (Rijswijk, Netherlands) (5,16–20). Cervix Vulva Vagina Penis Anus Oral cavity Oropharynx Larynx Austria Australia Argentina Belarus Bangladesh Bolivia Bosnia&Herzegovina China Brazil Croatia India Chile Czech Rep. Israel Colombia France Japan Equador Germany Kuwait Guatemala Greece Lebanon Honduras Italy New Zealand Mexico The Netherlands Algeria Pakistan Paraguay Poland Mali Philippines Peru Portugal Mozambique South Korea Uruguay Slovenia Nigeria Taiwan USA Spain Senegal Thailand Venezuela United Kingdom Uganda Turkey Figure 1. Anatomical sites included from each country. The number of cancer samples included at each anatomical site varies among countries. Table 1. General characteristics of included cases* General characteristics Cervix Vulva Vagina Anus Penis Oropharynx Oral cavity Larynx Regions (%) Eu (21%), Am Eu (53%), Am Eu (37%), Am (47%), Eu (34%), Am (51%), Eu (41%), Am (49%), Eu (74%), Am (16%), Eu (46%), Am (41%), Eu (48%), Am (38%), (39%), Af (9%), (22%), Af (1%), Af (5%), AO (11%) Af (4%), AO (11%) Af (2%), AO (8%) Af (1%), AO (9%) Af (5%), AO (8%) Af (7%), AO (7%) AO (31%) AO (24%) Period of diagnosis 1940–2010 1980–2011 1986–2011 1986–2011 1983–2011 1990–2012 1990–2012 1990–2012 Sex (%) Fem (66.3%), Fem (16.3%), Fem (37.3%), Fem (10.5%), Mal (31.7%) Mal (81.1%) Mal (61.8%) Mal (89.2%) Mean age (SD), y 51.3 (13.3) 68.5 (15.3) 61.3 (15.3) 62.8 (14.7) 64.1 (15.0) 61.0 (11.2) 61.4 (14.0) 61.8 (10.9) Histology, % SCC—90% SCC100WB—19% SCC100WB—31% SCC100WB—59% SCC100WB—23% SCC100WB—11% SCC100WB—3% SCC100WB—7% ADC—7% ADSC—2% SCC100NonWB—72% SCC100NonWB—51% SCC100NonWB -33% SCC100NonWB—64% SCC100NonWB—88% SCC100NonWB—96% SCC100NonWB—92% Other—2% SCC-Mixed—6% SCC-Mixed—4% SCC-Mixed—6% SCC-Mixed—7% SCC-Mixed—0% SCC-Mixed—0% SCCMixed—0% Other—3% Other—13% Other—2% Other—6% Other—1% Other—1% Other—1% No. Total 11 228 1709 408 496 1010 1090 1264 1042 HPV DNAþ No. positive (%) 9516 (84.8%) 488 (28.6%) 303 (74.3%) 438 (88.3%) 334 (33.1%) 271 (24.9%) 93 (7.4%) 59 (5.7%) Sin/mul/und % 93.0/6.3/0.7% 89.8/6.1/4.1% 95.7/4.0/0.3% 92.2/7.3/0.5% 89.2/9.0/1.8% 98.5/0.4/1.1% 87.1/5.4/7.5% 94.9/1.7/3.4% HPV DNAþ and (E6*I mRNA/p16INK4a) No. positive (%) NA 419 (24.6%) 289 (71.0%) 375 (82.6%) 280 (27.7%) 243 (22.4%) 55 (4.4%) 36 (3.5%) Sin/mul/und % NA 92.8/5.7/1.4% 96.2/3.8/0.0% 94.4/5.1/0.5% 90.0/8.9/1.1% 99.6/0.4/0.0% 96.4/3.6/0.0% 100.0/0.0/0.0% *ADC ¼ adenocarcinoma; ADSC ¼ adenosquamous carcinoma; Af ¼ Africa; AO ¼ Asia and Oceania; Am ¼ Americas; Eu ¼ Europe; Fem ¼ female; HPV ¼ human papillomavirus; Mal ¼ male; mul ¼ multiple infections; N ¼ number of cases; NA ¼ not analyzed; Other ¼ mainly ADC (for noncervical cancers, ADSC (for noncervical cancers, undifferentiated, and neuroendrocrine carcinomas; SCC ¼ squamous cell carcinoma; sin ¼ single infections; und ¼ undeter- mined infections; WB ¼ warty basaloid. HPV DNAþ & (E6*I mRNA/p16INK4a) ¼ cases that tested positive for HPV DNA and at least one of the following two markers: E6*I mRNA/p16INK4a. E6*I mRNA was performed on HPV DNA-posi- tive samples for types: 16/18/26/31/33/35/39/45/51/52/53/56/58/59/66/67/68/70/73/82. SCC100WB: SCC cases 100% with WB morphological features; SCC100NonWB: SCC cases 100% non-WB features; mixed histology includes SCC with variable percentages of WB and non-WB features. For more details, please see the methodological section from Castellsague  et al. 2016; de Sanjose et al. 2010; de Sanjose  et al. 2013; Alemany et al. 2014, Alemany et al. 2015, and Alemany et al. 2016 (5, 16–20). S. de Sanjose  et al. | 3 of 11 4 of 11 | JNCI Cancer Spectrum, 2019, Vol. 2, No. 4 Study Design prevalence was calculated as those positive among all tested samples. For the other cancer sites, RC estimates were based on The project is a cross-sectional study. Case recruitment proto- the combined positivity for HPV DNA and (E6*I mRNA or cols were previously described (5,16–20). Briefly, consecutive (or p16INK4a). HPV prevalence indicates those positive for HPV DNA randomly selected) FFPE specimens processed in pathology lab- and (E6*I mRNA or p16INK4a) among all those tested. For all sites, oratories were obtained from hospital pathology archives in information on multiple infections was added to single types in 50 countries (Figure 1). Other variables collected were sex, age at accordance with a proportional weighting attribution (22,23). diagnosis, year at diagnosis, and original histological diagnosis. RCs by region, sex, and age at diagnosis were determined. To This article uses information from 11 228 cervical, 496 anal, 408 evaluate differences among proportions, we used the most suit- vaginal, 1709 vulvar, 1010 penile, 1090 oropharyngeal, 1264 oral able test (chi-squared, Fisher), adjusting for multiple compari- cavity, and 1042 laryngeal cancer cases. Table 1 summarizes the sons (Bonferroni–Holms) when necessary. Trend test analysis main characteristics of the tumor series. for proportions was used to evaluate trends by age at diagnosis. The statistically significant P value was set at .05. We further estimated the potential preventable number of Pathology and Laboratory Procedures: FFPE Block cases per year by applying the estimated proportions attributed Processing, Histological Evaluation, and HPV Detection to HPV to the number of cases estimated to occur each year as and Genotyping reported by de Martel et al. (24). FFPE blocks were processed under strict conditions. At least five paraffin sections were systematically obtained for each block (sandwich method). First and last sections were used for histo- Results pathological evaluation after hematoxylin and eosin staining. Squamous cell carcinomas (SCCs) accounted for more than 90% The intermediate sections were used for HPV DNA and mRNA of cases reviewed, with the exception of vaginal cancer. Most testing. HPV DNA detection was performed by polymerase chain cases were from Europe and the Americas (Table 1). Cervical reaction (PCR) with SPF-10 broad-spectrum primers. The ampli- cancer cases were diagnosed at younger ages (mean age ¼ fied PCR products were tested for the presence of HPV DNA us- 51.1 years) compared to the other sites (mean ages ranging from ing a DNA enzyme immunoassay (DEIA) that recognized at least 61.0 to 68.5 years). 54 mucosal HPV genotypes. DNA quality was evaluated by PCR Overall HPV and type-specific prevalence results are testing for the human tubulin in all HPV DNA-negative vaginal, reported in Table 1 and Figures 2 and 3. The HPV prevalence for anal, and HNCs, and a subset of samples of cervical, vulvar, and all types combined was 84.8% in cervical cancer cases (HPV DNA penile cases (almost all HPV DNA-negative cervical cancer cases alone). For the other cancer sites, the HPV prevalence (HPV DNA were due to technical artifacts, and 10% of the vulvar and penile cancers were due to poor DNA quality). Vaginal, anal, and HNCs that were negative both for HPV DNA and tubulin were excluded from the final analyses. Amplimers testing positive for viral Cervix 84.8% DNA by DEIA were genotyped with a reverse hybridization line probe assay—LiPA25 that detects 25 HPV types (HPVs 6/11/16/ Anus 88.3% 18/31/33/34/35/39/40/42/43/44/45/51/52/53/54/56/58/59/66/68/70/ 82.6% 74) (21). Sequence analysis was performed to characterize HPV Vagina 74.3% DEIA-positive samples with unknown types and to disentangle HPVs 68/73 because the LiPA system is unable to separate these 71.0% types. If no HPV type could be attributed after DNA sequencing, 33.1% Penis the HPV type was labeled as undetermined. 27.7% With the exception of cervical cancer cases, all HPV DNA- positive samples, and a random selection of HPV DNA-negative Vulva 28.6% cases for quality control, were further subjected to HPV E6*I 24.6% mRNA detection, a biomarker of HPV viral transcriptional activ- Oropharynx 24.9% ity, and p16INK4a immunohistochemistry, a cellular surrogate 22.4% marker for HPV carcinogenic transformation. The E6*I mRNA as- say targets 20 HPV types (HPVs 16/18/26/31/33/35/39/45/51/52/ 7.4% Oral Cavity 53/56/58/59/66/67/68/70/73/82). For each case, type-specific E6*I 4.4% mRNA PCR was performed for all types detected by the SPF-10 5.7% PCR/DEIA/LiPA25 system that had at least one of the mRNA tar- Larynx geted types, and additionally for HPV 16. p16INK4a was detected 3.5% using the CINtec histology kit (clone E6H4, Roche mtm laborato- 0 10 20 30 40 50 60 70 80 90 100 ries AG, Germany) following the manufacturer’s protocol. A pat- Prevalence (%) tern of diffuse staining of more than 25% stained cells (nuclear and cytoplasmic) was considered positive. DNA+ HPV DNA+(E6*I mRNA/p16INK4a) 95%CI Figure 2. Overall HPV positivity for HPV DNA and HPV DNA þ (E6*I mRNA or Statistical Analysis p16INK4a) by anatomical site. 95% CI ¼ 95% confidence interval (one-sided, 97.5% CI calculated when appropriate; HPV ¼ human papillomavirus. For more details, For cervical cancer, RC of the nine types included in the nonava- please see the methodological section from Castellsague  et al. 2016; de Sanjose lent vaccine was expressed as the proportion of women positive  et al. 2013; Alemany et al. 2014, Alemany et al. 2015, and et al. 2010; de Sanjose for a specific type among all HPV DNA-positive samples. HPV Alemany et al. 2016 (5,16–20).  et al. | 5 of 11 S. de Sanjose Both sexes Female Male Cervix Vulva Vagina Anus Penis Oropharynx Oral Cavity Larynx 0 10 20 30 40 50 60 70 80 90 100 0 10 20 30 40 50 60 70 80 90 100 0 10 20 30 40 50 60 70 80 90 100 Prevalence (%) Both Bothsexes sexes Female Female Male Male Cervix Vulva Vagina Anus Penis Oropharynx Oral cavity Larynx 0 10 20 30 40 50 60 70 80 90 100 0 10 20 30 40 50 60 70 80 90 100 0 10 20 30 40 50 60 70 80 90 100 Relative Contribution (%) HPVs 16/18 HPVs 6/11 HPVs 31/33/45/52/58 Other HPVs 95%CI Prev (Any HPV) 95%CI RC (HPVs 16/18/31/33/45/52/58/6/11) Figure 3. Worldwide HPV prevalence and relative contribution in HPV-related cancers, by sex. 95% CI ¼ 95% confidence interval (one-sided, 97.5% CI calculated when appropriate); Cervix ¼ based on HPV DNA; HPV ¼ human papillomavirus; Other locations ¼ based on information on three markers (HPV DNA + (E6*I mRNA or p16INK4a)); Prev ¼ prevalence and type-specific relative contribution estimations. For more details, please see the methodological section from Castellsague  et al. 2016;  et al. 2013; Alemany et al. 2014, Alemany et al. 2015, and Alemany et al. 2016 (5, 16–20). de Sanjose et al. 2010; de Sanjose and “E6*I mRNAþ or p16INK4aþ”) for all types combined was to HPV 16 (90.9%, 88.5%, and 83.7%, respectively). RCs of HPVs 82.6% anal, 71.0% vaginal, 27.7% penile, 24.6% vulvar, 22.4% oro- 16/18 in other anogenital and HNCs was 79.0% in the vulva, fol- pharyngeal, 4.4% oral cavity, and 3.5% laryngeal cancers. When lowed by penis (76.6%), cervix (70.9%), vagina (64.3%), and larynx comparing HPV DNA detection estimates to the ones adding (52.8%). The additional contribution of HPVs 31/33/45/52/58 was E6*I mRNA and p16INK4a, a reduction of the proportion of cases higher in the larynx (22.2%), vagina (21.0%), and cervix (18.3%) attributed to HPV were higher for oral cavity and larynx (per- compared to other cancer sites (Tables 2 and 3). HPV 16 was by centage reduction ¼ 40.5% and 38.6%, respectively) than penis, far the most frequently detected genotype across all anatomical vulva, oropharynx, anus, and vagina (percentage reduc- sites. RCs by individually tested types are detailed in tion ¼ 16.3%, 14.0%, 10.0%, 6.4% and 4.4%, respectively), (P < .05 Supplementary Material Table 1 (available online). RC of the for oral cavity, larynx, penis, vulva, and anus). nine HPV types was significantly higher when taking into ac- Among HPV positives, RC of the nine types was 95.7% in the count the combined detection of HPV DNA, and E6*I mRNA or anus, followed by oropharynx (95.1%), vulva (92.8%), oral cavity p16INK4a for vulvar and oropharyngeal cancers as compared to (92.7%), cervix (89.3%), penis (88.1%), vagina (85.6%), and larynx HPV DNA detection alone (Tables 2 and 3). (77.8%) (Tables 2 and 3 and Figure 2). RC of HPVs 16/18 was espe- HPV RCs by region are provided in Figure 4 and cially prominent in oral cavity, oropharyngeal, and anal cancer Supplementary Material Tables 2 and 3 (available online). sites (90.9%, 90.5%, and 87.2%, respectively), mostly attributable HPV 16 was again the dominant type in HPV-related cancers in Table 2. Worldwide relative contribution of HPVs 16/18/31/33/45/52/58/6 and 11 in anogenital HPV-related cancers Vulva Vagina Anus Penis Cervix and (E6*I mRNA/ and (E6*I mRNA/ and (E6*I mRNA/ and (E6*I mRNA/ p16INK4a) p16INK4a) p16INK4a) p16INK4a) DNAþ(n ¼ 9516) DNAþ(n ¼ 488) (n ¼ 419) DNAþ(n ¼ 303) (n ¼ 289) DNAþ(n ¼ 438) (n ¼ 375) DNAþ(n ¼ 334) (n ¼ 280) HPV type RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) 6 of 11 | JNCI Cancer Spectrum, 2019, Vol. 2, No. 4 Combined nine HPV types† 89.3 (88.7 to 89.9) 87.1 (83.8 to 89.9) 92.8 (89.9 to 95.1) 85.3 (80.6 to 89) 85.6 (80.9 to 89.3) 95.3 (92.8 to 97.0) 95.7 (93.2 to 97.5) 84.4 (80.1 to 88.1) 88.1 (83.8 to 91.7) HPVs 16/18 70.9 (69.9 to 71.8) 72.6 (68.4 to 76.5) 79.0 (74.8 to 82.8) 63.7 (58.0 to 69.1) 64.3 (58.5 to 69.9) 84.3 (80.5 to 87.5) 87.2 (83.4 to 90.4) 70.2 (64.8 to 74.9) 76.6 (71.4 to 81.6) HPVs 31/33/45/52/58 18.3 (17.6 to 19.1) 13.0 (10.2 to 16.4) 13.6 (10.5 to 17.3) 20.1 (15.8 to 25.1) 21.0 (16.5 to 26.3) 8.0 (5.6 to 10.9) 8.0 (5.5 to 11.2) 9.1 (6.2 to 12.7) 10.7 (7.3 to 14.9) Specific HPV type: HPV 16 60.3 (59.4 to 61.3) 68.0 (63.7 to 72.2) 74.0 (69.5 to 78.1) 58.7 (53.0 to 64.3) 59.4 (53.6 to 65.2) 80.7 (76.8 to 84.4) 83.7 (79.6 to 87.3) 68.7 (63.3 to 73.5) 75.3 (69.9 to 80.3) HPV 18 10.5 (9.9 to 11.2) 4.6 (2.8 to 6.7) 5.0 (3.1 to 7.6) 5.0 (2.8 to 8.0) 4.9 (2.7 to 8.0) 3.6 (2.1 to 5.9) 3.5 (1.9 to 5.9) 1.5 (0.5 to 3.5) 1.4 (0.4 to 3.6) HPV 31 3.6 (3.2 to 4.0) 1.3 (0.5 to 2.7) 1.0 (0.3 to 2.4) 5.3 (3.0 to 8.4) 5.5 (3.2 to 8.8) 1.9 (0.8 to 3.6) 1.1 (0.3 to 2.7) 0.8 (0.2 to 2.6) 0.9 (0.2 to 3.1) HPV 33 3.7 (3.4 to 4.1) 5.9 (4.0 to 8.4) 6.5 (4.3 to 9.2) 4.8 (2.8 to 8.0) 4.7 (2.7 to 8.0) 2.7 (1.4 to 4.7) 3.1 (1.7 to 5.5) 2.9 (1.4 to 5.4) 3.2 (1.5 to 6.0) HPV 45 6.1 (5.6 to 6.6) 2.9 (1.6 to 4.8) 3.4 (1.8 to 5.5) 3.6 (1.8 to 6.4) 3.8 (1.9 to 6.7) 0.9 (0.2 to 2.3) 1.1 (0.3 to 2.7) 2.7 (1.2 to 5.1) 3.2 (1.5 to 6.0) HPV 52 2.7 (2.4 to 3.1) 1.8 (0.8 to 3.5) 1.7 (0.7 to 3.4) 2.9 (1.4 to 5.6) 3.1 (1.4 to 5.8) 0.7 (0.1 to 2.0) 0.5 (0.1 to 1.9) 1.5 (0.5 to 3.5) 1.9 (0.6 to 4.1) HPV 58 2.2 (1.9 to 2.5) 1.1 (0.3 to 2.4) 1.0 (0.3 to 2.4) 3.6 (1.8 to 6.4) 3.8 (1.9 to 6.7) 1.8 (0.8 to 3.6) 2.1 (0.9 to 4.2) 1.3 (0.3 to 3.0) 1.6 (0.4 to 3.6) HPV 6 0.1 (0.1 to 0.2) 1.1 (0.3 to 2.4) 0.3 (0.0 to 1.3) 1.0 (0.2 to 2.9) 0.0 (0.0 to 1.3) 1.8 (0.8 to 3.6) 0.5 (0.1 to 1.9) 3.7 (1.9 to 6.2) 0.7 (0.1 to 2.6) HPV 11 0.0 (0.0 to 0.1) 0.5 (0.0 to 1.5) 0.0 (0.0 to 0.9) 0.3 (0.0 to 1.8) 0.3 (0.0 to 1.9) 1.1 (0.4 to 2.6) 0.0 (0.0 to 1.0) 1.5 (0.5 to 3.6) 0.0 (0.0 to 1.3) †Nine HPV types includes the ones in the nonavalent HPV vaccine: HPVs 16/18/31/33/45/52/58/6/11. 95% CI ¼ 95% confidence interval (one-sided, 97.5% CI calculated when appropriate; DNAþ ¼ cases that tested positive for HPV DNA; E6*I mRNA/p16INK4a ¼ cases that tested positive for HPV DNAþ (E6*I mRNA positive or p16INK4a positive; HPV ¼ human papillomavirus; N ¼ number of cases; RC ¼ relative contribution. E6*I mRNA was performed in HPV-DNA- positive samples for types: HPVs 16/18/26/31/33/35/39/45/51/52/53/56/58/59/66/67/68/70/73/82. Type-specific RC estimations are estimated among HPV DNA positive and among HPV DNA positive þ (E6*I mRNA positive or p16INK4a positive). For cervix, RC estimations are calculated only among DNA positive. For more details, please see methodological section from de Sanjose et al. 2010; de Sanjose  et al. 2013; Alemany et al. 2014, Alemany et al. 2015, and Alemany et al. 2016 (16–20).  et al. | 7 of 11 S. de Sanjose Table 3. Worldwide relative contribution of HPVs 16/18/31/33/45/52/58/6 and 11 in head and neck HPV-related cancers Oropharynx Oral cavity Larynx and (E6*I mRNA/ and (E6*I mRNA/ and (E6*I mRNA/ p16INK4a) p16INK4a) p16INK4a) DNAþ(n ¼ 271) (n ¼ 243) DNAþ(n ¼ 93) (n ¼ 55) DNAþ(n ¼ 59) (n ¼ 36) HPV type RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) RC% (95% CI) Combined nine HPV types† 90.0 (85.8 to 93.3) 95.1 (91.5 to 97.4) 81.7 (72.4 to 89.0) 92.7 (82.4 to 98.0) 83.1 (71.0 to 91.6) 77.8 (60.8 to 89.9) HPVs 16/18 85.2 (80.4 to 89.2) 90.5 (86.1 to 93.9) 72.0 (61.8 to 80.9) 90.9 (80.0 to 97.0) 57.6 (44.1 to 70.4) 52.8 (35.5 to 69.6) HPVs 31/33/45/52/58 8.6 (3.8 to 16.2) 4.5 (2.3 to 8.0) 4.4 (2.3 to 7.6) 1.8 (0.0 to 9.7) 16.9 (8.4 to 29.0) 22.2 (10.1 to 39.2) Specific HPV type HPV 16 83.4 (78.4 to 87.6) 88.5 (83.8 to 92.2) 70.9 (60.6 to 79.9) 90.9 (80.0 to 97.0) 50.8 (37.5 to 64.1) 44.4 (27.9 to 61.9) HPV 18 1.8 (0.6 to 4.3) 2.1 (0.7 to 4.7) 1.1 (0.0 to 5.8) 0.0 (0.0 to 6.5) 6.8 (1.9 to 16.5) 8.3 (1.8 to 22.5) HPV 31 0.0 (0.0 to 1.4) 0.0 (0.0 to 1.5) 0.0 (0.0 to 3.9) 0.0 (0.0 to 6.5) 3.4 (0.4 to 11.7) 2.8 (0.1 to 14.5) HPV 33 3.3 (1.5 to 6.2) 3.3 (1.4 to 6.4) 0.0 (0.0 to 3.9) 0.0 (0.0 to 6.5) 3.4 (0.4 to 11.7) 2.8 (0.1 to 14.5) HPV 45 0.4 (0.0 to 2.0) 0.4 (0.0 to 2.3) 0.0 (0.0 to 3.9) 0.0 (0.0 to 6.5) 8.5 (2.8 to 18.7) 13.9 (4.7 to 29.5) HPV 52 0.0 (0.0 to 1.4) 0.0 (0.0 to 1.5) 7.6 (3.1 to 14.9) 0.0 (0.0 to 6.5) 0.0 (0.0 to 6.1) 0.0 (0.0 to 9.7) HPV 58 0.7 (0.1 to 2.6) 0.8 (0.1 to 2.9) 1.1 (0.0 to 5.8) 1.8 (0.0 to 9.7) 1.7 (0.0 to 9.1) 2.8 (0.1 to 14.5) HPV 6 0.4 (0.0 to 2.0) 0.0 (0.0 to 1.5) 0.0 (0.0 to 3.9) 0.0 (0.0 to 6.5) 6.8 (1.9 to 16.5) 2.8 (0.1 to 14.5) HPV 11 0.0 (0.0 to 1.4) 0.0 (0.0 to 1.5) 1.1 (0.0 to 5.8) 0.0 (0.0 to 6.5) 1.7 (0.0 to 9.1) 0.0 (0.0 to 9.7) †Nine HPV types includes the ones in nonavalent HPV vaccine: HPVs 16/18/31/33/45/52/58/6/11. 95% CI ¼ 95% confidence interval (one-sided, 97.5% CI calculated when appropriate); DNAþ ¼ cases that tested positive for HPV DNA; E6*I mRNA/p16INK4a ¼ cases that tested positive for HPV DNAþ (E6*I mRNA positive or p16INK4a positive; HPV ¼ human papillomavirus; N ¼ number of cases; RC ¼ relative contribution. E6*I mRNA was performed in HPV-DNA–positive samples for types: HPVs 16/18/26/31/ 33/35/39/45/51/52/53/56/58/59/66/67/68/70/73/82. Type-specific RC estimations are estimated among HPV DNA positive and among HPV DNA positive þ (E6*I mRNA pos- itive or p16INK4a positive). For more details, please see the methodological section from Castellsague  et al. 2016 (5). all regions. Within cervical cancer, HPV 16 RC ranged from estimates to be used to monitor the vaccine impact worldwide. 47.3% (Africa) to 65.5% (Europe); in vulvar cancer from 66.7% We present data from all HPV-related sites, updating the num- (Americas) to 87.5% (Africa); in vaginal cancer from 46.2% ber of the cervical series, and adding the evaluation of molecu- (Africa) to 66.9% (Europe); in anal cancer from 66.7% (Africa) to lar and immunohistochemistry markers of viral activity beyond 87.3% (Europe), in penile cancer from 71.4% (Africa) to 78.9% HPV DNA detection. In the absence of a unique carcinogenic (Europe), in oropharyngeal cancer from 78.1% (Americas) to pathway and of detection of specific infections leading to can- 94.7% (Asia and Oceania), in oral cavity cancer 90.9% (Americas cer, the addition of these markers increases our ability to attrib- and Europe), and in larynx cancer from 0.0% (Africa based on ute causality (8,11). Our data confirm the biological role of HPV small number of samples) to 58.3% (Europe). Although cumula- in the great majority of HPV DNA-positive anogenital and oro- tive RC of the nine types varied slightly by region, differences pharyngeal cancers, and to a much lesser extent in oral cavity were not statistically significant when comparing regions with and laryngeal cancers. world data. The only exception was observed in cervical cancer We have estimated that the combined detection of HPV DNA in Africa and Asia and Oceania (World—89.3%, Africa—86.3%, and (E6*I mRNA or p16INK4a) confirms that the addition of HPVs Asia and Oceania—91.2%; P < .05). 31/33/45/52/58 to the current vaccine types (HPVs 16/18/6/11), Differences by sex were not detected in the combined RC of could prevent almost 96% of the HPV-positive cancers of the HPVs 16/18, HPVs 31/33/45/52/58, or the nine HPVs for anal and anus, vulva (93%), cervix (90%), penis (88%), vagina (86%), oro- HNCs, except for a higher RC of HPVs 16/18 in male oropharynx pharynx (95%), oral cavity (93%), and larynx (78%). A marked (P < .05) (Figure 3). heterogeneity in the incremental HPV-related preventable frac- No specific nine HPV RC age trends were observed by cancer tion added by the nonavalent vaccine compared to bivalent and site, except in the cervix (Supplementary Material Table 4, avail- tetravalent vaccines was observed, ranging from 8.0% (anal) to able online), where the combined RC of the nine types decreased 21.0% (vaginal) in anogenital cancer sites, and from 1.8% (oral with age (P < .001); mainly explained by the decrease of HPVs 16/ cavity) to 22.2% (larynx) in HNC sites. 18/45 in older ages (more details in Serrano et al. 2012) (25). Similar to previous data, overall HPV prevalence was lower The potentially preventable fraction of the nonavalent HPV for vulvar (24.6%), penile (27.0%), oral cavity (4.4%), laryngeal vaccine would be around 50% worldwide, taking into account (5.7%), and oropharyngeal (22.4%) cancers, compared with vagi- our type-specific prevalence estimates and the estimated num- nal (71.0%), anal (82.6%), and cervical (84.8%) cancers (2). ber of incident cases at HPV-related cancer sites provided by de Therefore, overall the nonavalent vaccine could prevent 22.8% Martel et al. (24). These would include most cervical and anal of vulvar, 24.5% of penile, 60.7% of vaginal, 79.0% of anal, and cancers and a substantial fraction of vaginal, penile, vulvar, and 90% of cervical cancer cases, 21.3% of oropharynx, 4.0% of oral HNCs (Table 4). cavity, and 2.7% of larynx cancers cases. Some of our prevalence estimates are lower than those published in previous meta- analyses (6,26) or in series using similar protocols (27–30). These differences were expected, as we used a more restrictive proto- Discussion col in terms of HPV attribution by adding additional markers. This comprehensive analysis of the contribution of HPV vaccine This study included cases from numerous countries, some of types in selected anogenital and HNCs provides robust which had scarce data published. This is particularly relevant in 8 of 11 | JNCI Cancer Spectrum, 2019, Vol. 2, No. 4 Europe Americas Cervix (n=2058) Cervix (n=3564) Vulva (n=141) Vulva (n=135) Vagina (n=106) Vagina (n=139) Anus (n=142) Anus (n=184) Penis (n=119) Penis (n=142) Oropharynx (n=160) Oropharynx (n=64) Oral cavity (n=22) Oral cavity (n=33) Larynx (n=12) Larynx (n=22) Africa Asia and Oceania Cervix (n=838) Cervix (n=3056) Vulva (n=16) Vulva (n=127) Vagina (n=13) Vagina (n=31) Anus (n=9) Anus (n=40) Penis (n=7) Penis (n=12) Oropharynx Oropharynx (n=19) Oral cavity Oral cavity Larynx (n=2) Larynx 0 10 20 30 40 50 60 70 80 90 100 0 10 20 30 40 50 60 70 80 90 100 Relative Contribution (%) HPVs 16/18 HPVs 6/11 HPVs 31/33/45/52/58 95%CI HPVs 16/18/31/33/45/52/58/6/11 Figure 4. HPV relative contribution of types included in licensed HPV vaccines in HPV-related cancers, by region. 95% CI ¼ 95% confidence interval (one-sided, 97.5% CI calculated when appropriate); HPV ¼ human papillomavirus; n ¼ number of cancer cases represented in the region. Type-specific relative contribution estimations: Cervix ¼ based on HPV DNA positive; Other locations ¼ based on information on three markers (HPV DNA positive þ (E6*I mRNA or p16INK4a)). For more details, please  et al. 2016; de Sanjose et al. 2010; de Sanjose see the methodological section from Castellsague  et al. 2013; Alemany et al. 2014, Alemany et al. 2015, and Alemany et al. 2016 (5, 16–20). Table 4. Preventable fraction of human papillomavirus (HPV)-related cancer cases per year Cases attributable Cases attributable HPV-related cancer site Incident cases, No.* to HPV, No. Nine HPV types, % to HPV, No. Nine HPV types, % Cervix uteri 530 000 405 450† 76.5† 477 000‡ 90.0‡ Anus 40 000 31 600 79.0 31 600 79.0 Vulva 34 000 7752 22.8 7752 22.8 Vagina 15 000 9105 60.7 9105 60.7 Penis 26 000 6370 24.5 6370 24.5 Oropharynx 96 000 20 448 21.3 20 448 21.3 Oral cavity 200 000 8000 4.0 8000 4.0 Larynx 160 000 4320 2.7 4320 2.7 Total HPV-related sites 1 101 000 493 045 564 595 Potentially preventable cases, % 44.8% 51.3% *Source: de Martel, 2017 †Assumption: 85% HPV DNA-positivity and 90% relative contribution (RC) of nine HPV types. ‡Assumption: HPV is a necessary cause of invasive cervical cancer; and 90% RC of nine HPV types. HNC sites where a substantial variability in the HPV-related at- attributable fractions in HNCs have been recently reported (30). tribution has been observed across regions (2). Contrary to this, Although the exclusive use of HPV DNA results in higher attri- our series have a distinctive limitation on samples from areas bution to HPV (30), when adding other biomarkers the attribu- reported to have an increased trend of HPV-driven HNCs, such tion was reduced to 67.2% in oropharynx SCC, 5.9% in oral as the United States and Northern Europe, where higher cavity SCC, and 1.7% in larynx SCC (27–29).  et al. | 9 of 11 S. de Sanjose Worldwide incidence rates at HPV-related cancer sites cannot completely rule out that we are not excluding some other than the cervix are much lower than that observed for truly HPV-driven cases because of imperfect detection of the HPV cervical cancer. However, incidence rates in anal, vulvar, and DNA test, although we used the most sensitive test available to HPV-related HNCs are increasing, probably linked to changes date. Moreover, although HPV E6*I mRNA and p16INK4a together in sexual behavior, tobacco and alcohol consumption, and in with HPV DNA are well established markers in research and even cervical cancer screening policies (2–4).The large contribution in clinical settings for specifically attributing one case to be etio- of HPV infection, mainly of HPV 16, the absence of well- logically related to the virus, we are missing other potential established screening instruments, and algorithms for the approaches such as laser capture microdissection that can specif- detection of HPV-related cancer cases other than the cervix, ically analyze the markers in tumoral cells instead of the whole and the current evidence on HPV vaccine efficacy, reinforces paraffin-embedded tissue block section that may be even more the role that HPV vaccines can play in primary prevention of specific. However, we have applied the most suitable markers HPV-related lesions. Moreover, even if a higher impact of HPV analyses taking into account the design of the study, a large epi- vaccination would be expected in cervical and anal cancer, the demiological biorepository. Another limitation of the study was confirmed increasing trends of HNCs attributable to HPV could the small number of samples included from some regions (such be translated to a high number of cases prevented by vaccina- as Africa) and ages, thus restricting the validity of our results for tion despite the low proportion of HPV-related cases estimated certain regions and ages. Finally, the use of fresh tissue samples for these sites (2). would have been preferred but unrealistic to obtain when includ- HPV 16 was the most important oncogenic type (60% in ing such a large number of countries. Additionally, the studies anogenital cancer sites, and 44% to 91% in HNCs). The inclusion reflect the epidemiology of each country and the impact of poten- of additional markers increased the HPV 16 RC in vulvar, penile, tial variation in screening or other environmental modifications oral cavity, and oropharyngeal cancers, and confirmed the caus- in HPV attribution is difficult to fully evaluate in this study. ative role for other, less frequently occurring mucosal HPV Finally, for the estimated preventable fraction we make several types. Some non-HPV 16 types, usually involved in cervical can- vaccine effects assumptions: efficacy for all HPV-related cancers, cer, are also related to other cancers of the genital tract and long-term duration of the protection, among others. HNCs, but to a lesser extent. Although HPV18 is reported to be Our estimations suggest that if HPV vaccination programs the second most common type in cervical cancer, variations with the nonavalent HPV vaccine are effectively implemented, were observed in other cancer sites. HPV 33 follows HPV 16 in around 90% of cervical cancers and 50% of all HPV cancer sites vulvar and oropharyngeal cancers, HPV 31 in vaginal cancer, related to HPV could be prevented. HPVs 31/45 in penile cancer, HPVs 18/33 in anal cancer, HPV 58 in oral cavity cancer, and HPVs 18/45 in laryngeal cancers. However, results in oral cavity and larynx should be interpreted Funding with caution because of the limited number of cases included. This analysis was partially supported by Merck & Co Inc. HPVs 6/11, contribution is minimal in the overall burden of HPV-related cancers, although they still rank relatively high in Data used for this analysis are derived from the RIS HPV TT, penile and anal cancer. VVAP, and Head and Neck studies, which received funds We found little heterogeneity by geographical region, sex, from: Spanish public grants from the Instituto de Salud and age at diagnosis. Regional differences in the HPV RCs were Carlos III (CIBERESP CB06/02/0073 and CIBERONC CB16/12/ significant only for cervical and oropharyngeal cancers. For cer- 00401, FIS PI030240, PI061246, PI081535, PI1102096, PI1102104) vical cancer a significantly higher contribution was observed in cofunded by FEDER funds, European Regional Development Asia and Oceania, and for oropharyngeal cancer a significantly Fund (ERDF)—a way to build Europe; with the support of the higher contribution was observed in Europe compared to the Secretariat for Universities and Research of the Department Americas. However, further research will be necessary to pro- of Business and Knowledge of the Government of Catalonia; vide robust estimates by region for noncervical lesions. Overall, grants to support the activities of research groups (SGR 2017– HPV RC estimates were higher in women than in men, although 2019); grant number 2017SGR1085; Asociacio  n Espan ~ ola differences were significant only for oropharyngeal cancer. The Contra el Ca  ncer (personal grant to LA), Marato  de TV3 magnitude of RCs decreased with increasing age for all cancer Foundation (051530), European Commission (7th Framework sites. RC age trends were evaluated for the nine types combined. Programme Grant HEALTH-F2-2011-282562 HPV-AHEAD); A similar analysis for individual HPV types was not possible be- Lilly Foundation (Premio de Investigacio  n Biome dica cause of the limited number of cases in cancer sites other than Preclınica 2012 FXB); Stichting Pathologie On twikkelingen the cervix. Assessment of multiple infections was possible by Onderzoek Foundation (Netherlands) and from means of a proportional weighting attribution (22,23). A decrease GlaxoSmithKline (GSK) Biologicals, Sanofi Pasteur MSD, and in the proportion of cases with multiple infections was observed Merck, which had no role in the data collection, analysis, in- in oropharyngeal cancers. The proportion of cases with multiple terpretation of the results, or submission of the article for infections (0.4%) was 15 times lower than in cervical cancer publication. (6.3%). It is unknown why there is this difference in the propor- tion of multiple infections, but it could be explained by differen- Affiliations of authors: Cancer Epidemiology Research ces in cell susceptibility to HPV infection in different tissues. Program, ICO; Bellvitge Biomedical Research Institute Despite the large sample size and the strong design, this (IDIBELL), Gran Via de l’Hospitalet, 199-203, 08908 study is not free of limitations. The principal one is that while L’Hospitalet de Llobregat, Barcelona, Spain (SDS, BS, ST, BQ, the presence of HPV DNA was assessed in all tested samples, OC, MAP, FXB, LA); Centro de Investigacio  n Biome dica en the two additional biomarkers E6*I mRNA and p16INK4a were Red de Epidemiologıa y Salud Pu  blica (CIBERESP), Madrid, assessed only in HPV DNA-positive samples as well as in a Spain (SDS, LA); Centro de Investigacio  n Biome  dica en Red small fraction of HPV DNA-negative cases (of which 10% were de Ca ncer (CIBERONC), Madrid, Spain (BS, ST, BQ, OC, MAP, mRNA positive), and not for all HPV genotypes. Therefore, we FXB); PATH, Scale-Up Project Director, Sexual and 10 of 11 | JNCI Cancer Spectrum, 2019, Vol. 2, No. 4 Reproductive Health Global Program (SDS); Department of Francesc Xavier Bosch: received institutional research fund- Pathology, Hospital General de l’Hospitalet, Av. Josep Molins, ing from Merck and Seegene, occasional speaker fees from 29, 08906 L’Hospitalet de Llobregat, Barcelona, Spain (MA); Merck, GSK, and Roche, and occasional travel grants from Merck Department of Pathology, Hospital del Mar, Passeig Marıtim, and GSK. He also has received scientific advisory board fees, 25–29, 08003 Barcelona, Spain (BL); Department of Pathology, speakers fees, or travel grants from GSK, Merck, Inovio, IMS Hospital Universitari de Bellvitge, Carrer de la Feixa Llarga, Health, Abbott Laboratoires SA, Hologic and Roche; and unre- s/n, 08907 L’Hospitalet de Llobregat, Barcelona, Spain (AV); stricted institutional research grants from Merck, GSK and Seegene. Department of Dermatology, Hospital Universitari Vall  noma de Barcelona, Passeig de la Laia Alemany: received institutional research funding from d’Hebron, Universitat Auto Merck, GSK and Seegene. Vall d’Hebron, 119–129, 08035 Barcelona, Spain (CFP); All other authors declare no potential conflicts of interest. Division of Molecular Diagnostics of Oncogenic Infections, Research Program Infection, Inflammation and Cancer, German Cancer Research Center (DKFZ), ImNeuenheimer References Feld 280, 69120 Heidelberg, Germany (DH, GH, MP); 1. Ervik M, Lam F, Ferlay J, Mery L, Soerjomataram I, Bray F. Cancer Today. Lyon, Obstetrics and Gynecology Department, David Geffen School France: International Agency for Research on Cancer; 2016. http://gco.iarc.fr/ of Medicine at UCLA, Los Angeles, CA 90095-7363, USA (GH); today. Accessed August 17, 2017. 2. Plummer M, de Martel C, Vignat J, Ferlay J, Bray F, Franceschi S. Global burden Infections and Cancer Biology Group, IARC, WHO, 150 Cours of cancers attributable to infections in 2012: a synthetic analysis. Lancet Glob Albert Thomas, 69008 Lyon, France (MT); DDL Diagnostic Health. 2016;4(9):e609–e616. 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Combining routine morphology, Authors and Contributions p16(INK4a) immunohistochemistry, and in situ hybridization for the detection of human papillomavirus infection in penile carcinomas: a tissue microarray All authors made substantial contribution to the conception study using classifier performance analyses. Urol Oncol. 2014;32(2):171–177. and design of the work. Beatriz Serrano, Laia Alemany, and Sara 10. Mills AM, Dirks DC, Poulter MD, Mills SE, Stoler MH. HR-HPV E6/E7 mRNA in situ hybridization: validation against PCR, DNA in situ hybridization, and Tous were responsible for the data analysis and preparations of p16 immunohistochemistry in 102 samples of cervical, vulvar, anal, and the tables and figures. All authors contributed to the writing of head and neck neoplasia. Am J Surg Pathol. 2017;41(5):607–615. the article and Beatriz Serrano, Laia Alemany, and Silvia de 11. Venuti A, Paolini F. HPV detection methods in head and neck cancer. Head  were responsible for the preparation of the article for Neck Pathol. 2012;6(S1):63–74. Sanjose  Boily MC, et al. Population-level impact and herd effects 12. Drolet M, Be nard E, submission. Laia Alemany had full access to all the data in the following human papillomavirus vaccination programmes: a systematic re- study and takes responsibility for the integrity of the data and view and meta-analysis. Lancet Infect Dis. 2015;15(5):565–580. 13. Garland SM, Kjaer SK, Mun ~ oz N, et al. Impact and effectiveness of the quadri- the accuracy of the data analysis, and had the final responsibil- valent human papillomavirus vaccine: a systematic review of ten years of ity to submit the article for publication. All authors read and ap- real-world experience. Clin Infect Dis. 2016;63(4):519–527. proved the final article. No financial support for the project from 14. World Health Organization. Human Papillomavirus Vaccines WHO Position commercial partners was provided to Massimo Tommasino. Paper. Geneva, Switzerland: WHO; 2017. http://apps.who.int/iris/bitstream/ 10665/255353/1/WER9219.pdf?. Accessed August 17, 2017. 15. Bruni L, Diaz M, Barrionuevo-Rosas L, et al. Global estimates of human papil- lomavirus vaccination coverage by region and income level: a pooled analy- Conflicts of Interest sis. Lancet Glob Health. 2016;4(7):e453–e463. 16. de Sanjose S, Quint WG, Alemany L, et al. Human papillomavirus genotype Silvia de Sanjose : received institutional research funding from attribution in invasive cervical cancer: a retrospective cross-sectional world- Merck, GSK and Seegene. wide study. Lancet Oncol. 2010;11(11):1048–1056. 17. de Sanjose  S, Alemany L, Ordi J, et al. Worldwide human papillomavirus ge- Beatriz Serrano: received Institutional research funding from notype attribution in over 2000 cases of intraepithelial and invasive lesions Merck, GSK and Seegene. of the vulva. Eur J Cancer. 2013;49(16):3450–3461. Sara Tous: received institutional research funding from 18. Alemany L, Saunier M, Tinoco L, et al. Large contribution of human papillo- mavirus in vaginal neoplastic lesions: a worldwide study in 597 samples. Eur Merck, GSK and Seegene. J Cancer. 2014;50(16):2846–2854. Beatriz Quiro s: received institutional research funding from 19. Alemany L, Saunier M, Alvarado-Cabrero I, et al. Human papillomavirus DNA Merck, GSK and Seegene. prevalence and type distribution in anal carcinomas worldwide. Int J Cancer. 2015;136(1):98–107. Omar Clavero: received institutional research funding from 20. Alemany L, Cubilla A, Halec G, et al. Role of human papillomavirus in penile Merck, GSK and Seegene. carcinomas worldwide. Eur Urol. 2016;70(6):1078–1079.  Miquel Angel Pavo n: received institutional research funding 21. Geraets DT, Struijk L, Kleter B, et al. The original SPF10 LiPA25 algorithm is more sensitive and suitable for epidemiologic HPV research than the SPF10 from Merck, GSK and Seegene. INNO-LiPA Extra. J Virol Methods. 2015;215–216:22–29. Michael Pawlita: received institutional research funding 22. Insinga RP, Liaw K-L, Johnson LG, Madeleine MM. A systematic review of the from Roche and Qiagen. prevalence and attribution of human papillomavirus types among cervical,  et al. | 11 of 11 S. de Sanjose vaginal, and vulvar precancers and cancers in the United States. Cancer intraepithelial neoplasia of the vulva, vagina and anus: a meta-analysis. Int J Epidemiol Biomarkers Prev. 2008;17(7):1611–1622. Cancer. 2009;124(7):1626–1636. 23. Wentzensen N, Schiffman M, Dunn T, et al. Multiple human papillomavirus 27. Jordan RC, Lingen MW, Perez-Ordonez B, et al. Validation of methods for oro- genotype infections in cervical cancer progression in the study to understand pharyngeal cancer HPV status determination in US cooperative group trials. cervical cancer early endpoints and determinants. Int J Cancer. 2009;125(9): Am J Surg Pathol. 2012;36(7):945–954. 2151–2158. 28. 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