Carolina Porras

Reduced Prevalence of Oral Human Papillomavirus (HPV) 4 Years after Bivalent HPV Vaccination in a Randomized Clinical Trial in Costa Rica

Background Human papillomavirus (HPV) infection, particularly with type 16, causes a growing fraction of oropharyngeal cancers, whose incidence is increasing, mainly in developed countries. In a double-blind controlled trial conducted to investigate vaccine efficacy (VE) of the bivalent HPV 16/18 vaccine against cervical infections and lesions, we estimated VE against prevalent oral HPV infections 4 years after vaccination. Methods and Findings A total of 7,466 women 18–25 years old were randomized (1∶1) to receive the HPV16/18 vaccine or hepatitis A vaccine as control. At the final blinded 4-year study visit, 5,840 participants provided oral specimens (91·9% of eligible women) to evaluate VE against oral infections. Our primary analysis evaluated prevalent oral HPV infection among all vaccinated women with oral and cervical HPV results. Corresponding VE against prevalent cervical HPV16/18 infection was calculated for comparison. Oral prevalence of identifiable mucosal HPV was relatively low (1·7%). Approximately four years after vaccination, there were 15 prevalent HPV16/18 infections in the control group and one in the vaccine group, for an estimated VE of 93·3% (95% CI = 63% to 100%). Corresponding efficacy against prevalent cervical HPV16/18 infection for the same cohort at the same visit was 72·0% (95% CI = 63% to 79%) (p versus oral VE = 0·04). There was no statistically significant protection against other oral HPV infections, though power was limited for these analyses. Conclusions HPV prevalence four years after vaccination with the ASO4-adjuvanted HPV16/18 vaccine was much lower among women in the vaccine arm compared to the control arm, suggesting that the vaccine affords strong protection against oral HPV16/18 infection, with potentially important implications for prevention of increasingly common HPV-associated oropharyngeal cancer. ClinicalTrials.gov, Registry number NCT00128661

Durable Antibody Responses Following One Dose of the Bivalent Human Papillomavirus L1 Virus-Like Particle Vaccine in the Costa Rica Vaccine Trial

The Costa Rica HPV16/18 Vaccine Trial (CVT) showed that four-year vaccine efficacy against 12-month HPV16/18 persistent infection was similarly high among women who received one, two, or the recommended three doses of the bivalent HPV16/18 L1 virus-like particle (VLP) vaccine. Live-attenuated viral vaccines, but not simple-subunit vaccines, usually induce durable lifelong antibody responses after a single dose. It is unclear whether noninfectious VLP vaccines behave more like live-virus or simple-subunit vaccines in this regard. To explore the likelihood that efficacy will persist longer term, we investigated the magnitude and durability of antibodies to this vaccine by measuring HPV16- and HPV18-specific antibodies by VLP-ELISA using serum from enrollment, vaccination, and annual visits through four years in four vaccinated groups; one-dose (n = 78), two-doses separated by one month (n = 140), two doses separated by six months (n = 52), and three scheduled doses (n = 120, randomly selected). We also tested enrollment sera from n = 113 HPV16- or HPV18 L1-seropositive women prevaccination, presumably from natural infection. At four years, 100% of women in all groups remained HPV16/18 seropositive; both HPV16/18 geometric mean titers (GMT) among the extended two-dose group were non-inferior to the three-dose group, and ELISA titers were highly correlated with neutralization titers in all groups. Compared with the natural infection group, HPV16/18 GMTs were, respectively, at least 24 and 14 times higher among the two-dose and 9 and 5 times higher among one-dose vaccinees. Antibody levels following one-dose remained stable from month 6 through month 48. Results raise the possibility that even a single dose of HPV VLPs will induce long-term protection. Cancer Prev Res; 6(11); 1242–50. ©2013 AACR.

Human papillomavirus infection with multiple types: pattern of coinfection and risk of cervical disease.

OBJECTIVE We investigated coinfection patterns for 25 human papillomavirus (HPV) types and assessed the risk conferred by multiple HPV types toward cervical disease. METHODS Sexually active women (n=5,871) in the NCI-sponsored Costa Rica HPV Vaccine Trials prevaccination enrollment visit were analyzed. Genotyping for 25 HPVs was performed using SPF(10)/LiPA(25). We calculated odds ratios (ORs) to assess coinfection patterns for each genotype with 24 other genotypes. These ORs were pooled and compared with pair-specific ORs to identify genotype combinations that deviated from the pooled OR. We compared risk of CIN2+/HSIL+between multiple and single infections and assessed additive statistical interactions. RESULTS Of the 2478 HPV-positive women, 1070 (43.2%) were infected with multiple types. Multiple infections occurred significantly more frequently than predicted by chance. However, this affinity to be involved in a coinfection (pooled OR for 300 type-type combinations=2.2; 95% confidence interval [CI]=2.1-2.4) was not different across HPV type-type combinations. Compared with single infections, coinfection with multiple α9 species was associated with significantly increased risk of CIN2+(OR=2.2; 95% CI=1.1-4.6) and HSIL+(OR=1.6; 95% CI=1.1-2.4). However, disease risk was similar to the sum of estimated risk from individual types, with little evidence for synergistic interactions. CONCLUSIONS Coinfecting HPV genotypes occur at random and lead to cervical disease independently.

Rationale and design of a community-based double-blind randomized clinical trial of an HPV 16 and 18 vaccine in Guanacaste, Costa Rica.

We report the rationale, design, methods and details of participation of a community-based, double-blind, randomized clinical trial of an HPV 16 and 18 vaccine conducted in two provinces of Costa Rica to investigate the efficacy and population impact of the vaccine in the prevention of cervical cancer precursors. More than 24,000 women between 18 and 25 years of age were invited to participate and pre-screened for eligibility, with recruitment of 7466 women (30% of those pre-screened, 59% of those eligible) who were randomized to receive 3 doses of the HPV vaccine or hepatitis A vaccine as control. A complex protocol of data and specimen collection was applied, including an interview, pelvic exam for sexually active women, blood for serology and cell-mediated immunity, cervical secretions for local immunity and cells for HPV, Chlamydia trachomatis and gonorrhea testing. Eighty percent of the women received three doses, 12.4% two doses and 7.4% one dose. At visits, compliance with data and specimen collection was close to 100%. Baseline characteristics and age-specific prevalence of HPV and cervical neoplasia are reported. Overall prevalence of HPV was high (50%), with 8.3% of women having HPV 16 and 3.2% HPV 18. LSIL was detected in 12.7% of women at baseline and HSIL in 1.9%. Prevalence of Chlamydia was 14.2%. There was very good agreement in HPV detection between clinician-collected and self- collected specimens (89.4% agreement for all types, kappa 0.59). Follow up will continue with yearly or more frequent examinations for at least 4 years for each participant.

HPV16/18 L1 VLP vaccine induces cross-neutralizing antibodies that may mediate cross-protection

Human papillomavirus (HPV) L1 VLP-based vaccines are protective against HPV vaccine-related types; however, the correlates of protection have not been defined. We observed that vaccination with Cervarix™ induced cross-neutralizing antibodies for HPV types for which evidence of vaccine efficacy has been demonstrated (HPV31/45) but not for other types (HPV52/58). In addition, HPV31/45 cross-neutralizing titers showed a significant increase with number of doses (HPV31, p<0.001; HPV45, p<0.001) and correlated with HPV16/18 neutralizing titers, respectively. These findings raise the possibility that cross-neutralizing antibodies are effectors of cross-protection observed for the HPV16/18 vaccine.

Prevention of Persistent Human Papillomavirus Infection by an HPV16/18 Vaccine: A Community-Based Randomized Clinical Trial in Guanacaste, Costa Rica

Target groups for human papillomavirus (HPV) vaccination are controversial. We evaluated vaccine efficacy (VE) against 1-year persistent infection, stratified by age and sexual behavior, among young women in Costa Rica. We randomized 7,466 healthy women 18 to 25 years of age to HPV16/18 or hepatitis A vaccine (follow-up, 50.4 months). According-to-protocol (ATP) cohorts included compliant HPV-negative women; intention-to-treat (ITT) included all randomized women. ATP VE was 90.9% (95% CI, 82.0-95.9) against HPV16/18 infections, 44.5% against HPV31/33/45 (95% CI, 17.5-63.1), and 12.4% (95% CI, -3.2 to 25.6) against any oncogenic infection. Overall ITT VE against HPV16/18 infections was 49.0%, but ATP and ITT VE almost reached 100% in year 4 of follow-up. ATP efficacy against HPV16/18 was similar by age, but ITT VE was greatest among youngest women (68.9% among those 18-19 years of age; 21.8% among those 24-25 years of age) and 79.8% among virgins. Among previously unexposed women, vaccination is highly efficacious against HPV16/18 and partially against HPV31/33/45. Vaccination is most effective in women and girls before they initiate sexual activity, with programmatic and individual decision implications.

Common variants in immune and DNA repair genes and risk for human papillomavirus persistence and progression to cervical cancer.

BACKGROUND We examined host genetic factors to identify those more common in individuals whose human papillomavirus (HPV) infections were most likely to persist and progress to cervical intraepithelial neoplasia grade 3 (CIN3) and cancer. METHODS We genotyped 92 single-nucleotide polymorphisms (SNPs) from 49 candidate immune response and DNA repair genes obtained from 469 women with CIN3 or cancer, 390 women with persistent HPV infections (median duration, 25 months), and 452 random control subjects from the 10,049-woman Guanacaste Costa Rica Natural History Study. We calculated odds ratios and 95% confidence intervals (CIs) for the association of SNP and haplotypes in women with CIN3 or cancer and HPV persistence, compared with random control subjects. RESULTS A SNP in the Fanconi anemia complementation group A gene (FANCA) (G501S) was associated with increased risk of CIN3 or cancer. The AG and GG genotypes had a 1.3-fold (95% CI, 0.95-1.8-fold) and 1.7-fold (95% CI, 1.1-2.6-fold) increased risk for CIN3 or cancer, respectively (P(trend) = .008; referent, AA). The FANCA haplotype that included G501S also conferred increased risk of CIN3 or cancer, as did a different haplotype that included 2 other FANCA SNPs (G809A and T266A). A SNP in the innate immune gene IRF3 (S427T) was associated with increased risk for HPV persistence (P(trend) = .009). CONCLUSIONS Our results require replication but support the role of FANCA variants in cervical cancer susceptibility and of IRF3 in HPV persistence.

Common Genetic Variants and Risk for HPV Persistence and Progression to Cervical Cancer

HPV infrequently persists and progresses to cervical cancer. We examined host genetic factors hypothesized to play a role in determining which subset of individuals infected with oncogenic human papillomavirus (HPV) have persistent infection and further develop cervical pre-cancer/cancer compared to the majority of infected individuals who will clear infection. We evaluated 7140 tag single nucleotide polymorphisms (SNPs) from 305 candidate genes hypothesized to be involved in DNA repair, viral infection and cell entry in 416 cervical intraepithelial neoplasia 3 (CIN3)/cancer cases, 356 HPV persistent women (median: 25 months), and 425 random controls (RC) from the 10,049 women Guanacaste Costa Rica Natural History study. We used logistic regression to compute odds ratios and p-trend for CIN3/cancer and HPV persistence in relation to SNP genotypes and haplotypes (adjusted for age). We obtained pathway and gene-level summary of associations by computing the adaptive combination of p-values. Genes/regions statistically significantly associated with CIN3/cancer included the viral infection and cell entry genes 2′,5′ oligoadenylate synthetase gene 3 (OAS3), sulfatase 1 (SULF1), and interferon gamma (IFNG); the DNA repair genes deoxyuridine triphosphate (DUT), dosage suppressor of mck 1 homolog (DMC1), and general transcription factor IIH, polypeptide 3 (GTF2H4); and the EVER1 and EVER2 genes (p<0.01). From each region, the single most significant SNPs associated with CIN3/cancer were OAS3 rs12302655, SULF1 rs4737999, IFNG rs11177074, DUT rs3784621, DMC1 rs5757133, GTF2H4 rs2894054, EVER1/EVER2 rs9893818 (p-trends≤0.001). SNPs for OAS3, SULF1, DUT, and GTF2H4 were associated with HPV persistence whereas IFNG and EVER1/EVER2 SNPs were associated with progression to CIN3/cancer. We note that the associations observed were less than two-fold. We identified variations DNA repair and viral binding and cell entry genes associated with CIN3/cancer. Our results require replication but suggest that different genes may be responsible for modulating risk in the two critical transition steps important for cervical carcinogenesis: HPV persistence and disease progression.

Comparison of the SPF10-LiPA System to the Hybrid Capture 2 Assay for Detection of Carcinogenic Human Papillomavirus Genotypes among 5,683 Young Women in Guanacaste, Costa Rica

ABSTRACT The objective of this analysis was to compare the performance characteristics of two human papillomavirus (HPV) DNA detections assays, the Hybrid Capture 2 assay (HC2) and the SPF10 assay, for the detection of carcinogenic HPV. Data are from the enrollment visits of women who participated in the randomized, double-blind, placebo-controlled phase III HPV16/18 Vaccine Trial in Guanacaste, Costa Rica. We compared the results of HC2 and SPF10 testing of cervical specimens. Since the line probe assay (LiPA) detection system does not distinguish between HPV type 68 (HPV68; which is targeted by HC2) and HPV73 (which is not targeted by HC2), for SPF10-LiPA, we defined the carcinogenic HPV types as the 12 HC2-targeted types (types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, and 59), HPV68/73, and the HC2-cross-reactive, carcinogenic type HPV66. The kappa values and the performance characteristics for the detection of cervical abnormalities were ascertained. Paired observations were available for 5,683 sexually active, young women (median age, 21 years). The prevalence of carcinogenic HPV types was 35% (n = 1,962) by HC2 and 35% (n = 2,003) by SPF10-LiPA. There were no differences in the prevalence of carcinogenic HPV types by HC2 and SPF10-LiPA among women with normal, atypical squamous cells of undetermined significance and high-grade squamous intraepithelial lesion cytology. Among women with low-grade squamous intraepithelial lesion cytology, HC2 was more likely to test positive than SPF10-LiPA for the carcinogenic HPV types (87% and 79%, respectively; P = 0.001) as a result of HC2 cross-reactivity with HPV types 40, 43, 44, 53, 54, 60, 70, and 74. The crude agreement between the two assays was 88%, with a kappa value of 0.75 (95% confidence limits, 0.73 to 0.76). We observed very good agreement between HC2 and SPF10-LiPA for carcinogenic HPV type detection.

Evaluation of systemic and mucosal anti-HPV16 and anti-HPV18 antibody responses from vaccinated women

Ideal methods to monitor HPV neutralizing antibodies induced by vaccination have not been established yet. Here, we evaluated systemic and cervical antibody levels induced by HPV16/18 AS04-adjuvanted vaccine (GlaxoSmithKline Biologicals) using a secreted alkaline phosphatase neutralization assay (SEAP-NA) and enzyme-linked immunosorbent assay (ELISA). Serum and cervical secretions from 50 vaccinated women were used to assess (1) overall assay reproducibility; (2) inter-assay and inter-specimen correlation; (3) correlations between month 1 and month 12 titers. Strong correlations between SEAP-NA and ELISA were observed (serum anti-HPV16/18, rho=0.91/0.85; cervix anti-HPV16/18, rho=0.84/0.89). Systemic and cervical antibody measures also correlated well (rho range: 0.64-0.75); except at mid-cycle (rho range: 0.28-0.65). Correlations between antibody levels at 1 and 12 months following the start of vaccination were poor (rho range: 0.16-0.38). In conclusion, HPV16/18 VLP-based ELISA is a reliable and valid method to monitor anti-HPV16/18 neutralizing potential for the first year following vaccination; however, additional studies will be required to better define the effects of the time on cycle and patterns of antibody response over time following vaccination.