Circulation of human papillomavirus (HPV) genotypes in women from Córdoba, Argentina, with squamous intraepithelial lesions

Human papillomavirus (HPV) can induce a wide spectrum of squamous intraepithelial lesions (SIL) of varying severity. The aim of the present study was to establish the frequency of HPV infection and identify the genotypes circulating in women from Córdoba, Argentina, in relation to age and cytology. A total of 186 women, between 18 and 65 years old, with antecedents of SIL, underwent a pelvic examination and had cervical cells collected for cytology and HPV DNA detection. Ninety-six samples (51.6%) were positive for HPV detection, and sixty-three (65.6%) of them showed the presence of at least one HR-HPV. Low- and high-grade SIL showed significant association in patients younger than 35 years of age. We found 18 different genotypes, with a greater presence of HR-HPV. Genotypes 16 and 6 were the most frequent. Seven (7.3%) multiple infections, 85.7% of which had at least one HR-HPV, were detected. The detection of a large number of different HPV genotypes is a warning sign. It is thus necessary to strengthen the monitoring of the circulation of high-risk genotypes, currently less prevalent in intraepithelial lesions, as a control measure for the possible impact of the implementation of vaccines against genotypes 16 and 18.

MMP-9/RECK imbalance: a mechanism associated with high-grade cervical lesions and genital infection by Human Papillomavirus (HPV) and Chlamydia trachomatis

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Human papillomavirus (HPV) screening and cervical cancer burden. A Brazilian perspective

This review tackles the issues related to disease burden caused by cervical cancer (CC) and its precursor (CIN) lesions in Brazil. A special focus is given to new technologies with potential to interfere with the development of CC by reducing the high-risk human papillomavirus (hr-HPV)-induced lesions that remain a major public health burden in all developing countries where organized screening programs do not exist. Globally, 85 % of all incident CC and 50 % of CC deaths occur in the developing countries. Unfortunately, most regions of Brazil still demonstrate high mortality rates, ranking CC as the second most common cancer among Brazilian women. Recently, CC screening programs have been tailored in the country to enable early detection of CC precursor lesions and thereby reduce cancer mortality. A combination of HPV testing with liquid-based cytology (LBC) seems to be a promising new approach in CC screening, with high expectation to offer an adequate control of CC burden in this country.

Human Papillomavirus (HPV) Type Distribution in Females with Abnormal Cervical Cytology. A Correlation with Histological Study

The aim of this study was to determine human papillomavirus (HPV) types distribution in cervical preneoplasic lesions in a Southern Spanish population and their relationship between HPV type and grade of histopathological abnormality. Finally, 232 cervical samples from 135 women with previous cytological abnormalities were included in this study. Colposcopy studies and biopsies were performed. Haematoxylin-eosin stained slides were observed and detection of HPV DNA in cervical swabs was carried out with use of a polymerase chain reaction and microarrays technology. The relationship between the presence of HPV infection and diagnostic variables was evaluated. HPV 16 was the most common type followed by HPV 58, 51, 33 and 31. However, the two HPV types targeted in the prophylactic vaccines such as HPV type 16 and 18 were detected in only 37 (21.2%) and 2 (1.1%) cases respectively. Thirty-three (18.9%) of samples were infected with multiple types, the majority of them with two types. In addition, during the follow-up of patients many changes in type distribution were observed. Several studies will be necessary in order to evaluate the HPV type distribution for therapeutically and prophylactic purposes such as vaccine treatment. Also, because of the differences obtained depending of use of various DNA technologies, the performance of some comparative studies of the different methods from detection of HPV would be advisable in a high population of patients and with the most homogeneous conditions possible.

Human papillomavirus (HPV) vaccines as an option for preventing cervical malignancies: (how) effective and safe?

We carried out a systematic review of HPV vaccine pre- and post-licensure trials to assess the evidence of their effectiveness and safety. We find that HPV vaccine clinical trials design, and data interpretation of both efficacy and safety outcomes, were largely inadequate. Additionally, we note evidence of selective reporting of results from clinical trials (i.e., exclusion of vaccine efficacy figures related to study subgroups in which efficacy might be lower or even negative from peer-reviewed publications). Given this, the widespread optimism regarding HPV vaccines long-term benefits appears to rest on a number of unproven assumptions (or such which are at odd with factual evidence) and significant misinterpretation of available data. For example, the claim that HPV vaccination will result in approximately 70% reduction of cervical cancers is made despite the fact that the clinical trials data have not demonstrated to date that the vaccines have actually prevented a single case of cervical cancer (let alone cervical cancer death), nor that the current overly optimistic surrogate marker-based extrapolations are justified. Likewise, the notion that HPV vaccines have an impressive safety profile is only supported by highly flawed design of safety trials and is contrary to accumulating evidence from vaccine safety surveillance databases and case reports which continue to link HPV vaccination to serious adverse outcomes (including death and permanent disabilities). We thus conclude that further reduction of cervical cancers might be best achieved by optimizing cervical screening (which carries no such risks) and targeting other factors of the disease rather than by the reliance on vaccines with questionable efficacy and safety profiles.

Human papillomavirus (HPV) in mothers before and after delivery – a three year follow-up study.

Human papillomavirus (HPV) infections in mothers are important to consider since pregnancy may affect the outcome of the infection and the mother may transmit HPV to the child. This thesis is part of the 3-year Finnish Family HPV Study on HPV infection dynamics within 329 families. The presence of maternal HPV antibodies and HPV DNA in placenta, umbilical cord blood and breast milk was examined. In addition, genital and oral HR-HPV carriage was studied among mothers with one or two pregnancies. At enrollment, seropositivity to HPV 6, 11, 16, 18 and 45 was recorded in 53 %, 21 %, 35 %, 21 % and 9 % of the mothers, respectively. Age at sexual debut, number of sexual partners, a history of genital warts and antibodies to LR/HR-HPV predicted HR/LR-HPV-seropositivity. During follow-up 27 %, 14 %, 17 %, 17 % and 7 % of the mothers seroconverted to the tested HPV-types, respectively. Decay of HPV-antibodies was rare. The mother’s new pregnancy was of minor impact in the outcome of oral and cervical HR-HPV infections. HPV-DNA was present in 4.2 % and 3.5 % of the placentas and umbilical cord blood samples, and in 4.5 % and 19.7 % of the breast milk samples collected at day 3 and month 2 postpartum, respectively. HPV-positivity in placenta/cord blood was related to a history of abnormal pap-smears or genital warts, and raised the risk of the neonate being HPV-positive at birth. The mode of delivery did not predict the HPVstatus of neonate, placenta, or cord blood. HPV DNA in breast milk was associated with oral HPV status of the father, but not with HPV status of the neonate. The results indicate that exposure to HPV is common and that part of the exposure might take place already early in life. Contrary to the common claim, pregnancy is not a risk factor for HPV.

Outcomes of cervical human papillomavirus (HPV) infections among mothers in the Finnish Family HPV Study

To understand the natural history of cervical human papillomavirus (HPV)-infections, more information is needed on their genotype-specific prevalence, acquisition, clearance, persistence and progression. This thesis is part of the prospective Finnish Family HPV study. 329 pregnant women (mean age 25.5 years) were recruited during the third trimester of pregnancy and were followed up for 6 years. The outcomes of cervical HPV infections were evaluated among all the mothers participating in the study. Generalized estimating equation (GEE)-models and Poisson regression were used to estimate the risk factors of type-specific acquisition, clearance, persistence and progression of Species 7 and 9 HPV-genotypes. Independent protective factors against incident infections were higher number of life-time sexual partners, initiation of oral contraceptive use after age 20 years and becoming pregnant during FU. Older age and negative oral HR-HPV DNA status at baseline were associated with increased clearance, whereas higher number of current sexual partners decreased the probability of clearance. Early onset of smoking, practicing oral sex and older age increased the risk of type-specific persistence, while key predictors of CIN/SIL were persistent HR-HPV, abnormal Pap smear and new sexual partners. HPV16, together with multiple-type infections were the most frequent incident genotypes, most likely to remain persistent and least likely to clear. Collectively, LR-HPV types showed shorter incidence and clearance times than HR-HPV types. In multivariate models, different predictors were associated with these main viral outcomes, and there is some tentative evidence to suggest that oral mucosa might play a role in controlling some of these outcomes.

Effect of the human papillomavirus (HPV) quadrivalent vaccine in a subgroup of women with cervical and vulvar disease: Retrospective pooled analysis of trial data

Objectives To determine the effect of human papillomavirus (HPV) quadrivalent vaccine on the risk of developing subsequent disease after an excisional procedure for cervical intraepithelial neoplasia or diagnosis of genital warts, vulvar intraepithelial neoplasia, or vaginal intraepithelial neoplasia. Design Retrospective analysis of data from two international, double blind, placebo controlled, randomised efficacy trials of quadrivalent HPV vaccine (protocol 013 (FUTURE I) and protocol 015 (FUTURE II)). Setting Primary care centres and university or hospital associated health centres in 24 countries and territories around the world. Participants Among 17 622 women aged 15–26 years who underwent 1:1 randomisation to vaccine or placebo, 2054 received cervical surgery or were diagnosed with genital warts, vulvar intraepithelial neoplasia, or vaginal intraepithelial neoplasia. Intervention Three doses of quadrivalent HPV vaccine or placebo at day 1, month 2, and month 6. Main outcome measures Incidence of HPV related disease from 60 days after treatment or diagnosis, expressed as the number of women with an end point per 100 person years at risk. Results A total of 587 vaccine and 763 placebo recipients underwent cervical surgery. The incidence of any subsequent HPV related disease was 6.6 and 12.2 in vaccine and placebo recipients respectively (46.2% reduction (95% confidence interval 22.5% to 63.2%) with vaccination). Vaccination was associated with a significant reduction in risk of any subsequent high grade disease of the cervix by 64.9% (20.1% to 86.3%). A total of 229 vaccine recipients and 475 placebo recipients were diagnosed with genital warts, vulvar intraepithelial neoplasia, or vaginal intraepithelial neoplasia, and the incidence of any subsequent HPV related disease was 20.1 and 31.0 in vaccine and placebo recipients respectively (35.2% reduction (13.8% to 51.8%)). Conclusions Previous vaccination with quadrivalent HPV vaccine among women who had surgical treatment for HPV related disease significantly reduced the incidence of subsequent HPV related disease, including high grade disease.

Human papillomavirus (HPV) detection in lip squamous cell carcinoma: correlation with clinical aspects and risk factors

O papilomavírus humano (HPV) está associado a um largo espectro de lesões em humanos e tem sido ligado à carcinogênese oral. O objetivo deste estudo foi investigar a presença do DNA do HPV em pacientes com carcinoma espinocelular de lábio e correlacioná-la com aspectos clínicos e fatores de risco. Foram estudados 33 pacientes com carcinoma espinocelular de lábio. Destes, 30 pacientes foram positivos para o gene da beta-globina humana e então foram testados para o DNA do HPV com uso da reação em cadeia de polimerase em duas etapas (PCR e nPCR) com os oligonucleotídeos iniciadores MY11/MY09 e GP5+/ GP6+. O DNA do HPV foi detectado em 43,33% dos 30 pacientes analisados. Não houve associação com os fatores de risco analisados.

Longitudinal outcomes of high-risk human papillomavirus (HPV) infections as competing-risks events following cervical HPV test at baseline visit in the *NIS-LAMS** cohort

Background: The complex natural history of human papillomavirus (HPV) infections following a single HPV test can be modeled as competing-risks events (i.e., no-, transient- or persistent infection) in a longitudinal setting. The covariates associated with these compet ng events have not been previously assessed using competing-risks regression models. Objectives: To gain further insights in the outcomes of cervical HPV infections, we used univariate- and multivariate competing-risks regression models to assess the covariaies associated with these competing events. Study Design and Methods: Covariates associated with three competing outcomes (no-, transient- or persistent HR-HPV infection) were analysed in a sub-cohort of 1,865 women prospectively followed-up in the NIS (n = 3,187) and LAMS Study (n = 12,114). Results: In multivariate competing-risks models (with two other outcomes as competing events), permanently HR-HPV negative outcome was significantly predicted only by the clearance of ASCUS+Pap during FU, while three independent covariates predicted transient HR-HPV infections: i) number of recent (< 12 months) sexual partners (risk increased), ii) previous Pap screening history (protective), and history of previous CIN (increased risk). The two most powerful predictors of persistent HR-HPV infections were persistent ASCUS+Pap (risk increased), and previous Pap screening history (protective). In pair-wise comparisons, number of recent sexual partners and previous CIN history increase the probability of transient HR-HPV infection against the HR-HPV negative competing event, while previous Pap screening history is protective. Persistent ASCUS+Pap during FU and no previous Pap screening history are significantly associated with the persistent HR-HPV outcome (compared both with i) always negative, and ii) transient events), whereas multiparity is protective. Conclusions: Different covariates are associated with the three main outcomes of cervical HPV infections. The most significant covariates of each competing events are probably distinct enough to enable constructing of a risk-profile for each main outcome.

Covariates of high-risk human papillomavirus (HPV) infections are distinct for incident CIN1, CIN2 and CIN3 as disclosed by competing-risks regression models

Background: In addition to the oncogenic human papillomavirus (HPV), several cofactors are needed in cervical carcinogenesis, but whether the HPV covariates associated with incident i) CIN1 are different from those of incident ii) CIN2 and iii) CIN3 needs further assessment. Objectives: To gain further insights into the true biological differences between CIN1, CIN2 and CIN3, we assessed HPV covariates associated with incident CIN1, CIN2, and CIN3. Study Design and Methods: HPV covariates associated with progression to CIN1, CIN2 and CIN3 were analysed in the combined cohort of the NIS (n = 3,187) and LAMS study (n = 12,114), using competing-risks regression models (in panel data) for baseline HR-HPV-positive women (n = 1,105), who represent a sub-cohort of all 1,865 women prospectively followed-up in these two studies. Results: Altogether, 90 (4.8%), 39 (2.1%) and 14 (1.4%) cases progressed to CIN1, CIN2, and CIN3, respectively. Among these baseline HR-HPV-positive women, the risk profiles of incident GIN I, CIN2 and CIN3 were unique in that completely different HPV covariates were associated with progression to CIN1, CIN2 and CIN3, irrespective which categories (non-progression, CIN1, CIN2, CIN3 or all) were used as competing-risks events in univariate and multivariate models. Conclusions: These data confirm our previous analysis based on multinomial regression models implicating that distinct covariates of HR-HPV are associated with progression to CIN1, CIN2 and CIN3. This emphasises true biological differences between the three grades of GIN, which revisits the concept of combining CIN2 with CIN3 or with CIN1 in histological classification or used as a common end-point, e.g., in HPV vaccine trials.

Expression of Mitochondrial Non-coding RNAs (ncRNAs) Is Modulated by High Risk Human Papillomavirus (HPV) Oncogenes

The study of RNA and DNA oncogenic viruses has proved invaluable in the discovery of key cellular pathways that are rendered dysfunctional during cancer progression. An example is high risk human papillomavirus (HPV), the etiological agent of cervical cancer. The role of HPV oncogenes in cellular immortalization and transformation has been extensively investigated. We reported the differential expression of a family of human mitochondrial non-coding RNAs (ncRNAs) between normal and cancer cells. Normal cells express a sense mitochondrial ncRNA (SncmtRNA) that seems to be required for cell proliferation and two antisense transcripts (ASncmtRNAs). In contrast, the ASncmtRNAs are down-regulated in cancer cells. To shed some light on the mechanisms that trigger down-regulation of the ASncmtRNAs, we studied human keratinocytes (HFK) immortalized with HPV. Here we show that immortalization of HFK with HPV-16 or 18 causes down-regulation of the ASncmtRNAs and induces the expression of a new sense transcript named SncmtRNA-2. Transduction of HFK with both E6 and E7 is sufficient to induce expression of SncmtRNA-2. Moreover, E2 oncogene is involved in down-regulation of the ASncmtRNAs. Knockdown of E2 in immortalized cells reestablishes in a reversible manner the expression of the ASncmtRNAs, suggesting that endogenous cellular factors(s) could play functions analogous to E2 during non-HPV-induced oncogenesis.

Expression of BAG-1 and PARP-1 in Precursor Lesions and Invasive Cervical Cancer Associated with Human Papillomavirus (HPV)

Cervical cancer remains persistently the second most common malignancies among women worldwide, responsible for 500,000 new cases annually. Only in Brazil, the estimate is for 18,430 new cases in 2011. Several types of molecular markers have been studied in carcinogenesis including proteins associated with apoptosis such as BAG-1 and PARP-1. This study aims to demonstrate the expression of BAG-1 and PARP-1 in patients with low-grade squamous intraepithelial lesions (LSILs), high-grade squamous intraepithelial lesions (HSILs) and invasive squamous cell carcinomas (SCCs) of the uterine cervix and to verify a possible association with HPV infection. Fifty samples of LSILs, 50 samples of HSILs and 50 samples of invasive SCCs of the uterine cervix were analyzed by immunohistochemistry for BAG-1 and PARP-1 expression. PCR was performed to detect and type HPV DNA. BAG-1 expression levels were significantly different between LSILs and HSILs (p = 0,014) and between LSILs and SCCs (p = 0,014). In regards to PARP-1 expression, we found significant differences between the expression levels in HSILs and SCCs (p = 0,022). No association was found between BAG-1 expression and the presence of HPV. However, a significant association was found between PARP-1 expression and HPV positivity in the HSILs group (p = 0,021). In conclusion our research suggests that BAG-1 expression could contribute to the differentiation between LSIL and HSIL/SCC whereas PARP-1 could be useful to the differentiation between HSIL HPV-related and SCC. Further studies are needed to clarify the molecular aspects of the relationship between PARP-1 expression and HPV infection, with potential applications for cervical cancer prediction.

Human papillomavirus (HPV) and associated diseases: between applied diagnostic and basic research

Human Papillomavirus (HPV) is the cause of cervical cancers (among these, adenocarcinoma, AdCa) and is associated to a subgroup of oropharyngeal carcinomas (OPSCCs). Even if the risk for cancer development is linked to the infection by some viral genotypes, mainly HPV16 and 18, viral DNA alone seems not to be sufficient for diagnosis. Moreover, the role of the virus in OPSCCs has not been totally clarified yet. In the first part of the thesis, the performances concerning viral genotyping in clinical cervical samples of a new pyrosequencing-based test and a well-known hybridization-based assay have been compared. Similar results between the methods have been obtained. However, the former showed advantages in detecting intratype variants, higher specificity and a broader spectrum of detectable HPV types. The second part deals with the evaluation of virological markers (genotyping, viral oncoproteins expression, viral load, physical state and CpG methylation of HPV16 genome) in the diagnosis/prognosis of cervical AdCa and HPV-associated OPSCCs. HPV16 has been confirmed the most prevalent genotype in both the populations. Interestingly, the mean methylation frequency of viral DNA at the early promoter showed the tendency to be associated to invasion for cervical AdCa and to a worse prognosis for OPSCCs, suggesting a promising role as diagnostic/prognostic biomarker. The experiments of the third part were performed at the DKFZ in Heidelberg (Germany) and dealt with the analysis of the response to IFN-k transfection in HPV16-positive cervical cancer and head&neck carcinoma cell lines to evaluate its potential role as new treatment. After 24h, we observed increased IFN-b expression which lead to the up-regulation of genes involved in the antigens presentation pathway (MHC class I and immunoproteasome) and antiviral response as well, in particular in cervical cancer cell lines. This fact suggested also the presence of different HPV-mediated carcinogenic pathways between the two anatomical districts.