Anogenital Human Papillomavirus Research Articles

Penile Intraepithelial Neoplasia Is Frequent in HIV-Positive Men with Anal Dysplasia

Anogenital human papillomavirus (HPV)-infection is common in HIV-infected men who have sex with men (HIV+MSM). These patients have a strongly increased risk of HPV-induced anal cancer and its precursor lesion, anal intraepithelial neoplasia (AIN), and a moderately increased risk for penile cancer. Only limited data exist on penile intraepithelial neoplasia (PIN) in HIV+MSM. We determined the prevalence and evaluated the virologic characteristics of PIN and AIN in 263 HIV+MSM. In case of histologically confirmed PIN (and AIN), HPV-typing, HPV-DNA load determination, and immunohistochemical staining for p16(INK4a) were performed. PIN was detected in 11 (4.2%) and AIN in 156 (59.3%) patients. Ten PIN patients also had AIN within the observation period. Four clinical types of PINs could be distinguished. High-risk-alpha-HPV-DNA was found in 10 PIN lesions, with HPV16 being the most frequent type. Infections with multiple HPV-types were common. All high-grade lesions had high-risk-HPV-DNA-loads > or = 1 HPV-copy/beta-globin-gene-copy. Cutaneous beta-HPVs were found in PIN and AIN, but beta-HPV-DNA loads were very low, irrespective of the histological grade. p16(INK4a) Expression was detectable in all PIN lesions and correlated both with the histological grade and with high-risk HPV-DNA loads. In view of the PIN prevalence found in our study, all HIV+MSM should be screened for PIN in addition to AIN screening.

State-of-the-art of infections produced by human papillomavirus.

Anogenital human papillomavirus infection is the most frequent sexually transmitted disease. Around one hundred genotypes have been identified in humans, and 40 infect the genitalia and anal regions. Fifteen genotypes, classified as high-risk HPVs, are the necessary cause of cervical cancer and have been involved as carcinogenic agents for cancer of the vulva, vagina, anus, penis, and oropharyngeal cavity. Low-risk HPVs are the causative agents of genital warts, and recurrent respiratory papillomatosis in both men and women. The number of women harboring HPV-DNA worldwide is estimated to be 300 million. The recently introduced prophylactic HPV vaccines represent a hopeful strategy to prevent HPV infection and HPV-related diseases.

Impact of a Prophylactic Quadrivalent Human Papillomavirus (Types 6, 11, 16, 18) L1 Virus-Like Particle Vaccine in a Sexually Active Population of North American Women

Although most human papillomavirus (HPV) infections resolve without sequelae, some cause cervical, vulvar, vaginal, and anal cancers as well as precancerous lesions and genital warts. These lesions appear to be preventable by prophylactically administering a quadrivalent HPV-6/-11/-16/-18 vaccine to young women. This study estimated the clinical effects of vaccination in 5996 North American (US, Canada, Puerto Rico) women, most of whom were 16 to 25 years of age. Participants were culled from 5 clinical trials of nearly 22,000 women who received either quadrivalent HPV vaccine or its HPV-16 vaccine prototype. The influence of vaccination on the risk of anogenital HPV lesions was estimated in an intention-to-treat population. Women enrolled in the study had a median age of 20 years. A Papanicolaou test abnormality was present in 13% of participants. Approximately 3 of 4 women (76%) had negative test results for all 4 vaccine HPV types. After about 3 years of follow-up in the intention-to-treat population, regardless of HPV status at enrollment, vaccination lowered the rate of HPV-16- and HPV-18-related precancers by 66.4% (95% confidence interval [CI], 42.7–81.1%), and that of HPV-6/-11/-16/-18—related genital lesions by 57.7% (95% CI, 27.3%–76.3%). In general, the clinical benefit of vaccination was similar across the age range and across patterns of sexual behavior. Vaccination had a lesser effect in women who had a Papanicolaou test abnormality when vaccination began, many of whom already had HPV disease. The investigators believe that a catch-up vaccination program using the quadrivalent HPV vaccine is appropriate for North American women aged 16 to 25 years.

Impact of a prophylactic quadrivalent human papillomavirus (types 6, 11, 16, 18) L1 virus-like particle vaccine in a sexually active population of North American women

The purpose of this study was to inform policy regarding human papillomavirus (HPV) vaccination in North America. We measured the clinical impact of HPV-6/-11/-16/-18 vaccination in North American women. The study enrolled 21,954 women, the majority aged 16-25, across 5 studies of a quadrivalent HPV vaccine or its HPV-16 vaccine prototype. The North American subjects (n = 5996) were pooled from these trials, and the prevalence of HPV-6/-11/-16/-18 exposure was measured. The impact of vaccination on the burden of anogenital HPV lesions in an intention-to-treat population (regardless of enrollment HPV status) was calculated. At enrollment, the median age was 20 years; 13% of the women had had a Papanicolaou test abnormality, and 76% of the women had negative tests results for all 4 vaccine HPV types. With approximately 3 years of follow-up evaluations in the intention-to-treat population (regardless of enrollment HPV status), vaccination reduced the rate of HPV-16- and -18-related precancers and HPV-6/-11/-16/-18-related genital lesions by 66.4% (95% CI, 42.7%-81.1%) and 57.7% (95% CI, 27.3%-76.3%), respectively. The administration of HPV vaccine to sexually active North American women reduced the burden of HPV-6/-11/-16/-18-related disease. Catch-up vaccination programs in this population are warranted.

Human papillomavirus E1 helicase interacts with the WD repeat protein p80 to promote maintenance of the viral genome in keratinocytes.

Due to the limited coding capacity of their small genomes, human papillomaviruses (HPV) rely extensively on host factors for the completion of their life cycles. Accordingly, most HPV proteins, including the replicative helicase E1, engage in multiple protein interactions. The fact that conserved regions of E1 have not yet been ascribed a function prompted us to use tandem affinity protein purification (TAP) coupled to mass spectrometry to identify novel targets of this helicase. This method led to the discovery of a novel interaction between the N-terminal 40 amino acids of HPV type 11 (HPV11) E1 and the cellular WD repeat protein p80 (WDR48). We found that interaction with p80 is conserved among E1 proteins from anogenital HPV but not among cutaneous or animal types. Colocalization studies showed that E1 can redistribute p80 from the cytoplasm to the nucleus in a manner that is dependent on the E1 nuclear localization signal. Three amino acid substitutions in E1 proteins from HPV11 and -31 were identified that abrogate binding to p80 and its relocalization to the nucleus. In HPV31 E1, these substitutions reduced but did not completely abolish transient viral DNA replication. HPV31 genomes encoding two of the mutant E1 proteins were not maintained as episomes in immortalized primary keratinocytes, whereas one encoding the third mutant protein was maintained at a very low copy number. These findings suggest that the interaction of E1 with p80 is required for efficient maintenance of the viral episome in undifferentiated keratinocytes.

Risk Factors for Anogenital Human Papillomavirus Infection in Men

Human papillomavirus (HPV) is strongly associated with cervical and other anogenital cancers. Identification of risk factors for HPV infection in men may improve our understanding of HPV transmission and prevention. HPV testing for 37 types was conducted in 463 men 18-40 years old recruited from 2 US cities. The entire anogenital region and semen were sampled. A self-administered questionnaire was completed. Multivariate logistic regression aided the identification of independent risk factors for any HPV type, oncogenic HPV types, and nononcogenic HPV types. Prevalence was 65.4% for any HPV, 29.2% for oncogenic HPV, and 36.3% for nononcogenic HPV. Factors significantly associated with any HPV were smoking > or =10 cigarettes per day (odds ratio [OR], 2.3 [95% confidence interval {CI}, 1.0-5.3]) and lifetime number of female sex partners (FSPs) (OR for > or =21, 2.5 [95% CI, 1.3-4.6]), and factors significantly associated with oncogenic HPV were lifetime number of FSPs (OR for > or =21, 7.4 [95% CI, 3.4-16.3]) and condom use during the past 3 months (OR for more than half the time, 0.5 [95% CI, 0.3-0.8]). For nononcogenic HPV, a significant association was found for number of FSPs during the past 3 months (OR for > or =2, 2.9 [95% CI, 1.4-6.3]). Lifetime and recent number of FSPs, condom use, and smoking were modifiable risk factors associated with HPV infection in men.

Cutaneous Human Papillomaviruses Down-regulate AKT1, whereas AKT2 Up-regulation and Activation Associates with Tumors

Epithelial tumorigenesis has been linked to AKT up-regulation. Human papillomaviruses (HPV) cause anogenital cancers and anogenital HPV infection up-regulates AKT activity. Mounting evidence points to a role for cutaneous HPVs as etiologic factors in skin tumorigenesis. High-risk cutaneous beta HPVs have been linked to carcinogenesis in immunosuppressed patients, and high-risk cutaneous HPV8 genes enhance tumorigenesis in transgenic mice. We find that, in contrast to anogenital HPVs, cutaneous HPV8 early genes down-regulate epidermal AKT activity by down-regulating AKT1 isoform levels. This down-regulation occurs before papilloma formation or tumorigenesis and leads to cutaneous differentiation changes that may weaken the epidermal squame for viral release. We find that, in viral warts (papillomas) and HPV gene-induced epidermal tumors, AKT activity can be activated focally by up-regulation and phosphorylation of the AKT2 isoform. In squamous cell carcinomas (SCC), AKT1 down-regulation is also common, consistent with a viral influence, whereas AKT2 up-regulation is widespread. Activation of up-regulated AKT2 by serine phosphorylation associates with high-grade tumors. Our data suggest that AKT2 up-regulation is characteristic of SCC and that coincident AKT2 activation through serine phosphorylation correlates with malignancy. These findings highlight differences between the effects of anogenital and cutaneous HPV on epithelial AKT activity and furthermore show that AKT isoforms can behave differently during epidermal tumorigenesis. These findings also suggest AKT2 as a possible therapeutic tumor target in SCC.

Update: human papillomavirus infection remains highly prevalent and persistent among HIV-infected individuals

Human papillomavirus infections and human papillomavirus-associated anogenital tumors are more prevalent in HIV-infected than HIV-uninfected individuals. This review focuses on recent developments related to human papillomavirus burden in HIV-infected individuals; anogenital human papillomavirus types and type-specific differences in the natural history of human papillomavirus infections; the effect of highly active antiretroviral therapy on human papillomavirus infection; and novel human papillomavirus therapeutic interventions. There is a paucity of recent data on the effect of highly active antiretroviral therapy on human papillomavirus infection and its related anogenital abnormalities/cancer. Review articles on the molecular biology of human papillomavirus in HIV infection outline why, despite highly active antiretroviral therapy, anogenital tumors may continue to increase in this population. Studies continue to confirm the high prevalence of human papillomavirus infection and to define the different human papillomavirus types correlated with anogenital cytologic abnormalities, an important area in light of the development of an effective type-specific human papillomavirus vaccine. Anogenital human papillomavirus infection remains highly prevalent and persistent in HIV-infected individuals. HIV is associated with a wide diversity of human papillomavirus types and a high prevalence of anogenital cytologic abnormalities. The incidence of anogenital human papillomavirus-related cancers remains high in the highly active antiretroviral therapy era, raising concerns of human papillomavirus infections as a rising health burden among HIV-infected individuals. Interventions aimed at preventing human papillomavirus infections with vaccinations need to be evaluated in HIV-infected individuals.

High load for most high risk human papillomavirus genotypes is associated with prevalent cervical cancer precursors but only HPV16 load predicts the development of incident disease

Cervicovaginal human papillomavirus (HPV) viral load has been purported as a potential marker for the detection of high-grade cervical intraepithelial neoplasia or cancer (>/=CIN2). To examine disease association with type-specific viral load for the full-range of anogenital HPV infections, we conducted cross-sectional and prospective analyses of approximately 2,000 HPV-infected women from a 10,000-woman population-based study in Guanacaste, Costa Rica with 7 years of follow-up. Cervical specimens were tested for >40 HPV types using a MY09/MY11 L1 consensus primer PCR method with type-specific dot blot hybridization and PCR signal intensity as a measure of viral load. A positive association was observed between prevalent >/=CIN2 and high viral load compared to low viral load for women with baseline single HPV16 infections (OR = 19.2, 95% CI = 4.4-83.2) and single non-16 carcinogenic infections (OR = 9.2, 95% CI = 2.1-39.9). Inclusion of women with multiple HPV types did not substantially change these associations. In prospective follow-up, only women infected with HPV16 alone (OR = 27.2, 95% = 3.5-213.5) had a strong association between high viral load and incident >/=CIN2; non-16 carcinogenic high viral load was not associated with incident >/=CIN2 (OR = 0.7, 95% CI = 0.2-1.9). Single noncarcinogenic type viral load was not associated with increased risk of prevalent or incident >/=CIN2 (OR = 1.2 and 1.1, respectively). In conclusion, carcinogenic high viral load was associated with prevalent >/=CIN2; however HPV16 was uniquely associated with incident >/=CIN2. The extent to which these observations can be translated into clinical practice must be rigorously examined in the context of the method of viral load measurement and the type-specific differences observed for incident >/=CIN2.

The seroprevalence of IgG antibodies to human papillomavirus (HPV) types HPV‐16, HPV‐18, and HPV‐11 capsid‐antigens in mothers and their children

Human papillomavirus (HPV) types causing anogenital lesions and cancer are accepted as being sexually transmitted. The methods whereby children acquire these anogenital type HPV infections are unclear. The present study determined the prevalence of anti-HPV-16, HPV-11 and HPV-18 IgG antibodies in mothers and their children in an attempt to identify evidence of HPV transmission from mother to child. HPV virus-like particles (VLP) VLP-16, VLP-11 and VLP-18 were used in enzyme-linked immunosorbent assay to identify IgG antibodies in serum from 100 mothers and their 111 children. Antibodies to VLP-16, VLP-11 and VLP-18 were found in serum from 17%, 21% and 16% of mothers, respectively and seroprevalences were 9%, 11.7% and 9.9%, respectively amongst the children. Of the 111 children, 23 (20.7%) showed antibodies to one or more of the three HPV types tested. Seven of these (30.4%) HPV IgG positive children had the same antibodies to one or more HPV types as their mothers. The prevalence of HPV-11 was similar in children of seropositive compared with seronegative mothers (14% and 11%, respectively). The prevalence of HPV-16 and HPV-18 was higher in children of seropositive mothers compared with seronegative mothers (for HPV-16, 18% and 7%, respectively, P = 0.1, for HPV-18, 19% and 8%, respectively, P = 0.2). None of these differences were statistically significant indicating a lack of correlation between antibodies in mothers and children and no evidence to support vertical or horizontal mother to child transmission of HPV infection. Indications were of multiple sources of HPV infection in the children.

Recent Advances in the Search for Antiviral Agents against Human Papillomaviruses

Infection by human papillomavirus (HPV) is extremely common and associated with the development of benign warts or malignant lesions of the skin and mucosa. Infection by a high-risk (oncogenic) anogenital HPV type, most often through sexual contacts, is the starting point of virtually all cases of cervical cancers and the majority of anal cancers. The same viral types are also increasingly being linked with a subset of head-and-neck and non-melanoma skin cancers. Although prophylactic vaccines are now available to protect against the four types most commonly found in cervical and anal cancers (HPV16 and HPV18) and anogenital warts (HPV6 and HPV11), these neither protect against all genital HPVs nor are of therapeutic utility for already infected patients. Thus, the need for antiviral agents to treat HPV-associated diseases remains great, but none currently exist. This article reviews the recent progress made towards the development of antiviral agents to treat HPV infections, from target identification and validation to the discovery of lead compounds with therapeutic potential. Emphasis has been placed on novel low-molecular-weight compounds that antagonize HPV proteins or, alternatively, inhibit cellular proteins which have been usurped by papillomaviruses and are mediating their pathogenic effects.

HPV Infection in Men

While much is known about the natural history of cervical human papillomavirus (HPV) infection and its consequences, including cervical intraepithelial neoplasia and cervical cancer, relatively little is known about the natural history of anogenital HPV infection and diseases in men. In part this reflects difficulties in penile sampling and visual assessment of penile lesions. Anal HPV infection and disease also remain poorly understood. Although HPV is transmitted sexually and infects the genitals of both sexes, the cervix remains biologically more vulnerable to malignant transformation than does the penis or anus in men. An understanding of male HPV infection is therefore important in terms of reducing transmission of HPV to women and improving women's health. However, it is also important due to the burden of disease in men, who may develop both penile and anal cancer, particularly among HIV-positive men who have sex with men. Improved sampling techniques of the male genitalia and cohort studies in progress should provide important information on the natural history of anogenital HPV infection and disease in men, including risk factors for HPV acquisition and transmission. The impact of HPV vaccination in women on male anogenital HPV infection will also need to be assessed.

Relationships of Human Papillomavirus Type, Qualitative Viral Load, and Age with Cytologic Abnormality

Persistent cervical infections with carcinogenic human papillomaviruses (HPV) cause virtually all cervical cancer. Cytologic abnormalities are the manifestations of HPV infections used to identify women at risk. To compare the potential of the full range of anogenital HPV genotypes to induce cytopathic effects, we examined the influences of HPV type, viral load, and age on cytopathology among 1,222 women having a single HPV type at enrollment into a 10,000-woman population-based study in Costa Rica. Cervical specimens were tested for approximately 40 HPV types by MY09/MY11 L1 primer PCR and type-specific dot blot hybridization. Types were organized by phylogenetic species and cancer risk. PCR signal strength served as a qualitative surrogate for viral load. Overall, 24.8% [95% confidence interval (95% CI), 22.4-27.3] of single prevalent HPV infections had concurrent abnormalities (atypical squamous cells or worse) ranging from 0.0% to 80.0% based on HPV type. Noncarcinogenic alpha3/alpha15 types, although highly prevalent, uncommonly caused cytologic abnormalities (13.1%; 95% CI, 9.8-17.0). In contrast, one quarter to nearly one half of infections with a single major carcinogenic species type (alpha9/alpha11/alpha7/alpha5/alpha6) produced abnormalities. Greater abnormalities were observed with increasing qualitative viral load of carcinogenic types; fewer abnormalities were observed among older women (>54 years). A high percentage (46.2%) of detected abnormalities in women infected with HPV16 or related alpha9 types were high grade or worse, consistent with strong carcinogenicity, compared with 10.7% in women infected with alpha7 types, including HPV18, a major cause of adenocarcinoma. The lack of evident severe abnormalities associated with HPV18 and related HPV types might have implications for screening for poorly detected glandular and alpha7-related lesions.

PL9 The HPV‐genital (and oral?) infections – how are we standing now?

Human papillomavirus (HPV)‐associated genital pathology represents one of the major problems among STIs mostly due to the high recurrence rate, difficult eradication and oncogenic potential. Besides, the young, sexually active population in the generative period is mostly affected. Anogenital HPV infections are the most frequently diagnosed STIs of viral origin. However, oral affections caused (or induced?) by the HPV‐‘genital’ types seem to be rather rare and could be very sporadically associated with the genital lesions in the immunocompetent individuals in spite of the clear involvement of the oral mucosa in the (sexual) mode of transmission. HPV genital infections are also one of the most frequent diagnoses in the Sexually Transmitted Diseases (STD) Outpatient Clinic of the Department of Dermatology and Venereology of the Zagreb University Medical School. As the very careful and friendly‐orientated manner of taking the medical history and clinical examination is rather important in order to obtain the exact data (especially regarding the oro‐genital mode of transmission), the clinical variations are presented ranging from clinically invisible or poorly visible, ’asymptomatic’ lesions to the bizarre forms of giant condyloma of Buschke‐Löwenstein type. In spite of the fundamental importance of the clinical examination itself, we wanted to identify the HPV DNA type in these lesions. We wanted to evaluate the significance of viral tests (PCR, hybridization) for HPV‐induced, clinically visible lesions (condylomata acuminata, condylomata plana, Buschke‐Löwenstein) in men. According to our results, HPV 16 and 18 have been isolated from ’benign’ HPV‐associated genital lesions in 20% of patients, i.e. more than it is usually expected. Therefore, the diagnostic approach to HPV genital infections needs to be complex including HPV DNA typing whenever it seems appropriate. Different methods are presented for the treatment of genital warts, such as cryotherapy, podophyllotoxin, curettage, podophyllin, and imiquimod (in the smaller group, as compared to other treatment modalities). It can be concluded that no definite treatment method has been clearly found superior so far. Thus, treatment should be guided by the available resources, the experience of the provider and the preference of the patient. In general, it can be postulated that, over the last decade, the oncogenic properties of HPVs have been intensively studied. Significant progress has been achieved in the investigation of the HPV prevention. More than 35 types of HPV infect the genital tract; types 16 and 18 inducing about 70% of cervical cancer and high‐grade cervical (and not only cervical) intraepithelial neoplasia (CIN), and HPV 6 and 11 causing 90% of anogenital warts. A prophylactic vaccine that targets these types should thus substantially reduce the burden of HPV‐associated clinical diseases. The results of the most recent studies have clearly shown that a candidate quadrivalent HPV vaccine (6, 11, 16, and 18) was generally well tolerated, induced high‐titres of serum antibodies to HPV types, and effectively prevented acquisition of infection and clinical disease caused by common HPV types.

High‐risk human papillomavirus infection of the genital tract of women with a previous history or current high‐grade vulvar intraepithelial neoplasia

Human papillomavirus (HPV) infection is associated with high-grade vulvar intraepithelial neoplasia (VIN-3). The prevalence of anogenital HPV infection in women with previously treated VIN-3 has not been documented yet. This cross-sectional study compared high-risk HPV DNA detection rates in women with past (n = 30) and current (n = 22) VIN-3 to those without current or past VIN (n = 86). HPV DNA was detected in vulvar and cervical samples with Hybrid Capture 2 (HC-2). Smoking was associated in multivariate analysis with current VIN-3 (odds ratio (OR) 8.3, 95% confidence interval (CI) 2.0-8.2) and any VIN-3 history (OR 6.5, 95% CI 2.5-16.5). High-risk HPV DNA was found on the vulva of 64%, 33%, and 20% of women with current VIN-3, past VIN-3, and without previous or current VIN, respectively. After controlling for age and smoking, high-risk HPV vulvar infection was associated with cervical high-risk HPV infection (OR 8.6, 95% CI 2.8-26.5; P = 0.001). After controlling for age, HPV infection was more often multifocal in women with current VIN-3 compared to women with previous but no current VIN-3 lesion (OR 17.6, 95% CI 1.4-227.2). Multifocal vulvar HPV infection was detected in women with previous or active VIN-3. Longitudinal studies are required to determine if the multifocality of HPV infection on the vulva could explain the high recurrence rate of VIN-3.

Human Papillomavirus (HPV) Detection Among Human Immunodeficiency Virus–Infected Pregnant Thai Women: Implications for Future HPV Immunization

Human immunodeficiency virus (HIV)-infected women are at increased risk for developing cervical cancer and for infection with human papillomavirus (HPV). Prophylactic vaccines targeting HPV types 16 and 18 are being evaluated for efficacy among young women. The goal was to assess the prevalence of HPV among HIV-infected pregnant women in Bangkok and to evaluate the need for prophylactic HPV vaccines studies in this population. The study population consisted of 256 HIV-infected pregnant women who participated in a mother-to-child HIV transmission trial. Stored cervicovaginal lavage samples were tested for the presence of HPV DNA by polymerase chain reaction with PGMY09/11 primers and reverse line-blot hybridization for determination of anogenital HPV types. HPV prevalence was 35.5% (91/256); high-risk HPV prevalence was 23.4% (60/256). HPV type 16 or 18 was present in 8.2% (21/256). Almost half of all infections were multiple. Furthermore, overall HPV detection was associated with abnormal cervical cytology (P<0.001) and higher HIV-plasma viral load (P=0.007). Only one-quarter of HIV-infected pregnant women in Bangkok had high-risk HPV types; less than 10% had HPV types 16 or 18. As the HPV prevalence is expected to increase during HIV disease, prophylactic vaccines targeting HPV types 16 and 18 should be studied among HIV-infected women not yet infected with these HPV types and not previously exposed.

Methylation of the human papillomavirus-18 L1 gene: A biomarker of neoplastic progression?

Epigenetic transcriptional regulation plays an important role in the life cycle of human papillomaviruses (HPVs) and the carcinogenic progression of anogenital HPV associated lesions. We performed a study designed to assess the methylation status of the HPV-18 genome, specifically of the late L1 gene, the adjacent long control region (LCR), and part of the E6 oncogene in cervical specimens with a range of pathological diagnoses. In asymptomatic infections and infections with precancerous (precursor) lesions, HPV-18 DNA was mostly unmethylated, with the exception of four samples where hypermethylation of L1 was detected. In contrast, L1 sequences were strongly methylated in all cervical carcinomas, while the LCR and E6 remained unmethylated. HeLa cells, derived from a cervical adenocarcinoma, contain chromosomally integrated HPV-18 genomes. We found that L1 is hypermethylated in these cells, while the LCR and E6 are unmethylated. Treatment of HeLa cells with the methylation inhibitor 5-Aza-2′-deoxycytidine (5-Aza-CdR) led to the expected reduction of L1 methylation. After removal of 5-Aza-CdR, L1 methylation resumed and exceeded pretreatment levels. Unexpectedly, the LCR and E6 also became methylated under these conditions, albeit at lower levels than L1. We hypothesize that L1 is preferentially methylated after integration of the HPV genome into the cellular DNA, possibly since linearization prohibits its normal transcription, while the enhancer and promoter may be protected from methylation by transcription factors. Since our data suggest that HPV-18 L1 methylation can only be detected in carcinomas, except in some few precancerous lesions and asymptomatic infections, L1 methylation may constitute a powerful molecular marker for detecting this important step of neoplastic progression.

Development and evaluation of a highly sensitive human papillomavirus genotyping DNA chip

Objectives. Test of human papillomavirus (HPV) is a useful adjunctive tool of Pap smear to screen cervical cancer. We have developed a novel HPV genotyping DNA chip arrayed by multiple oligonucleotide probes of both L1 and E6/E7 gene sequence of 42 types of anogenital HPV. Methods. Consensus PCR products of L1 and E6/E7 gene sequences of HPV are hybridized to arrayed probes on the HPV chip and HPV genotypes are identified by fluorescence scanner. We have comparatively analyzed the value of HPV DNA chip and DNA sequencing in 100 cervical cancer tissues. Results. Overall, 98 cervical cancer tissues were found to harbor DNA sequences of high-risk type HPVs, of which 88 (89.8%) were detected by PCR-sequencing of L1 alone, 98 (100%) by PCR-sequencing of both L1 and E6/E7, and 98 (100%) by HPV DNA chip, respectively. All of the genotypes of HPV detected on sequencing analysis were also found on DNA chip analysis. HPV DNA chip was superior to direct DNA sequencing in detection of mixed infection. Conclusions. These results suggest that HPV DNA chip analysis in the present study is highly accurate for detection and genotyping of HPV and may have potential value as a robust, high-throughput screening test of uterine cervix cancer.

Anogenital and Respiratory Tract Human Papillomavirus Infections Among Children: Age, Gender, and Potential Transmission Through Sexual Abuse

To evaluate human papillomavirus (HPV) presentation among children <13 years of age and its association with suspected child sexual abuse (CSA), and to assess sexual abuse consideration among different clinical services treating these children. Records of children <13 years of age from 1985 to 2003 were selected for review if the children had a HPV-related International Classification of Diseases, Ninth Revision, code or had been examined in the CSA clinic. Abstracted data included demographic features, clinical findings, clinical services involved, age at diagnosis, age when care was first sought, and age when symptoms were first noted. HPV was identified by clinical examination and/or biopsy for 124 children, 40 with laryngeal lesions, 67 with anogenital lesions, 10 with oral lesions, and 7 with both anogenital and oral lesions. The mean age at HPV diagnosis was 4.0 +/- 2.9 years, compared with 6.4 +/- 3.0 years for 1565 HPV-negative children. Among 108 HPV cases with data for age when symptoms were first noted, the mean age was 3.3 +/- 2.9 years (median: 2.2 years) for children with anogenital and oral HPV and 2.4 +/- 2.3 years (median: 1.9 years) for children with laryngeal HPV. Among HPV-positive patients, 56% were female, compared with 82% of HPV-negative children. Fifty-five (73%) of 75 children with anogenital HPV infections were referred to the CSA clinic for evaluation, compared with none of 49 children with laryngeal or oral HPV infections treated by the otolaryngology service. Laryngeal cases presented earlier than anogenital and oral lesions. Abuse was considered at least possible for 17 of 55 children with any CSA evaluation. The mean age of likely abused, HPV-positive children was 6.5 +/- 3.8 years (median: 5.3 years), compared with 3.6 +/- 2.3 years (median: 2.6 years) for likely not abused, HPV-positive children. The likelihood of possible abuse as a source of HPV infection increased with age. The positive predictive value of HPV for possible sexual abuse was 36% (95% confidence interval: 13-65%) for children 4 to 8 years of age and 70% (95% confidence interval: 35-93%) for children >8 years of age. The data from this epidemiologic study of HPV suggest that many anogenital and laryngeal HPV infections among preadolescent children are a result of nonsexual horizontal transmission, acquired either perinatally or postnatally. It seems that many children >2 years of age acquire HPV infection from nonsexual contact. Different subspecialties vary greatly in their suspicion and evaluation of CSA. At this time, there remains no clear age below which sexual abuse is never a concern for children with anogenital HPV infections. Every case needs a medical evaluation to determine whether enough concern for abuse exists to pursue additional investigations.

Human papillomavirus DNA detection in women with primary abnormal cytology of the cervix: prevalence and distribution of HPV genotypes

In this study, we focus on the prevalence and occurrence of different anogenital human papillomavirus (HPV) genotypes in a first abnormal cervical screening test, and correlate HPV genotyping with the cytological diagnosis on thin-layer liquid-based preparations in routine gynaecological screening. Out of 780 abnormal smears, 513 tested positive for HPV. All 25 different HPV types were identified by Line Probe Assay. The prevalence of high-risk HPV types increased from 72% in atypical squamous cell of undetermined significance to 94.5% in high-grade intra-epithelial lesion (HSIL). Co-infection with multiple HPV types was predominantly found in HSIL (35.8%). In the HSIL group the most common HPV types were 16, 52, 51 and 31; type 18 was rarely present. The role of types 31, 51 and 52 should be considered in future studies on vaccine development.