Increase of human papillomavirus-16 E7-specific T helper type 1 response in peripheral blood of cervical cancer patients after radiotherapy

It has been suggested that tumour cell lysis by gamma-radiation induces a tumoral antigen release eliciting an immune response. It is not clear how a specific immune response in cervical cancer patients is developed after radiotherapy. This study is an attempt to investigate the role of the human papillomavirus type 16 (HPV-16) E7-specific T helper response before and after radiotherapy. Lymphocytes were isolated from 32 cervical cancer patients before and after radiotherapy and from 16 healthy women. They were stimulated for 12 hr with autologous HPV-16 E7-pulsed monocyte-derived dendritic cells or directly with HPV-16 E7 synthetic peptides: E7(51-70), E7(65-84) and E7(79-98). The cells were stained for CD4, CD69, intracellular interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) cytokines and analysed by flow cytometry. A specific CD4(+) CD69(+) IFN-gamma(+) immune response against HPV-16 E7(79-98) peptide was observed in 10 of 14 patients (71.4%) after treatment, compared with 4 of 14 (28.5%) before radiotherapy (P = 0.039); however, this response was not associated with a successful clinical response. Before treatment, 5 of 31 patients showed a HPV-16 E7(79-98)-specific T helper type 2 (Th2) response. Interestingly, this response was significantly associated with a decrease in disease-free survival (P = 0.027). These results suggest that a Th2-type cellular response could be useful as a predictor of recurrence and poor prognosis. An increase of the HPV-specific immune response was observed after radiotherapy; however, it is not enough to control completely the disease after treatment. Our results support that the E7-specific T-cell IFN-gamma response in cervical cancer patients, rather than reflecting the host's capability of controlling tumour growth, might be an indicator for disease severity.     

Identification of a novel human papillomavirus type 16 E1 gene variant with potentially reduced oncogenicity

The human papillomavirus (HPV) 16 genome has been studied extensively, although no study has focused on the E1 gene that is implicated in viral DNA replication. After analyzing the E1 region of HPV 16 genomes in 429 cervical samples, 11.2% were found to contain a 63 nucleotides duplication in this region. Sequence analysis of the E6 and the E7 regions has shown that all samples containing this duplication were related to E6‐G350 variant of the HPV 16 (Chi square test, P = 0.0012). A comparison of cervical lesion severity of the examinees having regular or variant E1 genes has shown that the variant group had a significantly (Fischer's exact test, P = 0.0401) lower percentage of high grade disease cases, suggesting that this particular duplication might reduce the oncogenic potential of HPV 16, and also might clarify the differences of E6‐G350 variant oncogenicity observed in European populations. Albeit, a decreased incidence of high grade cervical lesions can be linked to the prevalence of multiple HPV infection, the additional decrease of those cases with the variant E1 gene versus those without (10.5% and 18.6%, respectively) can only be ascribed to the effect of this particular HPV variant. Further research is needed to clarify the biology of these HPV 16 E1 variants. J. Med. Virol. 80:2134–2140, 2008. © 2008 Wiley‐Liss, Inc.     

Correlation between human papillomavirus infection and histopathological diagnosis of women in Northeast Brazil

Cervical carcinoma is the fourth leading cause of death among women worldwide. Epidemiological studies claim that human papillomavirus (HPV) infection is a necessary condition for cervical cancer development. Knowledge of the geographic distribution of HPV is important in guiding the introduction of prophylactic vaccines. This study analyzed the prevalence of HPV infection in cervical samples obtained from women with abnormal cervical histopathological diagnosis in Northeast Brazil. The study included an analysis of 211 women whose diagnosis was confirmed for cervical intraepithelial neoplasia type 1 (CIN-1), cervical intraepithelial neoplasia type 2 (CIN-2), cervical intraepithelial neoplasia type 3 (CIN-3), and cancer. The identification of the HPV genotypes was based on the polymerase chain reaction–restriction fragment length polymorphism technique. A total of 42.7% of the samples showed a single HPV infection, while 57.3% showed multiple infections. The most common genotypes detected were HPV-16, HPV-18, and HPV-31. HPV-16, HPV-31, HPV-35, and HPV-18 were the most common types in CIN-1 with a single infection. HPV-16 and HPV-18 were the most often found in CIN-2 with a single infection. HPV-16, HPV-18, and HPV-31 were the most detected in CIN-3 with a single infection. HPV-16 and HPV-31 were the most frequent in cancer with a single infection. Multiple infection with HPV-16 shows a 2.7 times greater risk of CIN-3 (P = .04). Multiple infections for HPV with HPV-16 and excluding the HPV18/31 types, were associated with CIN-3 (P = .01). The results allowed the detection and genotyping of HPV types circulating in the population studied. These findings must be taken into account when devising vaccination strategies against HPV.     

Parenteral and oral immunization with a plasmid DNA expressing the human papillomavirus 16-L1 gene induces systemic and mucosal antibodies and cytotoxic T lymphocyte responses.

The association of human papillomavirus (HPV) infection and cervical cancer has been demonstrated. The development of a prophylactic vaccine to protect against primary HPV infection may therefore be an efficient means to reduce the incidence of this cancer worldwide. To assess the capacity of a plasmid DNA that expresses the L1 gene of HPV type 16 to induce a protective immune response, mice were immunized by parenteral and oral routes. Animals that received the DNA vaccine intramuscularly, subcutaneously and orally, developed systemic anti-L1 IgG antibodies. Antibodies developed in mice vaccinated subcutaneously were detectable twelve months post-immunization. Specific IgA antibodies were also found in vaginal washes from immunized mice. Both systemic and local antibodies proved effective in a surrogate neutralization assay. Splenic T cells extracted from experimental mice showed cytotoxic T lymphocytes (CTL) activity mediated by CD8 + cells. Mice were challenged with a syngeneic melanoma cell line, engineered to express the HPV16-L1 protein, tumours in vaccinated animals showed slower growth rate, correlated directly with a longer survival of mice. The results suggest that the L1-based DNA vaccine may be useful for the prevention of primary infections by HPV16.     

Lineage analysis of human papillomavirus type 18 based on E6 region in cervical samples of Iranian women

Distinct human papillomavirus (HPV) 18 variants are thought to differ in oncogenic potential and geographic distribution. As such, understanding the regional variants of HPV 18 would be of great importance for evolutionary, epidemiological, and biological analysis. In this regard, the sequence variations of E6 gene were investigated to characterize more common variants of HPV 18 in normal cells, premalignant, and malignant samples collected from the cervix. In total, 99 samples of HPV 18 were analyzed by polymerase chain reaction and sequencing. In overall, lineages A was identified in all study subjects, among which sublineage A4 was dominant although the difference observed was not statistically significant with regard to different stages of disease. Sublineage A4 comprised 90.9% of samples and the remaining were belonged to sublineages A1, A2, A3, and A5 at the frequency of 6.1%, 1%, 1%, and 1%, respectively. In conclusion, our findings clearly highlight the sublineage A4 of HPV 18 as the most dominant variant in Iran.     

Association of human papillomavirus type 16 and its genetic variants with cervical lesion in Korea

Persistent human papillomavirus type 16 (HPV16) is the major risk factor for cervical cancer. HPV16 intratypic variants differ in their geographical distribution and oncogenic potential. This study aimed to analyze the distribution of HPV16 variants and their association with cervical lesion histopathology in Korean women. In total, 133 HPV16‐positive cervical samples from women admitted to Seoul National University Boramae Hospital were analyzed by sequencing E6, E7, and L1 genes and the long control region (LCR), and the variant distribution according to cervical lesion grade was determined. Isolates were grouped into a phylogenetic lineage, and A1‐3, A4, C, and D sublineages were detected in 54.1, 37.8, 0.7, and 7.4% of samples, respectively. The most commonly observed LCR variations were 7521G>A (91.5%), 7730A>C (59.6%), and 7842G>A (59.6%). Furthermore, A4 or D sublineage‐positive women had a higher risk for cervical cancer than women who were positive for A1–3. Among HPV phylogenetic clusters, A1–3 was the predominant sublineage, and within A1–3, the 350G polymorphism was highly frequent. These results differed from those of previous studies in Korea and other Asian countries. The findings suggest that cervical neoplasia incidence in HPV16‐infected patients could be affected by the distribution of HPV16 variants in the population.     

Detection of regulatory T cell phenotypic markers and cytokines in patients with human papillomavirus infection

Infection with human papillomavirus (HPV) is the main cause of cervical cancer. Viral persistence is considered the main risk factor for neoplastic progression and evidence suggests that regulatory T cells (Treg) play an important role in the failure of viral elimination. The aim of this study was to detect phenotypic markers of Treg and cytokines interleukin (IL)-10 and transforming growth factor (TGF)-β, in the cervical microenvironment of HPV-infected patients. One hundred and one samples of uterine cervix embedded in paraffin were analyzed. We used immunohistochemistry to examine the coexpression of the CD25/FOXP3 and CD4/TGF-β markers, and the expression of GITR and IL-10 in cells present in the cervical stroma. We detected a microenvironment composed of high proportions of CD25⁺FOXP3⁺, CD4⁺TGFβ⁺, IL-10⁺, and GITR⁺ cells in samples with high viral loads and severe lesions of HPV-infected patients. The abundance of these markers, indicative of the presence of Treg cells and immunosuppressive cytokines, was significantly associated with severe lesions and elevated viral loads in the examined samples. These results suggest that Treg cells may be involved in maintaining a microenvironment favorable for viral persistence and neoplastic progression. Our findings support those of previous studies that suggested that these markers could be used to predict HPV persistence and neoplastic progression, and as potential targets for immune response modulation.     

Prevalence and distribution of cervical human papillomavirus genotypes in women with cytological results from Sichuan province,China

The epidemiologic characteristics of human papillomavirus (HPV) genotypes vary by age, ethnicity, and geographic location, and the available data on HPV epidemiological characteristics with cytology results in Sichuan province are limited. Our research was conducted from June 2016 to July 2017. A total of 10 953 women getting HPV testing were enrolled. Liquid-based cytological and histological results were collected. The overall HPV infection rate was 24.1% in Sichuan province. The prevalence of high-risk HPV (hrHPV) was 19.9%. For hrHPV genotypes, HPV52 (15.5%) was the most prevalent genotype, followed by HPV16 (13.8%), HPV58 (13.3%), HPV51 (8.6%), HPV39 (8.1%), and HPV68 (7.8%). Among all HPV-positive women with a cytology or histology result, HPV16-positive women have the highest cervical intraepithelial neoplasia 1 (CIN1)+ prevalence (11.1%), followed by HPV18 and HPV33; HPV16-positive women also have the highest CIN2+ prevalence (9.3%), followed by HPV58 and HPV18. To date, this is the largest study done in the Sichuan province for HPV prevalence and subtype distribution with normal and abnormal cytological results. The age-specific prevalence in patients at gynecology clinics and other clinics is different. Besides, patients at the same age also have a different hrHPV prevalence and lrHPV prevalence. Our result revealed that in every 10 HPV16-positive women, there is approximately one women with CIN2, CIN3, or cervical cancer. A higher oncogenic potential of HPV58 than that of HPV52 was observed.     

Importance of specimen type in detecting human papillomavirus DNA from the female genital tract

HPV testing is a valuable tool in cervical cancer screening and efficacy assessment of HPV vaccines. Concordance of specimens from three sites for detection of HPV DNA in the female genital tract was evaluated. At a single visit, the following specimens were collected: an endo‐ecto‐cervical swab (EEC), labial/vulvar/perineal/perianal swab (LVPP) and cervicovaginal lavage (CVL). Specimens were evaluated with HPV6, HPV11, HPV16, and HPV18 type‐ and gene‐specific PCR assays. Of the 898 women evaluated at baseline, 232 were HPV PCR positive in at least one specimen. Of these, for HPV6, HPV11, HPV16, and HPV18, respectively, throughout: (a) 70.4%, 40.0%, 65.3%, and 64.1% tested three‐site positive; (b) 13.6%, 30.0%, 19.7%, and 18.8% tested two‐site positive; and (c) 16.4%, 30.0%, 15.0%, and 17.2% tested single‐site positive. For patients who tested single‐site positive for HPV6, HPV11, HPV16, or HPV18, respectively, the specimen was: LVPP in 92.3%, 33.3%, 68.2%, and 72.7%; EEC in 0.0%, 33.3%, 18.2%, and 9.1%; and CVL in 7.7%, 33.3%, 13.6%, and 18.2%. Combining results of swab specimens together increases detection of HPV6, HPV11, HPV16, and HPV 18, respectively, to 98.7%, 90.0%, 97.9%, and 96.9%. HPV DNA is detectable from all three sites using type‐specific PCR assays; most women who tested positive for a given HPV type were positive for that type in all three specimens. J. Med. Virol. 81:1620–1626, 2009. © 2009 Wiley‐Liss, Inc.     

Sequence variation of human papillomavirus type 16 E7 in preinvasive and invasive cervical neoplasias

Variation in the nucleotide sequence of the HPV 16 E7 gene in preinvasive cervical intra-epitherial neoplasia (CIN) and invasive cervical carcinoma specimens was analyzed. Direct DNA sequencing of PCR-amplified products with primers different from those used for PCR with 5-end labeling generated distinct sequence ladders with a low background, even in specimens containing relatively low copy numbers of HPV. Of 14 cervical neoplasias, 11 cases showed sequence diversity from prototype HPV16, and a total of 22 nucleotide exchanges were detected. Nine of these led to single amino acid exchanges: [Thr⁵] to [Lys⁵] in one case and [Asn²⁹] to [Ser²⁹] in eight cases. The [Ser²⁹] E7 was distributed uniformly among invasive carcinomas and precancerous legions, and was also found in a normal cervix. The [Lys⁵] E7 and [Ser²⁹] E7 had transforming potential similar to the prototype E7 assessed by cooperation with the activatedras gene in rat embryo fibroblasts.     

Cervical human papillomavirus among 19 753 women attending gynecological department of a major comprehensive hospital in north Anhui China 2013-2016: Implication for cervical cancer screening and prevention

Our study aimed to assess the prevalent, incident, and persistent infection, and clearance of HPV among 19 753 individual women attending the gynecological department at a major comprehensive hospital. HPV 16, 52, and 58 ranked top three types with the highest prevalence and incidence. The prevalence of high-risk (HR) HPV peaked among women aged 15 to 19 years, then sharply decreased with age, stabilized among women aged 25 to 44 years, and then surged again among women aged 45 years and older. HR HPV infection were more likely to be prevalent (15.9% vs 1.3%, P < 0.001), incident (17.3 vs 2.0 per 1000 person-months, P < 0.001), and persistent (33.0% vs 24.2%, P = 0.033), and less likely to clear (88 vs 115 per 1000 person-months, P = 0.040) compared to low-risk HPV types. The majority of women detected with HR HPV types did not retest within 12 months. Clinical guidelines on HPV DNA testing are needed and education and counseling about HPV infection and its implications for women detected with HPV at clinical settings are warranted.     

High prevalence of human papillomavirus type 58 in patients with cervical pre‐malignant lesions in southern Brazil

Cervical cancer is the second most common type of cancer in women worldwide. Several human papillomavirus (HPV) genotypes, sexual behavior, and socioeconomic profile represent major risk factors for the development of this carcinoma. Cervical invasive cancer is preceded by cellular abnormalities that can be identified by cytological or histological exams. In order to determine the prevalence and genotypes of HPV in women with abnormal cytology or histopathology, cervical cell samples from 256 patients were evaluated for the presence of HPV/DNA by polymerase chain reaction (PCR), followed by virus genotyping by restriction fragment length polymorphism (RFLP). A total of 113 samples (51.2%) were HPV/DNA positive. Viral genotyping showed that the most prevalent genotypes were HPV 16 (34.7%) and 58 (13.8%), followed by HPV 33 (9.72%), 11 (8.33%), 18 (5.55%), 53 (5.55%), and 6 (4.2%). Four samples (5.55%) exhibited multiple infections due to the great similarity of socioeconomic characteristics and sexual behavior of HPV positive women, it was not possible to establish a risk profile for female HPV infection. J. Med. Virol. 81:1270–1275, 2009. © 2009 Wiley‐Liss, Inc.     

Comparison of cervical and blood T-cell responses to human papillomavirus-16 in women with human papillomavirus-associated cervical intraepithelial neoplasia

Human papillomaviruses (HPVs) are obligate epithelial pathogens and typically cause localized mucosal infections. We therefore hypothesized that T-cell responses to HPV antigens would be greater at sites of pathology than in the blood. Focusing on HPV-16 because of its association with cervical cancer, the magnitude of HPV-specific T-cell responses at the cervix was compared with those in the peripheral blood by intracellular cytokine staining following direct ex vivo stimulation with both virus-like particles assembled from the major capsid protein L1, and the major HPV oncoprotein, E7. We show that both CD4(+) and CD8(+) T cells from the cervix responded to the HPV-16 antigens and that interferon-gamma (IFN-gamma) production was HPV type-specific. Comparing HPV-specific T-cell IFN-gamma responses at the cervix with those in the blood, we found that while CD4(+) and CD8(+) T-cell responses to L1 were significantly correlated between compartments (P = 0.02 and P = 0.05, respectively), IFN-gamma responses in both T-cell subsets were significantly greater in magnitude at the cervix than in peripheral blood (P = 0.02 and P = 0.003, respectively). In contrast, both CD4(+) and CD8(+) T-cell IFN-gamma responses to E7 were of similar magnitude in both compartments and CD8(+) responses were significantly correlated between these distinct immunological compartments (P = 0.04). We therefore show that inflammatory T-cell responses against L1 (but not E7) demonstrate clear compartmental bias and the magnitude of these responses do reflect local viral replication but that correlation of HPV-specific responses between compartments indicates their linkage.     

Prevalence and genotype distribution of human papillomavirus in non-neoplastic cervical tissue lesion: cervical erosion

Human papillomavirus (HPV) infection is the commonest sexually transmitted infection, which is associated with various clinical conditions, ranging from asymptomatic infection to malignant disease of the cervix. The aim of this study was to evaluate the prevalence and genotypic distribution of HPV in women with cervical erosion and to compare the results with those in women with a clinically normal cervix. A further aim was to establish the association between HPV infection and cervical cytology results in women with and without cervical erosion. Cervical samples were collected by liquid-based method and consecutively evaluated for the presence of HPV DNA and for cervical cytology. HPV DNA was tested by a nested polymerase chain reaction (PCR) and typed by reverse dot blot genotyping. Cytological classification was made according to Bethesda 2001 criteria. The overall HPV prevalence was 16.9%; HPV DNA was positive in 20.2% of women with cervical erosion and 12.8% in women with normal cervix (P < 0.05). Multiple infections were found in 34.1% of the HPV-positive women. Commonest types were HPV 18 (32.9%), HPV 16 (29.5%), HPV 54 (20.5%), and HPV 6 (17%). Cervical cytology results were abnormal for 5.2% of women with cervical erosion and for 1.3% with clinically normal cervix (P < 0.05). This study detected a high prevalence of HPV infection in women with cervical erosion compared to women with a normal cervix. This data may contribute to the HPV epidemiology in the southeastern Turkey. It is recommended that women with cervical erosion should be given priority in HPV screening programs.     

Detection of human papillomavirus in urine and cervical swabs from patients with invasive cervical cancer

Despite the high prevalence of both human papillomavirus (HPV) infections and cervical cancer among Zimbabwean women, the ability to test for HPV infection of the uterine cervix is limited by a lack of an easy sample collection method that does not require gynecological examination. The presence of HPVs in urine and cervical swab samples collected from 43 women who presented with invasive cervical cancer was investigated. HPV detection was done by means of degenerate primers in a nested polymerase chain reaction (PCR). Typing of HPVs was done using restriction fragment length polymorphism (RFLP) analysis. HPV was identified and typed in 98% (42/43) of cervical swabs and 72% (31/43) of paired urine samples. HPV type 16 was the most common (25/42, 59%), followed by types: 33 (13/42, 31%), 18 (6/42, 14%), and 31 (1/42, 2%). Type-specific concordance between cervical and urine samples was high (22/28, 79%). Therefore, the HPV types identified in urine samples in most cases represent the same HPV type infecting the cervical epithelium. The results suggest that urine may be a practical sample for testing of HPV urogenital infection. Further research is required before the detection of HPV in urine can be applied in the routine cervical screening programs.     

Prevalence of human papillomavirus genotypes and associated cervical squamous intraepithelial lesions in HIV-infected women in Botswana

Human papillomaviruses (HPV) constitute one of the most prevalent sexually transmitted infections and are the etiological agents for invasive cervical cancer, the predominant cancer among women in Botswana. However, the prevalence of HPV genotypes in Botswana has yet to be reported. One hundred thirty‐nine endocervical swabs were taken at baseline from HIV‐1 infected, HSV‐2 seropositive women enrolled in a longitudinal cohort study designed to assess the influence of herpes simplex virus‐2 (HSV‐2) infection on genital tract shedding of HIV‐1. Extracted DNA was evaluated for the presence of low‐risk and high‐risk HPV using the Roche Linear Array. Genotyping identified HPV in 95 of 139 women of which 61/95 were infected with high‐risk HPV and 56/95 with low‐risk HPV. The median number of genotypes was 2 (IQR: 1–4). The most prevalent HPV genotype in HIV‐infected women was HPV 58. Abnormal cervical cytology was detected in 87/127 women and was associated with contemporaneous HPV infection (RR = 1.43, 95% CI: 1.05–1.93; P = 0.02). HPV prevalence was high among HIV‐infected women with infection by multiple genotypes being widespread. The associations attributed to specific oncogenic HPV subtypes and cervical squamous intraepithelial lesions presented here provide critical information to inform future vaccine policy within Botswana. J. Med. Virol. 83:1689–1695, 2011. © 2011 Wiley‐Liss, Inc.     

Prevalence and genotype distribution of cervical human papillomavirus infection in Macao

Population‐specific epidemiological data on human papillomavirus (HPV) infection are essential for formulating strategies to prevent cervical cancer. The age‐specific prevalence of HPV infection was determined among 1,600 women enrolled for cervical screening in Macao. A U‐shaped age‐specific prevalence curve with a first peak (prevalence rate, 10%) at 20–25 years and a second peak (13%) at 51–55 years was observed. Co‐infections with multiple types were detected in 32.5% of HPV‐positive subjects and without significant variation among different age groups (P = 0.318). The majority (84.6%) of the positive samples harbored high‐ or probable high‐risk HPV types, and these types also exhibited a similar U‐shaped age‐specific prevalence curve. In contrast, low and unknown‐risk HPV types remained at a low prevalence (1.5–2.5%) throughout the age groups between 20 and 50 years, and with a small peak (4.5%) at 51–55 years. HPV 52 was the most common type found in 26.8% of positive samples, followed by HPV 16 (15.5%), HPV 68 (11.4%), HPV 18 and HPV 58 (8.9% each), HPV 54 (8.1%), HPV 53 (7.3%), HPV 39 (6.5%), HPV 33 and HPV 66 (5.7% each). In conclusion, because of the early peak of infection, vaccination and educational campaigns in Macao should start early and target at teenagers. The presence of a second peak containing mainly high‐risk HPV types in older women indicates the need to evaluate the cover of the cervical screening programme for older women. Further study to determine the contribution of HPV 52 in high‐grade cervical neoplasia and invasive cancers in Macao is warranted. J. Med. Virol. 82:1724–1729, 2010. © 2010 Wiley‐Liss, Inc.