Identification of human papillomavirus type 58 lineages and the distribution worldwide.

BACKGROUND: Human papillomavirus type 58 (HPV-58) accounts for a much higher proportion of cervical cancers in East Asia than other types. A classification system of HPV-58, which is essential for molecular epidemiological study, is lacking. METHODS AND RESULTS: This study analyzed the sequences of 401 isolates collected from 15 countries and cities. The 268 unique concatenated E6-E7-E2-E5-L1-LCR sequences that comprised 57% of the whole HPV-58 genome showed 4 distinct clusters. L1 and LCR produced tree topologies that best resembled the concatenated sequences and thus are the most appropriate surrogate regions for lineage classification. Moreover, short fragments from L1 (nucleotides 6014-6539) and LCR (nucleotides 7257-7429 and 7540-52) were found to contain sequence signatures informative for lineage identification. Lineage A was the most prevalent lineage across all regions. Lineage C was more frequent in Africa than elsewhere, whereas lineage D was more prevalent in Africa than in Asia. Among lineage A variants, sublineage A2 dominated in Africa, the Americas, and Europe, but not in Asia. Sublineage A1, which represents the prototype that originated from a patient with cancer, was rare worldwide except in Asia. CONCLUSIONS: HPV-58 can be classified into 4 lineages that show some degree of ethnogeographic predilection in distribution. The evolutionary, epidemiological, and pathological characteristics of these lineages warrant further study.

Transcriptional repression of WEE1 by Kruppel-like factor 2 is involved in DNA damage-induced apoptosis.

Human Kruppel-like factor 2 (KLF2) is a Cys(2)/His(2) zinc-finger-containing transcriptional factor, which is involved in multiple cellular pathways. Utilizing gene expression profiling to identify aberrantly expressed genes in ovarian cancer, we found that KLF2 was significantly and specifically downregulated in ovarian tumors. After reintroducing KLF2 into ovarian cancer cell lines, we observed decreased cell growth and increased sensitivity to DNA damage-induced apoptosis. Analysis of genes that could be potential targets of KLF2 revealed that KLF2 negatively regulated WEE1 expression. WEE1 encodes a tyrosine kinase that regulates the G2/M cell cycle transition. Expression of KLF2 markedly repressed the transcription of WEE1 by directly binding to an SP1/CPBP motif located between -252 bp and the start codon of the WEE1 promoter. Both activation and zinc-finger domains of KLF2 were required for this suppression of Wee1 expression. In addition, we demonstrated that Wee1 expression prevents cancer cells from undergoing apoptosis in response to DNA damage; however, this resistance was abolished by coexpression of KLF2, which inhibits WEE1 transcription. Thus, the level of WEE1 is regulated by KLF2 and enhanced KLF2 expression sensitizes cells to DNA damage-induced apoptosis.

Detection and quantitation of human papillomavirus DNA in primary tumour and lymph nodes of patients with early stage cervical carcinoma.

Fifteen Chinese women with early stage cervical squamous cell carcinoma (14 stage IB, one stage IIA) were retrospectively analysed for the correlation between human papillomavirus (HPV) load in primary tumour and the presence of HPV DNA in histologically tumour-free pelvic lymph nodes. HPV16 DNA was detected from majority (12/15) of primary tumours, with a viral load ranging from 12 to 1800 copies per cell. Of the 156 histologically tumour-free pelvic lymph nodes, 41 (26.3%) were positive for HPV DNA. The levels of viral load detected in histologically tumour-free lymph nodes were low and most were not detectable by the less sensitive consensus PCR GP5+/6+. Among patients without histological evidence of nodal involvement, the presence of HPV DNA in lymph nodes was associated with a significantly higher viral load in primary tumour (mean [interquartile range]=800 [600-1450] versus 40 [19-70] copies per cell, P=0.016). Three of the four patients with recurrence had histological evidence of lymph node metastases. In contrast, none of the seven patients with HPV DNA-positive lymph nodes but without histologically evidence of nodal involvement developed recurrence. The results of this study suggest that the presence of HPV DNA in histologically tumour-free lymph nodes do not have prognostic significance. The HPV DNA detected from lymph nodes may have originated from circulating necrotic tumour cells or those internalized by scavengers, which was easier to be detected when the viral load per tumour cell was high.

Antagonist-mediated down-regulation of Toll-like receptors increases the prevalence of human papillomavirus infection in systemic lupus erythematosus.

INTRODUCTION: Prevalence of an abnormal Papanicolaou smear was significantly increased in lupus patients in cross-sectional studies, associated with a higher prevalence of high-risk human papillomavirus (HPV) infection. The nucleic acid-specific Toll-like receptors (TLRs) locate at the endolysosomal compartments and trigger the induction of cytokines for the innate immune response. This study evaluated whether abnormal host innate immune response in lupus patients may enhance HPV persistence. METHODS: Protein levels of TLRs 3, 7, 8 and 9 in cervical epithelial cells of lupus patients and controls with or without HPV infection were assessed using flow cytometry. Characteristics associated with the differential expression of TLRs in systemic lupus erythematosus (SLE) were elucidated. The effect and interferon-stimulated genes (ISGs) (ISG15 and Mx-1) gene expressions were then measured in oncogenic HeLa (HPV18), CaSki (HPV) and C33A (HPV negative) cell lines using flow cytometry and quantitative real-time PCR. Ex vivo productions of cytokines and interferon-gamma (IFN-γ) upon TLR ligands stimulations were subsequently measured using cytometric bead array and ELISA. RESULTS: For subjects with HPV infection, levels of TLR3 and TLR7 were significantly lower in lupus patients compared with controls. Significantly decreased TLRs 7, 8 and 9 levels were observed in HPV-negative SLE compared to healthy controls. For SLE with and without HPV infection, TLR7 and 9 levels were significantly lower in infected SLE than those in HPV-negative patients. Independent explanatory variables associated with down-regulation of TLR7 level included HPV infection and a higher cumulative dose of prednisolone; while a higher cumulative dose of hydroxychloroquine and HPV infection were associated with down-regulation of TLR9 level. In cervical cell lines, TLRs 3, 7, 8, 9 protein levels and antiviral ISG15 and Mx-1 gene expressions were inhibited in two oncogenic HPV types. Functional data showed that the induction of pro-inflammatory cytokines by TLR ligands (R837, ssRNA and ODN2395) was greatly impaired in CaSki and HeLa than C33A cells. CONCLUSIONS: In conclusion, prednisolone and TLR antagonist (hydroxychloroquine) may down-regulate protein levels of TLR7 and TLR9 in lupus patients, thereby decreasing the innate immune response against HPV infection. Upon infection, HPV further down-regulate TLR7 and 9 levels for viral persistence. Furthermore, reduction of nucleic acid-sensing TLRs 7, 8 and 9 in carcinogenic HPVs ensures that the expression of inducible pro-inflammatory cytokines is minimized to prevent the expression of antiviral ISGs (ISG15 and Mx-1) on a biologically relevant antiviral response.

Clinical use of laparoscopic ultrasonography in detecting nodal metastasis in advanced-stage cervical carcinoma.

OBJECTIVE: To assess the clinical use of a laparoscopic ultrasound scan (LUS) to identify pelvic and para-aortic node metastasis in patients with advanced-stage cervical cancer. METHODS: After examination under general anesthesia and cystoscopy, LUS was used to examine the pelvic nodes of patients with advanced-stage cervical cancer. Abnormal nodes were excised before definitive treatment to confirm the nodal status. Patients without abnormal para-aortic nodes on preoperative computer tomography/magnetic resonance imaging in the past 3 years were surgically staged via laparoscopic extraperitoneal aortic node sampling, and the findings were correlated with LUS findings. The predictive values of abnormal pelvic nodes on LUS for pelvic and aortic node metastasis were determined. RESULTS: A total of 119 advanced-stage cervical cancer patients underwent LUS of pelvic nodes. Abnormal pelvic nodes were found in 62 (52.1%) patients, and metastasis was confirmed by histology in 38 (31.9%) patients. Three patients had micro-metastasis in para-aortic nodes, and all of these patients had abnormal pelvic lymph nodes on LUS. CONCLUSION: Abnormal pelvic nodes are commonly found on LUS in patients with advanced-stage cervical cancer, and selective excision biopsy is needed to confirm pelvic node metastasis. Surgical staging of aortic nodes might be considered for patients with abnormal pelvic nodes on LUS.

Natural history of cervical papilloma virus infection in systemic lupus erythematosus - a prospective cohort study.

OBJECTIVE: To ascertain the incidence, cumulative prevalence, persistence, and clearance of human papilloma virus (HPV) infection in patients with systemic lupus erythematosus (SLE), and to assess the risk factors for the acquisition and persistence of HPV infection. METHODS: One hundred forty-four patients with SLE were evaluated at 6-month intervals for up to 3 years. At each visit, a Pap test, a test for HPV DNA, and clinical assessment were performed. RESULTS: The cumulative prevalence of HPV infection increased significantly (12.5% at baseline to 25.0% after 3 years; p = 0.006). Regarding type-specific HPV infection, 18.8% patients experienced 68 incident infections. The cumulative prevalence of high-risk HPV infection (11.1% at baseline to 20.8% after 3 years; p = 0.02) and multiple HPV infection also increased significantly (6.9% at baseline to 16.7% after 3 years; p = 0.009). Half (33/68, 48.5%) of the incident infections persisted for >or= 6 months. Overall, 29/32 (90.6%) of the preexisting infection and 10/68 (14.7%) of the incident infections were cleared. Independent risk factors associated with incident HPV infection included younger age at first sexual intercourse (p = 0.025) and baseline Systemic Lupus International Collaborating Clinics score >or= 1 (p = 0.038). Independent risk factor associated with persistent HPV infection included preexisting HPV infection (p = 0.04) and multiple HPV infection during first incident infection (p = 0.02). CONCLUSION: High frequency of persistent HPV infection, especially high-risk and multiple HPV infection, may explain why squamous intraepithelial lesions occurred frequently in patients with SLE. Patients with high inflammatory burden are at risk of acquiring HPV infection.

HLA-DQB1 polymorphisms and risk for cervical cancer: a case-control study in a southern Chinese population.

OBJECTIVES AND METHODS: HLA II DQB1 polymorphisms have been shown to associate with cervical cancer risk, but results varied among different populations. In this study, the HLA DQB1 alleles among 221 southern Chinese women with cervical intraepithelial neoplasia grade III (CIN III)/invasive cervical carcinoma (ICC) were compared to 191 controls. RESULTS: The frequency of DQB1*03 was significantly lower among ICC overall as compared to controls (65.4% vs. 79.1%, odds ratio [95% confidence interval]: 0.50 [0.28-0.88], corrected p-value: 0.04). The protective association of DQB1*03 remained significant for human papillomavirus (HPV) 16-positive ICC, but not for HPV16-negative cases. This is in contrast to studies on European populations where DQB1*03 was associated with an increased risk for ICC. In the current study, 70.1% of the HPV16 isolates were Asian variants, and 28.0% were European variants. However, no significant association between HPV16 variant and DQB1*03 distribution was observed. HPV52 and HPV58 were found respectively in 16.3% and 10.0% of CIN III/ICC, which were higher compared to that of Europe and North America. Further analyses revealed a positive risk association between DQB1*06 and HPV58-positive CIN III/ICC (3.68 [1.37-9.92], corrected p-value: 0.012). CONCLUSION: The host genetics and the distribution of HPV types/variants may account for the observed differences among southern Chinese and other populations.

Expression of maspin in gestational trophoblastic disease.

BACKGROUND: Maspin is a tumor suppressor gene whose expression is altered in neoplasia and malignancies of many tissues. In the human placenta, the maspin gene is expressed in trophoblastic cells and might act as an inhibitory regulator of trophoblastic invasion. Hence, in gestational trophoblastic disease (GTD), where there is increased propensity for invasion in the trophoblastic tissue, we hypothesized that maspin expression would be decreased. The present study aimed at investigating the expression of maspin in GTD and its prognostic significance. METHODS: Using immunohistochemical staining, we firstly studied the expression of maspin in hydatidiform moles, with gestational age-matched normal first trimester placenta used as control. A total of 38 cases of hydatidiform moles were studied, including 20 complete moles (CM) and 18 partial moles (PM). Among them, 10 cases of the CM group and 8 cases of the PM group subsequently developed gestational trophoblastic neoplasia (GTN). Immunostaining was also performed on tissue from 4 cases of choriocarcinoma and 5 cases of placental site trophoblastic tumor. Reverse transcriptase-polymerase chain reaction (RT-PCR) was further performed on RNA extracted from 10 hydatidiform moles (5 with GTN and 5 without) and 6 normal first-trimester placentae. RESULTS: In all tissue sections, nuclear expression of immunostaining signal was demonstrated, mainly in the cytotrophoblasts. The percentage of trophoblastic nuclei stained in both complete and partial moles was significantly lower than that in normal first-trimester placenta (P < 0.001). However, there was no significant difference in immunostaining between complete and partial moles (P > 0.05). There was also significantly lower expression of maspin in those cases subsequently developing GTN than those which did not (P = 0.01). Immunostaining on choriocarcinoma and placental site trophoblastic tumor showed reduced expression of maspin in all the tumor cells. Reverse transcriptase-polymerase chain reaction revealed that the expression of maspin was consistently down-regulated in all the hydatidiform mole samples. CONCLUSIONS: Our results suggest that there is down-regulated expression of maspin in gestational trophoblastic diseases, and the down-regulation is more prominent in cases developing gestational trophoblastic neoplasia. This may play a role with prognostic significance in the pathogenesis and malignant transformation of hydatidiform moles.

Biases in human papillomavirus genotype prevalence assessment associated with commonly used consensus primers.

Consensus primers targeting human papillomaviruses (HPVs) have biases in sensitivity toward certain HPV types. We applied 3 primer sets (GP5+/6+, MY09/11, PGMY09/11) in parallel on 120 Chinese cervical cancer specimens. GP5+/6+ exhibited a poor sensitivity for HPV52, for which the prevalence among squamous cell cervical cancer was underestimated from 14.6% to 0%. The fact that HPV52 should rank second in prevalence among squamous cell cervical carcinoma in Hong Kong could be missed if GP5+/6+, a worldwide commonly used primer set, was selected for HPV detection. Biases in HPV type-specific sensitivity may result in misprioritization of vaccine candidates.

Expression of leptin and leptin receptors in gestational trophoblastic diseases.

OBJECTIVES: To investigate the expression profile of leptin and leptin receptors in gestational trophoblastic diseases (GTDs). METHODS: Using immunohistochemical staining on archival paraffin-embedded tissue sections, we studied the expression of leptin and leptin receptor in hydatidiform moles, with gestational age-matched normal first-trimester placenta used as control. A total of 38 cases of hydatidiform moles were studied, including 20 complete moles (CHMs) and 18 partial moles (PHMs). Among them, 10 cases of the CHM group and 8 cases of the PHM group subsequently developed residual trophoblastic disease (RTD). In addition, two cases of choriocarcinoma and three cases of placental site trophoblastic tumor (PSTT) were also studied. Reverse transcriptase-polymerase chain reaction (RT-PCR) was further performed using RNA extracted from frozen tissue (five CHMs, four PHMs and nine normal first-trimester placenta) to study the expression of leptin and individual leptin receptor isoforms at the transcription level. RESULTS: In all tissue sections, immunostaining signal was shown in the cytoplasmic compartment of cytotrophoblasts and syncytiotrophoblasts, with much stronger staining in the former. Significantly higher immunostaining intensity was shown for both leptin (P < 0.05) and leptin receptor (P < 0.001) in both CHMs and PHMs compared to normal first-trimester placenta. There was no significant difference between those cases subsequently developing RTD and those which did not (P > 0.05). In the choriocarcinoma and PSTT cases, intense immunostaining was found in the tumor cells. RT-PCR revealed that the expression of leptin and all leptin receptor isoforms were significantly higher in both CHMs and PHMs than in normal placenta (P < 0.05). CONCLUSIONS: There is up-regulated expression of leptin and leptin receptor in GTDs. However, there is no obvious correlation with the development of RTD. The exact role played by leptin and its receptors in the pathogenesis of GTDs awaits further investigations.