Norcantharidin Enhances the Antitumor Effect of 5-Fluorouracil by Inducing Apoptosis of Cervical Cancer Cells: Network Pharmacology, Molecular Docking, and Experimental Validation.

The high recurrence rate of cervical cancer is a leading cause of cancer deaths in women. 5-Fluorouracil (5-FU) is an antitumor drug used to treat many types of cancer, but its diminishing effectiveness and side effects limit its use. Norcantharidin (NCTD), a demethylated derivative of cantharidin, exhibits various biological activities. Here, we investigated whether NCTD could potentiate 5-FU to induce cervical cancer cell death. To assess the cell viability and synergistic effects of the drugs, cell counting kit-8 and colony formation assays were performed using HR-HPV-positive cervical cancer cell lines. Annexin V-FITC/PI staining and TUNEL assays were performed to confirm the induction of apoptosis. The synergistic effect of NCTD on the antitumor activity of 5-FU was analyzed using network pharmacology, molecular docking, and molecular dynamics simulations. Apoptosis-related proteins were examined using immunoblotting. The combination of NCTD and 5-FU was synergistic in cervical cancer cell lines. Network pharmacological analysis identified 10 common targets of NCTD and 5-FU for cervical cancer treatment. Molecular docking showed the strong binding affinity of both compounds with CA12, CASP9, and PTGS1. Molecular dynamics simulations showed that the complex system of both drugs with caspase-9 could be in a stable state. NCTD enhanced 5-FU-mediated cytotoxicity by activating apoptosis-related proteins. NCTD acts synergistically with 5-FU to inhibit cervical cancer cell proliferation. NCTD enhances 5-FU-induced apoptosis in cervical cancer cell lines via the caspase-dependent pathway.

Detection of antibody subclasses IgA, IgM and IgG against HPV L1 in HPV-positive oropharyngeal squamous cell carcinoma patients: a pilot study.

PURPOSE: Despite prognostic superiority of HPV-positive oropharyngeal squamous cell carcinoma (OPSCC), up to 25% of patients will suffer from recurrence within the first 5 years. Therefore, it is of great scientific interest to find relevant biomarkers to identify patients at risk. In this prospective observational study, we aimed to investigate the dynamics of HPV-L1 capsid protein specific antibody (AB) subclasses IgA, IgM, and IgG in HPV-positive OPSCC patients under therapy. METHODS: Serum samples from HPV-positive OPSCC patients, identified by positive p16-immunohistochemistry, were collected before and during tumor-specific therapy and 3-6 months during follow-up. They were analyzed for the presence of HPV-L1 AB subclasses IgA, IgM, and IgG using an HPV-L1-specific immuno-assay. Additionally, a PCR-based HPV-DNA detection from the tumor tissue was performed. RESULTS: Altogether, 33 patients with a mean follow-up of 55 months were included. Analysis of a total of 226 serum samples revealed that the most common L1-AB-subclass pattern was characterized by IgG > > IgA > IgM without significant fluctuation during the course of disease. Patients with excessive IgG levels tended to higher tumor stages and three out of three patients with disease recurrence showed increasing IgG AB titers beforehand. Seven patients showed an IgA dominance at diagnosis, which was associated with a better disease-free survival. CONCLUSION: Despite limited cases, our prospective pilot study revealed promising trends in HPV L1 AB subclasses and may contribute useful information for future risk stratification and post-treatment monitoring in HPV-positive OPSCC patients.

"This is really something: same place, same day result, same day treatment" women's experiences of testing positive for HPV and receiving same-day treatment in Papua New Guinea: an interpretative phenomenological analysis.

INTRODUCTION: Human papillomavirus (HPV) testing is transforming cervical screening globally. The World Health Organization (WHO) now recommends same-day HPV screen-and-treat for primary cervical screening in low- and middle-income countries (LMIC) but there is a lack of evidence on women's lived experience of testing positive for oncogenic HPV and receiving same-day treatment. This study aimed to address this knowledge gap among women participating in a same-day HPV screen-and-treat (HPV S&T) program in Papua New Guinea. METHODS: As part of a larger qualitative study, this paper explores the lived experiences of 26 women who tested positive for oncogenic HPV and were treated the same day. We analysed the data using the interpretative phenomenological analysis method. All data were managed using Nvivo 12.5. RESULTS: The interpretative phenomenological analysis led to three superordinate themes: 1) facing and alleviating initial worries, (2) transforming the disclosure process, and (3) connecting to their faith. Women's experiences of the same day HPV screen-and-treat were framed by initial emotional reactions to their positive HPV test result, and having access to treatment on the same day, which helped address their worries and fears, and transformed their experience of disclosing their test result and subsequent treatment to family and friends. CONCLUSION: This study shows that, while women experience similar initial emotional reactions, undergoing same day treatment quickly resolved the women's worries, making this program highly acceptable. Overall, women's engagement in the program confirmed its high acceptability and cultural congruence, leaving women feeling empowered and hopeful about their future, and the future of all Papua New Guinea women.

Facility volume and survival: Human papilloma virus positive oropharyngeal squamous cell carcinoma.

BACKGROUND: To analyze the impact of facility volume on survival for human papilloma virus positive oropharyngeal squamous cell carcinoma (HPV+ OPSCC) patients. METHODS: Patients treated for HPV+ OPSCC from 2010 to 2017 were queried from the National Cancer Database. Facilities of average annual case volume <50th percentile were categorized as low-volume (LV) and >95th percentile as high-volume (HV). RESULTS: 11,546 were included, with 10,305 patients (89.3 %) treated at LV and 1241 (10.7 %) at HV facilities. A greater proportion of cases involving resection of base of tongue and lingual tonsil were treated at HV (30.3 %) compared to LV (22.3 %) facilities (p < 0.001). Patients treated at a HV facility had greater percentage of clinical T4 (11.2 % vs. 8.6 %, p = 0.001) and N+ disease (90.5 % vs. 85.7 %, p < 0.001) patients. Survival analysis showed no statistically significant difference between five-year overall survival rates by facility volume (p = 0.388) for all patients. On multivariable analysis, facility volume was not associated with survival (HR: 0.968 [0.758-1.235], p = 0.791). These trends were found for both patients undergoing primary surgery or chemoradiotherapy. CONCLUSION: Our data indicates that patients with HPV+ OPSCC do not experience a survival benefit with treatment at HV facility, suggesting these patients may be adequately treated at LV centers.

Prognostic significance of extra-nodal extension and positive surgical margins in HPV positive oropharyngeal squamous cell carcinoma.

BACKGROUND: Whether extra-nodal extension (ENE+) and surgical margin positivity (margin+) are poor prognostic factors in HPV-associated (HPV+) oropharyngeal carcinoma (OPC) remains uncertain. RESULTS: Our study evaluated if microscopic ENE+ and/or margin+ are associated poorer recurrence free survival (RFS) and overall survival (OS) in HPV+ OPC. Patients were classified as high risk (ENE+ and/or margin+) or low risk (ENE- and margins-). Of a total of 176 patients HPV+ OPC, 81 underwent primary surgery and dad data on ENE and margin status. There was no statistically significant difference in RFS (p = 0.35) or OS (p = 0.13) for high-risk versus low-risk groups. Ongoing smoking (p = 0.023), alcohol use (p = 0.044) and advanced stage (p = 0.019) were associated with higher risk of recurrence. Only advanced stage (p-value <0.0001) was associated poorer overall survival. CONCLUSIONS: The presence of ENE+ and/or margin+ was not an independent predictor of poor RFS or OS in HPV+ OPC.

The functions of lncRNAs in the HPV-negative cervical cancer compared with HPV-positive cervical cancer.

Cervical cancer is one of the most common female malignancies. Human papillomaviruses (HPV) are the main causative agents of virtually all cervical carcinomas. Nevertheless, emerging evidence has demonstrated that a small proportion of cervical cancer patients are HPV negative. Long noncoding RNAs (lncRNAs) have been identified to play a crucial role in cervical cancer development. Here, this review describes the incidence and development of HPV-negative cervical cancer. Moreover, HPV-negative cervical cancers are more likely diagnosed at non-squamous type, older ages, more advanced stage and metastases, and associated with poorer prognosis as compared to HPV-positive cervical cancer. Furthermore, the significant role and functions of lncRNAs underlying HPV-negative cervical cancer is clarified.

TMEM45A Affects Proliferation, Apoptosis, Epithelial-Mesenchymal Transition, Migration, Invasion and Cisplatin Resistance of HPV-Positive Cervical Cancer Cell Lines.

To investigate the effects of transmembrane protein 45A (TMEM45A) on biological characteristics and cisplatin (DDP) resistance of cervical cancer cells. TMEM45A in cervical cancer cells and normal cervical epithelial cells (HCerEpiC) were quantified by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. HPV genotypes were identified by multiplex PCR. SiHa and HeLa cells were divided into Blank, shCTL, shTMEM45A-1, and shTMEM45A-2 groups, followed by Cell Counting Kit-8 (CCK-8), EdU, Annexin V-FITC/PI staining, Wound healing, and Transwell invasion assays, as well as qRT-PCR and Western blotting. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide) was employed to evaluate the impact of TMEM45A shRNA on cisplatin-resistant cervical cancer cells (SiHa/DDP and HeLa/DDP). Compared with HcerEpic cell, cervical cancer cells exhibited the upregulation of TMEM45A expression, especially in HPV-positive cell lines (CaSki, SiHa, HeLa). TMEM45A shRNA suppressed the proliferation of SiHa and HeLa cells, arrested cells at the S phase, and promoted cell apoptosis. TMEM45A shRNA inhibited the epithelial-mesenchymal transition (EMT), invasion, migration of SiHa and HeLa cells, accompanying by the downregulated Vimentin and N-cadherin with the upregulated E-cadherin. Moreover, SiHa/DDP and HeLa/DDP had higher TMEM45A expression than their parental SiHa and HeLa cells, respectively. And inhibiting TMEM45A can reduce the IC50 of SiHa/DDP cells and HeLa/DDP cells to cisplatin. Silencing TMEM45A can inhibit cell proliferation, invasion, migration and EMT, regulate cell cycle distribution, promote cell apoptosis, and reverse cisplatin resistance of HPV-positive cervical cancer cells, highlighting that inhibition of TMEM45A may be a therapeutic strategy for HPV-positive cervical cancer.

A cost-utility analysis comparing CT surveillance, PET-CT surveillance, and planned postradiation neck dissection for advanced nodal HPV-positive oropharyngeal cancer.

BACKGROUND: The cost utility of image-guided surveillance using computed tomography (CT) and positron emission tomography (PET)-CT to planned postradiation neck dissection (PRND) was compared for the management of advanced nodal human papillomavirus-positive oropharyngeal cancer following chemoradiation. METHODS: A universal payer perspective was adopted. A Markov model was designed to simulate four treatment approaches with 3-month cycles over a lifetime horizon: 1) CT surveillance, 2) standard PET-CT surveillance, 3) a novel PET-CT approach with repeat PET at 6 months postchemoradiation for equivocal responders, and 4) PRND. Parameters including probabilities of CT nodal progression/resolution, PET avidity, recurrence, and survival were obtained from the literature. Costs were reported in 2019 Canadian dollars and utilities were expressed in quality-adjusted life years (QALYs). Deterministic and probabilistic sensitivity analyses were performed to evaluate model uncertainty. RESULTS: PET-CT surveillance dominated CT surveillance and PRND in the base case scenario, and the novel PET-CT approach was the most cost-effective strategy across a wide range of variables tested in one-way sensitivity analysis. On probabilistic sensitivity analysis, novel PET-CT surveillance was the most cost-effective strategy in 78.1% of model iterations at a willingness-to-pay of $50,000/QALYs. Novel PET-CT surveillance resulted in a 49% lower rate of neck dissection compared with traditional PET-CT, and yielded an incremental benefit of 0.14 QALYs with average cost savings of $1309. CONCLUSIONS: Image-guided surveillance including PET-CT and CT are more cost effective than PRND. The novel PET-CT approach with repeat PET for equivocal responders was the dominant strategy and yielded both higher benefit and lower costs compared with standard PET-CT surveillance.

Comparison of different strategies for the triage to colposcopy of women tested high-risk HPV positive on self-collected cervicovaginal samples.

OBJECTIVE: To identify the optimal strategy for the triage of women who test high-risk (hr) HPV positive on self-collected cervicovaginal samples. METHODS: This is a diagnostic accuracy sub-analysis of the GRECOSELF study, which reported on HPV-DNA testing with self-sampling in Greece. More than 13,000 women, 25-60 years old, who resided in rural areas of Greece, provided a self-collected cervicovaginal sample. Samples were tested for HPV-DNA and HPV16/18 genotyping with the cobas® HPV test (Roche® Molecular Systems, Pleasanton, CA, USA). HrHPV positive women were referred for colposcopy. Prior to colposcopy a physician-collected sample was obtained for cytology. After colposcopy/biopsy, women were classified as having cervical disease or not, and treated accordingly. RESULTS: Out of 1070 hrHPV positive women, 773 were subjected to colposcopy. Seventeen triage strategies, combining HPV16/18 genotyping and cytology, were investigated. The strategy referring to colposcopy women positive for HPV16 regardless of the cytology report, and women positive for other hrHPVs, in case of a subsequent atypical squamous cells of undetermined significance or worse (ASCUS+) cytology report, presented optimal trade-off; sensitivity 96.36% [(95%CI: (91.41-100.0)], positive predictive value (PPV) 27.46% [95%CI: (21.16-33.76)], and number of colposcopies required to detect one case of Cervical Intraepithelial Neoplasia grade-2 or worse (CIN2+) 3.64. CONCLUSIONS: The optimal strategy for the triage to colposcopy of hrHPV positive women, detected with the cobas® HPV test on self-collected cervicovaginal samples, is referring all HPV16 positive women directly to colposcopy, and women positive for HPV18 or other hrHPVs only after an ASCUS or worse cytology report.

DOCK8 Serves as a Prognostic Biomarker and Is Related to Immune Infiltration in Patients With HPV Positive Head and Neck Squamous Cell Carcinoma.

PURPOSE: Dedicator of cytokinesis 8 (DOCK8) was reported to have a vital link to immunoregulation. However, the mechanisms by which it drives immune infiltration in cancer remain uncertain. We tried to assess the role of DOCK8 in patients with cancer, especially human papillomavirus (HPV)-positive head and neck squamous cell carcinoma (HNSCC). METHODS: Data on the expression and survival of DOCK8 in patients with various cancers were analyzed using the Oncomine and TIMER databases. The TIMER database assessed the relationship of DOCK8 with immune infiltration levels and various markers of multiple immune cells. Gene set enrichment analysis revealed tumor-associated biological processes related to DOCK8. ENCODE database was used to explore relevant transcription factors of DOCK8, and a PPI network was constructed using GENEMINIA. The expression and survival role of DOCK8 was confirmed in patients from independent GEO datasets. RESULTS: We determined that DOCK8 expression was upregulated or downregulated in various cancers unlike in healthy tissues. A high expression of DOCK8 was significantly correlated with a favorable prognosis in HPV-positive HNSCC and lung adenocarcinoma (LUAD). Furthermore, multivariate Cox regression analysis revealed that DOCK8 was an independent prognostic factor of HPV-positive HNSCC. Additionally, elevated DOCK8 expression was positively correlated with multiple immune cell infiltration levels and immune marker expression associated with particular immune cell subsets. Also, 14 pathways involved in immune activities and carcinogenesis, 22 potential TFs, and co-expression proteins of DOCK8 indicated DOCK8 to be related to tumor-associated biological processes. Ultimately, we verified that DOCK8 is upregulated and confers a favorable overall survival and progression-free survival status in patients with HPV-positive HNSCC. CONCLUSION: These results elucidate that high expression of DOCK8 indicates a favorable prognosis in patients with HPV-positive HNSCC as well as increased microenvironmental immune infiltration levels. It would provide new insights into the prognosis predicting and clinical regimen decision making in patients with HPV-positive HNSCC.

Epidermotropic metastasis of squamous cell carcinoma of the tonsil: A case report with molecular confirmation.

Metastasis of oropharyngeal squamous cell carcinoma (SCC) to skin is uncommon and portends a poor prognosis. Clinical history and histopathology are key to discerning between metastatic disease vs de novo SCC of the skin. We describe a case of an HPV+ tonsillar SCC in a 77-year-old male, with metastasis to the neck skin. This case is unique because of prominent in situ epidermal involvement on skin biopsy specimen, complicating the distinction between primary and secondary disease. The nature of the lesion was resolved using next-generation sequencing of both the primary oropharyngeal SCC and skin lesion biopsy specimens. Both tumors showed identical ATR D1639G somatic mutations, while the skin lesion contained an additional POLE F1366L mutation. Clonal evolution of metastatic lesions is a well-described phenomenon; comparing the genetic profiles of primary and metastatic specimens can be useful in evaluating the tumor origin as well as identifying targetable genetic aberrations.

Bazedoxifene, a GP130 Inhibitor, Modulates EMT Signaling and Exhibits Antitumor Effects in HPV-Positive Cervical Cancer.

Persistent HPV (Human Papillomavirus) infection is the primary cause of cervical cancer. Despite the development of the HPV vaccine to prevent infections, cervical cancer is still a fatal malignant tumor and metastatic disease, and it is often difficult to treat, so a new treatment strategy is needed. The FDA-approved drug Bazedoxifene is a novel inhibitor of protein-protein interactions between IL-6 and GP130. Multiple ligand simultaneous docking and drug repositioning approaches have demonstrated that an IL-6/GP130 inhibitor can act as a selective estrogen modulator. However, the molecular basis for GP130 activation in cervical cancer remains unclear. In this study, we investigated the anticancer properties of Bazedoxifene in HPV-positive cervical cancer cells. In vitro and in vivo experiments showed that Bazedoxifene inhibited cell invasion, migration, colony formation, and tumor growth in cervical cancer cells. We also confirmed that Bazedoxifene inhibits the GP130/STAT3 pathway and suppresses the EMT (Epithelial-mesenchymal transition) sub-signal. Thus, these data not only suggest a molecular mechanism by which the GP130/STAT3 pathway may promote cancer, but also may provide a basis for cervical cancer replacement therapy.

Combined use of cytology, p16 immunostaining and genotyping for triage of women positive for high-risk human papillomavirus at primary screening.

Human papillomavirus (HPV) testing is very sensitive for primary cervical screening but has low specificity. Triage tests that improve specificity but maintain high sensitivity are needed. Women enrolled in the experimental arm of Phase 2 of the New Technologies for Cervical Cancer randomized controlled cervical screening trial were tested for high-risk HPV (hrHPV) and referred to colposcopy if positive. hrHPV-positive women also had HPV genotyping (by polymerase chain reaction with GP5+/GP6+ primers and reverse line blotting), immunostaining for p16 overexpression and cytology. We computed sensitivity, specificity and positive predictive value (PPV) for different combinations of tests and determined potential hierarchical ordering of triage tests. A number of 1,091 HPV-positive women had valid tests for cytology, p16 and genotyping. Ninety-two of them had cervical intraepithelial neoplasia grade 2+ (CIN2+) histology and 40 of them had CIN grade 3+ (CIN3+) histology. The PPV for CIN2+ was >10% in hrHPV-positive women with positive high-grade squamous intraepithelial lesion (61.3%), positive low-grade squamous intraepithelial lesion (LSIL+) (18.3%) and positive atypical squamous cells of undetermined significance (14.8%) cytology, p16 positive (16.7%) and, hierarchically, for infections by HPV33, 16, 35, 59, 31 and 52 (in decreasing order). Referral of women positive for either p16 or LSIL+ cytology had 97.8% sensitivity for CIN2+ and women negative for both of these had a 3-year CIN3+ risk of 0.2%. Similar results were seen for women being either p16 or HPV16/33 positive. hrHPV-positive women who were negative for p16 and cytology (LSIL threshold) had a very low CIN3+ rate in the following 3 years. Recalling them after that interval and referring those positive for either test to immediate colposcopy seem to be an efficient triage strategy. The same applies to p16 and HPV16.

Stage I HPV-positive oropharyngeal cancer: Should all patients receive similar treatments?

BACKGROUND: Patients with clinical stage I human papillomavirus (HPV)-positive oropharyngeal squamous cell cancer (OPSCC) according to the American Joint Committee on Cancer (AJCC) eighth edition classification comprise a heterogeneous group formerly classified as stage I to stage IVA according to the seventh edition of the AJCC classification. These patients historically were treated with disparate treatment regimens, particularly with respect to the use of concurrent chemotherapy. METHODS: The National Cancer Data Base was queried for patients with AJCC eighth edition clinical stage I HPV-positive OPSCC (AJCC seventh edition stage T1-2N0-2bM0) who were diagnosed from 2010 to 2014 and underwent definitive radiotherapy. Concurrent chemotherapy with definitive radiotherapy was defined as chemotherapy administered within 7 days of the initiation of radiotherapy. RESULTS: The current analysis included 4473 patients with HPV-positive stage I OPSCC with a median follow-up of 36.3 months. A total of 3127 patients (69.9%) received concurrent chemotherapy. Concurrent chemotherapy was found to be associated with improved overall survival on multivariable analyses (hazard ratio [HR], 0.782; 95% CI, 0.645-0.948 [P = .012]). The effect of chemotherapy on survival varied based on lymph node involvement (P for interaction = .001). Specifically, chemotherapy was associated with improved survival for patients with lymph node-positive stage I disease (stage III-IVA according to the AJCC seventh edition: HR, 0.682; 95% CI, 0.557-0.835 [P < .001]), but not for patients with N0 disease (stage I-II according to the AJCC seventh edition: HR, 1.646; 95% CI, 1.011-2.681 [P = .05]). Similar results were noted among propensity score-matched cohorts. CONCLUSIONS: Treatment with concurrent chemotherapy was associated with improved overall survival for patients with lymph node-positive, but not lymph node-negative, AJCC eighth edition stage I HPV-positive OPSCC undergoing definitive radiotherapy, thereby supporting different treatment paradigms for these patients.

Evaluating the impact of smoking on disease-specific survival outcomes in patients with human papillomavirus-associated oropharyngeal cancer treated with transoral robotic surgery.

BACKGROUND: When treated nonsurgically with definitive chemoradiation, smokers with human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC) have a worse prognosis compared with their nonsmoking counterparts. To the authors' knowledge, the prognostic significance of smoking in surgically treated patients is unknown. METHODS: The current study is a retrospective case series of patients with HPV-positive OPSCC who underwent upfront transoral robotic surgery at a single institution from 2010 through 2017. Exclusion criteria were nonoropharyngeal primary tumors, histology other than SCC, HPV-negative tumors, previous history of head and neck cancer, and/or previous head and neck radiotherapy. Recurrence-free survival (RFS), overall survival, and disease-specific survival were compared using the Kaplan-Meier method and the log-rank test. Smoking history was categorized as never smokers (<1 pack-year), current smokers (smoking at the time of the cancer diagnosis), and former smokers. RESULTS: A total of 258 patients met the study criteria. The average age was 60 years, and approximately 87% of patients were male. A total of 148 patients (57.4%) were smokers whereas 110 (42.6%) reported never smoking. There were 44 active smokers (17.1%) and 104 former smokers (40.3%). The median follow-up was 3.23 years. There were 17 patients of disease recurrence. Smoking pack-year history was not found to be significant for RFS (hazard ratio, 1.01; 95% CI, 0.99-1.03 [P = .45]). There was no significant difference in RFS noted between never and ever smokers (92% vs 89.8%; P = .85) nor was there a difference observed between never, former, and current smokers (92% vs 91.5% vs 86.1%, respectively; P = .69). CONCLUSIONS: A smoking history is common in patients with HPV-positive OPSCC. In the current study, HPV-positive smokers were found to have excellent survival and locoregional control, similar to their nonsmoking counterparts. The results of the current study do not support the exclusion of smokers with early-stage, HPV-positive OPSCC from transoral robotic surgery-based deintensification trials.

Human papillomavirus in spermatozoa is efficiently removed by washing: a suitable approach for assisted reproduction.

RESEARCH QUESTION: Is it possible, by sperm-washing spermatozoa from clinically HPV-positive men, to obtain spermatozoa free of human papillomavirus (HPV) to be employed in assisted reproduction? DESIGN: This was an observational study performed on HPV-positive men. Freshly ejaculated semen was collected and readily processed by gradient separation followed by swim-up from the washed pellet. The resulting fractions were seminal plasma, cell pellet, round cells, non-motile spermatozoa and motile spermatozoa. All fractions were then tested for the presence of HPV DNA. RESULTS: Of the 15 clinically HPV-positive subjects, 67% were positive in at least one of the seminal fractions. If any postivity was detected, the plasma was always HPV positive. No consistent pattern was observed throughout different samples in the cell pellet, round cell and non-motile spermatozoa fractions. However, after the sperm-wash procedure, the fraction of motile spermatozoa was never found to be HPV-positive. CONCLUSIONS: The sperm-washing technique, which was previously successfully used to remove human immunodeficiency virus, can efficiently remove HPV from spermatozoa. However, the present study was conducted on a small population so a larger follow-up study is recommended. HPV screening should be performed in sperm samples and, upon HPV positivity, sperm-washing should be considered before assisted reproduction techniques are used.

[Evaluation the triage performance of p16/Ki-67 dual staining for HR-HPV positive women].

Objective: This study aimed to evaluate the clinical performance of p16/Ki-67 dual staining for triage high risk HPV (HR-HPV) infected women. Method: Target objects were women who infected HR-HPV and received colposcopy examination between April and December of 2016 at the Second Affiliated Hospital of Zhengzhou University. Gynecologists collected the cervical exfoliated cells from eligible women for p16/Ki-67 dual staining, LBC testing and HPV DNA testing. Histology diagnosis were used as gold standard. Sensitivities, specificities, positive predictive values (PPVs), negative predictive values (NPVs) of p16/Ki-67 dual staining, LBC testing and HPV16/18 testing for triage of HR-HPV positive population were calculated and compared. Results: A total of 295 HR-HPV infected women were selected, and the mean age was (44.29±11.48) years old. Positive rates of p16/Ki-67 dual staining, HPV16/18 testing and LBC testing were 70.17% (207), 56.95% (168) and 85.76% (253), respectively. When CIN2+as the endpoint, among the three triage methods, sensitivity of p16/Ki-67 dual staining was 90.00% (95%CI: 85.06%-93.43%), higher than the value of HPV 16/18 testing, but lower than the value of LBC testing. Specificity, PPV and NPV of p16/Ki-67 dual staining were the highest [71.58% (95%CI: 61.81%-79.67%), 86.96% (95%CI:81.69%-90.88%) and 77.27% (95%CI: 67.49%-84.78%)]. When detection for CIN3+, sensitivity of p16/Ki-67 dual staining was 92.90% (95%CI: 87.74%-95.99%), lower than the value of LBC testing, but higher than the value of HPV16/18 testing. Specificity of p16/Ki-67 dual staining was 55.00% (95%CI: 46.74%-63.00%), lower than the value of HPV16/18 testing, but higher than the value of LBC testing. PPV of p16/Ki-67 dual staining was 69.57% (95%CI: 62.99%-75.43%), lower than the value of HPV 16/18 testing, but higher than the value of LBC testing. NPV of p16/Ki-67 dual staining was 87.50% (95%CI: 78.99%-92.87%), higher than value of HPV 16/18 testing, but lower than the value of LBC testing. Conclusion: p16/Ki-67 dual staining has better clinical effects than HPV 16/18 testing and LBC testing for triage women with HR-HPV infection.

Targeted juglone blocks the invasion and metastasis of HPV-positive cervical cancer cells.

Human papillomaviruses (HPVs), for instance, HPV 16 and HPV 18, are concerned associated with cervical cancer. Thus, it is essential to suppress HPVs-in HPV-positive cervical cancer for treating cervical cancer. The purpose of this study was to explore the proposed molecular mechanisms, which that underlies the antintumor potential of juglone to treat of HPV-positive on cervical cancer cells. The results showed that juglone suppressed HPV-positive cell growth in a dose- and time-dependent way. In addition, cell invasion and metastasis were also inhibited by juglone. Nevertheless, when pin 1 was knocked down in HPV-positive cells, cell proliferation, invasion and metastasis were reduced. This study was designed to acquire an understanding of the mechanism of invasion and metastasis in HPV-positive cells suppressed by juglone. It provides evidence of the advantageous use of juglone in the future.

Juglone Inhibits Proliferation of HPV-Positive Cervical Cancer Cells Specifically.

Human papillomaviruses (HPVs), such as HPV 16 and HPV 18 are related to cervical cancer. Therefore, it is important to inhibit HPV-positive cervical cancer for treating cervical cancer. This study is aiming at investigating the proposed molecular mechanism, which underlies the antineoplastic potential of the aqueous extract of juglone of HPV-positive cervical cancer cells. According to the results, it is showed that, juglone prohibited HPV positive cervical cancer cells' growth through dose-dependent way. Nevertheless, when pin 1 was knocked down, the proliferation inhibition reduced. The detection of apoptosis and cell cycle also illustrated that juglone influenced HPV positive cells. Western blot expressed the influence mechanism that it affected the B-cell lymphoma 2 (Bcl-2) family and later activated the Caspase-depended apoptosis way. It is contributable for this study to understand the mechanism of inhibiting HPV positive cells by juglone and it also provides an effective strategy for the application of it in the future.

O-linked-N-acetylglucosamine transferase is associated with metastatic spread of human papillomavirus E6 and E7 oncoproteins to the lungs of mice.

High-risk human papilloma virus (HPV) 16/18 infections are often found in lung cancer. The cellular mechanisms involved in the metastatic spread of HPV-infected cervical cancer cells remain largely elusive. High O-linked-N-acetylglucosamine (O-GlcNAc) modification has also been observed in lung cancer. In the present study, we assessed the relationship between O-GlcNAc transferase (OGT) and HPV 16/18 E6/E7, or C-X-C chemokine receptor type 4 (CXCR4), in HeLa cells and in lungs of xenografted mice. Depleting OGT with an OGT-specific shRNA significantly decreased levels of E6 and E7 oncoproteins in HeLa cells and xenograft tumors, and reduced tumor formation in vivo. Western blotting and immunofluorescence analysis showed significantly decreased expression levels of E6, E7, and HCF-1 in the lungs of xenografted mice treated with an OGT-specific shRNA compared to those treated with non-targeting shRNA. Additionally, levels of E7 or OGT co-localized with Ki-67 were significantly decreased in the lungs of xenografted mice treated with OGT-specific shRNA compared to those treated with non-targeting shRNA. Moreover, levels of CXCR4 were significantly decreased in HeLa cells and in the lungs of xenografted mice treated with OGT-specific shRNA compared to those treated with non-targeting shRNA; this may be related to reduced adhesion or invasion of circulating HPV-positive tumor cells. These findings provide novel evidence that OGT functions in metastatic spread of HPV E6/E7-positive tumor cells to the lungs through E6/E7, HCF-1 and CXCR4, suggesting OGT might be a therapeutic target for HPV-positive lung cancer.