Human Papillomavirus E6 Expression Research Articles

Human Papillomavirus E7 and p16INK4a mRNA Multiplexed Quantification by a QuantiGeneTM Proof-of-Concept Assay Sensitively Detects Infection and Cervical Dysplasia Severity.

Persistent infection with human papillomavirus (HPV) can lead to cervical cancer (CxCa). During the progression to CxCa, the expression of HPV oncogenes E6 and E7 is upregulated. In turn, cellular proteins such as p16INK4a are also modulated. The combined detection of HPV oncogenes and cellular biomarkers indicative for dysplasia could be informative and convey better specificity than the current HPV tests that cannot discriminate transient infection from dysplastic changes. The QuantiGeneTM 2.0 Plex Assay platform was chosen for the effective multiplexing and quantitative detection of seven HPV-E7 mRNA targets (HPV6, 16, 18, 31, 45, 59, and 68) and the cellular mRNA of p16INK4a as a biomarker for HPV-induced transformation. Actin-beta (ACTB) and hypoxanthine-guanine phosphoribosyltransferase 1 (HPRT1) were included as reference markers. Sequences for the specific capture and detector probes were customized and developed by ThermoFisher and formulated as a QuantiGene proof-of-concept (QG-POC) plex-set. The crude lysates of the HPV-positive cervical cancer cell lines CaSki (HPV16), HeLa (HPV18), MRHI-215 (HPV45), Erin59 (HPV59), ME180 (HPV68), and the HPV-negative cell line C33A, as well as liquid-based cytology smear samples (n = 441) were analyzed. The study was a proof-of-concept evaluating the feasibility of the platform. Logistic regression and receiver operating characteristic (ROC) analyses were performed to test for the sensitivity and specificity of HPV detection and dysplastic stage discrimination. A QG-POC assay specifically and sensitively detects the HPV-E7 mRNA of seven different genotypes with an assay linearity between 20 and 13,000 cells. Cellular mRNA was detected from the crude lysates of cell lines and of cellular material from clinical liquid-based cytology smear samples. By combining HPV-E7 and p16INK4a expression normalized to ACTB, high-grade dysplasia (HCIN) and invasive cervical cancer (CxCa) were detectable, discriminable, and correlated to the biomarker expression strength. The ROC analysis from the multivariate logistic regression model including HPV-E7 and p16 INK4a resulted in an AUC of 0.74, at the optimal cut-off (sensitivity: 70.4%; specificity: 66.0%) for HCIN detection. CxCa was detected with an AUC of 0.77 (sensitivity: 81.8%, specificity: 77.4%). The QG-POC assay is sufficiently sensitive to detect and quantify HPV-E7 and cellular mRNA species. Multiplexing allows the specific detection of at least 10 analytes in a single reaction. Determining the abundance of E7 and p16INK4a transcripts when normalized to ACTB is informative about the presence of cervical dysplasia and potentially discriminates between low-grade and high-grade dysplasia and invasive cervical cancer. Further studies including more HPV genotypes and biomarkers are warranted.

Genome-Wide Profiling Reveals HPV Integration Pattern and Activated Carcinogenic Pathways in Penile Squamous Cell Carcinoma.

Simple SummaryPenile squamous cell carcinoma (PSCC) has been regarded as an HPV-related cancer for a long time. However, the integration pattern and carcinogenic pathways of HPV in PSCC remain unclear. The results of this study provide insights into the HPV-related carcinogenic mechanism in PSCC, which may be less prone to involvement in the traditional E6/E7 carcinogenic process, and are characterized by effects on the host genome, which result in the inactivation of tumor suppressors (CADM2, etc.) and the activation of oncogenes (KLF5, etc.), thus activating oncogenic signaling pathways (MAPK, JAK/STAT, etc.). This study could enhance our understanding of HPV integration and pave the way for subsequent HPV studies in PSCC.Human papillomavirus (HPV) is a significant etiologic driver of penile squamous cell carcinoma (PSCC). The integration pattern of HPV and its carcinogenic mechanism in PSCC remain largely unclear. We retrospectively reviewed 108 PSCC cases who received surgery between 2008 and 2017. Using high-throughput viral integration detection, we identified 35 HPV-integrated PSCCs. Unlike cervical cancer, the HPV E2 oncogene was not prone to involvement in integration. Eleven of the 35 (31.4%) HPV-integrated PSCCs harbored intact HPV E2; these tumors had lower HPV E6 and E7 expression and higher expression of p53 and pRb proteins than those with disrupted E2 did (p < 0.001 and p = 0.024). Integration breakpoints are preferentially distributed in or near host genes, including previously reported hotspots (KLF5, etc.) and newly identified hotspots (CADM2, etc.), which are mainly involved in oncogenic signaling pathways (MAPK, JAK/STAT, etc.). Regarding the phosphorylation levels of JNK, p38 was higher in HPV-positive tumors with MAPK-associated integration than those in HPV-positive tumors with other integration and those in HPV-negative tumors. In vitro, KLF5 knockdown inhibited proliferation and invasion of PSCC cells, while silencing CADM2 promoted migration and invasion. In conclusion, this study enhances our understanding of HPV-induced carcinogenesis in PSCC, which may not only rely on the E6/E7 oncogenes, but mat also affect the expression of critical genes and thus activate oncogenic pathways.

Direct Generation of Immortalized Erythroid Progenitor Cell Lines from Peripheral Blood Mononuclear Cells.

Reliable human erythroid progenitor cell (EPC) lines that can differentiate to the later stages of erythropoiesis are important cellular models for studying molecular mechanisms of human erythropoiesis in normal and pathological conditions. Two immortalized erythroid progenitor cells (iEPCs), HUDEP-2 and BEL-A, generated from CD34+ hematopoietic progenitors by the doxycycline (dox) inducible expression of human papillomavirus E6 and E7 (HEE) genes, are currently being used extensively to study transcriptional regulation of human erythropoiesis and identify novel therapeutic targets for red cell diseases. However, the generation of iEPCs from patients with red cell diseases is challenging as obtaining a sufficient number of CD34+ cells require bone marrow aspiration or their mobilization to peripheral blood using drugs. This study established a protocol for culturing early-stage EPCs from peripheral blood (PB) and their immortalization by expressing HEE genes. We generated two iEPCs, PBiEPC-1 and PBiEPC-2, from the peripheral blood mononuclear cells (PBMNCs) of two healthy donors. These cell lines showed stable doubling times with the properties of erythroid progenitors. PBiEPC-1 showed robust terminal differentiation with high enucleation efficiency, and it could be successfully gene manipulated by gene knockdown and knockout strategies with high efficiencies without affecting its differentiation. This protocol is suitable for generating a bank of iEPCs from patients with rare red cell genetic disorders for studying disease mechanisms and drug discovery.

E6-mediated activation of JNK drives EGFR signalling to promote proliferation and viral oncoprotein expression in cervical cancer

Human papillomaviruses (HPV) are a major cause of malignancy worldwide, contributing to ~5% of all human cancers including almost all cases of cervical cancer and a growing number of ano-genital and oral cancers. HPV-induced malignancy is primarily driven by the viral oncogenes, E6 and E7, which manipulate host cellular pathways to increase cell proliferation and enhance cell survival, ultimately predisposing infected cells to malignant transformation. Consequently, a more detailed understanding of viral-host interactions in HPV-associated disease offers the potential to identify novel therapeutic targets. Here, we identify that the c-Jun N-terminal kinase (JNK) signalling pathway is activated in cervical disease and in cervical cancer. The HPV E6 oncogene induces JNK1/2 phosphorylation in a manner that requires the E6 PDZ binding motif. We show that blockade of JNK1/2 signalling using small molecule inhibitors, or knockdown of the canonical JNK substrate c-Jun, reduces cell proliferation and induces apoptosis in cervical cancer cells. We further demonstrate that this phenotype is at least partially driven by JNK-dependent activation of EGFR signalling via increased expression of EGFR and the EGFR ligands EGF and HB-EGF. JNK/c-Jun signalling promoted the invasive potential of cervical cancer cells and was required for the expression of the epithelial to mesenchymal transition (EMT)-associated transcription factor Slug and the mesenchymal marker Vimentin. Furthermore, JNK/c-Jun signalling is required for the constitutive expression of HPV E6 and E7, which are essential for cervical cancer cell growth and survival. Together, these data demonstrate a positive feedback loop between the EGFR signalling pathway and HPV E6/E7 expression, identifying a regulatory mechanism in which HPV drives EGFR signalling to promote proliferation, survival and EMT. Thus, our study has identified a novel therapeutic target that may be beneficial for the treatment of cervical cancer.

Resveratrol inhibits the progression of cervical cancer by suppressing the transcription and expression of HPV E6 and E7 genes.

Resveratrol is a representative polyphenol of diet-derived putative cancer chemopreventive agents, which have attracted increasing interest in the cancer chemoprevention community. The inhibition of the action of human papillomavirus (HPV) E6 and E7 has been considered a key approach for cervical cancer therapy. Resveratrol has been shown to induce the apoptosis, and reduce both the viability and mitotic index of a number of cancer cell lines, including human cervical cancer cells. In the present study, it was confirmed that resveratrol inhibited the HPV E6 mRNA, HPV E6 protein and phosphorylated retinoblastoma protein (p-pRb1) levels, and increased the p53 protein levels in HeLa and Ca Ski cells, as well as in subcutaneous tumor tissue grown from HeLa cells. High-risk HPV uses a bicistronic RNA to control E6 and E7 genes simultaneously. On the whole, the present study demonstrates that resveratrol inhibits cervical cancer development by suppressing the transcription and translation of E6 and E7, and also by promoting the apoptosis and G1/S phase transition arrest. These findings may provide the basis for the development of resveratrol as a candidate for cervical cancer therapy.

Interleukin‑17A and heparanase promote angiogenesis and cell proliferation and invasion in cervical cancer.

Interleukin‑17A (IL‑17A) is a CD4 T-cell-derived pro-inflammatory cytokine that is involved in human cervical tumorigenesis. Heparanase (HPSE) is an endo-glycosidase expressed in mammals, which has been confirmed to be associated with cervical cancer invasion. In the present study, it was hypothesized that IL‑17A and HPSE are key proteins promoting tumor angiogenesis and cell proliferation and invasion in cervical cancer. The expression of IL‑17A and HPSE in cervical cancer tissues was detected by immunohistochemical staining. In addition, the expression of IL‑17A and HPSE was down- and upregulated via RNAi and human recombinant proteins, and MTT and Transwell assays were performed to examine cervical cancer cell proliferation and invasion, respectively. Flow cytometry analysis was also performed to detect cell cycle distribution, and the levels of target mRNA and protein were evaluated by reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. IL‑17A and HPSE were highly expressed in cervical cancer tissues, and microvessel density was notably higher in the IL‑17A-positive group. IL‑17A and/or HPSE recombinant protein promoted the proliferation and invasion of cervical cancer cells, increased the proportion of cells in the G2/M phase, and enhanced the mRNA and protein expression of human papillomavirus E6, P53, vascular endothelial growth factor and CD31, whereas downregulation of IL‑17A and/or HPSE exerted the opposite effects. Furthermore, downregulation of IL‑17A and/or HPSE was found to inhibit the expression of nuclear factor (NF)-κB P65. In summary, IL‑17A and HPSE may promote tumor angiogenesis and cell proliferation and invasion in cervical cancer, possibly via the NF-κB signaling pathway. These findings may lead to the identification of new diagnostic markers and therapeutic targets.

Abstract 679: ABI-231: A novel small molecule suppresses tumor growth and metastatic phenotypes of cervical cancer cells via targeting HPV E6 and E7

Abstract Objective: Cervical cancer is one of the most common and deadly cancers among women worldwide and is associated with persistent Human Papillomavirus (HPV) infection. Human papilloma virus (HPV) expressing E6 and E7 oncoproteins involved in carcinogenesis through their interactions with the p53 and pRB pathways, respectively. Therefore, non-toxic agents that have potential to inhibit the expression of E6 and E7 oncoproteins and their regulated oncogenic signaling pathways could be used in the management of cervical cancer. Microtubule targeting agents including paclitaxel, colchicine and vinca alkaloids are widely used in the treatment of various cancers but most of these agents have toxic side effects and develop chemoresistance. Herein, we investigated the potential anti-cancer effects of a novel tubulin targeting agent (ABI-231) in in vitro and in vivo model systems. Method: ABI-231 ((2-(1H-indol-3-yl)-1H-imidazol-4-yl)(3,4,5-trimethoxyphenyl))- methanone was synthesized in our home institution. Human cervical cancer cells (CaSki and SiHa) were used as in vitro model system. To determine the effect of ABI-231 on cell proliferation, migration and invasion, we performed MTS and wound healing assays. Effect of ABI-231 on the expression of HPV E6/E7 was determined by Western blot, qRT-PCR, and confocal microscopy. Xenograft study was performed to determine the effect of ABI-231 on cervical tumor growth. Results: ABI-231 treatment significantly (P&amp;lt;0.01) inhibited growth, clonogenic, invasive and migratory potential of cervical cancer cells. ABI-231 treatment was resulted in inhibition of HPV E6 and E7 expression at both mRNA and protein levels in both Caski and SiHa cells. ABI-231 inhibited phosphorylation of STAT3 at both Tyr705 and Ser727 residues. ABI-231 arrested cell cycle in G2/M phase as determined by flow cytometry and inhibited protein levels of cyclin B1, p21 and p27. Moreover, ABI-231 treatment SiHa and Caski was resulted in induction of apoptosis which was analyzed by enhance Annexin V staining. Western blot results demonstrated cleavage in PARP protein, Bid, and Bim. ABI-231 treatment also showed significant (P&amp;lt;0.01) inhibition of xenograft tumors in athymic nude mice. Excised xenograft tumors tissues were also analyzed for oncogenic signaling components which showed potent inhibition of aforementioned oncogenic signaling components compared to control. Conclusions: Taken together, our results demonstrate the potential anti-cancer efficacy of ABI-231 in in vivo and in vivo. ABI-231 can be explored as a potent therapeutic agent for the treatment of cervical cancer. Citation Format: Vivek K. Kashyap, Bilal B. Hafeez, Qinghai Wang, Neeraj Chauhan, Prashanth K. Nagesh, Nirnoy Dan, sonam kumari, Shabnam Malik, Saini Setua, Aditya Ganju, Murali M. Yallapu, Duane D. Miller, Wei Li, Meena Jaggi, Subhash C. Chauhan C. Chauhan. ABI-231: A novel small molecule suppresses tumor growth and metastatic phenotypes of cervical cancer cells via targeting HPV E6 and E7 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 679.

Inhibitory Effect of Vaginal Lactobacillus Supernatants on Cervical Cancer Cells.

Lactobacilli have been shown to inhibit the proliferation of several types of cancer cells, but the effects of vaginal Lactobacilli on cervical cancer cells have seldom been reported. We incubated Caski cells with supernatants of predominant strains in the vagina and investigated their effects on cell growth and the possible mechanisms. Cell-free supernatants of Lactobacillus crispatus, L. jensenii, and L. gasseri were prepared and purified. Caski cells were treated with various concentrations of Lactobacillus supernatants (LS). The effect of LS on cell growth was investigated using MTT assays. The influence of LS on the cell cycle and expression of human papillomavirus (HPV) E6 and E7 oncogenes was determined by flow cytometry and RT-PCR, respectively. LS-inhibited Caski cell proliferation caused morphological changes in a pH-independent manner. Flow cytometric analysis revealed that cells exposed to LS exhibited a significant increase of cell number in S phase and a strong decrease of cell number in G2/M phase. Expression of HPV E6 and E7 oncogenes, as well as CDK2 and cyclin A was decreased after treatment with LS, while expression of p21 was increased. Supernatants of L. crispatus, L. jensenii, and L. gasseri have inhibitory effects on the viability of cervical cancer cells via regulation of HPV oncogenes and cell cycle-related genes. Lactobacillus, as a promising treatment for cancer, is being assessed for its effect, and these results provide further evidence in this respect.

TAT-mediated si-hWAPL inhibits the invasion and metastasis of cervical cancer stem cells.

Human wings apart-like (hWAPL) is reported to have an association with cervical cancer. In the present study, the role of hWAPL in cervical cancer stem cells (CCSCs) was evaluated. Cervical tumorspheres were generated from cervical cancer tissues cultured in stem cell medium, and the expression of hWAPL by the tumorspheres was detected using immunohistochemistry. hWAPL expression levels in the tumorspheres were then upregulated using hWAPL adenoviral vectors or downregulated via the TAT-mediated knockdown of hWAPL and the effects on the tumorspheres were evaluated using colony formation, cell invasion and western blotting assays. The results demonstrated that the expression of hWAPL and human papillomavirus (HPV) was associated with the pluripotency of CCSCs, with hWAPL expression decreasing following the differentiation of cervical tumorspheres. Knockdown of hWAPL expression decreased HPV E6 expression and inhibited tumor invasion and colony formation. TAT-mediated knockdown of hWAPL with short interfering RNA significantly reduced tumor growth in nude mice. These results suggest that hWAPL is a marker of CCSC proliferation and is potentially a therapeutic target for cervical carcinoma through the downregulation of HPV E6.

Human papilloma virus (HPV) infection leads to the development of head and neck lesions but offers better prognosis in malignant Indian patients.

Head and neck cancers constitute a multifactorial global disease burden and are associated with human papilloma virus (HPV) as a possible risk factor. The aim of the study is to understand the relationship between HPV and the development of head and neck lesions in Indian patients. To this end, frequency of HPV was assessed in relation to different demographic and etiological features and correlated with patient survival. The prevalence of HPV significantly increased from mild dysplastic lesions (43.6%) to head and neck squamous cell carcinoma (HNSCC) stage IV (68.5%) with HPV 16 being pre-dominant in both dysplasia (43.8%) and HNSCC (61.5%). Similar trend was observed in increasing grades of the tumour. In invasive lesions, patients aged below the median age of onset showed significantly higher occurrence of HPV than those above it. Patients harbouring HPV showed a significantly better survival irrespective of age of onset. Likewise, better survival was observed in tobacco habit negative/HPV-positive patients, and as reflected in both univariate and multivariate analysis. Majority of the HPV 16-positive samples showed moderate/high nuclear expression of HPV E6 and E7 proteins in tumours and respective basal layer of adjacent normal tissues. Thus, our data indicate that frequent HPV infection, along with tobacco habit, is a pre-requisite factor for the development of HNSCC of Indian patients but offers a better survival even during tobacco usage, implicating its diagnostic and prognostic importance.

Identification and characterization of HPV-independent cervical cancers.

BackgroundHuman papillomavirus (HPV) initiates cervical cancer, and continuous expression of HPV oncogenes E6 and E7 is thought to be necessary to maintain malignant growth. Current therapies target proliferating cells, rather than specific pathways, and most experimental therapies specifically target E6/E7. We investigated the presence and expression of HPV in cervical cancer, to correlate HPV oncogene expression with clinical and molecular features of these tumors that may be relevant to new targeted therapies.ResultsWhile virtually all cervical cancers contained HPV DNA, and most expressed E6/E7 (HPV-active), a subset (8%) of HPV DNA-positive cervical cancers did not express HPV transcripts (HPV-inactive). HPV-inactive tumors occurred in older women (median 54 vs. 45 years, p = 0.02) and were associated with poorer survival (median 715 vs 3046 days, p = 0.0003). Gene expression profiles of HPV-active and -inactive tumors were distinct. HPV-active tumors expressed E2F target genes and increased AKT/MTOR signaling. HPV-inactive tumors had increased WNT/β-catenin and Sonic Hedgehog signaling. Substantial genome-wide differences in DNA methylation were observed. HPV-inactive tumors had a global decrease in DNA methylation; however, many promoter-associated CpGs were hypermethylated. Many inflammatory response genes showed promoter methylation and decreased expression. The somatic mutation landscapes were significantly different. HPV-active tumors carried few somatic mutations in driver genes, whereas HPV-inactive tumors were enriched for non-synonymous somatic mutations (p-value < 0.0000001) specifically targeting TP53, ARID, WNT, and PI3K pathways.Materials and MethodsThe Cancer Genome Atlas (TCGA) cervical cancer data were analyzed.ConclusionsMany of the gene expression changes and somatic mutations found in HPV-inactive tumors alter pathways for which targeted therapeutics are available. Treatment strategies focused on WNT, PI3K, or TP53 mutations may be effective against HPV-inactive tumors and could improve survival for these cervical cancer patients.

Anticancer and Cytotoxic Potential of Turmeric (Curcuma longa), Neem (Azadirachta indica), Tulasi (Ocimum sanctum) and Ginger (Zingiber officinale) Extracts on HeLa cell line

Anticancer and Cytotoxic Potential of Turmeric (Curcuma longa), Neem (Azadirachta indica), Tulasi (Ocimum sanctum) and Ginger (Zingiber officinale) Extracts on HeLa cell line

Transcriptional regulation of E-cadherin and oncoprotein E7 by valproic acid in HPV positive cell lines.

Valproic acid (VPA) has proven to be as one of the most promising useful drug with anticancer properties. In this study, we investigate the VPA effects on E-cadherin expression in HeLa, TC1, MKN45, and HCT116 cell lines. This study assesses the effects of VPA on human papillomavirus E7 expression in HPV positive cell lines. Cell lines were treated by 2 mmol/l VPA and expression of E-cadherin and E7 was analyzed by quantitative real-time PCR. Student's t test and ANOVA were used to determine changes in expression levels. The results revealed that mean of E-cadherin expression is increased by VPA 1.8 times in HCT116 and MKN45 cell lines, also the mean of E-cadherin mRNA levels is up-regulated 2.9 times in HeLa and TC1 cell lines. So, E-cadherin augmentation induced by VPA in HeLa and TC-1, HPV positive cell lines, is higher than HPV negative cell lines MKN45 and HCT116. The mean of HPV E7 expression is decreased by VPA, 4.6 times in in HeLa and TC-1 cell lines. This study demonstrates that re-expression of E-cadherin by VPA in HPV positive cell lines is more than HPV negative cell lines. Whereas, HPV E7 reduces the expression of E-cadherin, reduction of HPV E7 expression by VPA is related to more augmentation of E-cadherin in HPV positive cell lines. So, this study demonstrates that VPA has more anticancer properties in HPV positive cell lines, and could potentially be a promising candidate for cervical cancer treatment.

The Effect of Lactobacillus crispatus and Lactobacillus rhamnosusCulture Supernatants on Expression of Autophagy Genes and HPV E6 and E7 Oncogenes in The HeLa Cell Line.

The aim of this study was to clarify the mechanism by which lactobacilli exert their cytotoxic effects on cervical cancer cells. In addition, we aimed to evalu- ate the effect of lactobacilli on the expression of human papilloma virus (HPV) onco- genes. In this experimental study, using quantitative real-time polymer- ase chain reaction (PCR), we analyzed the expression of CASP3 and three autophagy genes [ATG14, BECN1 and alpha 2 catalytic subunit of AMPK (PRKAA2)] along with HPV18 E6 and E7 genes in HeLa cells before and after treatment with Lactobacillus crispatus and Lactobacillus rhamnosus culture supernatants. The expression of CASP3 and autophagy genes in HeLa cells was de- creased after treatment with lactobacilli culture supernatants. However, this de- crease was not significant for PRKAA2 when compared with controls. In addition, expression of HPV E6 was significantly decreased after treatment with lactobacilli culture supernatants. Lactobacilli culture supernatants can decrease expression of ATG14 and BECN1 as well as the HPV E6 oncogene. It has been demonstrated that the main changes occurring during cervical carcinogenesis in cell machinery can be reversed by suppression of HPV oncogenes. Therefore, downregulation of HPV E6 by lacto- bacilli may have therapeutic potential for cervical cancer. As the role of autophagy in cancer is complicated, further work is required to clarify the link between downregula- tion of autophagy genes and antiproliferative effects exerted by lactobacilli.

Expression of human papillomavirus E6 and E7 oncoprotein mRNA in women with low-grade squamous intraepithelial lesions or less.

We verified the prevalence of human papillomavirus (HPV) E6/E7 protein mRNA expression in patients with low-grade squamous intraepithelial lesions (LSILs) and negative cervicovaginal cytology. To investigate the relationship between mRNA expression and viral infection type, we assessed genotyping in single infections. Samples from 825 women were submitted to the E6/E7 survey. We noticed a larger percentage of E6/E7 mRNA expression in the atypical squamous cells of undetermined significance (ASC-US) and LSIL cytologies. Negative results of mRNA expression were in accordance with negative cytologies. In positive cases, the infection by a single HPV type was most common, with type 16 being most prevalent. The expression of mRNA was most prevalent in ASC-US and LSIL cytologies, compared with the negative cytology. The infection by a HPV type was more frequent in cases of positive expression, with HPV type 16 being found most frequently. Patients with LSIL cytologies had a higher percentage of multiple infections.

Human Papilloma Viruses and Breast Cancer.

Human papillomaviruses (HPV) may have a role in some breast cancers. The purpose of this study is to fill important gaps in the evidence. These gaps are: (i) confirmation of the presence of high risk for cancer HPVs in breast cancers, (ii) evidence of HPV infections in benign breast tissues prior to the development of HPV-positive breast cancer in the same patients, (iii) evidence that HPVs are biologically active and not harmless passengers in breast cancer. RNA-seq data from The Cancer Genome Atlas (TCGA) was used to identify HPV RNA sequences in breast cancers. We also conducted a retrospective cohort study based on polymerase chain reaction (PCR) analyses to identify HPVs in archival specimens from Australian women with benign breast biopsies who later developed breast cancer. To assess whether HPVs in breast cancer were biologically active, the expression of the oncogenic protein HPV E7 was assessed by immunohistochemistry (IHC). Thirty (3.5%) low-risk and 20 (2.3%) high-risk HPV types were identified in 855 breast cancers from the TCGA database. The high risk types were HPV 18 (48%), HPV 113 (24%), HPV 16 (10%), HPV 52 (10%). Data from the PCR cohort study indicated that HPV type 18 was the most common type identified in breast cancer specimens (55% of 40 breast cancer specimens) followed by HPV 16 (13%). The same HPV type was identified in both the benign and subsequent breast cancer in 15 patients. HPV E7 proteins were identified in 72% of benign breast specimens and 59% of invasive breast cancer specimens. There were four observations of particular interest: (i) confirmation by both NGS and PCR of the presence of high-risk HPV gene sequences in breast cancers, (ii) a correlation between high-risk HPV in benign breast specimens and subsequent HPV-positive breast cancer in the same patient, (iii) HPVs in breast cancer are likely to be biologically active (as shown by transcription of HPV DNA to RNA plus the expression of HPV E7 proteins), (iv) HPV oncogenic influences may occur early in the development of breast cancer.

SAHA inhibits the transcription initiation of HPV18 E6/E7 genes in HeLa cervical cancer cells

High risk human papillomavirus (HPV) is a well recognized causative agent of cervical cancer. Suberoylanilide hydroxamic acid (SAHA) is a potential anti-cervical cancer drug; however, its effect on the expression of HPV E6 and E7 genes remains unclear. Here, we show that, in SAHA treated HeLa cells, HPV18 E6 and E7 mRNA and protein levels were reduced, HPV18 promoter activity was decreased, and the association of RNP II with HPV18 promoter was diminished, suggesting that SAHA inhibited the transcription initiation of HPV18 E6 and E7 genes. In SAHA-treated HeLa, although the level of lysine 9-acetylated histone H3 in the whole cell extracts increased obviously, its enrichment on HPV18 promoter was significantly reduced which is correlated with the down-regulation of HPV E6 and E7.

Tanshinone IIA inhibits viral oncogene expression leading to apoptosis and inhibition of cervical cancer

Human papilloma virus (HPV) is the well-established etiological factor of cervical cancer. E6 and E7 oncoproteins expressed by HPV are known to inactivate tumor suppressor proteins p53 and pRb, respectively. Tanshinone IIA (Tan IIA) is a diterpenoid naphthoquinone found in the traditional Chinese medicine Danshen (Salvia sp.). Tan IIA has been shown to possess anti-tumor activity against several cancer types. In this study we show that Tan IIA potently inhibited proliferation of the human cervical cancer CaSki, SiHa, HeLa and C33a cells. Mechanistically in HPV positive CaSki cells, Tan IIA was found to (i) downregulate expression of HPV E6 and E7 genes and modulate associated proteins E6AP and E2F1, (ii) cause S phase cell cycle arrest, (iii) induce accumulation of p53 and alter expression of p53-dependent targets, (iv) modulate pRb and related proteins, and (v) cause p53-mediated apoptosis by moderating Bcl2, Bax, caspase-3, and PARP cleavage expressions. In vivo, Tan IIA resulted in over 66% reduction in tumor volume of cervical cancer xenograft in athymic nude mice. Tan IIA treated tumor tissues had lower expression of proliferation marker PCNA and changes in apoptosis targets were in agreement with in vitro studies, further confirming reduced proliferation and involvement of multiple targets behind anti-cancer effects. This is the first demonstration of Tan IIA to possess significant anti-viral activity by repressing HPV oncogenes leading to inhibition of cervical cancer. Together, our data suggest that Tan IIA can be exploited as a potent therapeutic agent for the prevention and treatment of cervical and other HPV-related cancers.

Cervical dysplasia: etiology, patogenesis, diagnosis and treatment

Cervical dysplasia is precancerous diseases and may transform into cervical cancer in 30% of cases. Development of epithelial dysplasia and cervical cancer is closely associated with the activity of the human papilloma virus. Such processes are needed for the formation of irreversible neoplasia: induction of the mechanisms of metabolic conversion of estradiol into 16α-ONE1; active expression of human papillomavirus E6 and E7 oncogenes; induction of multiple lesions in the chromosomal DNA of the infected cell; enhancement of biological processes, mediating carcinogenesis. Over time, selection of cell clones containing the integrated viral DNA is formed into a malignant tumor. Epigalin is a promise natural drug with antitumor mechanism of action for the prevention and accompanying therapy of the cervical pathology.

Inhibitors of differentiation‐1 promotes nitrosopyrrolidine‐induced transformation of HPV 16‐immortalized cervical epithelial cell

Our previous study implied a correlation between inhibitors of differentiation-1 (Id-1) and cervical cancer development. However, how Id-1 contributes to cervical carcinogenesis is unknown. In the present study, we used an in vitro transformation model to investigate the role of Id-1 in the transformation of cervical cells. Human papillomavirus (HPV)-immortalized cervical epithelial cells (H8) were successfully transformed by exposure to the carcinogen N-nitrosopyrrolidine (NPYR). The expression of both Id-1 RNA and protein was significantly increased in transformed H8 cells, suggesting a possible role of Id-1 in cervical cell transformation. Ectopic expression of Id-1 in H8 cells potentiated NPYR-induced cell transformation. In contrast, silencing of Id-1 suppressed NPYR-induced H8 cell transformation. In addition, the expression of HPV E6 and E7 oncoproteins was upregulated while that of the tumor suppressors p53 and pRb was suppressed after H8 cell transformation. Our results suggest that Id-1 plays an oncogenic role in HPV-related cervical carcinogenesis, which sheds light on cervical cancer development mechanisms and implies that Id-1 is a potential target for cervical cancer prevention and therapy.