Phosphorylation of RCC1 on Serine 11 Facilitates G1/S Transition in HPV E7-Expressing Cells.

Persistent infection of high-risk human papillomavirus (HR-HPV) plays a causal role in cervical cancer. Regulator of chromosome condensation 1 (RCC1) is a critical cell cycle regulator, which undergoes a few post-translational modifications including phosphorylation. Here, we showed that serine 11 (S11) of RCC1 was phosphorylated in HPV E7-expressing cells. However, S11 phosphorylation was not up-regulated by CDK1 in E7-expressing cells; instead, the PI3K/AKT/mTOR pathway promoted S11 phosphorylation. Knockdown of AKT or inhibition of the PI3K/AKT/mTOR pathway down-regulated phosphorylation of RCC1 S11. Furthermore, S11 phosphorylation occurred throughout the cell cycle, and reached its peak during the mitosis phase. Our previous data proved that RCC1 was necessary for the G1/S cell cycle progression, and in the present study we showed that the RCC1 mutant, in which S11 was mutated to alanine (S11A) to mimic non-phosphorylation status, lost the ability to facilitate G1/S transition in E7-expressing cells. Moreover, RCC1 S11 was phosphorylated by the PI3K/AKT/mTOR pathway in HPV-positive cervical cancer SiHa and HeLa cells. We conclude that S11 of RCC1 is phosphorylated by the PI3K/AKT/mTOR pathway and phosphorylation of RCC1 S11 facilitates the abrogation of G1 checkpoint in HPV E7-expressing cells. In short, our study explores a new role of RCC1 S11 phosphorylation in cell cycle regulation.

Involvement of p53-dependent apoptosis signal in antitumor effect of Colchicine on human papilloma virus (HPV)-positive human cervical cancer cells.

Colchicine, a plant-derived alkaloid with relatively low toxicity on normal human epidermal keratinocytes (HEKn), has selective inhibitory effect on the growth of CaSki (HPV16-positive) and HeLa (HPV18-positive) human cervical cancer cell lines via the induction of apoptosis. Colchicine (2.5, 5.0 and 10.0 ng/ml) significantly reduced the expression of human papilloma virus (HPV) 16 E6/E7 mRNA and protein in CaSki and HeLa cells. Moreover, reduced expression of E6 and E7 induced by Colchicine resulted in the up-regulation of tumor suppressor proteins, p53 and Rb, as well as down-regulation of phospho Rb (pRb) protein. In addition, Bax, cytosolic cytochrome c and cleaved caspase-3 protein were increased while Bcl-2 protein was decreased significantly by 48 h of Colchicine treatment. These results implied that Colchicine could be explored as a potent candidate agent for the treatment and prevention of HPV-associated cervical cancer without deleterious effects.

miR­218 functions as a tumor suppressor gene in cervical cancer.

Previous microRNA (miR) microarray analysis revealed that miR­218 is downregulated in cervical cancer tissues. The present study aimed to further evaluate the expression of miR­218 in cervical cancer specimens, determine the association between its expression with disease progression, and investigate the roles of miR­218 in cervical cancer cells. Tissue specimens were obtained from 80 patients with cervical squamous cell carcinoma, 30 patients with high­grade cervical intraepithelial neoplasia [(CIN) II/III] and 15 patients with low­grade CIN (CINI); in addition, 60 plasma samples were obtained from patients with cervical cancer, and 15 normal cervical tissue specimens and 30 plasma samples were obtained from healthy women. These samples were used for analysis of miR­218 expression via reverse transcription­-quantitative PCR. In addition, tumor cells were transfected with miR­218 mimics, human papillomavirus (HPV)16 E6/E7 small interfering RNA, or their respective negative controls to determine the viability, colony formation, migration and invasion of cells using MTT, colony formation, wound healing and Transwell assays, respectively. Target genes of miR­218 were bioinformatically predicted and analyzed using Gene Ontology (GO) terms. The results revealed that miR­218 was downregulated in the tumor tissues and plasma of patients with cervical cancer, with expression associated with the advanced clinicopathological characteristics of patients, including HPV positivity, tumor size, blood vessel invasion and lymph node metastasis. Furthermore, miR­218 overexpression reduced tumor cell viability and xenograft growth, and suppressed tumor cell migration and invasion. HPV was detected in 75% of the 80 patients with cervical cancer, and HPV positivity was inversely associated with miR­218 expression. In addition, bioinformatics analysis predicted that roundabout guidance receptor 1 (ROBO1) was a target gene of miR­218; miR­218 overexpression significantly reduced ROBO1 levels. Furthermore, GO analysis revealed that ROBO1 was involved in regulating cell proliferation, adhesion and migration, and the cell cycle. In conclusion, the findings of the present study suggested that miR­218 may possess antitumor activities in cervical cancer.

Human papillomavirus type 18 oncoproteins exert their oncogenicity in esophageal and tongue squamous cell carcinoma cell lines distinctly.

BACKGROUND: Increasing evidence indicates an etiological role of human papillomavirus (HPV) in head and neck cancers, particularly oropharyngeal squamous cell carcinoma (OPSCC). However, the association between HPV and other cancers, including esophageal and tongue remains unclear. This study delineated the molecular characteristics of HPV18 E6 and E7 in esophageal (EC109 and EC9706) and tongue (Tca83) cancer cell lines with reference to cervical cancer (HeLa). METHODS: We analysed the HPV transcription profiles of esophageal and tongue cancer cells through Next-generation RNA sequencing, and the role of HPV18 E6 and E7 in these cells was assessed via siRNA approach, Western blotting and immunofluorescence assays. RESULTS: Overall, the HPV transcription profiles of esophageal and tongue cancer cells mimicked that of cervical cancer cells, with notable disruption of E2, and expression of E6, spliced E6 (E6*), E7, E1 and L1 transcripts. As with cervical cancer cells, p53 and its downstream transactivation target, p21, were found to be the major targets of E6 in esophageal and tongue cancer cell lines. Intriguingly, E7 preferentially targeted p130 in the two esophageal cancer cell lines, instead of pRb as in cervical cancer. Tca83 exhibited an E7 to E6 transcript ratio comparable to HeLa (cervix), targeted the ERK1/2 and MMP2 pathways, and was dependent on E6 and E7 to survive and proliferate. In contrast, both the esophageal cancer cell lines were distinct from HeLa in these aspects. CONCLUSIONS: This is the first study that delineates transcript expression and protein interaction of HPV18 E6 and E7 in esophageal and tongue cancer cell lines, suggesting that HPV plays a role in inducing these cancers, albeit via distinct pathways than those observed in cervical cancer.

HPV16-transformed human keratinocytes depend on SIX1 expression for proliferation and HPV E6/E7 gene expression.

The homeodomain transcription factor SIX1 plays a critical role in embryogenesis, is not expressed in normal adult tissue, but is expressed in many malignancies, including cervical cancer. SIX1 drives the progression of HPV16-immortalized human keratinocytes (HKc/HPV16) toward malignancy: HKc/HPV16 express high levels of SIX1 mRNA and protein; overexpression of SIX1 in HKc/HPV16 produces pre-malignant, differentiation-resistant lines (HKc/DR); SIX1 overexpression in HKc/DR induces tumorigenicity. In this paper, we explore the consequences of inhibition of SIX1 expression in premalignant HKc/DR. Only partial inhibition of SIX1 expression could be obtained in HKc/DR by RNA interference. Decreased SIX1 expression (up to 80%) in HKc/DR resulted in slower proliferation, decreased HPV16-E6/E7 mRNA levels, and increased p53 protein levels. Gene expression changes induced in HKc/DR by anti-SIX1 shRNA were indicative of mesenchymal-epithelial transition (MET) and changes in TGF-beta signaling. We conclude that HPV16-transformed cells depend on SIX1 for survival, HPV16 E6/E7 gene expression and epithelial-mesenchymal transition.

High-risk human papillomavirus oncogenes disrupt the Fanconi anemia DNA repair pathway by impairing localization and de-ubiquitination of FancD2.

Persistent expression of high-risk HPV oncogenes is necessary for the development of anogenital and oropharyngeal cancers. Here, we show that E6/E7 expressing cells are hypersensitive to DNA crosslinking agent cisplatin and have defects in repairing DNA interstrand crosslinks (ICL). Importantly, we elucidate how E6/E7 attenuate the Fanconi anemia (FA) DNA crosslink repair pathway. Though E6/E7 activated the pathway by increasing FancD2 monoubiquitination and foci formation, they inhibited the completion of the repair by multiple mechanisms. E6/E7 impaired FancD2 colocalization with double-strand breaks (DSB), which subsequently hindered the recruitment of the downstream protein Rad51 to DSB in E6 cells. Further, E6 expression caused delayed FancD2 de-ubiquitination, an important process for effective ICL repair. Delayed FancD2 de-ubiquitination was associated with the increased chromatin retention of FancD2 hindering USP1 de-ubiquitinating activity, and persistently activated ATR/CHK-1/pS565 FancI signaling. E6 mediated p53 degradation did not hamper the cell cycle specific process of FancD2 modifications but abrogated repair by disrupting FancD2 de-ubiquitination. Further, E6 reduced the expression and foci formation of Palb2, which is a repair protein downstream of FancD2. These findings uncover unique mechanisms by which HPV oncogenes contribute to genomic instability and the response to cisplatin therapies.

Extracellular overproduction of E7 oncoprotein of Iranian human papillomavirus type 16 by genetically engineered Lactococcus lactis.

BACKGROUND: We aimed at constructing Lactococcus lactis strains expressing HPV-16 recombinant E7 (rE7) oncoprotein and examining its overproduction ability followed by optimizing batch and fed-batch fermentations. Thereafter, in order to assess the immunogenicity of recombinant L. lactis cells, C57BL/6 mice were immunized by oral gavage. RESULTS: The results suggested that recombinant strains harboring optiE7 and E7 genes produced a maximum of 4.84 and 1.91 µg/mL of rE7 in static flask experiments, while the corresponding strains gave a maximum yield of 35.49 and 14.24 µg/mL in batch experiments, respectively. Fed-batch study indicated that the concentration of rE7 protein significantly increased after feeding yeast extract plus GM17 medium. The rE7 production of the best performing strains was 2.09- and 1.48-fold higher than that of the strains during the batch fermentation. Furthermore, biomass levels were 1.98- and 1.92-fold higher than those in batch cultivation. Oral immunization of C57BL/6 mice with recombinant L. lactis produced significant specific IgG and IgA antibody responses in serum and vaginal fluids, respectively. Our outcomes suggest that vaccination with L. lactis expressing rE7 can generate significant protective effects against E7-expressing cell line. Also, our study provides evidence that the presence of large amounts of E7-specific CD4+ T helper and CD8+ T cell precursors was stimulated. Significantly higher frequencies of HPV-16 E7 specific IL-2- and IFN-γ-secreting T cells were detected in antigen-stimulated splenocytes and intestinal mucosal lymphocytes, when compared to the control groups. CONCLUSIONS: We conclude that optimization of culture conditions along with recombinant protein expression can highly stimulate both specific humoral and cell-mediated immune responses in mice after oral immunization. These promising results represent a step towards fast-tracking a vaccine against HPV-16-associated cervical cancer.

Characterization of HPV integration, viral gene expression and E6E7 alternative transcripts by RNA-Seq: A descriptive study in invasive cervical cancer.

Scarce data are available on the expression of papillomavirus genome and the frequency of alternatively spliced E6E7 mRNAs in invasive cervical cancer. We carried out a comprehensive characterization of HPV expression by RNA-Seq analysis in 22 invasive cervical cancer with HPV16 or HPV18, characterizing the presence of integrated/episomal viral DNA, the integration sites in human genome and the proportion of alternative splicing products of E6 and E7 genes. The expression patterns suggested the presence of episomal and/or integrated viral DNA, with integration detected in most tumors, frequently occurring within human genes in HPV18+ and in intergenic regions in HPV16+ tumors. Alternative splicing of E6E7 transcripts showed E6*I as the most frequent isoform for both viral types, followed by E6*II and E6/E7 (unspliced) transcripts in HPV16+, and by E6/E7 in HPV18+ tumors. Previously described E6*VI and E6*V transcript isoforms for HPV16, and E6*X for HPV18, were rare or not detected.

Short half-life of HPV16 E6 and E7 mRNAs sensitizes HPV16-positive tonsillar cancer cell line HN26 to DNA-damaging drugs.

Here we show that treatment of the HPV16-positive tonsillar cancer cell line HN26 with DNA alkylating cancer drug melphalan-induced p53 and activated apoptosis. Melphalan reduced the levels of RNA polymerase II and cellular transcription factor Sp1 that were associated with HPV16 DNA. The resulting inhibition of transcription caused a rapid loss of the HPV16 early mRNAs encoding E6 and E7 as a result of their inherent instability. As a consequence of HPV16 E6 and E7 down-regulation, the DNA damage inflicted on the cells by melphalan caused induction of p53 and activation of apoptosis in the HN26 cells. The BARD1-negative phenotype of the HN26 cells may have contributed to the failure to repair DNA damage caused by melphalan, as well as to the efficient apoptosis induction. Finally, nude mice carrying the HPV16 positive tonsillar cancer cells responded better to melphalan than to cisplatin, the chemotherapeutic drug of choice for tonsillar cancer. We concluded that the short half-life of the HPV16 E6 and E7 mRNAs renders HPV16-driven tonsillar cancer cells particularly sensitive to DNA damaging agents such as melphalan since melphalan both inhibits transcription and causes DNA damage.

Adenovirus-mediated transfer of HPV 16 E6/E7 antisense RNA combined with cisplatin inhibits cellular growth and induces apoptosis in HPV-positive head and neck cancer cells.

Human papillomavirus (HPV) infection has been identified as an etiologic factor of head and neck cancers (HNCs). We explored the potential use of antisense HPV RNA transcripts for gene therapy and its effect in combination with cisplatin (CDDP) for HPV-positive HNCs. We introduced the antisense RNA transcripts of the E6 and E7 genes of HPV type 16 into UM-SCC-47 cells harboring HPV 16 and YCU-T892 cells that were HPV-negative using a recombinant adenoviral vector, Ad-E6/E7-AS. We then analyzed the effects of the introduction of Ad-E7-AS on cell and tumor growth and the synergistic effect with CDDP in vitro and in vivo. After infection of Ad-E6/E7-AS, the cellular growth of UM-SCC-47 cells were suppressed, but not that of YCU-T892 cells. E7 protein expression was suppressed, and p53 and pRb protein expression increased after infection of Ad-E7-AS. Cell growth and tumorigenicity were greatly suppressed in combination with CDDP compared with Ad-E7-AS or CDDP treatment alone in vitro. Ad-E7-AS combined with CDDP treatment significantly reduced the volumes of established subcutaneous tumors. Transfection with HPV 16 E7 antisense RNA combined with CDDP treatment might be a potentially useful approach to the therapy of HPV 16-positive HNC.

Microparticles produced by human papillomavirus type 16 E7-expressing cells impair antigen presenting cell function and the cytotoxic T cell response.

High-risk, cancer-causing human papillomaviruses (HPV) cause infections of the epidermis that may progress to cancer, including cervical cancer. Viral persistence, contributed to by viral evasion of the host immune response, is associated with the likelihood of cancer developing. Langerhans cells (LCs) are the only professional antigen presenting cells located in the epidermis, therefore may influence the antiviral immune response. Microparticles, or microvesicles, are small membrane particles shed by cells that can exert effects on other cells at both a local and systemic level. We found increased numbers of microparticles were shed from human or mouse keratinocytes expressing the HPV16 E7 oncoprotein, compared with control keratinocytes. Co-culture of LCs with microparticles from E7-expressing cells suppressed the cytotoxic T cell response. We attributed this, at least in part, to the reduction in surface of CD40 and intracellular pro-inflammatory cytokine IL-12 p40 subunit that we measured in the LCs. The evidence provided here shows that co-culture of E7-microparticles with LCs inhibits antigen-specific cytotoxicity. This is an important finding, suggesting that microparticles from HPV-infected cells could suppress the T cell response by regulating LCs, potentially contributing to persistence of HPV infection and cancer.

Expression optimization of a cell membrane-penetrating human papillomavirus type 16 therapeutic vaccine candidate in Nicotiana benthamiana.

High-risk human papillomaviruses (hr-HPVs) cause cervical cancer, the fourth most common cancer in women worldwide. A HPV-16 candidate therapeutic vaccine, LALF32-51-E7, was developed by fusing a modified E7 protein to a bacterial cell-penetrating peptide (LALF): this elicited both tumour protection and regression in pre-clinical immunization studies. In the current study, we investigated the potential for producing LALF32-51-E7 in a plant expression system by evaluating the effect of subcellular localization and usage of different expression vectors and gene silencing suppressors. The highest expression levels of LALF32-51-E7 were obtained by using a self-replicating plant expression vector and chloroplast targeting, which increased its accumulation by 27-fold compared to cytoplasmic localization. The production and extraction of LALF32-51-E7 was scaled-up and purification optimized by affinity chromatography. If further developed, this platform could potentially allow for the production of a more affordable therapeutic vaccine for HPV-16. This would be extremely relevant in the context of developing countries, where cervical cancer and other HPV-related malignancies are most prevalent, and where the population have limited or no access to preventative vaccines due to their typical high costs.

Efficient production and optimization of E7 oncoprotein from Iranian human papillomavirus type 16 in Lactococcus lactis using nisin-controlled gene expression (NICE) system.

The present work was aimed at investigating the expression and optimization of a human papillomavirus (HPV) type 16 gene encoding oncoprotein E7 in Lactococcus lactis. We genetically engineered Lactococcus lactis using nisin-controlled gene expression (NICE) system pNZ8148 to express the native and codon optimized E7 oncogenes isolated from Iranian HPV-16. The results of optimizing fermentation showed, the concentration of produced protein was expressively improved by 10 ng/mL nisin after 3.5, and 4 h induction for NZ9000 harboring the codon-optimized, and native E7 respectively. Furthermore the recombinant NZ9000 strains expressed rE7 by maximum value of 4.7 (Codon-optimized), and 1.82 µg/mL (Native) in static flask experiments at initial glucose concentrations of 50 and 75 g/L respectively. The rE7 yield was further enriched in batch fermenter experiments using controlled pH. Thus, the overall production of rE7 under optimized conditions accumulated in the cytoplasm to nearly 33.25 µg/mL by L. lactis NZ9000 containing codon-optimized E7, which was over ∼2.7-fold higher compared to the NZ9000 having native E7 strain (12.01 µg/mL). Accordingly, the maximum biomass production was calculated 4.87, and 1.51 g/L respectively.

Analysis of The Cancer Genome Atlas sequencing data reveals novel properties of the human papillomavirus 16 genome in head and neck squamous cell carcinoma.

Human papillomavirus (HPV) DNA is detected in up to 80% of oropharyngeal carcinomas (OPC) and this HPV positive disease has reached epidemic proportions. To increase our understanding of the disease, we investigated the status of the HPV16 genome in HPV-positive head and neck cancers (HNC). Raw RNA-Seq and Whole Genome Sequence data from The Cancer Genome Atlas HNC samples were analyzed to gain a full understanding of the HPV genome status for these tumors. Several remarkable and novel observations were made following this analysis. Firstly, there are three main HPV genome states in these tumors that are split relatively evenly: An episomal only state, an integrated state, and a state in which the viral genome exists as a hybrid episome with human DNA. Secondly, none of the tumors expressed high levels of E6; E6*I is the dominant variant expressed in all tumors. The most striking conclusion from this study is that around three quarters of HPV16 positive HNC contain episomal versions of the viral genome that are likely replicating in an E1-E2 dependent manner. The clinical and therapeutic implications of these observations are discussed.

Tandemly Integrated HPV16 Can Form a Brd4-Dependent Super-Enhancer-Like Element That Drives Transcription of Viral Oncogenes.

UNLABELLED: In cancer cells associated with human papillomavirus (HPV) infections, the viral genome is very often found integrated into the cellular genome. The viral oncogenes E6 and E7 are transcribed from the viral promoter, and integration events that alter transcriptional regulation of this promoter contribute to carcinogenic progression. In this study, we detected highly enriched binding of the super-enhancer markers Brd4, MED1, and H3K27ac, visible as a prominent nuclear focus by immunofluorescence, at the tandemly integrated copies of HPV16 in cells of the cervical neoplasia cell line W12 subclone 20861. Tumor cells are often addicted to super-enhancer-driven oncogenes and are particularly sensitive to disruption of transcription factor binding to the enhancers. Treatment of 20861 cells with bromodomain inhibitors displaced Brd4 from the HPV integration site, greatly decreased E6/E7 transcription, and inhibited cellular proliferation. Thus, Brd4 activates viral transcription at this integration site, and strong selection for E6/E7 expression can drive the formation of a super-enhancer-like element to promote oncogenesis. IMPORTANCE: Oncogenic human papillomaviruses play an essential role in the development of cervical cancer, and growth of these cancer cells requires continued expression of the viral E6 and E7 oncogenes. Integration of the virus into the host genome often results in deregulation of E6 and E7 expression, which provides a selective growth advantage and increases genetic instability of infected cells. We show here that tandemly integrated copies of the viral genome can form a super-enhancer-like element that drives E6/E7 transcription. Targeted disruption of factors binding to this element decreases viral transcription and causes cell death. Thus, cancer cells that harbor integrated HPV could be targeted by therapeutics that disrupt super-enhancer function.

Role of Cdc6 in re-replication in cells expressing human papillomavirus E7 oncogene.

The E7 oncoprotein of high-risk human papillomavirus (HPV) types induces DNA re-replication that contributes to carcinogenesis; however, the mechanism is not fully understood. To better understand the mechanism by which E7 induces re-replication, we investigated the expression and function of cell division cycle 6 (Cdc6) in E7-expressing cells. Cdc6 is a DNA replication initiation factor and exhibits oncogenic activities when overexpressed. We found that in E7-expressing cells, the steady-state level of Cdc6 protein was upregulated and its half-life was increased. Cdc6 was localized to the nucleus and associated with chromatin, especially upon DNA damage. Importantly, downregulation of Cdc6 reduced E7-induced re-replication. Interestingly, the level of Cdc6 phosphorylation at serine 54 (S54P) was increased in E7-expressing cells. S54P was associated with an increase in the total amount of Cdc6 and chromatin-bound Cdc6. DNA damage-enhanced upregulation and chromatin binding of Cdc6 appeared to be due to downregulation of cyclin-dependent kinase 1 (Cdk1) as Cdk1 knockdown increased Cdc6 levels. Furthermore, Cdk1 knockdown or inhibition led to re-replication. These findings shed light on the mechanism by which HPV induces genomic instability and may help identify potential targets for drug development.

Epithelium Expressing the E7 Oncoprotein of HPV16 Attracts Immune-Modulatory Dendritic Cells to the Skin and Suppresses Their Antigen-Processing Capacity.

Antigen presenting cells (APCs) in skin can promote either antigen-specific effector functions or antigen tolerance, and thus determine clearance or persistence of cutaneous viral infections. Human papillomavirus (HPV) infections can persist in squamous epithelium in immunocompetent individuals, and some persisting HPV infections, particularly with HPV16, promote malignant epithelial transformation. Here, we investigate whether local expression of the HPV16 protein most associated with malignant transformation, HPV16-E7, affects the phenotype and function of APC subsets in the skin. We demonstrate an expanded population of Langerhans cells in HPV16-E7 transgenic skin with distinct cell surface markers which express immune-modulatory enzymes and cytokines not expressed by cells from non transgenic skin. Furthermore, HPV16-E7 transgene expression in keratinocytes attracts new APC subsets to the epidermis. In vivo migration and transport of antigen to the draining lymph node by these APCs is markedly enhanced in HPV16-E7 expressing skin, whereas antigen-processing, as measured by proteolytic cleavage of DQ-OVA and activation of T cells in vivo by APCs, is significantly impaired. These data suggest that local expression of HPV16-E7 in keratinocytes can contribute to persisting infection with this oncogenic virus, by altering the phenotype and function of local APCs.

Immunoexpression of HPV 16/18 E6 and E7 oncoproteins in high-grade cervical squamous intraepithelial lesions in HIV-positive women.

The aim of this study was to assess the immunoexpression of human papillomavirus genotypes 16 and 18 (E6 and E7) oncoproteins in cervical high-grade squamous intraepithelial lesions (HSIL) of human immunodeficiency virus (HIV)-positive women. These results were also compared to the persistence and/or recurrence of lesions after loop electrosurgical excision procedure. Cervical samples from 158 patients were divided into three groups according to the presence or absence of HSIL in women who were or were not HIV-positive. By using the tissue microarray technique, immunohistochemistry was performed to analyze the expression of HPV 16/18 E6 and E7 oncoproteins. Cervical samples from 95 HIV-positive women and 63 HIV-negative women were studied. A statistically significant difference was found in the immunoexpression of E6 and E7 oncoproteins in samples from HIV-positive women with HSIL and that of women with non-neoplastic tissue (P < 0.001). There was also a statistically significant correlation between the immunoexpression of E6 (P = 0.012) and E7 (P < 0.001) oncoproteins in lesion persistence among HIV-positive women. Within the limitations of this study, the immunoexpression of HPV 16/18 E6 and E7 oncoproteins may have prognostic value regarding lesion persistence in HIV-positive women.

From HPV-positive towards HPV-driven oropharyngeal squamous cell carcinomas.

The incidence of HPV-positive oropharyngeal squamous cell carcinoma (OPSCC), which is both biologically and clinically distinct from tobacco- and alcohol-related OPSCC, is dramatically increasing. The finding that individuals with HPV-positive local/regionally advanced OPSCC have a significantly better prognosis than their negative counterparts have led to efforts to de-escalate treatment in those patients to avoid serious side effects and to improve their long-term quality of life, while maintaining treatment efficacy. Identifying diagnostic tests that are able to distinguish cancers etiologically associated with HPV is thus becoming a pressing challenge for researchers. The purpose of this review is to provide an overview of the diagnostic tools presently available to evaluate HPV status in patients with OPSCC and, in particular, to discuss their strengths and weaknesses in identifying those infections that are the real driving force in the oropharyngeal carcinogenesis process.

Detection of active human papilloma virus-16 in head and neck cancers of Asian North Indian patients.

BACKGROUND: Head and neck cancers (HNC) are one of the most common cancers in India. Human papillomavirus (HPV) has been identified as an emerging risk factor for HNC. METHODS: The present study was carried out to determine the active form of HPV-16 using a combination of PCR, viral load determination, HPV-16 E7 mRNA expression, p16, p53, and pRB immuno-histochemistry (IHC). RESULTS: A total of 226 HNC patients were enrolled in the present study. Sixty-seven (29.7%) of HNC cases were found to be HPV DNA positive. Thirty-two (14%) cases were HPV-16 DNA positive and 20 (9%) cases expressed HPV-16 E7 mRNA. HPV-16 mRNA/p16 positive cases had significantly increased viral load and integrated HPV-16 DNA. In summary, of total HNC patients, 6% cases were positive for both HPV-16 DNA and p16, and 5% were positive for both E7 mRNA and p16 IHC. We observed similar HPV-16 DNA/E7mRNA prevalence in oropharynx and oral cavity sites, however, oropharynx SCC had significantly higher viral load. CONCLUSION: Our results show low prevalence of active HPV-16 in North Indian HNC patients. HPV-16 E7 mRNA expression correlated with p16 nuclear positivity and increased viral load. Therefore, E7 mRNA expression may be used as a good surrogate indicator for active form of HPV-16 infection.