Human Papillomaviruses Disease Research Articles

Human papillomavirus 16 replication converts SAMHD1 into a homologous recombination factor and promotes its recruitment to replicating viral DNA.

Human papillomaviruses (HPVs) are causative agents in around 5% of all human cancers. The development of anti-viral therapeutics depends upon an increased understanding of the viral life cycle. Here, we demonstrate that HPV16 replication converts sterile alpha motif and histidine-aspartic domain HD-containing protein 1 (SAMHD1) into a homologous recombination (HR) factor via phosphorylation. This phosphorylation promotes recruitment of SAMHD1 to viral DNA to assist with replication. A SAMHD1 mutant that mimics phosphorylation is hyper-recruited to viral DNA and attenuates viral replication. Expression of this mutant in HPV16-immortalized cells attenuates the growth of these cells, but not cells immortalized by the viral oncogenes E6/E7 alone. Finally, we demonstrate that the phosphatase inhibitor endothall promotes hyper-recruitment of endogenous SAMHD1 to HPV16 replicating DNA and can attenuate the growth of both HPV16-immortalized human foreskin keratinocytes (HFKs) and HPV16-positive head and neck cancer cell lines. We propose that phosphatase inhibitors represent a novel tool for combating HPV infections and disease.

A human papillomavirus 16 E2-TopBP1 dependent SIRT1-p300 acetylation switch regulates mitotic viral and human protein levels and activates the DNA damage response.

Human papillomaviruses (HPVs) are causative agents in around 5% of all human cancers. While there are prophylactic vaccines that will significantly alleviate HPV disease burden on future generations, there are currently no anti-viral strategies available for the treatment of HPV cancers. To generate such reagents, we must understand more about the HPV life cycle, and in particular about viral-host interactions. Here, we describe a novel mitotic complex generated by the HPV16 E2 protein interacting with the host protein TopBP1 that controls the function of the deacetylase SIRT1. The E2-TopBP1 interaction disrupts SIRT1 function during mitosis in order to enhance acetylation and stability of viral and host proteins. We also demonstrate that the E2-TopBP1 interaction activates the DDR. This novel complex is essential for the HPV16 life cycle and represents a novel anti-viral therapeutic target.

96. Human Papilloma Viruses Associated Diseases in a Cohort of Patients with Idiopathic CD4 Lymphopenia

BackgroundIdiopathic CD4 Lymphocytopenia (ICL) is a rare immunodeficiency characterized by an absolute CD4+ T count of < 300 cells/µL, in absence of HIV-infection or any other known cause. Patients with ICL have an increased risk of opportunistic infections. The prevalence, natural history, and spectrum of Human Papillomaviruses (HPV) associated diseases in ICL patients are unknown.MethodsICL patients were enrolled in a prospective observational study (N = 90). Demographic, clinical, and immunologic data were analyzed by nonparametric Methods. Immunophenotyping was performed by flow cytometry.ResultsThe median age of ICL patients was 48 years, 47% were women, and 92% were Caucasian. Sixty-five percent of patients had at least one opportunistic infection, with HPV being the most prevalent (34.4%), followed by cryptococcal disease (22%), shingles (15.5%), molluscum contagiosum (8.8%), Histoplasma capsulatum (4.4%), Mycobacterium avium complex (4.4%), and progressive multifocal encephalopathy (2.2%). HPV-related diseases were identified in 18 women and 13 men. ICL patients with HPV disease were younger compared with those without (median age 34 vs. 53.5 years, P < 0.0001). Nine (29%) had anogenital, 9 (29%) had a cutaneous disease (verruca plana, verrucous carcinoma, squamous cell carcinoma) while 13 (42%) had both anogenital and cutaneous disease. Patients with HPV-related disease were also more likely to have history of cryptococcal disease, shingles or molluscum (P = 0.036, P = 0.22 and 0.11, respectively). Thirteen patients had HPV-associated cancers: 7 both mucosal and skin and 3 either skin or mucosal malignancies. Patients with HPV-disease had lower CD4+ T cells (median CD4 70 vs. 114 cells/µL, P = 0.036). No differences were observed in the numbers of CD8+ T cells, B cells, NK cells, and levels of IgG between patients with and without HPV disease.ConclusionHPV-related disease represents the most common opportunistic infection in ICL patients. Patients with ICL and HPV disease are younger, have lower CD4s and high prevalence of HPV-associated malignancies. Therefore, for patients presenting early in life with severe HPV disease further immunological workup should be considered and for patients with ICL excessive screening for HPV-related malignancies should be a priority.Disclosures All Authors: No reported Disclosures.

Eurogin 2016 Roadmap: how HPV knowledge is changing screening practice.

Human papillomaviruses (HPVs) are the necessary cause of most cervical cancers, a large proportion of other anogenital cancers, and a subset of oropharyngeal cancers. The knowledge about HPV has led to development of novel HPV-based prevention strategies with important impact on clinical and public health practice. Two complementary reviews have been prepared following the 2015 Eurogin Conference to evaluate how knowledge about HPV is changing practice in HPV infection and disease control through vaccination and screening. This review focuses on screening for cervical and anal cancers in increasingly vaccinated populations. The introduction of HPV vaccines a decade ago has led to reductions in HPV infections and early cancer precursors in countries with wide vaccination coverage. Despite the high efficacy of HPV vaccines, cervical cancer screening will remain important for many decades. Many healthcare systems are considering switching to primary HPV screening, which has higher sensitivity for cervical precancers and allows extending screening intervals. We describe different approaches to implementing HPV-based screening efforts in different healthcare systems with a focus in high-income countries. While the population prevalence for other anogenital cancers is too low for population-based screening, anal cancer incidence is very high in HIV-infected men who have sex with men, warranting consideration of early detection approaches. We summarize the current evidence on HPV-based prevention of anal cancers and highlight important evidence gaps.

HPV disease transmission protection and control.

Human papillomaviruses (HPVs) represent a large collection of viral types associated with significant clinical disease of cutaneous and mucosal epithelium. HPV-associated cancers are found in anogenital and oral mucosa, and at various cutaneous sites. Papillomaviruses are highly species and tissue restricted, and these viruses display both mucosotropic, cutaneotropic or dual tropism for epithelial tissues. A subset of HPV types, predominantly mucosal, are also oncogenic and cancers with these HPV types account for more than 200,000 deaths world-wide. Host control of HPV infections requires both innate and adaptive immunity, but the viruses have developed strategies to escape immune detection. Viral proteins can disrupt both innate pathogen-sensing pathways and T-cell based recognition and subsequent destruction of infected tissues. Current treatments to manage HPV infections include mostly ablative strategies in which recurrences are common and only active disease is treated. Although much is known about the papillomavirus life cycle, viral protein functions, and immune responsiveness, we still lack knowledge in a number of key areas of PV biology including tissue tropism, site-specific cancer progression, codon usage profiles, and what are the best strategies to mount an effective immune response to the carcinogenic stages of PV disease. In this review, disease transmission, protection and control are discussed together with questions related to areas in PV biology that will continue to provide productive opportunities of discovery and to further our understanding of this diverse set of human viral pathogens.

Immunotherapy in new pre-clinical models of HPV-associated oral cancers

Cervical, anal, penile and a sub-set of head and neck (HN) tumors are critical health problems caused by high risk Human Papilloma Viruses (HPVs), like HPV type 16. No specific/effective pharmacological treatments exist. A valid preventive vaccination as well as the immunotherapy of persistent infections, pre-cancerous lesions or early-stage cancers could drive the HPV disease burden down. These treatments might be featured through low-cost platforms like those based on DNA and plant biotechnologies to produce tailored and enhanced formulations taking profit from the use of plants as bio-factories and as a source of immune-stimulators. Finally, and regardless of the formulation type, pre-clinical tests and models are crucial to foresee efficacy of immunotherapy before clinical trials. In this study, we created an orthotopic mouse model for HPV-related oral tumors, a subset of HN tumors for which no models have been generated before. The model was obtained by inducing the stable expression of the HPV16 E7 protein into the mouse oral squamous cell carcinoma (OSCC) AT-84 (AT-84 E7). The AT-84 E7 cells were injected into the mouth pavement of C3H mice via an extra-oral route to obtain orthotopic tumors. The model turned out to mimic the natural history of the human HPV oral cancer. From AT-84 E7, through engineering to express luciferase, the bioluminescent AT-84 E7-Luc cells were obtained for a fast and easy monitoring by imaging. The AT-84 E7 and the AT-84 E7-Luc tumors were used to test the efficacy of E7-based therapeutic vaccines that we had previously generated and that had been already proven to be active in mice against non-orthotopic E7-expressing tumors (TC-1 cells). In particular, we used genetic and plant-derived formulations based on attenuated HPV16 E7 variants either fused to plant virus genes with immunological activity or produced by tobacco plants. Mice were monitored by imaging allowing to test the size reduction of the mouth implanted experimental tumors in function of the different regimens used. The proposed tumor model is easy to handle and to reproduce and it is efficacious in monitoring immunotherapy. Furthermore, it is expected to be more predictive of clinical outcome of therapeutic vaccines than non-orthotopic models that are currently used. Finally, imaging offers unique opportunities to predict formulation efficacy through measuring tumor growth in vivo.

Abstract IA23: Cervical tissue T cells in HPV disease.

Abstract Despite the availability of preventative vaccines, human papillomaviruses (HPVs) cause more cancers than any other virus. Like other viral pathogens that cause human cancers, most HPV infections do not cause overt harm, let alone symptoms. Indeed, in immune-competent persons, most genital infections are eventually cleared. Nonetheless, the incidence of HPV-associated cancers in anatomic sites for which no screening algorithms have been validated is rising steadily. Preinvasive, intraepithelial lesions are associated with integration of the HPV genome into the host genome, and subsequent expression of the viral oncoproteins, E6 and E7, which are required for disease initiation and persistence. These true tumor-specific antigens provide non-“self” targets for immune therapies. The relatively indolent clinical course of preinvasive lesions, and tissue tropism of HPV infections provide opportunities to identify and treat early stage disease. In cervical HPV disease, a subset of high grade dysplastic lesions, cervical intraepithelial neoplasia grade 3 (CIN3), do in fact undergo complete regression, which is presumably immunologically mediated. However, T cell responses to viral antigens, measured in the blood, have not been informative with respect to immune responses in the cervical mucosa. Our team has been studying strategies to enhance the quality and magnitude of immune responses to HPV antigens. Ongoing clinical trials testing therapeutic vaccination and strategies to manipulate the lesion microenvironment to enhance homing and access of immune effector cells will be discussed. The design of these trials has been informed by studies of tissue-resident T cells in normal cervical mucosa, and by changes identified in dysplastic mucosa, both in lesions that regress, and in those that do not, in the course of a defined, prospective study window. Changes in the composition and distribution of tissue T cells postvaccination will be presented. Strategies to modify the stromal compartment of HPV lesions to promote lesion clearance will also be discussed. Citation Format: C.L. Trimble, J. Teague, T.C. Wu, R.A. Clark. Cervical tissue T cells in HPV disease. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology: Multidisciplinary Science Driving Basic and Clinical Advances; Dec 2-5, 2012; Miami, FL. Philadelphia (PA): AACR; Cancer Res 2013;73(1 Suppl):Abstract nr IA23.

The Fanconi Anemia Pathway Limits Human Papillomavirus Replication

High-risk human papillomaviruses (HPVs) deregulate epidermal differentiation and cause anogenital and head and neck squamous cell carcinomas (SCCs). The E7 gene is considered the predominant viral oncogene and drives proliferation and genome instability. While the implementation of routine screens has greatly reduced the incidence of cervical cancers which are almost exclusively HPV positive, the proportion of HPV-positive head and neck SCCs is on the rise. High levels of HPV oncogene expression and genome load are linked to disease progression, but genetic risk factors that regulate oncogene abundance and/or genome amplification remain poorly understood. Fanconi anemia (FA) is a genome instability syndrome characterized at least in part by extreme susceptibility to SCCs. FA results from mutations in one of 15 genes in the FA pathway, whose protein products assemble in the nucleus and play important roles in DNA damage repair. We report here that loss of FA pathway components FANCA and FANCD2 stimulates E7 protein accumulation in human keratinocytes and causes increased epithelial proliferation and basal cell layer expansion in the HPV-positive epidermis. Additionally, FANCD2 loss stimulates HPV genome amplification in differentiating cells, demonstrating that the intact FA pathway functions to restrict the HPV life cycle. These findings raise the possibility that FA genes suppress HPV infection and disease and suggest possible mechanism(s) for reported associations of HPV with an FA cohort in Brazil and for allelic variation of FA genes with HPV persistence in the general population.

Genital human papillomavirus infections: current and prospective therapies

Infection with human papillomaviruses (HPVs) is very common and associated with benign and malignant epithelial proliferations of skin and internal squamous mucosae. A subset of the mucosal HPVs are oncogenic and associated with 5 % of all cancers in men and women. There are two licensed prophylactic vaccines, both target HPV 16 and 18, the two most pathogenic, oncogenic types and one, additionally, targets HPV 6 and 11 the cause of genital warts. The approach of deliberate immunization with oncogenic HPV E6 and/or E7 proteins and the generation of antigen-specific cytotoxic T-cells as an immunotherapy for HPV-associated cancer and their high-grade pre-cancers has been tested with a wide array of potential vaccine delivery systems in Phase I/II trials with varying success. Understanding local viral and tumour immune evasion strategies is a prerequisite for the rational design of therapeutic vaccines for HPV-associated infection and disease, progress in this is discussed. There are no antiviral drugs for the treatment of HPV infection and disease. Current therapies are not targeted antiviral therapies, but either attempt physical removal of the lesion or induce inflammation and a bystander immune response. There has been recent progress in the identification and characterization of molecular targets for small molecule antagonists of the HPV proteins E1, E2 and E6 or their interactions with their cellular targets. Lead compounds that could disrupt E1-E2 protein-protein interactions have been discovered as have inhibitors of E6-E6-AP-binding interactions. Some of these compounds showed nanomolar affinities and high specificities and demonstrate the feasibility of this approach for HPV infections. These studies are, however, at an early phase and it is unlikely that any specific anti-HPV chemotherapeutic will be in the clinic within the next 10-20 years.

The humoral response to Gardasil over four years as defined by Total IgG and competitive Luminex immunoassay

Safe and effective vaccines against anogenital human papillomaviruses (HPV) are now available. These vaccines, composed of virus-like particles (VLPs) made from the L1 major capsid protein of specific HPV types, induce a polyclonal antibody response directed against specific conformational and linear epitopes displayed on the VLP. Numerous studies indicated the importance of neutralizing antibodies in protection from infection. However, our understanding of the antibody responses to these vaccines is not complete, and there is no established immune correlate of protection nor antibody threshold that correlates with protection against HPV infection or disease. In the current study, antibody responses of young women to Gardasil®, the quadrivalent HPV 6, 11, 16 and 18 L1 VLP vaccine (qHPV), were assessed through 48 months (M) in total IgG and competitive Luminex immunoassays (total IgG LIA and cLIA). The total IgG LIA was developed as a research assay to evaluate preclinical multivalent HPV VLP vaccine formulations. The cLIA simultaneously evaluates the antibody response to a unique conformational, neutralizing epitope on each of the four HPV types present in the quadrivalent vaccine; HPV 6, 11, 16 and 18. The same sera from women vaccinated with the qHPV vaccine were tested in both the total IgG LIA and the cLIA assays. The proportion of vaccinated women achieving seropositivity and the anti-HPV VLP total IgG and cLIA geometric mean titers (GMTs) were summarized at M7, M24, M48 based on the serostatus cut-points defined for each immunoassay. Overall, greater than 99% of subjects seroconverted to all four vaccine types in both assays; GMTs peaked at M7. For all four HPV types, regardless of the immunoassay used, the most significant decline in GMTs was observed between M7 and M24. By M24, the antibody titers had reached a plateau and minimal declines in antibody titers were observed between M24 and M48 for all four HPV types in both immunoassays. Testing the same sera, seropositivity for M48 HPV18 remained high (96.7%) in the total IgG LIA, but was 64.8% in the cLIA. The current study illustrates potential important differences in serologic assays utilized in the clinical trials of the two currently available HPV VLP vaccines (quadrivalent and bivalent). Differences in seropositivity status are attributed to the measurement parameters and sensitivity of the individual immunoassays and do not indicate reduced anti-HPV18 protective antibodies.

Indian women with higher serum concentrations of folate and vitamin B12 are significantly less likely to be infected with carcinogenic or high-risk (HR) types of human papillomaviruses (HPVs)

Studies conducted in the USA have demonstrated that micronutrients such as folate and vitamin B12 play a significant role in modifying the natural history of high-risk human papillomaviruses (HR-HPVs), the causative agent for developing invasive cervical cancer (CC) and its precursor lesions. The purpose of the current study was to investigate whether these micronutrients have similar effects on HR-HPV infections in Indian women. The associations between serum concentrations of folate and vitamin B12 and HR-HPV infections were evaluated in 724 women who participated in a CC screening study in the southern state of Andhra Pradesh, India. Serum folate and vitamin B12 concentrations were measured by using a competitive radio-binding assay. Digene hybrid capture 2 (HC2) assay results were used to categorize women into two groups, positive or negative for HR-HPVs. Unconditional logistic regression models specified a binary indicator of HC2 (positive/negative) as the dependent variable and serum folate concentrations combined with serum vitamin B12 concentrations as the independent predictor of primary interest. Models were fitted, adjusting for age, education, marital status, parity, type of fuel used for cooking and smoking status. Women with higher concentrations of serum folate (>6 ng/mL) and vitamin B12 (>356 pg/mL) were at lower risk of being positive for HR-HPVs compared to those with serum folate ≤6 ng/mL and serum vitamin B12 ≤ 356 pg/mL (odds ratio = 0.26; 95% confidence interval: 0.08-0.89; P = 0.03). These results demonstrated that improving folate and vitamin B12 status in Indian women may have a beneficial impact on the prevention of CC. Micronutrient based interventions for control of HR-HPV infections may represent feasible alternatives to vaccine based approaches to HPV disease prevention, which are currently unaffordable for use in resource limited areas in rural India.

Cervical Cancer and Human Papillomaviruses: Inactivation of Retinoblastoma and Other Tumor Suppressor Pathways

Infection with human papillomaviruses (HPVs) is a major public health burden worldwide and is associated with benign and malignant lesions of the skin and genital tract. HPV causes cervical cancer, which represents the second most prevalent cancer in women worldwide. Functions of the viral oncogenes E6 and E7 are essential for carcinogenesis and for support of the viral life cycle. We will begin by discussing the relationship between HPV infection and disease, followed by a review of E6 and E7 activities and their respective cellular targets. Particular emphasis will be placed on established and newly discovered mechanisms by which E7 inhibits members of the cellular retinoblastoma protein family. We will then describe how current research links the above molecular interactions to malignant transformation as well as to aspects of the viral life cycle in vitro and in vivo. As a result of decades of intense HPV research, promising therapies to prevent infection and to treat HPV associated cancers are now on the horizon. We will conclude our review by a description of potential gene therapeutic and hormonal approaches and of new developments in the design of effective vaccines.

Human papillomavirus virus-like particles are efficient oral immunogens when coadministered with Escherichia coli heat-labile enterotoxin mutant R192G or CpG DNA.

Certain human papillomaviruses (HPVs) cause most cervical cancer, which remains a significant source of morbidity and mortality among women worldwide. HPV recombinant virus-like particles (VLPs) are promising vaccine candidates for controlling anogenital HPV disease and are now being evaluated as a parenteral vaccine modality in human subjects. Vaccines formulated for injection generally are more costly, more difficult to administer, and less acceptable to recipients than are mucosally administered vaccines. Since oral delivery represents an attractive alternative to parenteral injection for large-scale human vaccination, the oral immunogenicity of HPV type 11 (HPV-11) VLPs in mice was previously investigated; it was found that a modest systemic neutralizing antibody response was induced (R. C. Rose, C. Lane, S. Wilson, J. A. Suzich, E. Rybicki, and A. L. Williamson, Vaccine 17:2129-2135, 1999). Here we examine whether VLPs of other genotypes may also be immunogenic when administered orally and whether mucosal adjuvants can be used to enhance VLP oral immunogenicity. We show that HPV-16 and HPV-18 VLPs are immunogenic when administered orally and that oral coadministration of these antigens with Escherichia coli heat-labile enterotoxin (LT) mutant R192G (LT R192G) or CpG DNA can significantly improve anti-VLP humoral responses in peripheral blood and in genital mucosal secretions. Our results also suggest that LT R192G may be superior to CpG DNA in this ability. These findings support the concept of oral immunization against anogenital HPV disease and suggest that clinical studies involving this approach may be warranted.

Association of human papillomavirus infection and disease with magnitude of human immunodeficiency virus type 1 (HIV-1) RNA plasma level among women with HIV-1 infection.

Ninety-three women with human immunodeficiency virus type 1 (HIV-1) infection were enrolled in a cross-sectional study to evaluate the relationship between plasma HIV-1 RNA levels and coincident cervical infection and disease caused by human papillomaviruses (HPVs). HIV-1 RNA plasma levels of >10,000 copies/mL were highly associated with the presence in cervical specimens of HPV DNA of oncogenic (high risk) virus genotypes (P=.006; relative risk, 2.57). In addition, similar HIV-1 RNA plasma levels were associated with abnormal Pap smears (P=.01; relative risk, 2.11). In this study, 81% of women with high-risk HPV cervical infection had abnormal Pap smears. Measurement of HIV-1 RNA plasma levels may help to identify a subgroup of HIV-1-infected women at increased risk for cervical HPV infection and disease.