An overview of human papillomaviruses and current vaccine strategies

Cervical cancer is one of the most common cancers in women worldwide, particularly in developing countries. The viral origin of cervical cancer has been proven beyond any reasonable doubt. Persistent infection with certain subsets of human papillomaviruses is recognized as a necessary cause for the development of cervical cancer. Persistence of oncogenic HPVs, immunodeficiency, high HPV viral load and cofactors like smoking, multiple sex partners and poor nutrition predispose to cervical cancer. Prophylactic vaccines using HPV virus-like particles containing capsid protein L1 have shown protection against disease in animals and are currently undergoing clinical trials. Therapeutic vaccines using HPV E6 and E7 proteins are also being investigated for their ability to remove residual infection.     

Preventative and therapeutic vaccines for cervical cancer

"High-risk" genotypes of the human papillomavirus (HPV), most commonly HPV genotype 16, are the primary etiologic agents of cervical cancer. Indeed HPV DNA is detected in 99% of cervical carcinomas. Thus, cervical cancer and other HPV-associated malignancies might be prevented or treated by the induction of the appropriate viral-antigen-specific immune responses. Transmission of papillomavirus may be prevented by the generation of antibodies to capsid proteins L1 and L2 that neutralize viral infection. HPV L1 virus-like particles (VLPs) show great promise as prophylactic HPV vaccines in ongoing clinical trials but L2-based preventative vaccines have yet to be tested in patients. Since the capsid proteins are not expressed at detectable levels by infected basal keratinocytes or in HPV-transformed cells, therapeutic vaccines generally target the nonstructural early viral antigens. Two HPV oncogenic proteins, E6 and E7, are critical to the induction and maintenance of cellular transformation and are co-expressed in the majority of HPV-containing carcinomas. Although other early viral antigens show promise for vaccination against papillomas, therapeutic vaccines targeting E6 and E7 may provide the best opportunity to control HPV-associated malignancies. Various candidate therapeutic HPV vaccines are currently being tested whereby E6 and/or E7 are administered in live vectors, as peptides or proteins, in nucleic acid form, as components of chimeric VLPs, or in cell-based vaccines. Encouraging results from experimental vaccination systems in animal models have led to several prophylactic and therapeutic vaccine clinical trials. Should this new generation of HPV preventative and therapeutic vaccines function in patients as demonstrated in animal models, oncogenic HPV infection and its associated malignancies could be controlled by vaccination. Importantly, recent advances in HPV detection and continued improvements in screening further enhance our opportunities to systematically eradicate HPV-associated malignancy.     

Human papillomavirus vaccines for the treatment of cervical cancer

Human papillomavirus (HPV) infection is a major cause of cervical cancer, the second most common cancer in women worldwide. Currently, a HPV L1-based virus-like particle has been approved as a prophylactic vaccine against HPV infection, which will probably lead to a reduction in cervical cancer incidence within a few decades. Therapeutic vaccines, however, are expected to have an impact on cervical cancer or its precursor lesions, by taking advantage of the fact that the regulatory proteins (E6 and E7) of HPV are expressed constantly in HPV-associated cervical cancer cells. Vaccine types targeting these regulatory proteins include the recombinant protein and DNA vaccines, peptide vaccines, dendritic-cell vaccines, and viral and bacterial vector deliveries of vaccines, and these may provide an opportunity to control cervical cancer. Further approaches incorporating these vaccine types with either conventional therapy modalities or the modulation of CD4(+) regulatory T cells appear to be more promising in achieving increased therapeutic efficacy. In this review, we summarize current and future therapeutic vaccine strategies against HPV-associated malignancies at the animal and clinical levels.     

Vaccination to prevent and treat cervical cancer

Human papillomaviruses (HPVs) are the primary etiologic agents of cervical cancer. Thus, cervical cancer and other HPV-associated malignancies might be prevented or treated by HPV vaccines. Transmission of papillomavirus may be prevented by the generation of antibodies to capsid proteins L1 and L2 that neutralize viral infection. However, because the capsid proteins are not expressed at detectable levels by infected basal keratinocytes or in HPV-transformed cells, therapeutic vaccines generally target nonstructural early viral antigens. Two HPV oncogenic proteins, E6 and E7, are critical to the induction and maintenance of cellular transformation and are coexpressed in the majority of HPV-containing carcinomas. Thus, therapeutic vaccines targeting E6 and E7 may provide the best option for controlling HPV-associated malignancies. Various candidate therapeutic HPV vaccines are currently being tested whereby E6 and/or E7 are administered in live vectors, as peptides or protein, in nucleic acid form, as components of chimeric virus-like particles, or in cell-based vaccines. Encouraging results from experimental vaccination systems in animal models have led to several prophylactic and therapeutic vaccine clinical trials. If these preventive and therapeutic HPV vaccines prove successful in patients, as they have in animal models, then oncogenic HPV infection and its associated malignancies may be controllable by vaccination.     

Developing HPV virus-like particle vaccines to prevent cervical cancer: a progress report.

BACKGROUND: the knowledge that sexually transmitted infection with one of a limited number of human papillomaviruses (HPVs) is a central cause of almost all cervical cancers affords the opportunity to prevent this common cancer through anti-viral vaccination. OBJECTIVE: the spectacular success of vaccines in preventing several other viral diseases offers hope that immunoprophylaxis against the relevant HPVs could lead to a major reduction in cervical cancer incidence. RESULTS AND CONCLUSION: the results of preclinical studies and early phase clinical trials of virus-like particle (VLP) based subunit vaccines have been very encouraging. However, unique aspects of papillomavirus biology and genital tract infections, and the lack of sexual a transmission model for papillomavirus, make it far from certain that effective prophylactic vaccination against genital HPV infection will be easily achieved. Future clinical efficacy trials will likely test the hypothesis that parenteral injection of VLPs can induce antibody mediated and type specific protection against genital tract HPV infection and subsequent development of premalignant neoplastic disease.     

Molecular mechanisms of cervical carcinogenesis by high‐risk human papillomaviruses: novel functions of E6 and E7 oncoproteins

Over the last two decades, since the initial discovery of human papillomavirus (HPV) type 16 and 18 DNAs in cervical cancers by Dr. Harald zur Hausen (winner of the Nobel Prize in Physiology or Medicine, 2008), the HPVs have been well characterised as causative agents for cervical cancer. Viral DNA from a specific group of HPVs can be detected in at least 90% of all cervical cancers and two viral genes, E6 and E7, are invariably expressed in HPV‐positive cervical cancer cells. Their gene products are known to inactivate the major tumour suppressors, p53 and retinoblastoma protein (pRB), respectively. In addition, one function of E6 is to activate telomerase, and E6 and E7 cooperate to effectively immortalise human primary epithelial cells. Though expression of E6 and E7 is itself not sufficient for cancer development, it seems to be either directly or indirectly involved in every stage of multi‐step carcinogenesis. Epidemiological and biological studies suggest the potential efficacy of prophylactic vaccines to prevent genital HPV infection as an anti‐cancer strategy. However, given the widespread nature of HPV infection and unresolved issues about the duration and type specificity of the currently available HPV vaccines, it is crucial that molecular details of the natural history of HPV infection as well as the biological activities of the viral oncoproteins be elucidated in order to provide the basis for development of new therapeutic strategies against HPV‐associated malignancies. This review highlights novel functions of E6 and E7 as well as the molecular mechanisms of HPV‐induced carcinogenesis. Copyright © 2009 John Wiley & Sons, Ltd.     

Recombinant vaccines for the prevention of human papillomavirus infection and cervical cancer

Carcinogenic human papillomaviruses (HPVs) that cause cervical cancer preferentially infect basal, metaplastic squamous cells of the transformation zone. If infection persists, and a vegetative infection ensues, a premalignant lesion may develop with the potential to progress into an invasive squamous cell carcinoma. Papillomavirus prophylactic vaccines target the systemic immune system for induction of neutralizing antibodies that protect the basal cells against infection. Because the carcinogenic HPVs are susceptible to neutralization by antibodies for 9–48 h after reaching the basal cells, both low and high titered HPV type-specific antibodies induced by HPV L1 and L2-based vaccines are highly efficacious. The greatest burden of HPV-associated cancers occurs in poor areas of the world where women do not have access to routine gynecological care. The burden of HIV/AIDS in these same regions of the world has added to the burden of HPV-associated disease. There is an urgent need for a cost-effective, broad-spectrum HPV prophylactic vaccine in developing countries, which necessitates substantial cost subsidization of the virus-like particle (VLP) based vaccines licensed in industrialized countries or an alternative approach with second-generation vaccines that are specifically designed for delivery to women in resource-poor communities.     

International standard reagents for HPV detection

Human papillomavirus is the commonest genital viral infection in healthy sexually active subjects, and the presence of chronic or persistent HPV types in genital cells may constitute a prognostic marker of underlying, or predict future HPV-associated diseases. A variety of novel tests for detecting the presence of oncogenic HPV types in biological specimens have been reported. These are based on the various stages of infection and viral life cycle. HPV infects squamous epithelium with expression of various gene products intimately linked to epithelial cell differentiation. Hence, there are basically three classes of detectable markers directly derived from HPVs: molecular markers based on detection of nucleic acid sequences, serological markers based on detection of antibodies against viral proteins, and cellular markers based on detection of proteins expressed intracellularly, upon either infection or carcinogenesis. The nature of various assays and the development of international standard reagents for qualitative and quantitative assessment of assay performance are outlined. There is an increasing demand to develop standard tools to assess the quality of HPV detection systems, for regulatory and clinical management purposes. International standard reagents for HPV will help defining the analytical sensitivity and specificity of various detection methods, and will allow assuring that laboratory services used to evaluate disease burden, HPV vaccines, and cancer prevention strategies are accurate and comparable worldwide. The advancement of prophylactic vaccine candidates against HPV infections and related diseases stresses the increasing importance of HPV assays in monitoring the impact of HPV vaccination on disease burden.     

HPV-vaccination against cervical carcinoma: will it really work?

Prophylactic HPV vaccination provides an opportunity to profoundly affect cervical cancer incidence worldwide. The quadrivalent HPV VLP 6, 11, 16, 18 vaccine (Gardasil) and the bivalent HPV VLP 16, 18 vaccine (Cervarix) are effective for prevention of HPV infection and cervical precancerous lesions. The quadrivalent vaccine is also effective for prevention of vulvar and vaginal lesions and genital warts. With the introduction of the vaccines general issues have to be raised such as optimal age for vaccination, duration of protection after vaccination, impact on cervical cancer screening, vaccination of males and feasibility of application to developing countries. The prospects of a vaccine which will protect against the most common viral sexually transmitted infection and thereby, protect against the complications of HPV infection such as cervical cancer is extremely appealing. The success of HPV vaccination as a major public health prevention opportunity, however, will entirely depend on efficient infrastructures to deliver the vaccines and on the acceptance by individuals, parents and health care providers.     

Analyses of human papillomavirus genotypes and viral loads in anogenital warts

Condylomata acuminata (genital warts) are the most common sexually transmitted viral diseases. These lesions are caused by infection with mucosal human papillomaviruses (HPVs). However, there is limited information on HPV strain distribution involved in the molecular pathogenesis of these lesions. To address this, the strain prevalence and the frequency of multiple HPV infections were determined in wart tissue obtained from 31 patients attending a wart clinic. These lesions were bisected and subjected to parallel DNA and mRNA extractions. HPV-type prevalence and incidence of multiple infections were determined by the Roche Linear Array assay. qPCR compared HPV 6, 11, 16, and 18 viral loads and RT-qPCR measured HPV 6 and 11 E6 genomic expression levels. Seventy-one percent of these samples were infected with multiple HPVs. Only one sample was negative for HPV 6 or 11 DNA. Forty-eight percent of samples were positive for a high risk (oncogenic) HPV. The results show that multiple infections in tissue are frequent and the subsequent analysis of HPV 6 and 11 E6 DNA viral loads suggested that other HPVs could be causing lesions. Further analysis of HPV 6/11 E6 mRNA levels showed that there was no discernable relationship between HPV 6 E6 DNA viral load and relative HPV 6 or 11 E6 mRNA levels thereby questioning the relevance of viral load to lesion causality.     

Human papillomavirus update with a particular focus on cervical disease

Human papillomavirus (HPV) is a common viral infection of squamous epithelial tissues, but its importance has only recently been recognised by the medical community. HPVs are now realised to consist of many genotypes and are associated with a diverse spectrum of clinical manifestations. Within the genital tract, some diseases have been recognised since antiquity; for example, genital warts which are caused by HPV types distinct from those causing genital cancer. However, others (such as cervical cancer), although recognised centuries ago as linked to sexual activity, have only been associated with oncogenic HPVs relatively recently, with the tools of molecular biology. We now understand that genital HPV infections are the most common sexually transmitted viral infections, are largely transient, asymptomatic and of no consequence. This virus manifests as more than just benign warts. Chronic carriage of with oncogenic genotypes (over years and in a minority of patients), together with other cofactors (host and/or exogenous) in complex pathways not totally understood, result in severe dysplasia or, ultimately, carcinogenesis. As it takes time for precursor lesions to develop and there are effective screening programmes for their detection and treatment, HPV-related neoplastic disease of the cervix is largely a preventable reproductive health issue of women. Yet, on a global scale, cervical cancer is the second most common cancer of women, with the majority of cases occurring in developing countries. Although HPV is noncultivatable by traditional diagnostic virological methods, successfully applied molecular biology techniques have underpinned development of vaccines which are now in phase II/III clinical trials. Successful vaccination ultimately has the greatest potential to impact upon the global burden of disease from genital HPV infection. However, the outcome from reduction in incidence of dysplasia and neoplasia will take years to eventuate; consequently, various cervical cancer prevention strategies still need to be endorsed and maintained in the meantime.     

HPV-assoziierte Plattenepithelkarzinogenese

About 7-8% of all human cancers are thought to be related to infections with high-risk (HR) human papilloma virus (HPV). Besides cervical cancer, especially squamous cell carcinomas of the anogenital and oropharyngeal regions are associated with HR-HPV. Transmission of HPV is due to sexual activity. Harald zur Hausen was awarded in 2008 with the Nobel price in medicine for the establishment of a causal link between certain HPV infections and cervical cancer. Meanwhile potent prophylactic vaccines are available to prevent infections with HPV-16 and HPV-18, the two most frequently observed HR HPV types worldwide. On molecular grounds a persistent HPV infection is the central risk factor for the development of HPV-associated neoplasias. Continued expression of the viral E6 and E7 oncogenes disrupts cell cycle control mechanisms in infected cells, thereby gaining limitless proliferative capacity and resistance against apoptotic signals. However acquisition of mutations and genomic instability might cause malignant transformation in these cells.     

Development of a Topical Protein Therapeutic for Human Papillomavirus and Associated Cancers

Human papillomaviruses (HPVs) are the causative agents of several disease states, including genital warts and cervical cancer. There are around 500 million cases of genital warts per annum worldwide and around 450,000 cases of cervical cancer. Although HPV vaccines should eventually reduce the incidence of these diseases, new and effective treatments are still urgently required. The E2 (early) proteins from some HPV types induce growth arrest and apoptosis, and these proteins could be used as therapeutics for HPV-induced disease. A major obstacle to this approach concerns the delivery of the protein to HPV-transformed cells and/or HPV-infected cells in vivo. One possible solution is to use recombinant viruses to deliver E2. Another possible solution is to use purified E2 proteins or E2 fusion proteins. The herpes simplex virus VP22 protein is one of a small number of proteins that have been shown to cross the cell membrane with high efficiency. VP22-E2 fusion proteins produced in bacterial cells are able to enter mammalian cells and induce apoptosis. This suggests that VP22-E2 fusion proteins could be topically applied as a treatment for HPV-induced diseases, most probably post-surgery. In this review, we discuss this and other approaches to the topical delivery of selective therapeutic agents against HPV-associated conditions.     

The prevalence of HPV-18 and variants of E6 gene isolated from cervical cancer patients in Taiwan.

BACKGROUND: Cervical cancer is the second most common cancer in women worldwide. It has been considered that human papillomavirus (HPV) is associated with cervical cancer. Currently, more than 80 different serotypes of HPV have been characterized and they are divided into low- and high-risk groups. The most common types that lead to cervical cancer are HPV-16 and -18. The viral oncogenes E6 and E7 are associated with the development of cervical cancer. In previous study, the variants of HPV-16 E6 gene have been reported. It suggests that variants may influence the morbidity of carcinogenesis, but the variant study on HPV-18 remains unknown. OBJECTIVES: To identify the variants of integrated HPV-18 E6 gene in the prevalent infection of HPV-18 of cervical cancer patients. STUDY DESIGN: 25 cervical cancer patients were clinically identified and the biopsies were obtained. The infectious HPV types were identified by PCR and Southern blotting analysis. The DNA fragments of the integrated HPV-18 E6 were amplified by PCR and cloned. The nucleotide sequences were obtained by sequencing. RESULTS: The prevalence of HPV infection in our 25 cases was HPV-18 (100%) and 7 out of these 25 cases (28%) were co-infected with HPV-16. The most dominant mutation among 25 tested patients was a silence mutation C183G of the E6 coding region. CONCLUSIONS: The prevalent HPV infectious serotype is HPV-18, which differs from the worldwide prevalent type. The identified HPV-18 E6 variants had a unique silence mutation located on C183G in E6 coding region.     

HPV vaccine: an overview of immune response,clinical protection,and new approaches for the future

Although long-term protection is a key-point in evaluating HPV-vaccine over time, there is currently inadequate information on the duration of HPV vaccine-induced immunity and on the mechanisms related to the activation of immune-memory. Longer-term surveillance in a vaccinated population is needed to identify waning immunity, evaluating any requirements for booster immunizations to assess vaccine efficacy against HPV-diseases. Current prophylactic vaccines have the primary end-points to protect against HPV-16 and 18, the genotypes more associated to cervical cancer worldwide. Nevertheless, data from many countries demonstrate the presence, at significant levels, of HPVs that are not included in the currently available vaccine preparations, indicating that these vaccines could be less effective in a particular area of the world. The development of vaccines covering a larger number of HPVs presents the most complex challenge for the future. Therefore, long term immunization and cross-protection of HPV vaccines will be discussed in light of new approaches for the future.     

Aetiology, pathogenesis, and pathology of cervical neoplasia.

Early epidemiological studies of cervical neoplasia suggested a causal relation with sexual activity and human papillomaviruses (HPVs) have emerged as prime suspects as venerally transmitted carcinogens. HPVs fall into two broad camps: low risk types, associated with cervical condylomas and CIN 1; and high risk types (mostly 16 and 18), found in 50-80% of CIN 2 and CIN 3 lesions, and 90% of cancers. This association with cancer is very strong, with odds ratios of > 15 (often much higher) in case-control studies that are methodologically sound. An infrequently detected third group of intermediate risk type HPVs is associated with all grades of CIN and occasionally with cancers. HPVs have also been detected in a wide range of asymptomatic controls, indicating that other events are required for development of neoplasia such as viral persistence and/or altered expression of viral genes, often following integration of the viral genome. This leaves the two major viral oncogenes, E6 and E7, directly coupled to viral enhancers and promoters, allowing their continued expression after integration. High risk HPV E7 proteins bind and inactivate the Rb protein, whereas E6 proteins bind p53 and direct its rapid degradation. A range of putative cofactors has been implicated in progression: HLA type, immunosuppression, sex steroid hormones, and smoking; most of these cofactors appear to influence progression to CIN 3. The natural history includes progression to CIN 3 in 10% of CIN 1 and 20% of CIN 2 cases, whereas at least 12% of CIN 3 cases progress to invasive carcinoma. Cervical glandular intraepithelial neoplasia (CGIN) often coexists with squamous CIN, and the premalignant potential of high grade CGIN is not in doubt, but the natural history of low grade CGIN remains uncertain. A high proportion of CGIN lesions and adenocarcinomas are HPV positive, and HPV18 has been implicated more in glandular than in squamous lesions. A strong clinical case for the application of HPV typing of cells recovered from cervical scrapes can be made; however, a rigorous cost-benefit analysis of introducing HPV typing into the cervical screening programme is required. Prophylactic and therapeutic HPV vaccines are under development. This article reviews the aetiology, pathogenesis, and pathology of cervical neoplasia, emphasising the role of HPVs.     

Human papillomavirus: current status and issues of vaccination

An association between human papillomavirus (HPV) infection and the development of cervical cancer was initially suggested over 30 years ago, and today there is clear evidence that certain subtypes of HPV are the causative agents of such malignancies. Papillomaviruses make up a vast family that comprises hundreds of different viruses. These viruses infect epithelia in humans and animals and cause benign hyperproliferative lesions, commonly called warts or papillomas, which can occasionally progress to squamous cell cancer. HPV infections are considered the most common among sexually transmitted diseases. One of the most prevalent cancer types induced by HPV (mostly types 16 and 18) is cervical cancer. Vaccination is the most effective means of preventing this infectious disease. These prophylactic vaccines, based on virus-like particles (VLPs), are extremely effective in providing protection from infection in almost 100 % of cases. VLP vaccines of HPV are subunit vaccines consisting only of the major viral capsid protein of HPV. There are two types of vaccine available: bivalent vaccine (against HPV-16/18) and quadrivalent vaccine (against HPV-6/11/16/18). Second-generation prophylactic HPV vaccines, currently in clinical trials, may hold several merits over the current bivalent and quadrivalent vaccines, such as protection against additional oncogenic HPV types, less dependence on cold-chain storage and distribution, and non-invasive methods of delivery.     

Prerequisites for human papillomavirus vaccine trial: results of feasibility studies.

BACKGROUND AND AIMS: Oncogenic human papillomaviruses (HPVs) are the major cause of cervical cancer and associated cancers. First generation preventive vaccines against HPVs are entering clinical trials. Therefore, it is time to consider prerequisites of field trials in Finland. RESULTS: Incidence of cervical cancer is increasing in young women which is not unexpected since risk taking behavior among young women has also increased. In the developed countries up to 44% of cervical cancer cases are attributable to HPV16 infection alone. However, high risk HPV types other than HPV16 and HPV18 are emerging in the population based on HPV DNA pilot screening studies. Annual attack rates among young women less than 25 years of age is 2.3% for HPV16 infection, and 0.03% for CIN3 invasive cervic cancer. Thus, sample size estimates for HPV vaccine efficacy trial are approximately 1000 when the endpoint is HPV16 infection, and approximately 15000 when the endpoint is >/=CIN3 or worse assuming that the vaccine efficacy is 70%. Both HPV vaccine trial acceptability and compliance rates at routine visits of the general Finnish female population are going to be high based on a pilot study. CONCLUSION: Prerequisites for large scale field trials on HPV vaccination are fulfilled in Finland.     

Progress in prophylactic and therapeutic vaccines for human papillomavirus infection

Virus-like particle (VLP) subunit vaccines composed of the major capsid protein L1 of the genital human papillomaviruses (HPVs) are now in Phase III clinical trials. The vaccines are immunogenic and safe and early results indicate efficacy. VLPs induce strong cell-mediated as well as humoral immune responses and chimeric VLPs including an HPV early protein may have therapeutic potential. Polynucleotide and recombinant viral vaccines encoding nonstructural viral proteins show therapeutic and prophylactic efficacy in animal models and are candidate immunotherapies for established low-grade benign genital infections. Vaccines designed to elicit cytotoxic T-lymphocytes specific for the HPV oncoproteins E6 and E7 show immunogenicity and efficacy in transplantable tumor models in rodents. In Phase I and II trials these vaccines are immunogenic and safe but show limited efficacy.