Joshua W.Wang, Chein-fu Hung,Warner K. Huh, Cornelia L. Trimble And Richard B.S. Roden 2016-02-29 03:26:08
Abstract Persistent infection by one of 15 high-risk human papillomavirus (hrHPV) types is a necessary but not sufficient cause of 5% of all human cancers. This provides a remarkable opportunity for cancer prevention via immunization. Since Harald zur Hausen's pioneering identification of hrHPV types 16 and 18, found in approximately 50% and 20% of cervical cancers, respectively, two prophylactic HPV vaccines containing virus-like particles (VLP) of each genotype have been widely licensed. These vaccines are beginning to affect infection and HPV-associated neoplasia rates after immunization campaigns in adolescents. Here, we review recent progress and opportunities to better preventHPV-associated cancers, including broadening immune protection to cover all hrHPV types, reducing the cost of HPV vaccines especially for developing countries that have the highest rates of cervical cancer, and immune-based treatment of established HPV infections. Screening based upon George Papanicolaou's cervical cytology testing, and more recently detection of hrHPVDNA/RNA, followed by ablative treatment of high-grade cervical intraepithelial neoplasia (CIN2/3) have substantially reduced cervical cancer rates, and we examine their interplay with immune-based modalities for the prevention and eventual elimination of cervical cancer and other HPV-related malignancies. Cancer Prev Res; 8(2); 95–104. 2014 AACR. Introduction Cervical cancer remains the third leading cause of cancer in women worldwide and virtually all cases can be attributed to infection via one of 15 high-risk HPVs (hrHPV; ref. 1). Importantly, significant fractions of other anogenital malignancies (vaginal, vulval, anal, and penile), as well as oropharyngeal and oral cancers (2), are also caused by HPV infection, predominantly by the HPV16 genotype. Infections with hrHPV and their associated neoplasia remain highly prevalent and although there are effective screening strategies available for detection of HPV infection at the cervix, importantly, HPV screening strategies are not routinely applied to other anatomic sites. Because 16% to 18% of cancer cases globally are caused by infections annually (3) and preventive vaccination against infectious agents ranks among the most cost-effective of medical interventions, the concept of "cancer immunoprevention" offers enormous promise to improve human health. Indeed, this promise is being realized with the introduction of preventive vaccines against Hepatitis B (HBV) vaccine and Human Papillomavirus (HPV) vaccines, and complements both cervical cytology screening efforts and chemoprevention via drug treatments for Helicobacter pylori, Hepatitis C, and HIV (1). Although prevention of liver cancers linked to HBVvia vaccinationhas already been demonstrated(4–6), it is too early to see the impact of HPV vaccination on cancer incidence although HPV disease rates are clearly dropping (7). Unfortunately, HPV vaccination uptake has been slow in some developed and many developing countries. The current high cost of the vaccine and need for administration to adolescents have hindered widespread introduction. Motivation for uptake among the general population has also been complicated by perceived concerns for safety, potential for increased promiscuity, lack of efficacy and need, but with continued scientific reporting of the benefits and safety of HPV vaccination, it is hopeful that these concerns will be resolved and implementation will improve. In this review, we discuss the potential of immunization for both prevention and treatment of HPV disease as primary and secondary cancer prevention strategies. Because of space limitations, it is not possible to cover these areas comprehensively and our intent is only to provide a concise summary of the most recent advances of over a century of progress (Fig. 1) that address immune prevention of HPV malignancies and how they could interface with current cervical cancer prevention efforts. Etiology of HPV and the Early Successes of HPV Vaccination More than 100 different HPV genotypes have been fully sequenced and can be generally divided into cutaneous and mucosal types. Although most infections are benign, those caused by a subset (approximately 15) of the mucosal HPV types can progress to malignancy and are considered "high risk" (hrHPV). The hrHPV are the primary etiologic agent of almost all cervical cancers (8–12). HPV16 (13) and HPV18 (14) are the most studied HRtypes as they cause approximately 70%of all cervical cancers (15). More recently, the link between mucosal HPV and cancers has been expanded to certain subsets of anal, vaginal, vulval, penile, and or opharyngeal cancers (9). HPV16 is also the cause of approximately 90% of HPVassociated cancers at these noncervical sites (9). The currently licensed HPV vaccines, Gardasil (Merck & Co.) And Cervarix (GlaxoSmithKline), are based on the major capsid protein L1, which has self-assembled into noninfectious virus-like particles (VLP; refs. 16, 17) and both target HPV16 and HPV18. While the mucosal hrHPV types have received the most attention, the "low risk" mucosal HPV types also produce disease with considerable morbidity, including recalcitrant anogenital warts, and life-threatening laryngeal papillomas (e.g., HPV6 and 11). Gardasil also prevents infection and disease associated with the two most prevalent types in benign genital warts (90%), HPV6 and HPV11. Randomized controlled trials for both Gardasil (FUTURE trials; refs. 18–22) and Cervarix (PATRICIA and the Costa Rica HPV vaccine trial, CVT; refs. 23–25) examined three immunizations in young women. The vaccines demonstrated high immunogenicity, excellent safety profiles and showed efficacy in preventing incident vaccine-related HPV infection as well as incident persistent infection related to vaccine HPV types. Several countries have adopted national HPV immunization programs, which have begun to bear fruit. For example, in 2007 Australia was one of the first countries to adopt such a campaign with Gardasil and a significant decline (<1% of women versus 10.5% before introduction of vaccines in 2006) in genital wart diagnoses in women (26, 27), as well as reduced cervical abnormalities in teenage girls (28) has been reported. Significant declines in the reporting of genital warts in men were also observed although this finding was attributed to herd-immunity rather than direct vaccination (29). Improving Access to Current Vaccines When the HPV vaccine was first introduced, the duration of immunity was also unknown. Therefore, pre-teens were considered the optimal population for immunization given the importance of vaccination before sexual debut (7). The targeting of 9- to 26-year-old patients, especially young adolescents, however, has complicated vaccination because it is infants that traditionally receive the majority of vaccinations and hence, compliance toward three doses has been an outstanding issue. In light of this, it is reasonable to consider exploring the safety and immunogenicity of HPV vaccines in infants. Alternatively, coadministration of current HPV vaccines with other childhood combination vaccines against multiple infectious agents should also be considered, as this potentially reduces the costs of vaccine administration. Indeed, studies suggest that, upon administration of an HPV vaccine with a second vaccine against another infectious agent, the antibody responses elicited by each are non-inferior to when either vaccine is administered alone (30–33). The cost of the two commercial vaccines has also limited vaccine uptake, especially in the developing world where >80% of cervical cancer cases occur. The vaccines were introduced at $120 dose, i.e., $360 total, although recent GAVI pricing of $4.50/dose for developing countries has been negotiated and is hopefully sustainable (34). On a national scale, the costs of HPV vaccination provide a significant barrier to mass vaccination programs especially as they are born in addition to ongoing cytologic screening programs. It is clearly important to understand how completion of an HPV vaccination regimen affects a patient's need for cytologic and/or HPV screening. Presently, because the vaccines target only two of 15 oncogenic types, cervical cytology screening continues. However, because HPV vaccination reduces the incidence of high-grade CIN, the predictive value and cost effectiveness of cytologic screening will drop in this population. This issue might potentially be addressed by implementing screening via HPV testing and increasing the screening interval. Several studies are examining whether fewer doses (2 doses versus 3) can be administered with acceptable protective efficacy and duration (35–41). Although more independent studies are required to ensure sustained efficacy and examine cross-protection of related types, the data thus far are supportive of a two-dose regimen. Such studies have also been examined by health authorities such as the WHOSAGE (Strategic Advisory Group of Expertson immunization) and the overall conclusions were that a two-dose prime-boost schedule within an interval of 6months is noninferior to the standard three-dose schedule (42). However, it is important to note that there were some findings suggesting poorer clinical efficacy with two doses that were attributed to a failure in controlling the intervals between the prime and booster immunization. WHO SAGE recommends adolescents within 9 and 13 years range for the two-dose regimen and that the interval between the prime and booster shot needs 6 months or else a third boost will be required. As a result of this, there is now considerable interest in the efficacy of a single-dose vaccination. Remarkably, in one recent study, a subset of patients that received only one dose of Cervarix still exhibited detectable antibody responses after 4 years and no evidence of breakthrough infection (43). While these findings must be interpreted cautiously, they suggest that a trial to specifically address the efficacy of a single dose may be warranted. Protection against More Genotypes The current HPV vaccines are not approved for protection against the nonvaccine hr HPV types that can also cause cervical cancer and indeed are currently responsible for approximately 30%of all cases (44) . Further analysis from both preclinical and the HPV vaccine trials showed that cross-protection against hr HPV types not directly targeted by the current HPV vaccines, is generally partial, limited to a few genotypes, and of unclear duration (45–47). Therefore, it remains an important goal to extend protection to all hr HPV types without substantially driving up the cost of immunization. Broad protection is especially important for immune-compromised individuals who suffer more disease associated with HPV types beyond those targeted by the current vaccines. HIVþ patients have much higher rates of multi-type infections and an increased risk for HPV-associated cancers (e.g., anal cancer in men; ref. 48) despite the introduction of the HAART therapy.Linkage studies have also shown a 2- to 22-fold increase in incidence of cervical cancer in HIVþ women compared with HIV+ women (49). Although B-cell responses are somewhat compromised in HIVþ patients and solid organ transplant recipients (SOTR), vaccination studies in other infectious diseases show they are still capable of generating effective neutralizing antibody responses (50–52). In light of this, several trials are ongoing to evaluate if this is also true for the current HPV vaccines (reviewed in ref. 52)). In one recently completed trial (53), all HIV+ women (HAART naive) were seropositive for HPV16 and 18 antibodies following vaccination with the bivalent vaccine although titers were overall lower than the healthy control group. The safety profile was also consistent with clinical experience with healthy women. It is worth emphasizing, however, that certain meta-analysis and population studies indicate thatHPV16 infection is underrepresented in HIVþ patients, suggesting that these populations acquire more distinct HPV genotypes or they experience more multi-type infections. Although this further complicates current HPV vaccine uptake as well as policy making decision as not all types are targeted by the current HPV vaccines, the findings also support the efforts tomake amore broadly protective HPV vaccine. An interesting avenue for exploration is targeting of the epidermodysplasia verruciformis (EV)–associated approximately 30 cutaneous types that have been proposed as a cofactor along with UV radiation in the development of nonmelanoma skin cancers (NMSC; refs. 54, 55). EV is a rare inherited condition that predisposes patients to widespread cutaneous warts and squamous cell carcinoma predominantly caused by HPV5 and HPV8. Both HIV and SOTRs have a higher risk of developing warts, keratotic skin lesions (e.g., actinic keratosis), and NMSC in association with EV type HPV infections (56, 57). However, the etiologic link toNMSC remains controversial and there are concerns over the timing of infection, which appears to occur throughout the lifespan. Recent studies in the mouse system nevertheless are encouraging and vaccination offers an approach to test for an etiologic role for EV type HPV in NMSC in these populations. Taken together, it is now clear that the field of HPV malignancy prevention via vaccination needs to address more types of HPV (i.e., not just HPV16 and 18). Second-Generation HPV Vaccines The need for broader protection against cancer-related HPV has spurred the efforts of many laboratories to develop next-generation vaccine candidates. Merck & Co., the manufacturers of Gardasil, has created a nonavalent vaccine (V503, targeting the seven most common oncogenic HPV types in cervical cancer and genital wart types HPV6 and 11), which has completed advanced phase III clinical trials (NCT00543543, NCT00943722, and NCT01651949). Preliminary results in a three-dose vaccine regimen report that the immune responses (with respect toHPV6, 11, 16, and 18) from V503 are noninferior to Gardasil and it was recently approved by the FDA (www.fda.gov/NewsEvents/Newsroom/PressAnnouncements/ucm426485.htm) (58). In addition, the nonavalent vaccine prevents approximately 97% of high-grade cervical, vulvar, and vaginal diseases caused by five additional oncogenic types and the vaccine is called Gardasil 9 (59). However, the cost of manufacturing this vaccine (and hence its subsequent pricing) is not expected to be cheaper than the current vaccines, and this price (estimated at $162/dose) likely remain as a limitation toward global implementation. In light of this, several groups have attempted to simplify the manufacturing process with alternative L1-VLP eukaryotic expression systems such as tobacco plants, yeast, or modified insect cells that secrete the VLPs, which together with local low-cost manufacturing of a generic product may drive down costs (reviewed in ref. 60). Delivery of L1 VLP via live recombinant vectors such a S. typhi, measles virus, or adenovirus also have great potential because this potentially offers simpler manufacturing, no need for needles/syringes, and potentially fewer doses, but enthusiasm is tempered by potential safety concerns with a live vaccine and preexisting immunity to the vector. In bacterial production systems, VLPs are not readily formed; rather, the expression of L1 proteins results in VLP subunits known as pentamers or capsomeres (61, 62). The neutralizing epitopes of VLP are preserved in capsomeres, and vaccination with capsomeres also induces strong neutralizing antibody responses in animals although lower than for L1-VLP (63). However, the use of an appropriate adjuvant with capsomeres can achieve neutralizing titers similar to VLPs (64). Vaccination with L1-expression vectors is another potential approach but efficient in vivo delivery and avoiding interference remain significant hurdles. Given the importance of broad immunity covering all hrHPV, several research groups, including ours, have investigated the HPV minor capsid protein L2 as a candidate antigen for second generation HPV vaccines. Studies with L2 have shown that the N-terminus contains several highly conserved protective epitopes and is required for several events during the infectious life cycle (reviewed in ref. 65). Importantly, L2 vaccination induces antibodies that can cross neutralize a large diverse range of HPV genotypes both in vitro and in vivo (66–69). Moreover, because L2's epitopes are linear, they can be readily expressed in E. coli. Unfortunately, coexpression of L1/L2 VLPs results in an L1- response only suggesting that L2 is immune subdominant in the context of a HPV virion (70, 71). To further complicate matters, vaccination of L2 alone although broadly neutralizing is still not as immunogenic as the L1-VLPs even with the use of a potent adjuvant. To boost immunogenicity, several groups have attempted several methods, including the concatenation of multiple L2 epitopes (72), using scaffolds (73), and alternative display methods of L2 on VLP platforms of papillomavirus (74, 75) or other viruses (76). Although some of these strategies have yielded higher immune responses, the overall responses were still below those of the current VLP, and it is not known if immune titers will be as long lasting as the current vaccine. The apparently weak response to L2 may reflect in part the use of an in vitro neutralization assay with poor sensitivity to L2-specific protective antibodies. Nevertheless, it appears that low titers of neutralizing serum antibody are sufficient to confer robust protection in preclinical animal models, suggesting that it may not be necessary to achieve similar titers as L1 VLP for durable immunity. Therapeutic HPV Vaccines and New Immunologic Considerations Because HPV is a very common sexually transmitted infection, there remains an urgent need for a therapy to effectively treat existing chronic HPV infections and disease (Fig. 2). No therapeutic activity has been demonstrated for the licensed HPV vaccines likely reflecting the absence of detectable L1 expression in HPV-transformed tumor cells or basal keratinocytes that harbor the infection. HPV viral oncoproteins E6 and E7 are required for the induction and maintenance of cellular transformation (77), and are consistently and specifically coexpressed in all infected cells, including HPV-associated cancers (78, 79). Although the targeting of other viral antigens like E1, E2, and E5 with vaccination is effective for therapy in animal models of disease, it is likely that tumor cells could escape immune responses by loss or downregulation of E1, E2, and E5 expression, but this is not possible for E6 or E7. Therefore, most therapeutic HPV vaccines being tested clinically target HPV E6 and/or E7. It is clear from natural history studies that most immune competent persons eventually clear HPV infections and lowgrade intraepithelial lesions, such that the virus becomes undetectable and the cervical histopathology returns to normal (Fig.2) . In contrast, the rate of spontaneous clearance is much lower in immune-compromised patients, suggesting that infection elicits a delayed but eventually effective anti-HPV immune response in the majority of patients with an intact immune system (80, 81). In fact, a subset of high-grade intraepithelial lesions does undergo complete regression (80), which is presumably immunologically mediated. Although immunotherapies should aim to enhance such antiviral immunity in those unable to clear HPV naturally, to date, no algorithms exist that can distinguish persons at risk for either high-grade dysplasias or invasive disease. Therefore, the standard of care for high-grade intraepithelial neoplasia is resection. In a landmark therapeutic clinical trial by Kenter and colleagues (82) wherein HPV16þ VIN3 patients were vaccinated with synthetic overlapping long peptides (SLP vaccines) covering HPV-16 E6 and E7, 9 out of 19 patients (47%) exhibited complete regression of the disease and HPV16-specific T-cell responses were detected. Although the regression rate was less than 50%, these findings suggest that it is possible to treat HPV-specific disease and induce complete regression via a vaccine-induced T-cell response. Unfortunately, monitoring of E6/E7-specific cellular immune responses has proven more complex than for the antibody responses to VLP vaccines (refs. 83–87; and reviewed in ref. 88 and ref. 89). To date, trials using a plethora of therapeutic vaccine platforms have elicited E6- or E7-specific cellular immune responses in the peripheral blood that were weak and often did not infiltrate the tumor regions. More importantly, even if HPV antigen-specific T-cell responses were detected in the peripheral blood, these responses did not always correlate with clinical response. Recently, Maldonado and colleagues (90) suggest that T-cell responses are sequestered in the lesion micro-environment, at the site of antigen. In patients with HPV16þ CIN2/3 primed twice with a DNA vaccine and boosted with a recombinant vaccinia 8 weeks before a standard therapeutic resection, they found that in subjects that had residual disease, clonally expanded, proliferating effector immune responses that were organized in lymphoid aggregates were localized in lesional mucosa. Intraepithelial CD8þ infiltrates were increased compared with prevaccination, and these infiltrates were associated with histologic features of apoptosis in dysplastic epithelial cells. Vaccinated subjects who had shared HLA alleles had shared T-cell receptors (TCR) in tissue T cells. The frequencies of these TCRs were variable in the peripheral blood, suggesting that the tissue responses were the result of a process of selection, as opposed to transudate. These observations raise the question of whether earlier vaccine studies in which HPV-specific T-cell responses in the blood were very weak or not detectable may have elicited local responses that were not measured. In the heterologous prime-boost vaccination study, within-subject comparisons of tissue samples obtained before and after vaccination suggested that previous studies may have been, in effect, censoring histologic endpoints. Because preinvasive HPV lesions are clinically indolent, and directly accessible, they present an opportunity to better understand mechanisms of disease clearance, and tissuelocalized obstacles to clearance. On the basis of earlier studies of tissue predictors of clinical outcomes in unvaccinated CIN2/3 lesions that demonstrated downregulated expression of adhesion molecules in the neovasculature associated with persistent lesions (91), clinical testing of peripheral vaccination with the heterologous prime-boost regimen, in concert with direct manipulation of the lesion microenvironment with topical TLR agonist is ongoing (NCT00788164). Clearly, there is much to be learnt about the mechanisms of targeting vaccination responses to the relevant site (87), and that it is beneficial to monitor cellular immune responses systemically and at the site of infection (92). The negative influence of the local tumor microenvironment on clinical responses to therapeutic vaccination is also becoming more recognized. Much work is currently focused on using different strategies to alter the local microenvironment to enhance immune surveillance against tumors (reviewed in ref. 93 and ref. 94). For example, the local application of the topical immune modulator imiquimod can alter the local microenvironment via activation of innate (TLR7) signaling and foster an effective immune response. Such local inflammatory responses likely enhance targeting to the relevant site and overcome local immune suppressive responses. Indeed, topical imiquimod treatment is partially effective against genital warts, cervical and vulvar neoplasia (95, 96). However, as the HPV disease enlarges and progresses, the effects of imiquimod become limited (97). This situation may require additional and systemic treatments to overcome more profound immune suppression in cancer such as combination therapy of HPV therapeutic vaccination with either chemotherapeutic agents or with low doses of radiation. Indeed, in preclinical models, conventional cytotoxic therapies either prime or enhance preexisting or HPV therapeutic vaccination induced antitumor-specific immune responses (98–100), but additional work to determine the optimal combinations and timing of administration is needed. Several factors also limit the action of HPV-specific cytotoxic T cells on infected cells. Immune-suppressive cells such as the T-regulatory cells (Tregs) or tumor-associated macrophages (TAM; refs. 101, 102) are also important contributors to immune-suppression locally in the tumor-microenvironment. Studies in cervical cancer also indicate a central role for CD4þ Tregs in immune evasion (86, 102). Currently, there is also much interest in using monoclonal antibody-based systemic therapies that target coinhibitory receptors such as CTLA-4 (cytotoxic Tlymphocyte- associated protein 4) or PD-1 (programmed celldeath protein 1) on CTLs. Recent studies show that the use of anti-CTLA-4 antibodies (e.g., ipilimumab or tremelimumab) in clinical trials in several cancers achieved complete clinical responses in some cases associated with the reactivation and infiltration of CTLs into the tumor bed. These findings in turn suggest the potential use of CTLA-4 inhibition in combination with HPV therapeutic vaccination to treat HPVþ cancer. A second potential target is the recent discovery of PD-1, another inhibitory member of the CD28/CTLA-4 family of coreceptors, which is expressed not only in T cells but other immune cells such as B cells, macrophages, and even NK-cells. Importantly, PD-1 ligands have been shown to be more highly expressed in tonsillar crypts as well as in both HPV-associated head and neck squamous cell cancer (HPV-HNSCC) tumor-associated macrophages and tumor cells. Concurrently, it was found that the majority of the CD8þ tumor-infiltrating CTLs had high expression of PD-1. Taken together, this provides evidence for the PD-1/PDL1 pathway in maintaining an immune-suppressive tumor microenvironment and provides a rationale for blocking this PD-1/PDL1 pathway to improve the immune response against HPVHNSCC (103, 104). Surprisingly, however, most cervical cancers are PD-L1–negative (105), implying differences in the immune microenvironment of cervical cancer and HPV-HNSCC. Endpoints In HPV vaccine studies, protection against or clearance of highgrade CIN (CIN2/3) is typically a primary endpoint as it is the recognized precursor lesion of cervical cancer (Fig. 2). However, because of the diagnostic variability for CIN2, many therapeutic studies now focus on CIN3 only to test clinical activity. Therapeutic effects of a vaccine are often delayed because of the time needed to develop an effective immune response as opposed to the more direct action of a small molecule. Therefore, it is critical in such studies to follow the endpoints over a sufficient period to capture the effects of vaccination. However, the safety of a delay in treatment of CIN2/3 with LEEP must be carefully considered because of the risk, albeit low over a limited period, of progression. Indeed, a follow-up of 19 weeks has been safely used, and delays of 9 months for LEEP treatment are taken in routine clinical practice for newly pregnant women with CIN2/3. Care must also be taken when interpreting an effective vaccine response as CIN2/ 3 patients exhibit significant rates of spontaneous regression (25%) and inflammation triggered by biopsy may influence responses(80). An alternative disease upon which to test HPV therapeutic vaccines is VIN2/3, given its lower regression rate (1.2%; refs. 106, 107), although this is much less prevalent than CIN2/3. Indeed, the low rate of spontaneous regression was used to support the absence of a control group in the landmark HPV16 E6/E7 SLP clinical trial (82). Similarly, much thought is needed in deciding the endpoints as well as what constitutes "efficacy" of current prophylactic vaccines. These regulatory and policy-making decisions profoundly affect vaccine development and uptake (ref. 108, reviewed in ref. 109). The use of CIN2/3 as the endpoint for the qualification of second-generation preventive HPV vaccines will greatly increase trial sizes over the use of persistent HPV DNA detection as an endpoint, and may impede their development and drive up cost of biosimilar vaccines. This issue is particularly important for vaccines intended to prevent HPVþ head and neck cancer because there is currently no precursor lesion and screening protocol defined, and it is neither feasible nor ethical to use cancer as an endpoint. However, robust protocols exist for detection of persistent oral hrHPV infection and warrant serious consideration as an endpoint. The use of immunologic endpoints such as L1 VLP ELISA or in vitro neutralization titers in serum might also be considered for noninferiority studies of biosimilar L1 VLP vaccines, although the correlate of protection has yet to be properly defined. Fortunately there has been minimal, if any evidence of breakthrough infection by vaccine types in appropriately immunized patients. However, this renders the determination of a minimal neutralizing titer associated with protection very difficult, although one possibility is to use the infection by nonvaccine types for which protection is partial and compare serum cross-neutralizing titers in these patients with those who are not infected. It will also be important to understand the role of memory B cells and the recall response in long-term protection (i.e., is there sufficient time for the inoculum to elicit a rapid and local antibody response if the local protective antibody level has waned below that required for sterilizing immunity), as this has also been suggested as an important factor. The measurement of relevant immune correlates for therapeutic HPV vaccines is much more controversial especially given the greater technical complexity of the assays, the diversity of effector cells, the importance of targeting of the antiviral responses to the lesion site, and the potential of an immune-suppressive local environment. These issues suggest the importance of monitoring the response locally, which creates significant technical hurdles over measurement of systemic immune responses in blood. The need for broad protection is becoming increasingly clear in light of differing prevalence of certain key hrHPV in different populations, notably HPV52 and 58, although HPV16 and 18 are the dominant types in cervical cancer worldwide. Although there is not clear evidence for competition between types and most mathematicalmodeling studies suggest genotype replacement is unlikely, one study has recently reported an increase in the prevalence of nonvaccine HPV types after vaccination (110). As there are insufficient data currently to further substantiate such findings (111), long-term follow-ups of vaccinated populations will be required to answer such questions. However, the development of highly multivalent L1 VLP or other L2-based second-generation HPV vaccines will further reduce concern for such issues. Screening for HPV-Associated Malignancies in the Era of HPV Vaccination In considering the impact of HPV vaccination, it is critical to address how it is best integrated with Pap screening. Vaccination with L1 VLPwill not render cervical screening redundant in the near term as it has no therapeutic effect. In addition, screening is not recommended forwomen ages >26 years (basedonthe assumption that they have had an active sexual history), and vaccination is recommended for 9 to 26-year olds, i.e., many older women have not benefitted from HPV vaccination. Furthermore, because of the limited hrHPV type specificity of the licensed vaccines, screening will still detect disease associated with nonvaccine types. However, these screening programs must be reevaluated as the predictive value and cost-effectiveness of screening will be significantly lower in vaccinated women (112; reviewed in ref. 113), and dramatically so with the advent of the nonavalent vaccine. To begin to address such issues, it has been proposed that primary screening be done via HPV DNA testing first followed by Pap cytology triage (reviewed in refs. 114, 115). These HPV DNA detection assays are more sensitive for disease as compared with PAP screening; however, in terms of determining true disease, the assays are at least 10% less specific than the Pap smear (116). Nonetheless, cohort studies have subsequently shown that there is actually minimal overdiagnosis when HPV DNA testing is used as the solemodality with cytology reserved for triage of HPV-positive women with increased screening intervals (e.g., once every 3 years; refs. 117, 118). Indeed, the U.S. FDA has recently approved the Roche cobas HPV test methodology as the first HPV DNA test for primary cervical cancer screening (there are current four FDAapproved assays but only one is approved for primary screening). The cobas test specifically identifies HPV 16 and HPV 18, while concurrently detecting 12 other types ofhigh-riskHPVs. Itwill beup to professional medical societies and organizations to determine how this HPV DNA test will be incorporated into current screening protocols. Logically, DNA testing combined with cytology will probably the best regimen for developed countries with robust healthcare infrastructure, and might be used to trigger treatment with a therapeutic HPV vaccine. Many resource-limited developing countries have adopted visual inspection by acetic acid (VIA)-only programs as it is a cheaper and immediate point-of-care approach compared with delayed HPV DNA/Pap testing in a central laboratory, although clearly less predictive and potentially problematic to combine with an immunotherapy without knowledge of the hrHPV genotype (assuming a type-specific immunotherapy). In contrast with the cervix, measures to detect and screen for HPV-associated oropharynx cancers have been lacking due to a lack of well-defined precursor lesion (reviewed in ref. 119). While primary prevention via vaccination should prove promising in the long term (120), there is currently no secondary prevention option. Recently, a strong association between HPV16 E6 serum antibodies and HPV-associated oropharynx cancers was reported (121) . Furthermore, these HPV16 E6 antibodies were present in a notable proportion of patients with HPV-associated oropharyngeal cancers for >10 years before diagnosis, but this approach currently lacks sufficient predictive value alone. HPV DNA testing in oral cavity specimens (122) is a particularly promising approach for screening, although more work is needed to understand the predictive value of this biomarker testing strategy for HPVþ HNSCC and its precursors. HPV malignancies of the oropharynx are predominantly due to a single genotype, HPV16, and thus an immunotherapeutic approach focused on HPV16 Tcell based vaccines could be the primary goal for immunotherapeutic control of these infections, particularly in those were have seroconverted to HPV16 E6 positivity. Concluding Remarks The finding that hrHPV infection is necessary although insufficient for the development of cervical cancer has driven tremendous advances in cancer immunoprevention, including three licensed prophylactic vaccines (Fig. 1). We anticipate further significant advances in broadening protection to all hrHPV, lowering the number of doses and cost of HPV vaccination, and eliciting therapeutic immunity. Such developments in immunotherapy of HPV will complement major advances in screening, notably HPV DNA, RNA and possibly oncoprotein testing as a first-line screening modality in the cervix and also possibly at noncervical sites. The advent of such opportunities will require significant changes in health policies for best implementation and realization of their potential to eliminate HPV-related cancer and drive down costs so that all may benefit. Disclosure of Potential Conflicts of Interest W. K. Huh is a consultant/advisory board member for Merck and TheraVax. C. L. Trimble is a consultant/advisoryboardmember forMerck.RichardRodenisan inventor of L2-related patents licensed to Shantha Biotechnics Ltd., Glaxo- SmithKline, PaxVax, Inc. and Acambis, Inc. Richard Roden has received research funding from Sanofi Pasteur, Shantha Biotechnics and GlaxoSmithKline and is a co-founder of and has an equity ownership interest in Papivax LLC. Richard Roden owns Papivax Biotech Inc. stock options and is a member of Papivax Biotech Inc.'s Scientific Advisory Board. Under a licensing agreement between Papivax Biotech, Inc. and the Johns Hopkins University, Richard Roden is entitled to royalties on technologies described in this review. This arrangement has been reviewed and approvedby theJohnsHopkinsUniversity in accordancewith its conflict of interest policies. No potential conflicts of interest were disclosed by the other authors. Grant Support This work was funded by Public Health Service grants P50 CA098252, RO1 CA133749, and CA118790 (grants.nih.gov/), and the V foundation (wwww.jimmyv.org/). Received September 23, 2014; revised November 6, 2014; accepted December 1, 2014; published OnlineFirst December 8, 2014. References Bray F, Ren JS, Masuyer E, Ferlay J. Global estimates of cancer prevalence for 27 sites in the adult population in 2008. Int J Cancer 2013;132:1133–45. D'Souza G, Kreimer AR, Viscidi R, Pawlita M, Fakhry C, Koch WM, et al.Case-control study of human papillomavirus and oropharyngeal cancer. N Engl J Med 2007;356:1944–56. De Martel C, Ferlay J, Franceschi S, Vignat J, Bray F, Forman D, et al. Global burden of cancers attributable to infections in 2008: a review and synthetic analysis. Lancet Oncol 2012;13:607–15. CDC. Disease Burden from Viral Hepatitis A, B, and Cin the United States. Cited May 5, 2014. Available from: www.cdc.gov/hepatitis/HBV/ StatisticsHBV.htm. Mast EE, Weinbaum CM, Fiore AE, Alter MJ, Bell BP, Finelli L, et al. A comprehensive immunization strategy to eliminate transmission of hepatitis B virus infection in the United States: recommendations of the Advisory Committee on Immunization Practices (ACIP) Part II: immunization of adults. MMWR Recomm Rep 2006;55:1–33; quiz CE1–4. Chang MH, Chen CJ, Lai MS, Hsu HM, Wu TC, Kong MS, et al. Universal hepatitis B vaccination in Taiwan and the incidence of hepatocellular carcinoma in children. Taiwan Childhood Hepatoma Study Group. N Engl J Med 1997;336:1855–9. RodriguezAC, SolomonD,HerreroR,HildesheimA,Gonzalez P,Wacholder S, et al. Impact of human papillomavirus vaccination on cervical cytology screening, colposcopy, and treatment. Am J Epidemiol 2013;178:752–60. Walboomers JMM, Jacobs MV, Manos MM, Bosch FX, Kummer JA, Shah KV, et al. Human papillomavirus is a necessary cause of invasive cervical cancer worldwide. J Pathol 1999;189:12–9. Bouvard V, Baan R, Straif K, Grosse Y, Secretan B, El Ghissassi F, et al. A review of human carcinogens–Part B: biological agents. Lancet Oncol 2009;10:321–2. Munoz N, Bosch FX, de Sanjose S, Herrero R, Castellsague X, Shah KV, et al. Epidemiologic classification of human papillomavirus types associated with cervical cancer. New Engl J Med 2003;348:518–27. Bosch FX, Lorincz A, Munoz N, Meijer CJLM, Shah KV. The causal relation between human papillomavirus and cervical cancer. J Clin Pathol 2002;55:244–65. Meisels A, Fortin R. Condylomatous lesions of the cervix and vagina. I. Cytologic patterns. Acta cytologica 1976;20:505–9. Durst M, Gissmann L, Ikenberg H, zur Hausen H. A papillomavirus DNA froma cervical carcinomaand its prevalence in cancer biopsy samples from different geographic regions. Proc Natl Acad Sci U S A 1983;80:3812–5. Boshart M, Gissmann L, Ikenberg H, Kleinheinz A, Scheurlen W, zur Hausen H. A new type of papillomavirus DNA, its presence in genital cancer biopsies and in cell lines derived from cervical cancer. EMBO J 1984;3:1151–7. IARC. IARC monograph on evaluation of carcinogenic risks of humans. Human Papillomaviruses; 1995. Vol. 64. Kirnbauer R, Booy F, Cheng N, Lowy DR, Schiller JT. Papillomavirus L1 major capsid protein self-assembles into virus-like particles that are highly immunogenic. Proc Natl Acad Sci U S A 1992;89:12180–4. Zhou J, Sun XY, Stenzel DJ, Frazer IH. Expression of vaccinia recombinant HPV16 L1 and L2ORFproteins in epithelial cells is sufficient for assembly of HPV virion-like particles. Virology 1991;185:251–7. Villa LL, Perez G, Kjaer SK, Paavonen J, Lehtinen M, Munoz N, et al. Quadrivalent vaccine against human papillomavirus to prevent highgrade cervical lesions. New Engl J Med 2007;356:1915–27. Garland SM, Hernandez-Avila M, Wheeler CM, Perez G, Harper DM, Leodolter S, et al. Quadrivalent vaccine against human papillomavirus to prevent anogenital diseases. New Engl J Med 2007;356:1928–43. Munoz N, Kjaer SK, Sigurdsson K, IversenOE, Hernandez-Avila M,Wheeler CM, et al. Impact of Human Papillomavirus (HPV)-6/11/16/18 vaccine on All HPV-associated genital diseases in young women. J Natl Cancer I 2010;102:325–39. Joura EA, Leodolter S, Hernandez-Avila M, Wheeler CM, Perez G, Koutsky LA, et al. Efficacy of a quadrivalent prophylactic human papillomavirus (types 6, 11, 16, and 18) L1 virus-like-particle vaccine against high-grade vulval and vaginal lesions: a combined analysis of three randomised clinical trials. Lancet 2007;369:1693–702. Dillner J, Kjaer SK, Wheeler CM, Sigurdsson K, Iversen OE, Hernandez- Avila M, et al. Four year efficacy of prophylactic human papillomavirus quadrivalent vaccine against low grade cervical, vulvar, and vaginal intraepithelial neoplasia and anogenital warts: randomised controlled trial. Brit Med J 2010;341:c3493. Paavonen J, Jenkins D, Bosch FX, Naud P, Salmeron J, Wheeler CM, et al. Efficacy of a prophylactic adjuvanted bivalent L1 virus-like-particle vaccine against infection with human papillomavirus types 16 and 18 in young women: an interim analysis of a phase III double-blind, randomised controlled trial. Lancet 2007;369:2161–70. Paovonen J, Naud P, Salmeron J, Wheeler CM, Chow SN, Apter D, et al. Efficacy of human papillomavirus (HPV)-16/18 AS04-adjuvanted vaccine against cervical infection and precancer caused by oncogenic HPV types (PATRICIA): final analysis of a double-blind, randomised study in young women. Lancet 2009;374:301–14. Hildesheim A, Herrero R, Wacholder S, Rodriguez AC, Solomon D, Bratti MC, et al. Effect of human papillomavirus 16/18 L1 viruslike particle vaccine among young women with preexisting infection - A randomized trial. Jama-J Am Med Assoc 2007;298:743–53. Ali H, Donovan B, Wand H, Read TRH, Regan DG, Grulich AE, et al. Genital warts in young Australians five years into national human papillomavirus vaccination programme: national surveillance data. Bmj-Brit Med J 2013;346:f2032. Read TR, Hocking JS, Chen MY, Donovan B, Bradshaw CS, Fairley CK. The near disappearance of genital warts in young women 4 years after commencing a national human papillomavirus (HPV) vaccination programme. Sex Transm Infect 2011;87:544–7. Gertig DM, Brotherton JML, Saville M. Measuring human papillomavirus (HPV) vaccination coverage and the role of the National HPV Vaccination Program Register, Australia. Sex Health 2011;8:171–8. Ali H, Guy RJ, Wand H, Read TRH, Regan DG, Grulich AE, et al. Decline in in-patient treatments of genital warts among young Australians following the national HPV vaccination program. BMC Infect Dis 2013;13:140. Leroux-Roels G, Haelterman E, Maes C, Levy J, De Boever F, Licini L, et al. Randomized trial of the immunogenicity and safety of the Hepatitis B vaccine given in an accelerated schedule coadministered with the human papillomavirus type 16/18 AS04-adjuvanted cervical cancer vaccine. Clin Vaccine Immunol 2011;18:1510–8. Schmeink CE, Bekkers RL, Josefsson A, Richardus JH, Berndtsson Blom K, David MP, et al. Co-administration of human papillomavirus-16/18 AS04-adjuvanted vaccine with hepatitis B vaccine: randomized study in healthy girls. Vaccine 2011;29:9276–83. Arguedas A, Soley C, Loaiza C, Rincon G, Guevara S, Perez A, et al. Safety and immunogenicity of one dose of MenACWY-CRM, an investigational quadrivalent meningococcal glycoconjugate vaccine, when administered to adolescents concomitantly or sequentially with Tdap and HPV vaccines. Vaccine 2010;28:3171–9. Wheeler CM, Harvey BM, Pichichero ME, Simon MW, Combs SP, Blatter MM, et al. Immunogenicity and safety of human papillomavirus-16/18 AS04-adjuvanted vaccine coadministered with tetanus toxoid, reduced diphtheria toxoid, and acellular pertussis vaccine and/or meningococcal conjugate vaccine to healthy girls 11 to 18 years of age: results from a randomized open trial. Pediatr Infect Dis J 2011;30:e225–34. Levin A, Wang SA, Levin C, Tsu V, Hutubessy R. Costs of introducing and delivering HPV vaccines in low and lower middle income countries: inputs for GAVI policy on introduction grant support to countries. PloS ONE 2014;9:e101114. Romanowski B, Schwarz TF, Ferguson LM, Peters K, DionneM, Schulze K, et al. Immunogenicity and safety of the HPV-16/18 AS04-adjuvanted vaccine administered as a 2-dose schedule compared with the licensed 3- dose schedule Results from a randomized study. Hum Vaccines 2011;7:1374–86. Schwarz TF, Huang LM, Medina DMR, Valencia A, Lin TY, Behre U, et al. Four-year follow-up of the immunogenicity and safety of the HPV-16/18 AS04-adjuvanted vaccine when administered to adolescent girls aged 10– 14 Years. J Adolescent Health 2012;50:187–94. Krajden M, Cook D, Yu A, Chow R, Mei W, McNeil S, et al. Human papillomavirus 16 (HPV 16) and HPV 18 antibody responses measured by pseudovirus neutralization and competitive luminex assays in a twoversus three-dose HPV vaccine trial. Clin Vaccine Immunol 2011;18: 418–23. Smolen KK, Gelinas L, Franzen L, Dobson S, Dawar M,Ogilvie G, et al. Age of recipient and number of doses differentially impact human B and T cell immune memory responses to HPV vaccination. Vaccine 2012;30: 3572–9. Dobson SRM, McNeil S, Dionne M, Dawar M, Ogilvie G, Krajden M, et al. Immunogenicity of 2 doses of HPV vaccine in younger adolescents vs 3 doses in young women a randomized clinical trial. Jama-J Am Med Assoc 2013;309:1793–802. Lazcano-Ponce E, Stanley M, Munoz N, Torres L, Cruz-Valdez A, Salmeron J, et al. Overcoming barriers to HPV vaccination: non-inferiority of antibody response to human papillomavirus 16/18 vaccine in adolescents vaccinated with a two-dose vs. a three-dose schedule at 21 months. Vaccine 2014;32:725–32. Herweijer E, Leval A, Ploner A, Eloranta S, Simard JF, Dillner J, et al. Association of varying number of doses of quadrivalent human papillomavirus vaccine with incidence of condyloma. JAMA 2014;311:597–603. WHO. Meeting of the strategic advisory group of experts on immunization, April 2014 – conclusions and recommendations. The Weekly Epidemiological Record (WER) 2014 [cited 89 21]; 221–36]. Available from: www.who.int/wer/2014/wer8921.pdf?ua¼1) Safaeian M, Porras C, Pan Y, Kreimer A, Schiller JT, Gonzalez P, et al. Durable antibody responses following one dose of the bivalent human papillomavirus L1 virus-like particle vaccine in the Costa Rica Vaccine Trial. Cancer Prev Res 2013;6:1242–50. Jenkins D. A review of cross-protection against oncogenic HPVby an HPV- 16/18 AS04-adjuvanted cervical cancer vaccine: importance of virological and clinical endpoints and implications for mass vaccination in cervical cancer prevention. Gynecol Oncol 2008;110:S18–25. Malagon T, Drolet M, Boily MC, Franco EL, Jit M, Brisson J, et al. Crossprotective efficacy of two human papillomavirus vaccines: a systematic review and meta-analysis. Lancet Infect Dis 2012;12:781–9. Wheeler CM, Castellsague X, Garland SM, Szarewski A, Paavonen J, Naud P, et al. Cross-protective efficacy of HPV-16/18 AS04-adjuvanted vaccine against cervical infection and precancer caused by non-vaccine oncogenic HPV types: 4-year end-of-study analysis of the randomised, double-blind PATRICIA trial. Lancet Oncol 2012;13:100–10. Harro CD, Pang YY, Roden RB, Hildesheim A, Wang Z, Reynolds MJ, et al. Safety and immunogenicity trial in adult volunteers of a human papillomavirus 16 L1 virus-like particle vaccine. J Natl Cancer Inst 2001;93:284–92. Franceschi S, Lise M, Clifford GM, Rickenbach M, Levi F, Maspoli M, et al. Changing patterns of cancer incidence in the early- and late-HAART periods: the Swiss HIV Cohort Study. British journal of cancer 2010; 103:416–22. Palefsky JM, Minkoff H, Kalish LA, Levine A, Sacks HS, Garcia P, et al. Cervicovaginal human papillomavirus infection in human immunodeficiency virus-1 (HIV)-positive and high-risk HIV-negative women. J Natl Cancer Inst 1999;91:226–36. Janoff EN, Hardy WD, Smith PD, Wahl SM. Humoral recall responses in HIV infection. Levels, specificity, and affinity of antigen-specific IgG. J Immunol 1991;147:2130–5. Arpadi SM, Markowitz LE, BaughmanAL, Shah K,Adam H, Wiznia A, et al. Measles antibody in vaccinated human immunodeficiency virus type 1- infected children. Pediatrics 1996;97:653–7. Kroon FP, van Dissel JT, de Jong JC, van Furth R. Antibody response to influenza, tetanus and pneumococcal vaccines in HIV-seropositive individuals in relation to the number of CD4þ lymphocytes. AIDS 1994; 8:469–76. Denny L, Hendricks B, Gordon C, Thomas F, Hezareh M, Dobbelaere K, et al. Safety and immunogenicity of the HPV-16/18 AS04-adjuvanted vaccine in HIV-positive women in South Africa: a partially-blind randomised placebo-controlled study. Vaccine 2013;31:5745–53. Asgari MM, Kiviat NB, Critchlow CW, Stern JE, Argenyi ZB, Raugi GJ, et al. Detection of human papillomavirus DNA in cutaneous squamous cell carcinoma among immunocompetent individuals. J Invest Dermatol 2008;128:1409–17. Feltkamp MCW, Broer R, di Summa FM, Struijk L, van der Meijden E, Verlaan BPJ, et al. Seroreactivity to epidermodysplasia verruciformisrelated human papillomavirus types is associated with nonmelanoma skin cancer. Cancer Res 2003;63:2695–700. Bavinck JNB, Euvrard S, Naldi L, Nindl I, Proby CM, Neale R, et al. Keratotic skin lesions and other risk factors are associated with skin cancer in organ-transplant recipients: a case-control study in the Netherlands, United Kingdom, Germany, France, and Italy. J Invest Dermatol 2007; 127:1647–56. Bavinck JNB, Plasmeijer EI, Feltkamp MCW. Beta-papillomavirus infection and skin cancer. J Invest Dermatol 2008;128:1355–8. Van Damme PVT, Brodszki N, Diez-Domingo J, Icardi G, Petersen L-K, Tran C, Thomas S, Baudin M. Immunogenicity and safety of a novel 9- valent hpv vaccine in girls 9–15 years of age compared to the quadrivalent vaccine. Eurogin Florence: Italy; 2013. Joura EV503–001 study team. Efficacy and immunogenicity of a novel 9- valent hpv l1 virus-like particle vaccine in 16- to 26-year-old women. Eurogin Florence: Italy; 2013. Wang JW, Roden RBS. Virus-like particles for the prevention of human papillomavirus-associated malignancies. Expert Rev Vaccines 2013; 12:129–41. Chen XJS, Casini G, Harrison SC, Garcea RL. Papillomavirus capsid protein expression in Escherichia coli: purification and assembly of HPV11 and HPV16 L1. J Mol Biol 2001;307:173–82. Schadlicha L, Senger T, Kirschning CJ, Muller M, Gissmann L. Refining HPV 16 L1 purification from E. coli: reducing endotoxin contaminations and their impact on immunogenicity. Vaccine 2009;27:1511–22. Schadlich L, Senger T, Gerlach B, Mucke N, Klein C, Bravo IG, et al. Analysis of modified human papillomavirus type 16 L1 capsomeres: the ability to assemble into larger particles correlates with higher immunogenicity. J Virol 2009;83:7690–705. Thones N, Muller M. Oral immunization with different assembly forms of the HPV 16 major capsid protein L1 induces neutralizing antibodies and cytotoxic T-lymphocytes. Virology 2007;369:375–88. Wang JW, Roden RB. L2, the minor capsid protein of papillomavirus. Virology 2013;445:175–86. Gambhira R, Jagu S, Karanam B, Gravitt PE, Culp TD, Christensen ND, et al. Protection of rabbits against challenge with rabbit papillomaviruses by immunization with the N terminus of human papillomavirus type 16 minor capsid antigen L2. J Virol 2007;81:11585–92. Roden RB, Weissinger EM, Henderson DW, Booy F, Kirnbauer R, Mushinski JF, et al. Neutralization of bovine papillomavirus by antibodies to L1 and L2 capsid proteins. J Virol 1994;68:7570–4. Gambhira R, Karanam B, Jagu S, Roberts JN, Buck CB, Bossis I, et al. A protective and broadly cross-neutralizing epitope of human papillomavirus L2. J Virol 2007;81:13927–31. Kawana Y, Kawana K, Yoshikawa H, Taketani Y, Yoshiike K, Kanda T. Human papillomavirus type 16 minor capsid protein l2 N-terminal region containing a common neutralization epitope binds to the cell surface and enters the cytoplasm. J Virol 2001;75:2331–6. Wang JW, Jagu S, Kwak K, Wang C, Peng S, Kirnbauer R, et al. Preparation and properties of a papillomavirus infectious intermediate and its utility for neutralization studies. Virology 2014;449:304–16. Roden RB, Yutzy Wht, Fallon R, Inglis S, Lowy DR, Schiller JT. Minor capsid protein of human genital papillomaviruses contains subdominant, cross-neutralizing epitopes. Virology 2000;270:254–7. Jagu S, KaranamB,Gambhira R,Chivukula SV,Chaganti RJ, LowyDR, et al. Concatenated multitype L2 fusion proteins as candidate prophylactic panhuman papillomavirus vaccines. J Natl Cancer I 2009;101:782–92. Seitz H, Dantheny T, Burkart F, Ottonello S, Muller M. Influence of oxidation and multimerization on the immunogenicity of a thioredoxin- l2 prophylactic papillomavirus vaccine. Clin Vaccine Immunol 2013;20:1061–9. Kondo K, Ochi H, Matsumoto T, Yoshikawa H, Kanda T. Modification of human papillomavirus-like particle vaccine by insertion of the crossreactive L2-epitopes. J Med Virol 2008;80:841–6. Schellenbacher C, Kwak K, Fink D, Shafti-Keramat S, Huber B, Jindra C, et al. Efficacy of RG1-VLP vaccination against infections with genital and cutaneous human papillomaviruses. J Invest Dermatol 2013;133: 2706–13. Tumban E, Peabody J, Peabody DS, Chackerian B. A pan-HPV vaccine based on bacteriophage PP7 VLPs displaying broadly cross-neutralizing epitopes from the HPV minor capsid protein, L2. PloS ONE 2011;6: e23310. Von Knebel Doeberitz M, Rittmuller C, zur Hausen H, Durst M. Inhibition of tumorigenicity of cervical cancer cells in nude mice by HPV E6-E7 antisense RNA. Int J Cancer 1992;51:831–4. Schwarz E, Freese UK, Gissmann L, Mayer W, Roggenbuck B, Stremlau A, et al. Structure and transcription of human papillomavirus sequences in cervical-carcinoma cells. Nature 1985;314:111–4. Munger K, Phelps WC, Bubb V, Howley PM, Schlegel R. The E6 and E7 genes of the human papillomavirus type 16 together are necessary and sufficient for transformation of primary human keratinocytes. J Virol 1989;63:4417–21. Trimble CL, Piantadosi S, Gravitt P, Ronnett B, Pizer E, Elko A, et al. Spontaneous regression of high-grade cervical dysplasia: effects of human papillomavirus type and HLA phenotype. Clin Cancer Res 2005;11: 4717–23. Schlecht NF, Platt RW, Duarte-Franco E, Costa MC, Sobrinho JP, Prado JC, et al. Human papillomavirus infection and time to progression and regression of cervical intraepithelial neoplasia. J Natl Cancer Inst 2003;95:1336–43. Kenter GG, Welters MJ, Valentijn AR, Lowik MJ, Berends-van der Meer DM, Vloon AP, et al. Vaccination against HPV-16 oncoproteins for vulvar intraepithelial neoplasia. N Engl J Med 2009;361:1838–47. Nakagawa M, Stites DP, Palefsky JM, Kneass Z, Moscicki AB. CD4-positive and CD8-positive cytotoxic T lymphocytes contribute to human papillomavirus type 16 E6 and E7 responses. Clin Diagn Lab Immunol 1999;6:494–8. De Jong A, van Poelgeest MI, van der Hulst JM, Drijfhout JW, Fleuren GJ, Melief CJ, et al. Human papillomavirus type 16-positive cervical cancer is associated with impaired CD4þT-cell immunity against early antigens E2 and E6. Cancer Res 2004;64:5449–55. Van Poelgeest MI, van Seters M, van Beurden M, Kwappenberg KM, Heijmans-Antonissen C, Drijfhout JW, et al. Detection of human papillomavirus (HPV) 16-specific CD4þ T-cell immunity in patients with persistent HPV16-induced vulvar intraepithelial neoplasia in relation to clinical impact of imiquimod treatment. Clin Cancer Res 2005;11: 5273–80. Van der Burg SH, Piersma SJ, de Jong A, van der Hulst JM, Kwappenberg KM, van den Hende M, et al. Association of cervical cancer with the presence of CD4 þregulatory T cells specific for human papillomavirus antigens. Proc Natl Acad Sci U S A 2007;104:12087–92. Piersma SJ, Jordanova ES, van Poelgeest MI, Kwappenberg KM, van der Hulst JM, Drijfhout JW, et al. High number of intraepithelial CD8þ tumor-infiltrating lymphocytes is associated with the absence of lymph node metastases in patients with large early-stage cervical cancer. Cancer Res 2007;67:354–61. Stern PL, van der Burg SH, Hampson IN, Broker TR, Fiander A, Lacey CJ, et al. Therapy of human papillomavirus-related disease. Vaccine 2012;30: F71–82. Tran NP, Hung CF, Roden R,WuTC. Control of HPV infection and related cancer through vaccination. Recent Results Canc 2014;193:149–71. Maldonado L, Teague JE, Morrow MP, Jotova I, Wu TC, Wang CG, et al. Intramuscular therapeutic vaccination targeting HPV16 induces T cell responses that localize in mucosal lesions. Sci Transl Med 2014;6:221ra13. Trimble CL, Clark RA, Thoburn C, Hanson NC, Tassello J, Frosina D, et al. Human papillomavirus 16-associated cervical intraepithelial neoplasia in humans excludes CD8 T cells from dysplastic epithelium. J Immunol 2010;185:7107–14. Trimble CL, Peng SW, Thoburn C, Kos F, Wu TC. Naturally occurring systemic immune responses to HPV antigens do not predict regression of CIN2/3. Cancer Immunol Immun 2010;59:799–803. Vanneman M, Dranoff G. Combining immunotherapy and targeted therapies in cancer treatment. Nat Rev Cancer 2012;12:237–51. Pardoll DM. The blockade of immune checkpoints in cancer immunotherapy. Nat Rev Cancer 2012;12:252–64. Van Seters M, van Beurden M, ten Kate FJW, Beckmann I, Ewing PC, Eijkemans MJC, et al. Treatment of vulvar intraepithelial neoplasia with topical imiquimod. New Engl J Med 2008;358:1465–73. Grimm C, Polterauer S, Natter C, Rahhal J, Hefler L, Tempfer CB, et al. Treatment of cervical intraepithelial neoplasia with topical imiquimod: a randomized controlled trial. Obstet Gynecol 2012;120:152–9. Pachman DR, Barton DL, Clayton AC, McGovern RM, Jefferies JA, Novotny PJ, et al. Randomized clinical trial of imiquimod: an adjunct to treating cervical dysplasia. Am J Obstet Gynecol 2012;206:42 e1–7. Tseng CW, Hung CF, Alvarez RD, Trimble C, Huh WK, Kim D, et al. Pretreatment with cisplatin enhances E7-specific CD8þ T-Cell-mediated antitumor immunity induced by DNA vaccination. Clin Cancer Res 2008;14:3185–92. Lee SY, KangTH,Knoff J,Huang Z, Soong RS, AlvarezRD, et al. Intratumoral injection of therapeutic HPV vaccinia vaccine following cisplatin enhances HPV-specific antitumor effects. Cancer Immun 2013;62:1175–85. Lee SY, Huang Z, Kang TH, Soong RS, Knoff J, Axenfeld E, et al. Histone deacetylase inhibitor AR-42 enhances E7-specific CD8(þ) T cell-mediated antitumorimmunity induced by therapeutic HPVDNAvaccination. Jmol Med 2013;91:1221–31. Lepique AP, Daghastanli KRP, Cuccovia IM, Villa LL. HPV16 Tumor associated macrophages suppress antitumor T cell responses. Clin Cancer Res 2009;15:4391–400. Loddenkemper C, Hoffmann C, Stanke J, Nagorsen D, Baron U, Olek S, et al. Regulatory (FOXP3(þ)) T cells as target for immune therapy of cervical intraepithelial neoplasia and cervical cancer. Cancer Sci 2009;100:1112–7. Lyford-Pike S, Peng S, Young GD, Taube JM, Westra WH, Akpeng B, et al. Evidence for a role of the PD-1:PD-L1 pathway in immune resistance of HPV-associated head and neck squamous cell carcinoma. Cancer Res 2013;73:1733–41. Badoual C, Hans S, Merillon N, Van Ryswick C, Ravel P, Benhamouda N, et al. PD-1-expressing tumor-infiltrating T cells are a favorable prognostic biomarker in HPV-associated head and neck cancer. Cancer Res 2013;73:128–38. Karim R, Jordanova ES, Piersma SJ, Kenter GG, Chen LP, Boer JM, et al. Tumor-expressed B7-H1 and B7-DC in relation to PD-1þT-cell infiltration and survival of patients with cervical carcinoma. Clin Cancer Res 2009;15:6341–7. Bourgault Villada I, Moyal Barracco M, Ziol M, Chaboissier A, Barget N, Berville S, et al. Spontaneous regression of grade 3 vulvar intraepithelial neoplasia associated with human papillomavirus-16-specific CD4(þ) and CD8(þ) T-cell responses. Cancer Res 2004;64:8761–6. Van Seters M, van Beurden M, de Craen AJM. Is the assumed natural history of vulvar intraepithelial neoplasia III based on enough evidence? A systematic review of 3322 published patients. Gynecol Oncol 2005;97: 645–51. Malmqvist E, Helgesson G, Lehtinen J, Natunen K, Lehtinen M. The ethics of implementing human papillomavirus vaccination in developed countries. Med Health Care Phil 2011;14:19–27. Fisher WA. Understanding human papillomavirus vaccine uptake. Vaccine 2012;30:F149–56. Tabrizi SN, Brotherton JML, Kaldor JM, Skinner SR, Cummins E, Liu B, et al. Fall in human papillomavirus prevalence following a national vaccination program. J Infect Dis 2012;206:1645–51. Pons-Salort M, Thiebaut ACM, Guillemot D, Favre M, Delarocque-Astagneau E.HPV genotype replacement: too early to tell. Lancet Infect Dis 2013;13:1012-. Cong X, Cox DD, Cantor SB. Bayesian meta-analysis of Papanicolaou smear accuracy. Gynecol Oncol 2007;107:S133–7. Franco EL, Cuzick J, Hildesheim A, de Sanjose S. Chapter 20: Issues in planning cervical cancer screening in the era of HPV vaccination. Vaccine 2006;24:S3/171–7. Cuzick J, Arbyn M, Sankaranarayanan R, Tsu V, Ronco G, Mayrand MH, et al. Overview of human papillomavirus-based and other novel options for cervical cancer screening in developed and developing countries. Vaccine 2008;26:K29–41. Bosch FX. Human papillomavirus: science and technologies for the elimination of cervical cancer. Expert Opin Pharmacother 2011;12: 2189–204. Molijn A, Kleter B, Quint W, van Doorn LJ. Molecular diagnosis of human papillomavirus (HPV) infections. J Clin Virol 2005;32:S43–51. Bulkmans NW, Berkhof J, Rozendaal L, van Kemenade FJ, Boeke AJ, Bulk S, et al. Human papillomavirus DNA testing for the detection of cervical intraepithelial neoplasia grade 3 and cancer: 5-year follow-up of a randomised controlled implementation trial. Lancet 2007;370: 1764–72. Naucler P, Ryd W, Tornberg S, Strand A, Wadell G, Elfgren K, et al. Human papillomavirus and Papanicolaou tests to screen for cervical cancer. N Engl J Med 2007;357:1589–97. Kreimer AR. Prospects for prevention of HPV-driven oropharynx cancer. Oral oncology 2014;50:555–9. Herrero R, Quint W, Hildesheim A, Gonzalez P, Struijk L, Katki HA, et al. Reduced prevalence of oral human papillomavirus (HPV) 4 years after bivalent HPV vaccination in a randomized clinical trial in Costa Rica. PloS ONE 2013;8:e68329. Kreimer AR, Johansson M, Waterboer T, Kaaks R, Chang-Claude J, Drogen D, et al. Evaluation of human papillomavirus antibodies and risk of subsequent head and neck cancer. J Clin Oncol 2013;31: 2708–15. Ribeiro KB, Levi JE, Pawlita M, Koifman S, Matos E, Eluf-Neto J, et al. Low human papillomavirus prevalence in head and neck cancer: results from two large case-control studies in high-incidence regions. Int J Epidemiol 2011;40:489–502.
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