An improved understanding of the natural history of HPV, its interaction with the host immune system, and the distinct molecular alterations underlying HPV-positive cancers, are fueling development of new drugs, particularly immunotherapies geared toward generating an HPV-specific immune response.
Although unprecedented improvements in our ability to diagnose and treat cancer have driven a decline in incidence and mortality rates for most cancers in the United States, notable exceptions to this trend include oropharyngeal, anal, and vulvar cancers in patients who harbor the human papillomavirus (HPV).
While HPV is most famously the cause of cervical cancers, it is now known to be associated with a variety of other tumor types in which HPV-associated disease is on the rise. Cervical cancer provides proof that HPV-associated disease can be curbed by effective screening and preventive vaccination, but many patients still die despite these advances.
There is a pressing need to develop novel treatments to replace the use of highly invasive and morbid standard therapies. An improved understanding of the natural history of HPV, its interaction with the host immune system, and the distinct molecular alterations underlying HPV-positive cancers, are fueling development of new drugs, particularly immunotherapies geared toward generating an HPV-specific immune response (Table).
This figure illustrates elements of the tumor microenvironment that potentially could support immunebased therapies against HPV-associated cancers: the adaptive immune cells CD4, CD8, and Treg; innate immune cells such as dendritic cells (DC) and natural killer cells (NKT); and the regulatory cytokines IL-10, TGFβ, IFγ-. The goal would be to generate immune cells to prevent reinfection (inset).
Bergot AS, Kassianos A, Frazer IH, Mittal D. New approaches to immunotherapy for HPV associated cancers. Cancers (Basel). 2011;3(3):3461-3495.
A Cancer-Causing Virus
In the mid-1970s, German virologist Harald zur Hausen first made the causative link between HPV and cervical cancer, a discovery that resulted in his receiving a Nobel Prize. HPVs are a large group of related viruses responsible for more than 170 different types of HPV that preferentially infect the mucosa of the genitals, upper respiratory tract, or skin.
It is alpha-HPVs, which infect the mucosal tissue and are most commonly spread through direct and indirect sexual contact, that have gained notoriety for their ability to cause cancer.
Source: Schiller JT, Lowy DR. Understanding and learning from the sucess of prophylactic human papillomavirus vaccines. Nat Rev Microbiol. 2012;10(10):681-692.
Courtesy National Cancer Institute
These strains are categorized by their oncogenic potential as high-, intermediate- and low-risk, with HPV16 and HPV18 being most highly oncogenic. Up to 90% of sexually active individuals will be infected with at least one type of HPV at some point in their lives and around half of those cases will involve high-risk subtypes. Worldwide, the virus is responsible for around 5% of all human cancers.
Our understanding of precisely how HPV causes cancer is still evolving, and one of the most significant conundrums facing researchers is why not everyone infected with HPV develops cancer. The vast majority of patients may not even realize they are infected, since the virus is typically cleared by the immune system in less than 2 years. But when the virus persists for longer and the immune response fails, cancer arises.
HPV infects epithelial cells and hijacks the cellular machinery to generate viral proteins. The HPV genome is arranged into eight known genes: six early genes (E1, E2, E4, E5, E6, and E7) and two late genes (L1 and L2). (Figure). The proteins encoded by these genes promote viral replication and assembly by driving many of the hallmarks of cancer, such as excessive growth, promoting angiogenesis, and avoidance of apoptosis.
Courtesy National Cancer Institute
The E6 and E7 proteins are the most significant drivers of cancer and their bestcharacterized role in carcinogenesis involves suppression of the p53 and retinoblastoma protein (pRb) tumor suppressor pathways. Removal of the tumor suppressive activities of these proteins contributes to the genetic instability that is a central characteristic of cancer. Other mechanisms of HPV-induced oncogenesis are beginning to be uncovered, notably the integration of the HPV genome into the host genome and the ability of HPV to suppress the immune response.
Curbing Cervical Cancer
HPV is most widely known as the cause of the cancer to which it was first linked—cervical cancer. Almost all cases are caused by HPV, with 70% associated with high-risk subtypes HPV16 and HPV18. Persistent HPV infection can lead to the development of low and highgrade cervical intraepithelial neoplasia (CIN), a kind of precancerous state, which in many cases progresses to invasive cervical carcinoma if left untreated.
Since cervical cancer is the second-leading cause of cancer death in women, significant resources have been invested in prevention. Indeed, cervical cancer screening remains one of the most successful screening efforts ever developed. Although Pap smear remains the most frequently used screening tool, new liquid- based cytology technologies have become available. Clinical practice guidelines now recommend co-testing with an HPV DNA test (such as the recently FDA-approved Cobas HPV Test) to identify strains of the virus in addition to cellular abnormalities.
In 2006, the FDA approved Gardasil, a quadrivalent vaccine that protects against HPV types 6, 11, 16, and 18; last year, the agency approved Gardasil-9, which protects against five additional HPV types (31, 33, 45, 52, and 58). The bivalent (HPV16 and 18) vaccine Cervarix was approved in 2009. Despite some controversy over the use of the vaccines, combined with effective screening, they have contributed to a substantial decrease in the rates of cervical cancer.
In the past decade, a number of other cancers have been linked with HPV and, while the rates of cervical cancer have been declining, these other HPV-associated cancers are rapidly rising. Most significantly, a link between head and neck squamous cell carcinoma (HNSCC) and HPV was observed in 2005. Historically, HNSCC was predominantly caused by alcohol and tobacco use, but clinicians noticed an increasing trend in development of HNSCC in seemingly young, healthy individuals. In a landmark study, researchers found that people with HNSCC were 15 times more likely to be infected with HPV in their mouths or throats. Approximately 70% of oropharyngeal cancers are caused by HPV in much the same way that the virus leads to cervical cancer, although the reasons for the increased incidence in HPV-associated HNSCC are unclear.
HPV has also been linked to approximately 95% of anal cancers, 75% of vaginal cancers, 69% of vulvar cancers, and 63% of penile cancers. HPV DNA has been detected in various other patients with cancer, and there are hints that it may be a risk factor for more common tumor types including lung, breast, bladder, and brain cancer. In addition, a number of reports have demonstrated that various nonmelanoma skin cancer types contain the DNA of beta-HPV (a subtype that infects cutaneous sites). However, a causative association for these cancer types has not yet been clearly demonstrated.
Need for Novel Therapies
Despite the success of preventive strategies in cervical cancer, vaccination rates in the United States are far from adequate; just over half of eligible girls and only 35% of boys received at least one shot, according to 2013 figures. Additionally, vaccines do not protect against all potentially cancer-causing HPV strains, and currently there is no evidence that vaccination protects against other forms of HPV-associated cancers.
The result is that many patients are still diagnosed with HPV-associated cancers and preventive vaccines are not effective in treating disease.
The cure rate for cancers identified at an early stage or in a precancerous state is high for both major types of HPV-associated cancer. Patients are treated with standard therapies, including surgery, chemotherapy, and radiation therapy.
However, these treatments can be associated with high morbidity, particularly in the case of HNSCC, and are not effective in patients with more advanced disease, meaning that there is a great need for novel treatment strategies.
Immunotherapies in Development
The most substantial investment has been in the development of therapeutic vaccines and immunotherapies, which are designed to target HPV-associated viral proteins in order to boost the host immune response against HPV-infected cancer cells.
While preventive vaccines are designed to induce antibody and complement-induced immunity, therapeutic vaccines aim to induce cell-mediated immunity that allows them to fight active infection. The vast majority of therapeutic vaccines thus far have been developed to target the most abundant oncogenic proteins—E6 and E7 from the HPV16 and 18 strains.
A variety of different vaccine types have been developed, including live, attenuated vaccines produced by modifying a disease-producing bacteria or virus. Among the most advanced is ADXS11-011, which uses a live attenuated bacterial strain designed to secrete HPV16-E7. Phase II clinical trials are planned or ongoing in patients with anal cancer (NCT02399813), persistent or recurrent cervical cancer (NCT01266460), and oropharyngeal cancer prior to robotic surgery (NCT02002182).
Inactivated vaccines have also been developed that are protein-based, DNA-based, and peptide-based. Only a few protein-based vaccines have reached clinical development; they use the HPV16 and 18 E7 proteins (ProCervix) or a fusion of the L2, E6, and E7 proteins from HPV16 (TA-CIN) to stimulate an immune response. Tthough some clinical efficacy has been observed, there are potential safety issues associated with using oncogenic proteins in vaccines.
Inovio Pharmaceuticals, Inc is heavily invested in the development of therapeutic vaccines for HPV-associated cancers and has developed a number of DNA-based vaccines. MedImmune, a development arm of AstraZeneca, has entered into a collaboration with Inovio to develop its vaccines.
The advantage of using DNA is that it is safer and easier to produce, but still capable of inducing strong immune responses. The development of DNA vaccines has been greatly advanced by the introduction of electroporation, the application of brief electrical pulses to the vaccination site that increase uptake of the DNA plasmid.
VGX-3100 and INO-3112 (in which VGX-3100 is combined with the DNA-based immune activator INO-9012), which both target the HPV16 and 18 subtypes, are the company’s lead products. VGX-3100 monotherapy is being developed primarily for the treatment of CIN (NCT01304524) and the results of this phase II study were recently reported at the 29th Annual Meeting of the Society for Immunotherapy of Cancer. It demonstrated regression and virological clearance in 43 of 107 women with CIN (40.2%, compared with 14.3% in the placebo group). The company has said it expects to begin a phase III study in 2016.
INO-3112 is being investigated in both cervical cancer and HNSCC (NCT02172911, NCT02501278, and NCT02163057). At the World Vaccine Congress 2015 in April, the company announced prelininary data from the first four patients treated in a phase I/II study in patients with HPV-associated HNSCC. The vaccine generated strong CD8-positive T-cell responses in three of the four patients. Meanwhile, the company’s third DNA-based vaccine, INO-3106, targets HPV6 and is currently being evaluated in a phase I trial as monotherapy and in combination with INO-9012 in patients with aerodigestive malignancies.
Several peptide-based vaccines, which also offer safety advantages over protein-based vaccines and are easier to produce, are another category under investigation. The results of a phase I trial evaluating the HPV16 peptide vaccine GL-0810, in which four doses of the vaccine were administered subcutaneously, were recently published. Among five patients who received all four doses, 80% developed antigen-specific T-cell and antibody responses. Median progression- free survival (PFS) and overall survival (OS) were 80 and 196 days, respectively. There were no dose-limiting toxicities and adverse events were predominantly grade 1.
AIN indicates anal intraepithelial neoplasia; ASCUS, atypical squamous cells of undetermined significance; CIN, cervical intraepithelial neoplasia; HIV, human immunodeficiency virus; HNSCC, head and neck squamous cell carcinoma; HPV, human papillomavirus; LSIL, low-grade squamous intraepithelial lesion; TCR, T-cell receptor.
PepCan is a peptide-based vaccine made up of four peptides covering the entire HPV16 E6 protein. It will be evaluated in a phase II study in combination with the Candida albicans skin test reagent Candin, which is used as an adju-vant, in patients with high-grade CIN (NCT02481414). The results of a phase I trial in patients with biopsy-proven CIN 2/3 were presented at the 2015 American Society of Clinical Oncology Annual Meeting (NCT00569231). Among six people treated at four dose levels (50, 100, 250, and 500 μg per peptide), with four injections administered intradermally every 3 weeks, 65% experienced vaccineinduced immune responses. The best histological response was at 50 μg, with a regression rate of 83%. There were no dose-limiting toxicities and the most common adverse events were injection site reactions.
Adoptive Cell Therapy
Other immunotherapies being evaluated in HPV-associated cancers include adoptive cell therapy (ACT), in which the T cells of the immune system are isolated and manipulated outside the body to direct their cell-killing activity against a specific target, in this case the HPV viral proteins, then reinfused into a patient. The results of a trial in patients with cervical cancer treated with ACT-targeting HPV16 and 18 E6 and E7 proteins were recently published in the Journal of Clinical Oncology. Among nine patients previously treated with platinum-based chemotherapy or chemoradiation therapy who received a single infusion of tumor-infiltrating lymphocytes followed by aldesleukin (IL-2), there were two ongoing complete responses of 15 months and 22 months and one partial response of 3 months with tolerable toxicity.
In recent years it has become increasingly clear that simply targeting the immune response against HPV viral proteins may not be sufficient for therapeutic efficacy. Researchers have discovered that HPV has a complicated relationship with the host immune system and, like cancer cells, it is able to “hide” via a variety of mechanisms of immunosuppression.
Combination therapy not only with standard treatments and between vaccines with different mechanisms of action, but also with inhibitors of immune checkpoints such as programmed death-1 (PD1) and its ligand (PD-L1) may become increasingly important.
Early clinical combination trials involving inhibitors include the PD-1 inhibitor nivolumab, which is being paired with the peptide vaccine ISA-101 (NCT02426892) in patients with HPV-positive incurable solid tumors, and the PD-L1 inhibitor durvalumab (MEDI4736), which is being tested with ADXS11-001 in HPV-positive HNSCC (NCT02291055). Results are eagerly anticipated.
Jane de Lartigue, PhD, is a freelance medical writer and editor based in New Haven, Connecticut.