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Immunology Group - The Human Papilloma Virus

Persistent infection by ‘high risk' genotypes of human papilloma virus (HPV) is necessary but not sufficient for the development of over 98% of cervical cancers. Thus the development of vaccines that prevent HPV transmission represent an important opportunity to prevent cervical cancer. There are several prophylactic HPV vaccine formulations based upon L1 virus-like particles (VLPs) currently in phase III trials and recently released data are extremely promising. It seems clear that vaccination programmes that target adolescents prior to any exposure have great potential to reduce and ultimately eliminate cervix cancer. However, many practical issues surrounding implementation of these vaccines in different countries still need to be finalized including, who and when to vaccinate, duration of protection and integration with current screening programs. The vaccines currently being evaluated target the two most prevalent high risk HPV types which are responsible for approximately 70% of cervical cancers. To increase the breadth of protection, it is likely that L1 VLPs of other viral subtypes must be included, although vaccines targeting the conserved regions of the L2 minor capsid protein warrant further exploration in this regard. In addition the vaccines nearing licensing will not combat established HPV-related disease and a therapeutic vaccine, of which there are several candidates in early stages of development, would be desirable ( Winters et al 2006).

While HPV is necessary, it is not sufficient for development of invasive cancer. This requires multiple steps involving the selection of additional genetic changes, some of which contribute to evasion of the immune response. Such immune escape may limit the patients that will benefit from immunotherapies even for local disease (Brady et al 2000; Bontkes et al 1998). Our focus has been to test vaccines which can be used to treat existing infection and cancers. We have completed clinical trials of a recombinant vaccinia virus expressing HPV16/18 E6/E7 oncogenes (TA-HPV) in women with early stage cervical cancer (Kaufmann et al 2002) or vulval intraepithelial neoplasia (Davidson et al 2003). These have established the vaccine safety and immunogenicity. Importantly, clinical responses were seen but were significantly correlated to the higher levels of lesion-associated CD4+, CD8+ and CDla+ immune cells in responders compared to non-responders at time of vaccination. Local immune infiltration may be a critical factor in potential responsiveness to vaccine therapy in HPV-associated neoplasia and should be carefully monitored in future placebo-controlled trials of immunotherapy for VIN.

Delivery of cure in this and other HPV-associated lesions may depend on complete viral clearance or at least sustained anti-HPV immunity and a single vaccination may, therefore, not be enough. Heterologous prime-boost vaccination schedules employing TA-HPV in combination with TA-CIN, an HPV 16 L2E6E7 fusion protein, may offer advantages over the use of either agent alone for the immunotherapy of HPV 16-associated VIN. A small study of VIN patients who received 3 booster vaccinations with TA-CIN between 7-15 months after the TA-HPV vaccination demonstrated HPV 16-specific proliferative T cell and/or serological activity but there was no direct correlation between immunological and clinical responses (Davidson et al 2004). The reciprocal order of immunisation has also been tested in 29 women with high grade LGIN. This regimen is immunogenic with both proliferative and interferon- ELISPOT responses biased to HPV16 E6 but again there is no simple relationship between induction of systemic HPV16-specific immunity and clinical outcome (Smyth et al 2004). The patients treated in these studies have HPV associated lesions that can show evidence of altered HLA class I expression thereby facilitating immune escape from any CTL generated by the vaccination. Our other work has shown that HLA polymorphism is a factor in both susceptibility and progression of HPV associated disease (Brady et al 1999; Davidson et al 2003) with additional genetic changes including amplification of F-box protein Skp2 present at 5p13 (Dowen et al 2003) and functional deletion of TSLC1 important for cervical cancer development (Steenbergen et al 2004).

Current trials with Professor Henry Kitchener, St Mary's Hospital Manchester are looking at combination treatments with the first approach stimulation of local innate immunity. Thus 20 patients with high grade vulval intraepithelial neoplasia (VIN) have received treatment with imiquimod followed by photodynamic therapy (PDT); another group is receiving imiquimod plus HPV oncogene vaccination (collaboration with CRT and Xenova). The imiquimod acts partly through inducing a local inflammatory response, and the PDT or vaccination have the potential to stimulate T cell mediated immunity effecting resolution of persistent VIN. The aim is to improve the immunogenicity of the vaccination and the clinical outcome will be measured against important local immune and genetic factors. Furthermore, we will characterize the complex array of infiltrating immune cells and their cytokine products within the vulval skin in relation to clinical outcome. Of particular interest is FOXP3, a transcription factor necessary for the development and function of CD4 + CD25 + T regulatory cells. Understanding the equilibrium between viral and immunological factors is essential to provide the appropriate tools to evoke therapeutic immunity. In collaboration with CRT and Johns Hopkins Medical Center we are developing a clinical protocol for testing the potential for using TA-CIN as a vaccine to generate cross neutralizing antibodies against multiple high risk HPV types as well as therapeutic immunity versus HPV oncogenes.

Consistent with the loss of E2 regulation of oncogene function in tumours, over-expression of E2 in cervical cancer cells can repress E6 and E7 expression, resulting in apoptotic cell death and growth suppression. Work with Bristol University has shown that HPV 16 E2 can induce apoptosis in both HPV transformed and non-transformed cell lines (Webster et al 2000) This E2-induced apoptosis is abrogated by a transdominant negative mutant of p53 or by over-expression of HPV 16 E6, but is increased by over-expression of wild type p53. The apoptotic function depends on the N-terminal not the DNA binding properties of E2-dimers and occurs via a p53-dependent pathway. If E2 could be efficiently delivered to an HPV-associated lesion, a direct induction of apoptosis and concomitant immunisation could combine to clear the virus and establish protection from any subsequent infection. More recent studies have shown that a mutant of the E2 protein that fails to bind p53 is capable of inducing apoptosis in HPV-transformed cells but is unable to cause apoptosis in non-HPV transformed cells. This provides the basis for selective therapy of HPV positive epithelial cells. Current experiments are aimed at investigating in vitro and in vivo induction of apoptosis by VP22 E2 fusion protein (Roeder et al 2004; Green et al 2006).