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Abstract

Human papillomaviruses (HPV) are responsible for formation of tumors in the anogenital and oropharyngeal mucosa. The most common malignancy among these cancers is cervical cancer, affecting about 570,000 women every year. Although prophylactic vaccines against several high-risk types of HPV, such as HPV16, have been existing for some time, vaccination rates remain low and cancer incidence rates high. In our group ‘Immunotherapy and immunoprevention’ we are therefore working on a therapeutic vaccine for treatment of HPV16-derived lesions and tumors. So far, no therapeutic vaccines against HPV-induced malignancies have been approved for clinical use even though there is a plethora of promising preclinical studies. These studies have mainly examined the impact of vaccinations with murine MHC-restricted epitopes on subcutaneous tumors. The tumors of patients, however, present human MHC (HLA)-restricted epitopes and are located in the patients’ mucosae, which are restricted for the general circulation of T cells. Herein I present solutions to these problems by first establishing a novel orthotopic (vaginal) tumor model, consisting of a cell line immortalized by the viral oncogenes E6 and E7 and presenting HLA-A2:01-restricted epitopes on its surface for use in syngeneic MHC-humanized A2.DR1 mice. Second, I examined methods to facilitate migration of T cells into the cervicovaginal mucosa for eventual application in patients. For the establishment of the tumor model I first successfully transfected E6+E7+ A2.DR1 lung cells with the activated oncoprotein HRASG12V and firefly luciferase. These cells were specifically killed by HPV16-specific T cells, as well as being tumorigenic and detectable in vivo. This new cell line, E6/7-lucA2, depends on the HPV16 proteins E6 and E7 for its survival and was verified to present our lead epitope E711-19 on its MHC molecules. The cell line was then used to establish a novel orthotopic tumor model in the vaginal mucosa of A2.DR1 mice. This tumor model will ultimately be utilized for testing different therapeutic vaccination approaches. Therefore, I investigated methods to induce a robust vaccination-induced HPV16-specific CD8+ T cell response in the vaginal mucosa. Different exclusively intravaginally delivered vaccine compounds did not elicit a detectable immune response while methods with prior induction of a systemic T cell response were more successful. However, it turned out that the level of mucosal CD8+ T cells just correlates with the level of blood CD8+ T cells and is not detectably influenced by application of immunomodulators such as chemokines or TLR agonists. Taken together, the HPV16 E6/E7-dependent orthotopic, MHC-humanized tumor model E6/7-lucA2 in A2.DR1 mice constitutes a setting for exploration of therapeutic HPV vaccination approaches, which is mirroring the clinical situation much closer than previously available models.