Abstract 578: Investigation of the effect of hypoxia on presentation of HPV16-derived antigens - implications for therapeutic vaccine design

Human papillomavirus type 16 (HPV16) is a causal factor of ~50% of cervical cancers, and of ~95% of extra-cervical (other anogenital and oropharyngeal) HPV-mediated cancers. The aim of our group is to develop a therapeutic vaccine against HPV16-induced malignancies. To find optimal vaccine antigens, it is important to assess the effect of the tumor microenvironment on viral antigen presentation. Hypoxia has been reported to regulate HLA class-I expression levels in the context of various tumors. Thus, the aim of the present study was to explore the effect of hypoxia on various components of the antigen processing machinery (APM) in HPV16-positive tumor cells and on the HPV16 E6- and E7-derived HLA class-I epitope repertoire. We are investigating the effect of hypoxia on cervical cancer cells transformed with HPV16 variant lineages (European, Asian-American and African) correlated with differences in oncogenic potential. This is achieved by comparing cells cultured under hypoxia (1.2% O 2 ) or normoxia (21% O 2 ). We investigate E6 protein levels by immunoblotting and HLA-A2 surface expression by flow cytometry. Subsequently, quantitative PCR and immunoblotting are used to assess whether components of the APM are affected by hypoxia. Cervical cancer cell lines transformed by sublineages of the HPV16 European variant (CaSki and SNU17) showed a decrease in E6 protein levels upon hypoxic treatment. HLA-A2 levels were increased in SNU17 cells (HPV16 European Asian prototype), but no change was observed in CaSki cells (European Prototype 2). First qPCR experiments of SNU17 cells showed changes in additional components of the APM at the mRNA level, which still have to be assessed at the protein level. Cell lines transformed by HPV16 variants other than European will be subjected to the same analysis. Potential changes in the HPV16 epitope repertoire will be investigated using a targeted mass spectrometry-based epitope detection strategy established in the lab. These investigations may yield novel insights into HPV16 immune evasion pathways. Furthermore, the identification of epitopes that are presented under both normoxic and hypoxic conditions will provide optimal candidates for therapeutic vaccine design against HPV-induced malignancies. Citation Format: Nitya Mohan, Maria Bonsack, Jonas Forster, Alina Steinbach, Renata Blatnik, Mogjiborahman Salek, Angelika B. Riemer. Investigation of the effect of hypoxia on presentation of HPV16-derived antigens - implications for therapeutic vaccine design [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 578.