High-risk human papillomaviruses (HPVs), such as HPV16, cause over 500.000 cervical, anogenital and oropharyngeal cancer cases annually. The oncoproteins E6 and E7 are responsible of the transforming potential of high-risk HPVs. These proteins are indispensable for the induction and maintenance of the malignant phenotype, making them ideal targets for therapeutic HPV vaccines.
In order to develop an efficient therapeutic HPV vaccine, it is essential to target E6 and E7 derived epitopes that are present on the surface of cancer cells. By using MS3 mass spectrometry for direct epitope identification, we found that the HLA-A2-restricted HPV16 epitope E711-19 is naturally processed and presented in all tested HPV16+ cell lines. Moreover, E711-19 elicited strong CTL responses in vitro. Therefore, we used this epitope as model antigen to determine the optimal vaccine formulation for anti-HPV cytotoxic T cell (CTL) induction in vivo.
Short peptide epitopes (like E711-19) are poorly immunogenic when administered alone without appropriate immunostimulatory signals. Potent and proper activation of antigen presenting cells, mainly dendritic cells (DCs), is crucial for efficient priming of antigen-specific CTLs. DCs activation can be achieved by including adjuvants such as toll-like receptor (TLR) ligands.
In this study, we investigated the effects of TLR3 (Poly I:C), TLR4 (MPLA), TLR7/8 (R848) and TLR9 (CpG ODNs) ligands on the maturation and activation of human and murine DCs in vitro. Moreover, we analyzed the adjuvant effects of MPLA and R848 in vivo using HLA-A2.1-/HLA-DR1-transgenic H-2 class I-/class II-knockout mice. Mice were subcutaneously vaccinated with E711-19 emulsified in Montanide ISA51 in association with either MPLA alone, R848 alone, or MPLA and R848 in combination. The properties of TLR ligands as immunostimulatory components were investigated by detailed analysis of the induced immune responses 12 days after immunization.
We found that TLR4 and TLR7/8 ligands (alone and in combination) were most efficient in activating DCs in vitro, demonstrated by increased expression of CD80 on human DCs and CD40 on murine DCs. In the in vivo experiments, E711-19 induced antigen-specific T cells when coinjected with the adjuvant ISA 51. Adding MPLA to ISA51 further significantly increased the percentage of antigen-specific T cells.
This study validated the use of this complex humanized mouse model to evaluate the immunogenicity of HPV epitopes in vivo. Additionally, it demonstrated that adding MPLA to an emulsion-based adjuvant (ISA51) can significantly enhance immunogenicity of epitope-based therapeutic HPV vaccines. More vaccine formulations, including other TLR ligands, are currently being tested for in vivo immunogenicity. The most immunogenic formulations will be evaluated for anti-tumor efficacy.
Citation Format: Hadeel Khallouf, Jasmin Mangei, Renata Blatnik, Stephanie Hoppe, Alina Steinbach, Angelika B. Riemer. Evaluation of toll-like receptor ligands as adjuvants for an epitope-based therapeutic HPV vaccine. [abstract]. In: Proceedings of the AACR Special Conference: Tumor Immunology and Immunotherapy: A New Chapter; December 1-4, 2014; Orlando, FL. Philadelphia (PA): AACR; Cancer Immunol Res 2015;3(10 Suppl):Abstract nr B32.
- ©2015 American Association for Cancer Research.