In the development of peptide- or protein-based therapeutic cancer vaccines, suboptimal priming of antigen-specific CD8+ cytotoxic T lymphocytes (CTL) due to insufficient MHC class I presentation remains a major problem. Adjuvants are therefore important to improve cross-presentation on MHC class I and to prime CTLs. In this study, we implemented the endosomal escape method photochemical internalisation (PCI) in order to enhance the cytosolic delivery of a human papilloma virus long (HPV-L) vaccination peptide in immunocompetent mice in combination with the Toll-like receptor 3 (TLR3) agonist Poly(I:C) as adjuvant (double stranded RNA mimicking virus infections). In C57BL/6 mice with established dermal syngeneic tumours of the HPV TC-1 cancer model, PCI-mediated vaccination with a synthetic HPV-L derived from HPV16 in combination with poly(I:C) resulted in significant activation of antigen-specific CD8+ T-cells and caused anti-tumour effects. In comparison to PCI alone, combining intratumoural PCI, HPV-L and poly(I:C) resulted in complete responses and cure of all mice (8/8) treated as compared to mice treated with either HPV-L and poly(I:C) (0/8) or PCI and HPV-L (0/8). In conclusion, PCI strongly improved immunity of TLR3-agonist adjuvanted polypeptide antigens. The PCI-based vaccination strategy has a promising potential in the development of therapeutic cancer vaccines and warrant further development towards clinical testing.