Background: The DNAJB1-PRKACA fusion transcript is the driver of tumor pathogenesis in fibrolamellar hepatocellular carcinoma (FL-HCC) as well as in other tumor entities, e.g. oncocytic neoplasms of the pancreas and bile duct, thus representing a broad target for novel therapies in cancer. Recently, the DNAJB1-PRKACA fusion protein was identified as a source for HLA-presented neoepitopes that can be targeted by T-cell-based immunotherapy (Bauer et al. Nat. Commun., 2022). The DNAJB1-PRKACA fusion-derived neoepitope FusionVAC-22 is in silico predicted to bind to 1,290 HLA class II alleles and contains HLA class I ligands from 13 of the 20 most frequent HLA class I alleles that cover 93.8% of the world population with at least one HLA allotype, enabling the broad application of FusionVAC-22-based immunotherapies. The first application of FusionVAC-22-based peptide vaccines adjuvanted with the TLR1/2 agonist XS15 emulsified in Montanide ISA 51 VG in two FL-HCC patients was well tolerated and showed the induction of profound and long-lasting T-cell responses. Of note, T-cell responses were accompanied by progression-free survival of both patients for so far 32 months and 13 months, respectively. Methods: Based on these encouraging results, we established a Phase I open-label, multicentric clinical trial to evaluate the immunogenicity along with safety and toxicity, as well as first signs of efficacy of the FusionVAC-22 based peptide vaccine, combined with the immune checkpoint inhibitor (ICI) atezolizumab, in 20 patients with locally advanced or metastatic FL-HCC or other malignant diseases that carry the DNAJB1-PRKACA fusion transcript. Further, key eligibility criteria include the absence of autoimmune phenomena due to prior immunotherapy agents (≥ grade 3) as well as tissue or organ allografts. The FusionVAC-22-based peptide vaccine is applied twice in a 4-weekly interval with the opportunity of a booster vaccination after 11 months. Atezolizumab application starts on day 15 after first vaccination and is continued 4-weekly for 1 year, followed by a 6-month follow-up. The primary objectives of the trial are immunogenicity, in terms of induction of peptide specific T-cell responses until 28 days after second vaccination, as well as safety and toxicity of the peptide vaccine in combination with ICI. Safety assessment is based on the frequency of adverse events according to CTCAE v5.0. Clinical efficacy is determined by iRECIST assessment on imaging. Furthermore, disease control rate, quality of life as well as overall and progression free survival will be assessed. Clinical trial information: NCT05937295.