We established a platform for the design of patient-individual peptide vaccination cocktails by combination of whole exome sequencing of tumor and normal tissue with in silico epitope prediction algorithms for individual patient HLA types.
Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy. Standard chemotherapy is a successful treatment in 80% of patients, only about 20% develop a relapse, however these patients have a dismal prognosis. Prevention of relapse after first-line chemotherapy or stem cell transplantation (SCT) is therefore mandatory. Accumulation of somatic mutations is one characteristic feature of malignant cells. These single nucleotide variants (SNVs) can lead to altered amino acid sequences of the translated proteins, which in turn can be presented as antigenic peptides on HLA molecules of the malignant cells. Mutated peptides represent ideal T cell targets as they are true neoantigens, specific for the tumor, and should not have been subject to central tolerance selection mechanisms. A peptide vaccination composed of mutated T cell epitopes specific for individual patient tumors is therefore a promising approach to prevent relapse in high-risk patients. For this purpose we detect nonsynonymous mutations by whole exome and transcriptome sequencing of patient leukemic blasts and normal reference tissue. HLA binding peptides harboring the altered amino acids are subsequently predicted in silico by algorithms SYFPEITHI, NetMHC and NetMHCpan for the patients' individual HLA type.
Whole exome sequencing was performed for 17 c-ALL, 2 cortical T-ALL and 1 pro-B-ALL sample pairs. ALL-specific SNVs, as well as insertions and deletions (InDels) were identified using a comparative bioinformatics pipeline. The determined variants were further validated by deep sequencing in 9/20 patients so far, with an average of 12 (+/- 8) validated mutations per patient. For all patients with validated variants, MHC class I and MHC class II epitopes could be predicted successfully.
We applied the platform for 3 patients based on compassionate need and designed individual peptide vaccines. One patient underwent haploidentical SCT with relapsed c-ALL, a second patient received autologous SCT with ependymoma and the third patient got allogeneic SCT with pro-B ALL. In all cases validated mutations could be identified and epitope prediction was performed for MHC I & II binders. The predicted peptides were synthesized and vaccination cocktails were formulated. The vaccination schedule provides 16 vaccinations over 33 weeks using GM-CSF and Imiquimod as adjuvant. The vaccination was generally well tolerated. Response to the vaccination was monitored by detection of T cells recognizing the vaccinated peptides occurring over time in peripheral blood of the patients. Monitoring was performed for each vaccination time point by prestimulation with the peptides and subsequent intracellular cytokine staining (ICS) of T cells and FACS analysis. In all 3 patients we could detect a developing CD4+ T cell response against the vaccinated mutated MHC II binding peptides.
Whole exome sequencing of pediatric ALL patients is feasible and yields a small amount of validated mutations per patients. However, these few mutations seem sufficient to predict HLA-binding peptides that are immunogenic when vaccinated and elicit specific T cell responses in patients. Moreover, the platform is not limited to ALL/leukemia but can also be applied for solid tumor patients.
Citation Format: Armin Rabsteyn, Christina Kyzirakos, Christopher Schröder, Marc Sturm, Christopher Mohr, Mathias Walzer, Ulrike Pflückhahn, Michael Walter, Magdalena Feldhahn, Karoline Laske, Michael Bonin, Martin Ebinger, Stefan Stevanovic, Peter Bauer, Oliver Kohlbacher, Cecile Gouttefangeas, Hans-Georg Rammensee, Rupert Handgretinger, Peter Lang. iVacALL: A personalized peptide-vaccination design platform for pediatric acute lymphoblastic leukemia patients based on patient-individual tumor-specific variants. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A113.
- ©2016 American Association for Cancer Research.