Introduction: Dendritic cell (DC) vaccines currently in phase III trials for the treatment of glioblastoma use autologous tumor lysate (ATL) as an antigen source but it remains unclear what the targets of this personalized therapy are directed against in each patient. We hypothesized that the target antigens are truly tumor-specific, such as mutations and gene fusions found in tumor that create stable peptide-MHC complexes compared with the normal sequences.
Methods: We performed RNA sequencing on the GL261 mouse glioma cell line. Mutations were identified with the Missense Mutation and Frameshift Location Reporter (MMuFLR) software developed by our group and analyzed by NetMHC3.4 to predict which mutated sequences have enhanced peptide-MHC binding. 9 such candidate neoantigens were identified and were confirmed by IGV and Sanger sequencing. To evaluate whether ATL DC vaccination induced neoantigen-specific T cell responses, tumor-infiltrating lymphocytes (TILs) were isolated from GL261 tumor-bearing, ATL DC vaccine-treated mice by percoll gradient. TILs were co-cultured with DCs pulsed with peptides containing the neoantigen sequences and subjected to flow cytometry and cytotoxicity analysis.
Results: 5x10^5-2x10^6 TILs were recovered from vaccine-treated animals, whereas <10^5 were recovered from controls, indicating a T cell response to the vaccine. TILs antitumor activity as measured by the xCELLigence cytotoxicity assay. TILs were co-cultured with DCs pulsed with synthetic peptides containing the neoantigen sequences. 15-25% of tumor-infiltrating CD8+ T cells from tumor lysate-pulsed DC vaccine-treated mice upregulated CD69, an early marker of T cell activation in an antigen-specific manner. These cells also demonstrated elevated expression of PD-1, a marker of antigen-experienced cells.
Conclusions: Together, these studies indicate that in this murine model, autologous tumor lysate DC vaccines are capable of priming CD8+ T cell responses directed at tumor-specific mutations. Further studies will evaluate these findings in glioblastoma patients treated with DC vaccines.
Patient Care: It helps elucidate the mechanisms by which a promising new treatment for glioblastoma works, which may in turn, lead to improvements in the creation of future treatments for this devastating disease.
Learning Objectives: By the conclusion of this session, participants should be able to:
1. Explain the rationale for immunotherapy with autologous tumor lysate dendritic cell vaccination as a treament for glioblastoma
2. Discuss the scientific underpinnings of how the treatment might work, namely targeting individual, tumor-specific mutations.
References: 1. Rathe SK, Johnson JE, Silverstein KA, Erdmann JJ, Watson AL, Popescu FE, et al. MMuFLR: missense mutation and frameshift location reporter. Bioinformatics. 2013;29:2353-4.