Figure 1.

Oncolytic virotherapy elicits specific CD8 T-cell responses against a defined panel of neoepitopes in PD-1-resistant CMT64 tumors. (a) Work flow for prediction and identification of neoepitope-specific CD8 T-cell responses in CMT64 cells. (b) C57BL/6 mice with established s.c. CMT64-tumors were treated twice with αPD-1 antibody (days 0 and 3) or with intratumoral injection of hTertAd. Tumor-free and tumor-bearing mice without treatment served as controls. Splenocytes were harvested 1 week after the first treatment and used for analysis of neoepitope-specific CD8 T-cell responses. For this screen, 44 minimal epitopes from CMT64 sequencing were synthesized as minimal peptides to detect neoepitope-specific responses in IFNγ-ELISpot assays. CD8 T-cell responses observable in all investigated individuals are marked by an asterisk. Responses that were restricted to a subset of mice are marked with “s”. The graph shows representative results from three mice of a total number of eight (n = 8 per group). (c) Ndufs1-V491A-specific CD8 T-cell responses were determined in tumor-free mice treated with a PD-1 blocking antibody and compared to responses in CMT64-tumor-bearing mice (n = 5 per group). Irrelevant peptides served as control. (d) To investigate the ability of neoepitope-specific CD8 T-cell responses to distinguish between tumor-associated neoepitopes and corresponding wild-type epitopes, splenocytes from mice after virotherapy treatment were stimulated with mutated epitope motifs or with corresponding wild-type peptides, respectively. Pooled splenocytes were used for analysis (n = 5 animals). (e) Tabular overview containing a list of detected virotherapy-responsive neoepitopes and additional key information.