Figure 6.

Cytotoxic response triggered by CSF-470 vaccination in pt#006 PBMC samples. Pt#006-T cells lysis was evaluated by the in vitro calcein release assay as described under Methods. As effector cells, autologous PBMC were stimulated during 12 days with CSF-470 vaccine lysate (A) autologous tumor lysate (B), both in a 3:1 ratio (PBMC: tumor cell lysate), a pool of the 9 identified neoAgs peptides (C) or the same pool of 9 neoAgs plus OKT3 and a feeder layer, as described under Methods (D); every peptide was tested at a 10 μg/ml concentration. 5 × 10⁴ target cells/well were seeded in 96-well plates. Different effector: target ratios were tested in quadruplicate. (E) T cells stimulated by the autologous tumor cell lysate were analyzed by flow cytometry at days 0 and 11 after stimulation and prior to the Pt#006-T cells lysis assay, to identify CD4+ and CD8+ T cell's phenotype: activated effector cells (CD45RO⁻HLA DR+), memory subsets: effector memory (CD45RO⁺CCR7–); central memory (CD45RO+ CCR7+) cells and effector exhausted (PD-1+) cells in the different PBMC samples (PRE-P3).