Figure 5.

A MOS potency assay for T-Cell therapies. (A) TILs cannot penetrate traditional Matrigel. (B) TILs can penetrate MOS and adhere to tumor cells. Immune cells were stained with Cytolight Red dye before images taken using Incucyte. (C) Increased killing indicated by Annexin V was observed in MOS treated with autologous TILs. (D) Representative images of MOS killing by TILs in MOS (indicated by Annexin V dye). (E) Activated PBMCs induce MOS killing (indicated by Annexin V Green dye). (F) Representative images of MOS killing by PBMCs. (G) Representative images to illustrate an imaging analysis pipeline that identifies droplet area to minimize background noise from outside immune cells. (H) Quantification analysis suggest PBMCs induce MOS killing (indicated by Caspase 3/7 dye). (I) ESK1* enhanced PBMC-induced tumor cell killing compared to the DP47 (CD3 only TCB). (J) Representative images of induced death of ESK1* treated MOS combined with PBMCs. White arrows indicating lung cancer tumorspheres within MOS. Compared to ESK1*, the negative control TCB, DP47, did not induce significant apoptosis of tumorspheres within MOS. (K) Dotted plot suggests ESK1* induces PBMC-mediated lung tumor MOS death in seven patient cases (p<0.005). (M) Image of dsRed expressing vector infection on MOS derived from lung tumor (3 days post infection). Significant higher gene expression of HLA-A2 (N) and antigen expression (O) were observed in HLA-A2-infected MOS. (P) HLA-A2-infected MOS underwent higher cell death than matched uninfected MOS in the presence of ESK1* and activated PBMCs (as indicated by Annexin V dye).