Figure 4.

CAd12_PD-L1 MSC improves HER.2 CAR-T cytotoxicity in 3D tumor spheroids

(A) 2 x 10³ tumor cells were added to 96-well agarose-coated plates. 24 h after spheroid generation, 400 eflor 450 labeled uninfected of CAd_RFP infected MSCs were added to culture. Fluorescent images were acquired 24 h after MSC addition. Phase images are shown for tumor-only control. Scale bar represents 100 μM. (B) RFP virus detected in H1650 GFP tumor spheroids on day 2 and day 5 post MSC addition by confocal microscopy post 400 MSC addition. Scale bar represents 100 μM. RFP expression was measured overtime by Incucyte imaging system indicated by total red object integrated intensity (RCU/image). (C) 2 x 10³ cell H1650 tumor spheroids were generated. 24 h later, 200 CAd-infected MSCs were added followed by 12 NTR or 12 HER2 CAR-T cells 48 h later. Tumor cell cytotoxicity was measured by Annexin V AF594 (Red) staining overtime detected by Incucyte imaging analysis. p = 0.0186 determined by linear regression analysis, n = 4 T cell donors.