Figure 6 from Human 3D Ovarian Cancer Models Reveal Malignant Cell–Intrinsic and –Extrinsic Factors That Influence CAR T-cell Activity

Inhibition of TGFβ signaling reduced ECM density and stimulated CAR T-cell activity in G164/FB gels. A, M3C, COL1A1, and FN1 staining and quantification of G164/FB gels treated with TGFβR inhibitor (TGFβR inhi). Scale bar, 50 μm. B, TWOMBLI analysis showing the heatmaps and %HDM for M3C, COL1A1, and fibronectin (FN1) in G164/FB gels treated with TGFβR inhibitor. Scale bar, 100 μm. C, Number of CD3⁺ CAR T cells at the peripheral and core regions of G164/FB collagen gels treated with TGFβR inhibitor. Two different CAR T-cell and FB donors were used for this experiment. D, Heatmaps generated using Definiens Tissue Studio showing the distribution of CAR T cells in G164/FB collagen gels treated with TGFβR inhibitor. Representative images from two/three gels per two replicates. Scale bar, 500 μm. E, IHC staining and quantification for caspase-3 (Casp3) on G164/FB gels treated with TGFβR inhibitor. Scale bar, 50 μm. F, Expression of FAP on G164/FB gels treated with TGFβR inhibitor. Scale bar, 50 μm. G, Behavior of CAR T cells in G164/FB gels treated with TGFβR inhibitor. Representative images from three repeats. Blue, FN1; green, CAR T cells; red, EpCAM. Scale bar, 50 μm. Speed and length of movement of migrating (vehicle, 83; TGFβR inhibitor, 62) CAR T cells. H, Number of CD3⁺ (left) and Casp3⁺ (right) on G164/FB gels treated with TGFβR inhibitor and aCCL2. A–C, E, F, and H, Data plotted as mean ± SD of two/three gels per two replicates. Statistics performed using unpaired t test (A, B, and F) and two-way ANOVA (C, E, and H).