Fig. 5
From: Tailoring a novel colorectal cancer stem cell-targeted therapy by inhibiting the SMYD3/c-MYC axis
SMYD3 as a potential therapeutic target in CRC-SCs. a Quantification of cell viability by CellTiter-Glo in patient-derived CRC-SCs (#C108 and #C109) treated or not with EM127 (10 μM) for 72 h. b Quantification of cell death by Trypan blue staining in patient-derived CRC-SCs (#C108 and #C109) treated or not with EM127 (10 μM) for 72 h. c Invasive ability of growth factor-starved patient-derived CRC-SCs (#C108 and #C109) placed in the inner chamber of transwell plates and treated or not with EM127 (10 μM) for 16 h. Invading cells were fixed and counted under a fluorescence microscope. Scale bar: 200 μm. d Colony-forming ability of patient-derived CRC-SCs (#C108 and #C109) seeded onto double-layer soft agar and treated or not with EM127 (10 μM) for 72 h. Data represent the area of the evaluated colonies during four weeks. e Flow cytometry analysis (left panel) and corresponding bar plot representation (right panel) of Ki67 expression in patient-derived CRC-SCs (#C108 and #C109) treated or not with EM127 (10 μM) for 72 h. f Flow cytometry analysis (left panel) and corresponding bar plot representation (right panel) of annexin V staining in patient-derived CRC-SCs (#C108 and #C109) treated or not with EM127 (10 μM) for 72 h. The graph summarizes the percentage of apoptotic cells (early + late). g Immunoblot analysis of Cleaved PARP, Cleaved Caspase 3, and c-MYC K158/K163Me levels in patient-derived CRC-SCs (#C108 and #C109) treated or not with EM127 (10 μM) for 72 h. GAPDH was used as a loading control. h−n Characterization of PDTOs treated or not with EM127 (10 μM) for 72 h. PDTO brightfield imaging and live and dead staining (green: live cells; red: dead cells) (h, scale bar: 200 μm), number (i), area (j), relative intensity of red fluorescence (k), hematoxylin and eosin (H&E) staining (l, scale bar: 50 μm), TUNEL staining (green) with nuclei counterstained with DAPI (blue) (m, scale bar: 50 μm), and ddPCR analysis for stemness-related c-MYC target genes (n). *p < 0.05 EM127-treated vs untreated. Where applicable, data are expressed as means ± SD of 3 independent experiments
