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(A) hMOs were grown for three to seven days in growth medium followed by 24 h treatment with 10 µM of BV6, Birinapant (Bi), ASTX-660 (ASTX) and LCL-161 (LCL), respectively, or DMSO as vehicle control. Subsequently, CellTiter-Glo® viability assays were performed and the values were normalized to non-treated controls. Error bars represent standard deviation. * p < 0.05, ** p< 0.01, *** p < 0.005. (B) hMOs, treated as in (A) were harvested and Western blotting was performed with antibodies recognizing <t>cIAP1,</t> cIAP2, XIAP as well as total and cleaved caspase-3. Vinculin served as loading control. Representative blots of three independent experiments are shown.
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(A) hMOs were grown for three to seven days in growth medium followed by 24 h treatment with 10 µM of BV6, Birinapant (Bi), ASTX-660 (ASTX) and LCL-161 (LCL), respectively, or DMSO as vehicle control. Subsequently, CellTiter-Glo® viability assays were performed and the values were normalized to non-treated controls. Error bars represent standard deviation. * p < 0.05, ** p< 0.01, *** p < 0.005. (B) hMOs, treated as in (A) were harvested and Western blotting was performed with antibodies recognizing cIAP1, cIAP2, XIAP as well as total and cleaved caspase-3. Vinculin served as loading control. Representative blots of three independent experiments are shown.

Journal: bioRxiv

Article Title: Inhibition of IAPs induces programmed cell death and inflammatory signaling in patient-derived metastatic breast cancer organoids

doi: 10.1101/2024.08.28.610103

Figure Lengend Snippet: (A) hMOs were grown for three to seven days in growth medium followed by 24 h treatment with 10 µM of BV6, Birinapant (Bi), ASTX-660 (ASTX) and LCL-161 (LCL), respectively, or DMSO as vehicle control. Subsequently, CellTiter-Glo® viability assays were performed and the values were normalized to non-treated controls. Error bars represent standard deviation. * p < 0.05, ** p< 0.01, *** p < 0.005. (B) hMOs, treated as in (A) were harvested and Western blotting was performed with antibodies recognizing cIAP1, cIAP2, XIAP as well as total and cleaved caspase-3. Vinculin served as loading control. Representative blots of three independent experiments are shown.

Article Snippet: The following antibodies were used in this study: α-Vinculin (V9131, Sigma), α-RIPK1 (610459, BD Biosciences), α-phospho-RIPK1 S166 (657465, Cell Signaling Technologies), α-RIPK3 (13526, Cell Signaling Technologies), α-phoshpo-RIPK3 S227 (ab209384, Abcam), α-MLKL (14993, Cell Signaling Technologies), α-phospho-MLKL S358 (91689, Cell Signaling Technologies), α-caspase-3 (9662S, Cell Signaling Technologies), α-cleaved-caspase-3 (9661, Cell Signaling Technologies), α-cIAP1 (AF8181, R&D Systems), α-cIAP2 (3130, Cell Signaling Technologies), α-XIAP (610716, BD Biosciences).

Techniques: Control, Standard Deviation, Western Blot