Figure 3

OSI-906 mediates apoptosis by a TGFβ/PKA dependent manner. FET cells treated with OSI-906 (1 μM) activated Caspase 3/7. Caspase 3 inhibitor C3i (20 μM) was used to inhibit Caspase 3 activation (A). Caspase 3/7 activation by OSI-906 is PKA dependent as activation of Caspase 3/7 was abrogated with prior treatment of FET cells with PKA inhibitor H89 (15 μM) (B). FET cells treated with TGFβRI inhibitor ALK5i (400 nM) showed abrogation of Caspase 3/7 activation. Pretreatment with ALK5i followed by OSI-906 showed similar results (C). Activation of PKA by OSI-906 was abrogated in the presence of 25 μM AKAP inhibitor Ht31 (D). Caspase 3/7 activation by OSI-906 is AKAP dependent as OSI-906 mediated Caspase 3/7 activation was abrogated with prior treatment of FET cells with Ht31 (25 μM) (E). All experiments were repeated three times independently with 3 replicas per repeat for the ANOVA analysis (* P < 0.001).