A kinome inhibitor screen implicates adhesion and growth factor signaling in cellular recovery after caspase activation

Abstract

Apoptosis is a common form of regulated cell death and requires cysteine-aspartic proteases called effector caspases. Caspase activation triggers positive feedback, leading to the idea that apoptosis is irreversible. However, we and others have demonstrated that cancer cells can survive effector caspase activation and become more aggressive and drug-resistant as a result. Despite the profound implications of apoptotic reversal, also known as anastasis, for both regenerative medicine and cancer therapy, the molecular pathways that enable cells to survive executioner caspase activation remain largely unmapped. To systematically dissect this phenomenon, we developed a quantitative screening platform that combines inducible caspase activation with kinome-wide pharmacological profiling. This approach uniquely allowed us to: (1) identify pharmacological modulators of post-caspase survival, (2) identify specific kinases regulating post-caspase survival, and (3) distinguish general toxicity from anastasis effects. This approach implicated regulators of cell adhesion and the cytoskeleton, consistent with the known rounding of apoptotic cells and respreading during recovery. Growth factor signaling also emerged from the analysis. In addition to its expected effects on unstressed cells, fetal bovine serum markedly increased anastasis. Some growth factor combinations were more effective than individual ones at recapitulating the serum effect. Similarly, pleiotropic kinase inhibitors were generally more effective than selective ones. Nevertheless, selective Rho kinase inhibition significantly enhanced anastasis whereas Akt inhibition impaired it, suggesting that these kinases serve as central nodes that integrate multiple upstream inputs. Beyond identifying specific kinase targets, our work provides a framework for anti-anastasis therapies that could prevent cancer cell recovery after chemotherapy.