Abstract 3495: Tracking reversal of apoptosis after caspase activation in cancer cells
Authors: Tang Ho Lam , Tang Ho Man , Fung Ming Chiu , Hardwick J. Marie 
Published: July 2016

Using a novel caspase biosensor, we demonstrate that the late stages of apoptosis are reversible events in whole animals. Apoptosis is generally assumed to be a one-way process due to rapid activation of caspases that dismantle cellular components ending in cell suicide. Activation of apoptotic pathways is an important therapeutic strategy for treating cancer. Although some tumor types exhibit dramatic initial responses to such therapies, they inevitably recur, leading to treatment failure. One potential contributor to tumor recurrence is the failure to complete apoptotic cell death once the death pathway has been activated. We showed that cancer cells can reverse apoptosis at late stages (Anastasis, Greek for “Rising to Life”), even after passing through critical steps previously thought to be the “point of no return” such as cytochrome c release and caspases-3 activation. Simply by removing an apoptotic stimulus from cultured cells before they have completed the dying process, a high percent of cells can recover and continue to divide. We adopted the term anastasis to describe cell recovery from the brink of death. These in vitro observations suggest that anastasis may also occur in vivo, possibly allowing tumor cells to escape anti-cancer therapies. However, the major challenge to demonstrating anastasis in whole animals is that recovered cells appear morphologically indistinguishable from normal cells that never activated their cell death pathway. Therefore, we engineered the Drosophila CaspaseTracker, a novel caspase biosensor that permanently labels cells that have ever experienced caspase activation. This biosensor system is composed of 3 genetically encoded components, (1) a caspase-activatable transcription factor Gal4, and (2) G-TRACE, which is composed of Gal4-inducable red fluorescent protein (UAS-RFP), Gal4-inducable FLP recombinase (UAS-Flpase), and Flpase reporter (Ubi-LoxP-Stop-LoxP-nucGFP). Upon caspase activation, the linker tethering Gal4 in the cytoplasm is cleaved by caspases. Subsequently, Gal4 enters the nucleus where it drives the expression of FLP recombinase that removes the stop codon in nuclear-targeted GFP, robustly inducing biosensor activity in egg chambers and likely in the stem cell compartment. Following a recovery period after treatment with a death stimulus, normal healthy egg chambers reappear and continue to express biosensor activity, which is not observed in the absence of a death stimulus or with a control version of the same biosensor that has a single Asp to Ala point mutation abolishing cleavage and activation of the biosensor by caspases. Our study suggests that CaspaseTracker would be a useful tool to track reversal of apoptosis in cancer cells in live animals.

Citation Format: Ho Lam Tang, Ho Man Tang, Ming Chiu Fung, J. Marie Hardwick. Tracking reversal of apoptosis after caspase activation in cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3495.