In Vitro Cell Death Determination for Drug Discovery: A Landscape Review of Real Issues

Méry, Benoite; Guy, Jean-Baptiste; Vallard, Alexis; Espenel, Sophie; Ardail, Dominique; Rodriguez-Lafrasse, Claire; Rancoule, Chloé; Magné, Nicolas

Resistance to death is a major characteristic of cancer cell, hence the emergence of considerable interest in developing therapeutic strategies that could interfere with cellular death decisions. Cell death plays a major role in the homeostasis of tissues and allows the elimination of transformed cells. The deregulation of cell death in cancer implies that effective drugs have to kill cells outright and not only inhibit their growth to provide complete clinical responses. Besides, resistance to cell death during tumorigenesis may also explain tumor relapse following chemotherapeutic treatment. Thus, cell death-related assays are of primary importance in the field of oncology for drug discoveries. 1 Obviously, cell death is not unequivocal and can occur through many options. 2 Beyond semantic reasons, the ability to distinguish all forms of cell death is also clinically relevant to develop specific therapeutic drugs suitable for every cell death processes. Indeed, as the response to anticancer therapy is modulated by certain forms of cell death, specific modalities of cell death have to be identified by drug discovery assays. 3 Consequently, different in vitro methods for identifying dead cells in cell cultures have been described, including detection of end-stage cell death, irrespective of its type, as well as processes associated with a particular subroutine cell death. 4 If apoptosis, also referred to as programmed cell death, is considered as the most appropriate indicator of cell death, other categories of cell death pathways, including necrosis, autophagic cell death, and anoikis, should be of interest. As a matter of fact, if a recent classification has defined 8 different forms of cell death which may involve various molecular signaling pathways, some researchers describe that at least 11 pathways of cell death exist and that apoptotic cells frequently undergo secondary necrosis under in vitro conditions. 5, 6 It appears that apoptosis and necrosis may represent the 2 extreme types of cell death on a wide range of cell death modes. Therefore, it might be difficult to distinguish all of these specific death conditions monolithically, as differences may not be as distinct as it appears. Furthermore, appropriate cell death assays must be rigorously selected for the experimental setting under investigation; choosing the right assay tool and the appropriate end point to measure cell death is a condition sine qua non to generate meaningful data and remains a challenging task. 7 Indeed, cell death can be approached from a variety of viewpoints, including measurement of dead cells (cytotoxicity assay), quantification of live cells (viability assay), and, ultimately, determination of the accurate mechanism involved. Therefore, the plethora of strategies for monitoring cell death requires a selective choice of the best suited cell death assay according to the study aim, but a single method may not be sufficient to determine a type of cell death. 8 This review provides an overview of cell death assays currently used in vitro for estimation of cell death so as to assess Journal of Cell Death new potential chemotherapeutics and discusses the ins and outs of these scientific approaches.