Development of in vitro 3D assays for functional evaluation of immunogenic cell death

Abstract

In vitro evaluation of cancer drug efficacy is an important process to screen cancer drug candidates. Conventional cancer drug screening methods based on the measurement of cancer cell cytotoxicity using 2-dimensional (2D) culture can provide limited information as cancer drug can not only trigger cancer cell death but also initiate anti-cancer immune responses by inducing immunogenic cell death (ICD) of cancer cells. New drug evaluation methods that can assess the effects of cancer drugs on ICD as well as cancer cell death need to be developed. As a first step to address this issue, we developed a 3-dimensional (3D) platform that allow monitoring dynamic cancer cell-dendritic cell (DC) interactions with cancer drug treatment. DCs are immune cells that engulfs dying/dead cancer cells to initiate anti-cancer immune responses, thus interactions between cancer cells and DCs critical to evaluating ICD. To develop this platform, we first tested various cytotoxicity assays and identified 2D and 3D IC50s of various cancer drugs, of which ~ 50% of cancer cells die, for cancer cells and DCs. Then, cancer cells in 3D collagen gels were treated with various concentrations of cancer drugs, and DCs were added to sides of collagen gel blocks. DCs were recruited into the collage gels, presumably by ATP released from dying cancer cells, and interacted with cancer cells to engulf them. DC migration and interaction with cancer cells were monitored by performing live cell imaging. This 3D platform will be useful in evaluating ICD of cancer drugs and studying basic mechanisms of ICD.