MIL-99

The 3D co-culture model we describe was developed to study the potency of immune checkpoint inhibitors. In the assay, the first step is to provide 3D cell growth of endometrial carcinoma RL95-2 cells on a hydrogel, LifeGel ® , and then to co-culture cancer cells with PBMCs isolated from healthy blood donors. After 5 days of the co-culture, in the presence of anti-PD-1/PD-L1 or anti-VISTA immune checkpoint inhibitors, the cancer cells’ viability is measured using a colorimetric assay and fluorescence readout. The key features of the presented model are (1) a direct 3D co-culture model that involves cancer cells and immune cells represented by PBMCs, (2) the use of LifeGel ® hydrogel, where cancer cells form spheroids on the surface, allowing immune cells and larger molecules, such as monoclonal antibodies, to penetrate the cancerous structures, and (3) the possible reactivation of immune cells by immune checkpoint inhibitors may be observed via killing/destroying effects on cancer structures.