Immunotherapy has emerged as a promising approach to treating several forms of cancer. Use of immune cells, such as natural killer (NK) cells, along with small molecule drugs and antibodies through antibody dependent cell-mediated cytotoxicity (ADCC) has been investigated as a potential combination therapy for some difficult to treat solid tumors.
Immunotherapy has emerged as a promising approach to treating several forms of cancer. Use of immune cells, such as natural killer (NK) cells, along with small molecule drugs and antibodies through antibody dependent cell-mediated cytotoxicity (ADCC) has been investigated as a potential combination therapy for some difficult to treat solid tumors. Nevertheless, there remains a need to develop tools that support co-culture of target cancer cells and effector immune cells in a contextually relevant three-dimensional (3D) environment to provide a rapid means to screen for and optimize ADCC-drug combinations. To that end, we have developed a high throughput 330 micropillar-microwell sandwich platform that enables 3D co-culture of NK92-CD16 cells with pancreatic (MiaPaCa-2) and breast cancer cell lines (MCF7 and MDA-MB-231). The platform successfully mimicked hypoxic conditions found in a tumor microenvironment (Figure 1) and was used to demonstrate NK-cell mediated cell cytotoxicity in combination with two monoclonal antibodies; Trastuzumab and Atezolizumab (Figure 2). The platform was also used to show dose response behavior of target cancer cells with reduced EC50 values for paclitaxel (an anti-cancer chemotherapeutic) when treated with both NK cells and antibody. Such a platform may be used to develop more personalized cancer therapies using patient-derived cancer cells. [Gopal et al. Commun. Biol, 4, 893, 2020].
Figure 1. Generation of 3D tumor spheroid micropillar array. (a) Photo of 330-micropillar/well chip for effector cell-mediated cytotoxicity. Diameters of micropillar and microwell array spots are 1 and 1.9 mm, respectively. (b) Generation of 3D tumor spheroid by printing high density cells (~2500 cells/250 nL) in Matrigel, followed by culturing cancer cells for up to 8 days. (c) Expression of HIF-1α inside MiaPaCa-2 3D tumor spheroids. Confocal microscopy images of MiaPaCa-2 3D tumor spheroids stained with Hoechst 33342 and labeled with HIF-1α antibody via immunofluorescence. Top views of cross-sectional images in a 3D tumor spheroid at different focal planes (top, middle, and bottom) using Imaris software. A green arrow symbol orients the viewing direction for the cross-sections reconstructed from Z stacks. (d) 3D view of a 3D tumor spheroid after staining nucleus and labeling HIF1α.
Figure 2. High-content images of 3D tumor spheroid micropillar array using the 330-micropillar/microwell chip sandwich platform. (a) Scanned images of the chip containing tumor spheroids including MCF-7, MiaPaCa-2, and MDA-MB-231 spheroids after treating NK92-CD16 cells combining with doxorubicin (DOX), paclitaxel (PTX), and Trastuzumab (TRA). (b) Cytotoxic activity of NK92-CD16 cells (effector) combining with DOX, PTX, and Trastuzumab was quantified by live 3D tumor spheroid (target) images stained with Calcein-AM. Percent cytotoxicity was calculated by NK cell-mediated killing / maximum killing by Saponin. (c) Magnified images of 3D tumor spheroids after treating with NK92-CD16 cells (E:T = 10 : 1) in comparison with control (no treatment of NK92-CD16 cells). Error bars indicate Mean ± SD for three different replicates. *p ≤ 0.05, **p ≤ 0.01,***p ≤ 0.001, ****p ≤ 0.0001, ns-not significant (unpaired t-test)
Current Collaborator:
Bosung Ku – MBD Korea, Ltd.
