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• W3215501982 abstract "Abstract Background Most high-throughput screening (HTS) systems for the cytotoxic effect of chimeric antigen receptor (CAR) T cells on tumor cells rely on two-dimensional cell culture that does not suitably recapitulate the tumor microenvironment (TME). Tumor spheroids can recapitulate TME and have been used for cytotoxicity assays of CAR T cells. However, a major obstacle to the use of tumor spheroids for cytotoxicity assays is the difficulty in separating unbound CAR T and dead tumor cells from spheroids. Here, we present a three-dimensional hanging spheroid plate (3DHSP), which facilitates the formation of spheroids and the separation of unbound and dead cells from spheroids during cytotoxicity assays. Results The 3DHSP is a 24-well plate, with each well composed of a hanging dripper, spheroid wells, and waste wells. In the dripper, a tumor spheroid was formed and mixed with CAR T cells. In 3DHSP, droplets containing the spheroids were deposited into the spheroid separation well, where unbound and dead T and tumor cells were separated from the spheroid through a gap into the waste well by tilting the 3DHSP by more than 20°. Human epidermal growth factor receptor 2 (HER2)-positive tumor cells (BT474 and SKOV3) formed spheroids of approximately 300–350 mm in diameter after 2 d in the 3DHSP. The cytotoxic effects of T cells engineered to express CAR recognizing HER2 (HER2-CAR T cells) on these spheroids were directly measured by optical imaging, without the use of live and dead fluorescent staining of the cells. Our results suggest that the 3DHSP could be incorporated into an HTS system to screen for CARs that enable T cells to kill spheroids formed from a specific tumor type with high efficacy or for spheroids consisting of tumor types that can be killed efficiently by T cells bearing a specific CAR. Conclusions The results suggest that the 3DHSP could be incorporated into an HTS system for the cytotoxic effects of CAR T cells on tumor spheroids." @default.
• W3215501982 created "2021-12-06" @default.
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• W3215501982 date "2021-11-23" @default.
• W3215501982 modified "2023-09-23" @default.
• W3215501982 title "3D Hanging Spheroid Plate for High-Throughput CAR T Cell Cytotoxicity Assay" @default.
• W3215501982 doi "https://doi.org/10.21203/rs.3.rs-1087944/v1" @default.
• W3215501982 hasPublicationYear "2021" @default.
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