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Optimization of (CAR) T cell design guided by sensitive sssessment of B cell killing

Jul 10, 2017

Wednesday, July 12, 2017, 9 pm CEST.

Dan MacLeod Ph.D., Precision BioSciences, Inc.  

At Precision BioSciences, we have developed an allogeneic method using our proprietary ARCUS gene-editing technology to generate “universal” CAR T cells from healthy donors. These cells are engineered to eliminate expression of the endogenous T cell receptor making them safe for transfer to unrelated patients. By integrating the CAR transgene at a single genomic site, we can tightly control the level of CAR expression on the cell surface. We further optimize the cells by designing new CAR T constructs that include modifications to the extracellular binding domains as well as the intracellular signaling domains. Evaluating the impact of these new modifications requires sensitive methods to assay CAR T cell function. Traditional methods for evaluating cytotoxicity cannot effectively predict in vivo activity of a CAR T product; this is because they assess short-term killing potential rather than the ability to expand, persist, and exhibit serial killing. We have used the ACEA xCELLigence instrument for both B Cell Killing Assays and Leukemic Cell Killing Assays to test multiple CAR constructs with modified ectodomains, endodomains, and other modifications, and we have found this platform sensitive enough to detect subtle yet significant differences in performance not observed in traditional killing assays. Furthermore, the sensitivity of the instrument has enabled us to evaluate CAR constructs using cell numbers that are orders of magnitude lower than other assays, greatly enhancing our throughput and capabilities.

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Related technologies: Real-time, label free cell analysis

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Riccardo Pasculli

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Agilent technologies

Agilent provides xCELLigence impedance-based, label-free, real time cell analysis system and NovoCyte flow cytometers.

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Located inside a tissue culture incubator, the RTCA DP Analyzer has three integrated stations for E-Plates 16 or CIM-Plates 16

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Located inside a tissue culture incubator, it is capable of switching any one of the wells on the E-Plate 96 to the RTCA Analyzer for impedance measurement

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Located inside a tissue culture incubator, it is capable of switching any one of the wells on any of six E-Plates to the RTCA Analyzer for impedance measurement

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Instrument can be integrated with automated liquid handling platforms or a robotic plate handler from different vendors

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