Abstract
Cell-based immunotherapy has achieved preclinical success in certain types of cancer patients, with a few approved cell-based products for clinical use. These achievements revitalized the field of cell engineering/ immunotherapy and brought attention to the opportunities that cell-based immunotherapeutics can offer to patients. On the other hand, obvious indications emphasize the need for a better understanding of the biological mechanisms involved in the immune response. This knowledge may not only ameliorate safety and efficacy, but also determine the possibilities and limitations in use of immune cell engineering for cancer treatment, and facilitate developing novel immunotherapeutic strategies. Recently developed technology based on CRISPR-dCas9 has an immense potential to systematically uncover genetic mechanisms by identifying subsets of essential genes involved in interactions of cancer cells with the immune system. This chapter will present a reliable and reproducible general protocol for the application of genome-wide sgRNA gene-editing tools in the recently established two-cell type co-culture, consisting of immune cells as effectors and cancer cells as targets, utilizing CRISPRi/a-dCas9-based technology.
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Acknowledgments
We thank Dr. Jonathan Weissman at UCSF for providing research materials and expert advice. This work was supported by the Yale Cancer Center, NIH R21CA198561, NIH R21AI121993, and the Alliance for Cancer Gene Therapy.
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Zhang, J., Späth, S.S., Katz, S.G. (2020). Genome-Wide CRISPRi/a Screening in an In Vitro Coculture Assay of Human Immune Cells with Tumor Cells. In: Katz, S., Rabinovich, P. (eds) Cell Reprogramming for Immunotherapy. Methods in Molecular Biology, vol 2097. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0203-4_15
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DOI: https://doi.org/10.1007/978-1-0716-0203-4_15
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