CRISPR Screening of CAR T Cells and Cancer Stem Cells Reveals Critical Dependencies for Cell-Based Therapies

• Published in: Cancer discovery 2021-05, Vol.11 (5), p.1192-1211
• Main Authors: Wang, Dongrui, Prager, Briana C, Gimple, Ryan C, Aguilar, Brenda, Alizadeh, Darya, Tang, Hongzhen, Lv, Deguan, Starr, Renate, Brito, Alfonso, Wu, Qiulian, Kim, Leo J Y, Qiu, Zhixin, Lin, Peng, Lorenzini, Michael H, Badie, Behnam, Forman, Stephen J, Xie, Qi, Brown, Christine E, Rich, Jeremy N
• Format: Article
• Language: English
• Subjects: Brain Neoplasms - genetics; Brain Neoplasms - pathology; Cell Line, Tumor; Cell- and Tissue-Based Therapy; Clustered Regularly Interspaced Short Palindromic Repeats; Glioblastoma - genetics; Glioblastoma - pathology; Humans; Neoplastic Stem Cells - metabolism; Receptors, Chimeric Antigen - genetics
• ISSN: 2159-8274; 2159-8290
• DOI: 10.1158/2159-8290.CD-20-1243

Glioblastoma (GBM) contains self-renewing GBM stem cells (GSC) potentially amenable to immunologic targeting, but chimeric antigen receptor (CAR) T-cell therapy has demonstrated limited clinical responses in GBM. Here, we interrogated molecular determinants of CAR-mediated GBM killing through whole-genome CRISPR screens in both CAR T cells and patient-derived GSCs. Screening of CAR T cells identified dependencies for effector functions, including TLE4 and IKZF2. Targeted knockout of these genes enhanced CAR antitumor efficacy. Bulk and single-cell RNA sequencing of edited CAR T cells revealed transcriptional profiles of superior effector function and inhibited exhaustion responses. Reciprocal screening of GSCs identified genes essential for susceptibility to CAR-mediated killing, including RELA and NPLOC4, the knockout of which altered tumor-immune signaling and increased responsiveness of CAR therapy. Overall, CRISPR screening of CAR T cells and GSCs discovered avenues for enhancing CAR therapeutic efficacy against GBM, with the potential to be extended to other solid tumors. SIGNIFICANCE: Reciprocal CRISPR screening identified genes in both CAR T cells and tumor cells regulating the potency of CAR T-cell cytotoxicity, informing molecular targeting strategies to potentiate CAR T-cell antitumor efficacy and elucidate genetic modifications of tumor cells in combination with CAR T cells to advance immuno-oncotherapy. .