Table 1.
CRISPR screens in T cells to identify genes for cancer immunotherapy
| Conditions | Cells | Libraries | Methods | Purpose | Hit genes | References |
|---|---|---|---|---|---|---|
| ex vivo | IL13Rα2-targeted CAR T cells | Addgene #73179 | CRISPR loss of function screens | CAR-T effector activity, cytotoxic potency | TLE4 IKZF2 | Wang, D., et al. (2021)[113] |
| ex vivo | Primary human T cells | Addgene #73178 | CRISPR loss of function screens | T cell proliferation | SOCS1 TCEB2 RASA2 CBLB | Shifrut, E., et al. (2018)[114] |
| ex vivo | Primary human T cells | lentiSAM v2 | Coupled CRISPR activation and CRISPR interference screen | Gene networks controlling IL-2 and IFN-g production | VAV1 CD28 LCP2 LAT MALT1 BCL10 TRAF6 | Schmidt, R., et al. (2022)[115] |
| ex vivo | Primary mouse CD8+ T cells | mm10dgLib | dead-guide RNA (dgRNA)-based CRISPR activation screen | CAR-T killing and in vivo efficacy in cancers | PRODH2 | Ye, L., et al. (2022)[116] |
| in vivo | P14-transgenic Cas9-expressing CD45.1+ LSK cells (mouse) | A library of 110 sgRNAs targeting 21 genes relevant to T cell biology and 50 non-targeting control sgRNAs | CHIME: CHimeric IMmune Editing, | CD8+ T cell responses to LCMV Clone 13 infection | Ptpn2 | LaFleur, M. W., et al. (2019)[96] |
| in vivo and ex vivo | Primary mouse CD8+ T cells | The genome-scale mouse T cell CRISPR library (MKO) | CRISPR loss of function screens | T cell activation, tumour infiltration and degranulation | Dhx37 | Dong, M. B., et al. (2019)[117] |
| in vivo | Primary mouse CD8+ T cells | sgRNA library targeting 1658 genes, with 6628 sgRNAs and 1000 nontargeting controls | AAV–SB-CRISPR screens | T cells effector functions, and potent of killing tumour cells | Lag3 Mgat5 Pdia3 Emp1 | Ye, L., et al. (2019)[118] |
| ex vivo | Primary human CD4+ T cells | Human genome-wide sgRNA library targeting 18,360 protein-coding genes | Guide Swap pooled CRISPR enrichment and depletion screens | Validation of guide swap in depletion and enrichment screens in CD4 + T cells | MDM2 WEE1 EEF2 TOP2A | Ting, P. Y., et al. (2018)[119] |
| ex vivo | 68–41 murine T cell line | Lentiviral gRNA library (87,898 gRNAs/19,150 genes) | CRISPR loss of function screens | T cell activation | Fut8 | Okada, M., et al. (2017)[120] |
| ex vivo | Primary mouse T helper cells | Addgene #67988 | CRISPR loss of function screens | T helper cell activation and differentiation | PPARG | Henriksson, J., et al. (2019) [121] |
| in vivo | Mouse CD8+ T cells | Pooled gRNA library of 3017 metabolism-associated genes | CRISPR loss of function screens | CD8+ T cell fate decision | Pofut1 | Huang, H., et al. (2021)[122] |
| in vivo | Mouse CD8+ T cells | Two lentiviral sub-libraries of single-guide sgRNAs targeting 3017 metabolic enzymes, small molecule transporters and metabolism-related transcriptional regulators | CRISPR loss of function screens | T cell longevity and antitumour response | REGNASE-1 | Wei, J., et al. (2019)[123] |
| in vivo | Antigen-experienced mouse CD4+ T cells | A genome-wide knockout (GWKO) sgRNA lentiviral library (18,400 genes, 90,000 sgRNAs) | CRISPR loss of function screens | The expansion of activated CD4+ T cells | SOCS1 | Sutra Del Galy, A., et al. (2021)[97] |
| in vivo | Primary mouse CD8+ T cells | Retroviral sgRNA library targeting 271 transcription factors | CRISPR loss of function screens (OpTICS) | T cell effector activity | Fli1 | Chen, Z., et al. (2021)[124] |
| in vivo and ex vivo | Chronic stimulated CD8+ T cells | Addgene #104861 | CRISPR loss of function screens | T cell exhaustion | Arid1a | Belk, J. A., et al. (2022)[125] |
| ex vivo | Jurkat T cells | sgRNA library consists of over 250,000 total sgRNAs targeting every unique Refseq annotated (hg19) protein coding isoform with up to 12 sgRNAs, plus 7700 NTCs | CRISPR loss of function screens | T cell activation | FAM49B | Shang, W., et al. (2018)[126] |
| ex vivo | Human CD4+ and CD8+ T cells | Lentiviral library of human ORFs containing nearly 12,000 full-length genes | CRISPR gain of function screens | T cell proliferation | LTBR | Legut, M., et al. (2022)[127] |
| in vivo | Mouse OT-1 CD8+ T cells | scCRISPR library containing 180 TFs with 720 sgRNAs and 80 NTC sgRNAs | CRISPR loss of function screens | CD8+ cytotoxic T cells differentiation and Tex cell reinvigoration | IKZF1 ETS1 | Zhou, P., et al. (2023)[128] |
| ex vivo | Primary human CD8+ T cells | Published highly optimized library that encodes gRNAs and Cas9 (Wang et al. 2017) | CRISPR loss of function screens | T cell exhaustion | SNX9 | Trefny, M. P., et al. (2023)[129] |
| ex vivo | Primary mouse CD8+ T cells | sgRNA library from Benchling or Brie, adding up to a total of 2298 sgRNAs | CRISPR loss of function screens | T cell adhesion, migration, homeostasis | RASA3 | Johansen, K. H., et al. (2022)[130] |
| ex vivo | Primary human T cells | Brunello sgRNA library | CRISPR loss of function screens (SLICE) | T cell dysfunction | RASA2 | Carnevale, J., et al. (2022)[131] |
| ex vivo | Primary mouse Treg cells | A library of 489 nuclear factors on the basis of optimized sgRNA sequences from the Brie library | CRISPR loss of function screens | Treg instability | Usp22 Rnf20 | Cortez, J. T., et al. (2020)[98] |
| in vivo | Primary mouse CD4+ T cells | Addgene, PooledLibrary #73632 | CRISPR loss of function screens | T cell differentiation and function | MTHFD2 | Sugiura, A., et al. (2022)[132] |
| ex vivo | Primary mouse Treg cells | LentiCRISPRv2-Brie | CRISPR loss of function screens | Treg cell function and anti-tumour immunity | Brd9-containing ncBAF complex | Loo, C. S., et al. (2020)[133] |
| ex vivo | Primary human Treg cells | Cas9 RNPs pooled containing 40 selected candidate TFs of Treg cell identity | CRISPR loss of function screens | Treg cell function | HIVEP2 SATB1 | Schumann, K., et al. (2020)[134] |
| in vivo | Primary mouse CD4+ T cells | sgRNA metabolic library | CRISPR loss of function screens | Tfh cell differentiation | ETNK1 PCYT2 SELENOI | Fu, G., et al. (2021)[135] |
| ex vivo | Primary mouse CD4+ T cells | Lentiviral Brie sgRNA library (78,633 sgRNAs targeting 19,674 genes) | CRISPR loss of function screens | T cell priming and Treg suppressive function | SEC31A CCDC101 | Long, L., et al. (2021)[136] |
| in vivo | Primary mouse CD4+ T cells | ~ 400 SgRNAs from Brie library targeting ~ 80 genes | CRISPR loss of function screens | Tfh cell development, Tfh versus Th1 decisions | HIF-1α | Huang, B., et al. (2022)[137] |
| ex vivo | Human T cells | sgRNA library targeting 829 genes | CRISPR loss of function screens | T cell exhaustion | IKZF1 | Gayathri, B., et al. (2022)[138] |
| in vivo and ex vivo | Primary mouse CD8+ T cells | sgRNA library with 78,633 sgRNAs targeting 19,674 genes | CRISPR loss of function screens | CD8+ T cell expansion and anti-tumour immunity | Roquin | Zhao, H., et al. (2021)[139] |
AAV adeno-associated virus, Arid1a AT-rich interactive domain-containing protein 1A, Dhx37 DEAH-box helicase 37, Fli1, friend leukaemia integration 1 transcription factor, LCMV lymphocytic choriomeningitis virus, LTBR lymphotoxin beta receptor, HIF-1α Hypoxia-inducible factor 1-alpha, RASA2 RAS P21 Protein Activator 2, RASA3 RAS P21 Protein Activator 3, Pofut1 Protein O-Fucosyltransferase 1, PPARG Peroxisome proliferator-activated receptor gamma, Fut8 Fucosyltransferase 8, SOCS1 Suppressor of cytokine signalling 1, FAM49B Family with Sequence Similarity 49, Member B; SNX9 Sorting Nexin 9, IKZF1 IKAROS family zinc finger protein 1, ETS1 ETS Proto-Oncogene 1, Ptpn2 Protein Tyrosine Phosphatase Non-Receptor Type 2, PRODH2 Proline Dehydrogenase 2, MALT1 Mucosa-Associated Lymphoid Tissue Lymphoma Translocation Protein 1, BCL10 B-cell lymphoma/leukaemia 10, TRAF6 TNF Receptor Associated Factor 6, VAV1 Vav Guanine Nucleotide Exchange Factor 1, LCP2 Lymphocyte Cytosolic Protein 2, LAT Linker For Activation Of T Cells, CBLB Cbl Proto-Oncogene B, TLE4 TLE Family Member 4, IKZF2 IKAROS Family Zinc Finger 2, Usp22 Ubiquitin Specific Peptidase 22, Rnf20 Ring Finger Protein 20, MTHFD2 Methylenetetrahydrofolate Dehydrogenase 2, HIVEP2 Human Immunodeficiency Virus Type I Enhancer-Binding Protein 2,SATB1 Special AT-Rich Sequence Binding Protein 1, ETNK1, Ethanolamine Kinase 1, PCYT2 Phosphate Cytidylyltransferase 2, SELENOI Selenoprotein I, SEC31A SEC31 Homolog A, CCDC101 Roquin (RC3H1) Ring finger and CCCH-type domains 1