genome-wide h3 human crispr ko library  (Addgene inc)

Journal: Journal for immunotherapy of cancer

Article Title: Genome-wide CRISPR/Cas9 screen reveals factors that influence the susceptibility of tumor cells to NK cell-mediated killing.

doi: 10.1136/jitc-2024-010699

Figure Lengend Snippet: Figure 1 Genome-wide CRISPR/Cas9 screening identifies genes involved in NKp44-mediated specific killing of tumor cells by KHYG-1 cells. (A) Genome-wide CRISPR/Cas9 screening design. HCT-116 Cas9+ cells were transduced with the knockout sgRNA Brunello library. Collection of mutant cells was subjected to lysis by WT KHYG-1 or NKp44-deficient KHYG-1 in the presence of α-NKp44 mAb or not (Round 1). Mutant cells that survived lysis by KHYG-1 were subjected to a second round (Round 2) of co-culture with WT KHYG-1 (condition 1), in the presence of α-NKp44 mAb (condition 2) or with NKp44-deficient KHYG-1 (condition 3). The abundance of sgRNA in the collection, in the surviving cells in Round 1 and in the surviving cells in Round 2 were determined by sequencing (see Material and methods). (B) Venn diagram of hits obtained with KHYG-1 co-culture (condition 1, see online supplemental table 1), KHYG-1+αNKp44mAb (condition 2, see online supplemental table 2) and NKp44- deficient KHYG-1 cells (condition 3, see online supplemental table 3). (B) Scatter plot showing the ranking of hits enriched in the NKp44-dependent killing of tumor cells by KHYG-1 cells by MAGeCK score and false discovery rate (see online supplemental table 4). FDR, false discovery rate; mAb, monoclonal antibody; sgRNA, single guide RNA; WT, wild-type.

Article Snippet: Human Brunello CRISPR KO pooled library was provided by the CRISPR Screen LabTech platform (Addgene #73178).

Techniques: Genome Wide, CRISPR, Transduction, Knock-Out, Mutagenesis, Lysis, Co-Culture Assay, Sequencing

Journal: Journal for immunotherapy of cancer

Article Title: Genome-wide CRISPR/Cas9 screen reveals factors that influence the susceptibility of tumor cells to NK cell-mediated killing.

doi: 10.1136/jitc-2024-010699

Figure Lengend Snippet: Figure 5 Relative requirement of NK cell effector functions for NK cell-mediated tumor lysis over time. WT and Fas-deficient HCT-116 Cas9+ cells were co-cultured with WT KHYG-1 or perforin-deficient KHYG-1 cells or with IL-2 activated primary NK cells from four donors, as indicated. The extent of specific lysis was determined in a standard 4-hour assay (short-term) (A) or as the CRISPR screen condition (long-term, 72 hours) (B). P values: ****<0.0001; **=0.024 in (A) and **=0.0037 in (B). Unpaired t-test. Data result from the pool of two to three independent experiments. IL, interleukin; KO, knockout; NK, natural killer; WT, wild-type.

Article Snippet: Human Brunello CRISPR KO pooled library was provided by the CRISPR Screen LabTech platform (Addgene #73178).

Techniques: Lysis, Cell Culture, CRISPR, Knock-Out