Fig. 1: Generation process and evaluation of CRISPR/Cas9 knockout for CAR33-KLRC1ko-NK cells. | Nature Communications

Fig. 1: Generation process and evaluation of CRISPR/Cas9 knockout for CAR33-KLRC1ko-NK cells.

From: CRISPR/Cas9 editing of NKG2A improves the efficacy of primary CD33-directed chimeric antigen receptor natural killer cells

Fig. 1: Generation process and evaluation of CRISPR/Cas9 knockout for CAR33-KLRC1ko-NK cells.

a Scheme of the in vitro generation of primary CAR33-KLRC1ko-NK cells. b Exemplary flow cytometry plots of CAR33 and NKG2A expression on modified NK cells. c Flow cytometry-based CAR33 surface expression analysis over time (n = 6–9). Mean ± SD. CAR33 vs CAR33-KLRC1ko: d14 (n = 0.1158), d21 (n = 0.1296), d28 (n = 0.8095). d Flow cytometric analysis of NKG2A expression following CRISPR/Cas9 knockout of the KLRC1 gene (n = 7). Mean ± SD. e Frequency of KLRC1 disruption on genomic level was evaluated by Inference of CRISPR Edits (ICE) and T7E1 assay for CAR-transduced (+CAR) and control NK cells (−CAR) (n = 3). Mean ± SD. f Insertion/deletion (indel) distribution profiles were shown for ICE analysis from three different donors (D1, D2, D3). g Expansion-fold and workflow of non-transduced (NT)-NK, KLRC1ko-NK, CAR33-NK and CAR33-KLRC1ko-NK cells generation in the presence of IL-2 (500 U/mL) and IL-15 (10 ng/ml Miltenyi Biotec or 50 ng/mL CellGenix) (n = 7). Mean ± SD (ns = 0.9991). Statistical analysis was performed using two-way ANOVA (c, g) and paired Student’s t test (d, e). The entity of n is biological replicates (from different healthy donors) (c–e, g).

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