Journal: Molecular Therapy Oncology

Article Title: Co-editing of NKG2A and FAS increases long-term cytotoxic capacity and persistence of CAR NK cells

doi: 10.1016/j.omton.2026.201126

Figure Lengend Snippet: NKG2A-edited CAR NK cells eliminate CD19-negative target cells resistant to conventional CAR NK cells (A) Specific lysis of 721.45 CD19 KO cells by engineered NK cells (left) or CAR NK cells (right). Cells were co-cultured for 4–5 h at an effector to target ratio of 5:1. (B) Schematic of repetitive stimulation assay. 0.1 × 10 6 NK or CAR NK cells were seeded at day 0, and 0.1 × 10 6 CD19-positive 721.45 cells (target cells) were added every 2–3 days for 14 days. IL-2 was added 2×/week until day 12. Target cells added on day 7 were labeled with PKH26 and on day 14 with PKH67. Data shown in (C–H) were generated using this setup. (C) Expression of CD19 (histograms) and relative MFI of CD19 on total target cells (live CD56 − ) and representative flow cytometry plots on day 14 after co-culture with NK (top) or CAR NK (bottom) cells. Relative MFI is calculated by dividing by MFI of CD19 on target cells cultured alone. (D) Representative expression of PKH26 (target cells added day 7) and PKH67 (target cells added day 14) on total target cells (live CD56 − ) after co-culture with NK or CAR NK cells for 14 days. (E and F) Frequency of remaining live PKH26 + target cells (added on day 7) on day 14 after co-culture with NK cells (E) or CAR NK cells (F). Target cells alone (721.45 only) were not included in the statistical analysis. (G and H) Frequency of remaining live PKH67 + target cells (added on day 14 and co-cultured for 4–5 h before readout) after co-culture with NK cells (G) or CAR NK cells (H). Target cells alone (721.45 only) were not included in the statistical analysis. (A–H) n = 6–9 (individual donors). One-way ANOVA. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001.

Article Snippet: EBV-transformed LCLs from allogeneic human fetal liver tissue were generated by isolating CD19 + cells by magnetic-activated cell sorting with CD19 beads (Miltenyi, Cat. 130-050-301) and then infected with EBV B95-8, produced as previously described at a multiplicity of infection (MOI) of 0.05–0.15.

Techniques: Lysis, Cell Culture, Labeling, Generated, Expressing, Flow Cytometry, Co-Culture Assay

Journal: Molecular Therapy Oncology

Article Title: Co-editing of NKG2A and FAS increases long-term cytotoxic capacity and persistence of CAR NK cells

doi: 10.1016/j.omton.2026.201126

Figure Lengend Snippet: Increased cytotoxicity of NKG2A-edited CAR NK cells against CD19-negative BCP-ALL PDX (A) CD19 and (B) HLA-E expression of BCP-ALL PDX by flow cytometry. (C) Specific lysis of PDX1 and PDX2 after co-culture with control NK cells (NK mock) or CAR NK cells (CAR mock). Co-culture at E:T ratio 5:1 for 4 h. (D) Specific lysis of PDX1 after 4 h co-culture at 1:1 E:T ratio. (E) Data from (D) shown as relative increase in killing of engineered CAR NK cells compared with controls (CAR mock). (F–H) Specific lysis of PDX2 after 20 h co-culture at 5:1 E:T ratio with NK (F) or CAR NK (G) cells. (H) Data from (G) shown as relative increase in killing of engineered CAR NK cells compared with controls (CAR mock). (C) n = 3 (individual donors), paired t test. (D–G) n = 3 (individual donors), repeated measures one-way ANOVA. Engineered NK cells shown in this figure were generated from bulk NK cells (not NKG2A + NK cells) as the starting population. ∗ p < 0.05.

Article Snippet: EBV-transformed LCLs from allogeneic human fetal liver tissue were generated by isolating CD19 + cells by magnetic-activated cell sorting with CD19 beads (Miltenyi, Cat. 130-050-301) and then infected with EBV B95-8, produced as previously described at a multiplicity of infection (MOI) of 0.05–0.15.

Techniques: Expressing, Flow Cytometry, Lysis, Co-Culture Assay, Control, Generated

Journal: Molecular Therapy Oncology

Article Title: Co-editing of NKG2A and FAS increases long-term cytotoxic capacity and persistence of CAR NK cells

doi: 10.1016/j.omton.2026.201126

Figure Lengend Snippet: NKG2A-edited CAR NK cells eliminate CD19-negative target cells resistant to conventional CAR NK cells (A) Specific lysis of 721.45 CD19 KO cells by engineered NK cells (left) or CAR NK cells (right). Cells were co-cultured for 4–5 h at an effector to target ratio of 5:1. (B) Schematic of repetitive stimulation assay. 0.1 × 10 6 NK or CAR NK cells were seeded at day 0, and 0.1 × 10 6 CD19-positive 721.45 cells (target cells) were added every 2–3 days for 14 days. IL-2 was added 2×/week until day 12. Target cells added on day 7 were labeled with PKH26 and on day 14 with PKH67. Data shown in (C–H) were generated using this setup. (C) Expression of CD19 (histograms) and relative MFI of CD19 on total target cells (live CD56 − ) and representative flow cytometry plots on day 14 after co-culture with NK (top) or CAR NK (bottom) cells. Relative MFI is calculated by dividing by MFI of CD19 on target cells cultured alone. (D) Representative expression of PKH26 (target cells added day 7) and PKH67 (target cells added day 14) on total target cells (live CD56 − ) after co-culture with NK or CAR NK cells for 14 days. (E and F) Frequency of remaining live PKH26 + target cells (added on day 7) on day 14 after co-culture with NK cells (E) or CAR NK cells (F). Target cells alone (721.45 only) were not included in the statistical analysis. (G and H) Frequency of remaining live PKH67 + target cells (added on day 14 and co-cultured for 4–5 h before readout) after co-culture with NK cells (G) or CAR NK cells (H). Target cells alone (721.45 only) were not included in the statistical analysis. (A–H) n = 6–9 (individual donors). One-way ANOVA. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001.

Article Snippet: EBV-transformed LCLs from allogeneic human fetal liver tissue were generated by isolating CD19 + cells by magnetic-activated cell sorting with CD19 beads (Miltenyi, Cat. 130-050-301) and then infected with EBV B95-8, produced as previously described at a multiplicity of infection (MOI) of 0.05–0.15.

Techniques: Lysis, Cell Culture, Labeling, Generated, Expressing, Flow Cytometry, Co-Culture Assay

Journal: Molecular Therapy Oncology

Article Title: Co-editing of NKG2A and FAS increases long-term cytotoxic capacity and persistence of CAR NK cells

doi: 10.1016/j.omton.2026.201126

Figure Lengend Snippet: Increased cytotoxicity of NKG2A-edited CAR NK cells against CD19-negative BCP-ALL PDX (A) CD19 and (B) HLA-E expression of BCP-ALL PDX by flow cytometry. (C) Specific lysis of PDX1 and PDX2 after co-culture with control NK cells (NK mock) or CAR NK cells (CAR mock). Co-culture at E:T ratio 5:1 for 4 h. (D) Specific lysis of PDX1 after 4 h co-culture at 1:1 E:T ratio. (E) Data from (D) shown as relative increase in killing of engineered CAR NK cells compared with controls (CAR mock). (F–H) Specific lysis of PDX2 after 20 h co-culture at 5:1 E:T ratio with NK (F) or CAR NK (G) cells. (H) Data from (G) shown as relative increase in killing of engineered CAR NK cells compared with controls (CAR mock). (C) n = 3 (individual donors), paired t test. (D–G) n = 3 (individual donors), repeated measures one-way ANOVA. Engineered NK cells shown in this figure were generated from bulk NK cells (not NKG2A + NK cells) as the starting population. ∗ p < 0.05.

Article Snippet: EBV-transformed LCLs from allogeneic human fetal liver tissue were generated by isolating CD19 + cells by magnetic-activated cell sorting with CD19 beads (Miltenyi, Cat. 130-050-301) and then infected with EBV B95-8, produced as previously described at a multiplicity of infection (MOI) of 0.05–0.15.

Techniques: Expressing, Flow Cytometry, Lysis, Co-Culture Assay, Control, Generated