Figure 4.
Mouse Asxl1–/–Ezh2–/– dKO cells are highly sensitive to PARPi. (A) The bar chart (left) shows the percentage of cells with ≥10 γH2AX foci for the indicated cell type with or without PARPi treatment. Representative images (right) of γH2AX foci (green) and their merge with DAPI (4′,6-diamidino-2-phenylindole; blue) are shown. (B) Comet assay was performed after treatment with or without PARPi on in vitro–deleted Asxl–/–Ezh2–/– dKO c-kit+ or WT c-kit+ cells and head length, tail length, and tail moment are shown in the violin plots. (C) Example comet pictures for control and PARPi-treated cells are shown. (D-E) The relative number of colonies for in vitro–deleted Asxl1–/–Ezh2–/– c-kit+ cells (n = 4) and WT c-kit+ cells (n = 3) (D) and Asxl1–/–Ezh2–/– Runx1KD MDS (n = 2) and MLL-transformed cells (n = 2) (E) with or without PARPi treatment are shown (left). Representative colony pictures are shown (right). (F) The dot plot shows the cell numbers relative to untreated control (blue dotted line) for Asxl1–/–Ezh2–/– dKO CLL (n = 4) and TCL1 CLL (n = 4) after 1 or 2 days of PARPi treatment. (G) The bar chart shows the percentage of cells with ≥10 γH2AX foci upon in vitro treatment with or without PARPi. (H) Comet assay was performed after treatment with or without PARPi on Asxl–/–Ezh2–/– dKO CLL or TCL1 CLL cells and head length, tail length, and tail moment are shown in the violin plots. (I) Example comet pictures for control and PARPi-treated cells are shown. (J) Schematic diagram of the experimental plan to treat tertiary Asxl1–/–Ezh2–/– dKO CLL cells in vivo with the indicated drugs (vehicle, 10%; HBC, n = 9; olaparib, 50 mg/kg; olaparib, n = 10) (top). Kaplan-Meier survival curves of Asxl1–/–Ezh2–/– dKO CLL mice treated with vehicle or olaparib treatment indicated (bottom). (K) Weight of the spleens of mice at the end of experiment for the indicated groups. Vehicle, n = 9; Olaparib, n = 10. (L) Representative photos of the spleens from Asxl1–/–Ezh2–/– dKO CLL mice treated with indicated drugs. Plots show mean ± SEM, ∗∗∗P < .001, ∗∗∗∗P < .001, unpaired t test. FDR, false discovery rate; FITC, fluorescein isothiocyanate; NES, normalized enrichment score; ns, not significant.

Mouse Asxl1–/–Ezh2–/– dKO cells are highly sensitive to PARPi. (A) The bar chart (left) shows the percentage of cells with ≥10 γH2AX foci for the indicated cell type with or without PARPi treatment. Representative images (right) of γH2AX foci (green) and their merge with DAPI (4′,6-diamidino-2-phenylindole; blue) are shown. (B) Comet assay was performed after treatment with or without PARPi on in vitro–deleted Asxl–/–Ezh2–/– dKO c-kit+ or WT c-kit+ cells and head length, tail length, and tail moment are shown in the violin plots. (C) Example comet pictures for control and PARPi-treated cells are shown. (D-E) The relative number of colonies for in vitro–deleted Asxl1–/–Ezh2–/– c-kit+ cells (n = 4) and WT c-kit+ cells (n = 3) (D) and Asxl1–/–Ezh2–/– Runx1KD MDS (n = 2) and MLL-transformed cells (n = 2) (E) with or without PARPi treatment are shown (left). Representative colony pictures are shown (right). (F) The dot plot shows the cell numbers relative to untreated control (blue dotted line) for Asxl1–/–Ezh2–/– dKO CLL (n = 4) and TCL1 CLL (n = 4) after 1 or 2 days of PARPi treatment. (G) The bar chart shows the percentage of cells with ≥10 γH2AX foci upon in vitro treatment with or without PARPi. (H) Comet assay was performed after treatment with or without PARPi on Asxl–/–Ezh2–/– dKO CLL or TCL1 CLL cells and head length, tail length, and tail moment are shown in the violin plots. (I) Example comet pictures for control and PARPi-treated cells are shown. (J) Schematic diagram of the experimental plan to treat tertiary Asxl1–/–Ezh2–/– dKO CLL cells in vivo with the indicated drugs (vehicle, 10%; HBC, n = 9; olaparib, 50 mg/kg; olaparib, n = 10) (top). Kaplan-Meier survival curves of Asxl1–/–Ezh2–/– dKO CLL mice treated with vehicle or olaparib treatment indicated (bottom). (K) Weight of the spleens of mice at the end of experiment for the indicated groups. Vehicle, n = 9; Olaparib, n = 10. (L) Representative photos of the spleens from Asxl1–/–Ezh2–/– dKO CLL mice treated with indicated drugs. Plots show mean ± SEM, ∗∗∗P < .001, ∗∗∗∗P < .001, unpaired t test. FDR, false discovery rate; FITC, fluorescein isothiocyanate; NES, normalized enrichment score; ns, not significant.

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