Therapeutic targeting of DDR pathways. (A) Schematic representation of the ATM/Chk2 and ATR/Chk1 pathways in response to different types of DNA damage and the investigated compounds targeting different components within these pathways. (B) GRmax values of 8 compounds targeting different components of the DDR across 22 TCL cell lines after 72-hour treatment. GRmax >0 indicate partial cytostatic response, GRmax = 0 indicate full cytostatic response, GRmax <0 indicate partial cytotoxic response, GRmax = −1 indicate full cytotoxic effect. GR50 values represent mean values from at least 3 independent experiments. (C) ZMAD scores of ATM and ATR pathway genes obtained from a previously performed genome-wide CRISPR screen, with negative ZMAD scores (red colors) indicating a high and positive ZMAD scores (blue color) indicating low genetic dependency. (D-F) Cellular effects of WEE1 inhibition after 24 hours treatment with adavosertib: (D) Cell cycle analysis after adavosertib treatment. (E) MPM2 positivity by flow cytometry after adavosertib treatment. Significance was assessed with a 2-way analysis of variance (ANOVA) test after Tukey using correction for multiple comparison. (F) Immunoblot of proteins affected by WEE1 inhibition. (G) Drug response of TCL cell lines measured in relative IC50 values vs mutation status. Left: IC50 values of the MDM2i RG-7112 in TP53 wild type vs TP53 mutated cell lines. Middle: IC50 values of the WEE1 inhibitor (WEE1i) adavosertib in TP53 wild type vs TP53 mutated cell lines. Right: IC50 values of the WEE1i adavosertib in cell lines harboring loss of function alterations in the ATM/ATR pathways (ATM, ATR, Chek1, RPA1/2, and NBN) vs cell lines without such mutations. Significance was assessed with an unpaired 2-tailed t test. IC50 values are mean values from at least 3 independent experiments. ∗∗∗P ≤ .001, ∗∗∗∗P ≤ .0001. BER, base excision repair; DSB, DNA double-strand break; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HR, homologous recombination; MDM2i, MDM2 inhibitor; NHEJ, nonhomologous end joining; ns, nonsignificant; SSB, DNA single-strand breaks; WT, wild type.
Figure 3.

Therapeutic targeting of DDR pathways. (A) Schematic representation of the ATM/Chk2 and ATR/Chk1 pathways in response to different types of DNA damage and the investigated compounds targeting different components within these pathways. (B) GRmax values of 8 compounds targeting different components of the DDR across 22 TCL cell lines after 72-hour treatment. GRmax >0 indicate partial cytostatic response, GRmax = 0 indicate full cytostatic response, GRmax <0 indicate partial cytotoxic response, GRmax = −1 indicate full cytotoxic effect. GR50 values represent mean values from at least 3 independent experiments. (C) ZMAD scores of ATM and ATR pathway genes obtained from a previously performed genome-wide CRISPR screen, with negative ZMAD scores (red colors) indicating a high and positive ZMAD scores (blue color) indicating low genetic dependency. (D-F) Cellular effects of WEE1 inhibition after 24 hours treatment with adavosertib: (D) Cell cycle analysis after adavosertib treatment. (E) MPM2 positivity by flow cytometry after adavosertib treatment. Significance was assessed with a 2-way analysis of variance (ANOVA) test after Tukey using correction for multiple comparison. (F) Immunoblot of proteins affected by WEE1 inhibition. (G) Drug response of TCL cell lines measured in relative IC50 values vs mutation status. Left: IC50 values of the MDM2i RG-7112 in TP53 wild type vs TP53 mutated cell lines. Middle: IC50 values of the WEE1 inhibitor (WEE1i) adavosertib in TP53 wild type vs TP53 mutated cell lines. Right: IC50 values of the WEE1i adavosertib in cell lines harboring loss of function alterations in the ATM/ATR pathways (ATM, ATR, Chek1, RPA1/2, and NBN) vs cell lines without such mutations. Significance was assessed with an unpaired 2-tailed t test. IC50 values are mean values from at least 3 independent experiments. ∗∗∗P ≤ .001, ∗∗∗∗P ≤ .0001. BER, base excision repair; DSB, DNA double-strand break; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HR, homologous recombination; MDM2i, MDM2 inhibitor; NHEJ, nonhomologous end joining; ns, nonsignificant; SSB, DNA single-strand breaks; WT, wild type.

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