Figure 4 from Replication Stress Is an Actionable Genetic Vulnerability in Desmoplastic Small Round Cell Tumors

Abstract

<p>EWS–WT1 is a determinant of DSRCT cells’ sensitivity to PARPi and ATRi. <b>A,</b> Western blot of EWS–WT1 in JN1 and R cells transfected with either siCNTRL or siEWS–WT1. Whole-cell lysates were generated 48 hours after transfection. <b>B–E,</b> Dose–response survival curves of JN1 or R cells exposed to PARPi talazoparib (<b>B</b> and <b>C</b>) or ATRi M4344 (<b>D</b> and <b>E</b>) for 7 days in the presence or absence of siRNA-mediated silencing of EWS–WT1. Mean ± SD; <i>n</i> = 3. <b>F</b> and <b>G,</b> Quantification of γH2AX in JN1 cells exposed to DMSO control, PARPi talazoparib, ATRi M4344, or a combination of both for 72 hours, in the presence or absence of siRNA-mediated silencing of EWS–WT1. Cisplatin was used as the positive control. A minimum of 500 nuclei was analyzed per condition. Violin plots show the absolute number of foci per nucleus. Thick line, median; thin lines, bottom and top quartiles; two-way ANOVA and <i>post hoc</i> Dunn test. <b>H</b> and <b>I,</b> Western blots of pCHK1, CHK1, pRPA2, RPA2, γH2AX, H2AX, and EWS–WT1 in JN1 (<b>H</b>) or R (<b>I</b>) cells exposed to DMSO control, PARPi talazoparib (Tala), ATRi M4344, or a combination of both for 48 hours, in the presence or absence of siRNA-mediated silencing of EWS–WT1. ****, <i>P</i> < 0.0001; ns, not significant. </p>