Purpose: The importance of cellular context to the synergy of DNA Damage Response (DDR) targeted agents is important for tumors with mutations in DDR pathways, but less well-established for tumors driven by oncogenic transcription factors. In this study, we exploit the widespread transcriptional dysregulation of the EWS-FLI1 transcription factor to identify an effective DDR targeted combination therapy for Ewing Sarcoma (ES). Experimental Design: We used matrix drug screening to evaluate synergy between a DNA-PK inhibitor (M9831) or an ATR inhibitor (berzosertib) and chemotherapy. The combination of berzosertib and cisplatin was selected for broad synergy, mechanistically evaluated for ES selectivity, and optimized for in vivo schedule. Results: Berzosertib combined with cisplatin demonstrates profound synergy in multiple ES cell lines at clinically achievable concentrations. The synergy is due to loss of expression of the ATR downstream target CHEK1, loss of cell cycle checkpoints, and mitotic catastrophe. Consistent with the goals of the project, EWS-FLI1 drives the expression of CHEK1 and five other ATR pathway members. The loss of CHEK1 expression is not due to transcriptional repression and instead caused by degradation coupled with suppression of protein translation. The profound synergy is realized in vivo with a novel optimized schedule of this combination in subsets of ES models leading to durable complete responses in 50% of animals bearing two different ES xenografts. Conclusion: These data exploit EWS-FLI1 driven alterations in cell context to broaden the therapeutic window of berzosertib and cisplatin to establish a promising combination therapy and a novel in vivo schedule.

中文翻译：

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Cell Context 是 ATR 抑制和顺铂联合治疗尤文肉瘤的第三个协同轴

目的：细胞环境对 DNA 损伤反应 (DDR) 靶向药物协同作用的重要性对于 DDR 通路突变的肿瘤很重要，但对于由致癌转录因子驱动的肿瘤则不太确定。在这项研究中，我们利用 EWS-FLI1 转录因子广泛的转录失调来确定针对尤文肉瘤 (ES) 的有效 DDR 靶向联合疗法。实验设计：我们使用基质药物筛选来评估 DNA-PK 抑制剂 (M9831) 或 ATR 抑制剂 (berzosertib) 与化疗之间的协同作用。选择 berzosertib 和顺铂的组合以实现广泛的协同作用，对 ES 选择性进行机械评估，并针对体内方案进行优化。结果：Berzosertib 与顺铂组合在多种 ES 细胞系中以临床可达到的浓度表现出深刻的协同作用。这种协同作用是由于 ATR 下游靶点 CHEK1 表达缺失、细胞周期检查点缺失和有丝分裂灾难造成的。与该项目的目标一致，EWS-FLI1 驱动 CHEK1 和其他五个 ATR 通路成员的表达。 CHEK1 表达的丧失不是由于转录抑制，而是由降解加上蛋白质翻译抑制引起的。通过在 ES 模型子集中这种组合的新颖优化方案在体内实现了深刻的协同作用，导致 50% 携带两种不同 ES 异种移植物的动物产生持久的完全反应。结论：这些数据利用 EWS-FLI1 驱动的细胞环境改变来扩大 berzosertib 和顺铂的治疗窗口，从而建立一种有前途的联合疗法和一种新颖的体内方案。