Ziener Judy, Henao-Restrepo Julián Andrés, Leonhardi Johanna, Sturm Max-Johann, Becker Sabine, Morales-Prieto Diana M, Milde Till, Beck James F, Sonnemann Jürgen
Department of Paediatric and Adolescent Medicine, Jena University Hospital, Friedrich Schiller University Jena, Jena, Germany.
Research Centre Lobeda, Jena University Hospital, Friedrich Schiller University Jena, Jena, Germany.
BMC Cancer. 2025 Feb 17;25(1):277. doi: 10.1186/s12885-025-13691-2.
Ewing's sarcoma is a childhood bone and soft tissue cancer with poor prognosis. Treatment outcomes for Ewing's sarcoma patients have improved only modestly over the past decades, making the development of new treatment strategies paramount. In this study, the combined targeting of ribonucleotide reductase (RNR) and WEE1 was explored for its effectiveness against Ewing's sarcoma cells.
The RNR inhibitor triapine and the WEE1 inhibitors adavosertib and ZN-c3 were tested in p53 wild-type and p53 mutant Ewing's sarcoma cells. The combination of adavosertib with the PARP inhibitors olaparib and veliparib was tested for comparison. Combinatorial effects were determined by flow cytometric analyses of cell death, loss of mitochondrial membrane potential and DNA fragmentation as well as by caspase 3/7 activity assay, immunoblotting and real-time RT-PCR. The drug interactions were assessed using combination index analysis.
RNR and WEE1 inhibitors were weakly to moderately effective on their own, but highly effective in combination. The combination treatments were similarly effective in p53 wild-type and p53 mutant cells. They synergistically induced cell death and cooperated to elicit mitochondrial membrane potential decay, to activate caspase 3/7 and to trigger DNA fragmentation, evidencing the induction of the apoptotic cell death cascade. They also cooperated to boost CHK1 phosphorylation, indicating augmented replication stress after combination treatment. In comparison, the combination of adavosertib with PARP inhibitors produced weaker synergistic effects.
Our findings show that combined inhibition of RNR and WEE1 was effective against Ewing's sarcoma in vitro. They thus provide a rationale for the evaluation of the potential of combined targeting of RNR and WEE1 in Ewing's sarcoma in vivo.
尤因肉瘤是一种儿童期骨与软组织癌症，预后较差。在过去几十年中，尤因肉瘤患者的治疗效果仅略有改善，因此开发新的治疗策略至关重要。在本研究中，探索了联合靶向核糖核苷酸还原酶（RNR）和WEE1对尤因肉瘤细胞的有效性。
在p53野生型和p53突变型尤因肉瘤细胞中测试了RNR抑制剂曲安西滨以及WEE1抑制剂adavosertib和ZN-c3。测试了adavosertib与聚腺苷二磷酸核糖聚合酶（PARP）抑制剂奥拉帕尼和维利帕尼的联合用药效果以作比较。通过对细胞死亡、线粒体膜电位丧失和DNA片段化的流式细胞术分析以及半胱天冬酶3/7活性测定、免疫印迹和实时逆转录聚合酶链反应来确定联合效应。使用联合指数分析评估药物相互作用。
RNR和WEE1抑制剂单独使用时效果较弱至中等，但联合使用时效果显著。联合治疗在p53野生型和p53突变型细胞中同样有效。它们协同诱导细胞死亡，并共同引起线粒体膜电位衰减，激活半胱天冬酶3/7并引发DNA片段化，证明诱导了凋亡细胞死亡级联反应。它们还共同促进了CHK1磷酸化，表明联合治疗后复制应激增强。相比之下，adavosertib与PARP抑制剂的联合产生的协同效应较弱。
我们的研究结果表明，联合抑制RNR和WEE1在体外对尤因肉瘤有效。因此，它们为在体内评估联合靶向RNR和WEE1治疗尤因肉瘤的潜力提供了理论依据。
