Abstract：
Radiotherapy is one of the standard therapeutic regimens for medulloblastoma (MB). Tumor cells utilize DNA damage repair (DDR) mechanisms to survive and develop resistance during radiotherapy. It has been found that targeting DDR sensitizes tumor cells to radiotherapy in several types of cancer, but whether and how DDR pathways are involved in the MB radiotherapy response remain to be determined. Single-cell RNA sequencing was carried out on 38 MB tissues, followed by expression enrichment assays. Fanconi anemia group D2 gene (FANCD2) expression was evaluated in MB samples and public MB databases. The function of FANCD2 in MB cells was examined using cell counting assays (CCK-8), clone formation, lactate dehydrogenase activity, and in mouse orthotopic models. The FANCD2-related signaling pathway was investigated using assays of peroxidation, a malondialdehyde assay, a reduced glutathione assay, and using FerroOrange to assess intracellular iron ions (Fe2+ ). Here, we report that FANCD2 was highly expressed in the malignant sonic hedgehog (SHH) MB subtype (SHH-MB). FANCD2 played an oncogenic role and predicted worse prognosis in SHH-MB patients. Moreover, FANCD2 knockdown markedly suppressed viability, mobility, and growth of SHH-MB cells and sensitized SHH-MB cells to irradiation. Mechanistically, FANCD2 deficiency led to an accumulation of Fe2+ due to increased divalent metal transporter 1 expression and impaired glutathione peroxidase 4 activity, which further activated ferroptosis and reduced proliferation of SHH-MB cells. Using an orthotopic mouse model, we observed that radiotherapy combined with silencing FANCD2 significantly inhibited the growth of SHH-MB cell-derived tumors in vivo. Our study revealed FANCD2 as a potential therapeutic target in SHH-MB and silencing FANCD2 could sensitize SHH-MB cells to radiotherapy via inducing ferroptosis. © 2024 The Pathological Society of Great Britain and Ireland.
摘要：
放射治疗是髓母细胞瘤（MB）的标准治疗方案之一。肿瘤细胞利用DNA损伤修复(DDR)机制在放疗期间存活并产生抗性。已经发现，靶向DDR使肿瘤细胞对几种类型的癌症的放疗敏感，但DDR通路是否以及如何参与MB放疗反应仍有待确定。对38个MB组织进行单细胞RNA测序，然后是表达富集测定。在MB样品和公共MB数据库中评估范可尼贫血组D2基因(FANCD2)表达。使用细胞计数测定法(CCK-8)检查MB细胞中FANCD2的功能,克隆形成,乳酸脱氢酶活性，和小鼠原位模型。FANCD2相关的信号通路使用过氧化试验进行了研究，丙二醛检测,还原型谷胱甘肽测定,并使用FerroOrange评估细胞内铁离子(Fe2+)。这里,我们报告FANCD2在恶性音效刺猬（SHH）MB亚型（SHH-MB）中高表达。FANCD2在SHH-MB患者中发挥致癌作用并预测预后较差。此外，FANCD2敲低明显抑制了生存能力，移动性,和SHH-MB细胞的生长以及SHH-MB细胞对辐射的敏感性。机械上,由于二价金属转运蛋白1表达增加和谷胱甘肽过氧化物酶4活性受损，FANCD2缺乏导致Fe2积累。这进一步激活了铁凋亡并减少了SHH-MB细胞的增殖。使用原位小鼠模型，我们观察到放疗联合沉默FANCD2在体内显著抑制SHH-MB细胞源性肿瘤的生长。我们的研究表明，FANCD2是SHH-MB的潜在治疗靶点，沉默FANCD2可以通过诱导铁凋亡使SHH-MB细胞对放疗敏感。©2024英国和爱尔兰病理学会。
