PO.ET08.01 · 实验与分子治疗
解析放射敏感性确定MDM2为TP53野生型肺腺癌协同放射增敏的可靶向驱动因素
Deciphering radiation sensitivity identifies MDM2 as a targetable driver of synergistic radiosensitization in TP53 wild-type lung adenocarcinoma
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摘要 Abstract
中文摘要
背景：对电离辐射（IR）的抗性仍是有效放疗的主要障碍。除了作为p53负性调节因子的经典作用外，MDM2对放射抗性的更广泛贡献仍未得到充分理解。鉴于肺腺癌（LUAD）对放疗表现出异质性应答，界定放射敏感性的分子决定因素或可揭示可操作的靶点。在此背景下，我们试图识别此类决定因素，并确定MDM2是否代表一个能够增强LUAD放疗疗效的关键且可治疗靶向的调节因子。
方法：使用照射后集落形成生存分析，将LUAD细胞系（A549、NCI-H460、NCI-H650、NCI-H1573）分为IR敏感和IR不敏感两组。整合转录组学图谱和DepMap CRISPR依赖性数据集，以揭示放射敏感性的潜在分子决定因素。在具有不同TP53背景的LUAD细胞系中，测试了单独或与IR联合的药理学MDM2抑制（idasanutlin）。使用集落形成实验、3D球体培养和异种移植评估在2D、3D和体内环境中的治疗应答。
结果：集落形成生存分析揭示了LUAD细胞系中存在不同的IR敏感和IR不敏感亚群。整合分析表明，MDM2依赖性在TP53野生型IR不敏感细胞中明显更高，提示存在一个MDM2驱动放射抗性的p53功能完整背景。与此预测一致，单独MDM2抑制（idasanutlin单药）在p53野生型细胞中诱导的生长抑制比在p53突变细胞中更为显著。IR-idasanutlin联合产生了强劲的协同细胞毒性，深度降低了集落形成生存，并引起球体缩小，伴随EthD-1荧光强度显著升高，提示细胞死亡增加。在NCI-H460异种移植模型中，idasanutlin（15 mpk，口服，每日一次）联合IR（4 Gy）产生了最高的肿瘤生长抑制（TGI=77.1%，p<0.001）且无体重下降，显示出强大的体内协同疗效和耐受性。这些发现凸显了MDM2阻断作为p53功能完整肿瘤放射增敏策略的转化潜力。
结论：我们的发现表明，通过整合组学分析揭示的MDM2依赖性代表了p53功能完整LUAD中IR抗性背后的一个治疗脆弱点。idasanutlin与放疗联合可有效克服这种抗性，凸显MDM2作为一个稳健且可操作的靶点，可增强由高MDM2活性驱动的癌症的放射应答。
查看英文原文 English abstract
Background: Resistance to ionizing radiation (IR) remains a major obstacle to effective radiotherapy. Beyond its classical role as a negative regulator of p53, MDM2's broader contribution to radioresistance remains insufficiently understood. Given that lung adenocarcinoma (LUAD) exhibits heterogeneous responses to radiotherapy, defining molecular determinants of radiation sensitivity may reveal actionable targets. In this context, we sought to identify such determinants and to determine whether MDM2 represents a critical and therapeutically targetable regulator capable of enhancing radiotherapy efficacy in LUAD.
Methods: LUAD cell lines (A549, NCI-H460, NCI-H650, NCI-H1573) were stratified into IR-sensitive and IR-insensitive groups using clonogenic survival following irradiation. Transcriptomic profiles and DepMap CRISPR dependency datasets were integrated to uncover potential molecular determinants of radiation sensitivity. Pharmacologic MDM2 inhibition (idasanutlin) was tested alone or in combination with IR across LUAD cell lines with distinct TP53 backgrounds. Clonogenic assays, 3D spheroid cultures, and xenografts were used to assess treatment responses in 2D, 3D, and in vivo settings.
Results: Clonogenic survival analysis revealed distinct IR-sensitive and IR-insensitive subsets among LUAD cell lines. Integrated analyses demonstrated that MDM2 dependency was markedly higher in TP53 wild-type IR-insensitive cells, suggesting a p53-proficient context for MDM2-driven radioresistance. In line with this prediction, MDM2 inhibition alone (idasanutlin monotherapy) induced more pronounced growth inhibition in p53 wild-type compared with p53-mutant cells. The IR-idasanutlin combination produced robust synergistic cytotoxicity, profoundly reducing clonogenic survival and causing spheroid shrinkage accompanied by markedly elevated EthD-1 fluorescence intensity, indicating increased cell death. In an NCI-H460 xenograft model, idasanutlin (15 mpk, p.o. daily) combined with IR (4 Gy) yielded the highest tumor growth inhibition (TGI = 77.1%, p < 0.001) without body-weight loss, demonstrating strong in vivo synergistic efficacy and tolerability. These findings underscore the translational potential of MDM2 blockade as a radiosensitizing strategy in p53-proficient tumors.
Conclusions: Our findings demonstrate that MDM2 dependency, revealed through integrative omics analyses, represents a therapeutic vulnerability underlying IR-resistance in p53-proficient LUAD. Combining idasanutlin with radiotherapy effectively overcomes this resistance, highlighting MDM2 as a robust and actionable target for enhancing radiation response in cancers driven by high MDM2 activity.
利益披露 Disclosure
C. Maeng, None..
J. Hwang, None..
J. Ahn, None..
S. Shin, None.