Department of Radiation Oncology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, United States.
Department of Comparative Medicine, Medical University of South Carolina, Charleston, SC 29425, United States.
Radiother Oncol. 2024 Nov;200:110531. doi: 10.1016/j.radonc.2024.110531. Epub 2024 Sep 11.
Overcoming radioresistance is a critical challenge in pancreatic ductal adenocarcinoma (PDAC). Our study investigates the targeting of Cyclin-dependent kinase-1 (CDK1) through genetic and pharmaceutical inhibition to radiosensitize PDAC cells.
Mass spectrometry and phosphoproteomics were used to analyze engineered radiation-resistant PDAC cell lines (MIA PaCa-2 and PANC-1) compared to parental controls. The TCGA PDAC database was queried for clinical outcomes and patients were dichotomized based on the median CDK1 mRNA expression. We generated a microRNA-based TET-on inducible shRNA to inhibit CDK1 expression in two PDAC cell lines. We used an orthotopic model of PDAC to test the radiation sensitivity of PDAC tumors with or without doxycycline treatment. We targeted CDK1 activation with a selective CDK1 inhibitor, RO-3306, followed by in vitro experiments employing immunoblotting, immunocytochemistry, and clonogenic assays.
Phosphoproteomics analysis revealed that phospho-CDK1 (Tyr15) was significantly elevated in the resistant clones. We found that high CDK1 expression was associated with worse OS in PDAC patients. Radiation exposure increased CDK1 phosphorylation. In MIA PaCa-2 and PANC-1 cells, CDK1 inhibition synergized with radiation therapy to delay tumor growth in vivo. CDK1 inhibition via. RO-3306 resulted in a significant shift of cells into the G2/M phase and disrupted DNA repair after radiation exposure. In vitro, pre-treatment with RO-3306 led to enhanced radiosensitivity of PDAC cells.
CDK1 plays a crucial role in PDAC radioresistance. Targeting CDK1 with radiotherapy holds promise for further investigation in PDAC treatment.
克服放射性抵抗是胰腺导管腺癌（PDAC）的一个关键挑战。我们的研究通过基因和药物抑制来靶向细胞周期蛋白依赖性激酶-1（CDK1），以放射增敏 PDAC 细胞。
质谱和磷酸化蛋白质组学用于分析与亲本对照相比的工程化辐射抗性 PDAC 细胞系（MIA PaCa-2 和 PANC-1）。TCGA PDAC 数据库被查询以获得临床结果，并根据 CDK1 mRNA 表达的中位数将患者分为两组。我们生成了一种基于 microRNA 的 TET-on 诱导性 shRNA，以抑制两种 PDAC 细胞系中的 CDK1 表达。我们使用 PDAC 原位模型来测试 PDAC 肿瘤在有无强力霉素治疗下的放射敏感性。我们使用选择性 CDK1 抑制剂 RO-3306 靶向 CDK1 激活，然后进行体外实验，包括免疫印迹、免疫细胞化学和集落形成实验。
磷酸化蛋白质组学分析表明，磷酸化 CDK1（Tyr15）在抗性克隆中显著升高。我们发现 CDK1 高表达与 PDAC 患者的 OS 较差相关。辐射暴露增加了 CDK1 的磷酸化。在 MIA PaCa-2 和 PANC-1 细胞中，CDK1 抑制与放射治疗协同作用，延迟体内肿瘤生长。通过 RO-3306 抑制 CDK1 导致细胞显著进入 G2/M 期，并在辐射暴露后破坏 DNA 修复。在体外，RO-3306 的预处理导致 PDAC 细胞的放射敏感性显著增强。
CDK1 在 PDAC 的放射性抵抗中起关键作用。用放射治疗靶向 CDK1 有望进一步研究 PDAC 的治疗。
