DYRK1A in pancreatic cancer

Although radiation therapy has recently made great advances in cancer treatment, the majority of patients diagnosed with pancreatic cancer (PC) cannot achieve satisfactory outcomes due to in-trinsic and acquired radioresistance. Identifying the molecular mechanisms that impair the effi-cacy of radiotherapy and targeting these pathways are essential to improve the radiation re-sponse of PC patients. Our goal is to identify sensitive targets for pancreatic cancer radiotherapy (RT) using the kinome-wide CRISPR-Cas9 loss-of-function screen, and to enhance the therapeutic effect through the development and application of targeted inhibitors combined with radiother-apy. We transduced pancreatic cancer cells with a protein kinase library, 2D and 3D library cells were daily irradiated with a single dose of up to 2 Gy for 4 weeks at a total of 40 Gy using an X-ray generator. Sufficient DNA was collected for next-generation deep sequencing to identify candidate genes. In this study, we identified several cell cycle checkpoint kinases and DNA damage related kinases in 2D- and 3D-cultivated cells, including DYRK1A, whose loss of func-tion sensitizes cells to radiotherapy. Additionally, we demonstrated that harmine-targeted sup-pression of DYRK1A used in conjunction with radiotherapy increases DNA double strand breaks (DSBs) and impairs homologous repair (HR), resulting in more cancer cell death. Our results support the use of CRISPR-Cas9 screening to identify new therapeutic targets, develop radiosensi-tizer, and provide novel strategies for overcoming the tolerance of pancreatic cancer to radio-therapy.

尽管放射治疗在癌症治疗领域近年来取得了长足进展，但大多数确诊胰腺癌（PC）的患者因固有及获得性放射抵抗，无法获得满意的治疗效果。阐明削弱放射治疗疗效的分子机制并靶向调控此类通路，是改善胰腺癌患者放射应答效果的关键所在。本研究旨在通过全激酶组CRISPR-Cas9功能缺失筛选，鉴定胰腺癌放射治疗（RT）的敏感靶点，并通过开发并应用靶向抑制剂联合放射疗法，提升治疗效果。我们将蛋白激酶文库转导至胰腺癌细胞中，随后使用X射线发生器对二维（2D）及三维（3D）培养的文库细胞每日进行单次照射，单次剂量最高为2 Gy，持续4周，总照射剂量达40 Gy。收集足够的DNA用于下一代深度测序，以筛选鉴定候选基因。本研究在二维及三维培养的细胞中鉴定出多种细胞周期检查点激酶与DNA损伤相关激酶，其中包括双特异性酪氨酸磷酸化调节激酶1A（DYRK1A），该基因的功能缺失可使细胞对放射治疗增敏。此外，本研究证实，采用哈尔明靶向抑制DYRK1A并联合放射治疗，可增加DNA双链断裂（DSBs）的数量并削弱同源重组修复（HR）功能，最终导致更多癌细胞死亡。本研究结果证实，可通过CRISPR-Cas9筛选鉴定新型治疗靶点、开发放射增敏剂，并为破解胰腺癌的放射抵抗提供全新策略。