DataSheet_6_Identification of Key Genes Mutations Associated With the Radiosensitivity by Whole Exome Sequencing in Pancreatic Cancer.xlsx

BackgroundPancreatic cancer (PC) is one of the most lethal human cancers, and radiation therapy (RT) is an important treating option. Many patients diagnosed with PC do not achieve objective responses because of the existence of intrinsic and acquired radioresistance. Therefore, biomarkers, which predict radiotherapy benefit in PC, are eagerly needed to be identified.
MethodsWhole-exome sequencing of six pancreatic ductal adenocarcinoma patients (PDAC) (three with a good response and three with a poor response) who had received radical surgery and then radiotherapy has been performed as standard of care treatment. Somatic and germline variants and the mutational signatures were analyzed with bioinformatics tools and public databases. Functional enrichment and pathway-based protein-protein interaction analyses were utilized to address the possibly mechanism in radioresistance. MTT, LDH, and colony formation assay were applied to evaluate cell growth and colony formation ability.
ResultsIn the present study, somatic mutations located in 441 genes were detected to be radiosensitivity-related loci. Seventeen genes, including the Smad protein family members (SMAD3 and SMAD4), were identified to influence the radiosensitivity in PDAC. The SMAD3 and SMAD4 genes mutate differently between radiosensitive and radioresistant PDAC patients. Mutation of SMAD3 potentiates the effects of ionizing radiation (IR) on cell growth and colony formation in PDAC cells, whereas mutation of SMAD4 had the opposite effects. SMAD3 and SMAD4 regulate the radiosensitivity of PDAC, at least in part, by P21 and FOXO3a, respectively.
ConclusionsThese results indicate that mutations of SMAD3 and SMAD4 likely cause the difference of response to radiotherapy in PDAC, which might be considered as the biomarkers and potential targets for the radiotherapy of pancreatic cancer.

背景：胰腺癌（Pancreatic Cancer, PC）是人类致死性最高的恶性肿瘤之一，放射治疗（Radiation Therapy, RT）是其重要的治疗手段。由于固有放射抵抗与获得性放射抵抗的存在，多数确诊胰腺癌的患者无法获得客观治疗应答。因此，亟需筛选可预测胰腺癌患者放疗获益的生物标志物。
方法：本研究按照标准临床诊疗流程，对6例接受根治性手术联合术后放疗的胰腺导管腺癌（Pancreatic Ductal Adenocarcinoma, PDAC）患者开展全外显子组测序，其中放疗应答良好与较差的患者各3例。采用生物信息学工具及公共数据库分析体细胞变异、种系变异及突变特征；通过功能富集分析与基于通路的蛋白质相互作用分析，探究放射抵抗的潜在分子机制。采用MTT实验、LDH释放实验及集落形成实验，评估细胞增殖能力与集落形成能力。
结果：本研究共检测到441个基因的体细胞突变与放射敏感性相关。筛选得到17个可影响胰腺导管腺癌放射敏感性的基因，其中包括Smad蛋白家族成员SMAD3与SMAD4。放疗敏感与放疗抵抗的PDAC患者中，SMAD3与SMAD4的突变模式存在显著差异。在PDAC细胞中，SMAD3突变可增强电离辐射（Ionizing Radiation, IR）对细胞增殖与集落形成的抑制作用，而SMAD4突变则呈现相反效应。SMAD3与SMAD4分别通过调控P21与FOXO3a通路，部分介导胰腺导管腺癌的放射敏感性调控。
结论：本研究结果表明，SMAD3与SMAD4的突变差异可能是导致PDAC患者放疗应答异质性的关键因素，二者有望作为胰腺癌放疗的潜在生物标志物与治疗靶点。