Sui Xin, Geng Jian-Hao, Li Yong-Heng, Zhu Guang-Ying, Wang Wei-Hu
Key Laboratory of Carcinogenesis and Translational Research (Ministry of Education/Beijing), Department of Radiation Oncology, Peking University Cancer Hospital and Institute, Beijing, China,
Department of Radiation Oncology, National Clinical Research Center for Respiratory Disease, Center for Respiratory Disease, Lung Cancer Center, China-Japan Friendship Hospital, Peking University Health Science Center, Beijing, China,
Cancer Manag Res. 2018 Oct 26;10:5009-5018. doi: 10.2147/CMAR.S176084. eCollection 2018.
Radiotherapy is a major treatment method for patients with non-small cell lung cancer (NSCLC). However, the presence of radioresistant cancer stem cells (CSCs) may be associated with disease relapse or a poor outcome after radiotherapy. Voltage-gated calcium channel α2δ1 subunit (encoded by the gene ) isoform 5 is a marker of CSCs in hepatocellular carcinoma. This study aimed to investigate the radiosensitivity of α2δ1-high cells in NSCLC cell lines.
NSCLC cell lines A549, H1975, H1299, and PC9 were used. -knockdown and -overexpressing cell lines were established by lentiviral infection. Colony formation assay was performed to determine radiosensitivity. Sphere formation assay in serum-free medium was performed to evaluate self-renewal capacity. Proteins associated with DNA damage repair were analyzed by immunofluorescence or Western blot. The monoclonal antibody of α2δ1 was applied alone or in combination with radiation either in vitro or in vivo to determine the anti-tumor effect of the antibody.
α2δ1-high cells showed greater sphere-forming efficiency than α2δ1-low cells and were relatively resistant to radiation. knockdown in A549 cells enhanced radiosensitivity, whereas overexpression in PC9 and H1975 cells reduced radiosensitivity, suggesting that α2δ1 imparted radioresistance to NSCLC cells. Analysis of proteins involved in DNA damage repair suggested that α2δ1 enhanced the efficiency of DNA damage repair. The monoclonal antibody of α2δ1 had a synergistic effect with that of radiation to block the self-renewal of α2δ1-high cells and enhanced the radiosensitivity of α2δ1-positive cells in colony formation assays. The combination of the α2δ1 antibody with radiation repressed A549 xenograft growth in vivo.
α2δ1 enhances radioresistance in cancer stem-like cells in NSCLC. The α2δ1 monoclonal antibody sensitizes α2δ1-high cells to radiation, suggesting that the antibody may be used to improve the treatment outcome when combined with radiation in NSCLC.
放射治疗是非小细胞肺癌（NSCLC）患者的主要治疗方法。然而，放射抗性癌干细胞（CSCs）的存在可能与疾病复发或放射治疗后的不良预后相关。电压门控钙通道α2δ1亚基（由该基因编码）异构体5是肝细胞癌中CSCs的标志物。本研究旨在探讨NSCLC细胞系中α2δ1高表达细胞的放射敏感性。
使用NSCLC细胞系A549、H1975、H1299和PC9。通过慢病毒感染建立α2δ1基因敲低和过表达细胞系。进行集落形成试验以确定放射敏感性。在无血清培养基中进行成球试验以评估自我更新能力。通过免疫荧光或蛋白质印迹分析与DNA损伤修复相关的蛋白质。单独或与辐射联合应用α2δ1单克隆抗体，在体外或体内确定该抗体的抗肿瘤作用。
α2δ1高表达细胞比α2δ1低表达细胞表现出更高的成球效率，并且对辐射相对抗性。A549细胞中α2δ1基因敲低增强了放射敏感性，而PC9和H1975细胞中α2δ1过表达降低了放射敏感性，表明α2δ1赋予NSCLC细胞放射抗性。对参与DNA损伤修复的蛋白质分析表明，α2δ1提高了DNA损伤修复效率。α2δ1单克隆抗体与辐射具有协同作用，可阻断α2δ1高表达细胞的自我更新，并在集落形成试验中增强α2δ1阳性细胞的放射敏感性。α2δ1抗体与辐射联合可抑制体内A549异种移植瘤的生长。
α2δ1增强NSCLC中癌干细胞样细胞的放射抗性。α2δ1单克隆抗体使α2δ1高表达细胞对辐射敏感，表明该抗体与NSCLC放射治疗联合应用时可能用于改善治疗效果。
