Cao Kaiyue, Pan Yunzhi, Yu Long, Shu Xiong, Yang Jing, Sun Linxin, Sun Lichao, Yang Zhihua, Ran Yuliang
National Cancer Center/Cancer Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100021, P.R. China.
Int J Oncol. 2017 Feb;50(2):587-596. doi: 10.3892/ijo.2016.3818. Epub 2016 Dec 22.
Cancer stem cells (CSCs) are a rare subset of cancer cells that play a significant role in cancer initiation, spreading, and recurrence. In this study, a subpopulation of lung cancer stem-like cells (LCSLCs) was identified from non-small cell lung carcinoma cell lines, SPCA-1 and A549, using serum-free suspension sphere-forming culture method. A monoclonal antibody library was constructed using immunized BLAB/c mice with the multipotent CSC cell line T3A-A3. Flow cytometry analysis showed that 33 mAbs targeted antigens can be enriched in sphere cells compared with the parental cells of SPCA-1 and A549 cell lines. Then, we performed functional antibody screening including sphere-forming inhibiting and invasion inhibiting assay. The results showed that two antibodies, 12C7 and 9B8, notably suppressed the self-renewal and invasion of LCSLCs. Fluorescence-activated cell sorting (FACs) found that the positive cells recognized by mAbs, 12C7 or 9B8, displayed features of LCSLCs. Interestingly, we found that these two antibodies recognized different subsets of cells and their combination effect was superior to the individual effect both in vitro and in vivo. Tissue microarrays were applied to detect the expression of the antigens targeted by these two antibodies. The positive expression of 12C7 and 9B8 targeted antigen was 84.4 and 82.5%, respectively, which was significantly higher than that in the non-tumor lung tissues. In conclusion, we screened two potential therapeutic antibodies that target different subsets of LCSLCs.
癌症干细胞（CSCs）是癌细胞中的一个稀有亚群，在癌症的起始、扩散和复发中起着重要作用。在本研究中，采用无血清悬浮成球培养法，从非小细胞肺癌细胞系SPCA-1和A549中鉴定出肺癌干细胞样细胞（LCSLCs）亚群。用多能CSC细胞系T3A-A3免疫BALB/c小鼠，构建单克隆抗体文库。流式细胞术分析显示，与SPCA-1和A549细胞系的亲本细胞相比，33种单克隆抗体靶向的抗原可在成球细胞中富集。然后，我们进行了功能抗体筛选，包括成球抑制和侵袭抑制试验。结果显示，两种抗体12C7和9B8显著抑制了LCSLCs的自我更新和侵袭。荧光激活细胞分选（FACs）发现，单克隆抗体12C7或9B8识别的阳性细胞表现出LCSLCs的特征。有趣的是，我们发现这两种抗体识别不同的细胞亚群，它们的联合作用在体外和体内均优于单独作用。应用组织芯片检测这两种抗体靶向抗原的表达。12C7和9B8靶向抗原的阳性表达分别为84.4%和82.5%，显著高于非肿瘤肺组织。总之，我们筛选出了两种靶向LCSLCs不同亚群的潜在治疗性抗体。
