人乳腺癌细胞特异结合多肽的筛选与鉴定  

Screening and Identification of Human Breast Cancer Cell Specific Binding Polypeptides

作　　者：赵瑞霞 郑静 杨光梅 白雪 张丹莹 车皖琳 聂国朝[2] 侯颖春[1] Zhao Ruixia;Zheng Jing;Yang Guangmei;Bai Xue;Zhang Danying;Che Wanlin;Nie Guochao;Hou Yingchun(School of Life Sciences,Shaanxi Normal University,Xi′an,Shanxi 710119;Key Laboratory of Agricultural Resource Chemistry and Biotechnology,Yulin Normal University,Yulin,Guangxi 537000)

机构地区：[1]陕西师范大学生命科学学院,陕西西安710119 [2]玉林师范学院广西农产资源化学与生物技术重点实验室,广西玉林537000

出　　处：《玉林师范学院学报》2024年第3期1-10,共10页Journal of Yulin Normal University

基　　金：国家自然科学基金(2196104);玉林师范学院高层次人才项目(G2021ZK16,G2023ZK12);玉林师范学院大学生创新创业训练科研项目(S202210606117,S202210606118,S202210606120,202310606027)

摘　　要：乳腺癌(Breast cancer,BC)是女性最常见的恶性肿瘤之一,发病率和死亡率都很高.当前BC临床在早期诊断和靶向化疗方面仍具有许多不足,迫切需要开发新的BC靶向诊断和靶向化疗新方法.为此,寻找BC细胞特异性结合的分子片段具有重要意义.本文利用噬菌体肽库Biopanning技术,以人BC MCF-7细胞株为靶细胞,人胚肾HEK293细胞株为非特异性吸附细胞,进行四轮Biopanning,从末次洗脱库随机挑取60个噬菌体克隆,通过ELISA筛选阳性噬菌体克隆,对获得的36个阳性噬菌体克隆进行测序,获得7组共有(Consensus)序列克隆群,随后每组选1个OD_(450)最高克隆,以MCF-7细胞和BC组织作免疫荧光鉴定,筛选和验证了Consensus序列中的最佳序列.结果表明,S35噬菌体克隆所代表的Consensus序列(CSB1)对MCF-7细胞及BC组织结合特异性和敏感性最佳,随即命名该序列为CSB1.生物信息学分析表明,肽CSB1靶向5-脂氧合酶激活蛋白(FLAP),该蛋白与BC的进展和预后有关.本研究筛选鉴定的CSB1序列在未来开发BC分子影像检测试剂,特别是研发BC靶向化疗药物递送新方法的过程中具有突出的潜在应用价值.Breast cancer(BC)is a popular malignancy in women with high incidence rate and mortality.At present,there are still many deficiencies in the early diagnosis and targeted chemotherapy of BC.It is urgently needed to develop new methods of the targeting diagnosis and chemotherapy for BC.Therefore,the selection and identification of the specifically binding molecular fragments to BC cells are of great significance.This research performed four rounds of Biopannings,using phage peptide library Biopanning technology to put human BC MCF-7 cell line as target cells and human embryonic kidney HEK293 cell line as non-specific adsorbent cells.60 phage clones were randomly selected from the final elution library,and positive phage clones were screened by ELISA.The 36 positive phage clones obtained were sequenced to obtain 7 sets of consensus sequence clone groups.Subsequently,one clone with the highest OD_(450) was selected from each group,and immunofluorescence identification was performed on MCF-7 cells and BC tissues to screen and verify the best sequence in the Consensus sequence.The results showed that the Consensus sequence(CSB1)represented by the S35 phage clone had the best binding specificity and sensitivity to MCF-7 cells and BC tissues,and was subsequently named CSB1.Bioinformatics analysis shows that peptide CSB1 targets 5-lipoxygenase activating protein(FLAP),which is associated with the progression and prognosis of breast cancer.The CSB1 sequence screened and identified in this study has outstanding potential application value in the future development of BC molecular imaging detection reagents,especially in the development of new methods for BC targeted chemotherapy drug delivery.

关 键 词：乳腺癌 噬菌体展示肽库 靶向多肽 肿瘤靶向诊治 

分 类 号：Q945[生物学—植物学]