人结肠癌血管特异结合短肽的噬菌体肽库筛选及鉴定

目的：通过建立小鼠肾包膜下荷人结肠癌模型、利用噬菌体肽库体内筛选技术,筛选并鉴定能够与人结肠癌血管内皮细胞特异结合的噬菌体呈现短肽.方法：制备免疫抑制小鼠肾包膜下人结肠癌移植瘤动物模型.将随机环七肽库通过尾静脉注入荷瘤鼠体内,回收归巢于肿瘤组织的噬菌体,进行4轮体内筛选,随机挑取20个克隆进行测序.细胞ELISA实验初步鉴定阳性噬菌体的内皮细胞结合能力.免疫细胞化学染色鉴定阳性噬菌体克隆与共培养内皮细胞的结合能力.免疫荧光技术检测短肽与共培养内皮细胞的特异性结合能力.结果：18个克隆测序正确,它们呈现2种氨基酸序列,命名为CV1及CV2,其呈现噬菌体称为pCV1及pCV2,其中CV2/pCV2重复次数多.细胞ELISA结果显示pCV2与Co-HUVECs（与结肠癌细胞共培养的人脐静脉内皮细胞）的结合明显高于HUVECs（人脐静脉内皮细胞）,有显著的差异性结合,而pCV1在Co-HUVECs,HUVECs上结合均较少,无差异性结合.免疫荧光染色结果显示,FITC-CV2特异性结合于Co-HUVECs的胞膜与核周胞质.结论：得到两个能特异性结合于结肠癌移植瘤的噬菌体单克隆pCV1及pCV2.pCV2具有特异结合结肠癌血管的能力,有可能用于结肠癌血管靶向治疗.

Selection and identification of a specific binding peptide targeting to blood vessel of human colon cancer

AIM:To select,identify and analyze a peptide binding specifically to blood vessels of human colon cancer by phage displayed peptide library in vivo.METHODS:Animal models were established using sub-renal capsular assay(SRCA) in immunosupressed mice implanted with human colon cancer xenografts.The phage displayed peptide library was injected intravenously into mice.After 4 rounds of selection,20 clones were picked up randomly and sequenced individually.The binding ability to Co-HUVECs of the positive phage cl...