Selection of tumor-binding ligands in cancer patients with phage display libraries
Selection of tumor-binding ligands in cancer patients with phage display libraries
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DOI：
10.1158/0008-5472.can-05-4441
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发表时间：
2006-08-01
期刊：
影响因子：
11.2
通讯作者：
Thomas, Christian
中科院分区：
文献类型：
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作者：
Krag, David N.;Shukla, Girja S.;Thomas, Christian
噬菌体展示已在体外和动物模型中被广泛用于产生配体和识别与癌症相关的靶点。我们在这里报道了在癌症患者中使用噬菌体展示文库来识别肿瘤靶向配体。8例IV期乳腺癌、黑色素瘤和胰腺癌患者静脉注射噬菌体展示随机多肽或ScFv文库(1.6×10(8)-1×10(11)转导单位/kg)，30min后切除肿瘤，恢复肿瘤归巢噬菌体。在三名患者中，使用从同一患者的肿瘤中回收和扩增的噬菌体重复淘洗是可能的。在多达三次输液中，没有观察到包括过敏反应在内的严重副作用。患者出现抗噬菌体抗体，在连续给药的10天协议窗口内达到次最大水平。所有患者均可获得肿瘤噬菌体。使用基于过滤器的ELISA法，从患者的子集中鉴定出几个克隆，这些克隆与来自同一患者的肿瘤结合，在该克隆中恢复了克隆。通过免疫染色、生物测定和实时荧光定量聚合酶链式反应(Real-time PCR)等方法证实了克隆与肿瘤的结合。与相同组织结构的非癌症细胞系和癌细胞系的结合研究显示了肿瘤结合克隆的特异性。对肿瘤归巢多肽克隆的插入序列分析显示有几个基序，表明克隆在人类肿瘤中非随机积累。这是第一个报道的一系列癌症患者接受噬菌体文库进行肿瘤靶向配体的连续淘洗。无毒性和恢复具有良好特性的克隆的能力是在癌症患者中进一步研究这项技术的第一步。
Phage display has been used extensively in vitro and in animal models to generate ligands and to identify cancer-relevant targets. We report here the use of phage-display libraries in cancer patients to identify tumor-targeting ligands. Eight patients with stage IV cancer, including breast, melanoma, and pancreas, had phage-displayed random peptide or scFv library (1.6 x 10(8)-1 x 10(11) transducing units/kg) administered i.v.; tumors were excised after 30 minutes; and tumor-homing phage were recovered. In three patients, repeat panning was possible using phage recovered and amplified from that same patient's tumor. No serious side effects, including allergic reactions, were observed with up to three infusions. Patients developed antiphage antibodies that reached a submaximal level within the 10-day protocol window for serial phage administration. Tumor phage were recoverable from all the patients. Using a filter-based ELISA, several clones from a subset of the patients were identified that bound to a tumor from the same patient in which clones were recovered. The clone-binding to tumor was confirmed by immunostaining, bioassay, and real-time PCR-based methods. Binding studies with noncancer and cancer cell lines of the same histology showed specificity of the tumor-binding clones. Analysis of insert sequences of tumor-homing peptide clones showed several motifs, indicating nonrandom accumulation of clones in human tumors. This is the first reported series of cancer patients to receive phage library for serial panning of tumor targeting ligands. The lack of toxicity and the ability to recover clones with favorable characteristics are a first step for further research with this technology in cancer patients.