Department of Pharmacy , The 960 Hospital of Chinese People's Liberation Army , Jinan 250031 , China.
Section of Thoracic Surgery, Department of Surgery , University of Michigan Medical School , Ann Arbor , Michigan 48109 , United States.
Mol Pharm. 2019 May 6;16(5):2199-2213. doi: 10.1021/acs.molpharmaceut.9b00185. Epub 2019 Apr 18.
Tumor targeting agents are being developed for early tumor detection and therapeutics. We previously identified the peptide SNFYMPL (SNF*) and demonstrated its specific binding to human esophageal specimens of high-grade dysplasia (HGD) and adenocarcinoma with imaging ex vivo. Here, we aim to identify the target for this peptide and investigate its potential applications in imaging and drug delivery. With SNF* conjugated affinity chromatography, mass spectrum, Western blot, enzyme-linked immunosorbent assay (ELISA), and molecular docking, we found that the epithelial cell adhesion molecule (EpCAM) was the potential target of SNF*. Next, we showed that FITC-labeled SNF* (SNF*-FITC) colocalized with EpCAM antibody on the surface of esophageal adenocarcinoma cells OE33, and SNF*-FITC binding patterns significantly changed after EpCAM knockdown or exogenous EpCAM transfection. With the data from TCGA, we demonstrated that EpCAM was overexpressed in 17 types of cancers. Using colon and gastric adenocarcinoma cells and tissues as examples, we found that SNF*-FITC bound in a pattern was colocalized with EpCAM antibody, and the SNF* binding did not upregulate the EpCAM downstream Wnt signals. Subsequently, we conjugated SNF* with our previously constructed poly(histidine)-PEG/DSPE copolymer micelles. SNF* labeling significantly improved the micelle binding with colon and gastric adenocarcinoma cells in vitro, and enhanced the antitumor effects and decreased the toxicities of the micelles in vivo. In conclusion, we identified and validated SNF* as a specific peptide for EpCAM. The future potential use of SNF* peptide in multiple tumor surveillance and tumor-targeted therapeutics was demonstrated.
肿瘤靶向剂正在被开发用于早期肿瘤检测和治疗。我们之前鉴定了肽 SNFYMPL（SNF*），并证明其在体外显影中能够特异性结合人食管高级别异型增生（HGD）和腺癌标本。在这里，我们旨在确定该肽的靶标，并研究其在成像和药物输送方面的潜在应用。通过 SNF缀合亲和层析、质谱、Western blot、酶联免疫吸附测定（ELISA）和分子对接，我们发现上皮细胞黏附分子（EpCAM）是 SNF的潜在靶标。接下来，我们表明 FITC 标记的 SNF*（SNF*-FITC）与食管腺癌细胞 OE33 表面的 EpCAM 抗体共定位，并且 EpCAM 敲低或外源性 EpCAM 转染后 SNF*-FITC 结合模式发生明显变化。利用 TCGA 中的数据，我们证明 EpCAM 在 17 种癌症中过表达。使用结肠和胃腺癌细胞和组织作为示例，我们发现 SNF*-FITC 结合模式与 EpCAM 抗体共定位，并且 SNF结合不会上调 EpCAM 下游 Wnt 信号。随后，我们将 SNF与我们之前构建的聚（组氨酸）-PEG/DSPE 共聚胶束偶联。SNF标记显著提高了胶束与结肠和胃腺癌细胞在体外的结合能力，并增强了胶束在体内的抗肿瘤作用和降低了毒性。总之，我们鉴定并验证了 SNF作为 EpCAM 的特异性肽。展示了 SNF*肽在多种肿瘤监测和肿瘤靶向治疗中的未来潜在用途。
