Cd13Targeted Fluorescein-NGR-gadolinium Agent For Tumor Fluorescent And MR Imaging:Design, Synthesis And In Vitro Assay

Background and Objective:The CD13receptor of the tumor cellular membrane and neovascular endothelium plays an important role in the growth, proliferation, infiltration and metastasis of the tumor. In present study, NGR peptide ligands specifically targeting CD13receptor of the tumor was synthesized and conjugated with gadolinium and fluorescein,with the purpose to develop a bimodal contrast agent for fluorescence and MR imaging. Bioactivity and targeting ability of the compound in vitro were evaluated.Materials and Methods:The expression of CD13on the tumor cellular membrane was detected using immunohistochemistry. The experiment consists of experimental group (hepatocellular carcinoma of human being, HePG2and sarcoma of human being, HT1080) and control group (colon carcinoma of human being, HT29). The contrast agent was synthesized through chemical coupling. At first, fluorescent-NGR was synthesized, and gadolinium was coupled with it afterward. Some physical and chemical parameters of the compound were determined, and its targeting ability was evaluated using cellular fluorescence assay in vitro. a. experimental group:fluorescent-NGR was dispensed to glass sheet stained with cell HePG2and HT1080of experimental group and glass sheet stained with cell HT29of control group, respectively, and the fluorescence intensity was observed. b. the ability of NGR targets CD13was assayed through antagonistic experiment in vitro. Two batch experiments were conducted:batch one:HT1080antagonistic group, NGR was dispensed to glass sheet before being stained with cell HT1080, after sealing CD13, fluorescent-NGR was dispensed with equivalent volume as used in previous experimental group, and the fluorescence intensity was observed, respectively; batch two:HePG2antagonistic group, NGR was dispensed to glass sheet before being stained with cell HePG2, then dispensed fluorescent-NGR, with equivalent volume as used in experimental group (this time). a. assays for binding ability of fluorescent-NGR-gadolinium to cell of high CD13expression and cell of low CD13expression:glass sheet stained with HT1080of experimental group and glass sheet stained with HT29of control group were dispensed fluorescent-NGR-gadolinium, respectively, and their fluorescence intensities were observed, respectively. b. assays for bioactivity of fluorescent-NGR-gadolinium in vitro through antagonistic experiment. The assays were conducted in HT1080of experimental group and HT1080of antagonistic group. fluorescent-NGR-gadolinium was dispensed with gradually decreasing volume to the HT1080of experimental group staining on glass sheet; NGR was dispensed with gradually increasing volume to the HT1080of antagonistic group staining on glass sheet at first, then dispensed fluorescent-NGR-gadolinium to it with volume as used in the experimental group (this time), and their fluorescence intensities were observed and compared.Result:High CD13expression was detected in HePG2and HT1080of experimental group using histochemistry; low CD13expression was detected in HT29of control group; fluorescent-NGR coupled with gadolinium well in vitro; distinctive fluorescence was detected in HePG2and HT1080of experimental group that had been dispensed with fluorescent-NGR, and weak fluorescence was detected in HT29of control group. Fluorescence intensity of the cells in experimental group was significantly higher than that in control group and antagonistic group (P<0.005); Fluorescence intensity of the cells for assay of fluorescent-NGR-gadolinium in HT1080of experimental group was significantly higher than that in HT29of control group and HT1080of antagonistic group (P<0.005)...