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目的:采用固相法人工合成异硫氰酸荧光素(fluorescein isothiocyanate,FITC)标记的新型肿瘤归巢穿膜肽t Ly P-1(CGNKRTR),并于体外检测其肿瘤归巢及穿膜特性。方法:采用Na-芴甲氧羰基(Fmoc)法固相合成t Ly P-1,并用FITC标记N端。高效液相色谱及质谱仪检测其纯度及分子量;激光共聚焦显微镜及流式细胞仪观察其肿瘤归巢及穿膜能力;CCK8(cell counting kit-8)评价其细胞毒性。结果:合成的FITC-t Ly P-1纯度为99.14%,分子量为1 334.7;激光共聚焦显微镜可见FITCt Ly P-1在MDA-MB-231组细胞内分布多于HUVEC组,流式细胞仪检测MDA-MB-231组细胞内荧光强度值高于HUVEC组;CCK8检测不同浓度的FITC-t Ly P-1对细胞活性没有明显影响(P>0.05)。结论:采用固相法成功合成FITC标记的新型肿瘤归巢穿膜肽t Ly P-1,具有乳腺癌MDA-MB-231肿瘤靶向性和穿膜性,能作为一种潜在的肿瘤靶向药物运输载体。
OBJECTIVE: To establish a novel tumor-associated transmembrane peptide t Ly P-1 (CGNKRTR) labeled with fluorescein isothiocyanate (FITC) synthesized by solid-phase method and detect its tumor homing and transmembrane properties in vitro . Methods: t Ly P-1 was synthesized by Na-Fmoc method and labeled with FITC. Purity and molecular weight were determined by HPLC and mass spectrometry. Laser confocal microscopy and flow cytometry were used to observe the tumor homing and transmembrane capacity. CCK8 (cell counting kit-8) was used to evaluate the cytotoxicity. Results: The purity of FITC-t Ly P-1 was 99.14% and the molecular weight was 1 334.7. Confocal laser scanning microscopy showed that FITCt Ly P-1 was more distributed in MDA-MB-231 group than in HUVEC group. Flow cytometry The intracellular fluorescent intensity of MDA-MB-231 group was higher than that of HUVEC group. The detection of different concentrations of FITC-t Ly P-1 by CCK8 had no significant effect on cell activity (P> 0.05). CONCLUSION: FITC-labeled novel tumor homing transmembrane peptide, t Ly P-1, has been successfully synthesized by solid-phase method. It possesses the tumor targeting and transmembrane properties of breast cancer MDA-MB-231 and can be used as a potential tumor target Drug delivery vehicle.