The synthesis and study of 3PRGD^sub 2^-Pyro for targeted photosensitizer

Objectives: Photodynamic therapy (PDT) is a promising treatment for cancer. Photosensitizer (PS) plays an important role in photodynamic therapy. A hydrosoluble and targeted PS was synthesized by modifying pheophorbide a (Pyro) with E[PEG4-c(RGDfK)]2(3PRGD2), In addition, its application value was further investigated in this study. Methods: High Performance Liquid Chromatography (HPLC) and Mass Spectrum (MS) were used to analysis the purity and molecular weight of PS (3PRGD2 -Pyro). A549 cells was treated with Pyro, Pyro +PDT, 3PRGD2-Pyro and 3PRGD2-Pyro +PDT, respectively. Then the cell viability and 50% inhibition concentration were calculated by MTT assay. The confocal laser scanning microscopy was used to research the targeting in vitro of 3PRGD2-Pyro. The distribution of 3PRGD2-Pyro in vivo was evaluated by in vivo imaging system with subcutaneous A549 xenograft mice injected Pyro and 3PRGD2-Pyro. In vivo, we administered PBS, Pyro +PDT, 3PRGD2-Pyro +PDT through vena caudalis to each group of athymic nude mice with A549 tumors. And the volume of tumors were examined to detect the therapeutic effect of PDT. Result: The purity of 3PRGD2-Pyro was greater than 98%. In addition, the water solubility of 3PRGD2-Pyro was much better than Pyro. In MTT assay, 3PRGD2 -Pyro exhibited significant photocytotoxicity in A549 cells and lower darktoxicity. Pyro modified with 3PRGD2 reduced the survival rate of cells, and the 50%inhibition concentration for 3PRGD2-Pyro is 0.1μM,which is much lower than the 50%inhibition concentration for Pyro (0.3μM). The confocal laser scanning microscopy revealed that 3PRGD2-Pyro is well targeted to the cytomembrane of A549 cells. Through in vivo imaging system, it can be observed obviously that 3PRGD2-Pyro was abundant in tumor, while the uptake of Pyro in tumor was lower than that of 3PRGD2-Pyro. For the therapeutic effect of PDT, 3PRGD2-Pyro +PDT group can obviously reduced the tumor size, and the tumor disappeared after a week’s treatment. In addition, the decrease of tumor size was slower in Pyro +PDT group when compared to 3PRGD2-Pyro +PDT group, suggesting Pyro laser radiation constrain the tumor growth. Conclusions: In summary, targeted and hydrosoluble 3PRGD2-Pyro with lower darktoxicty exhibited satisfactory effect in photodynamic therapy, which should be of great potential in photosensitizer.