| 4,5,6,7-Tetrachloro-2’,4’,5’,7-tetraiodofluorescein (Rose Bengal, RB) is a derivative of fluorescein, while it can act as a photosensitizer which can produce reactive oxygen species (ROS) under light of specific wavelength. In this way, it causes damage to pathological cells and has been applied as photosensitizer for phtotdynamic therapy of many diseases. Rescent research has shown that it also exerts intrinsic cytotoxicity to tumor cells in the absence of light which makes it can act as a chemotherapeutic agent in the treatment of cancer. It was found in clinical trial that both injected and non-injected melanoma lesions in patients regressed after intralesional treatment of RB because of the systemic immune response. The research data have shown a bright future for RB in many cancer treatments. However, RB is a anionic molecule with low molecular weight, which makes it hard to cross cell membrane, and it is cleared rapidly from the blood after injection, and thus it can only be used by intraregional injection. These drawbacks have limited its clinical application.In this study, a derivative of RB with amino group was synthesized and conjugated to linear poly L-glutamic acid (PGA), a common drug delivery carrier. The polymer-drug conjugate (PGA-RB) we designed contained the hydrophobic part of RB and the hydrophilic part of glutamic acid residues, therefore it self-assembled into nanoparticles with grafting ratio of 8%, diameter of 160 nm and zeta potential of-30 mV in water. In this way, PGA-RB nanoparticles were prepared and its toxicity in the absence of light was equal to RB, but its phototoxiciy index was much higher than RB by 27.8,5.8, and 4.8 times on Bcap-37, A549 and HepG2, respectively. In the mechanism study, we found that PGA-RB possessed many advantages over RB such as more uptake and ROS generation in tumor cells compared to RB by flow cytometry analysis, perinuclear ditribution rather than even distribution by laser confocal microscope observation. PGA-RB also had much longer blood circulation time, which made its intravenous treatment possible. Therefore, we did in vivo antitumor activity experiment on nude mice model (Bcap-37) by intravenous injection, and found that PGA-RB showed higher treatment efficiency than RB. Altogether, the modification we did to RB had significantly improved its anti-cancer performance.Furthermore, we intended to verify if the oxydation reaction of luciferein catalyzed by luciferase (LUC) can be a reliable light source for excitation of photoseitizers in photodynamic therapy of cancers, so we immobilized luciferase with the PGA-RB nanoparticle by the controlling crosslinking degree. The PGA-RB-LUC nanoparticle remained its catalystic ability and accumulated in tumor cells as PGA-RB did. However, the in vitro antitumor study results showed luciferase cannot generate enough light for photodynamic therapy. |
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