The Superior Anticancer Effect Of Paclitaxel-nanoparticles In Tumor Diseases

BACKGROUNDSMalignant tumor is one of the common disease with the second most lethal in the world,which causes 1.269‰ of the total fatality in China.Paclitaxel(Ptx)is a first line drug of cancer chemotherapy.Ptx blocks cell mitosis by inhibiting depolymerization of cellular microtubule.Tetrandrine(Tet)is a bibenzylisoquinoline alkaloid which extracted from traditional Chinese medicine Tetradra.Previous experiments of our group demonstrated that Tet induced intracellular oxidative stress and suppressed proliferation of cancer cells.At present,defects of paclitaxel reagents,such as nonspecific toxic effect and low solution in water,still limited the clinical application.Nanotechnology were explored in many studied to delivery chemotherapeutics more efficient and less toxicity.OBJECTIVESThis study tried to synthesize self-assembled "carrier free" Ptx nanoparticles and ?ROS responsive fractured? Ptx-Tet-m PEG-TK supramolecular nanoparticles.Further experiments explored the antitumor efficacy and molecular mechanisms of these characteristic nanoparticles.METHODSFor Ptx-NPs in part 1,MTT assay and clonogenic assay were tested to analyze the cytotoxicity in A549 lung adenocarcinoma cells.Ptx-NPs induced apoptosis was observed through flow cytometer and fluorescence microscope.Western blot was used to study apoptosis proteins expression.Finally,in vivo lung cancer xenograft model showed the antitumor efficacy of Ptx-NPs.In part 2,the microscopic morphology of P/T-NPs was studied by Transmission Electron Microscopy(TEM).Mean diameter and zeta potential of the nanospheres were analyzed by Dynamic Laser scattering(DLS).High performance liquid chromatography(HPLC)was used to examine the loading efficiency of P/T-NPs.In vitro cytotoxicity was studied in lung adenocarcinoma cells A549 and gastric adenocarcinoma cells SGC-7901 by MTT assay.Intracellular oxidative stress was tested through flow cytometer.Autophagy was measured via flow cytometer,fluorescence observation and electron microscope.Further proteins expression of autophagy was studied by LC3? transfected cells and western blot.Autophagy inhibitor 3MA and antioxidant ROS were co-incubated with P/T-NPs to examine the relationship between ROS,autophagy and anticancer effect.RESULTS AND DISCUSSIONSTwo kinds of Ptx loaded nano-delivery systems were synthesized.In the first part,we successfully obtained Ptx-SA-PEG nanoparticles in accordance with previous work.In MTT test and clonogenic assay,Ptx-NPs treatment exhibited more effective inhibition of tumor compared with free Ptx.Flow cytometer and fluorescence showed that more apoptosis was induced by Ptx-NPs.Moreover,Ptx-NPs induced the upregulation of apoptosis-proteins expression.The subcutaneous xenograft model showed superior antitumor effect of Ptx-NPs over free Ptx.The conjugation of Ptx with SA and PEG made it possible for the "carrier-free" nanoparticles,which exerted a more stable and durable antitumor effect via cell apoptosis both in vitro and in vivo,providing a potential strategy to improve the clinical therapy of Ptx.In the second part,we designed a novel supramolecular nanomaterial which conjugated Ptx and Tet with ?ROS sensitive? thioketal(TK)to improve synergistic drug delivery in intracellular high ROS area.The physical characteristics of P/T-NPs,such as median size,zeta potential and drug release curve,were successfully verified.The stronger cytotoxicity and antitumor effect of P/T-NPs were analyzed in lung adenocarcinoma cells A549 and gastric adenocarcinoma cells SGC-7901.P/T-NPs treatment induced more cell autophagy and modulated autophagic proteins.Furthermore,both 3-MA and NAC could decrease the cytotoxicity of drugs.NAC also inhibited cell autophagy which induced by P/T-NPs.Therefore,the ?ROS sensitive? conjugated Ptx-Tet-TK nanoparticles demonstrated a superior cancer suppressive effect associated with autophagic cell death and could be a promising antitumor agent for many cancer treatment with more efficiency.CONCLUSIONSIn the current study,we successfully synthesized two nanoparticles with special characteristics.Both Ptx-NPs and P/T-NPs showed superior therapeutic effect compared with free drugs.Therefore,further exploration of nano-delivery system is a novel field for designing new reagent of paclitaxel and other drugs with potential clinical application.